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Sample records for cx43 gap junction

  1. Sulforaphane counteracts aggressiveness of pancreatic cancer driven by dysregulated Cx43-mediated gap junctional intercellular communication

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    Zhang, Yiyao; Isayev, Orkhan; Heilmann, Katharina; Schoensiegel, Frank; Liu, Li; Nessling, Michelle; Richter, Karsten; Labsch, Sabrina; Nwaeburu, Clifford C.; Mattern, Juergen; Gladkich, Jury; Giese, Nathalia; Werner, Jens; Schemmer, Peter; Gross, Wolfgang; Gebhard, Martha M.; Gerhauser, Clarissa; Schaefer, Michael; Herr, Ingrid

    2014-01-01

    The extreme aggressiveness of pancreatic ductal adenocarcinoma (PDA) has been associated with blocked gap junctional intercellular communication (GJIC) and the presence of cancer stem cells (CSCs). We examined whether disturbed GJIC is responsible for a CSC phenotype in established and primary cancer cells and patient tissue of PDA using interdisciplinary methods based in physiology, cell and molecular biology, histology and epigenetics. Flux of fluorescent dyes and gemcitabine through gap junctions (GJs) was intact in less aggressive cells but not in highly malignant cells with morphological dysfunctional GJs. Among several connexins, only Cx43 was expressed on the cell surface of less aggressive and GJIC-competent cells, whereas Cx43 surface expression was absent in highly malignant, E-cadherin-negative and GJIC-incompetent cells. The levels of total Cx43 protein and Cx43 phosphorylated at Ser368 and Ser279/282 were high in normal tissue but low to absent in malignant tissue. si-RNA-mediated inhibition of Cx43 expression in GJIC-competent cells prevented GJIC and induced colony formation and the expression of stem cell-related factors. The bioactive substance sulforaphane enhanced Cx43 and E-cadherin levels, inhibited the CSC markers c-Met and CD133, improved the functional morphology of GJs and enhanced GJIC. Sulforaphane altered the phosphorylation of several kinases and their substrates and inhibition of GSK3, JNK and PKC prevented sulforaphane-induced CX43 expression. The sulforaphane-mediated expression of Cx43 was not correlated with enhanced Cx43 RNA expression, acetylated histone binding and Cx43 promoter de-methylation, suggesting that posttranslational phosphorylation is the dominant regulatory mechanism. Together, the absence of Cx43 prevents GJIC and enhances aggressiveness, whereas sulforaphane counteracts this process, and our findings highlight dietary co-treatment as a viable treatment option for PDA. PMID:24742583

  2. HPV16 E6 Controls the Gap Junction Protein Cx43 in Cervical Tumour Cells

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    Peng Sun

    2015-10-01

    Full Text Available Human papillomavirus type 16 (HPV16 causes a range of cancers including cervical and head and neck cancers. HPV E6 oncoprotein binds the cell polarity regulator hDlg (human homologue of Drosophila Discs Large. Previously we showed in vitro, and now in vivo, that hDlg also binds Connexin 43 (Cx43, a major component of gap junctions that mediate intercellular transfer of small molecules. In HPV16-positive non-tumour cervical epithelial cells (W12G Cx43 localised to the plasma membrane, while in W12T tumour cells derived from these, it relocated with hDlg into the cytoplasm. We now provide evidence that E6 regulates this cytoplasmic pool of Cx43. E6 siRNA depletion in W12T cells resulted in restoration of Cx43 and hDlg trafficking to the cell membrane. In C33a HPV-negative cervical tumour cells expressing HPV16 or 18 E6, Cx43 was located primarily in the cytoplasm, but mutation of the 18E6 C-terminal hDlg binding motif resulted in redistribution of Cx43 to the membrane. The data indicate for the first time that increased cytoplasmic E6 levels associated with malignant progression alter Cx43 trafficking and recycling to the membrane and the E6/hDlg interaction may be involved. This suggests a novel E6-associated mechanism for changes in Cx43 trafficking in cervical tumour cells.

  3. Functional expression of gap junction gene Cx43 and the myogenic differentiation of rhabdomyosarcoma cells

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    林仲翔; 张志谦; 韩亚玲; C.C.G.Naus; K.R.Yu; H.Holtzer

    1995-01-01

    Rhabdomyosarcoma (RD) cells express low levels of the gap junction protein connexin 43 (Cx43), and its mRNA, and display very weak gap junctional intercellular communication (GJIC) as detected by Cx43 immunofluorescence, slot-blot and dye-transfer methods. These cells grow rapidly and show aberrant and incomplete myogenic differentiation. To investigate the role of gap junctions in these cells, the expression of Cx43 with relation to cell growth and myogenic differentiation in RD single-cell subclones-RDL3 and RDL6 is studied. The subclone RDL3 grows slowly and displays better myogenic differentiation. The expression of Cx43, its mRNA and the GJIC in RDL3 is comparable to that in normal myoblasts. Another subclone RDL6 which grows rapidly, but is poorly differentiated, expresses very low levels of Cx43 and its mRNA, and very weak GJIC. By using the calcium phosphate precipitate transfection technique, a full-length cDNA-encoding Cx43 and a pSV2neo have been introduced into the RDL6 cells. Several stably

  4. Interacting Network of the Gap Junction (GJ) Protein Connexin43 (Cx43) is Modulated by Ischemia and Reperfusion in the Heart.

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    Martins-Marques, Tania; Anjo, Sandra Isabel; Pereira, Paulo; Manadas, Bruno; Girão, Henrique

    2015-11-01

    The coordinated and synchronized cardiac muscle contraction relies on an efficient gap junction-mediated intercellular communication (GJIC) between cardiomyocytes, which involves the rapid anisotropic impulse propagation through connexin (Cx)-containing channels, namely of Cx43, the most abundant Cx in the heart. Expectedly, disturbing mechanisms that affect channel activity, localization and turnover of Cx43 have been implicated in several cardiomyopathies, such as myocardial ischemia. Besides gap junction-mediated intercellular communication, Cx43 has been associated with channel-independent functions, including modulation of cell adhesion, differentiation, proliferation and gene transcription. It has been suggested that the role played by Cx43 is dictated by the nature of the proteins that interact with Cx43. Therefore, the characterization of the Cx43-interacting network and its dynamics is vital to understand not only the molecular mechanisms underlying pathological malfunction of gap junction-mediated intercellular communication, but also to unveil novel and unanticipated biological functions of Cx43. In the present report, we applied a quantitative SWATH-MS approach to characterize the Cx43 interactome in rat hearts subjected to ischemia and ischemia-reperfusion. Our results demonstrate that, in the heart, Cx43 interacts with proteins related with various biological processes such as metabolism, signaling and trafficking. The interaction of Cx43 with proteins involved in gene transcription strengthens the emerging concept that Cx43 has a role in gene expression regulation. Importantly, our data shows that the interactome of Cx43 (Connexome) is differentially modulated in diseased hearts. Overall, the characterization of Cx43-interacting network may contribute to the establishment of new therapeutic targets to modulate cardiac function in physiological and pathological conditions. Data are available via ProteomeXchange with identifier PXD002331.

  5. Diabetes Increases Cryoinjury Size with Associated Effects on Cx43 Gap Junction Function and Phosphorylation in the Mouse Heart

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    Joseph A. Palatinus

    2016-01-01

    Full Text Available Diabetic patients develop larger myocardial infarctions and have an increased risk of death following a heart attack. The poor response to myocardial injury in the diabetic heart is likely related to the many metabolic derangements from diabetes that create a poor substrate in general for wound healing, response to injury and infection. Studies in rodents have implicated a role for the gap junction protein connexin 43 (Cx43 in regulating the injury response in diabetic skin wounds. In this study, we sought to determine whether diabetes alters Cx43 molecular interactions or intracellular communication in the cryoinjured STZ type I diabetic mouse heart. We found that epicardial cryoinjury size is increased in diabetic mice and this increase is prevented by preinjury insulin administration. Consistent with these findings, we found that intercellular coupling via gap junctions is decreased after insulin administration in diabetic and nondiabetic mice. This decrease in coupling is associated with a concomitant increase in phosphorylation of Cx43 at serine 368, a residue known to decrease channel conductance. Taken together, our results suggest that insulin regulates both gap junction-mediated intercellular communication and injury propagation in the mouse heart.

  6. Levonorgestrel Inhibits Human Endometrial Cell Proliferation through the Upregulation of Gap Junctional Intercellular Communication via the Nuclear Translocation of Ser255 Phosphorylated Cx43

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    Xiaomiao Zhao

    2015-01-01

    Full Text Available Objects. To assess whether LNG exerts antiproliferation effects on human endometrial cells through changes of GJIC function and the phosphorylated Cx43. Methods. Cell proliferation and apoptosis of human endometrial stromal cells (HESCs and glandular cells (HEGCs treated with LNG in a dose- and time-dependent manner. GJIC change and further total Cx43 and serine 368 and 255 phosphorylated Cx43 were measured. Results. 5 × 10−5 mol/L LNG revealed a time-dependent inhibition of cell proliferation and an increase of apoptosis in both HESCs and HEGCs. Furthermore, these cells demonstrated a significant GJIC enhancement upon treatment with 5 × 10−5 mol/L for 48 hours. The effects of LNG were most noticeable in HESCs rather than in HEGCs. Associated with these changes, LNG induced a relative increase in total Cx43 in a time-dependent manner but not Ser368 phosphorylated Cx43. Moreover, laser scanning confocal microscope confirmed the increased expression of total Cx43 in the cytoplasm and, interestingly, the nuclear translocation of Ser255 phosphorylated Cx43. Conclusions. LNG likely inhibits the proliferation and promotes apoptosis in HESCs and HEGCs though an increase in gap junction permeability in vitro, which is achieved through the upregulation of Cx43 expression and the translocation of serine 255 phosphorylated Cx43 from the plasma to the nuclear compartment.

  7. Gap junctional protein Cx43 is involved in the communication between extracellular vesicles and mammalian cells

    NARCIS (Netherlands)

    Soares, Ana Rosa; Martins-Marques, Tania; Ribeiro-Rodrigues, Teresa; Ferreira, Joao Vasco; Catarino, Steve; Pinho, Maria Joao; Zuzarte, Monica; Anjo, Sandra Isabel; Manadas, Bruno; Sluijter, Joost P. G.; Pereira, Paulo; Girao, Henrique

    2015-01-01

    Intercellular communication is vital to ensure tissue and organism homeostasis and can occur directly, between neighbour cells via gap junctions (GJ), or indirectly, at longer distances, through extracellular vesicles, including exosomes. Exosomes, as intercellular carriers of messenger molecules, m

  8. Lentivirus-mediated RNAi knockdown of the gap junction protein, Cx43, attenuates the development of vascular restenosis following balloon injury.

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    Han, Xiao-Jian; Chen, Min; Hong, Tao; Zhu, Ling-Yu; He, Dan; Feng, Jiu-Geng; Jiang, Li-Ping

    2015-04-01

    Percutaneous coronary intervention [PCI or percutaneous transluminal coronary angioplasty (PTCA)] has been developed into a mature interventional treatment for atherosclerotic cardiovascular disease. However, the long-term therapeutic effect is compromised by the high incidence of vascular restenosis following angioplasty, and the underlying mechanisms of vascular restenosis have not yet been fully elucidated. In the present study, we investigated the role of the gap junction (GJ) protein, connexin 43 (Cx43), in the development of vascular restenosis. To establish vascular restenosis, rat carotid arteries were subjected to balloon angioplasty injury. At 0, 7, 14 and 2 days following balloon injury, the arteries were removed, and the intimal/medial area of the vessels was measured to evaluate the degree of restenosis. We found that the intimal area gradually increased following balloon injury. Intimal hyperplasia and restenosis were particularly evident at 14 and 28 days after injury. In addition, the mRNA and protein expression of Cx43 was temporarily decreased at 7 days, and subsequently increased at 14 and 28 days following balloon injury, as shown by RT-PCR and western blot analysis. To determine the involvement of Cx43 in vascular restenosis, the lentivirus vector expressing shRNA targeting Cx43, Cx43-RNAi-LV, was used to silence Cx43 in the rat carotid arteries. The knockdown of Cx43 effectively attenuated the development of intimal hyperplasia and vascular restenosis following balloon injury. Thus, our data indicate the vital role of the GJ protein, Cx43, in the development of vascular restenosis, and provide new insight into the pathogenesis of vascular restenosis. Cx43 may prove to be a novel potential pharmacological target for the prevention of vascular restenosis following PCI.

  9. Keratitis-Ichthyosis-Deafness syndrome-associated Cx26 mutants produce nonfunctional gap junctions but hyperactive hemichannels when co-expressed with wild type Cx43

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    García, Isaac E.; Maripillán, Jaime; Jara, Oscar; Ceriani, Ricardo; Palacios-Muñoz, Angelina; Ramachandran, Jayalakshimi; Olivero, Pablo; Pérez-Acle, Tomás; González, Carlos; Sáez, Juan C.; Contreras, Jorge E.; Martínez, Agustín D.

    2015-01-01

    Mutations in Cx26 gene are found in most cases of human genetic deafness. Some mutations produce syndromic deafness associated with skin disorders, like Keratitis Ichthyosis Deafness syndrome (KID). Because in the human skin Cx26 is co-expressed with other connexins, like Cx43 and Cx30, and since KID syndrome is inherited as autosomal dominant condition, it is possible that KID mutations change the way Cx26 interacts with other co-expressed connexins. Indeed, some Cx26 syndromic mutations showed gap junction dominant negative effect when co-expressed with wild type connexins, including Cx26 and Cx43. The nature of these interactions and the consequences on hemichannels and gap junction channels functions remain unknown. In this study we demonstrate that syndromic mutations at the N-terminus segment of Cx26, change connexin oligomerization compatibility, allowing aberrant interactions with Cx43. Strikingly, heteromeric oligomer formed by Cx43/Cx26 (syndromic mutants) show exacerbated hemichannel activity, but nonfunctional gap junction channels; this also occurs for those Cx26 KID mutants that do not show functional homomeric hemichannels. Heterologous expression of these hyperactive heteromeric hemichannels increases cell membrane permeability, favoring ATP release and Ca2+ overload. The functional paradox produced by oligomerization of Cx43 and Cx26 KID mutants could underlie the severe syndromic phenotype in human skin. PMID:25625422

  10. Keratitis-ichthyosis-deafness syndrome-associated Cx26 mutants produce nonfunctional gap junctions but hyperactive hemichannels when co-expressed with wild type Cx43.

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    García, Isaac E; Maripillán, Jaime; Jara, Oscar; Ceriani, Ricardo; Palacios-Muñoz, Angelina; Ramachandran, Jayalakshmi; Olivero, Pablo; Perez-Acle, Tomas; González, Carlos; Sáez, Juan C; Contreras, Jorge E; Martínez, Agustín D

    2015-05-01

    Mutations in Cx26 gene are found in most cases of human genetic deafness. Some mutations produce syndromic deafness associated with skin disorders, like the Keratitis-Ichthyosis-Deafness syndrome (KID). Because in the human skin connexin 26 (Cx26) is co-expressed with other connexins, like Cx43 and Cx30, and as the KID syndrome is inherited as autosomal dominant condition, it is possible that KID mutations change the way Cx26 interacts with other co-expressed connexins. Indeed, some Cx26 syndromic mutations showed gap junction dominant negative effect when co-expressed with wild-type connexins, including Cx26 and Cx43. The nature of these interactions and the consequences on hemichannels and gap junction channel (GJC) functions remain unknown. In this study, we demonstrate that syndromic mutations, at the N terminus segment of Cx26, change connexin oligomerization compatibility, allowing aberrant interactions with Cx43. Strikingly, heteromeric oligomer formed by Cx43/Cx26 (syndromic mutants) shows exacerbated hemichannel activity but nonfunctional GJCs; this also occurs for those Cx26 KID mutants that do not show functional homomeric hemichannels. Heterologous expression of these hyperactive heteromeric hemichannels increases cell membrane permeability, favoring ATP release and Ca(2+) overload. The functional paradox produced by oligomerization of Cx43 and Cx26 KID mutants could underlie the severe syndromic phenotype in human skin.

  11. Engineered Cx26 variants established functional heterotypic Cx26/Cx43 and Cx26/Cx40 gap junction channels.

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    Karademir, Levent B; Aoyama, Hiroshi; Yue, Benny; Chen, Honghong; Bai, Donglin

    2016-05-15

    Gap junction (GJ) channels mediate direct intercellular communication and are composed of two docked hemichannels (connexin oligomers). It is well documented that the docking and formation of GJs are possible only between compatible hemichannels (or connexins). The mechanisms of heterotypic docking compatibility are not fully clear. We aligned the protein sequences of docking-compatible and -incompatible connexins with that of connexin26 (Cx26). We found that two docking hydrogen bond (HB)-forming residues on the second extracellular domain (E2) of Cx26 and their equivalent residues are well conserved within docking-compatible connexins, but different between docking-incompatible connexins. Replacing one or both of these residues of Cx26 into the corresponding residues in the docking incompatible connexins (K168V, N176H or K168V-N176H) increased the formation of morphological and functional heterotypic GJs with connexin43 (Cx43) or connexin40 (Cx40), indicating that these two residues are important for docking incompatibility between Cx26 and these connexins. Our homology structure models predict that both HBs and hydrophobic interactions at the E2 docking interface are important docking mechanisms in heterotypic Cx26 K168V-N176H/Cx43 GJs and probably other docking compatible connexins. Revealing the key residues and mechanisms of heterotypic docking compatibility will assist us in understanding why these putative docking residues are hotspots of disease-linked mutants. © 2016 The Author(s). published by Portland Press Limited on behalf of the Biochemical Society.

  12. Expression of the gap junction gene connexin43 (Cx43) in preimplantation bovine embryos derived in vitro or in vivo.

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    Wrenzycki, C; Herrmann, D; Carnwath, J W; Niemann, H

    1996-09-01

    In this study we have examined the presence of mRNA encoding connexin 43 (Cx43) in bovine embryos derived in vivo and in vitro. Cumulus-oocyte complexes, immature and matured oocytes liberated from cumulus cells, zygotes, 2-4-cell and 8-16-cell embryos, morulae, blastocysts and hatched blastocysts were produced in vitro from ovaries obtained from an abattoir using TCM 199 supplemented with hormones and 10% oestrous cow serum for maturation. Cumulus-oocyte complexes matured for 24 h were exposed to bull spermatozoa for 19 h and then cultured in TCM 199 supplemented with 10% oestrous cow serum to the desired developmental stage. Morulae and blastocysts derived in vivo were collected from superovulated donor cows. Total RNA was extracted from pools of 60-200 bovine oocytes or embryos using a modified phenol-chloroform extraction method and analysed by reverse transcriptase polymerase chain reaction. Before reverse transcription, aliquots of DNase-digested embryonic RNA were tested by polymerase chain reaction using bovine-specific primers to control for residual genomic DNA contamination. DNA-free, total RNA was reverse transcribed after preincubation with the Cx43 specific 3'primer. The resultant cDNA was amplified by polymerase chain reaction using Cx43 specific primers that define a 516 bp fragment of Cx43. The reverse transcriptase polymerase chain reaction product was verified by restriction enzyme analysis with Alu I and sequencing. Assays were repeated at least twice for each developmental stage and provided identical results between replicates. Cx43 transcripts were detected in bovine morulae and blastocysts grown in vivo. In contrast, whereas the early in vitro stages from cumulus-oocyte complexes to morulae expressed Cx43, blastocysts and hatched blastocysts did not have detectable concentrations of mRNA from this gene. Restriction enzyme cutting revealed three fragments of the predicted size (139, 177, 200 bp). The amplified product showed 100% identity

  13. On the role of the gap junction protein Cx43 (GJA1 in human cardiac malformations with Fallot-pathology. a study on paediatric cardiac specimen.

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    Aida Salameh

    Full Text Available INTRODUCTION: Gap junction channels are involved in growth and differentiation. Therefore, we wanted to elucidate if the main cardiac gap junction protein connexin43 (GJA1 is altered in patients with Tetralogy of Fallot or double-outlet right ventricle of Fallot-type (62 patients referred to as Fallot compared to other cardiac anomalies (21 patients referred to as non-Fallot. Patients were divided into three age groups: 0-2years, 2-12years and >12years. Myocardial tissue samples were collected during corrective surgery and analysis of cell morphology, GJA1- and N-cadherin (CDH2-distribution, as well as GJA1 protein- and mRNA-expression was carried out. Moreover, GJA1-gene analysis of 16 patients and 20 healthy subjects was performed. RESULTS: Myocardial cell length and width were significantly increased in the oldest age group compared to the younger ones. GJA1 distribution changed significantly during maturation with the ratio of polar/lateral GJA1 increasing from 2.93±0.68 to 8.52±1.41. While in 0-2years old patients ∼6% of the lateral GJA1 was co-localised with CDH2 this decreased with age. Furthermore, the changes in cell morphology and GJA1-distribution were not due to the heart defect itself but were significantly dependent on age. Total GJA1 protein expression decreased during growing-up, whereas GJA1-mRNA remained unchanged. Sequencing of the GJA1-gene revealed only few heterozygous single nucleotide polymorphisms within the Fallot and the healthy control group. CONCLUSION: During maturation significant changes in gap junction remodelling occur which might be necessary for the growing and developing heart. In our study point mutations within the Cx43-gene could not be identified as a cause of the development of TOF.

  14. 缝隙连接蛋白Cx43在SD大鼠电点燃癫痫模型中的表达%Expression of gap junction connexin43 in the SD rats electrical kindling of epilepsy model

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    宋建华; 谢锡驹

    2012-01-01

    Objective To investigate the role of connexin (Cx) in pathogenesis of epilepsy. Methods Thirty - two male SD rats were randomly divided into 4 groups and 4 models were set up. The behaviors of all SD rats were observed meanwhile the EEGs were recorded. Before the end Cx43 in hippocampus was detected with Western blot. The relationship between gap junction connexin and the formation of epilepsy was observed through the intervention of medicine to the kindling - model. Results Rats displayed typical epileptic behaviors,and epileptic discharges were observed in the epilepsy model group. The expression of connexin 43 in the hippocampal tissue was significantly enhanced in the epilepsy model group,but reduced by carbenoxolone and phenytoin sodium. Carbenoxolone and phenytoin sodium reduced frequency and amplitude of epileptic waves. The increasing expression of Cx43 was inhibited. Conclusion Gap junction Cx43 is the material base for the formation of epilepsy. The inhibition of excessive expression of Cx43 can reduce the formation of abnormal gap junction. Consequently the formation of epilepsy is inhibited.%目的 探讨缝隙连接蛋白(Cx)在癫痫形成中的作用.方法 将32只雄性SD大鼠随机分为4组,分别造模.观察SD大鼠行为,记录脑电图.通过免疫蛋白印迹的方法检测Cx43在大鼠海马区的表达变化.通过药物对癫痫电点燃模型成模过程的干预来观察癫痫形成与缝隙连接蛋白间的关系.结果 模型组大鼠出现癫痫发作.蛋白印迹显示Cx43的表达异常增高.药物干预的2组SD大鼠模型痫性发作滞后,发作频率降低,程度减轻,痫性脑电波波幅降低,蛋白印迹显示Cx43的表达增高受到抑制.结论 缝隙连接蛋白是癫痫形成的物质基础.抑制Cx43的过度表达,减少异常缝隙连接的形成,癫痫形成也受到抑制,从而预防癫痫产生.

  15. Correlations of differentially expressed gap junction connexins Cx26, Cx30, Cx32, Cx43 and Cx46 with breast cancer progression and prognosis.

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    Ivett Teleki

    Full Text Available Connexins and their cell membrane channels contribute to the control of cell proliferation and compartmental functions in breast glands and their deregulation is linked to breast carcinogenesis. Our aim was to correlate connexin expression with tumor progression and prognosis in primary breast cancers.Meta-analysis of connexin isotype expression data of 1809 and 1899 breast cancers from the Affymetrix and Illumina array platforms, respectively, was performed. Expressed connexins were also monitored at the protein level in tissue microarrays of 127 patients equally representing all tumor grades, using immunofluorescence and multilayer, multichannel digital microscopy. Prognostic correlations were plotted in Kaplan-Meier curves and tested using the log-rank test and cox-regression analysis in univariate and multivariate models.The expression of GJA1/Cx43, GJA3/Cx46 and GJB2/Cx26 and, for the first time, GJA6/Cx30 and GJB1/Cx32 was revealed both in normal human mammary glands and breast carcinomas. Within their subfamilies these connexins can form homo- and heterocellular epithelial channels. In cancer, the array datasets cross-validated each other's prognostic results. In line with the significant correlations found at mRNA level, elevated Cx43 protein levels were linked with significantly improved breast cancer outcome, offering Cx43 protein detection as an independent prognostic marker stronger than vascular invasion or necrosis. As a contrary, elevated Cx30 mRNA and protein levels were associated with a reduced disease outcome offering Cx30 protein detection as an independent prognostic marker outperforming mitotic index and necrosis. Elevated versus low Cx43 protein levels allowed the stratification of grade 2 tumors into good and poor relapse free survival subgroups, respectively. Also, elevated versus low Cx30 levels stratified grade 3 patients into poor and good overall survival subgroups, respectively.Differential expression of Cx43 and Cx

  16. Effects of Banxia Xiexin Decoction and Its Components on Protein Expression of Cx43 in Gap Junction in Gastric Tissue of Electrogastric Dysrhythmias Rats%半夏泻心汤及其拆方对胃电节律失常大鼠胃组织缝隙连接蛋白Cx43表达的影响

    Institute of Scientific and Technical Information of China (English)

    李翠; 张冬梅; 娄利霞; 赵明镜; 李丽娜; 陈萌

    2015-01-01

    Objective To discuss compatibility regularity of Banxia Xiexin Decoction adjusting gastric mobility. Methods Ninty healthy SD male rats were divided into normal group (10 rats) and model group (80 rats). Rat models of electrogastric dysrhythmias were established. After the evaluation of gastric electrophysiology, Banxia Xiexin Decoction were divided into Xinkai, Kujiang, Ganbu, Xinkaikujiang, Xinkaiganbu, and Kujiangganbu groups according to the compatibility regularity of Banxia Xiexin Decoction, 10 rats for each group. Rats in each medication administration group received gavage with same concentration but different volume for 4 weeks. Each group was analyzed for gastric electrical parameters after administration. The expressions of Cx43 and Cx43 mRNA in the gastric tissue of rats in each group were analyzed by immunohistochemistry and real-time fluorescent quantitative PCR. Results Compare with the normal group, the expressions of Cx43 and Cx43 mRNA in the gastric tissue of rats increased significantly in the model group. Compared with the model group, variable coefficient of electrogastric slow wave frequency in all medication administration groups decreased obviously. Banxia Xiexin Decoction and its components had decreased the expression of Cx43 and Cx43 mRNA to some extent, among which Xinkaiganbu group had best effects. Conclusion The mechanism of Banxia Xiexin Decoction adjusting gastric motility maybe related to its function of decreasing the expressions of Cx43 and Cx43 mRNA, as to adjust disordered electrogastric rhythm.%目的:探讨半夏泻心汤调节胃运动的配伍规律。方法选取90只健康SD雄性大鼠,分为正常组10只和模型组80只。复制大鼠胃电节律失常模型,经胃电生理学指标评价后,根据半夏泻心汤的配伍特点,将其拆分为辛开、苦降、甘补、辛开苦降、辛开甘补、苦降甘补组,每组10只。各给药组按相同药物浓度不等体积连续灌胃4周。给药后进

  17. Connexin 43 mimetic peptide Gap27 reveals potential differences in the role of Cx43 in wound repair between diabetic and non-diabetic cells

    OpenAIRE

    Pollok, Simone; Pfeiffer, Ann-Catherine; Lobmann, Ralf; Wright, Catherine S; Moll, Ingrid; Martin, Patricia E M; Johanna M Brandner

    2010-01-01

    Abstract During early wound healing (WH) events Connexin 43 (Cx43) is down-regulated at wound margins. In chronic wound margins, including diabetic wounds, Cx43 expression is enhanced suggesting that down-regulation is important for WH. We previously reported that the Cx43 mimetic peptide Gap27 blocks Cx43 mediated intercellular communication and promotes skin cell migration of infant cells in vitro. In the present work we further investigated the molecular mechanism of Gap27 action and its t...

  18. Hindered Diffusion through an Aqueous Pore Describes Invariant Dye Selectivity of Cx43 Junctions☆

    OpenAIRE

    Heyman, Nathanael S.; Burt, Janis M.

    2008-01-01

    The permselectivity (permeance/conductance) of Cx43-comprised gap junctions is a variable parameter of junctional function. To ascertain whether this variability in junctional permselectivity is explained by heterogeneous charge or size selectivity of the comprising channels, the permeance of individual Cx43 gap junctions to combinations of two dyes differing in either size or charge was determined in four cell types: Rin43, NRKe, HeLa43, and cardiac myocytes. The results show that Cx43 junct...

  19. [Remodeling of cardiac gap junctions and arrhythmias].

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    Yu, Zhi-Bin; Sheng, Juan-Juan

    2011-12-25

    In the heart, gap junctions mediate electrical and chemical coupling between adjacent cardiomyocytes, forming the cell-to-cell pathways for orderly spread of the wave of electrical excitation responsible for a functional syncytium. Three principal connexins are expressed in cardiomyocytes, connexin 43 (CX43), CX40, and CX45. CX43 predominates in ventricular muscle cells. Most of the gap junctions, assembled from CX43, are located at the intercalated discs, often with larger junctional plaques at the disc periphery. The gap junctions are rarely distributed to the sides of the cardiomyocyte. The ischemia-reperfusion, cardiac hypertrophy, heart failure, hypercholesterolemia, and diabetes mellitus induce gap junction remodeling. The gap junction remodeling induced by above-mentioned diseases shows similar characteristics, including down-regulation of CX43, reduction in gap junction plaque size, increased heterogeneity and lateralization of gap junction distribution, and dephosphorylation of CX43. The elevated angiotensin II concentration in local myocardium may play an important role in the gap junction remodeling. The down-regulation of CX43 and lateralization of gap junction distribution alter anisotropic spread of the impulse of ventricular myocardium. The dephosphorylation of CX43 not only reduces electrical conductance, but also decreases permeability of chemicals between cardiomyocytes. The lateralization of gap junctions may increase the number of hemichannels formed by CX43. The opening of hemichannels induces ATP efflux and Na(+) influx, which forms a delayed after-depolarization. The gap junction remodeling in pathological condition produces arrhythmia substrate in the ventricles. In this review, the current knowledge on the relationship between the remodeling of cardiac gap junctions and arrhythmias were summarized.

  20. Connexin 43 mimetic peptide Gap27 reveals potential differences in the role of Cx43 in wound repair between diabetic and non-diabetic cells.

    Science.gov (United States)

    Pollok, Simone; Pfeiffer, Ann-Catherine; Lobmann, Ralf; Wright, Catherine S; Moll, Ingrid; Martin, Patricia E M; Brandner, Johanna M

    2011-04-01

    During early wound healing (WH) events Connexin 43 (Cx43) is down-regulated at wound margins. In chronic wound margins, including diabetic wounds, Cx43 expression is enhanced suggesting that down-regulation is important for WH. We previously reported that the Cx43 mimetic peptide Gap27 blocks Cx43 mediated intercellular communication and promotes skin cell migration of infant cells in vitro. In the present work we further investigated the molecular mechanism of Gap27 action and its therapeutic potential to improve WH in skin tissue and diabetic and non-diabetic cells. Ex vivo skin, organotypic models and human keratinocytes/fibroblasts of young and old donors and of diabetic and non-diabetic origin were used to assess the impact of Gap27 on cell migration, proliferation, Cx43 expression, localization, phosphorylation and hemichannel function. Exposure of ex vivo WH models to Gap27 decreased dye spread, accelerated WH and elevated cell proliferation. In non-diabetic cell cultures Gap27 decreased dye uptake through Cx hemichannels and after scratch wounding cells showed enhanced migration and proliferation. Cells of diabetic origin were less susceptible to Gap27 during early passages. In late passages these cells showed responses comparable to non-diabetic cells. The cause of the discrepancy between diabetic and non-diabetic cells correlated with decreased Cx hemichannel activity in diabetic cells but excluded differences in Cx43 expression, localization and Ser368-phosphorylation. These data emphasize the importance of Cx43 in WH and support the concept that Gap27 could be a beneficial therapeutic to accelerate normal WH. However, its use in diabetic WH may be restricted and our results highlight differences in the role of Cx43 in skin cells of different origin.

  1. Acetylation mediates Cx43 reduction caused by electrical stimulation

    Science.gov (United States)

    Meraviglia, Viviana; Azzimato, Valerio; Colussi, Claudia; Florio, Maria Cristina; Binda, Anna; Panariti, Alice; Qanud, Khaled; Suffredini, Silvia; Gennaccaro, Laura; Miragoli, Michele; Barbuti, Andrea; Lampe, Paul D.; Gaetano, Carlo; Pramstaller, Peter P.; Capogrossi, Maurizio C.; Recchia, Fabio A.; Pompilio, Giulio; Rivolta, Ilaria; Rossini, Alessandra

    2015-01-01

    Communication between cardiomyocytes depends upon Gap Junctions (GJ). Previous studies have demonstrated that electrical stimulation induces GJ remodeling and modifies histone acetylases (HAT) and deacetylases (HDAC) activities, although these two results have not been linked. The aim of this work was to establish whether electrical stimulation modulates GJ-mediated cardiac cell-cell communication by acetylation-dependent mechanisms. Field stimulation of HL-1 cardiomyocytes at 0.5 Hz for 24 hours significantly reduced Connexin43 (Cx43) expression and cell-cell communication. HDAC activity was down-regulated whereas HAT activity was not modified resulting in increased acetylation of Cx43. Consistent with a post-translational mechanism, we did not observe a reduction in Cx43 mRNA in electrically stimulated cells, while the proteasomal inhibitor MG132 maintained Cx43 expression. Further, the treatment of paced cells with the HAT inhibitor Anacardic Acid maintained both the levels of Cx43 and cell-cell communication. Finally, we observed increased acetylation of Cx43 in the left ventricles of dogs subjected to chronic tachypacing as a model of abnormal ventricular activation. In conclusion, our findings suggest that altered electrical activity can regulate cardiomyocyte communication by influencing the acetylation status of Cx43. PMID:26264759

  2. The role of the Cx43 C-terminus in GJ plaque formation and internalization.

    Science.gov (United States)

    Wayakanon, Praween; Bhattacharjee, Rajib; Nakahama, Ken-ichi; Morita, Ikuo

    2012-04-06

    Connexin 43 (Cx43) is a major gap junction (GJ) protein found in many mammalian cell types. The C-terminal (CT) domain of Cx43 has unique characteristics in terms of amino acid (aa) sequence and its length differs from other connexins. This CT domain can be associated with protein partners to regulate GJ assembly and degradation, which results in the direct control of gap junction intercellular communication (GJIC). However, the essential roles of the CT regions involved in these mechanisms have not been fully elucidated. In this study, we aimed to investigate the specific regions of Cx43CT involved in GJ formation and internalization. Wild type Cx43((382aa)) and 10 CT truncated mutants were stably expressed in HeLa cells as GFP or DsRed tagged proteins. First, we found that the deletion of 235-382aa from Cx43 resulted in failure to make GJ and establish GJIC. Second, the Cx43 with 242-382aa CT deletion could form functional GJs and be internalized as annular gap junctions (AGJs). However, the plaques consisting of Cx43 with CT deletions (Δ242-382aa to Δ271-382aa) were longer than the plaques consisting of Cx43 with CT deletions (Δ302-382aa). Third, co-culture experiments of cells expressing wild type Cx43((382)) with cells expressing Cx43CT mutants revealed that the directions of GJ internalization were dependent on the length of the respective CT. Moreover, a specific region, 325-342aa residues of Cx43, played an important role in the direction of GJ internalization. These results showed the important roles of the Cx43 C-terminus in GJ expression and its turnover.

  3. Mutations of Cx43 that affect B cell spreading in response to BCR signaling

    Directory of Open Access Journals (Sweden)

    Letitia Falk

    2014-07-01

    Full Text Available The gap junction (GJ protein connexin 43 (Cx43 is both necessary and sufficient for B cell receptor (BCR-mediated cell spreading. To address how Cx43 mediates this effect, we blocked its function genetically, by expressing mutants of Cx43, and pharmacologically, by using chemical inhibitors. While various point mutations of Cx43 inhibited B cell spreading, treatment with channel blocking drugs did not, suggesting that this response was independent of channel function. The critical region of Cx43 appears to be the cytoplasmic carboxyl-terminal (CT domain, which has previously been shown to be important for B cell spreading. Consistent with this, mutations of either tyrosine 247 or 265 found in the CT were sufficient to inhibit spreading. Thus Cx43 may influence B cell spreading by mechanisms requiring protein binding to, or modification of, these sites in the CT tail.

  4. Gap junctions - guards of excitability.

    Science.gov (United States)

    Stroemlund, Line Waring; Jensen, Christa Funch; Qvortrup, Klaus; Delmar, Mario; Nielsen, Morten Schak

    2015-06-01

    Cardiomyocytes are connected by mechanical and electrical junctions located at the intercalated discs (IDs). Although these structures have long been known, it is becoming increasingly clear that their components interact. This review describes the involvement of the ID in electrical disturbances of the heart and focuses on the role of the gap junctional protein connexin 43 (Cx43). Current evidence shows that Cx43 plays a crucial role in organizing microtubules at the intercalated disc and thereby regulating the trafficking of the cardiac sodium channel NaV1.5 to the membrane.

  5. Pathological implications of Cx43 down-regulation in human colon cancer.

    Science.gov (United States)

    Ismail, Rehana; Rashid, Rabiya; Andrabi, Khurshid; Parray, Fazl Q; Besina, Syed; Shah, Mohd Amin; Ul Hussain, Mahboob

    2014-01-01

    Connexin 43 is an important gap junction protein in vertebrates and is known for its tumor suppressive properties. Cx43 is abundantly expressed in the human intestinal epithelial cells and muscularis mucosae. To explore the role of Cx43 in the genesis of human colon cancer, we performed the expression analysis of Cx43 in 80 cases of histopathologically confirmed and clinically diagnosed human colon cancer samples and adjacent control tissue and assessed correlations with clinicopathological variables. Western blotting using anti-Cx43 antibody indicated that the expression of Cx43 was significantly down regulated (75%) in the cancer samples as compared to the adjacent control samples. Moreover, immunohistochemical analysis of the tissue samples confirmed the down regulation of the Cx43 in the intestinal epithelial cells. Cx43 down regulation showed significant association (pcancer. Our data demonstrated that loss of Cx43 may be an important event in colon carcinogenesis and tumor progression, providing significant insights about the tumor suppressive properties of the Cx43 and its potential as a diagnostic marker for colon cancer.

  6. The role of the Cx43 C-terminus in GJ plaque formation and internalization

    Energy Technology Data Exchange (ETDEWEB)

    Wayakanon, Praween; Bhattacharjee, Rajib [Department of Cellular Physiological Chemistry, Graduate School, Tokyo Medical and Dental University, 1-5-45 Yushima, Bunkyo-ku, Tokyo 113-8549 (Japan); Global Center of Excellence Program, International Research Center for Molecular Science in Tooth and Bone Diseases, Tokyo Medical and Dental University, 1-5-45 Yushima, Bunkyo-ku, Tokyo 113-8549 (Japan); Nakahama, Ken-ichi, E-mail: nakacell@tmd.ac.jp [Department of Cellular Physiological Chemistry, Graduate School, Tokyo Medical and Dental University, 1-5-45 Yushima, Bunkyo-ku, Tokyo 113-8549 (Japan); Morita, Ikuo [Department of Cellular Physiological Chemistry, Graduate School, Tokyo Medical and Dental University, 1-5-45 Yushima, Bunkyo-ku, Tokyo 113-8549 (Japan); Global Center of Excellence Program, International Research Center for Molecular Science in Tooth and Bone Diseases, Tokyo Medical and Dental University, 1-5-45 Yushima, Bunkyo-ku, Tokyo 113-8549 (Japan)

    2012-04-06

    Highlights: Black-Right-Pointing-Pointer Cx43-GFP or -DsRed fusion proteins were expressed in HeLa cells. Black-Right-Pointing-Pointer Roles of C-terminus were examined using various mutants. Black-Right-Pointing-Pointer Gap junction plaque size was dependent on the length of C-terminus. Black-Right-Pointing-Pointer C-terminus dependent gap junction plaque internalization was observed. -- Abstract: Connexin 43 (Cx43) is a major gap junction (GJ) protein found in many mammalian cell types. The C-terminal (CT) domain of Cx43 has unique characteristics in terms of amino acid (aa) sequence and its length differs from other connexins. This CT domain can be associated with protein partners to regulate GJ assembly and degradation, which results in the direct control of gap junction intercellular communication (GJIC). However, the essential roles of the CT regions involved in these mechanisms have not been fully elucidated. In this study, we aimed to investigate the specific regions of Cx43CT involved in GJ formation and internalization. Wild type Cx43{sub (382aa)} and 10 CT truncated mutants were stably expressed in HeLa cells as GFP or DsRed tagged proteins. First, we found that the deletion of 235-382aa from Cx43 resulted in failure to make GJ and establish GJIC. Second, the Cx43 with 242-382aa CT deletion could form functional GJs and be internalized as annular gap junctions (AGJs). However, the plaques consisting of Cx43 with CT deletions ({Delta}242-382aa to {Delta}271-382aa) were longer than the plaques consisting of Cx43 with CT deletions ({Delta}302-382aa). Third, co-culture experiments of cells expressing wild type Cx43{sub (382)} with cells expressing Cx43CT mutants revealed that the directions of GJ internalization were dependent on the length of the respective CT. Moreover, a specific region, 325-342aa residues of Cx43, played an important role in the direction of GJ internalization. These results showed the important roles of the Cx43 C-terminus in GJ

  7. THE EFFECT OF TRANSFECTED CX43 GENE ON THE GJIC AND PROLIFERATION OF GLIOMA CELLS

    Institute of Scientific and Technical Information of China (English)

    浦佩玉; 夏之柏; 黄强; 王春艳; 王广秀

    2002-01-01

    Objective: To evaluate the effect of Cx43 gene on gap junction intercellular communication (GJIC) and proliferation of glioma cells. Methods: Cx43 cDNA was transfected into TJ905 human glioblastoma cells using lipofectamine. The expression of Cx43 was identified by Northern blot analyses, in situ hybridization and immunohistochemistry. MTT assay and average number of AgNORs (Argyrophlic nuclear organizer regions) were used to determine the cell proliferation. TUNEL method was used for detection of cell apoptosis, and scrape loading and dye tranfer method for examination of GJIC. Results: The Cx43 expression was greatly upregulated when Cx43 gene was transfected into TJ905 glioma cells. The cell proliferation was inhibited while the cell apoptosis was not increased and GJIC was significantly restored in the glioma cells tranfected with Cx43 gene. Conclusion: Cx43 gene has an inhibitory effect on the glioma cell proliferation, but no effect on induction of cell apoptosis. The restoration of GJIC may be the major mechanism involved in its effect. Cx43 gene can be the candidate for gene therapy of gliomas.

  8. Phosphorylation on Ser-279 and Ser-282 of connexin43 regulates endocytosis and gap junction assembly in pancreatic cancer cells

    Science.gov (United States)

    Johnson, Kristen E.; Mitra, Shalini; Katoch, Parul; Kelsey, Linda S.; Johnson, Keith R.; Mehta, Parmender P.

    2013-01-01

    The molecular mechanisms regulating the assembly of connexins (Cxs) into gap junctions are poorly understood. Using human pancreatic tumor cell lines BxPC3 and Capan-1, which express Cx26 and Cx43, we show that, upon arrival at the cell surface, the assembly of Cx43 is impaired. Connexin43 fails to assemble, because it is internalized by clathrin-mediated endocytosis. Assembly is restored upon expressing a sorting-motif mutant of Cx43, which does not interact with the AP2 complex, and by expressing mutants that cannot be phosphorylated on Ser-279 and Ser-282. The mutants restore assembly by preventing clathrin-mediated endocytosis of Cx43. Our results also document that the sorting-motif mutant is assembled into gap junctions in cells in which the expression of endogenous Cx43 has been knocked down. Remarkably, Cx43 mutants that cannot be phosphorylated on Ser-279 or Ser-282 are assembled into gap junctions only when connexons are composed of Cx43 forms that can be phosphorylated on these serines and forms in which phosphorylation on these serines is abolished. Based on the subcellular fate of Cx43 in single and contacting cells, our results document that the endocytic itinerary of Cx43 is altered upon cell–cell contact, which causes Cx43 to traffic by EEA1-negative endosomes en route to lysosomes. Our results further show that gap-junctional plaques formed of a sorting motif–deficient mutant of Cx43, which is unable to be internalized by the clathrin-mediated pathway, are predominantly endocytosed in the form of annular junctions. Thus the differential phosphorylation of Cx43 on Ser-279 and Ser-282 is fine-tuned to control Cx43’s endocytosis and assembly into gap junctions. PMID:23363606

  9. Regulation of connexin43 gap junctional communication by phosphatidylinositol 4,5-bisphosphate

    NARCIS (Netherlands)

    van Zeijl, Leonie; Ponsioen, Bas; Giepmans, Ben N G; Ariaens, Aafke; Postma, Friso R; Várnai, Péter; Balla, Tamas; Divecha, Nullin; Jalink, Kees; Moolenaar, Wouter H

    2007-01-01

    Cell-cell communication through connexin43 (Cx43)-based gap junction channels is rapidly inhibited upon activation of various G protein coupled receptors; however, the mechanism is unknown. We show that Cx43-based cell-cell communication is inhibited by depletion of phosphatidylinositol 4,5-bisphosp

  10. Protein kinase C-dependent regulation of connexin43 gap junctions and hemichannels

    DEFF Research Database (Denmark)

    Alstrøm, Jette Skov; Stroemlund, Line Waring; Nielsen, Morten Schak

    2015-01-01

    and allow transport of molecules such as fluorescent dyes and ATP. A range of phosphorylated amino acids have been detected in the C-terminus of Cx43 and their physiological role has been intensively studied both in the gap junctional form of Cx43 and in its hemichannel configuration. We present the current...

  11. Inflammatory conditions induce gap junctional communication between rat Kupffer cells both in vivo and in vitro

    Science.gov (United States)

    Eugenín, Eliseo A.; González, Hernán E.; Sánchez, Helmuth A.; Brañes, María C.; Sáez, Juan C.

    2007-01-01

    Connexin43 (Cx43), a gap junction protein subunit, has been previously detected in Kupffer cells (KCs) during liver inflammation, however, KCs phagocytose cell debris that may include Cx43 protein, which could explain the detection of Cx43 in KCs. We determined that KCs express Cx43 and form gap junctions both in vivo and in vitro. In liver sections of animals treated with LPS, Cx43 was detected at ED2+ cells interfaces, indicating formation of GJ between KCs in vivo. In vitro, unstimulated KCs cultures did not form functional GJs, and expressed low levels of Cx43 that showed a diffuse intracellular distribution. In contrast, KCs treated with LPS plus IFN-γ, expressed a greater amount of Cx43 at both the, protein and mRNA levels, and showed Cx43 at cell-cell contacts associated with higher dye coupling. In conclusion, activation of KCs in vivo or in vitro resulted in enhanced Cx43 expression levels and formation of GJ that might play relevant roles during liver inflammation. PMID:17900549

  12. Gap junction intercellular communication mediated by connexin43 in astrocytes is essential for their resistance to oxidative stress.

    Science.gov (United States)

    Le, Hoa T; Sin, Wun Chey; Lozinsky, Shannon; Bechberger, John; Vega, José Luis; Guo, Xu Qiu; Sáez, Juan C; Naus, Christian C

    2014-01-17

    Oxidative stress induced by reactive oxygen species (ROS) is associated with various neurological disorders including aging, neurodegenerative diseases, as well as traumatic and ischemic insults. Astrocytes have an important role in the anti-oxidative defense in the brain. The gap junction protein connexin43 (Cx43) forms intercellular channels as well as hemichannels in astrocytes. In the present study, we investigated the contribution of Cx43 to astrocytic death induced by the ROS hydrogen peroxide (H2O2) and the mechanism by which Cx43 exerts its effects. Lack of Cx43 expression or blockage of Cx43 channels resulted in increased ROS-induced astrocytic death, supporting a cell protective effect of functional Cx43 channels. H2O2 transiently increased hemichannel activity, but reduced gap junction intercellular communication (GJIC). GJIC in wild-type astrocytes recovered after 7 h, but was absent in Cx43 knock-out astrocytes. Blockage of Cx43 hemichannels incompletely inhibited H2O2-induced hemichannel activity, indicating the presence of other hemichannel proteins. Panx1, which is predicted to be a major hemichannel contributor in astrocytes, did not appear to have any cell protective effect from H2O2 insults. Our data suggest that GJIC is important for Cx43-mediated ROS resistance. In contrast to hypoxia/reoxygenation, H2O2 treatment decreased the ratio of the hypophosphorylated isoform to total Cx43 level. Cx43 has been reported to promote astrocytic death induced by hypoxia/reoxygenation. We therefore speculate the increase in Cx43 dephosphorylation may account for the facilitation of astrocytic death. Our findings suggest that the role of Cx43 in response to cellular stress is dependent on the activation of signaling pathways leading to alteration of Cx43 phosphorylation states.

  13. Hindered Diffusion through an Aqueous Pore Describes Invariant Dye Selectivity of Cx43 Junctions☆

    Science.gov (United States)

    Heyman, Nathanael S.; Burt, Janis M.

    2008-01-01

    Abstract The permselectivity (permeance/conductance) of Cx43-comprised gap junctions is a variable parameter of junctional function. To ascertain whether this variability in junctional permselectivity is explained by heterogeneous charge or size selectivity of the comprising channels, the permeance of individual Cx43 gap junctions to combinations of two dyes differing in either size or charge was determined in four cell types: Rin43, NRKe, HeLa43, and cardiac myocytes. The results show that Cx43 junctions are size- but not charge-selective and that both selectivities are constant parameters of junctional function. The consistency of dye selectivities indicates that the large continuum of measured junctional permselectivities cannot be ascribed to an equivalent continuum of individual channel selectivities. Further, the relative dye permeance sequence of NBD-M-TMA ∼ Alexa 350 > Lucifer yellow > Alexa 488 ≫ Alexa 594 (Stokes radii of 4.3 Å, 4.4 Å, 4.9 Å, 5.8 Å, and 7.4 Å, respectively) and the conductance sequence of KCl > TEACl ∼ Kglutamate are well described by hindered diffusion through an aqueous pore with radius ∼10 Å and length 160 Å. The permselectivity and dye selectivity data suggest the variable presence in Cx43-comprised junctions of conductive channels that are either dye-impermeable or dye-permeable. PMID:17921206

  14. Interaction of c-Src with gap junction protein connexin-43. Role in the regulation of cell-cell communication

    NARCIS (Netherlands)

    Giepmans, B N; Hengeveld, T; Postma, F R; Moolenaar, W H

    2001-01-01

    Cell-cell communication via connexin-43 (Cx43)-based gap junctions is transiently inhibited by certain mitogens, but the underlying regulatory mechanisms are incompletely understood. Our previous studies have implicated the c-Src tyrosine kinase in mediating transient closure of Cx43-based gap junct

  15. Benzalkonium chloride suppresses rabbit corneal endothelium intercellular gap junction communication.

    Directory of Open Access Journals (Sweden)

    Zhenhao Zhang

    Full Text Available Gap junction intercellular communication (GJIC plays a critical role in the maintenance of corneal endothelium homeostasis. We determined if benzalkonium chloride (BAK alters GJIC activity in the rabbit corneal endothelium since it is commonly used as a drug preservative in ocular eyedrop preparations even though it can have cytotoxic effects.Thirty-six adult New Zealand albino rabbits were randomly divided into three groups. BAK at 0.01%, 0.05%, and 0.1% was applied twice daily to one eye of each of the rabbits in one of the three groups for seven days. The contralateral untreated eyes were used as controls. Corneal endothelial morphological features were observed by in vivo confocal microscopy (IVCM. Immunofluorescent staining resolved changes in gap junction integrity and localization. Western blot analysis and RT-PCR evaluated changes in levels of connexin43 (Cx43 and tight junction zonula occludens-1 (ZO-1 gene and protein expression, respectively. Cx43 and ZO-1 physical interaction was detected by immunoprecipitation (IP. Primary rabbit corneal endothelial cells were cultured in Dulbecco's Modified Eagle Medium (DMEM containing BAK for 24 hours. The scrape-loading dye transfer technique (SLDT was used to assess GJIC activity.Topical administration of BAK (0.05%, 0.1% dose dependently disrupted corneal endothelial cell morphology, altered Cx43 and ZO-1 distribution and reduced Cx43 expression. BAK also markedly induced increases in Cx43 phosphorylation status concomitant with decreases in the Cx43-ZO-1 protein-protein interaction. These changes were associated with marked declines in GJIC activity.The dose dependent declines in rabbit corneal endothelial GJIC activity induced by BAK are associated with less Cx43-ZO-1 interaction possibly arising from increases in Cx43 phosphorylation and declines in its protein expression. These novel changes provide additional evidence that BAK containing eyedrop preparations should be used with caution to

  16. Intracellular Cleavage of the Cx43 C-Terminal Domain by Matrix-Metalloproteases: A Novel Contributor to Inflammation?

    Directory of Open Access Journals (Sweden)

    Marijke De Bock

    2015-01-01

    Full Text Available The coordination of tissue function is mediated by gap junctions (GJs that enable direct cell-cell transfer of metabolic and electric signals. GJs are formed by connexin (Cx proteins of which Cx43 is most widespread in the human body. Beyond its role in direct intercellular communication, Cx43 also forms nonjunctional hemichannels (HCs in the plasma membrane that mediate the release of paracrine signaling molecules in the extracellular environment. Both HC and GJ channel function are regulated by protein-protein interactions and posttranslational modifications that predominantly take place in the C-terminal domain of Cx43. Matrix metalloproteases (MMPs are a major group of zinc-dependent proteases, known to regulate not only extracellular matrix remodeling, but also processing of intracellular proteins. Together with Cx43 channels, both GJs and HCs, MMPs contribute to acute inflammation and a small number of studies reports on an MMP-Cx43 link. Here, we build further on these reports and present a novel hypothesis that describes proteolytic cleavage of the Cx43 C-terminal domain by MMPs and explores possibilities of how such cleavage events may affect Cx43 channel function. Finally, we set out how aberrant channel function resulting from cleavage can contribute to the acute inflammatory response during tissue injury.

  17. Neurological manifestations of oculodentodigital dysplasia: a Cx43 channelopathy of the central nervous system?

    Directory of Open Access Journals (Sweden)

    Marijke eDe Bock

    2013-09-01

    Full Text Available The coordination of tissue function is mediated by gap junctions (GJs that enable direct cell-cell transfer of metabolic and electric signals. GJs are formed by connexins of which Cx43 is most widespread in the human body. In the brain, Cx43 GJs are mostly found in astroglia where they coordinate the propagation of Ca2+ waves, spatial K+ buffering and distribution of glucose. Beyond its role in direct intercellular communication, Cx43 also forms unapposed, non-junctional hemichannels in the plasma membrane of glial cells. These allow the passage of several neuro- and gliotransmitters that may, combined with downstream paracrine signaling, complement direct GJ communication among glial cells and sustain glial-neuronal signaling. Mutations in the GJA1 gene encoding Cx43 have been identified in a rare, mostly autosomal dominant syndrome called oculodentodigital dysplasia (ODDD. ODDD patients display a pleiotropic phenotype reflected by eye, hand, teeth and foot abnormalities, as well as craniofacial and bone malformations. Remarkably, neurological symptoms such as dysarthria, neurogenic bladder (manifested as urinary incontinence, spasticity or muscle weakness, ataxia, and epilepsy are other prominent features observed in ODDD patients. Over 10 mutations detected in patients diagnosed with neurological disorders are associated with altered functionality of Cx43 GJs/hemichannels, but the link between ODDD-related abnormal channel activities and neurologic phenotype is still elusive. Here, we present an overview on the nature of the mutants conveying structural and functional changes of Cx43 channels and discuss available evidence for aberrant Cx43 GJ and hemichannel function. In a final step, we examine the possibilities of how channel dysfunction may lead to some of the neurological manifestations of ODDD.

  18. TLR2 mediates gap junctional intercellular communication through connexin-43 in intestinal epithelial barrier injury.

    Science.gov (United States)

    Ey, Birgit; Eyking, Annette; Gerken, Guido; Podolsky, Daniel K; Cario, Elke

    2009-08-14

    Gap junctional intercellular communication (GJIC) coordinates cellular functions essential for sustaining tissue homeostasis; yet its regulation in the intestine is not well understood. Here, we identify a novel physiological link between Toll-like receptor (TLR) 2 and GJIC through modulation of Connexin-43 (Cx43) during acute and chronic inflammatory injury of the intestinal epithelial cell (IEC) barrier. Data from in vitro studies reveal that TLR2 activation modulates Cx43 synthesis and increases GJIC via Cx43 during IEC injury. The ulcerative colitis-associated TLR2-R753Q mutant targets Cx43 for increased proteasomal degradation, impairing TLR2-mediated GJIC during intestinal epithelial wounding. In vivo studies using mucosal RNA interference show that TLR2-mediated mucosal healing depends functionally on intestinal epithelial Cx43 during acute inflammatory stress-induced damage. Mice deficient in TLR2 exhibit IEC-specific alterations in Cx43, whereas administration of a TLR2 agonist protects GJIC by blocking accumulation of Cx43 and its hyperphosphorylation at Ser368 to prevent spontaneous chronic colitis in MDR1alpha-deficient mice. Finally, adding the TLR2 agonist to three-dimensional intestinal mucosa-like cultures of human biopsies preserves intestinal epithelial Cx43 integrity and polarization ex vivo. In conclusion, Cx43 plays an important role in innate immune control of commensal-mediated intestinal epithelial wound repair.

  19. Regulation of gap-junction protein connexin 43 by β-adrenergic receptor stimulation in rat cardiomyocytes

    Institute of Scientific and Technical Information of China (English)

    Yi XIA; Kai-zheng GONG; Ming XU; You-yi ZHANG; Ji-hong GUO; Yao SONG; Ping ZHANG

    2009-01-01

    Aim:β-adrenergic receptor (β-AR) agonists are among the most potent factors regulating cardiac electrophysiological properties.Connexin 43 (Cx43),the predominant gap-junction protein in the heart,has an indispensable role in modulating cardiac electric activities by affecting gap-junction function.The present study investigates the effects of short-term stimulation of β-AR subtypes on Cx43 expression and gap junction intercellular communication (GJIC) function.Methods:The level of Cx43 expression in neonatal rat cardiomyocytes (NRCM) was detected by a Western blotting assay.The GJIC function was evaluated by scrape loading/dye transfer assay.Results:Stimulation of β-AR by the agonist isoproterenol for 5 min induces the up-regulation of nonphosphorylated Cx43 protein level,but not total Cx43.Selective β2-AR inhibitor ICI 118551,but not β-AR inhibitor CGP20712,could fully abolish the effect.Moreover,pretreatment with both protein kinase A inhibitor H89 and G,protein inhibitor pertussis toxin also inhibited the isoproterenol-induced increase of nonphosphorylated Cx43 expression.Isoproterenol-induced up-regulation of nonphosphorylated Cx43 is accompanied with enhanced GJIC function.Conclusion:Taken together,β2-AR stimulation increases the expression of nonphosphorylated Cx43,thereby enhancing the gating function of gap junctions in cardiac myocytes in both a protein kinase A-and G1-dependent manner.

  20. The gap junction protein connexin43 interacts with the second PDZ domain of the zona occludens-1 protein

    NARCIS (Netherlands)

    Giepmans, B N; Moolenaar, W H

    1998-01-01

    Gap junctions mediate cell-cell communication in almost all tissues and are composed of channel-forming integral membrane proteins, termed connexins [1-3]. Connexin43 (Cx43) is the most widely expressed and the most well-studied member of this family. Cx43-based cell-cell communication is regulated

  1. Cx43, ZO-1, alpha-catenin and beta-catenin in cataractous lens epithelial cells

    Indian Academy of Sciences (India)

    Anshul I Arora; Kaid Johar; Devarshi U Gajjar; Darshini A Ganatra; Forum B Kayastha; Anuradha K Pal; Alpesh R Patel; Rajkumar S; Abhay R Vasavada

    2012-12-01

    Specimens of the anterior lens capsule with an attached monolayer of lens epithelial cells (LECs) were obtained from patients (=52) undergoing cataract surgery. Specimens were divided into three groups based on the type of cataract: nuclear cataract, cortical cataract and posterior subcapsular cataract (PSC). Clear lenses (=11) obtained from donor eyes were used as controls. Expression was studied by immunofluorescence, real-time PCR and Western blot. Statistical analysis was done using the student’s -test. Immunofluorescence results showed punctate localization of Cx43 at the cell boundaries in controls, nuclear cataract and PSC groups. In the cortical cataract group, cytoplasmic pools of Cx43 without any localization at the cell boundaries were observed. Real-time PCR results showed significant up-regulation of Cx43 in nuclear and cortical cataract groups. Western blot results revealed significant increase in protein levels of Cx43 and significant decrease of ZO-1 in all three cataract groups. Protein levels of alpha-catenin were decreased significantly in nuclear and cortical cataract group. There was no significant change in expression of beta-catenin in the cataractous groups. Our findings suggest that ZO-1 and alpha-catenin are important for gap junctions containing Cx43 in the LECs. Alterations in cell junction proteins may play a role during formation of different types of cataract.

  2. Gap junction protein connexin-43 interacts directly with microtubules

    NARCIS (Netherlands)

    Giepmans, B N; Verlaan, I; Hengeveld, T; Janssen, H; Calafat, J; Falk, M M; Moolenaar, W H

    2001-01-01

    Gap junctions are specialized cell-cell junctions that mediate intercellular communication. They are composed of connexin proteins, which form transmembrane channels for small molecules [1, 2]. The C-terminal tail of connexin-43 (Cx43), the most widely expressed connexin member, has been implicated

  3. Ischaemia-induced autophagy leads to degradation of gap junction protein connexin43 in cardiomyocytes.

    Science.gov (United States)

    Martins-Marques, Tania; Catarino, Steve; Zuzarte, Monica; Marques, Carla; Matafome, Paulo; Pereira, Paulo; Girão, Henrique

    2015-04-15

    GJIC (gap junction intercellular communication) between cardiomyocytes is essential for synchronous heart contraction and relies on Cx (connexin)-containing channels. Increased breakdown of Cx43 has been often associated with various cardiac diseases. However, the mechanisms whereby Cx43 is degraded in ischaemic heart remain unknown. The results obtained in the present study, using both HL-1 cells and organotypic heart cultures, show that simulated ischaemia induces degradation of Cx43 that can be prevented by chemical or genetic inhibitors of autophagy. Additionally, ischaemia-induced degradation of Cx43 results in GJIC impairment in HL-1 cells, which can be restored by autophagy inhibition. In cardiomyocytes, ubiquitin signals Cx43 for autophagic degradation, through the recruitment of the ubiquitin-binding proteins Eps15 (epidermal growth factor receptor substrate 15) and p62, that assist in Cx43 internalization and targeting to autophagic vesicles, via LC3 (light chain 3). Moreover, we establish that degradation of Cx43 in ischaemia or I/R (ischaemia/reperfusion) relies upon different molecular players. Indeed, degradation of Cx43 during early periods of ischaemia depends on AMPK (AMP-activated protein kinase), whereas in late periods of ischaemia and I/R Beclin 1 is required. In the Langendorff-perfused heart, Cx43 is dephosphorylated in ischaemia and degraded during I/R, where Cx43 degradation correlates with autophagy activation. In summary, the results of the present study provide new evidence regarding the molecular mechanisms whereby Cx43 is degraded in ischaemia, which may contribute to the development of new strategies that aim to preserve GJIC and cardiac function in ischaemic heart.

  4. The Cx43-like connexin protein Cx40.8 is differentially localized during fin ontogeny and fin regeneration.

    Science.gov (United States)

    Gerhart, Sarah V; Eble, Diane M; Burger, R Michael; Oline, Stefan N; Vacaru, Ana; Sadler, Kirsten C; Jefferis, Rebecca; Iovine, M Kathryn

    2012-01-01

    Connexins (Cx) are the subunits of gap junctions, membraneous protein channels that permit the exchange of small molecules between adjacent cells. Cx43 is required for cell proliferation in the zebrafish caudal fin. Previously, we found that a Cx43-like connexin, cx40.8, is co-expressed with cx43 in the population of proliferating cells during fin regeneration. Here we demonstrate that Cx40.8 exhibits novel differential subcellular localization in vivo, depending on the growth status of the fin. During fin ontogeny, Cx40.8 is found at the plasma membrane, but Cx40.8 is retained in the Golgi apparatus during regeneration. We next identified a 30 amino acid domain of Cx40.8 responsible for its dynamic localization. One possible explanation for the differential localization is that Cx40.8 contributes to the regulation of Cx43 in vivo, perhaps modifying channel activity during ontogenetic growth. However, we find that the voltage-gating properties of Cx40.8 are similar to Cx43. Together our findings reveal that Cx40.8 exhibits differential subcellular localization in vivo, dependent on a discrete domain in its carboxy terminus. We suggest that the dynamic localization of Cx40.8 differentially influences Cx43-dependent cell proliferation during ontogeny and regeneration.

  5. The Cx43-like connexin protein Cx40.8 is differentially localized during fin ontogeny and fin regeneration.

    Directory of Open Access Journals (Sweden)

    Sarah V Gerhart

    Full Text Available Connexins (Cx are the subunits of gap junctions, membraneous protein channels that permit the exchange of small molecules between adjacent cells. Cx43 is required for cell proliferation in the zebrafish caudal fin. Previously, we found that a Cx43-like connexin, cx40.8, is co-expressed with cx43 in the population of proliferating cells during fin regeneration. Here we demonstrate that Cx40.8 exhibits novel differential subcellular localization in vivo, depending on the growth status of the fin. During fin ontogeny, Cx40.8 is found at the plasma membrane, but Cx40.8 is retained in the Golgi apparatus during regeneration. We next identified a 30 amino acid domain of Cx40.8 responsible for its dynamic localization. One possible explanation for the differential localization is that Cx40.8 contributes to the regulation of Cx43 in vivo, perhaps modifying channel activity during ontogenetic growth. However, we find that the voltage-gating properties of Cx40.8 are similar to Cx43. Together our findings reveal that Cx40.8 exhibits differential subcellular localization in vivo, dependent on a discrete domain in its carboxy terminus. We suggest that the dynamic localization of Cx40.8 differentially influences Cx43-dependent cell proliferation during ontogeny and regeneration.

  6. Mouse Hepatitis Virus Infection Remodels Connexin43-Mediated Gap Junction Intercellular Communication In Vitro and In Vivo.

    Science.gov (United States)

    Basu, Rahul; Banerjee, Kaveri; Bose, Abhishek; Das Sarma, Jayasri

    2015-12-16

    Gap junctions (GJs) form intercellular channels which directly connect the cytoplasm between neighboring cells to facilitate the transfer of ions and small molecules. GJs play a major role in the pathogenesis of infection-associated inflammation. Mutations of gap junction proteins, connexins (Cxs), cause dysmyelination and leukoencephalopathy. In multiple sclerosis (MS) patients and its animal model experimental autoimmune encephalitis (EAE), Cx43 was shown to be modulated in the central nervous system (CNS). The mechanism behind Cx43 alteration and its role in MS remains unexplored. Mouse hepatitis virus (MHV) infection-induced demyelination is one of the best-studied experimental animal models for MS. Our studies demonstrated that MHV infection downregulated Cx43 expression at protein and mRNA levels in vitro in primary astrocytes obtained from neonatal mouse brains. After infection, a significant amount of Cx43 was retained in endoplasmic reticulum/endoplasmic reticulum Golgi intermediate complex (ER/ERGIC) and GJ plaque formation was impaired at the cell surface, as evidenced by a reduction of the Triton X-100 insoluble fraction of Cx43. Altered trafficking and impairment of GJ plaque formation may cause the loss of functional channel formation in MHV-infected primary astrocytes, as demonstrated by a reduced number of dye-coupled cells after a scrape-loading Lucifer yellow dye transfer assay. Upon MHV infection, a significant downregulation of Cx43 was observed in the virus-infected mouse brain. This study demonstrates that astrocytic Cx43 expression and function can be modulated due to virus stress and can be an appropriate model to understand the basis of cellular mechanisms involved in the alteration of gap junction intercellular communication (GJIC) in CNS neuroinflammation. We found that MHV infection leads to the downregulation of Cx43 in vivo in the CNS. In addition, results show that MHV infection impairs Cx43 expression in addition to gap junction

  7. Gap Junctions

    Science.gov (United States)

    Nielsen, Morten Schak; Axelsen, Lene Nygaard; Sorgen, Paul L.; Verma, Vandana; Delmar, Mario; Holstein-Rathlou, Niels-Henrik

    2013-01-01

    Gap junctions are essential to the function of multicellular animals, which require a high degree of coordination between cells. In vertebrates, gap junctions comprise connexins and currently 21 connexins are known in humans. The functions of gap junctions are highly diverse and include exchange of metabolites and electrical signals between cells, as well as functions, which are apparently unrelated to intercellular communication. Given the diversity of gap junction physiology, regulation of gap junction activity is complex. The structure of the various connexins is known to some extent; and structural rearrangements and intramolecular interactions are important for regulation of channel function. Intercellular coupling is further regulated by the number and activity of channels present in gap junctional plaques. The number of connexins in cell-cell channels is regulated by controlling transcription, translation, trafficking, and degradation; and all of these processes are under strict control. Once in the membrane, channel activity is determined by the conductive properties of the connexin involved, which can be regulated by voltage and chemical gating, as well as a large number of posttranslational modifications. The aim of the present article is to review our current knowledge on the structure, regulation, function, and pharmacology of gap junctions. This will be supported by examples of how different connexins and their regulation act in concert to achieve appropriate physiological control, and how disturbances of connexin function can lead to disease. © 2012 American Physiological Society. Compr Physiol 2:1981-2035, 2012. PMID:23723031

  8. Gap junctions.

    Science.gov (United States)

    Nielsen, Morten Schak; Axelsen, Lene Nygaard; Sorgen, Paul L; Verma, Vandana; Delmar, Mario; Holstein-Rathlou, Niels-Henrik

    2012-07-01

    Gap junctions are essential to the function of multicellular animals, which require a high degree of coordination between cells. In vertebrates, gap junctions comprise connexins and currently 21 connexins are known in humans. The functions of gap junctions are highly diverse and include exchange of metabolites and electrical signals between cells, as well as functions, which are apparently unrelated to intercellular communication. Given the diversity of gap junction physiology, regulation of gap junction activity is complex. The structure of the various connexins is known to some extent; and structural rearrangements and intramolecular interactions are important for regulation of channel function. Intercellular coupling is further regulated by the number and activity of channels present in gap junctional plaques. The number of connexins in cell-cell channels is regulated by controlling transcription, translation, trafficking, and degradation; and all of these processes are under strict control. Once in the membrane, channel activity is determined by the conductive properties of the connexin involved, which can be regulated by voltage and chemical gating, as well as a large number of posttranslational modifications. The aim of the present article is to review our current knowledge on the structure, regulation, function, and pharmacology of gap junctions. This will be supported by examples of how different connexins and their regulation act in concert to achieve appropriate physiological control, and how disturbances of connexin function can lead to disease. © 2012 American Physiological Society. Compr Physiol 2:1853-1872, 2012.

  9. Connexin26 Mutations Causing Palmoplantar Keratoderma and Deafness Interact with Connexin43, Modifying Gap Junction and Hemichannel Properties.

    Science.gov (United States)

    Shuja, Zunaira; Li, Leping; Gupta, Shashank; Meşe, Gülistan; White, Thomas W

    2016-01-01

    Mutations in GJB2 (connexin [Cx]26) cause either deafness or deafness associated with skin diseases. That different disorders can be caused by distinct mutations within the same gene suggests that unique channel activities are influenced by each class of mutation. We have examined the functional characteristics of two human mutations, Cx26-H73R and Cx26-S183F, causing palmoplantar keratoderma (PPK) and deafness. Both failed to form gap junction channels or hemichannels when expressed alone. Coexpression of the mutants with wild-type Cx43 showed a transdominant inhibition of Cx43 gap junction channels, without reductions in Cx43 protein synthesis. In addition, the presence of mutant Cx26 shifted Cx43 channel gating and kinetics toward a more Cx26-like behavior. Coimmunoprecipitation showed Cx43 being pulled down more efficiently with mutant Cx26 than wild-type, confirming the enhanced formation of heteromeric connexons. Finally, the formation of heteromeric connexons resulted in significantly increased Cx43 hemichannel activity in the presence of Cx26 mutants. These findings suggest a common mechanism whereby Cx26 mutations causing PPK and deafness transdominantly influence multiple functions of wild-type Cx43. They also implicate a role for aberrant hemichannel activity in the pathogenesis of PPK and further highlight an emerging role for Cx43 in genetic skin diseases.

  10. Gap junction reduction in cardiomyocytes following transforming growth factor-β treatment and Trypanosoma cruzi infection

    Directory of Open Access Journals (Sweden)

    Mariana C Waghabi

    2009-12-01

    Full Text Available Gap junction connexin-43 (Cx43 molecules are responsible for electrical impulse conduction in the heart and are affected by transforming growth factor-β (TGF-β. This cytokine increases during Trypanosoma cruzi infection, modulating fibrosis and the parasite cell cycle. We studied Cx43 expression in cardiomyocytes exposed or not to TGF-β T. cruzi, or SB-431542, an inhibitor of TGF-β receptor type I (ALK-5. Cx43 expression was also examined in hearts with dilated cardiopathy from chronic Chagas disease patients, in which TGF-β signalling had been shown previously to be highly activated. We demonstrated that TGF-β treatment induced disorganised gap junctions in non-infected cardiomyocytes, leading to a punctate, diffuse and non-uniform Cx43 staining. A similar pattern was detected in T. cruzi-infected cardiomyocytes concomitant with high TGF-β secretion. Both results were reversed if the cells were incubated with SB-431542. Similar tests were performed using human chronic chagasic patients and we confirmed a down-regulation of Cx43 expression, an altered distribution of plaques in the heart and a significant reduction in the number and length of Cx43 plaques, which correlated negatively with cardiomegaly. We conclude that elevated TGF-β levels during T. cruzi infection promote heart fibrosis and disorganise gap junctions, possibly contributing to abnormal impulse conduction and arrhythmia that characterise severe cardiopathy in Chagas disease.

  11. Gap junction reduction in cardiomyocytes following transforming growth factor-beta treatment and Trypanosoma cruzi infection.

    Science.gov (United States)

    Waghabi, Mariana C; Coutinho-Silva, Robson; Feige, Jean-Jacques; Higuchi, Maria de Lourdes; Becker, David; Burnstock, Geoffrey; Araújo-Jorge, Tânia C de

    2009-12-01

    Gap junction connexin-43 (Cx43) molecules are responsible for electrical impulse conduction in the heart and are affected by transforming growth factor-beta (TGF-beta). This cytokine increases during Trypanosoma cruzi infection, modulating fibrosis and the parasite cell cycle. We studied Cx43 expression in cardiomyocytes exposed or not to TGF-beta T. cruzi, or SB-431542, an inhibitor of TGF-beta receptor type I (ALK-5). Cx43 expression was also examined in hearts with dilated cardiopathy from chronic Chagas disease patients, in which TGF-beta signalling had been shown previously to be highly activated. We demonstrated that TGF-beta treatment induced disorganised gap junctions in non-infected cardiomyocytes, leading to a punctate, diffuse and non-uniform Cx43 staining. A similar pattern was detected in T. cruzi-infected cardiomyocytes concomitant with high TGF-beta secretion. Both results were reversed if the cells were incubated with SB-431542. Similar tests were performed using human chronic chagasic patients and we confirmed a down-regulation of Cx43 expression, an altered distribution of plaques in the heart and a significant reduction in the number and length of Cx43 plaques, which correlated negatively with cardiomegaly. We conclude that elevated TGF-beta levels during T. cruzi infection promote heart fibrosis and disorganise gap junctions, possibly contributing to abnormal impulse conduction and arrhythmia that characterise severe cardiopathy in Chagas disease.

  12. Sox4 mediates Tbx3 transcriptional regulation of the gap junction protein Cx43

    NARCIS (Netherlands)

    Boogerd, C.J.; Wong, L.Y.; van den Boogaard, M.; Bakker, M.A.J.; Tessadori, F.; Bakkers, J.; 't Hoen, P.A.C.; Moorman, A.F.; Christoffels, V.M.; Barnett, P.

    2011-01-01

    Tbx3, a T-box transcription factor, regulates key steps in development of the heart and other organ systems. Here, we identify Sox4 as an interacting partner of Tbx3. Pull-down and nuclear retention assays verify this interaction and in situ hybridization reveals Tbx3 and Sox4 to co-localize

  13. The connexin43 carboxyl terminus and cardiac gap junction organization.

    Science.gov (United States)

    Palatinus, Joseph A; Rhett, J Matthew; Gourdie, Robert G

    2012-08-01

    The precise spatial order of gap junctions at intercalated disks in adult ventricular myocardium is thought vital for maintaining cardiac synchrony. Breakdown or remodeling of this order is a hallmark of arrhythmic disease of the heart. The principal component of gap junction channels between ventricular cardiomyocytes is connexin43 (Cx43). Protein-protein interactions and modifications of the carboxyl-terminus of Cx43 are key determinants of gap junction function, size, distribution and organization during normal development and in disease processes. Here, we review data on the role of proteins interacting with the Cx43 carboxyl-terminus in the regulation of cardiac gap junction organization, with particular emphasis on Zonula Occludens-1. The rapid progress in this area suggests that in coming years we are likely to develop a fuller understanding of the molecular mechanisms causing pathologic remodeling of gap junctions. With these advances come the promise of novel approach to the treatment of arrhythmia and the prevention of sudden cardiac death. This article is part of a Special Issue entitled: The Communicating junctions, composition, structure and characteristics. Copyright © 2011 Elsevier B.V. All rights reserved.

  14. Interfering amino terminal peptides and functional implications for heteromeric gap junction formation

    Directory of Open Access Journals (Sweden)

    Richard David Veenstra

    2013-05-01

    Full Text Available Connexin43 (Cx43 is widely expressed in many different tissues of the human body. In cells of some organs, Cx43 is co-expressed with other connexins (Cx, including Cx46 and Cx50 in lens, Cx40 in atrium, Purkinje fibers, and the blood vessel wall, Cx45 in heart, and Cx37 in the ovary. Interactions with the co-expressed connexins may have profound functional implications. The abilities of Cx37, Cx45, Cx46, and Cx50 to function in heteromeric gap junction combinations with Cx43 are well documented. Different studies disagree regarding the ability of Cx43 and Cx40 to produce functional heteromeric gap junctions with each other. We review previous studies regarding the heteromeric interactions of Cx43. The possibility of negative functional interactions between the cytoplasmic pore-forming amino terminal (NT domains of these connexins was assessed using pentameric connexin sequence-specific NT domain (iNT peptides applied to cells expressing homomeric Cx40, Cx37, Cx45, Cx46, and Cx50 gap junctions. A Cx43 iNT peptide corresponding to amino acids 9 to 13 (Ac-KLLDK-NH2 specifically inhibited the electrical coupling of Cx40 gap junctions in a transjunctional (Vj voltage-dependent manner without affecting the function of homologous Cx37, Cx46, Cx50, and Cx45 gap junctions. A Cx40 iNT (Ac-EFLEE-OH peptide counteracted the Vj-dependent block of Cx40 gap junctions, whereas a similarly charged Cx50 iNT (Ac-EEVNE-OH peptide did not, suggesting that these NT domain interactions are not solely based on electrostatics. These data are consistent with functional Cx43 heteromeric gap junction formation with Cx37, Cx45, Cx46, and Cx50 and suggest that Cx40 uniquely experiences functional suppressive interactions with a Cx43 NT domain sequence. These findings present unique functional implications about the heteromeric interactions between Cx43 and Cx40 that may influence cardiac conduction in atrial myocardium and the specialized conduction system.

  15. Effects of Angiotensin Ⅱ on Expression of the Gap Junction Channel Protein Connexin 43 in Neonatal Rat Ventricular Myocytes

    Institute of Scientific and Technical Information of China (English)

    Jun Yang; Wei Wu

    2007-01-01

    To study the effects of angiotensin Ⅱ,as a mediator of cardiac hypertrophy,on expression of connexin 43 (Cx43) in cultured neonatal rat ventricular myocytes and correlation of expression of Cx43 and cardiomyocyte hypertrophy.Methods Cardiomyocytes were isolated from newborn SD rats.Angiotensin Ⅱ was added into the media to induce myocyte hypertrophy.Cultures were exposed to 10 ~6 mol/L angiotensin Ⅱ for 72 h,Cx43 expression was characterized by RT-PCR and Immunofluorescence methods.Results Immunofluorescence analysis revealed decreased Cx43 immunoreactivity in cells treated for 72 h with angiotensin Ⅱ.RT-PCR analysis demonstrated there was an obvious decrease of Cx43 mRNA level in cells exposed to angiotensin Ⅱ for 72 h.The changes of expression of connexin 43 were related to its entrance into S phase of the cell cycle.Cultured neonatal rat cardiomyocytes were exposed for 72 h to increase concentrations of angiotensin Ⅱ ( 1.0 × 10-9 ~ 1.0 × 10-6mol/L),resulting in significantly decreased Cx43 expression.Conclusions Angiotensin Ⅱ leads to a concentration-dependent decrease in Cx43 protein in cultured neonatal rat ventricular myocytes by decreasing Cx43 mRNA synthesis.Signal transduction pathways activated by angiotensin Ⅱ under pathophysiologic conditions of cardiac hypertrophy could initiate remodeling of gap junctions.

  16. Activation of L-type calcium channels is required for gap junction-mediated intercellular calcium signaling in osteoblastic cells

    DEFF Research Database (Denmark)

    Jørgensen, Niklas Rye; Teilmann, Stefan Cuoni; Henriksen, Zanne

    2003-01-01

    of extracellular calcium, plasma membrane depolarization by high extracellular potassium, and the L-type voltage-operated calcium channel inhibitor, nifedipine. In contrast, all these treatments enhanced the spread of P2 receptor-mediated ICW in UMR rat osteoblastic cells. Using UMR cells transfected to express Cx......43 (UMR/Cx43) we confirmed that nifedipine sensitivity of ICW required Cx43 expression. In human osteoblastic cells, gap junction-dependent ICW also required activation of L-type calcium channels and influx of extracellular calcium....

  17. Gap junction protein connexin43 exacerbates lung vascular permeability.

    Directory of Open Access Journals (Sweden)

    James J O'Donnell

    Full Text Available Increased vascular permeability causes pulmonary edema that impairs arterial oxygenation and thus contributes to morbidity and mortality associated with Acute Respiratory Distress Syndrome and sepsis. Although components of intercellular adhesive and tight junctions are critical for maintaining the endothelial barrier, there has been limited study of the roles of gap junctions and their component proteins (connexins. Since connexins can modulate inflammatory signaling in other systems, we hypothesized that connexins may also regulate pulmonary endothelial permeability. The relationships between connexins and the permeability response to inflammatory stimuli were studied in cultured human pulmonary endothelial cells. Prolonged treatment with thrombin, lipopolysaccharide, or pathological cyclic stretch increased levels of mRNA and protein for the major connexin, connexin43 (Cx43. Thrombin and lipopolysaccharide both increased intercellular communication assayed by transfer of microinjected Lucifer yellow. Although thrombin decreased transendothelial resistance in these cells, the response was attenuated by pretreatment with the connexin inhibitor carbenoxolone. Additionally, the decreases of transendothelial resistance produced by either thrombin or lipopolysaccharide were attenuated by reducing Cx43 expression by siRNA knockdown. Both carbenoxolone and Cx43 knockdown also abrogated thrombin-induced phosphorylation of myosin light chain. Taken together, these data suggest that increased lung vascular permeability induced by inflammatory conditions may be amplified via increased expression of Cx43 and intercellular communication among pulmonary endothelial cells.

  18. Specificity of gap junction communication among human mammary cells and connexin transfectants in culture.

    Science.gov (United States)

    Tomasetto, C; Neveu, M J; Daley, J; Horan, P K; Sager, R

    1993-07-01

    In a previous paper (Lee et al., 1992), it was shown that normal human mammary epithelial cells (NMEC) express two connexin genes, Cx26 and Cx43, whereas neither gene is transcribed in a series of mammary tumor cell lines (TMEC). In this paper it is shown that normal human mammary fibroblasts (NMF) communicate and express Cx43 mRNA and protein. Transfection of either Cx26 or Cx43 genes into a tumor line, 21MT-2, induced the expression of the corresponding mRNAs and proteins as well as communication via gap junctions (GJs), although immunofluorescence demonstrated that the majority of Cx26 and Cx43 proteins present in transfected TMEC was largely cytoplasmic. Immunoblotting demonstrated that NMEC, NMF, and transfected TMEC each displayed a unique pattern of posttranslationally modified forms of Cx43 protein. The role of different connexins in regulating gap junction intercellular communication (GJIC) was examined using a novel two-dye method to assess homologous and heterologous communication quantitatively. The recipient cell population was prestained with a permanent non-toxic lipophilic dye that binds to membranes irreversibly (PKH26, Zynaxis); and the donor population is treated with a GJ-permeable dye Calcein, a derivative of fluorescein diacetate (Molecular Probes). After mixing the two cell populations under conditions promoting GJ formation, cells were analyzed by flow cytometry to determine the percentage of cells containing both dyes. It is shown here that Cx26 and Cx43 transfectants display strong homologous communication, as do NMEC and NMF. Furthermore, NMEC mixed with NMF communicate efficiently, Cx26 transfectants communicate with NMEC but not with NMF, and Cx43 transfectants communicate with NMF. Communication between Cx26 TMEC transfectants and NMEC was asymetrical with preferential movement of calcein from TMEC to NMEC. Despite the presence of Cx43 as well as Cx26 encoded proteins in the GJs of NMEC, few Cx43 transfectants communicated with NMEC

  19. Transient suppression of gap junctional intercellular communication after exposure to 100-nanosecond pulsed electric fields.

    Science.gov (United States)

    Steuer, Anna; Schmidt, Anke; Labohá, Petra; Babica, Pavel; Kolb, Juergen F

    2016-12-01

    Gap junctional intercellular communication (GJIC) is an important mechanism that is involved and affected in many diseases and injuries. So far, the effect of nanosecond pulsed electric fields (nsPEFs) on the communication between cells was not investigated. An in vitro approach is presented with rat liver epithelial WB-F344 cells grown and exposed in a monolayer. In order to observe sub-lethal effects, cells were exposed to pulsed electric fields with a duration of 100ns and amplitudes between 10 and 20kV/cm. GJIC strongly decreased within 15min after treatment but recovered within 24h. Gene expression of Cx43 was significantly decreased and associated with a reduced total amount of Cx43 protein. In addition, MAP kinases p38 and Erk1/2, involved in Cx43 phosphorylation, were activated and Cx43 became hyperphosphorylated. Immunofluorescent staining of Cx43 displayed the disassembly of gap junctions. Further, a reorganization of the actin cytoskeleton was observed whereas tight junction protein ZO-1 was not significantly affected. All effects were field- and time-dependent and most pronounced within 30 to 60min after treatment. A better understanding of a possible manipulation of GJIC by nsPEFs might eventually offer a possibility to develop and improve treatments.

  20. Construction of Mouse Melanoma Cell Models of Wild-type Overexpression Connexin 43 and Site-specific Mutant Connexin 43%过表达及定点突变缝隙连接蛋白Cx43小鼠黑色素瘤细胞模型的构建

    Institute of Scientific and Technical Information of China (English)

    李玢; 张广献; 刘娟; 赵青; 谭宇蕙; 吴映雅; 易华; 杜标炎

    2014-01-01

    目的:构建小鼠黑色素瘤细胞(B16)过表达野生型及点突变型缝隙连接蛋白43(Connexin43, Cx43)细胞模型,为以缝隙连接(Gap Junction, GJ)为靶点的中药复方、中药单药及药物单体的研究提供可靠阳性对照和阴性对照。方法构建野生型Cx43、突变型Cx43G21R、突变型Cx43G138R重组荧光蛋白融合慢病毒表达质粒,用定点突变技术获得Cx43G21R和Cx43G138R突变体,对上述3种质粒进行双酶切和测序鉴定,并分别包装病毒感染B16细胞,使B16细胞过表达野生型Cx43、突变型Cx43G21R、突变型Cx43G138R。 Western blot检测Cx43蛋白表达水平变化,荧光示踪法观察缝隙连接通讯(Gap Junction Intercellular Communication, GJIC)功能变化。结果①酶切及测序证明,成功构建 pLVCx43-mCherry、 pLVCx43-mCherryG21R、 pLVCx43-mCherryG138R重组荧光蛋白融合慢病毒表达质粒。②成功感染B16细胞并筛选稳定过表达Cx43Cx43G21R、Cx43G138R细胞株, Western blot检测显示上述细胞株Cx43蛋白表达均高于对照组。③过表达野生型Cx43后B16细胞GJIC功能较对照组增强;过表达突变型Cx43后B16细胞GJIC功能较对照组减弱。结论过表达野生型Cx43可增强B16细胞GJIC功能,过表达突变型Cx43可抑制B16细胞GJIC功能。%Objective To construct mouse B16 melanoma cell models of wild-type overexpression Cx43 and site-specific mutant Cx43, thus to provide reliable positive and negative control for the study of gap junction targeted Chinese herbal formula, Chinese medicine ingredient, and drug monomer. Methods We constructed the recombined fluorescent protein(mCherry) infused with Lentivirus expression plasmid of wild-type Cx43(pLVCx43-mCherry), mutant R (pLVCx43G21R-mCherry) and mutant Cx43G138R(pLVCx43 G138R-mCherry). The Cx43G21R and Cx43G138R mutants were obtained by site-specific mutagenesis. The three kinds of plasmid were performed double restriction

  1. High-dose insulin inhibits gap junction intercellular communication in vascular smooth muscle cells.

    Science.gov (United States)

    Bian, Ou; Zhang, Haishan; Guan, Qigang; Sun, Yingxian; Zeng, Dingyin

    2015-07-01

    Gap junction intercellular communication (GJIC) is important in mediating intercellular substance and signal transmission. Connexin (Cx)43 is a major component involved in GJIC in vascular tissue and its abnormal expression is closely associated with various vascular diseases. Insulin resistance is the central component of metabolic syndrome, and high doses of insulin can affect vascular function through multiple pathways, resulting in cardiovascular disease. However, the effects of insulin on GJIC function and connexin (Cx)43 expression in vascular smooth muscle cells (VSMCs) remain unclear. Following treatment of VSMCs with different doses of insulin, a fluorescence recovery after photobleaching (FRAP) assay was performed to evaluate GJIC function in treated VSMCs. The results showed that high‑dose insulin suppressed GJIC function. Western blot assays further demonstrated that high‑dose insulin induced the phosphorylation of Cx43 at s368 and downregulated the expression of Cx43. H2O2 release assays demonstrated that high‑dose insulin treatment significantly elevated the cellular H2O2 level. In addition, compared with cells treated with high‑dose insulin, pretreatment with catalase significantly restored the cellular GJIC function, decreased the phosphorylation level of Cx43 at s368, and enhanced Cx43 expression. In conclusion, these data indicate that high‑dose insulin inhibits cellular GJIC function through the oxidative stress‑activated signaling pathway. This phenomenon may also constitute a potential mechanism underlying the pathogenesis of insulin resistance and its complications.

  2. Microtubule-assisted altered trafficking of astrocytic gap junction protein connexin 43 is associated with depletion of connexin 47 during mouse hepatitis virus infection.

    Science.gov (United States)

    Basu, Rahul; Bose, Abhishek; Thomas, Deepthi; Das Sarma, Jayasri

    2017-09-08

    Gap junctions (GJs) are important for maintenance of CNS homeostasis. GJ proteins, connexin 43 (Cx43) and connexin 47 (Cx47), play a crucial role in production and maintenance of CNS myelin. Cx43 is mainly expressed by astrocytes in the CNS and forms gap junction intercellular communications between astrocytes-astrocytes (Cx43-Cx43) and between astrocytes-oligodendrocytes (Cx43-Cx47). Mutations of these connexin (Cx) proteins cause dysmyelinating diseases in humans. Previously, it has been shown that Cx43 localization and expression is altered due to mouse hepatitis virus (MHV)-A59 infection both in vivo and in vitro; however, its mechanism and association with loss of myelin protein was not elaborated. Thus, we explored potential mechanisms by which MHV-A59 infection alters Cx43 localization and examined the effects of viral infection on Cx47 expression and its association with loss of the myelin marker proteolipid protein. Immunofluorescence and total internal reflection fluorescence microscopy confirmed that MHV-A59 used microtubules (MTs) as a conduit to reach the cell surface and restricted MT-mediated Cx43 delivery to the cell membrane. Co-immunoprecipitation experiments demonstrated that Cx43-β-tubulin molecular interaction was depleted due to protein-protein interaction between viral particles and MTs. During acute MHV-A59 infection, oligodendrocytic Cx47, which is mainly stabilized by Cx43 in vivo, was down-regulated, and its characteristic staining remained disrupted even at chronic phase. The loss of Cx47 was associated with loss of proteolipid protein at the chronic stage of MHV-A59 infection. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  3. Hyperthermia Differently Affects Connexin43 Expression and Gap Junction Permeability in Skeletal Myoblasts and HeLa Cells

    Directory of Open Access Journals (Sweden)

    Ieva Antanavičiūtė

    2014-01-01

    Full Text Available Stress kinases can be activated by hyperthermia and modify the expression level and properties of membranous and intercellular channels. We examined the role of c-Jun NH2-terminal kinase (JNK in hyperthermia-induced changes of connexin43 (Cx43 expression and permeability of Cx43 gap junctions (GJs in the rabbit skeletal myoblasts (SkMs and Cx43-EGFP transfected HeLa cells. Hyperthermia (42°C for 6 h enhanced the activity of JNK and its target, the transcription factor c-Jun, in both SkMs and HeLa cells. In SkMs, hyperthermia caused a 3.2-fold increase in the total Cx43 protein level and enhanced the efficacy of GJ intercellular communication (GJIC. In striking contrast, hyperthermia reduced the total amount of Cx43 protein, the number of Cx43 channels in GJ plaques, the density of hemichannels in the cell membranes, and the efficiency of GJIC in HeLa cells. Both in SkMs and HeLa cells, these changes could be prevented by XG-102, a JNK inhibitor. In HeLa cells, the changes in Cx43 expression and GJIC under hyperthermic conditions were accompanied by JNK-dependent disorganization of actin cytoskeleton stress fibers while in SkMs, the actin cytoskeleton remained intact. These findings provide an attractive model to identify the regulatory players within signalosomes, which determine the cell-dependent outcomes of hyperthermia.

  4. Hyperthermia differently affects connexin43 expression and gap junction permeability in skeletal myoblasts and HeLa cells.

    Science.gov (United States)

    Antanavičiūtė, Ieva; Mildažienė, Vida; Stankevičius, Edgaras; Herdegen, Thomas; Skeberdis, Vytenis Arvydas

    2014-01-01

    Stress kinases can be activated by hyperthermia and modify the expression level and properties of membranous and intercellular channels. We examined the role of c-Jun NH2-terminal kinase (JNK) in hyperthermia-induced changes of connexin43 (Cx43) expression and permeability of Cx43 gap junctions (GJs) in the rabbit skeletal myoblasts (SkMs) and Cx43-EGFP transfected HeLa cells. Hyperthermia (42°C for 6 h) enhanced the activity of JNK and its target, the transcription factor c-Jun, in both SkMs and HeLa cells. In SkMs, hyperthermia caused a 3.2-fold increase in the total Cx43 protein level and enhanced the efficacy of GJ intercellular communication (GJIC). In striking contrast, hyperthermia reduced the total amount of Cx43 protein, the number of Cx43 channels in GJ plaques, the density of hemichannels in the cell membranes, and the efficiency of GJIC in HeLa cells. Both in SkMs and HeLa cells, these changes could be prevented by XG-102, a JNK inhibitor. In HeLa cells, the changes in Cx43 expression and GJIC under hyperthermic conditions were accompanied by JNK-dependent disorganization of actin cytoskeleton stress fibers while in SkMs, the actin cytoskeleton remained intact. These findings provide an attractive model to identify the regulatory players within signalosomes, which determine the cell-dependent outcomes of hyperthermia.

  5. Antofine-induced connexin43 gap junction disassembly in rat astrocytes involves protein kinase Cβ.

    Science.gov (United States)

    Huang, Yu-Fang; Liao, Chih-Kai; Lin, Jau-Chen; Jow, Guey-Mei; Wang, Hwai-Shi; Wu, Jiahn-Chun

    2013-03-01

    Antofine, a phenanthroindolizidine alkaloid derived from Cryptocaryachinensis and Ficusseptica in the Asclepiadaceae milkweed family, is cytotoxic for various cancer cell lines. In this study, we demonstrated that treatment of rat primary astrocytes with antofine induced dose-dependent inhibition of gap junction intercellular communication (GJIC), as assessed by scrape-loading 6-carboxyfluorescein dye transfer. Levels of Cx43 protein were also decreased in a dose- and time-dependent manner following antofine treatment. Double-labeling immunofluorescence microscopy showed that antofine (10ng/ml) induced endocytosis of surface gap junctions into the cytoplasm, where Cx43 was co-localized with the early endosome marker EEA1. Inhibition of lysosomes or proteasomes by co-treatment with antofine and their respective specific inhibitors, NH4Cl or MG132, partially inhibited the antofine-induced decrease in Cx43 protein levels, but did not inhibit the antofine-induced inhibition of GJIC. After 30min of treatment, antofine induced a rapid increase in the intracellular Ca(2+) concentration and activation of protein kinase C (PKC)α/βII, which was maintained for at least 6h. Co-treatment of astrocytes with antofine and the intracellular Ca(2+) chelator BAPTA-AM prevented downregulation of Cx43 and inhibition of GJIC. Moreover, co-treatment with antofine and a specific PKCβ inhibitor prevented endocytosis of gap junctions, downregulation of Cx43, and inhibition of GJIC. Taken together, these findings indicate that antofine induces Cx43 gap junction disassembly by the PKCβ signaling pathway. Inhibition of GJIC by antofine may undermine the neuroprotective effect of astrocytes in CNS.

  6. Photoperiod-Dependent Effects of 4-tert-Octylphenol on Adherens and Gap Junction Proteins in Bank Vole Seminiferous Tubules

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    Anna Hejmej

    2013-01-01

    Full Text Available In the present study we evaluated in vivo and in vitro effects of 4-tert-octylphenol (OP on the expression and distribution of adherens and gap junction proteins, N-cadherin, β-catenin, and connexin 43 (Cx43, in testes of seasonally breeding rodents, bank voles. We found that in bank vole testes expression and distribution of N-cadherin, β-catenin, and Cx43 were photoperiod dependent. Long-term treatment with OP (200 mg/kg b.w. resulted in the reduction of junction proteins expressions (P<0.05, P<0.01 and their delocalization in the testes of males kept in long photoperiod, whereas in short-day animals slight increase of Cx43 (P<0.05, N-cadherin, and β-catenin (statistically nonsignificant levels was observed. Effects of OP appeared to be independent of FSH and were maintained during in vitro organ culture, indicating that OP acts directly on adherens and gap junction proteins in the testes. An experiment performed using an antiestrogen ICI 182,780 demonstrated that the biological effects of OP on β-catenin and Cx43 involve an estrogen receptor-mediated response. Taken together, in bank vole organization of adherens and gap junctions and their susceptibility to OP are related to the length of photoperiod. Alterations in cadherin/catenin and Cx43-based junction may partially result from activation of estrogen receptor α and/or β signaling pathway.

  7. GAP junctional communication in brain secondary organizers.

    Science.gov (United States)

    Bosone, Camilla; Andreu, Abraham; Echevarria, Diego

    2016-06-01

    Gap junctions (GJs) are integral membrane proteins that enable the direct cytoplasmic exchange of ions and low molecular weight metabolites between adjacent cells. They are formed by the apposition of two connexons belonging to adjacent cells. Each connexon is formed by six proteins, named connexins (Cxs). Current evidence suggests that gap junctions play an important part in ensuring normal embryo development. Mutations in connexin genes have been linked to a variety of human diseases, although the precise role and the cell biological mechanisms of their action remain almost unknown. Among the big family of Cxs, several are expressed in nervous tissue but just a few are expressed in the anterior neural tube of vertebrates. Many efforts have been made to elucidate the molecular bases of Cxs cell biology and how they influence the morphogenetic signal activity produced by brain signaling centers. These centers, orchestrated by transcription factors and morphogenes determine the axial patterning of the mammalian brain during its specification and regionalization. The present review revisits the findings of GJ composed by Cx43 and Cx36 in neural tube patterning and discuss Cx43 putative enrollment in the control of Fgf8 signal activity coming from the well known secondary organizer, the isthmic organizer. © 2016 The Authors. Development, Growth & Differentiation published by John Wiley & Sons Australia, Ltd on behalf of Japanese Society of Developmental Biologists.

  8. Roles of gap junctions, connexins and pannexins in epilepsy

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    Shanthini eMylvaganam

    2014-05-01

    Full Text Available Enhanced gap junctional communication (GJC between neurons is considered a major factor underlying the neuronal synchrony driving seizure activity. In addition, the hippocampal sharp wave ripple complexes, associated with learning and seizures, are diminished by GJC blocking agents. Although gap junctional blocking drugs inhibit experimental seizures, they all have other nonspecific actions. Besides interneuronal GJC between dendrites, inter-axonal and inter-glial GJC is also considered important for seizure generation. Interestingly, in most studies of cerebral tissue from animal seizure models and from human patients with epilepsy, there is up-regulation of glial, but not neuronal gap junctional mRNA and protein. Significant changes in the expression and post-translational modification of the astrocytic connexin Cx43, and Panx1 were observed in an in vitro Co++ seizure model, further supporting a role for glia in seizure-genesis, although the reasons for this remain unclear. Further suggesting an involvement of astrocytic GJC in epilepsy, is the fact that the expression of astrocytic Cx mRNAs (Cxs 30 and 43 is several fold higher than that of neuronal Cx mRNAs (Cxs 36 and 45, and the number of glial cells outnumber neuronal cells in mammalian hippocampal and cortical tissue. Pannexin expression is also increased in both animal and human epileptic tissues. Specific Cx43 mimetic peptides, Gap 27 and SLS, inhibit the docking of astrocytic connexin Cx43 proteins from forming intercellular gap junctions, diminishing spontaneous seizures. Besides GJs, Cx membrane hemichannels in glia and Panx membrane channels in neurons and glia are also inhibited by gap junctional pharmacological blockers. Although there is no doubt that connexin-based gap junctions and hemichannels, and pannexin-based membrane channels are related to epilepsy, the specific details of how they are involved and how we can modulate their function for therapeutic purposes remain to

  9. Syndromic and non-syndromic disease-linked Cx43 mutations.

    Science.gov (United States)

    Laird, Dale W

    2014-04-17

    There are now at least 14 distinct diseases linked to germ line mutations in the 21 genes that encode the connexin (Cx) family of gap junction proteins. This review focuses on the links between germ-line mutations in the gene encoding Cx43 (GJA1) and the human disease termed oculodentodigital dysplasia (ODDD). This disease is clinically characterized by soft tissue fusion of the digits, abnormal craniofacial bone development, small eyes and loss of tooth enamel. However, the disease is considerably more complex and somewhat degenerative as patients often suffer from other syndromic effects that include incontinence, glaucoma, skin diseases and neuropathies that become more pronounced during aging. The challenge continues to be understanding how distinct Cx43 gene mutations cause such a diverse range of tissue phenotypes and pathophysiological changes while other Cx43-rich organs are relatively unaffected. This review will provide an overview of many of these studies and distill some themes and outstanding questions that need to be addressed in the coming years. Copyright © 2014 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

  10. Pharmacological enhancement of cardiac gap junction coupling prevents arrhythmias in canine LQT2 model.

    Science.gov (United States)

    Quan, Xiao-Qing; Bai, Rong; Lu, Jia-Gao; Patel, Chinmay; Liu, Nian; Ruan, Yanfei; Chen, Bo-Di; Ruan, Lei; Zhang, Cun-Tai

    2009-01-01

    Gap junctions contribute to the transmural heterogeneity of repolarization in the normal heart and under conditions of prolonged QT interval in the diseased heart. This study examined whether enhancing of gap junction coupling can reduce transmural dispersion of repolarization (TDR) and prevent torsade de pointes (TdP) in a canine LQT2 model. Canine left ventricular wedge preparations were perfused with delayed rectifier potassium current (IKr) blocker d-sotalol to mimic LQT2 and the antiarrhythmic peptide 10 (AAP10) was used as a gap junction coupling enhancer. As compared with the control group, the LQT2 group had significantly augmented TDR and higher incidence of TdP associated with increased nonphosphorylated connexin 43 (Cx43). AAP10 prevented augmentation of TDR and induction of TdP while rescuing Cx43 phosphorylation. There was no significant change in the quantity and spatial distribution of Cx43. These data indicate that gap junction enhancer AAP10 can prevent augmentation of TDR and suppress TdP by preventing dephosphorylation of Cx43 in a LQT2 model.

  11. Early disruption of glial communication via connexin gap junction in multiple sclerosis, Baló's disease and neuromyelitis optica.

    Science.gov (United States)

    Masaki, Katsuhisa

    2015-10-01

    Multiple sclerosis (MS), neuromyelitis optica (NMO), and Baló's disease (BD) are inflammatory demyelinating diseases of the CNS. We previously reported anti-aquaporin-4 (anti-AQP4) antibody-dependent AQP4 loss occurs in some NMO patients, while antibody-independent AQP4 astrocytopathy can occur in heterogeneous demyelinating conditions, including MS, NMO and BD. To investigate the relationship between astrocytopathy and demyelination, we focused on connexins (Cxs), which form gap junctions (GJs) between astrocytes and oligodendrocytes and maintain homeostasis in the CNS. We evaluated expression of astrocytic Cx43/Cx30 and oligodendrocytic Cx47/Cx32 in autopsied materials from MS, NMO and BD patients. Astrocytic Cx43 and oligodendrocytic Cx32/Cx47 expressions were significantly diminished in both demyelinated and preserved myelin layers in all BD samples. In the leading edge of BD lesions, Cx43 and AQP4 loss preceded Cx32/Cx47 loss. Half of the NMO and MS samples showed preferential loss of astrocytic Cx43 expression in actively demyelinating and chronic active lesions, where heterotypic Cx43/Cx47 astrocyte-oligodendrocyte GJs were lost. Cases with Cx43 loss were significantly associated with rapid disease progression, regardless of the disease phenotype. Pathologically, Cx43 loss was frequently accompanied by distal oligodendrogliopathy. Our findings suggest that Cx43 astrocytopathy can occur in MS, BD and NMO. Moreover, astrocytic Cx43 loss may be associated with disease aggressiveness and distal oligodendrogliopathy in demyelinating conditions. Early disruption of glial communications via GJs may cause loss of glia syncytium, thereby inducing oligodendroglial damage and myelin loss. Inhibition of Cx hemichannels and restoration of GJs may be a possible therapeutic target for demyelinating disorders. © 2015 Japanese Society of Neuropathology.

  12. Human cytomegalovirus immediate early proteins promote degradation of connexin 43 and disrupt gap junction communication: implications for a role in gliomagenesis.

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    Khan, Zahidul; Yaiw, Koon-Chu; Wilhelmi, Vanessa; Lam, Hoyin; Rahbar, Afsar; Stragliotto, Giuseppe; Söderberg-Nauclér, Cecilia

    2014-01-01

    A lack of gap junctional intercellular communication (GJIC) is common in cancer. Many oncogenic viruses have been shown to downregulate the junctional protein connexin 43 (Cx43) and reduce GJIC. Human cytomegalovirus (HCMV) is a ubiquitous, species-specific betaherpesvirus that establishes life-long latency after primary infection. It encodes two viral gene products, immediate early (IE) proteins IE1 and IE2, which are crucial in viral replication and pathogenesis of many diseases. Emerging evidence demonstrates that HCMV DNA and proteins are highly prevalent in glioblastoma multiforme (GBM) and in other tumors, but HCMV's role in tumorigenesis remains obscure. In the present study, we examined the effects of HCMV infection on Cx43 expression and GJIC as well as the viral mechanism mediating the effects in human GBM cells and tissue samples. We found that HCMV downregulated Cx43 protein, resulting in disruption of functional GJIC as assayed by fluorescent dye transfer assay. We show that both HCMV-IE72 and IE86 mediate downregulation of Cx43 by silencing RNA targeting either IE72 or IE86 coupled with ganciclovir. This finding was further validated by transfection with expression vectors encoding IE72 or IE86, and we show that viral-mediated Cx43 depletion involved proteasomal degradation. Importantly, we also observed that the Cx43 protein levels and IE staining correlated inversely in 10 human GBM tissue specimens. Thus, HCMV regulates Cx43 expression and GJIC, which may contribute to gliomagenesis.

  13. Targeting Cx43 and N-cadherin, which are abnormally upregulated in venous leg ulcers, influences migration, adhesion and activation of Rho GTPases.

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    Ariadna Mendoza-Naranjo

    Full Text Available BACKGROUND: Venous leg ulcers can be very hard to heal and represent a significant medical need with no effective therapeutic treatment currently available. PRINCIPAL FINDINGS: In wound edge biopsies from human venous leg ulcers we found a striking upregulation of dermal N-cadherin, Zonula Occludens-1 and the gap junction protein Connexin43 (Cx43 compared to intact skin, and in stark contrast to the down-regulation of Cx43 expression seen in acute, healing wounds. We targeted the expression of these proteins in 3T3 fibroblasts to evaluate their role in venous leg ulcers healing. Knockdown of Cx43 and N-cadherin, but not Zonula Occludens-1, accelerated cell migration in a scratch wound-healing assay. Reducing Cx43 increased Golgi reorientation, whilst decreasing cell adhesion and proliferation. Furthermore, Connexin43 and N-cadherin knockdown led to profound effects on fibroblast cytoskeletal dynamics after scratch-wounding. The cells exhibited longer lamelipodial protrusions lacking the F-actin belt seen at the leading edge in wounded control cells. This phenotype was accompanied by augmented activation of Rac-1 and RhoA GTPases, as revealed by Förster Resonance Energy Transfer and pull down experiments. CONCLUSIONS: Cx43 and N-cadherin are potential therapeutic targets in the promotion of healing of venous leg ulcers, by acting at least in part through distinct contributions of cell adhesion, migration, proliferation and cytoskeletal dynamics.

  14. Presence of Cx43 in extracellular vesicles reduces the cardiotoxicity of the anti-tumour therapeutic approach with doxorubicin

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    Tania Martins-Marques

    2016-09-01

    Full Text Available Extracellular vesicles (EVs are major conveyors of biological information, mediating local and systemic cell-to-cell communication under physiological and pathological conditions. These endogenous vesicles have been recognized as prominent drug delivery vehicles of several therapeutic cargoes, including doxorubicin (dox, presenting major advantages over the classical approaches. Although dox is one of the most effective anti-tumour agents in the clinical practice, its use is very often hindered by its consequent dramatic cardiotoxicity. Despite significant advances witnessed in the past few years, more comprehensive studies, supporting the therapeutic efficacy of EVs, with decreased side effects, are still scarce. The main objective of this study was to evaluate the role of the gap junction protein connexin43 (Cx43 in mediating the release of EV content into tumour cells. Moreover, we investigated whether Cx43 improves the efficiency of dox-based anti-tumour treatment, with a concomitant decrease of cardiotoxicity. In the present report, we demonstrate that the presence of Cx43 in EVs increases the release of luciferin from EVs into tumour cells in vitro and in vivo. In addition, using cell-based approaches and a subcutaneous mouse tumour model, we show that the anti-tumour effect of dox incorporated into EVs is similar to the administration of the free drug, regardless the presence of Cx43. Strikingly, we demonstrate that the presence of Cx43 in dox-loaded EVs reduces the cardiotoxicity of the drug. Altogether, these results bring new insights into the concrete potential of EVs as therapeutic vehicles and open new avenues toward the development of strategies that help to reduce unwanted side effects.

  15. Gap junction proteins in the light-damaged albino rat.

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    Guo, Cindy X; Tran, Henry; Green, Colin R; Danesh-Meyer, Helen V; Acosta, Monica L

    2014-01-01

    Changes in connexin expression are associated with many pathological conditions seen in animal models and in humans. We hypothesized that gap junctions are important mediators in tissue dysfunction and injury processes in the retina, and therefore, we investigated the pattern of connexin protein expression in the light-damaged albino rat eye. Adult Sprague-Dawley rats were exposed to intense light for 24 h. The animals were euthanized, and ocular tissue was harvested at 0 h, 6 h, 24 h, 48 h, and 7 days after light damage. The tissues were processed for immunohistochemistry and western blotting to analyze the expression of the gap junction proteins in the light-damaged condition compared to the non-light-damaged condition. Cell death was detected using the terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) technique. Intense light exposure caused increased TUNEL labeling of photoreceptor cells. Immunocytochemistry revealed that connexin 36 (Cx36) was significantly increased in the inner plexiform layer and Cx45 was significantly decreased in the light-damaged retina. The pattern of Cx36 and Cx45 labeling returned to normal 7 days after light damage. Cx43 significantly increased in the RPE and the choroid in the light-damaged tissue, and decreased but not significantly in the retina. This elevated Cx43 expression in the choroid colocalized with markers of nitration-related oxidative stress (nitrotyrosine) and inflammation (CD45 and ionized calcium-binding adaptor molecule-1) in the choroid. The results suggest that connexins are regulated differently in the retina than in the choroid in response to photoreceptor damage. Changes in connexins, including Cx36, Cx43, and Cx45, may contribute to the damage process. Specifically, Cx43 was associated with inflammatory damage. Therefore, connexins may be candidate targets for treatment for ameliorating disease progression.

  16. 17β estradiol regulation of connexin 43-based gap junction and mechanosensitivity through classical estrogen receptor pathway in osteocyte-like MLO-Y4 cells.

    KAUST Repository

    Ren, Jian

    2013-04-01

    Connexin 43 (Cx43) plays an essential role in osteocyte mechanotransduction. Although estrogen involves in the adaptive responses of bone cells to mechanical loadings, its effects on osteocytic Cx43-based gap junction intercellular communication (GJIC) remain obscure. We found that 17β estradiol (E2) up-regulated Cx43, and enhanced GJIC in osteocyte-like MLO-Y4 cells in fluorescence recovery after photobleaching (FRAP) assay. Combination of E2 pre-treatment and oscillating fluid flow (OFF) further enhanced Cx43 expression and mitogen-activated protein kinase (MAPK) phosphorylation, comparing to E2 or OFF treatment alone. Both blocking of classical estrogen receptors (ERα/β) by fulvestrant and ERα knockdown by small interfering RNA inhibited E2-mediated Cx43 increase, while a GPR30-specific agonist G-1 failed to promote Cx43 expression. Our results suggest that the presence of E2 enhanced Cx43-based GJIC mainly via ERα/β pathway, and sensitized osteocytes to mechanical loading. © 2012 Elsevier Inc. All rights reserved.

  17. A Cell-Based High-Throughput Assay for Gap Junction Communication Suitable for Assessing Connexin 43-Ezrin Interaction Disruptors Using IncuCyte ZOOM.

    Science.gov (United States)

    Dukic, Aleksandra R; McClymont, David W; Taskén, Kjetil

    2017-01-01

    Connexin 43 (Cx43), the predominant gap junction (GJ) protein, directly interacts with the A-kinase-anchoring protein (AKAP) Ezrin in human cytotrophoblasts and a rat liver epithelial cells (IAR20). The Cx43-Ezrin-protein kinase (PKA) complex facilitates Cx43 phosphorylation by PKA, which triggers GJ opening in cytotrophoblasts and IAR20 cells and may be a general mechanism regulating GJ intercellular communication (GJIC). Considering the importance of Cx43 GJs in health and disease, they are considered potential pharmaceutical targets. The Cx43-Ezrin interaction is a protein-protein interaction that opens possibilities for targeting with peptides and small molecules. For this reason, we developed a high-throughput cell-based assay in which GJIC can be assessed and new compounds characterized. We used two pools of IAR20 cells, calcein loaded and unloaded, that were mixed and allowed to attach. Next, GJIC was monitored over time using automated imaging via the IncuCyte imager. The assay was validated using known GJ inhibitors and anchoring peptide disruptors, and we further tested new peptides that interfered with the Cx43-Ezrin binding region and reduced GJIC. Although an AlphaScreen assay can be used to screen for Cx43-Ezrin interaction inhibitors, the cell-based assay described is an ideal secondary screen for promising small-molecule hits to help identify the most potent compounds.

  18. Cx43 Mediates Resistance against MPP+-Induced Apoptosis in SH-SY5Y Neuroblastoma Cells via Modulating the Mitochondrial Apoptosis Pathway

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    In-Su Kim

    2016-11-01

    Full Text Available Neuronal apoptosis in the substantia nigra par compacta (SNpc appears to play an essential role in the pathogenesis of Parkinson’s disease. However, the mechanisms responsible for the death of dopaminergic neurons are not fully understood yet. To explore the apoptotic mechanisms, we used a well-known parkinsonian toxin, 1-methyl-4-phenylpyridine (MPP+, to induce neuronal apoptosis in the human dopaminergic SH-SY5Y cell line. The most common method of interaction between cells is gap junctional intercellular communication (GJIC mediated by gap junctions (GJs formed by transmembrane proteins called connexins (Cx. Modulation of GJIC affects cell viability or growth, implying that GJIC may have an important role in maintaining homeostasis in various organs. Here, we hypothesized that increasing the level of the gap junction protein Cx43 in SH-SY5Y neuroblastoma cells could provide neuroprotection. First, our experiments demonstrated that knocking down Cx43 protein by using Cx43-specific shRNA in SH-SY5Y neuroblastoma cells potentiated MPP+-induced neuronal apoptosis evident from decreased cell viability. In another experiment, we demonstrated that over-expression of Cx43 in the SH-SY5Y cell system decreased MPP+-induced apoptosis based on the MTT assay and reduced the Bax/Bcl-2 ratio and the release of cytochrome C based on Western blot analysis. Taken together, our results suggest that Cx43 could mediate resistance against MPP+-induced apoptosis in SH-SY5Y neuroblastoma cells via modulating the mitochondrial apoptosis pathway.

  19. Altered Expression of Connexin-43 and Impaired Capacity of Gap Junctional Intercellular Communication in Prostate Cancer Cells

    Institute of Scientific and Technical Information of China (English)

    XING Yifei; XIAO Yajun; ZENG FuQing; ZHAO Jun; XIAO Chuanguo; XIONG Ping; FENG Wei

    2007-01-01

    Connexin-43 (Cx43) expression in prostate cancer (PCa) cells and the potency of gap junctional intercellular communication (GJIC) in the cells were investigated, with an attempt to elucidate the reason why the so-called "bystander effect" mediated by thymidine kinase (TK) suicide gene therapy on PCa cells is not of significance and to explore the role of GJIC in PCa carcinogenesis.mRNA and protein expression of Cx43 in a PCa cell line PC-3m was detected by reverse-transcription polymerase chain reaction (RT-PCR) and strapt-avidin-biotin-enzyme complex (SABC) immunohistochemical staining, and inherent GJIC of PC-3m cells was assayed by scrape-loading and dye transfer (SLDT) assay. The expression of Cx43 in human normal and malignant prostate tissues was determined by SABC immunohistochemistry as well. It was found that Cx43 mRNA and protein expression in PC-3m cells was slightly reduced as compared with positive controls and the location of Cx43 protein was aberrant in cytoplasm rather than on membrane. Assessment of paraffin sections demonstrated that the expression of Cx43 protein in PCa cells was abnormally located and markedly diminished as compared with normal prostatic epithelial ones, displaying a negative correlation to the pathological grade (χ2=4.025, P<0.05). Additionally, capacity of inherent GJIC in PC-3m cells was disrupted, which was semi-quantified as (+) or (-). It was indicated that both down-regulated expression of Cx43 mRNA and aberrant location of Cx43 protein participated in the mechanisms leading to deficient GJIC in PC-3m cells. Lack of efficient GJIC is a molecular event, which may contribute not only to limited extent of "bystander effect", but also to initiation and progression of prostatic neoplasm.

  20. Gap junction remodeling and cardiac arrhythmogenesis in a murine model of oculodentodigital dysplasia.

    Science.gov (United States)

    Kalcheva, Nellie; Qu, Jiaxiang; Sandeep, Nefthi; Garcia, Luis; Zhang, Jie; Wang, Zhiyong; Lampe, Paul D; Suadicani, Sylvia O; Spray, David C; Fishman, Glenn I

    2007-12-18

    Gap junction channels are required for normal cardiac impulse propagation, and gap junction remodeling is associated with enhanced arrhythmic risk. Oculodentodigital dysplasia (ODDD) is a multisystem syndrome due to mutations in the connexin43 (Cx43) gap junction channel gene. To determine the effects of a human connexin channelopathy on cardiac electrophysiology and arrhythmogenesis, we generated a murine model of ODDD by introducing the disease-causing I130T mutant allele into the mouse genome. Cx43 abundance was markedly reduced in mutant hearts with preferential loss of phosphorylated forms that interfered with trafficking and assembly of gap junctions in the junctional membrane. Dual whole-cell patch-clamp studies showed significantly lower junctional conductance between neonatal cell pairs from mutant hearts, and optical mapping of isolated-perfused hearts with voltage-sensitive dyes demonstrated significant slowing of conduction velocity. Programmed electrical stimulation revealed a markedly increased susceptibility to spontaneous and inducible ventricular tachyarrhythmias. In summary, our data demonstrate that the I130T mutation interferes with Cx43 posttranslational processing, resulting in diminished cell-cell coupling, slowing of impulse propagation, and a proarrhythmic substrate.

  1. Activation of L-type calcium channels is required for gap junction-mediated intercellular calcium signaling in osteoblastic cells

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    Jorgensen, Niklas Rye; Teilmann, Stefan Cuoni; Henriksen, Zanne; Civitelli, Roberto; Sorensen, Ole Helmer; Steinberg, Thomas H.

    2003-01-01

    The propagation of mechanically induced intercellular calcium waves (ICW) among osteoblastic cells occurs both by activation of P2Y (purinergic) receptors by extracellular nucleotides, resulting in "fast" ICW, and by gap junctional communication in cells that express connexin43 (Cx43), resulting in "slow" ICW. Human osteoblastic cells transmit intercellular calcium signals by both of these mechanisms. In the current studies we have examined the mechanism of slow gap junction-dependent ICW in osteoblastic cells. In ROS rat osteoblastic cells, gap junction-dependent ICW were inhibited by removal of extracellular calcium, plasma membrane depolarization by high extracellular potassium, and the L-type voltage-operated calcium channel inhibitor, nifedipine. In contrast, all these treatments enhanced the spread of P2 receptor-mediated ICW in UMR rat osteoblastic cells. Using UMR cells transfected to express Cx43 (UMR/Cx43) we confirmed that nifedipine sensitivity of ICW required Cx43 expression. In human osteoblastic cells, gap junction-dependent ICW also required activation of L-type calcium channels and influx of extracellular calcium.

  2. Gap junction intercellular communication: a review of a potential platform to modulate craniofacial tissue engineering.

    Science.gov (United States)

    Rossello, Ricardo A; Kohn, David H

    2009-02-01

    Defects in craniofacial tissues, resulting from trauma, congenital abnormalities, oncologic resection or progressive deforming diseases, may result in aesthetic deformity, pain and reduced function. Restoring the structure, function and aesthetics of craniofacial tissues represents a substantial clinical problem in need of new solutions. More biologically-interactive biomaterials could potentially improve the treatment of craniofacial defects, and an understanding of developmental processes may help identify strategies and materials that can be used in tissue engineering. One such strategy that can potentially advance tissue engineering is cell-cell communication. Gap junction intercellular communication is the most direct way of achieving such signaling. Gap junction communication through connexin-mediated junctions, in particular connexin 43 (Cx43), plays a major role bone development. Given the important role of Cx43 in controlling development and differentiation, especially in bone cells, controlling the expression of Cx43 may provide control over cell-to-cell communication and may help overcome some of the challenges in craniofacial tissue engineering. Following a review of gap junctions in bone cells, the ability to enhance cell-cell communication and osteogenic differentiation via control of gap junctions is discussed, as is the potential utility of this approach in craniofacial tissue engineering. Copyright 2008 Wiley Periodicals, Inc.

  3. Human Amniotic Fluid Cells Form Functional Gap Junctions with Cortical Cells

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    Anna Jezierski

    2012-01-01

    Full Text Available The usage of stem cells is a promising strategy for the repair of damaged tissue in the injured brain. Recently, amniotic fluid (AF cells have received a lot of attention as an alternative source of stem cells for cell-based therapies. However, the success of this approach relies significantly on proper interactions between graft and host tissue. In particular, the reestablishment of functional brain networks requires formation of gap junctions, as a key step to provide sufficient intercellular communication. In this study, we show that AF cells express high levels of CX43 (GJA1 and are able to establish functional gap junctions with cortical cultures. Furthermore, we report an induction of Cx43 expression in astrocytes following injury to the mouse motor cortex and demonstrate for the first time CX43 expression at the interface between implanted AF cells and host brain cells. These findings suggest that CX43-mediated intercellular communication between AF cells and cortical astrocytes may contribute to the reconstruction of damaged tissue by mediating modulatory, homeostatic, and protective factors in the injured brain and hence warrants further investigation.

  4. Oligodendrocyte gap junction loss and disconnection from reactive astrocytes in multiple sclerosis gray matter.

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    Markoullis, Kyriaki; Sargiannidou, Irene; Schiza, Natasa; Roncaroli, Federico; Reynolds, Richard; Kleopa, Kleopas A

    2014-09-01

    Gap junctions are essential for glial cell function and have been increasingly implicated in multiple sclerosis (MS). Because increasing cortical abnormalities correlate with disease progression and cognitive dysfunction, we examined the expression of oligodendrocytic connexin32 (Cx32) and Cx47 and their astrocytic partners Cx30 and Cx43 in cortical lesions and normal-appearing gray matter (NAGM) in MS patients. Postmortem brain tissue samples from 9 MS cases were compared with 10 controls using real-time polymerase chain reaction, immunoblot, and immunohistochemical analyses. Connexin32 and Cx47 gap junction formation in oligodendrocytes was reduced within lesions, whereas Cx32 loss also extended to NAGM. In contrast, astrocytic Cx30 expression was increased within cortical lesions, whereas Cx43 was elevated in both lesions and NAGM. Diffuse microglial activation and marked astrogliotic changes accompanied these connexin abnormalities. Increased expression of Cx43 correlated with inflammatory load (r = 0.828, p = 0.042), whereas Cx32 expression correlated with longer disease duration and, therefore, milder course (r = 0.825, p = 0.043). Thus, there is a loss of intramyelin and intercellular oligodendrocyte gap junctions in MS gray matter lesions and NAGM, whereas interastrocytic gap junctions are increased, reflecting astrogliosis. These changes correlate with inflammation and disease duration and suggest that disconnection of oligodendrocytes from reactive astrocytes may play a role in failed remyelination and disease progression.

  5. Gap junction signalling mediated through connexin-43 is required for chick limb development.

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    Makarenkova, H; Patel, K

    1999-03-15

    During chick limb development the gap junction protein Connexin-43 (Cx43) is expressed in discrete spatially restricted domains in the apical ectodermal ridge (AER) and mesenchyme of the zone of polarising activity. Antisense oligonucleotides (ODNs) were used to investigate the role of Connexin-43 (Cx43) in the development of the chick limb bud. We have used unmodified ODNs in Pluronic F-127 gel, which is liquid at low temperature but sets at room temperature and so remains situated at the point of application. As a mild surfactant, the gel increases antisense ODN penetration and supplies ODNs to the embryo continually for 12-18 h. We have shown a strong decrease in Cx43 protein expression after application of specific antisense oligonucleotides but the abundance of a closely related protein, Connexin-32 (Cx32), was not affected. Application of antisense Cx43 ODNs at stages 8-15 HH before limb outgrowth resulted in dramatic limb phenotypes. About 40% of treated embryos exhibited defects such as truncation of the limb bud, fragmentation into two or more domains, or complete splitting of the limb bud into two or three branches. Molecular analysis of antisense treated embryos failed to detect Shh or Bmp-2 in anterior structures and suggested that extra lobes seen in nicked and split limbs were not a result of establishment of new signalling centres as found after the application of FGF to the flank. However, examination of markers for the AER showed a number of abnormalities. In severely truncated specimens we were unable to detect the expression of either Fgf-4 or Fgf-8. In both nicked and split limbs the expression of these genes was discontinuous. Down-regulation of Cx43 after the antisense application could be comparable to AER removal and results in distal truncation of the limb bud. Taken together these data suggest the existence of a feedback loop between the FGFs and signalling mediated by Cx43.

  6. Mono-Heteromeric Configurations of Gap Junction Channels Formed by Connexin43 and Connexin45 Reduce Unitary Conductance and Determine both Voltage Gating and Metabolic Flux Asymmetry

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    Guoqiang Zhong

    2017-05-01

    Full Text Available In cardiac tissues, the expression of multiple connexins (Cx40, Cx43, Cx45, and Cx30.2 is a requirement for proper development and function. Gap junctions formed by these connexins have distinct permeability and gating mechanisms. Since a single cell can express more than one connexin isoform, the formation of hetero-multimeric gap junction channels provides a tissue with an enormous repertoire of combinations to modulate intercellular communication. To study further the perm-selectivity and gating properties of channels containing Cx43 and Cx45, we studied two monoheteromeric combinations in which a HeLa cell co-transfected with Cx43 and Cx45 was paired with a cell expressing only one of these connexins. Macroscopic measurements of total conductance between cell pairs indicated a drastic reduction in total conductance for mono-heteromeric channels. In terms of Vj dependent gating, Cx43 homomeric connexons facing heteromeric connexons only responded weakly to voltage negativity. Cx45 homomeric connexons exhibited no change in Vj gating when facing heteromeric connexons. The distributions of unitary conductances (γj for both mono-heteromeric channels were smaller than predicted, and both showed low permeability to the fluorescent dyes Lucifer yellow and Rhodamine123. For both mono-heteromeric channels, we observed flux asymmetry regardless of dye charge: flux was higher in the direction of the heteromeric connexon for MhetCx45 and in the direction of the homomeric Cx43 connexon for MhetCx43. Thus, our data suggest that co-expression of Cx45 and Cx43 induces the formation of heteromeric connexons with greatly reduced permeability and unitary conductance. Furthermore, it increases the asymmetry for voltage gating for opposing connexons, and it favors asymmetric flux of molecules across the junction that depends primarily on the size (not the charge of the crossing molecules.

  7. Effects of all-trans-retinoic acid on the expression and tyrosine phosphorylation of gap junction connexin 43 in HeLa cell line and its significance

    Institute of Scientific and Technical Information of China (English)

    CHEN Bi-liang; MA Xiang-dong; XIN Xiao-yan; WANG De-tang; WANG Chun-mei

    2001-01-01

    Objective: To investigate the signal transduction mechanism of gap junctional genes connexin43 in human cervical carcinogenesis. Methods: Human cervical carcinoma cell line HeLa was cultured and treated by all-trans-retinoic acid (ATRA). Flow cytometer (FCM) was employed to detect expression of Cx43 protein in HeLa cells. Fluo-3 AM loading and laser scanning confocal microscope (LSCM) were used to measure the concentrations of intracellular calcium ([Ca2+]i) in HeLa cells. Phosphorylation on tyrosine of connexin43 protein was examined by immunoblot. Results: The positive rate of Cx43 protein increased from 1.9% in untreated HeLa cells to 26.3% in RA-treated HeLa cells as shown by FCM. [Ca2+]i was 35.73 nmol/L in untreated HeLa cells which was increased to 58.16 nmol/L in ATRA-treated cells.Immunoblot showed that ATRA-treated HeLa cells had phosphorylation on tyrosine in Cx43 protein whereas untreated cells had not. Conclusions: Carcinogenesis of human cervical carcinoma is related with the abnormal expression of cx43gene and disorder of signal transduction manifested as the decrease of [Ca2+]i and post-translation phosphorylation on tyrosine of Cx43 protein. The anti-tumor effect of ATRA in HeLa cells might be due to the up-regulation of cx43 gene and its signal transduction pathway.

  8. Gap Junctions Contribute to Ictal/Interictal Genesis in Human Hypothalamic Hamartomas.

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    Wu, Jie; Gao, Ming; Rice, Stephen G; Tsang, Candy; Beggs, John; Turner, Dharshaun; Li, Guohui; Yang, Bo; Xia, Kunkun; Gao, Fenfei; Qiu, Shenfeng; Liu, Qiang; Kerrigan, John F

    2016-06-01

    Human hypothalamic hamartoma (HH) is a rare subcortical lesion associated with treatment-resistant epilepsy. Cellular mechanisms responsible for epileptogenesis are unknown. We hypothesized that neuronal gap junctions contribute to epileptogenesis through synchronous activity within the neuron networks in HH tissue. We studied surgically resected HH tissue with Western-blot analysis, immunohistochemistry, electron microscopy, biocytin microinjection of recorded HH neurons, and microelectrode patch clamp recordings with and without pharmacological blockade of gap junctions. Normal human hypothalamus tissue was used as a control. Western blots showed increased expression of both connexin-36 (Cx36) and connexin-43 (Cx43) in HH tissue compared with normal human mammillary body tissue. Immunohistochemistry demonstrated that Cx36 and Cx43 are expressed in HH tissue, but Cx36 was mainly expressed within neuron clusters while Cx43 was mainly expressed outside of neuron clusters. Gap-junction profiles were observed between small HH neurons with electron microscopy. Biocytin injection into single recorded small HH neurons showed labeling of adjacent neurons, which was not observed in the presence of a neuronal gap-junction blocker, mefloquine. Microelectrode field recordings from freshly resected HH slices demonstrated spontaneous ictal/interictal-like discharges in most slices. Bath-application of gap-junction blockers significantly reduced ictal/interictal-like discharges in a concentration-dependent manner, while not affecting the action-potential firing of small gamma-aminobutyric acid (GABA) neurons observed with whole-cell patch-clamp recordings from the same patient's HH tissue. These results suggest that neuronal gap junctions between small GABAergic HH neurons participate in the genesis of epileptic-like discharges. Blockade of gap junctions may be a new therapeutic strategy for controlling seizure activity in HH patients.

  9. Gap Junctions Contribute to Ictal/Interictal Genesis in Human Hypothalamic Hamartomas

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    Jie Wu

    2016-06-01

    Full Text Available Human hypothalamic hamartoma (HH is a rare subcortical lesion associated with treatment-resistant epilepsy. Cellular mechanisms responsible for epileptogenesis are unknown. We hypothesized that neuronal gap junctions contribute to epileptogenesis through synchronous activity within the neuron networks in HH tissue. We studied surgically resected HH tissue with Western-blot analysis, immunohistochemistry, electron microscopy, biocytin microinjection of recorded HH neurons, and microelectrode patch clamp recordings with and without pharmacological blockade of gap junctions. Normal human hypothalamus tissue was used as a control. Western blots showed increased expression of both connexin-36 (Cx36 and connexin-43 (Cx43 in HH tissue compared with normal human mammillary body tissue. Immunohistochemistry demonstrated that Cx36 and Cx43 are expressed in HH tissue, but Cx36 was mainly expressed within neuron clusters while Cx43 was mainly expressed outside of neuron clusters. Gap-junction profiles were observed between small HH neurons with electron microscopy. Biocytin injection into single recorded small HH neurons showed labeling of adjacent neurons, which was not observed in the presence of a neuronal gap-junction blocker, mefloquine. Microelectrode field recordings from freshly resected HH slices demonstrated spontaneous ictal/interictal-like discharges in most slices. Bath-application of gap-junction blockers significantly reduced ictal/interictal-like discharges in a concentration-dependent manner, while not affecting the action-potential firing of small gamma-aminobutyric acid (GABA neurons observed with whole-cell patch-clamp recordings from the same patient's HH tissue. These results suggest that neuronal gap junctions between small GABAergic HH neurons participate in the genesis of epileptic-like discharges. Blockade of gap junctions may be a new therapeutic strategy for controlling seizure activity in HH patients.

  10. Classical swine fever virus down-regulates endothelial connexin 43 gap junctions.

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    Hsiao, Hsiang-Jung; Liu, Pei-An; Yeh, Hung-I; Wang, Chi-Young

    2010-07-01

    Classical swine fever is a contagious disease of pigs characterized by fatal hemorrhagic fever. Classical swine fever virus (CSFV) induces the expression of pro-inflammatory and pro-coagulant factors of vascular endothelial cells and establishes a long-term infection. This study aimed to understand the effect of CSFV on endothelial connexin 43 (Cx43) expression and gap junctional intercellular coupling (GJIC). Porcine aortic endothelial cells were infected with CSFV at different multiplicity of infection for 48 h. Semi-quantitative RT-PCR, immunoconfocal microscopy, and Western blotting showed that the transcription and translation of Cx43 were reduced, and this was associated with an attenuation of GJIC. This decrease occurred in a time-dependent manner. An ERK inhibitor (PD98059), a JNK inhibitor (SP600125), and proteasome/lysosome inhibitors all significantly reversed the reduction in Cx43 protein levels without any influence on the titer of progeny virus. In addition, CSFV activated ERK and JNK in a time-dependent manner and down-regulated Cx43 promoter activity, mainly through decreased AP2 binding. This effect was primarily caused by the replication of CSFV rather than a consequence of cytokines being induced by CSFV infection of endothelial cells.

  11. Expression of gap junction proteins connexins 26, 30, and 43 in Dupuytren's disease.

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    Holzer, Lukas A; Cör, Andrej; Holzer, Gerold

    2014-02-01

    Dupuytren's disease (DD) is a benign fibroproliferative process of the palmar aponeurosis showing similarities to wound healing. Communication of cells involved in wound healing is mediated by the composition of gap junction (GJ) proteins. We investigated the expression of 3 GJ proteins, connexins 26, 30, and 43 (Cx26, Cx30, and Cx43) in DD. Fragments of Dupuytren's tissue from 31 patients (mean age 56 (30-76) years, 24 male) were analyzed immunohistochemically and compared to control tissue for expression of the GJ proteins Cx26, Cx30, and Cx43 and also alfa-smooth muscle actin (α-SMA). 14 of 31 samples could be attributed to the involutional phase (α-SMA positive) whereas 17 samples had to be considered cords in the residual phase (α-SMA negative). Expression of Cx26 and Cx43 was seen in 12 of the 14 samples from the involutional phase, and Cx30 was seen in 7 of these. Only 4 of the 17 samples from the residual phase showed any Cx, and there was none in the controls. The high expression of GJ proteins Cx26, Cx30, and Cx43 in α-SMA positive myofibroblast-rich nodules, which are characteristic of the active involutional phase of DD, suggests that connexins could be a novel treatment target for the treatment of DD.

  12. Role of gap junction and connexin-43 in hypoxic-ischemic brain damage

    Institute of Scientific and Technical Information of China (English)

    Jieying Lin; Niyang Lin

    2006-01-01

    OBJECTEVE:Gap junctin (GJ)is the structural basis for direct intercellular communication of nerve cells . Connexin(Cx) is the protein subunit for constructling GJ channel. Among them, Cx43is closely related with nervous system. Both Cx43 and nervous system play an important role in the pathophysiological development of hypoxic-ischemic injury. We are in attempt to investigate GJ,Cx43 and their correlations with hypoxic-ischemic brain damage by research.DATA SOURCES:Using the terms "brain gap junction"in English and "gap junction"in Chinese, we searched the Medline database and Chinese BioMedical Literature Database as well as China Hospital Knowledge Database to identify the articles published from 1996 to 2006 about GJ and brain hypoxic-ischemic injury.STUDY SELECTION:The articles were selected firstly and abstracts of 250 articles were read thuugh.Articles in which the experimental design met randomized controlled principle were included,and study articles and case reports with repetitve contents were excluded.DATA EXTRACTION:Among 53 included correlative articles, 23 were excluded for repetitive contents and the other 30 were analyzed.DATA SYNTHESIS:GJ,widely esistling in nervous system,plays a key role in maintainling normal differentiation and development as well as physiological function brain tissue.GJ channel is a hydrophilic,low-selectivity and lowohmic channel, which can provide direct channel for intercellular substance transmission and information communication. It plays an important role in the differentiation and development of nerve cells and regulation of physiological function,The funtions of GJ channel are regulated by many factors,which invilved intracellular Ph value, Ca2+concentration, ATP concentration, phosphorylation of Cx, transchannel pressure,some neurohormonal factors,regulatory factors of protein and so on. Cx43 is the main component of GJ channel in the brain tissues. Its expression in the brain tissue of mammal is the strongest

  13. Rescue of perfluorooctanesulfonate (PFOS)-mediated Sertoli cell injury by overexpression of gap junction protein connexin 43

    Science.gov (United States)

    Li, Nan; Mruk, Dolores D.; Chen, Haiqi; Wong, Chris K. C.; Lee, Will M.; Cheng, C. Yan

    2016-07-01

    Perfluorooctanesulfonate (PFOS) is an environmental toxicant used in developing countries, including China, as a stain repellent for clothing, carpets and draperies, but it has been banned in the U.S. and Canada since the late 2000s. PFOS perturbed the Sertoli cell tight junction (TJ)-permeability barrier, causing disruption of actin microfilaments in cell cytosol, perturbing the localization of cell junction proteins (e.g., occluden-ZO-1, N-cadherin-ß-catenin). These changes destabilized Sertoli cell blood-testis barrier (BTB) integrity. These findings suggest that human exposure to PFOS might induce BTB dysfunction and infertility. Interestingly, PFOS-induced Sertoli cell injury associated with a down-regulation of the gap junction (GJ) protein connexin43 (Cx43). We next investigated if overexpression of Cx43 in Sertoli cells could rescue the PFOS-induced cell injury. Indeed, overexpression of Cx43 in Sertoli cells with an established TJ-barrier blocked the disruption in PFOS-induced GJ-intercellular communication, resulting in the re-organization of actin microfilaments, which rendered them similar to those in control cells. Furthermore, cell adhesion proteins that utilized F-actin for attachment became properly distributed at the cell-cell interface, resealing the disrupted TJ-barrier. In summary, Cx43 is a good target that might be used to manage PFOS-induced reproductive dysfunction.

  14. Contribution of intracellular calcium and pH in ischemic uncoupling of cardiac gap junction channels formed of connexins 43, 40, and 45: a critical function of C-terminal domain.

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    Giriraj Sahu

    Full Text Available Ischemia is known to inhibit gap junction (GJ mediated intercellular communication. However the detail mechanisms of this inhibition are largely unknown. In the present study, we determined the vulnerability of different cardiac GJ channels formed of connexins (Cxs 43, 40, and 45 to simulated ischemia, by creating oxygen glucose deprived (OGD condition. 5 minutes of OGD decreased the junctional conductance (Gj of Cx43, Cx40 and Cx45 by 53±3%, 64±1% and 85±2% respectively. Reduction of Gj was prevented completely by restricting the change of both intracellular calcium ([Ca(2+]i and pH (pHi with potassium phosphate buffer. Clamping of either [Ca(2+]i or pHi, through BAPTA (2 mM or HEPES (80 mM respectively, offered partial resistance to ischemic uncoupling. Anti-calmodulin antibody attenuated the uncoupling of Cx43 and Cx45 significantly but not of Cx40. Furthermore, OGD could reduce only 26±2% of Gj in C-terminus (CT truncated Cx43 (Cx43-Δ257. Tethering CT of Cx43 to the CT-truncated Cx40 (Cx40-Δ249, and Cx45 (Cx45-Δ272 helped to resist OGD mediated uncoupling. Moreover, CT domain played a significant role in determining the junction current density and plaque diameter. Our results suggest; OGD mediated uncoupling of GJ channels is primarily due to elevated [Ca(2+]i and acidic pHi, though the latter contributes more. Among Cx43, Cx40 and Cx45, Cx43 is the most resistant to OGD while Cx45 is the most sensitive one. CT of Cx43 has major necessary elements for OGD induced uncoupling and it can complement CT of Cx40 and Cx45.

  15. Role of Myoendothelial Gap Junctions in the Regulation of Human Coronary Artery Smooth Muscle Cell Differentiation by Laminar Shear Stress

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    Zongqi Zhang

    2016-07-01

    Full Text Available Background/Aims: Smooth muscle cells may dedifferentiate into the synthetic phenotype and promote atherosclerosis. Here, we explored the role of myoendothelial gap junctions in phenotypic switching of human coronary artery smooth muscle cells (HCASMCs co-cultured with human coronary artery endothelial cells (HCAECs exposed to shear stress. Methods: HCASMCs and HCAECs were seeded on opposite sides of Transwell inserts, and HCAECs were exposed to laminar shear stress of 12 dyn/cm2 or 5 dyn/cm2. The myoendothelial gap junctions were evaluated by using a multi-photon microscope. Results: In co-culture with HCAECs, HCASMCs exhibited a contractile phenotype, and maintained the expression of differentiation markers MHC and H1-calponin. HCASMCs and HCAECs formed functional intercellular junctions, as evidenced by colocalization of connexin(Cx40 and Cx43 on cellular projections inside the Transwell membrane and biocytin transfer from HCAECs to HCASMCs. Cx40 siRNA and 18-α-GA attenuated protein expression of MHC and H1-calponin in HCASMCs. Shear stress of 5 dyn/cm2 increased Cx43 and decreased Cx40 expression in HCAECs, and partly inhibited biocytin transfer from HCAECs to HCASMCs, which could be completely blocked by Cx43 siRNA or restored by Cx40 DNA transfected into HCAECs. The exposure of HCAECs to shear stress of 5 dyn/cm2 promoted HCASMC phenotypic switching, manifested by morphological changes, decrease in MHC and H1-calponin expression, and increase in platelet-derived growth factor (PDGF-BB release, which was partly rescued by Cx43 siRNA or Cx40 DNA or PDGF receptor signaling inhibitor. Conclusions: The exposure of HCAECs to shear stress of 5 dyn/cm2 caused the dysfunction of Cx40/Cx43 heterotypic myoendothelial gap junctions, which may be replaced by homotypic Cx43/Cx43 channels, and induced HCASMC transition to the synthetic phenotype associated with the activation of PDGF receptor signaling, which may contribute to shear stress

  16. Gap junction hemichannels contribute to the generation of diarrhoea during infectious enteric disease.

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    Guttman, Julian Andrew; Lin, Ann En-Ju; Li, Yuling; Bechberger, John; Naus, Christian C; Vogl, A Wayne; Finlay, B Brett

    2010-02-01

    The attaching and effacing (A/E) pathogens enterohaemorrhagic Escherichia coli, enteropathogenic E coli and Citrobacter rodentium colonise intestinal tracts, attach to enterocytes, collapse infected cell microvilli and alter numerous host cell processes during infection. Enterocyte alterations result in numerous small molecules being released from host cells that likely contribute to diarrhoeal phenotypes observed during these infections. One possible route for small molecules to be released from intestinal cells may be through functional gap junction hemichannels. Here we examine the involvement of these hemichannels during the diarrhoeal disease caused by A/E pathogens in vivo. Mice were infected with the diarrhoea-causing murine A/E pathogen C rodentium for 7 days. Connexin43 (Cx43) protein levels and immunolocalization in the colon were initially used to determine alterations during A/E bacterial infections in vivo. Connexin mimetic peptides and connexin permeable tracer molecules were used to gage the presence and function of unpaired connexin hemichannels. The role of Cx43 in diarrhoea generation was assessed by comparing infections of wild-type mice to Cx43 mutant mice and determining the water abundance in the colonic luminal material. We demonstrate that Cx43 protein levels are increased in colonocytes during in vivo A/E bacterial infections, resulting in functionally open connexon hemichannels in apical membranes of infected cells. moreover, infected Cx43 +/- mice do not suffer from diarrhoeal disease. This study provides the first evidence that functional connexon hemichannels can occur in the intestine and are a novel molecular mechanism of water release during infectious diarrhoea.

  17. Inhibition of gap junction intercellular communication is involved in silica nanoparticles-induced H9c2 cardiomyocytes apoptosis via the mitochondrial pathway.

    Science.gov (United States)

    Du, Zhong-Jun; Cui, Guan-Qun; Zhang, Juan; Liu, Xiao-Mei; Zhang, Zhi-Hu; Jia, Qiang; Ng, Jack C; Peng, Cheng; Bo, Cun-Xiang; Shao, Hua

    2017-01-01

    Gap junction intercellular communication (GJIC) between cardiomyocytes is essential for synchronous heart contraction and relies on connexin-containing channels. Connexin 43 (Cx43) is a major component involved in GJIC in heart tissue, and its abnormal expression is closely associated with various cardiac diseases. Silica nanoparticles (SNPs) are known to induce cardiovascular toxicity. However, the mechanisms through which GJIC plays a role in cardiomyocytes apoptosis induced by SNPs remain unknown. The aim of the present study is to determine whether SNPs-decreased GJIC promotes apoptosis in rat cardiomyocytes cell line (H9c2 cells) via the mitochondrial pathway using CCK-8 Kit, scrape-loading dye transfer technique, Annexin V/PI double-staining assays, and Western blot analysis. The results showed that SNPs elicited cytotoxicity in H9c2 cells in a time- and concentration-dependent manner. SNPs also reduced GJIC in H9c2 cells in a concentration-dependent manner through downregulation of Cx43 and upregulation of P-Cx43. Inhibition of gap junctions by gap junction blocker carbenoxolone disodium resulted in decreased survival and increased apoptosis, whereas enhancement of the gap junctions by retinoic acid led to enhanced survival but decreased apoptosis. Furthermore, SNPs-induced apoptosis through the disrupted functional gap junction was correlated with abnormal expressions of the proteins involved in the mitochondrial pathway-related apoptosis such as Bcl-2/Bax, cytochrome C, Caspase-9, and Caspase-3. Taken together, our results provide the first evidence that SNPs-decreased GJIC promotes apoptosis in cardiomyocytes via the mitochondrial pathway. In addition, downregulation of GJIC by SNPs in cardiomyocytes is mediated through downregulation of Cx43 and upregulation of P-Cx43. These results suggest that in rat cardiomyocytes cell line, GJIC plays a protective role in SNPs-induced apoptosis and that GJIC may be one of the targets for SNPs-induced biological

  18. Computational simulations of asymmetric fluxes of large molecules through gap junction channel pores.

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    Mondal, Abhijit; Appadurai, Daniel A; Akoum, Nazem W; Sachse, Frank B; Moreno, Alonso P

    2017-01-07

    Gap junction channels are formed out of connexin isoforms, which enable molecule and ion selective diffusion amongst neighboring cells. HeLa cells expressing distinct connexins (Cx) allow the formation of heterotypic channels, where we observed a molecular charge-independent preferential flux of large fluorescent molecules in the Cx45 to Cx43 direction. We hypothesize that the pore's shape is a significant factor along-side charge and transjunctional voltages for this asymmetric flux. To test this hypothesis, we developed a 3D computational model simulating Brownian diffusion of large molecules in a gap junction channel pore. The basic pore contour was derived from x-ray crystallographic structures of Cx43 and Cx26 and approximated using basic geometric shapes. Lucifer yellow dye molecules and cesium counter-ions were modeled as spheres using their respective Stokes radii. Our simulation results from simple diffusion and constant concentration gradient experiments showed that only charged particles yield asymmetric fluxes in heterotypic pores. While increasing the inner mouth size resulted in a near-quadratic rise in flux, the rise was asymptotic for outer mouth radii increase. Probability maps and average force per particle per pore section explain the asymmetric flux with variation in pore shape. Furthermore, the simulation results are in agreement with our in vitro experimental results with HeLa cells in Cx43-Cx45 heterotypic configurations. The presence of asymmetric fluxes can help us to understand effects of the molecular structure of the pore and predict potential differences in vivo.

  19. Researches on Regulation Mechanism of Connexin 43 Expression in Reproductive System by Sexual Hormones%性激素对生殖系统Cx43表达调控机制的研究进展

    Institute of Scientific and Technical Information of China (English)

    孙馥箐; 段华; 汪沙

    2015-01-01

    由连接蛋白43 (connexin 43,Cx43)构成的细胞间间隙连接(gap junction,GJ)是介导细胞间直接的物质、能量交换及电、化学信号耦合的重要通路,其可保证细胞间功能活动的协调一致,在生殖系统细胞发育、分化及成熟等生理过程及功能活动中发挥非常重要的调控作用.性激素可在转录及翻译等层面调节生殖系统细胞Cx43的结构及表达,一方面通过核受体基因组机制调节Cx43基因的转录,另一方面通过非基因组快速信号传导通路机制调节Cx43的磷酸化水平,共同影响细胞间隙连接通讯(gapjunction intracellular communication,GJIC),进一步干预生殖系统细胞的病理、生理过程及功能活动.近年来,性激素在心血管系统病生理状态下对连接蛋白调节的变化及机制研究较为成熟,而在子宫、卵巢等生殖器官中,对性激素通过调节连接蛋白及间隙连接进而影响细胞间信息流通的研究较少,其作用机制并不清晰.故本文结合近年来文献,综述性激素对生殖系统细胞Cx43表达及GJIC的调控机制的研究进展,旨在为今后深入地研究提供可行的思路.%Gap junction (G J) composed of connexin 43 (Cx43) is a kind of cell-cell junction which significantly mediates the direct interchange of metabolites,energy and electrochemical signals from one cell to another.GJ ensures the coordination of intercellular activities and plays a vital role in the regulation of cell development,differentiation,maturation and functional activities in the reproductive system.Sexual hormones can influence the pathophysiological processes and functional activities of the reproductive cells by regulating gap junction intracellular communication (GJIC),which depends on the expression of Cx43.On one hand,sexual hormones regulate Cx43 gene transcription via the nuclear receptor mechanism,on the other hand,sexual hormones regulate Cx43 phosphorylation level through the signal

  20. Layer specific changes of astroglial gap junctions in the rat cerebellar cortex by persistent Borna Disease Virus infection.

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    Köster-Patzlaff, Christiane; Hosseini, Seyed Mehdi; Reuss, Bernhard

    2008-07-11

    Neonatal Borna Disease Virus (BDV) infection of the Lewis rat brain, leads to Purkinje cell degeneration, in association with astroglial activation. Since astroglial gap junctions (GJ) are known to influence neuronal degeneration, we investigated BDV dependent changes in astroglial GJ connexins (Cx) Cx43, and Cx30 in the Lewis rat cerebellum, 4, and 8 weeks after neonatal infection. On the mRNA level, RT-PCR demonstrated a BDV dependent increase in cerebellar Cx43, and a decrease in Cx30, 8, but not 4 weeks p.i. On the protein level, Western blot analysis revealed no overall upregulation of Cx43, but an increase of its phosphorylated forms, 8 weeks p.i. Cx30 protein was downregulated. Immunohistochemistry revealed a BDV dependent reduction of Cx43 in the granular layer (GL), 4 weeks p.i. 8 weeks p.i., Cx43 immunoreactivity recovered in the GL, and was induced in the molecular layer (ML). Cx30 revealed a BDV dependent decrease in the GL, both 4, and 8 weeks p.i. Changes in astroglial Cxs correlated not with expression of the astrogliotic marker GFAP, which was upregulated in radial glia. With regard to functional coupling, primary cerebellar astroglial cultures, revealed a BDV dependent increase of Cx43, and Cx30 immunoreactivity and in spreading of the GJ permeant dye Lucifer Yellow. These results demonstrate a massive, BDV dependent reorganization of astroglial Cx expression, and of functional GJ coupling in the cerebellar cortex, which might be of importance for the BDV dependent neurodegeneration in this brain region.

  1. Gap junctions in the control of vascular function.

    Science.gov (United States)

    Figueroa, Xavier F; Duling, Brian R

    2009-02-01

    Direct intercellular communication via gap junctions is critical in the control and coordination of vascular function. In the cardiovascular system, gap junctions are made up of one or more of four connexin proteins: Cx37, Cx40, Cx43, and Cx45. The expression of more than one gap-junction protein in the vasculature is not redundant. Rather, vascular connexins work in concert, first during the development of the cardiovascular system, and then in integrating smooth muscle and endothelial cell function, and in coordinating cell function along the length of the vessel wall. In addition, connexin-based channels have emerged as an important signaling pathway in the astrocyte-mediated neurovascular coupling. Direct electrical communication between endothelial cells and vascular smooth muscle cells via gap junctions is thought to play a relevant role in the control of vasomotor tone, providing the signaling pathway known as endothelium-derived hyperpolarizing factor (EDHF). Consistent with the importance of gap junctions in the regulation of vasomotor tone and arterial blood pressure, the expression of connexins is altered in diseases associated with vascular complications. In this review, we discuss the participation of connexin-based channels in the control of vascular function in physiologic and pathologic conditions, with a special emphasis on hypertension and diabetes.

  2. Adipocytes in both brown and white adipose tissue of adult mice are functionally connected via gap junctions: implications for Chagas disease.

    Science.gov (United States)

    Burke, Shoshana; Nagajyothi, Fnu; Thi, Mia M; Hanani, Menachem; Scherer, Philipp E; Tanowitz, Herbert B; Spray, David C

    2014-11-01

    Adipose tissue serves as a host reservoir for the protozoan Trypanosoma cruzi, the causative organism in Chagas disease. Gap junctions interconnect cells of most tissues, serving to synchronize cell activities including secretion in glandular tissue, and we have previously demonstrated that gap junctions are altered in various tissues and cells infected with T. cruzi. Herein, we examined the gap junction protein connexin 43 (Cx43) expression in infected adipose tissues. Adipose tissue is the largest endocrine organ of the body and is also involved in other physiological functions. In mammals, it is primarily composed of white adipocytes. Although gap junctions are a prominent feature of brown adipocytes, they have not been explored extensively in white adipocytes, especially in the setting of infection. Thus, we examined functional coupling in both white and brown adipocytes in mice. Injection of electrical current or the dye Lucifer Yellow into adipocytes within fat tissue spread to adjacent cells, which was reduced by treatment with agents known to block gap junctions. Moreover, Cx43 was detected in both brown and white fat tissue. At thirty and ninety days post-infection, Cx43 was downregulated in brown adipocytes and upregulated in white adipocytes. Gap junction-mediated intercellular communication likely contributes to hormone secretion and other functions in white adipose tissue and to nonshivering thermogenesis in brown fat, and modulation of the coupling by T. cruzi infection is expected to impact these functions. Copyright © 2014. Published by Elsevier Masson SAS.

  3. Impact of obesity on 7,12-dimethylbenz[a]anthracene-induced altered ovarian connexin gap junction proteins in female mice.

    Science.gov (United States)

    Ganesan, Shanthi; Nteeba, Jackson; Keating, Aileen F

    2015-01-01

    The ovarian gap junction proteins alpha 4 (GJA4 or connexin 37; CX37), alpha 1 (GJA1 or connexin 43; CX43) and gamma 1 (GJC1 or connexin 45; CX45) are involved in cell communication and folliculogenesis. 7,12-dimethylbenz[a]anthracene (DMBA) alters Cx37 and Cx43 expression in cultured neonatal rat ovaries. Additionally, obesity has an additive effect on DMBA-induced ovarian cell death and follicle depletion, thus, we investigated in vivo impacts of obesity and DMBA on CX protein levels. Ovaries were collected from lean and obese mice aged 6, 12, 18, or 24 wks. A subset of 18 wk old mice (lean and obese) were dosed with sesame oil or DMBA (1mg/kg; ip) for 14days and ovaries collected 3days thereafter. Cx43 and Cx45 mRNA and protein levels decreased (Pobese ovaries. Cx37 mRNA and antral follicle protein staining intensity were reduced (Pobesity while total CX37 protein was reduced (Pobese ovaries. Cx43 mRNA and total protein levels were decreased (Pobese ovaries while basal protein staining intensity was reduced (Pobese controls. Cx45 mRNA, total protein and protein staining intensity level were decreased (Pobesity. These data support that obesity temporally alters gap junction protein expression and that DMBA-induced ovotoxicity may involve reduced gap junction protein function. Copyright © 2014 Elsevier Inc. All rights reserved.

  4. Effect of the gap junction blocker 1-heptanol on chondrogenic differentiation of mouse bone marrow mesenchymal stem cells in vitro

    Institute of Scientific and Technical Information of China (English)

    Liu Ouyang; Yukun Zhang; Shuhua Yang; Shunan Ye; Weihua Xu

    2009-01-01

    Objective:To investigate the effect of the gap junction blocker 1-heptanol on the in vitro chondrogenic differentiation of mouse bone marrow mesencbymal stem cells(MSCs) following induction by GDF-5. Methods:MSCs were isohted from mouse bone marrow and cultured in vitro. After 3 passages cells were induced to undergo chondrogenic differentiation with recombinant human GDF-5(100 ng/ml), with or without 1-heptanol(2.5 μ mol/L). The effect of 1-heptanol on MSCs proliferation was investigated using the MTT assay. Type Ⅱ collagen mRNA and protein were examined by RT-PCR and immunocytochernistry respectively, and the sulfate glycosaminoglycan was assessed by Alcian blue dye staining. Connexin43(Cx43) protein was examined by western blotting. Results:GDF-5 induced proliferation and chondrogenic differentiation of MSCs. While 1-heptanol treatment had no effect on this proliferation, it inhibited the expression of both type Ⅱ collagen mRNA and protein. The Alcian blue staining revealed that 1-heptanol also inhibited the deposition of the typical cartilage extracellular matrix promoted by recombinant GDF-5. Western blotting demonstrated that 1-heptanol had no effect on the expression of Cx43. Conclusion:Tbese results suggest that mouse bone marrow MSCs can be differentiated into a chondrogenic phenotype by GDF-5 administration in vitro. While the gap junction blocker, 1-heptanol, did not reduce gap junction Cx43, these intercellular communication pathways clearly played an important functional role in GDF-5-induced cartilage differentiation.

  5. Human articular chondrocytes express multiple gap junction proteins: differential expression of connexins in normal and osteoarthritic cartilage.

    Science.gov (United States)

    Mayan, Maria D; Carpintero-Fernandez, Paula; Gago-Fuentes, Raquel; Martinez-de-Ilarduya, Oskar; Wang, Hong-Zhang; Valiunas, Virginijus; Brink, Peter; Blanco, Francisco J

    2013-04-01

    Osteoarthritis (OA) is the most common joint disease and involves progressive degeneration of articular cartilage. The aim of this study was to investigate if chondrocytes from human articular cartilage express gap junction proteins called connexins (Cxs). We show that human chondrocytes in tissue express Cx43, Cx45, Cx32, and Cx46. We also find that primary chondrocytes from adults retain the capacity to form functional voltage-dependent gap junctions. Immunohistochemistry experiments in cartilage from OA patients revealed significantly elevated levels of Cx43 and Cx45 in the superficial zone and down through the next approximately 1000 μm of tissue. These zones corresponded with regions damaged in OA that also had high levels of proliferative cell nuclear antigen. An increased number of Cxs may help explain the increased proliferation of cells in clusters that finally lead to tissue homeostasis loss. Conversely, high levels of Cxs in OA cartilage reflect the increased number of adjacent cells in clusters that are able to interact directly by gap junctions as compared with hemichannels on single cells in normal cartilage. Our data provide strong evidence that OA patients have a loss of the usual ordered distribution of Cxs in the damaged zones and that the reductions in Cx43 levels are accompanied by the loss of correct Cx localization in the nondamaged areas. Copyright © 2013 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

  6. Cytokine effects on gap junction communication and connexin expression in human bladder smooth muscle cells and suburothelial myofibroblasts.

    Directory of Open Access Journals (Sweden)

    Marco Heinrich

    Full Text Available BACKGROUND: The last decade identified cytokines as one group of major local cell signaling molecules related to bladder dysfunction like interstitial cystitis (IC and overactive bladder syndrome (OAB. Gap junctional intercellular communication (GJIC is essential for the coordination of normal bladder function and has been found to be altered in bladder dysfunction. Connexin (Cx 43 and Cx45 are the most important gap junction proteins in bladder smooth muscle cells (hBSMC and suburothelial myofibroblasts (hsMF. Modulation of connexin expression by cytokines has been demonstrated in various tissues. Therefore, we investigate the effect of interleukin (IL 4, IL6, IL10, tumor necrosis factor-alpha (TNFα and transforming growth factor-beta1 (TGFβ1 on GJIC, and Cx43 and Cx45 expression in cultured human bladder smooth muscle cells (hBSMC and human suburothelial myofibroblasts (hsMF. METHODOLOGY/PRINCIPAL FINDINGS: HBSMC and hsMF cultures were set up from bladder tissue of patients undergoing cystectomy. In cytokine stimulated cultured hBSMC and hsMF GJIC was analyzed via Fluorescence Recovery after Photo-bleaching (FRAP. Cx43 and Cx45 expression was assessed by quantitative PCR and confocal immunofluorescence. Membrane protein fraction of Cx43 and Cx45 was quantified by Dot Blot. Upregulation of cell-cell-communication was found after IL6 stimulation in both cell types. In hBSMC IL4 and TGFβ1 decreased both, GJIC and Cx43 protein expression, while TNFα did not alter communication in FRAP-experiments but increased Cx43 expression. GJ plaques size correlated with coupling efficacy measured, while Cx45 expression did not correlate with modulation of GJIC. CONCLUSIONS/SIGNIFICANCE: Our finding of specific cytokine effects on GJIC support the notion that cytokines play a pivotal role for pathophysiology of OAB and IC. Interestingly, the effects were independent from the classical definition of pro- and antiinflammatory cytokines. We conclude, that

  7. Impaired astrocytic gap junction coupling and potassium buffering in a mouse model of tuberous sclerosis complex.

    Science.gov (United States)

    Xu, Lin; Zeng, Ling-Hui; Wong, Michael

    2009-05-01

    Abnormalities in astrocytes occur in the brains of patients with Tuberous Sclerosis Complex (TSC) and may contribute to the pathogenesis of neurological dysfunction in this disease. Here, we report that knock-out mice with Tsc1 gene inactivation in glia (Tsc1(GFAP)CKO mice) exhibit decreased expression of the astrocytic connexin protein, Cx43, and an associated impairment in gap junction coupling between astrocytes. Correspondingly, hippocampal slices from Tsc1(GFAP)CKO mice have increased extracellular potassium concentration in response to stimulation. This impaired potassium buffering can be attributed to abnormal gap junction coupling, as a gap junction inhibitor elicits an additional increase in potassium concentration in control, but not Tsc1(GFAP)CKO slices. Furthermore, treatment with a mammalian target of rapamycin inhibitor reverses the deficient Cx43 expression and impaired potassium buffering. These findings suggest that Tsc1 inactivation in astrocytes causes defects in astrocytic gap junction coupling and potassium clearance, which may contribute to epilepsy in Tsc1(GFAP)CKO mice.

  8. The relationship between gap junctional remodeling and human atrial fibrillation

    Institute of Scientific and Technical Information of China (English)

    李大强; 冯义柏; 张会琴

    2004-01-01

    @@ Atrial fibrillation (AF) is currently the most common cardiac tachyarrhythmia in clinical practice. AF has a tendency to become more persistent over time. Progression of an underlying disease is one explanation. Another possible explanation is electrical, structural, and gap junctional remodeling of the atrium by repetitive induction of AF.1 The expression level and distribution of it have close relation with the conduction velocity of electrical activation in the atrium. The aim of the present study was to investigate the alternations of the expression and distribution of (connexin 40, Cx 40) and (connexin 43, Cx 43) in the right atrial appendages of the patients with AF by laser confocal scanning microscopy and Western blot technique.

  9. Impact of obesity on 7,12-dimethylbenz[a]anthracene-induced altered ovarian connexin gap junction proteins in female mice

    Energy Technology Data Exchange (ETDEWEB)

    Ganesan, Shanthi, E-mail: shanthig@iastate.edu; Nteeba, Jackson, E-mail: nteeba@iastate.edu; Keating, Aileen F., E-mail: akeating@iastate.edu

    2015-01-01

    The ovarian gap junction proteins alpha 4 (GJA4 or connexin 37; CX37), alpha 1 (GJA1 or connexin 43; CX43) and gamma 1 (GJC1 or connexin 45; CX45) are involved in cell communication and folliculogenesis. 7,12-dimethylbenz[a]anthracene (DMBA) alters Cx37 and Cx43 expression in cultured neonatal rat ovaries. Additionally, obesity has an additive effect on DMBA-induced ovarian cell death and follicle depletion, thus, we investigated in vivo impacts of obesity and DMBA on CX protein levels. Ovaries were collected from lean and obese mice aged 6, 12, 18, or 24 wks. A subset of 18 wk old mice (lean and obese) were dosed with sesame oil or DMBA (1 mg/kg; ip) for 14 days and ovaries collected 3 days thereafter. Cx43 and Cx45 mRNA and protein levels decreased (P < 0.05) after 18 wks while Cx37 mRNA and protein levels decreased (P < 0.05) after 24 wks in obese ovaries. Cx37 mRNA and antral follicle protein staining intensity were reduced (P < 0.05) by obesity while total CX37 protein was reduced (P < 0.05) in DMBA exposed obese ovaries. Cx43 mRNA and total protein levels were decreased (P < 0.05) by DMBA in both lean and obese ovaries while basal protein staining intensity was reduced (P < 0.05) in obese controls. Cx45 mRNA, total protein and protein staining intensity level were decreased (P < 0.05) by obesity. These data support that obesity temporally alters gap junction protein expression and that DMBA-induced ovotoxicity may involve reduced gap junction protein function. - Highlights: • Ovarian gap junction proteins are affected by ovarian aging and obesity. • DMBA exposure negatively impacts gap junction proteins. • Altered gap junction proteins may contribute to infertility.

  10. Methamphetamine compromises gap junctional communication in astrocytes and neurons.

    Science.gov (United States)

    Castellano, Paul; Nwagbo, Chisom; Martinez, Luis R; Eugenin, Eliseo A

    2016-05-01

    Methamphetamine (meth) is a central nervous system (CNS) stimulant that results in psychological and physical dependency. The long-term effects of meth within the CNS include neuronal plasticity changes, blood-brain barrier compromise, inflammation, electrical dysfunction, neuronal/glial toxicity, and an increased risk to infectious diseases including HIV. Most of the reported meth effects in the CNS are related to dysregulation of chemical synapses by altering the release and uptake of neurotransmitters, especially dopamine, norepinephrine, and epinephrine. However, little is known about the effects of meth on connexin (Cx) containing channels, such as gap junctions (GJ) and hemichannels (HC). We examined the effects of meth on Cx expression, function, and its role in NeuroAIDS. We found that meth altered Cx expression and localization, decreased GJ communication between neurons and astrocytes, and induced the opening of Cx43/Cx36 HC. Furthermore, we found that these changes in GJ and HC induced by meth treatment were mediated by activation of dopamine receptors, suggesting that dysregulation of dopamine signaling induced by meth is essential for GJ and HC compromise. Meth-induced changes in GJ and HC contributed to amplified CNS toxicity by dysregulating glutamate metabolism and increasing the susceptibility of neurons and astrocytes to bystander apoptosis induced by HIV. Together, our results indicate that connexin containing channels, GJ and HC, are essential in the pathogenesis of meth and increase the sensitivity of the CNS to HIV CNS disease. Methamphetamine (meth) is an extremely addictive central nervous system stimulant. Meth reduced gap junctional (GJ) communication by inducing internalization of connexin-43 (Cx43) in astrocytes and reducing expression of Cx36 in neurons by a mechanism involving activation of dopamine receptors (see cartoon). Meth-induced changes in Cx containing channels increased extracellular levels of glutamate and resulted in higher

  11. Effects of mechanical strain on the function of Gap junctions in osteocytes are mediated through the prostaglandin EP2 receptor.

    Science.gov (United States)

    Cherian, Priscilla P; Cheng, Benxu; Gu, Sumin; Sprague, Eugene; Bonewald, Lynda F; Jiang, Jean X

    2003-10-31

    Osteocytes embedded in the matrix of bone are thought to be mechanosensory cells that translate mechanical strain into biochemical signals that regulate bone modeling and remodeling. We have shown previously that fluid flow shear stress dramatically induces prostaglandin release and COX-2 mRNA expression in osteocyte-like MLO-Y4 cells, and that prostaglandin E2 (PGE2) released by these cells functions in an autocrine manner to regulate gap junction function and connexin 43 (Cx43) expression. Here we show that fluid flow regulates gap junctions through the PGE2 receptor EP2 activation of cAMP-dependent protein kinase A (PKA) signaling. The expression of the EP2 receptor, but not the subtypes EP1,EP3, and EP4, increased in response to fluid flow. Application of PGE2 or conditioned medium from fluid flow-treated cells to non-stressed MLO-Y4 cells increased expression of the EP2 receptor. The EP2 receptor antagonist, AH6809, suppressed the stimulatory effects of PGE2 and fluid flow-conditioned medium on the expression of the EP2 receptor, on Cx43 protein expression, and on gap junction-mediated intercellular coupling. In contrast, the EP2 receptor agonist butaprost, not the E1/E3 receptor agonist sulprostone, stimulated the expression of Cx43 and gap junction function. Fluid flow conditioned medium and PGE2 stimulated cAMP production and PKA activity suggesting that PGE2 released by mechanically stimulated cells is responsible for the activation of cAMP and PKA. The adenylate cyclase activators, forskolin and 8-bromo-cAMP, enhanced intercellular connectivity, the number of functional gap junctions, and Cx43 protein expression, whereas the PKA inhibitor, H89, inhibited the stimulatory effect of PGE2 on gap junctions. These studies suggest that the EP2 receptor mediates the effects of autocrine PGE2 on the osteocyte gap junction in response to fluid flow-induced shear stress. These data support the hypothesis that the EP2 receptor, cAMP, and PKA are critical components

  12. Functional role of gap junctions in cytokine-induced leukocyte adhesion to endothelium in vivo

    Science.gov (United States)

    Véliz, Loreto P.; González, Francisco G.; Duling, Brian R.; Sáez, Juan C.; Boric, Mauricio P.

    2008-01-01

    To assess the hypothesis that gap junctions (GJs) participate on leukocyte-endothelium interactions in the inflammatory response, we compared leukocyte adhesion and transmigration elicited by cytokine stimulation in the presence or absence of GJ blockers in the hamster cheek pouch and also in the cremaster muscle of wild-type (WT) and endothelium-specific connexin 43 (Cx43) null mice (Cx43e−/−). In the cheek pouch, topical tumor necrosis factor-α (TNF-α; 150 ng/ml, 15 min) caused a sustained increment in the number of leukocytes adhered to venular endothelium (LAV) and located at perivenular regions (LPV). Superfusion with the GJ blockers 18-α-glycyrrhetinic acid (AGA; 75 μM) or 18-β-glycyrrhetinic acid (50 μM) abolished the TNF-α-induced increase in LAV and LPV; carbenoxolone (75 μM) or oleamide (100 μM) reduced LAV by 50 and 75%, respectively, and LPV to a lesser extent. None of these GJ blockers modified venular diameter, blood flow, or leukocyte rolling. In contrast, glycyrrhizin (75 μM), a non-GJ blocker analog of AGA, was devoid of effect. Interestingly, when AGA was removed 90 min after TNF-α stimulation, LAV started to rise at a similar rate as in control. Conversely, application of AGA 90 min after TNF-α reduced the number of previously adhered cells. In WT mice, intrascrotal injection of TNF-α (0.5 μg/0.3 ml) increased LAV (fourfold) and LPV (threefold) compared with saline-injected controls. In contrast to the observations in WT animals, TNF-α stimulation did not increase LAV or LPV in Cx43e−/− mice. These results demonstrate an important role for GJ communication in leukocyte adhesion and transmigration during acute inflammation in vivo and further suggest that endothelial Cx43 is key in these processes. PMID:18599597

  13. 戊四氮点燃癫痫大鼠海马Cx32和Cx43表达及甘珀酸对其表达的影响%The expression of Cx32 and Cx43 in the hippocampus of PTZ-kindled epileptic rats

    Institute of Scientific and Technical Information of China (English)

    兰莉; 兰怡; 段丽; 曹荣

    2013-01-01

    目的 探讨点燃癫痫大鼠神经元、星形胶质细胞及其缝隙蛋白的变化,甘珀酸(CBX)对缝隙连接的影响.方法 30只SD雄性大鼠随机分为3组:对照组(NS组)、点燃组(K组)、点燃后干预组(KCBX组),每组10只大鼠.NS组每日腹腔注射生理盐水.K组和KCBX组大鼠腹腔注射戊四氮(PTZ)[35mg/(kg·d)]至点燃,再分别腹腔注射生理盐水、CBX(10mg/kg)干预3d.观察3组大鼠惊厥行为变化,并观察Cx32、Cx43在海马内的表达变化.结果 ①PTZ点燃癫痫大鼠出现自发性抽搐,KCBX组自发性抽搐次数明显减少(P<0.05).②点燃癫痫大鼠海马Cx32、Cx43的表达增加;CBX干预后,Cx32、Cx43的表达减少(P<0.05).结论 CBX抑制点燃大鼠Cx32、Cx43的表达和癫痫活动,提示缝隙连接在癫癎的发生发展过程中具有重要的作用.%Objective To investigate the changes of neurons, astrocytes and their connexin in PTZ-kindled rats as well as the effect of carbenoxolone (CBX) on gap junction. Methods Totally 30 male SD rats were randomly divided into normal saline (NS) group, kindled (K) group and KCBX group, with 10 rats in each. NS group rats were intraperitoneally injected (i.p.) with saline daily. Rats in KCBX group and K group received PTZ [i.p. 35 mg/kg · d)] to establish epilepsy model of kindled rats. After kindling, KCBX group rats were injected with CBX (i.p. 10 mg/kg) and K group rats NS for three days. Seizure changes were analyzed for all the groups. Immunohistochemistry method was used to observe the expressions of Cx32 and Cx43 in the hippocampus of the rats. Results Spontaneous seizures appeared in PTZ-kindled epileptic rats. Accumulative times of seizures were significantly fewer in KCBX group rats than in K group rats (P<0. 05). The expressions of Cx32 and Cx43 increased significantly in the hippocampus of K group rats, but decreased in KCBX group rats (P<0. 05). Conclusion Gap junction blocker CBX inhibits not only the expressions of Cx32 and Cx43

  14. Gap junction connexins in female reproductive organs: implications for women's reproductive health.

    Science.gov (United States)

    Winterhager, Elke; Kidder, Gerald M

    2015-01-01

    Connexins comprise a family of ~20 proteins that form intercellular membrane channels (gap junction channels) providing a direct route for metabolites and signalling molecules to pass between cells. This review provides a critical analysis of the evidence for essential roles of individual connexins in female reproductive function, highlighting implications for women's reproductive health. No systematic review has been carried out. Published literature from the past 35 years was surveyed for research related to connexin involvement in development and function of the female reproductive system. Because of the demonstrated utility of genetic manipulation for elucidating connexin functions in various organs, much of the cited information comes from research with genetically modified mice. In some cases, a distinction is drawn between connexin functions clearly related to the formation of gap junction channels and those possibly linked to non-channel roles. Based on work with mice, several connexins are known to be required for female reproductive functions. Loss of connexin43 (CX43) causes an oocyte deficiency, and follicles lacking or expressing less CX43 in granulosa cells exhibit reduced growth, impairing fertility. CX43 is also expressed in human cumulus cells and, in the context of IVF, has been correlated with pregnancy outcome, suggesting that this connexin may be a determinant of oocyte and embryo quality in women. Loss of CX37, which exclusively connects oocytes with granulosa cells in the mouse, caused oocytes to cease growing without acquiring meiotic competence. Blocking of CX26 channels in the uterine epithelium disrupted implantation whereas loss or reduction of CX43 expression in the uterine stroma impaired decidualization and vascularization in mouse and human. Several connexins are important in placentation and, in the human, CX43 is a key regulator of the fusogenic pathway from the cytotrophoblast to the syncytiotrophoblast, ensuring placental growth

  15. Inhibition of gap junction intercellular communication is involved in silica nanoparticles-induced H9c2 cardiomyocytes apoptosis via the mitochondrial pathway

    Directory of Open Access Journals (Sweden)

    Du ZJ

    2017-03-01

    Full Text Available Zhong-jun Du,1 Guan-qun Cui,2 Juan Zhang,1 Xiao-mei Liu,3 Zhi-hu Zhang,1 Qiang Jia,1 Jack C Ng,4 Cheng Peng,1,4 Cun-xiang Bo,1 Hua Shao1 1Department of Toxicology, Shandong Academy of Occupational Health and Occupational Medicine, Shandong Academy of Medical Sciences, 2Department of Respiratory Medicine, Qilu Children’s Hospital of Shandong University, Jinan, 3Department of Radiation Chemistry and Toxicology, School of Public Health, Jilin University, Changchun, People’s Republic of China; 4National Research Centre for Environmental Toxicology-Entox, The University of Queensland, Brisbane, QLD, Australia Abstract: Gap junction intercellular communication (GJIC between cardiomyocytes is essential for synchronous heart contraction and relies on connexin-containing channels. Connexin 43 (Cx43 is a major component involved in GJIC in heart tissue, and its abnormal expression is closely associated with various cardiac diseases. Silica nanoparticles (SNPs are known to induce cardiovascular toxicity. However, the mechanisms through which GJIC plays a role in cardiomyocytes apoptosis induced by SNPs remain unknown. The aim of the present study is to determine whether SNPs-decreased GJIC promotes apoptosis in rat cardiomyocytes cell line (H9c2 cells via the mitochondrial pathway using CCK-8 Kit, scrape-loading dye transfer technique, Annexin V/PI double-staining assays, and Western blot analysis. The results showed that SNPs elicited cytotoxicity in H9c2 cells in a time- and concentration-dependent manner. SNPs also reduced GJIC in H9c2 cells in a concentration-dependent manner through downregulation of Cx43 and upregulation of P-Cx43. Inhibition of gap junctions by gap junction blocker carbenoxolone disodium resulted in decreased survival and increased apoptosis, whereas enhancement of the gap junctions by retinoic acid led to enhanced survival but decreased apoptosis. Furthermore, SNPs-induced apoptosis through the disrupted functional gap junction

  16. Gap Junction Enhancer Potentiates Cytotoxicity of Cisplatin in Breast Cancer Cells.

    Science.gov (United States)

    Ding, Ying; Nguyen, Thu Annelise

    2012-11-01

    Cisplatin is one of the most widely used anti-cancer drugs due to its ability to damage DNA and induce apoptosis. However, increasing reports of side effects and drug resistance indicate the limitation of cisplatin in cancer therapeutics. Recent studies showed that inhibition of gap junctions diminishes the cytotoxic effect and contributes to drug resistance. Therefore, identification of molecules that counteract gap junctional inhibition without decreasing the anti-cancer effect of cisplatin could be used in combinational treatment, potentiating cisplatin efficacy and preventing resistance. This study investigates the effects of combinational treatment of cisplatin and PQ1, a gap junction enhancer, in T47D breast cancer cells. Our results showed that combinational treatment of PQ1 and cisplatin increased gap junctional intercellular communication (GJIC) as well as expressions of connexins (Cx26, Cx32 and Cx43), and subsequently decreased cell viability. Ki67, a proliferation marker, was decreased by 75% with combinational treatment. Expressions of pro-apoptotic factors (cleaved caspase-3/-8/-9 and bax) were increased by the combinational treatment with PQ1 and cisplatin; whereas, the pro-survival factor, bcl-2, was decreased by the combinational treatment. Our study demonstrates for the first time that the combinational treatment with gap junction enhancers can counteract cisplatin induced inhibition of gap junctional intercellular communication and reduction of connexin expression, thereby increasing the efficacy of cisplatin in cancer cells.

  17. Illuminating Myocyte-Fibroblast Homotypic and Heterotypic Gap Junction Dynamics Using Dynamic Clamp.

    Science.gov (United States)

    Brown, Tashalee R; Krogh-Madsen, Trine; Christini, David J

    2016-08-23

    Fibroblasts play a significant role in the development of electrical and mechanical dysfunction of the heart; however, the underlying mechanisms are only partially understood. One widely studied mechanism suggests that fibroblasts produce excess extracellular matrix, resulting in collagenous septa that slow propagation, cause zig-zag conduction paths, and decouple cardiomyocytes, resulting in a substrate for cardiac arrhythmia. An emerging mechanism suggests that fibroblasts promote arrhythmogenesis through direct electrical interactions with cardiomyocytes via gap junction (GJ) channels. In the heart, three major connexin (Cx) isoforms, Cx40, Cx43, and Cx45, form GJ channels in cell-type-specific combinations. Because each Cx is characterized by a unique time- and transjunctional voltage-dependent profile, we investigated whether the electrophysiological contributions of fibroblasts would vary with the specific composition of the myocyte-fibroblast (M-F) GJ channel. Due to the challenges of systematically modifying Cxs in vitro, we coupled native cardiomyocytes with in silico fibroblast and GJ channel electrophysiology models using the dynamic-clamp technique. We found that there is a reduction in the early peak of the junctional current during the upstroke of the action potential (AP) due to GJ channel gating. However, effects on the cardiomyocyte AP morphology were similar regardless of the specific type of GJ channel (homotypic Cx43 and Cx45, and heterotypic Cx43/Cx45 and Cx45/Cx43). To illuminate effects at the tissue level, we performed multiscale simulations of M-F coupling. First, we developed a cell-specific model of our dynamic-clamp experiments and investigated changes in the underlying membrane currents during M-F coupling. Second, we performed two-dimensional tissue sheet simulations of cardiac fibrosis and incorporated GJ channels in a cell type-specific manner. We determined that although GJ channel gating reduces junctional current, it does not

  18. An atrial-fibrillation-linked connexin40 mutant is retained in the endoplasmic reticulum and impairs the function of atrial gap-junction channels

    Directory of Open Access Journals (Sweden)

    Yiguo Sun

    2014-05-01

    Full Text Available Connexin40 (Cx40-containing gap-junction channels are expressed in the atrial myocardium and provide a low-resistance passage for rapid impulse propagation. A germline mutation in the GJA5 gene, which encodes Cx40, resulting in a truncated Cx40 (Q49X was identified in a large Chinese family with lone (idiopathic atrial fibrillation (AF. This mutation co-segregated with seven AF probands in an autosomal-dominant way over generations. To test the hypothesis that this Cx40 mutant affects the distribution and function of atrial gap junctions, we studied the Q49X mutant in gap-junction-deficient HeLa and N2A cells. The Q49X mutant, unlike wild-type Cx40, was typically localized in the cytoplasm and failed to form gap-junction plaques at cell-cell interfaces. When the Q49X mutant was co-expressed with Cx40 or Cx43, the mutant substantially reduced the gap-junction plaque formation of Cx40 and Cx43. Electrophysiological studies revealed no electrical coupling of cell pairs expressing the mutant alone and a significant decrease in the coupling conductance when the mutant was co-expressed with Cx40 or Cx43. Further colocalization experiments with the organelle residential proteins indicate that Q49X was retained in the endoplasmic reticulum. These findings provide evidence that the Q49X mutant is capable of impairing gap-junction distribution and function of key atrial connexins, which might play a role in the predisposition to and onset of AF.

  19. Segregated Foxc2, NFATc1 and Connexin expression at normal developing venous valves, and Connexin-specific differences in the valve phenotypes of Cx37, Cx43, and Cx47 knockout mice.

    Science.gov (United States)

    Munger, Stephanie J; Geng, Xin; Srinivasan, R Sathish; Witte, Marlys H; Paul, David L; Simon, Alexander M

    2016-04-15

    Venous valves (VVs) are critical for unidirectional blood flow from superficial and deep veins towards the heart. Congenital valve aplasia or agenesis may, in some cases, be a direct cause of vascular disease, motivating an understanding of the molecular mechanisms underlying the development and maintenance of VVs. Three gap junction proteins (Connexins), Cx37, Cx43, and Cx47, are specifically expressed at VVs in a highly polarized fashion. VVs are absent from adult mice lacking Cx37; however it is not known if Cx37 is required for the initial formation of valves. In addition, the requirement of Cx43 and Cx47 for VV development has not been studied. Here, we provide a detailed description of Cx37, Cx43, and Cx47 expression during mouse vein development and show by gene knockout that each Cx is necessary for normal valve development. The valve phenotypes in the knockout lines exhibit Cx-specific differences, however, including whether peripheral or central VVs are affected by gene inactivation. In addition, we show that a Cx47 null mutation impairs peripheral VV development but does not affect lymphatic valve formation, a finding of significance for understanding how some CX47 mutations cause inherited lymphedema in humans. Finally, we demonstrate a striking segregation of Foxc2 and NFATc1 transcription factor expression between the downstream and upstream faces, respectively, of developing VV leaflets and show that this segregation is closely associated with the highly polarized expression of Cx37, Cx43, and Cx47. The partition of Foxc2 and NFATc1 expression at VV leaflets makes it unlikely that these factors directly cooperate during the leaflet elongation stage of VV development.

  20. Gap Junctions Contribute to the Regulation of Walking-Like Activity in the Adult Mudpuppy (Necturus Maculatus).

    Science.gov (United States)

    Lavrov, Igor; Fox, Lyle; Shen, Jun; Han, Yingchun; Cheng, Jianguo

    2016-01-01

    Although gap junctions are widely expressed in the developing central nervous system, the role of electrical coupling of neurons and glial cells via gap junctions in the spinal cord in adults is largely unknown. We investigated whether gap junctions are expressed in the mature spinal cord of the mudpuppy and tested the effects of applying gap junction blocker on the walking-like activity induced by NMDA or glutamate in an in vitro mudpuppy preparation. We found that glial and neural cells in the mudpuppy spinal cord expressed different types of connexins that include connexin 32 (Cx32), connexin 36 (Cx36), connexin 37 (Cx37), and connexin 43 (Cx43). Application of a battery of gap junction blockers from three different structural classes (carbenexolone, flufenamic acid, and long chain alcohols) substantially and consistently altered the locomotor-like activity in a dose-dependent manner. In contrast, these blockers did not significantly change the amplitude of the dorsal root reflex, indicating that gap junction blockers did not inhibit neuronal excitability nonselectively in the spinal cord. Taken together, these results suggest that gap junctions play a significant modulatory role in the spinal neural networks responsible for the generation of walking-like activity in the adult mudpuppy.

  1. Gap Junctions Contribute to the Regulation of Walking-Like Activity in the Adult Mudpuppy (Necturus Maculatus.

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    Igor Lavrov

    Full Text Available Although gap junctions are widely expressed in the developing central nervous system, the role of electrical coupling of neurons and glial cells via gap junctions in the spinal cord in adults is largely unknown. We investigated whether gap junctions are expressed in the mature spinal cord of the mudpuppy and tested the effects of applying gap junction blocker on the walking-like activity induced by NMDA or glutamate in an in vitro mudpuppy preparation. We found that glial and neural cells in the mudpuppy spinal cord expressed different types of connexins that include connexin 32 (Cx32, connexin 36 (Cx36, connexin 37 (Cx37, and connexin 43 (Cx43. Application of a battery of gap junction blockers from three different structural classes (carbenexolone, flufenamic acid, and long chain alcohols substantially and consistently altered the locomotor-like activity in a dose-dependent manner. In contrast, these blockers did not significantly change the amplitude of the dorsal root reflex, indicating that gap junction blockers did not inhibit neuronal excitability nonselectively in the spinal cord. Taken together, these results suggest that gap junctions play a significant modulatory role in the spinal neural networks responsible for the generation of walking-like activity in the adult mudpuppy.

  2. Gap junction communication involved in brain protection following focal ischemia and reperfusion in rats

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    Fei Wang; Jian Hai; Yuhong Jing

    2009-01-01

    BACKGROUND: Studies have suggested that gap junctions not only modulate the fate of the neocortex, but are also involved in maintaining homeostasis in the mature brain. However, the neuroprotective effects of gap junction communication following brain ischemic injury remain poorly understood. OBJECTIVE: To investigate the neuroprotective effects and possible mechanisms of gap junction communication following focal ischemia and reperfusion. DESIGN, TIME AND SETTING: A randomized, controlled, animal experiment was performed at the School of Basic Medical Sciences of Lanzhou University between June 2007 and May 2008.MATERIALS: Rabbit polyclonal anti-connexin 43 (Cx43) and gap junction blocking agent octanol were purchased from Sigma, USA;mouse monoclonal anti-rat glial fibrillary acidic protein (GFAP) was provided by Santa Cruz, USA;mouse monoclonal anti-rat CD11b was produced by Abcam, England. METHODS: A total of 52 adult, male, Sprague Dawley rats were randomly assigned to three groups: sham-operated (n=12), vehicle control (n=20), and octanol-treated (n=20). Brain ischemia and reperfusion were induced by transient middle cerebral artery occlusion (MCAO) in vehicle control and octanol-treated groups, while no MCAO was administered to the sham-operated group. In the octanol-treated group, 5 mmol/kg octanol was dissolved in dimethyl sulfoxide (0.005% v/v) and was intraperitoneally injected 30 minutes prior to ischemic onset. Sham-operated and vehicle groups received equivalent volumes of dimethyl sulfoxide. MAIN OUTCOME MEASURES: Infarct volumes in ipsilateral striatum after MCAO were measured using cresyl violet dye;GFAP, CD11b, and Cx43 expression in the ipsilateral striatum following MCAO were detected by immunohistochemistry;Western blot analysis was employed to determine Cx43 and GFAP expression. RESULTS: At 1 and 3 days following MCAO and reperfusion, ipsilateral striatum infarct volumes in the octanol group were significantly greater than in the vehicle group

  3. Modulation of connexin expression and gap junction communication in astrocytes by the gram-positive bacterium S. aureus.

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    Esen, Nilufer; Shuffield, Debbie; Syed, Mohsin M D; Kielian, Tammy

    2007-01-01

    Gap junctions establish direct intercellular conduits between adjacent cells and are formed by the hexameric organization of protein subunits called connexins (Cx). It is unknown whether the proinflammatory milieu that ensues during CNS infection with S. aureus, one of the main etiologic agents of brain abscess in humans, is capable of eliciting regional changes in astrocyte homocellular gap junction communication (GJC) and, by extension, influencing neuron homeostasis at sites distant from the primary focus of infection. Here we investigated the effects of S. aureus and its cell wall product peptidoglycan (PGN) on Cx43, Cx30, and Cx26 expression, the main Cx isoforms found in astrocytes. Both bacterial stimuli led to a time-dependent decrease in Cx43 and Cx30 expression; however, Cx26 levels were elevated following bacterial exposure. Functional examination of dye coupling, as revealed by single-cell microinjections of Lucifer yellow, demonstrated that both S. aureus and PGN inhibited astrocyte GJC. Inhibition of protein synthesis with cyclohexamide (CHX) revealed that S. aureus directly modulates, in part, Cx43 and Cx30 expression, whereas Cx26 levels appear to be regulated by a factor(s) that requires de novo protein production; however, CHX did not alter the inhibitory effects of S. aureus on astrocyte GJC. The p38 MAPK inhibitor SB202190 was capable of partially restoring the S. aureus-mediated decrease in astrocyte GJC to that of unstimulated cells, suggesting the involvement of p38 MAPK-dependent pathway(s). These findings could have important implications for limiting the long-term detrimental effects of abscess formation in the brain which may include seizures and cognitive deficits. Copyright 2006 Wiley-Liss, Inc.

  4. Changes of transmural heterogeneity of Cx43 expression in acute myocardial ischemia reperfusion

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    Jiang-Hua Zhong; Shi-Juan Lu; Xiao-Pan Chen; Qi Zuo; Zheng-Wang Liu; Ding-Jun Sun; Miao Wu

    2016-01-01

    Objective:To observe the change of transmural heterogeneity of Cx43 protein among three myocardial layers and explore physiological mechanisms of malignant ventricular arrhythmia (MVA) in myocardial ischemia reperfusion (MIR).Methods: Twenty rabbits were randomly divided into MIR group (n=10) which rabbits with MIR were made in and sham group (n=10). 90% monophasic action potential repolarization duration (APD90), transmural dispersion of repolarization (TDR) and Cx43 protein (Cx43-pro) and mRNA (Cx43-Cq) expression among three myocardial layers were measured in both groups.Results:Compared with APD90 and TDR among three myocardial layers in the sham group, those in the MIR group were significant increased (P<0.01), which showed that transmural dispersion of repolarization increased significantly in MIR. Compared to those in the sham group, all three myocardial Cx43-pro significantly decreased (P<0.05) and△Cx43-pro obviously increased (P<0.05) in the MIR group which indicated transmural expression heterogeneity of Cx43 protein enlarged in MIR. 3. Cx43-Cq in all myocardium were significantly reduced (P<0.05) and△Cx43-Cq was significantly increased (P<0.05) in the MIR group compared to those in the sham group, which proved that transmural heterogeneity of Cx43 mRNA raised in MIR.Conclusions:Transmural heterogeneity of Cx43 expression among three myocardial layers in MIR significantly increased, which may enlarge dispersion of repolarization and prone to MVA.

  5. Heart Rate and Extracellular Sodium and Potassium Modulation of Gap Junction Mediated Conduction in Guinea Pigs.

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    Entz, Michael; George, Sharon A; Zeitz, Michael J; Raisch, Tristan; Smyth, James W; Poelzing, Steven

    2016-01-01

    Recent studies suggested that cardiac conduction in murine hearts with narrow perinexi and 50% reduced connexin43 (Cx43) expression is more sensitive to relatively physiological changes of extracellular potassium ([K(+)]o) and sodium ([Na(+)]o). Determine whether similar [K(+)]o and [Na(+)]o changes alter conduction velocity (CV) sensitivity to pharmacologic gap junction (GJ) uncoupling in guinea pigs. [K(+)]o and [Na(+)]o were varied in Langendorff perfused guinea pig ventricles (Solution A: [K(+)]o = 4.56 and [Na(+)]o = 153.3 mM. Solution B: [K(+)]o = 6.95 and [Na(+)]o = 145.5 mM). Gap junctions were inhibited with carbenoxolone (CBX) (15 and 30 μM). Epicardial CV was quantified by optical mapping. Perinexal width was measured with transmission electron microscopy. Total and phosphorylated Cx43 were evaluated by western blotting. Solution composition did not alter CV under control conditions or with 15μM CBX. Decreasing the basic cycle length (BCL) of pacing from 300 to 160 ms decreased CV uniformly with both solutions. At 30 μM CBX, a change in solution did not alter CV either longitudinally or transversely at BCL = 300 ms. However, reducing BCL to 160 ms caused CV to decrease more in hearts perfused with Solution B than A. Solution composition did not alter perinexal width, nor did it change total or phosphorylated serine 368 Cx43 expression. These data suggest that the solution dependent CV changes were independent of altered perinexal width or GJ coupling. Action potential duration was always shorter in hearts perfused with Solution B than A, independent of pacing rate and/or CBX concentration. Increased heart rate and GJ uncoupling can unmask small CV differences caused by changing [K(+)]o and [Na(+)]o. These data suggest that modulating extracellular ionic composition may be a novel anti-arrhythmic target in diseases with abnormal GJ coupling, particularly when heart rate cannot be controlled.

  6. Heart Rate and Extracellular Sodium and Potassium Modulation of Gap Junction Mediated Conduction in Guinea Pigs

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    Michael eEntz

    2016-02-01

    Full Text Available Background: Recent studies suggested that cardiac conduction in murine hearts with narrow perinexi and 50% reduced connexin43 (Cx43 expression is more sensitive to relatively physiological changes of extracellular potassium ([K+]o and sodium ([Na+]o. Purpose: Determine whether similar [K+]o and [Na+]o changes alter conduction velocity (CV sensitivity to pharmacologic gap junction (GJ uncoupling in guinea pigs.Methods: [K+]o and [Na+]o were varied in Langendorff perfused guinea pig ventricles (Solution A: [K+]o=4.56 and [Na+]o=153.3 mM. Solution B: [K+]o=6.95 and [Na+]o=145.5 mM. Gap junctions were inhibited with carbenoxolone (CBX (15 and 30 μM. Epicardial CV was quantified by optical mapping. Perinexal width was measured with transmission electron microscopy. Total and phosphorylated Cx43 were evaluated by western blotting. Results: Solution composition did not alter CV under control conditions or with 15M CBX. Decreasing the basic cycle length (BCL of pacing from 300 to 160ms decreased CV uniformly with both solutions. At 30 M CBX, a change in solution did not alter CV either longitudinally or transversely at BCL=300ms. However, reducing BCL to 160ms caused CV to decrease more in hearts perfused with Solution B than A. Solution composition did not alter perinexal width, nor did it change total or phosphorylated serine 368 Cx43 expression. These data suggest that the solution dependent CV changes were independent of altered perinexal width or GJ coupling. Action potential duration was always shorter in hearts perfused with Solution B than A, independent of pacing rate and/or CBX concentration. Conclusions: Increased heart rate and GJ uncoupling can unmask small CV differences caused by changing [K+]o and [Na+]o. These data suggest that modulating extracellular ionic composition may be a novel anti-arrhythmic target in diseases with abnormal GJ coupling, particularly when heart rate cannot be controlled.

  7. Improvement of cardiac function and reversal of gap junction remodeling by Neuregulin-1β in volume-overloaded rats with heart failure

    Institute of Scientific and Technical Information of China (English)

    Xue-Hui Wang; Xiao-Zhen Zhuo; Ya-Juan Ni; Min Gong; Ting-Zhong Wang; Qun Lu; Ai-Qun Ma

    2012-01-01

    Objective We performed experiments using Neuregulin-1β (NRG-1β) treatment to determine a mechanism for the protective role derived from its beneficial effects by remodeling gap junctions (GJs) during heart failure (HF). Methods Rat models of HF were established by aortocaval fistula. Forty-eight rats were divided randomly into the HF (HF, n = 16), NRG-1β treatment (NRG, n = 16), and sham operation (S, n = 16) group. The rats in the NRG group were administered NRG-1β (10 μg/kg per day) for 7 days via the tail vein, whereas the other groups were injected with the same doses of saline. Twelve weeks after operation, Connexin 43 (Cx43) expression in single myocytes obtained from the left ventricle was determined by immunocytochemistry. Total protein was extracted from frozen left ventricular tissues for immunoblotting assay, and the ultrastructure of myocytes was observed by transmission electron microscopy. Results Compared with the HF group, the cardiac function of rats in the NRG group was markedly improved, irregular distribution and deceased Cx43 expression were relieved. The ultrastructure of myocytes was seriously damaged in HF rats, and NRG-1β reduced these pathological damages. Conclusions Short-term NRG-1β treatment can rescue pump failure in experimental models of volume overload-induced HF, which is related to the recovery of GJs structure and the improvement of Cx43 expression.

  8. Selective permeability of gap junction channels.

    Science.gov (United States)

    Goldberg, Gary S; Valiunas, Virginijus; Brink, Peter R

    2004-03-23

    Gap junctions mediate the transfer of small cytoplasmic molecules between adjacent cells. A family of gap junction proteins exist that form channels with unique properties, and differ in their ability to mediate the transfer of specific molecules. Mutations in a number of individual gap junction proteins, called connexins, cause specific human diseases. Therefore, it is important to understand how gap junctions selectively move molecules between cells. Rules that dictate the ability of a molecule to travel through gap junction channels are complex. In addition to molecular weight and size, the ability of a solute to transverse these channels depends on its net charge, shape, and interactions with specific connexins that constitute gap junctions in particular cells. This review presents some data and interpretations pertaining to mechanisms that govern the differential transfer of signals through gap junction channels.

  9. Neural progenitor cells isolated from the subventricular zone present hemichannel activity and form functional gap junctions with glial cells

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    Rocío eTalaverón

    2015-10-01

    Full Text Available The postnatal subventricular zone lining the walls of the lateral ventricles contains neural progenitor cells (NPCs that generate new olfactory bulb interneurons. Communication via gap junctions between cells in the subventricular zone is involved in NPC proliferation and in neuroblast migration towards the olfactory bulb. Subventricular zone NPCs can be expanded in vitro in the form of neurospheres that can be used for transplantation purposes after brain injury. We have previously reported that neurosphere-derived NPCs form heterocellular gap junctions with host glial cells when they are implanted after mechanical injury. To analyze functionality of NPC-glial cell gap junctions we performed dye coupling experiments in co-cultures of subventricular zone NPCs with astrocytes or microglia. Neurosphere-derived cells expressed mRNA for at least the hemichannel/gap junction channel proteins connexin 26 (Cx26, Cx43, Cx45 and pannexin 1. Dye coupling experiments revealed that gap junctional communication occurred among neurosphere cells (incidence of coupling: 100%. Moreover, hemichannel activity was also detected in neurosphere cells as evaluated in time-lapse measurements of ethidium bromide uptake. Heterocellular coupling between NPCs and glial cells was evidenced in co-cultures of neurospheres with astrocytes (incidence of coupling: 91.0 ± 4.7% or with microglia (incidence of coupling: 71.9 ± 6.7%. Dye coupling in neurospheres and in co-cultures was inhibited by octanol, a gap junction blocker. Altogether, these results suggest the existence of functional hemichannels and gap junction channels in postnatal subventricular zone neurospheres. In addition, they demonstrate that subventricular zone-derived NPCs can establish functional gap junctions with astrocytes or microglia. Therefore, cell-cell communication via gap junctions and hemichannels with host glial cells might subserve a role in the functional integration of NPCs after implantation in

  10. Neural progenitor cells isolated from the subventricular zone present hemichannel activity and form functional gap junctions with glial cells.

    Science.gov (United States)

    Talaverón, Rocío; Fernández, Paola; Escamilla, Rosalba; Pastor, Angel M; Matarredona, Esperanza R; Sáez, Juan C

    2015-01-01

    The postnatal subventricular zone (SVZ) lining the walls of the lateral ventricles contains neural progenitor cells (NPCs) that generate new olfactory bulb interneurons. Communication via gap junctions between cells in the SVZ is involved in NPC proliferation and in neuroblast migration towards the olfactory bulb. SVZ NPCs can be expanded in vitro in the form of neurospheres that can be used for transplantation purposes after brain injury. We have previously reported that neurosphere-derived NPCs form heterocellular gap junctions with host glial cells when they are implanted after mechanical injury. To analyze functionality of NPC-glial cell gap junctions we performed dye coupling experiments in co-cultures of SVZ NPCs with astrocytes or microglia. Neurosphere-derived cells expressed mRNA for at least the hemichannel/gap junction channel proteins connexin 26 (Cx26), Cx43, Cx45 and pannexin 1 (Panx1). Dye coupling experiments revealed that gap junctional communication occurred among neurosphere cells (incidence of coupling: 100%). Moreover, hemichannel activity was also detected in neurosphere cells as evaluated in time-lapse measurements of ethidium bromide uptake. Heterocellular coupling between NPCs and glial cells was evidenced in co-cultures of neurospheres with astrocytes (incidence of coupling: 91.0 ± 4.7%) or with microglia (incidence of coupling: 71.9 ± 6.7%). Dye coupling in neurospheres and in co-cultures was inhibited by octanol, a gap junction blocker. Altogether, these results suggest the existence of functional hemichannels and gap junction channels in postnatal SVZ neurospheres. In addition, they demonstrate that SVZ-derived NPCs can establish functional gap junctions with astrocytes or microglia. Therefore, cell-cell communication via gap junctions and hemichannels with host glial cells might subserve a role in the functional integration of NPCs after implantation in the damaged brain.

  11. Neural progenitor cells isolated from the subventricular zone present hemichannel activity and form functional gap junctions with glial cells

    Science.gov (United States)

    Talaverón, Rocío; Fernández, Paola; Escamilla, Rosalba; Pastor, Angel M.; Matarredona, Esperanza R.; Sáez, Juan C.

    2015-01-01

    The postnatal subventricular zone (SVZ) lining the walls of the lateral ventricles contains neural progenitor cells (NPCs) that generate new olfactory bulb interneurons. Communication via gap junctions between cells in the SVZ is involved in NPC proliferation and in neuroblast migration towards the olfactory bulb. SVZ NPCs can be expanded in vitro in the form of neurospheres that can be used for transplantation purposes after brain injury. We have previously reported that neurosphere-derived NPCs form heterocellular gap junctions with host glial cells when they are implanted after mechanical injury. To analyze functionality of NPC-glial cell gap junctions we performed dye coupling experiments in co-cultures of SVZ NPCs with astrocytes or microglia. Neurosphere-derived cells expressed mRNA for at least the hemichannel/gap junction channel proteins connexin 26 (Cx26), Cx43, Cx45 and pannexin 1 (Panx1). Dye coupling experiments revealed that gap junctional communication occurred among neurosphere cells (incidence of coupling: 100%). Moreover, hemichannel activity was also detected in neurosphere cells as evaluated in time-lapse measurements of ethidium bromide uptake. Heterocellular coupling between NPCs and glial cells was evidenced in co-cultures of neurospheres with astrocytes (incidence of coupling: 91.0 ± 4.7%) or with microglia (incidence of coupling: 71.9 ± 6.7%). Dye coupling in neurospheres and in co-cultures was inhibited by octanol, a gap junction blocker. Altogether, these results suggest the existence of functional hemichannels and gap junction channels in postnatal SVZ neurospheres. In addition, they demonstrate that SVZ-derived NPCs can establish functional gap junctions with astrocytes or microglia. Therefore, cell-cell communication via gap junctions and hemichannels with host glial cells might subserve a role in the functional integration of NPCs after implantation in the damaged brain. PMID:26528139

  12. Communication of Ca(2+) signals via tunneling membrane nanotubes is mediated by transmission of inositol trisphosphate through gap junctions.

    Science.gov (United States)

    Lock, Jeffrey T; Parker, Ian; Smith, Ian F

    2016-10-01

    Tunneling membrane nanotubes (TNTs) are thin membrane projections linking cell bodies separated by many micrometers, which are proposed to mediate signaling and even transfer of cytosolic contents between distant cells. Several reports describe propagation of Ca(2+) signals between distant cells via TNTs, but the underlying mechanisms remain poorly understood. Utilizing a HeLa M-Sec cell line engineered to upregulate TNTs we replicated previous findings that mechanical stimulation elicits robust cytosolic Ca(2+) elevations that propagate to surrounding, physically separate cells. However, whereas this was previously interpreted to involve intercellular communication through TNTs, we found that Ca(2+) signal propagation was abolished - even in TNT-connected cells - after blocking ATP-mediated paracrine signaling with a cocktail of extracellular inhibitors. To then establish whether gap junctions may enable cell-cell signaling via TNTs under these conditions, we expressed sfGFP-tagged connexin-43 (Cx43) in HeLa M-Sec cells. We observed robust communication of mechanically-evoked Ca(2+) signals between distant but TNT-connected cells, but only when both cells expressed Cx43. Moreover, we also observed communication of Ca(2+) signals evoked in one cell by local photorelease of inositol 1,4,5-trisphosphate (IP3). Ca(2+) responses in connected cells began after long latencies at intracellular sites several microns from the TNT connection site, implicating intercellular transfer of IP3 and subsequent IP3-mediated Ca(2+) liberation, and not Ca(2+) itself, as the mediator between TNT-connected, Cx43-expressing cells. Our results emphasize the need to control for paracrine transmission in studies of cell-cell signaling via TNTs and indicate that, in this cell line, TNTs do not establish cytosolic continuity between connected cells but rather point to the crucial importance of connexins to enable communication of cytosolic Ca(2+) signals via TNTs.

  13. Persistent Borna Disease Virus infection changes expression and function of astroglial gap junctions in vivo and in vitro.

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    Köster-Patzlaff, Christiane; Hosseini, Seyed Mehdi; Reuss, Bernhard

    2007-12-12

    Neonatal Borna Disease Virus (BDV) infection of the Lewis rat brain leads to dentate gyrus (DG) degeneration, underlying mechanisms are not fully understood. Since astroglial gap junction (GJ) coupling is known to influence neurodegenerative processes, the question arose whether persistent BDV infection influences astroglial connexins (Cx) Cx43 and Cx30 in the hippocampal formation (HiF) of Lewis rats. RT-PCR and Western blot analysis of forebrain (FB) samples revealed a virus dependent reduction of both Cx types 8 but not 4 weeks post infection (p.i.). Immunohistochemistry revealed an increase of Cx43 in the DG and a decrease in the CA3 region 4 and 8 weeks p.i. Cx30, which was detectable only 8 weeks p.i., revealed a BDV dependent increase in DG and CA3 regions. BDV dependent astrogliosis as revealed by immunodetection of glial fibrillary acidic protein (GFAP) correlated not with astroglial connexin expression. With regard to functional coupling as revealed by scrape loading, BDV infection resulted in increased spreading of the GJ permeant dye Lucifer yellow in primary hippocampal astroglial cultures, and in increased expression of Cx43 and Cx30 as revealed by immunocytochemistry. In conclusion, persistent BDV infection of the Lewis rat brain leads to changes in astroglial Cx expression both in vivo and in vitro and of functional coupling in vitro. Distribution and time course of these changes suggest them to be a direct result of neurodegeneration in the DG and an indirect effect of neuronal deafferentiation in the CA3 region.

  14. Global ischemia-induced increases in the gap junctional proteins connexin 32 (Cx32) and Cx36 in hippocampus and enhanced vulnerability of Cx32 knock-out mice.

    Science.gov (United States)

    Oguro, K; Jover, T; Tanaka, H; Lin, Y; Kojima, T; Oguro, N; Grooms, S Y; Bennett, M V; Zukin, R S

    2001-10-01

    Gap junctions are conductive channels that connect the interiors of coupled cells. In the hippocampus, GABA-containing hippocampal interneurons are interconnected by gap junctions, which mediate electrical coupling and synchronous firing and thereby promote inhibitory transmission. The present study was undertaken to examine the hypothesis that the gap junctional proteins connexin 32 (Cx32; expressed by oligodendrocytes, interneurons, or both), Cx36 (expressed by interneurons), and Cx43 (expressed by astrocytes) play a role in defining cell-specific patterns of neuronal death in hippocampus after global ischemia in mice. Global ischemia did not significantly alter Cx32 and Cx36 mRNA expression and slightly increased Cx43 mRNA expression in the vulnerable CA1, as assessed by Northern blot analysis and in situ hybridization. Global ischemia induced a selective increase in Cx32 and Cx36 but not Cx43 protein abundance in CA1 before onset of neuronal death, as assessed by Western blot analysis. The increase in Cx32 and Cx36 expression was intense and specific to parvalbumin-positive inhibitory interneurons of CA1, as assessed by double immunofluorescence. Protein abundance was unchanged in CA3 and dentate gyrus. The finding of increase in connexin protein without increase in mRNA suggests regulation of Cx32 and Cx36 expression at the translational or post-translational level. Cx32(Y/-) null mice exhibited enhanced vulnerability to brief ischemic insults, consistent with a role for Cx32 gap junctions in neuronal survival. These findings suggest that Cx32 and Cx36 gap junctions may contribute to the survival and resistance of GABAergic interneurons, thereby defining cell-specific patterns of global ischemia-induced neuronal death.

  15. Neuroinflammation leads to region-dependent alterations in astrocyte gap junction communication and hemichannel activity.

    Science.gov (United States)

    Karpuk, Nikolay; Burkovetskaya, Maria; Fritz, Teresa; Angle, Amanda; Kielian, Tammy

    2011-01-12

    Inflammation attenuates gap junction (GJ) communication in cultured astrocytes. Here we used a well-characterized model of experimental brain abscess as a tool to query effects of the CNS inflammatory milieu on astrocyte GJ communication and electrophysiological properties. Whole-cell patch-clamp recordings were performed on green fluorescent protein (GFP)-positive astrocytes in acute brain slices from glial fibrillary acidic protein-GFP mice at 3 or 7 d after Staphylococcus aureus infection in the striatum. Astrocyte GJ communication was significantly attenuated in regions immediately surrounding the abscess margins and progressively increased to levels typical of uninfected brain with increasing distance from the abscess proper. Conversely, astrocytes bordering the abscess demonstrated hemichannel activity as evident by enhanced ethidium bromide (EtBr) uptake that could be blocked by several pharmacological inhibitors, including the connexin 43 (Cx43) mimetic peptide Gap26, carbenoxolone, the pannexin1 (Panx1) mimetic peptide (10)Panx1, and probenecid. However, hemichannel opening was transient with astrocytic EtBr uptake observed near the abscess at day 3 but not day 7 after infection. The region-dependent pattern of hemichannel activity at day 3 directly correlated with increases in Cx43, Cx30, Panx1, and glutamate transporter expression (glial L-glutamate transporter and L-glutamate/L-aspartate transporter) along the abscess margins. Changes in astrocyte resting membrane potential and input conductance correlated with the observed changes in GJ communication and hemichannel activity. Collectively, these findings indicate that astrocyte coupling and electrical properties are most dramatically affected near the primary inflammatory site and reveal an opposing relationship between the open states of GJ channels versus hemichannels during acute infection. This relationship may extend to other CNS diseases typified with an inflammatory component.

  16. Expression of Cx43-related microRNAs in patients with tetralogy of Fallot

    Institute of Scientific and Technical Information of China (English)

    Yao Wu; Xiao-Jing Ma; Hui-Jun Wang; Wen-Can Li; Long Chen; Duan Ma; Guo-Ying Huang

    2014-01-01

    Background: Abnormal expression of connexin 43 (Cx43) has been reported to play an important role in the development of conotrunccal anomalies. However, less is known about the underlying reason for its abnormal expression. MicroRNAs (miRNAs), as an important part of gene expression regulation, have been implicated in some cardiac diseases. This study aimed to investigate the expression of Cx43 and its related miRNAs in patients with tetralogy of Fallot (TOF), and illustrate the potential role of abnormal miRNAs regulation to Cx43 expression in the pathology of TOF. Methods: Real-time polymerase chain reaction was used to detect the expression of Cx43 and 10 Cx43-related miRNAs in the myocardium from 30 TOF patients and 10 normal controls. Immunohistochemistry was used to detect Cx43 protein expression. Putative miRNA binding sites in the 3'UTR of Cx43 were examined in 200 TOF patients and 200 healthy individuals, using Sanger sequencing, to exclude sequence variations resulting in binding diffi culties of miRNAs. Results: Cx43 mRNA and protein expression in the myocardium tissue was significantly increased in TOF patients. The expression of MiR-1 and 206 was significantly decreased in the TOF patients as compared with the controls (P0.05). No meaningful sequence variation was detected in the putative miR1/206 binding sites in the 3'UTR of Cx43. Conclusions: This study indicated that miR-1 and 206 is down-regulated in TOF patients, which may cause an up-regulation of Cx43 protein's synthesis. It provided a clue that miR-1 and 206 might be involved in the pathogenesis of TOF, additional experiments are needed to determine if in fact, miR-1 and 206 contribute substantially to TOF.

  17. Gap junctions in the inner ear: comparison of distribution patterns in different vertebrates and assessement of connexin composition in mammals.

    Science.gov (United States)

    Forge, Andrew; Becker, David; Casalotti, Stefano; Edwards, Jill; Marziano, Nerissa; Nevill, Graham

    2003-12-08

    The distribution and size of gap junctions (GJ) in the sensory epithelia of the inner ear have been examined in a reptile (gecko), birds (chicken and owl), and mammals (mouse, guinea pig, gerbil, and bat), and the connexin composition of GJs in the mammalian inner ear has been assessed. Freeze fracture revealed a common pattern of GJ distribution in auditory and vestibular sensory epithelia in the different vertebrate classes. In all these tissues, GJs are numerous, often occupying more than 25% of the plasma membrane area of supporting cells and sometimes composed of more than 100,000 channels. Screening for 12 members of the connexin family in the mammalian inner ear by RT-PCR, Western blotting, and immunohistochemistry revealed four connexin isotypes, cx26, cx30, cx31, and cx43, in the cochlea and three, cx26, cx30, and cx43, in the vestibular organs. With antibodies characterised for their specificity, cx26 and cx30 colocalised in supporting cells of the organ of Corti, in the basal cell region of the stria vascularis, and in type 1 fibrocytes of the spiral ligament. No other connexin was detected in these regions. Cx31 was localised among type 2 fibrocytes below the spiral prominence, a region where cx30 was not expressed and cx26 expression appeared to be low. Cx43 was detected only in the region of "tension fibrocytes" lining the inner aspect of the otic capsule. This suggests separate functional compartments in the cochlea. In addition to cx26 and cx30, cx43 was detected in supporting cells of the vestibular sensory epithelia. Where cx26 and cx30 were colocalised, double immunogold labelling of thin sections showed both cx26 and cx30 evenly distributed in individual GJ plaques, a pattern consistent with the presence of heteromeric connexons. Coimmunoprecipitation of cochlear membrane proteins solubilised with a procedure that preserves the oligomeric structure of connexons confirmed the presence of heteromeric cx26/cx30 connexons. Heteromeric cx26/cx30

  18. Gap junctions and connexin-interacting proteins

    NARCIS (Netherlands)

    Giepmans, Ben N G

    2004-01-01

    Gap junctions form channels between adjacent cells. The core proteins of these channels are the connexins. Regulation of gap junction communication (GJC) can be modulated by connexin-associating proteins, such as regulatory protein phosphatases and protein kinases, of which c-Src is the best-studied

  19. Gap junctions and connexin-interacting proteins

    NARCIS (Netherlands)

    Giepmans, Ben N G

    2004-01-01

    Gap junctions form channels between adjacent cells. The core proteins of these channels are the connexins. Regulation of gap junction communication (GJC) can be modulated by connexin-associating proteins, such as regulatory protein phosphatases and protein kinases, of which c-Src is the

  20. Gap junctions and connexin-interacting proteins

    NARCIS (Netherlands)

    Giepmans, Ben N G

    2004-01-01

    Gap junctions form channels between adjacent cells. The core proteins of these channels are the connexins. Regulation of gap junction communication (GJC) can be modulated by connexin-associating proteins, such as regulatory protein phosphatases and protein kinases, of which c-Src is the best-studied

  1. Connexin 30 expression and frequency of connexin heterogeneity in astrocyte gap junction plaques increase with age in the rat retina.

    Directory of Open Access Journals (Sweden)

    Hussein Mansour

    Full Text Available We investigated age-associated changes in retinal astrocyte connexins (Cx by assaying Cx numbers, plaque sizes, protein expression levels and heterogeneity of gap junctions utilizing six-marker immunohistochemistry (IHC. We compared Wistar rat retinal wholemounts in animals aged 3 (young adult, 9 (middle-aged and 22 months (aged. We determined that retinal astrocytes have gap junctions composed of Cx26, -30, -43 and -45. Cx30 was consistently elevated at 22 months compared to younger ages both when associated with parenchymal astrocytes and vascular-associated astrocytes. Not only was the absolute number of Cx30 plaques significantly higher (P<0.05 but the size of the plaques was significantly larger at 22 months compared to younger ages (p<0.05. With age, Cx26 increased significantly initially, but returned to basal levels; whereas Cx43 expression remained low and stable with age. Evidence that astrocytes alter connexin compositions of gap junctions was demonstrated by the significant increase in the number of Cx26/Cx45 gap junctions with age. We also found gap junctions comprised of 1, 2, 3 or 4 Cx proteins suggesting that retinal astrocytes use various connexin protein combinations in their gap junctions during development and aging. These data provides new insight into the dynamic and extensive Cx network utilized by retinal astrocytes for communication within both the parenchyma and vasculature for the maintenance of normal retinal physiology with age. This characterisation of the changes in astrocytic gap junctional communication with age in the CNS is crucial to the understanding of physiological aging and age-related neurodegenerative diseases.

  2. Effect of calcium channel blocker on gap junctional connexin 43 in infarcted myocardium in rats%钙通道阻滞剂对梗死心肌中不同部位心肌连接蛋白43表达的影响

    Institute of Scientific and Technical Information of China (English)

    杨永健; 速晓华; 唐兵; 李德; 杨大春

    2011-01-01

    目的:研究L和L/T型钙通道阻制剂对梗死心脏不同部位(坏死区域、肥厚区域等)心肌组织中连接蛋白43(Cx43)的影响.方法:随机将大鼠48只分为假手术组、心肌梗死(MI)组、阿莫地平(L型钙通道阻滞剂)组和米贝拉地尔(L/T型钙通道阻滞剂)组(每组n=12只).通过结扎大鼠左冠状动脉建立MI模型,术前7 d,上述4个组分别用安慰剂、L型钙通道阻滞剂阿莫地平4 mg/(kg·d)和L/T型钙通道阻滞剂米贝拉地尔10 mg/(kg·d).术后1、3、7 d,分别检测左心室游离壁(LVFW,梗死区)、心室间隔(IS,肥厚区)和右心室壁(Rv),正常心肌组织中Cx43蛋白的表达.术后7 d显微直视下测LVFW处MI病灶的大小、IS的厚度及左心室的大小.结果:IS中Cx43蛋白表达于术后1、3、7 d呈逐渐增加的趋势;LVFW中Cx43蛋白的表达于术后1、3、7 d时均处于低水平,与对照组相比差异显著(P<0.05).RV中Cx43蛋白的表达于术后1、3、7 d无显著差异,与对照组相比也无显著性差异.米贝拉地尔能明显地抑制LVFW心肌组织中Cx43表达的下调,缩小MI病灶;阿莫地平则抑制肥厚心肌中Cx43蛋白的表达,明显抑制IS的肥厚.结论:MI病理过程中,梗死病灶内Cx43的表达下调,肥厚组织中Cx43的表达上调.L和L/T型钙通道阻滞剂均能减轻心肌重构与选择性地调节心肌组织中Cx43的表达有关.%AIM: To investigate the effect of cardiac L- and L/T-type Ca2+ channels on gap junctional connexin 43 ( Cx43 ) in myocardium infarcted heart remodeling of rats. METHODS: Rat myocardium infarction model was established by permanent ligation of the left coronary artery. Infarcted rats were treated with oral placebo, amlodipine [ L-channel blockade, 4 mg/( kg · day) ] or mibefradil [ L/T-channel blockade, 10 mg/(kg·day] beginning 7 days before induction of myocardial infarction (MI). Protein levels of Cx43 were measured 1, 3 and 7 days postcoronary occlusion in the noninfarcted and

  3. Loss of cadherin-binding proteins β-catenin and plakoglobin in the heart leads to gap junction remodeling and arrhythmogenesis.

    Science.gov (United States)

    Swope, David; Cheng, Lan; Gao, Erhe; Li, Jifen; Radice, Glenn L

    2012-03-01

    Arrhythmic right ventricular cardiomyopathy (ARVC) is a hereditary heart muscle disease that causes sudden cardiac death (SCD) in young people. Almost half of ARVC patients have a mutation in genes encoding cell adhesion proteins of the desmosome, including plakoglobin (JUP). We previously reported that cardiac tissue-specific plakoglobin (PG) knockout (PG CKO) mice have no apparent conduction abnormality and survive longer than expected. Importantly, the PG homolog, β-catenin (CTNNB1), showed increased association with the gap junction protein connexin43 (Cx43) in PG CKO hearts. To determine whether β-catenin is required to maintain cardiac conduction in the absence of PG, we generated mice lacking both PG and β-catenin specifically in the heart (i.e., double knockout [DKO]). The DKO mice exhibited cardiomyopathy, fibrous tissue replacement, and conduction abnormalities resulting in SCD. Loss of the cadherin linker proteins resulted in dissolution of the intercalated disc (ICD) structure. Moreover, Cx43-containing gap junction plaques were reduced at the ICD, consistent with the arrhythmogenicity of the DKO hearts. Finally, ambulatory electrocardiogram monitoring captured the abrupt onset of spontaneous lethal ventricular arrhythmia in the DKO mice. In conclusion, these studies demonstrate that the N-cadherin-binding partners, PG and β-catenin, are indispensable for maintaining mechanoelectrical coupling in the heart.

  4. Modulatory effects of cAMP and PKC activation on gap junctional intercellular communication among thymic epithelial cells

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    Neves-dos-Santos Sandra

    2010-01-01

    Full Text Available Abstract Background We investigated the effects of the signaling molecules, cyclic AMP (cAMP and protein-kinase C (PKC, on gap junctional intercellular communication (GJIC between thymic epithelial cells (TEC. Results Treatment with 8-Br-cAMP, a cAMP analog; or forskolin, which stimulates cAMP production, resulted in an increase in dye transfer between adjacent TEC, inducing a three-fold enhancement in the mean fluorescence of coupled cells, ascertained by flow cytometry after calcein transfer. These treatments also increased Cx43 mRNA expression, and stimulated Cx43 protein accumulation in regions of intercellular contacts. VIP, adenosine, and epinephrine which may also signal through cyclic nucleotides were tested. The first two molecules did not mimic the effects of 8-Br-cAMP, however epinephrine was able to increase GJIC suggesting that this molecule functions as an endogenous inter-TEC GJIC modulators. Stimulation of PKC by phorbol-myristate-acetate inhibited inter-TEC GJIC. Importantly, both the enhancing and the decreasing effects, respectively induced by cAMP and PKC, were observed in both mouse and human TEC preparations. Lastly, experiments using mouse thymocyte/TEC heterocellular co-cultures suggested that the presence of thymocytes does not affect the degree of inter-TEC GJIC. Conclusions Overall, our data indicate that cAMP and PKC intracellular pathways are involved in the homeostatic control of the gap junction-mediated communication in the thymic epithelium, exerting respectively a positive and negative role upon cell coupling. This control is phylogenetically conserved in the thymus, since it was seen in both mouse and human TEC preparations. Lastly, our work provides new clues for a better understanding of how the thymic epithelial network can work as a physiological syncytium.

  5. Autophagy and gap junctional intercellular communication inhibition are involved in cadmium-induced apoptosis in rat liver cells

    Energy Technology Data Exchange (ETDEWEB)

    Zou, Hui [College of Veterinary Medicine, Yangzhou University, and Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou, Jiangsu, 225009 (China); Zhuo, Liling [College of Life Science, Zaozhuang University, Zaozhuang, Shandong, 277160 (China); Han, Tao; Hu, Di; Yang, Xiaokang; Wang, Yi; Yuan, Yan; Gu, Jianhong; Bian, Jianchun; Liu, Xuezhong [College of Veterinary Medicine, Yangzhou University, and Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou, Jiangsu, 225009 (China); Liu, Zongping, E-mail: liuzongping@yzu.edu.cn [College of Veterinary Medicine, Yangzhou University, and Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou, Jiangsu, 225009 (China)

    2015-04-17

    Cadmium (Cd) is known to induce hepatotoxicity, yet the underlying mechanism of how this occurs is not fully understood. In this study, Cd-induced apoptosis was demonstrated in rat liver cells (BRL 3A) with apoptotic nuclear morphological changes and a decrease in cell index (CI) in a time- and concentration-dependent manner. The role of gap junctional intercellular communication (GJIC) and autophagy in Cd-induced apoptosis was investigated. Cd significantly induced GJIC inhibition as well as downregulation of connexin 43 (Cx43). The prototypical gap junction blocker carbenoxolone disodium (CBX) exacerbated the Cd-induced decrease in CI. Cd treatment was also found to cause autophagy, with an increase in mRNA expression of autophagy-related genes Atg-5, Atg-7, Beclin-1, and microtubule-associated protein light chain 3 (LC3) conversion from cytosolic LC3-I to membrane-bound LC3-II. The autophagic inducer rapamycin (RAP) prevented the Cd-induced CI decrease, while the autophagic inhibitor chloroquine (CQ) caused a further reduction in CI. In addition, CBX promoted Cd-induced autophagy, as well as changes in expression of Atg-5, Atg-7, Beclin-1 and LC3. CQ was found to block the Cd-induced decrease in Cx43 and GJIC inhibition, whereas RAP had opposite effect. These results demonstrate that autophagy plays a protective role during Cd-induced apoptosis in BRL 3A cells during 6 h of experiment, while autophagy exacerbates Cd-induced GJIC inhibition which has a negative effect on cellular fate. - Highlights: • GJIC and autophagy is crucial for biological processes. • Cd exposure causes GJIC inhibition and autophagy increase in BRL 3A cells. • Autophagy protects Cd induced BRL 3A cells apoptosis at an early stage. • Autophagy exacerbates Cd-induced GJIC inhibition. • GJIC plays an important role in autophagy induced cell death or survival.

  6. Gap junction communication in myelinating glia.

    Science.gov (United States)

    Nualart-Marti, Anna; Solsona, Carles; Fields, R Douglas

    2013-01-01

    Gap junction communication is crucial for myelination and axonal survival in both the peripheral nervous system (PNS) and central nervous system (CNS). This review examines the different types of gap junctions in myelinating glia of the PNS and CNS (Schwann cells and oligodendrocytes respectively), including their functions and involvement in neurological disorders. Gap junctions mediate intercellular communication among Schwann cells in the PNS, and among oligodendrocytes and between oligodendrocytes and astrocytes in the CNS. Reflexive gap junctions mediating transfer between different regions of the same cell promote communication between cellular compartments of myelinating glia that are separated by layers of compact myelin. Gap junctions in myelinating glia regulate physiological processes such as cell growth, proliferation, calcium signaling, and participate in extracellular signaling via release of neurotransmitters from hemijunctions. In the CNS, gap junctions form a glial network between oligodendrocytes and astrocytes. This transcellular communication is hypothesized to maintain homeostasis by facilitating restoration of membrane potential after axonal activity via electrical coupling and the re-distribution of potassium ions released from axons. The generation of transgenic mice for different subsets of connexins has revealed the contribution of different connexins in gap junction formation and illuminated new subcellular mechanisms underlying demyelination and cognitive defects. Alterations in metabolic coupling have been reported in animal models of X-linked Charcot-Marie-Tooth disease (CMTX) and Pelizaeus-Merzbarcher-like disease (PMLD), which are caused by mutations in the genes encoding for connexin 32 and connexin 47 respectively. Future research identifying the expression and regulation of gap junctions in myelinating glia is likely to provide a better understanding of myelinating glia in nervous system function, plasticity, and disease. This

  7. Increasing gap junctional coupling: a tool for dissecting the role of gap junctions.

    Science.gov (United States)

    Axelsen, Lene Nygaard; Haugan, Ketil; Stahlhut, Martin; Kjølbye, Anne-Louise; Hennan, James K; Holstein-Rathlou, Niels-Henrik; Petersen, Jørgen Søberg; Nielsen, Morten Schak

    2007-03-01

    Much of our current knowledge about the physiological and pathophysiological role of gap junctions is based on experiments where coupling has been reduced by either chemical agents or genetic modification. This has brought evidence that gap junctions are important in many physiological processes. In a number of cases, gap junctions have been implicated in the initiation and progress of disease, and experimental uncoupling has been used to investigate the exact role of coupling. The inverse approach, i.e., to increase coupling, has become possible in recent years and represents a new way of testing the role of gap junctions. The aim of this review is to summarize the current knowledge obtained with agents that selectively increase gap junctional intercellular coupling. Two approaches will be reviewed: increasing coupling by the use of antiarrhythmic peptide and its synthetic analogs and by interfering with the gating of gap junctional channels.

  8. Gap junction intercellular communication and benzene toxicity.

    Science.gov (United States)

    Rivedal, Edgar; Witz, Gisela; Leithe, Edward

    2010-03-19

    Aberrant regulation of gap junction intercellular communication (GJIC) has been linked to several human diseases, including cancer and abnormal hematopoietic development. Benzene exposure has been shown to cause hematotoxicity and leukemia, but the underlying mechanisms involved remain unclear. We have observed that several metabolites of benzene have the ability to block gap junction intercellular communication. The ring-opened trans,trans-muconaldehyde (MUC) was found to be the most potent inhibitor of gap junction channels. MUC was found to induce cross-linking of the gap junction protein connexin43, which seemed to be responsible for the induced inhibition of GJIC. Glutaraldehyde, which has a similar molecular structure as MUC, was found to possess similar effects on gap junctions as MUC, while the mono-aldehyde formaldehyde shows lower potency, both as a connexin cross-linker, and as an inhibitor of GJIC. Both glutaraldehyde and formaldehyde have previously been associated with induction of leukemia and disturbance of hematopoiesis. Taken together, the data support a possible link between the effect of MUC on gap junctions, and the toxic effects of benzene. Copyright (c) 2009 Elsevier Ireland Ltd. All rights reserved.

  9. Gap junctions: structure and function (Review).

    Science.gov (United States)

    Evans, W Howard; Martin, Patricia E M

    2002-01-01

    Gap junctions are plasma membrane spatial microdomains constructed of assemblies of channel proteins called connexins in vertebrates and innexins in invertebrates. The channels provide direct intercellular communication pathways allowing rapid exchange of ions and metabolites up to approximately 1 kD in size. Approximately 20 connexins are identified in the human or mouse genome, and orthologues are increasingly characterized in other vertebrates. Most cell types express multiple connexin isoforms, making likely the construction of a spectrum of heteromeric hemichannels and heterotypic gap junctions that could provide a structural basis for the charge and size selectivity of these intercellular channels. The precise nature of the potential signalling information traversing junctions in physiologically defined situations remains elusive, but extensive progress has been made in elucidating how connexins are assembled into gap junctions. Also, participation of gap junction hemichannels in the propagation of calcium waves via an extracellular purinergic pathway is emerging. Connexin mutations have been identified in a number of genetically inherited channel communication-opathies. These are detected in connexin 32 in Charcot Marie Tooth-X linked disease, in connexins 26 and 30 in deafness and skin diseases, and in connexins 46 and 50 in hereditary cataracts. Biochemical approaches indicate that many of the mutated connexins are mistargeted to gap junctions and/or fail to oligomerize correctly into hemichannels. Genetic ablation approaches are helping to map out a connexin code and point to specific connexins being required for cell growth and differentiation as well as underwriting basic intercellular communication.

  10. The relation of Cx43 and NMDA to visceral sensitization in rats with irritable bowel syndrome

    Directory of Open Access Journals (Sweden)

    Jing-yu ZHANG

    2016-01-01

    Full Text Available Objective  To study the relationship between connexin 43 (Cx43 and N-methyl-D-aspartate (NMDA receptors and visceral sensitization in the rats with irritable bowel syndrome (IBS. Methods  Thirty rats were gavaged with Triehinella spiralis to reproduce the IBS model. These rats were randomly divided into IBS group, IBS+colon distension group, and IBS+STI-571+colon distension group, and other groups of normal rats were randomized into normal group and normal+colon distension group, with 10 rats in each group. Immunofluorescent double staining were used to observe the expressions of intestine Cx43 and sacral NMDA re ceptors of rats in all the groups. Results  The Cx43 and sacral NMDA expressions in the normal group, normal+colon distension group and IBS group showed no significant changes (P>0.05, however, Cx43 and sacral NMDA expressions were significantly higher in IBS rats with colon distension as compared with those in normal group, normal+colon distension group, and IBS group (P<0.05, while they were significantly lower in the IBS+STI-571+colon distension group after STI-571 intervention (P<0.05. Conclusion  Cx43 and sacral NMDA may be the most important factor of visceral sensitization in IBS rats. DOI: 10.11855/j.issn.0577-7402.2015.12.02

  11. Increasing gap junctional coupling: a tool for dissecting the role of gap junctions

    DEFF Research Database (Denmark)

    Axelsen, Lene Nygaard; Haugan, Ketil; Stahlhut, Martin;

    2007-01-01

    . In a number of cases, gap junctions have been implicated in the initiation and progress of disease, and experimental uncoupling has been used to investigate the exact role of coupling. The inverse approach, i.e., to increase coupling, has become possible in recent years and represents a new way of testing......Much of our current knowledge about the physiological and pathophysiological role of gap junctions is based on experiments where coupling has been reduced by either chemical agents or genetic modification. This has brought evidence that gap junctions are important in many physiological processes...... the role of gap junctions. The aim of this review is to summarize the current knowledge obtained with agents that selectively increase gap junctional intercellular coupling. Two approaches will be reviewed: increasing coupling by the use of antiarrhythmic peptide and its synthetic analogs...

  12. Gap junction diseases of the skin.

    Science.gov (United States)

    van Steensel, M A M

    2004-11-15

    Gap junctions are intercellular channels that allow the passage of water, ions, and small molecules. They are involved in quick, short-range messaging between cells and are found in skin, nervous tissue, heart, and muscle. An increasing number of hereditary skin disorders appear to be caused by mutations in one of the genes coding for the constituent proteins of gap junctions, known as connexins. In this review, the currently known connexin disorders that feature skin abnormalities are described: keratitis-ichthyosis deafness syndrome, erythrokeratoderma variabilis, Vohwinkel's syndrome, and a novel disorder called hypotrichosis-deafness syndrome. What is known about the pathogenesis of these disorders is discussed and related to gap junction physiology. (c) 2004 Wiley-Liss, Inc.

  13. Regulation of Hemichannels and Gap Junction Channels by Cytokines in Antigen-Presenting Cells

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    Pablo J. Sáez

    2014-01-01

    Full Text Available Autocrine and paracrine signals coordinate responses of several cell types of the immune system that provide efficient protection against different challenges. Antigen-presenting cells (APCs coordinate activation of this system via homocellular and heterocellular interactions. Cytokines constitute chemical intercellular signals among immune cells and might promote pro- or anti-inflammatory effects. During the last two decades, two membrane pathways for intercellular communication have been demonstrated in cells of the immune system. They are called hemichannels (HCs and gap junction channels (GJCs and provide new insights into the mechanisms of the orchestrated response of immune cells. GJCs and HCs are permeable to ions and small molecules, including signaling molecules. The direct intercellular transfer between contacting cells can be mediated by GJCs, whereas the release to or uptake from the extracellular milieu can be mediated by HCs. GJCs and HCs can be constituted by two protein families: connexins (Cxs or pannexins (Panxs, which are present in almost all APCs, being Cx43 and Panx1 the most ubiquitous members of each protein family. In this review, we focus on the effects of different cytokines on the intercellular communication mediated by HCs and GJCs in APCs and their impact on purinergic signaling.

  14. Neuroprotection in the treatment of glaucoma--A focus on connexin43 gap junction channel blockers.

    Science.gov (United States)

    Chen, Ying-Shan; Green, Colin R; Danesh-Meyer, Helen V; Rupenthal, Ilva D

    2015-09-01

    Glaucoma is a form of optic neuropathy and a common cause of blindness, affecting over 60 million people worldwide with an expected rise to 80 million by 2020. Successful treatment is challenging due to the various causes of glaucoma, undetectable symptoms at an early stage and inefficient delivery of drugs to the back of the eye. Conventional glaucoma treatments focus on the reduction of elevated intraocular pressure (IOP) using topical eye drops. However, their efficacy is limited to patients who suffer from high IOP glaucoma and do not address the underlying susceptibility of retinal ganglion cells (RGC) to degeneration. Glaucoma is known as a neurodegenerative disease which starts with RGC death and eventually results in damage of the optic nerve. Neuroprotective strategies therefore offer a novel treatment option for glaucoma by not only preventing neuronal loss but also disease progression. This review firstly gives an overview of the pathophysiology of glaucoma as well as current treatment options including conventional and novel delivery strategies. It then summarizes the rational for neuroprotection as a novel therapy for glaucomatous neuropathies and reviews current potential neuroprotective strategies to preserve RGC, with a focus on connexin43 (Cx43) gap junction channel blockers.

  15. ATP Is Required and Advances Cytokine-Induced Gap Junction Formation in Microglia In Vitro

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    Pablo J. Sáez

    2013-01-01

    Full Text Available Microglia are the immune cells in the central nervous system. After injury microglia release bioactive molecules, including cytokines and ATP, which modify the functional state of hemichannels (HCs and gap junction channels (GJCs, affecting the intercellular communication via extracellular and intracellular compartments, respectively. Here, we studied the role of extracellular ATP and several cytokines as modulators of the functional state of microglial HCs and GJCs using dye uptake and dye coupling techniques, respectively. In microglia and the microglia cell line EOC20, ATP advanced the TNF-α/IFN-γ-induced dye coupling, probably through the induction of IL-1β release. Moreover, TNF-α/IFN-γ, but not TNF-α plus ATP, increased dye uptake in EOC20 cells. Blockade of Cx43 and Panx1 HCs prevented dye coupling induced by TNF-α/IFN-γ, but not TNF-α plus ATP. In addition, IL-6 prevented the induction of dye coupling and HC activity induced by TNF-α/IFN-γ in EOC20 cells. Our data support the notion that extracellular ATP affects the cellular communication between microglia through autocrine and paracrine mechanisms, which might affect the timing of immune response under neuroinflammatory conditions.

  16. Red paprika (Capsicum annuum L.) and its main carotenoids, capsanthin and β-carotene, prevent hydrogen peroxide-induced inhibition of gap-junction intercellular communication.

    Science.gov (United States)

    Kim, Ji-Sun; Lee, Woo-Moon; Rhee, Han Cheol; Kim, Suna

    2016-07-25

    This study was conducted to investigate the protective effect of red paprika extract (RPE) and its main carotenoids, namely, capsanthin (CST) and β-carotene (BCT), on the H2O2-induced inhibition of gap-junction intercellular communication (GJIC) in WB-F344 rat liver epithelial cells (WB cells). We found that pre-treatment with RPE, CST and BCT protected WB cells from H2O2-induced inhibition of GJIC. RPE, CST and BCT not only recovered connexin 43 (Cx43) mRNA expression but also prevented phosphorylation of Cx43 protein by H2O2 treatment. RPE attenuated the phosphorylation of ERK, p38 and JNK, whereas pre-treatment with CST and BCT only attenuated the phosphorylation of ERK and p38 and did not affect JNK in H2O2-treated WB cells. RPE, CST and BCT significantly suppressed the formation of reactive oxygen species (ROS) in H2O2-treated cells compared to untreated WB cells. These results suggest that dietary intake of red paprika might be helpful for lowering the risk of diseases caused by oxidative stress.

  17. Antiproliferative Action of Conjugated Linoleic Acid on Human MCF-7 Breast Cancer Cells Mediated by Enhancement of Gap Junctional Intercellular Communication through Inactivation of NF-κB

    Directory of Open Access Journals (Sweden)

    Md. Abdur Rakib

    2013-01-01

    Full Text Available The major conjugated linoleic acid (CLA isomers, c9,t11-CLA and t10,c12-CLA, have anticancer effects; however, the exact mechanisms underlying these effects are unknown. Evidence suggests that reversal of reduced gap junctional intercellular communication (GJIC in cancer cells inhibits cell growth and induces cell death. Hence, we determined that CLA isomers enhance GJIC in human MCF-7 breast cancer cells and investigated the underlying molecular mechanisms. The CLA isomers significantly enhanced GJIC of MCF-7 cells at 40 μM concentration, whereas CLA inhibited cell growth and induced caspase-dependent apoptosis. CLA increased connexin43 (Cx43 expression both at the transcriptional and translational levels. CLA inhibited nuclear factor-κB (NF-κB activity and enhanced reactive oxygen species (ROS generation. No significant difference was observed in the efficacy of c9,t11-CLA and t10,c12-CLA. These results suggest that the anticancer effect of CLA is associated with upregulation of GJIC mediated by enhanced Cx43 expression through inactivation of NF-κB and generation of ROS in MCF-7 cells.

  18. Gap junction diseases of the skin.

    NARCIS (Netherlands)

    Steensel, M.A.M. van

    2004-01-01

    Gap junctions are intercellular channels that allow the passage of water, ions, and small molecules. They are involved in quick, short-range messaging between cells and are found in skin, nervous tissue, heart, and muscle. An increasing number of hereditary skin disorders appear to be caused by muta

  19. Gap junction diseases of the skin.

    NARCIS (Netherlands)

    Steensel, M.A.M. van

    2004-01-01

    Gap junctions are intercellular channels that allow the passage of water, ions, and small molecules. They are involved in quick, short-range messaging between cells and are found in skin, nervous tissue, heart, and muscle. An increasing number of hereditary skin disorders appear to be caused by muta

  20. Reduced heterogeneous expression of Cx43 results in decreased Nav1.5 expression and reduced sodium current that accounts for arrhythmia vulnerability in conditional Cx43 knockout mice.

    NARCIS (Netherlands)

    Jansen, J.; Noorman, M.; Musa, H.; Stein, M.; Jong, S. de; Nagel, R. van der; Hund, T.J.; Mohler, P.J.; Vos, M.E. de; Veen, T.A. van; Bakker, J.M. de; Delmar, M.; Rijen, H.V. van

    2012-01-01

    BACKGROUND: Reduced expression of connexin43 (Cx43) and sodium channel (Nav1.5) and increased expression of collagen (fibrosis) are important determinants of impulse conduction in the heart. OBJECTIVE: To study the importance and interaction of these factors at very low Cx43 expression, inducible Cx

  1. Nonspecific effects of the gap junction blocker mefloquine on fast hippocampal network oscillations in the adult rat in vitro.

    Science.gov (United States)

    Behrens, C J; Ul Haq, R; Liotta, A; Anderson, M L; Heinemann, U

    2011-09-29

    It has been suggested that gap junctions are involved in the synchronization during high frequency oscillations as observed during sharp wave-ripple complexes (SPW-Rs) and during recurrent epileptiform discharges (REDs). Ripple oscillations during SPW-Rs, possibly involved in memory replay and memory consolidation, reach frequencies of up to 200 Hz while ripple oscillations during REDs display frequencies up to 500 Hz. These fast oscillations may be synchronized by intercellular interactions through gap junctions. In area CA3, connexin 36 (Cx36) proteins are present and potentially sensitive to mefloquine. Here, we used hippocampal slices of adult rats to investigate the effects of mefloquine, which blocks Cx36, Cx43 and Cx50 gap junctions on both SPW-Rs and REDs. SPW-Rs were induced by high frequency stimulation in the CA3 region while REDs were recorded in the presence of the GABA(A) receptor blocker bicuculline (5 μM). Both, SPW-Rs and REDs were blocked by the gap junction blocker carbenoxolone. Mefloquine (50 μM), which did not affect stimulus-induced responses in area CA3, neither changed SPW-Rs nor superimposed ripple oscillations. During REDs, 25 and 50 μM mefloquine exerted only minor effects on the expression of REDs but significantly reduced the amplitude of superimposed ripples by ∼17 and ∼54%, respectively. Intracellular recordings of CA3 pyramidal cells revealed that mefloquine did not change their resting membrane potential and input resistance but significantly increased the afterhyperpolarization following evoked action potentials (APs) resulting in reduced probability of AP firing during depolarizing current injection. Similarly, mefloquine caused a reduction in AP generation during REDs. Together, our data suggest that mefloquine depressed RED-related ripple oscillations by reducing high frequency discharges and not necessarily by blocking electrical coupling.

  2. BRMS1 and Cx43 expression in fine needle aspiration thyroid cancer tissue and their correlation with tumor malignancy

    Institute of Scientific and Technical Information of China (English)

    Jian-Guo Sheng; Bin Wang; Zong-Ping Diao; Kun-Kun Cao; Sai Zhang; Zheng-Guo Pu

    2016-01-01

    Objective:To study the BRMS1 and Cx43 expression in fine needle aspiration thyroid cancer tissue and their correlation with tumor malignancy.Methods:Patients undergoing thyroid fine needle aspiration biopsy in our hospital from April 2012 to October 2015 were selected for study, 60 patients with thyroid cancer and 60 patients with benign thyroid tumor were screened after pathological diagnosis, biopsy tissue was collected to determine the expression of BRMS1 and Cx43, and serum specimens were collected to determin Gal-3, CEACAM1, MMP2 and MMP9 content.Results: mRNA levels and positive expression rate of BRMS1 andCx43in thyroid cancer tissue were significantly lower than those in benign thyroid tumor tissue; mRNA levels ofBRMS1andCx43in thyroid cancer tissue with different pathological types and tumor diameters were not different, mRNA level ofCx43in thyroid cancer tissue with TNM III-IV stage was significantly lower than that in thyroid cancer tissue with TNM I-II stage, mRNA levels ofBRMS1 in thyroid cancer tissue with different TNM stages were not different, and mRNA levels ofBRMS1andCx43in thyroid carcinoma tissue with lymph node metastasis were significantly lower than those in thyroid carcinoma tissue without lymph node metastasis; serum Gal-3, CEACAM1, MMP2 and MMP9 levels in patients with positive BRMS1 and Cx43 expression in thyroid cancer tissue were significantly lower than those in patients with negative BRMS1 and Cx43 expression in thyroid cancer tissue.Conclusions:Lower expression of BRMS1 and Cx43 in fine needle aspiration thyroid cancer tissue is associated with the distant metastasis and malignant degree of tumor, and lower expression of Cx43 is also associated with the growth of tumor and cancer cell proliferation.

  3. Gap junction channels and cardiac impulse propagation.

    Science.gov (United States)

    Desplantez, Thomas; Dupont, Emmanuel; Severs, Nicholas J; Weingart, Robert

    2007-08-01

    The role of gap junction channels on cardiac impulse propagation is complex. This review focuses on the differential expression of connexins in the heart and the biophysical properties of gap junction channels under normal and disease conditions. Structural determinants of impulse propagation have been gained from biochemical and immunocytochemical studies performed on tissue extracts and intact cardiac tissue. These have defined the distinctive connexin coexpression patterns and relative levels in different cardiac tissues. Functional determinants of impulse propagation have emerged from electrophysiological experiments carried out on cell pairs. The static properties (channel number and conductance) limit the current flow between adjacent cardiomyocytes and thus set the basic conduction velocity. The dynamic properties (voltage-sensitive gating and kinetics of channels) are responsible for a modulation of the conduction velocity during propagated action potentials. The effect is moderate and depends on the type of Cx and channel. For homomeric-homotypic channels, the influence is small to medium; for homomeric-heterotypic channels, it is medium to strong. Since no data are currently available on heteromeric channels, their influence on impulse propagation is speculative. The modulation by gap junction channels is most prominent in tissues at the boundaries between cardiac tissues such as sinoatrial node-atrial muscle, atrioventricular node-His bundle, His bundle-bundle branch and Purkinje fibers-ventricular muscle. The data predict facilitation of orthodromic propagation.

  4. Gap Junctions: The Claymore for Cancerous Cells

    Directory of Open Access Journals (Sweden)

    Ailar Nakhlband

    2011-07-01

    Full Text Available Introduction: Gap junctions play an important role in the cell proliferation in mammalian cells as well as carcinogenesis. However, there are controversial issues about their role in cancer pathogenesis. This study was designed to evaluate genotoxicity and cytotoxicity of Carbenoxolone (CBX as a prototype of inter-cellular gap junction blocker in MCF7 and BT20 human breast cancer cells. Methods: The MCF7and BT20 human breast cancer cell lines were cultivated, and treated at designated confluency with different doses of CBX. Cellular cytotoxicity was examined using standard colorimetric assay associated with cell viability tests. Gene expression evaluation was carried out using real time polymerase chain reaction (PCR. Results: MCF7 and BT20 cells were significantly affected by CBX in a dose dependent manner in cell viability assays. Despite varying expression of genes, down regulation of pro- and anti-apoptotic genes was observed in these cells. Conclusion: Based upon this investigation, it can be concluded that CBX could affect both low and high proliferative types of breast cancer cell lines and disproportionate down regulation of both pre- and anti-apoptotic genes may be related to interacting biomolecules, perhaps via gap junctions.

  5. Fisetin increases the cytotoxicity of cisplatin by enhancing gap junctional intercellular communication%漆黄素通过缝隙连接影响顺铂细胞毒性

    Institute of Scientific and Technical Information of China (English)

    杨克凡; 范丽霞; 彭悦霞; 王琴; 陶亮

    2015-01-01

    Aim To investigate the effect of fisetin on the cytotoxicity of cisplatin and its mechanisms. Meth-ods CCK-8 was used to assay the toxicity of fisetin.“Parachute” dye-coupling assay was used to examine dye spread through gap junctions in U87 cells. “Stand-ard colony-forming assay” was used to examine cell survivals of U87 treated with cisplatin with fisetin. Western blot was performed to detect the expression level of Cx43 . Results Fisetin had no cytotoxic effect on U87 cells at the concentration of 0~1μmol · L-1 . Parachute assay demonstrated that fisetin enhanced dye-coupling through gap juntions in U87 cells. Stand-ard Colony forming assay showed that cisplatin inhibi- ted the clonogenic survivals of U87 cells and fisetin could enhance cisplatin-induced inhibitory effect of clo-nogenic survivals on cells with gap junctional formation more than on cells without gap junctional formation. Conclusion These rusults demonstrate that fisetin could increase the sensitivity of glioma cells to cisplatin through the enhancement of gap junctional intercellular communication but this may not be related to the ex-pression of Cx43 .%目的:体外观察漆黄素对顺铂细胞毒性作用影响及其机制。方法 CCK-8法观察不同浓度漆黄素对人胶质瘤U87细胞的毒性;用细胞接种荧光示踪法测定不同浓度漆黄素对U87细胞缝隙连接( GJ )功能的影响;标准细胞集落形成分析法观察顺铂的毒性及漆黄素对顺铂毒性的影响;用Western blot法研究漆黄素在影响GJIC( GJ intercellular com-munication)功能浓度范围内对 Cx43表达的影响。结果CCK-8法显示漆黄素在小于1μmol·L-1的浓度范围内无细胞毒性;细胞接种荧光示踪法显示漆黄素浓度越高, U87细胞GJ通讯的荧光传递功能越强;标准细胞集落形成分析法显示,20μmol·L-1顺铂能够抑制U87细胞的集落形成,而且在有GJ形成的细胞顺铂对细胞集落形成的抑制作用显著

  6. Tanshinone IIA increases the bystander effect of herpes simplex virus thymidine kinase/ganciclovir gene therapy via enhanced gap junctional intercellular communication.

    Science.gov (United States)

    Xiao, Jianyong; Zhang, Guangxian; Qiu, Pengxiang; Liu, Xijuan; Wu, Yingya; Du, Biaoyan; Li, Jiefen; Zhou, Jing; Li, Jingjing; Tan, Yuhui

    2013-01-01

    The bystander effect is an intriguing phenomenon by which adjacent cells become sensitized to drug treatment during gene therapy with herpes simplex virus thymidine kinase/ganciclovir (HSV-tk/GCV). This effect is reported to be mediated by gap junctional intercellular communication (GJIC), and therefore, we postulated that upregulation of genes that facilitate GJIC may enhance the HSV-tk/GCV bystander effect. Previous findings have shown Tanshinone IIA (Tan IIA), a chemical substance derived from a Chinese medicine herb, promotes the upregulation of the connexins Cx26 and Cx43 in B16 cells. Because gap junctions are formed by connexins, we hypothesized that Tan IIA might increase GJIC. Our results show that Tan IIA increased GJIC in B16 melanoma cells, leading to more efficient GCV-induced bystander killing in cells stably expressing HSV-tk. Additionally, in vivo experiments demonstrated that tumors in mice with 10% HSV-tk positive B16 cells and 90% wild-type B16 cells became smaller following treatment with the combination of GCV and Tan IIA as compared to GCV or Tan IIA alone. These data demonstrate that Tan IIA can augment the bystander effect of HSV-tk/GCV system through increased gap junction coupling, which adds strength to the promising strategy that develops connexins inducer to potentiate the effects of suicide gene therapy.

  7. Tanshinone IIA increases the bystander effect of herpes simplex virus thymidine kinase/ganciclovir gene therapy via enhanced gap junctional intercellular communication.

    Directory of Open Access Journals (Sweden)

    Jianyong Xiao

    Full Text Available The bystander effect is an intriguing phenomenon by which adjacent cells become sensitized to drug treatment during gene therapy with herpes simplex virus thymidine kinase/ganciclovir (HSV-tk/GCV. This effect is reported to be mediated by gap junctional intercellular communication (GJIC, and therefore, we postulated that upregulation of genes that facilitate GJIC may enhance the HSV-tk/GCV bystander effect. Previous findings have shown Tanshinone IIA (Tan IIA, a chemical substance derived from a Chinese medicine herb, promotes the upregulation of the connexins Cx26 and Cx43 in B16 cells. Because gap junctions are formed by connexins, we hypothesized that Tan IIA might increase GJIC. Our results show that Tan IIA increased GJIC in B16 melanoma cells, leading to more efficient GCV-induced bystander killing in cells stably expressing HSV-tk. Additionally, in vivo experiments demonstrated that tumors in mice with 10% HSV-tk positive B16 cells and 90% wild-type B16 cells became smaller following treatment with the combination of GCV and Tan IIA as compared to GCV or Tan IIA alone. These data demonstrate that Tan IIA can augment the bystander effect of HSV-tk/GCV system through increased gap junction coupling, which adds strength to the promising strategy that develops connexins inducer to potentiate the effects of suicide gene therapy.

  8. Dephosphorylation of cardiomyocyte Cx43 is associated with myocardial ischemia and reperfusion injury

    Institute of Scientific and Technical Information of China (English)

    Zhijuan Cao; Xuan Xu; Linli Que; Qi Chen; Yuehua Li

    2009-01-01

    Objective:Myocardial ischemia/reperfusion(I/R) injury is the leading cause of death in the world. However, the details of the mechanism of its pathophysioiogy are still unknown. The present study was designed to investigate the role of connexin 43(Cx63) in acute models of myocardial I/R injury. Methods: Male C57BL/6 mice were subjected to myocardial ischemia(45 rain) followed by reperfusion(4 hrs) in vivo. The whole operation was monitored using a two-lead ECG. Hearts were harvested and the level of protein was assessed by western blot analysis. Haematoxylin and Eosin(HE) staining was used to detect the extent of neutrophil infiltration. The expression level of IL-6 was detected by ELISA. Results: A murine myocardial I/R injury model was constructed successfully. Phosphorylated Cx43 decreased 83.45% while non-phosphorylated Cx43 increased 1.62- fold in the myocardium after I/R injury. Neutrophil infiltration and the expression of the inflammatory cytokine IL-6 increased in the myocardium following I/R. Conclusion: During myocardial I/R injury, cardiomyocyte Cx43 is dephosphorylated, and this may be associated with an inflammatory response.

  9. Gap junctional communication in osteocytes is amplified by low intensity vibrations in vitro.

    Directory of Open Access Journals (Sweden)

    Gunes Uzer

    Full Text Available The physical mechanism by which cells sense high-frequency mechanical signals of small magnitude is unknown. During exposure to vibrations, cell populations within a bone are subjected not only to acceleratory motions but also to fluid shear as a result of fluid-cell interactions. We explored displacements of the cell nucleus during exposure to vibrations with a finite element (FE model and tested in vitro whether vibrations can affect osteocyte communication independent of fluid shear. Osteocyte like MLO-Y4 cells were subjected to vibrations at acceleration magnitudes of 0.15 g and 1 g and frequencies of 30 Hz and 100 Hz. Gap junctional intracellular communication (GJIC in response to these four individual vibration regimes was investigated. The FE model demonstrated that vibration induced dynamic accelerations caused larger relative nuclear displacement than fluid shear. Across the four regimes, vibrations significantly increased GJIC between osteocytes by 25%. Enhanced GJIC was independent of vibration induced fluid shear; there were no differences in GJIC between the four different vibration regimes even though differences in fluid shear generated by the four regimes varied 23-fold. Vibration induced increases in GJIC were not associated with altered connexin 43 (Cx43 mRNA or protein levels, but were dependent on Akt activation. Combined, the in silico and in vitro experiments suggest that externally applied vibrations caused nuclear motions and that large differences in fluid shear did not influence nuclear motion (<1% or GJIC, perhaps indicating that vibration induced nuclear motions may directly increase GJIC. Whether the increase in GJIC is instrumental in modulating anabolic and anti-catabolic processes associated with the application of vibrations remains to be determined.

  10. Gap junctions in the nervous system.

    Science.gov (United States)

    Rozental, R; Giaume, C; Spray, D C

    2000-04-01

    Synapses are classically defined as close connections between two nerve cells or between a neuronal cell and a muscle or gland cell across which a chemical signal (i.e., a neurotransmitter) and/or an electrical signal (i.e., current-carrying ions) can pass. The definition of synapse was developed by Charles Sherrington and by Ramon y Cajal at the beginning of this century and refined by John Eccles and Bernard Katz 50 years later; in this collection of papers, the definition of synapses is discussed further in the chapter by Mike Bennett. who provided the first functional demonstration of electrical transmission via gap junction channels between vertebrate neurons. As is evidenced by the range of topics covered in this issue, research dealing with gap junctions in the nervous system has expanded enormously in the past decade, major findings being that specific cell types in the brain expresses specific types of connexins and that expression patterns coincide with tissue compartmentalization and function and that these compartments change during development.

  11. Cx43和Pax3在人胚胎食管肌层组织中的表达及意义%Expression and Significance of Cx43 and Pax3 in the Esophagus of a Human Embryo

    Institute of Scientific and Technical Information of China (English)

    张剑; 刘学红

    2009-01-01

    应用免疫组织化学SABC法检测第2,3,4三个月胎龄段,Cx43和Pax3蛋白在人胚胎食管组织中的表达.结果表明:第2个月胚龄时,Cx43和Pax3在食管肌层肌细胞中呈阳性表达,在肌间神经丛处神经细胞呈阴性表达;第3个月胎龄段,Cx43和Pax3在食管肌层肌细胞阳性表达与第2个月胚龄相似,在肌间神经丛处大部分神经细胞呈阳性表达;第4个月胎龄段,Cx43和Pax3在食管肌层神经细胞和肌细胞均呈弱阳性表达.因此,Cx43和Pax3蛋白与人胚胎早期食管肌层组织细胞的生长发育关系密切.%The objective of the paper is to explore the patterns of Connexin43 (Cx43) and Pairedbox3 (Pax3) protein expressions in the esophageal muscular layers of a human embryo.The SABC method in Immunohistochemistry has been employed to examine the expressions of Cx43 and Pax3 proteins in the muscular layers of the esophagus in human embryos from the second to fourth months of gestation.The following are the results:In the second month of gestation,their muscle cells of the esophageal muscular layers are positive while the nerve cells in the myenteric plexus are negative;In the third month,the Cx43 and Pax3 expressions in the muscle cells in the esophageal muscular layers are similar to those of the second month,with most nerve cells positive in the myenteric plexus;In the fourth month,the Cx43 and Pax3 protein expressions are weakly positive both in the muscle cells and nerve cells of the esophageal muscular layers.A conclusion is reached:Their Cx43 and Pax3 proteins are closely related to the growth and development of the cells and tissues in the esophageal muscular layers in human embryos.

  12. The Expressions of Survivin, PTEN and Cx43 Gene in Skin Basal Cell Carcinoma%Survivin、PTEN和Cx43基因在皮肤基底细胞癌中的表达

    Institute of Scientific and Technical Information of China (English)

    杜金锋; 郭瑞珍

    2009-01-01

    背景与目的:探讨Survivin、PTEN和Cx43基因与皮肤基底细胞癌(basal cell carcinoma,BCC)的相关性.材料与方法:采用免疫组织化学S-P法分别检测35例皮肤BCC、10例正常皮肤组织中Survivin、PTEN和Cx43蛋白的表达;运用核酸分子原位杂交技术检测Cx43 mRNA的表达.应用图像分析系统采集分析图像,分别检测BCC和正常皮肤组织中各项检测指标的表达水平(阳性面积)和表达强度(光密度).结果:Survivin蛋白在正常皮肤中呈阴性表达,在BCC中呈弱阳性表达,其差异有统计学意义(P<0.05):PTEN蛋白在正常皮肤组织中呈强阳性表达,在BCC中呈阳性或弱阳性表达,其差异具有统计学意义(P<0.05).而Cx43蛋白和Cx43 mRNA在正常皮肤组织中的表达均呈强阳性,而在BCC组织中均呈阳性或弱阳性,差异均具有统计学意义(P均<0.01).相关性分析显示,BCC组织中,PTEN蛋白的表达与Cx43蛋白呈正相关(r=0.519,P<0.01),Cx43蛋白的表达与Cx43 mRNA也呈正相关(r=0.732,P<0.01). 结论:PTEN蛋白、Cx43蛋白和Cx43 mRNA在BCC中的低表达;Survivin蛋白在BCC中的高表达,可能在BCC发生发展的过程中发挥重要作用.

  13. Supercritical CO(2)-extracted tomato Oleoresins enhance gap junction intercellular communications and recover from mercury chloride inhibition in keratinocytes.

    Science.gov (United States)

    Leone, Antonella; Zefferino, Roberto; Longo, Cristiano; Leo, Lucia; Zacheo, Giuseppe

    2010-04-28

    A nutritionally relevant phytochemical such as lycopene, found in tomatoes and other fruits, has been proposed to have health-promoting effects by modulating hormonal and immune systems, metabolic pathways, and gap junction intercellular communication (GJIC). This work analyzes lycopene extracts, obtained from tomato and tomato added with grape seeds by using a safe and environmentally friendly extraction process, based on supercritical carbon dioxide technology (S-CO(2)). Analysis of the innovative S-CO(2)-extracted oleoresins showed peculiar chemical composition with high lycopene concentration and the presence of other carotenoids, lipids, and phenol compounds. The oleoresins showed a higher in vitro antioxidant activity compared with pure lycopene and beta-carotene and the remarkable ability to enhance the GJIC and to increase cx43 expression in keratinocytes. The oleoresins, (0.9 microM lycopene), were also able to overcome, completely, the GJIC inhibition induced by 10 nM HgCl(2), mercury(II) chloride, suggesting a possible action mechanism.

  14. Structure, regulation and function of gap junctions in liver

    Science.gov (United States)

    Maes, Michaël; Decrock, Elke; Wang, Nan; Leybaert, Luc; da Silva, Tereza Cristina; Veloso Alves Pereira, Isabel; Jaeschke, Hartmut; Cogliati, Bruno; Vinken, Mathieu

    2016-01-01

    Gap junctions are a specialized group of cell-to-cell junctions that mediate direct intercellular communication between cells. They arise from the interaction of 2 hemichannels of adjacent cells, which in turn are composed of 6 connexin proteins. In liver, gap junctions are predominantly found in hepatocytes and play critical roles in virtually all phases of the hepatic life cycle, including cell growth, differentiation, liver-specific functionality and cell death. Liver gap junctions are directed through a broad variety of mechanisms ranging from epigenetic control of connexin expression to posttranslational regulation of gap junction activity. This paper reviews established and novel aspects regarding the architecture, control and functional relevance of liver gap junctions. PMID:27001459

  15. Ascorbic acid 6-palmitate suppresses gap-junctional intercellular communication through phosphorylation of connexin 43 via activation of the MEK-ERK pathway.

    Science.gov (United States)

    Lee, Kyung Mi; Kwon, Jung Yeon; Lee, Ki Won; Lee, Hyong Joo

    2009-01-15

    Although the health benefits of dietary antioxidants have been extensively studied, their potential negative effects remain unclear. L-Ascorbic acid 6-palmitate (AAP), a synthetic derivative of ascorbic acid (AA), is widely used as an antioxidant and preservative in foods, vitamins, drugs, and cosmetics. Previously, we found that AA exerted an antitumor effect by protecting inhibition of gap-junctional intercellular communication (GJIC), which is closely associated with tumor progression. In this study, we examined whether AAP, an amphipathic derivative of AA, has chemopreventive effects using a GJIC model. AAP and AA exhibited dose-dependent free radical-scavenging activities and inhibited hydrogen peroxide (H(2)O(2))-induced intracellular reactive oxygen species (ROS) production in normal rat liver epithelial cells. Unexpectedly, however, AAP did not protect against the inhibition of GJIC induced by H(2)O(2); instead, it inhibited GJIC synergistically with H(2)O(2). AAP inhibited GJIC in a dose-dependent and reversible manner. This inhibitory effect was not due to the conjugated lipid structure of AAP, as treatment with palmitic acid alone failed to inhibit GJIC under the same conditions. The inhibition of GJIC by AAP was restored in the presence of mitogen-activated protein kinase/extracellular signal-regulated kinase (ERK) kinase (MEK) inhibitor U0126, but not in the presence of other signal inhibitors and antioxidant (PKC inhibitors, EGFR inhibitor, NADPH oxidase inhibitor, catalase, vitamin E, or AA), indicating the critical involvement of MEK signaling in the GJIC inhibitory activity of AAP. Phosphorylation of ERK and connexin 43 (Cx43) was observed following AAP treatment, and this was reversed by U0126. These results suggest that the AAP-induced inhibition of GJIC is mediated by the phosphorylation of Cx43 via activation of the MEK-ERK pathway. Taken together, our results indicate that AAP has a potent carcinogenic effect, and that the influence of dietary

  16. Glial connexins and gap junctions in CNS inflammation and disease.

    Science.gov (United States)

    Kielian, Tammy

    2008-08-01

    Gap junctions facilitate direct cytoplasmic communication between neighboring cells, facilitating the transfer of small molecular weight molecules involved in cell signaling and metabolism. Gap junction channels are formed by the joining of two hemichannels from adjacent cells, each composed of six oligomeric protein subunits called connexins. Of paramount importance to CNS homeostasis are astrocyte networks formed by gap junctions, which play a critical role in maintaining the homeostatic regulation of extracellular pH, K+, and glutamate levels. Inflammation is a hallmark of several diseases afflicting the CNS. Within the past several years, the number of publications reporting effects of cytokines and pathogenic stimuli on glial gap junction communication has increased dramatically. The purpose of this review is to discuss recent observations characterizing the consequences of inflammatory stimuli on homocellular gap junction coupling in astrocytes and microglia as well as changes in connexin expression during various CNS inflammatory conditions.

  17. Role of the gap junctions in the contractile response to agonists in pulmonary artery from two rat models of pulmonary hypertension

    Directory of Open Access Journals (Sweden)

    Dahan Diana

    2011-03-01

    Full Text Available Background Pulmonary hypertension (PH is characterized by arterial vascular remodelling and alteration in vascular reactivity. Since gap junctions are formed with proteins named connexins (Cx and contribute to vasoreactivity, we investigated both expression and role of Cx in the pulmonary arterial vasoreactivity in two rat models of PH. Methods Intrapulmonary arteries (IPA were isolated from normoxic rats (N, rats exposed to chronic hypoxia (CH or treated with monocrotaline (MCT. RT-PCR, Western Blot and immunofluorescent labelling were used to study the Cx expression. The role of Cx in arterial reactivity was assessed by using isometric contraction and specific gap junction blockers. Contractile responses were induced by agonists already known to be involved in PH, namely serotonin, endothelin-1 and phenylephrine. Results Cx 37, 40 and 43 were expressed in all rat models and Cx43 was increased in CH rats. In IPA from N rats only, the contraction to serotonin was decreased after treatment with 37-43Gap27, a specific Cx-mimetic peptide blocker of Cx 37 and 43. The contraction to endothelin-1 was unchanged after incubation with 40Gap27 (a specific blocker of Cx 40 or 37-43Gap27 in N, CH and MCT rats. In contrast, the contraction to phenylephrine was decreased by 40Gap27 or 37-43Gap27 in CH and MCT rats. Moreover, the contractile sensitivity to high potassium solutions was increased in CH rats and this hypersensitivity was reversed following 37-43Gap27 incubation. Conclusion Altogether, Cx 37, 40 and 43 are differently expressed and involved in the vasoreactivity to various stimuli in IPA from different rat models. These data may help to understand alterations of pulmonary arterial reactivity observed in PH and to improve the development of innovative therapies according to PH aetiology.

  18. Infection by Cx43 adenovirus increased chemotherapy sensitivity in human gastric cancer BGC-823 cells: not involving in induction of cell apoptosis.

    Science.gov (United States)

    Liu, Dan; Zhou, Hongfeng; Wu, Jin; Liu, Wentao; Li, Yongqing; Shi, Guangyue; Yue, Xiaolong; Sun, Xiwen; Zhao, Yanbin; Hu, Xiaowei; Wang, Tianjiao; Zhang, Xufeng

    2015-12-15

    There is a lower basal expression of Connexin43 (Cx43) in human gastric cancer BGC-823 cells. In the present study, BGC-823 cells were transfected with recombinant Cx43 adenovirus plasmid vector, and we explored the influences of Cx43 expression on cell proliferation, chemo-sensitivity, colony forming ability, invasion ability and apoptosis. Moreover, we also determined the expression of Pgp, Cx43, as well as apoptosis-related proteins (bcl-2, bax, caspase3 and caspase 9). MTT assay was performed to determine the proliferation of BGC-823 cells before and after Cx43 transfection. The influences of Cx43 infection on sensitivity of chemotherapy (including Doxorubicin, fluorouracil, oxaliplatin) were detected by MTT assay. Expression levels of Pgp, Cx43, as well as apoptosis-related proteins (bcl-2, bax, caspase-3 and caspase-9) in BGC-823 cells were determined by Western blotting analysis before and after the infection with Cx43 adenovirus. MDR expression was determined by RT-PCR before and after Cx43 infection. Invasive ability was detected by invasion chamber. Influence of Cx43 adenovirus infection on apoptosis of BGC-823 cells was determined by flow cytometry. After infection by Cx43 adenovirus, colony forming rate and invasive ability of BGC-823 cells were decreased. Flow cytometry results revealed that cell apoptosis were insignificantly increased. The data of MTT assay revealed that infection with Cx43 adenovirus, cell proliferation ability decreased and sensitivity to chemotherapy drugs (including doxorubicin, fluorouracil, oxaliplatin) increased. Results of Western blotting analysis revealed that increasing expression levels of Cx43, decreasing expression levels of Pgp, and insignificant changes of bcl-2, bax, caspase3 and caspase 9 were detected. RT-PCR revealed the expression of MDR1 gene, the gene encoding Pgp, decreased significantly (pinfected with Cx43-IRES2-EGFP recombinant adenovirus vector. Colony formation, invasive ability and cell proliferation all

  19. Gap Junctions and Cancer: Communicating for 50 Years’

    Science.gov (United States)

    Aasen, Trond; Mesnil, Marc; Naus, Christian C.; Lampe, Paul D.; Laird, Dale W.

    2017-01-01

    Fifty years ago, tumour cells were found to lack electrical coupling, leading to the hypothesis that loss of direct intercellular communication is commonly associated with cancer onset and progression. Subsequent studies linked this phenomenon to gap junctions composed of connexin proteins. While many studies support the notion that connexins are tumour suppressors, recent evidence suggests that, in some tumour types, they may facilitate specific stages of tumour progression through both junctional and non-junctional signalling pathways. This Timeline article highlights the milestones connecting gap junctions to cancer, and underscores important unanswered questions, controversies and therapeutic opportunities in the field. PMID:27782134

  20. O tabagismo está associado com a remodelação de junções comunicantes no coração de ratos: explicação do paradoxo dos fumantes? Smoking is associated with remodeling of gap junction in the rat heart: smoker's paradox explanation?

    Directory of Open Access Journals (Sweden)

    Rosangela Novo

    2013-03-01

    reperfusion. OBJECTIVE: Thus, this study aimed to analyze the effects of exposure to tobacco smoke on intensity, distribution or phosphorylation of connexin 43 in the rat heart. METHODS: Wistar rats weighing 100 g were randomly allocated into 2 groups: 1 Control (n = 25; 2 Exposed to tobacco smoke (ETS, n = 23. After 5 weeks, left ventricular morphometric analysis, immunohisthochemistry and western blotting for connexin 43 (Cx43 were performed. RESULTS: Collagen volume fraction, cross-sectional areas, and ventricular weight were not statistically different between control and ETS. ETS showed lower stain intensity of Cx43 at intercalated disks (Control: 2.32 ± 0.19; ETS: 1.73 ± 0.18; p = 0.04. The distribution of CX43 at intercalated disks did not differ between the groups (Control: 3.73 ± 0.12; ETS: 3.20 ± 0.17; p = 0.18. ETS rats showed higher levels of dephosphorylated form of Cx43 (Control: 0.45 ± 0.11; ETS: 0.90 ± 0.11; p = 0.03. On the other hand, total Cx43 did not differ between control and ETS groups (Control: 0.75 ± 0.19; ETS: 0.93 ± 0.27; p = 0.58. CONCLUSION: Exposure to tobacco smoke resulted in cardiac gap junction remodeling, characterized by alterations in the quantity and phosphorylation of the Cx43, in rats hearts. This finding could explain the smoker's paradox observed in some studies.

  1. C-kit、SCF、Cx43在先天性巨结肠中的表达及作用%Expression and correlation study of C-kit, SCF and Cx43 in hirschsprung's disease

    Institute of Scientific and Technical Information of China (English)

    谢丽微; 夏丽娜; 赵志光

    2013-01-01

    Objective:To explore the action of C-kit,SCF and Cx43 in the pathogenesis of HD by examining the expression and distribution of C-kit,SCF and Cx43.Methods:The colon specimens of 68 cases of HD and 8 cases of accidental death without digestive tract malformations (control) were collected.The case group composed of a expansion group in which the ganglion cells were normal in the intestinal wall and a narrow group in which the ganglion cells were absent in the intestinal wall according to diagnosis by HE.Immunohistochemical staining was used to detect the proteins of C-Kit,SCF and Cx43,and hybridization in situ to detect the mRNA of C-Kit in all specimens.Results:Expression of C-kit,SCF and Cx43 in the expansion group was not significantly different from the control group (P > 0.05),while expression of C-kit,SCF and Cx43 in narrow group decreased significantly compared with the control group (P < 0.05).Conclusion:The development of Hirschsprung's disease is associated with the decreased expression of C-kit,SCF and Cx43.%目的:探讨酪氨酸激酶膜受体基因(C-kit)、干细胞因子(SCF)与缝隙连接蛋白(Cx43)在先天性巨结肠发病机制中的作用.方法:收集68例先天性巨结肠(HD)患儿手术切除标本的存档蜡块,根据HE染色结果,将神经节细胞正常的扩张段标本设为扩张组,无神经节细胞的狭窄段标本设为狭窄组.并收集8例无消化道畸形新生儿尸检结肠组织作为对照.用免疫组织化学EliVision法检测C-kit、SCF及Cx43蛋白的表达,原位杂交法检测 C-kit mRNA的表达.结果:与对照组比较,HD扩张组中C-kit、SCF、Cx43的表达差异无统计学意义(P>0.05),狭窄组中C-kit、SCF、Cx43的表达差异有统计学意义(P<0.05).结论:先天性巨结肠的发生与C-kit、SCF及Cx43表达减少有关.

  2. Gap junction modulation and its implications for heart function.

    Science.gov (United States)

    Kurtenbach, Stefan; Kurtenbach, Sarah; Zoidl, Georg

    2014-01-01

    Gap junction communication (GJC) mediated by connexins is critical for heart function. To gain insight into the causal relationship of molecular mechanisms of disease pathology, it is important to understand which mechanisms contribute to impairment of gap junctional communication. Here, we present an update on the known modulators of connexins, including various interaction partners, kinases, and signaling cascades. This gap junction network (GJN) can serve as a blueprint for data mining approaches exploring the growing number of publicly available data sets from experimental and clinical studies.

  3. Gap junction modulation and its implications for heart function

    Directory of Open Access Journals (Sweden)

    Stefan eKurtenbach

    2014-02-01

    Full Text Available Gap junction communication mediated by connexins is critical for heart function. To gain insight into the causal relationship of molecular mechanisms of disease pathology, it is important to understand which mechanisms contribute to impairment of gap junctional communication. Here, we present an update on the known modulators of connexins, including various interaction partners, kinases and signaling cascades. This gap junction network can serve as a blueprint for data mining approaches exploring the growing number of publicly available data sets from experimental and clinical studies.

  4. Gap junction disorders of myelinating cells.

    Science.gov (United States)

    Kleopa, Kleopas A; Orthmann-Murphy, Jennifer; Sargiannidou, Irene

    2010-01-01

    Gap junctions (GJs) are channels that allow the diffusion of ions and small molecules across apposed cell membranes. In peripheral nerves, Schwann cells express the GJ proteins connexin32 (Cx32) and Cx29, which have distinct localizations. Cx32 forms GJs through non-compact myelin areas, whereas Cx29 forms hemichannels in the innermost layers of myelin apposing axonal Shaker-type K+ channels. In the CNS, rodent oligodendrocytes express Cx47, Cx32 and Cx29. Cx47 is expressed by all types of oligodendrocytes both in the white and grey matter and forms GJs on cell bodies and proximal processes, as well as most of the intercellular channels with astrocytes. Cx32 is expressed mostly by white matter oligodendrocytes and is localized in the myelin sheath of large diameter fibers. Cx29, and its human ortholog Cx31.3, appear to be restricted to oligodendrocytes that myelinate small caliber fibers, likely forming hemichannels. The importance of intercellular and intracellular GJs in myelinating cells are demonstrated by human disorders resulting from mutations affecting GJ proteins. The X-linked Charcot Marie Tooth disease (CMT1X) is caused by hundreds of mutations affecting Cx32. Patients with CMT1X present mainly with a progressive peripheral neuropathy, which may be accompanied by CNS myelin dysfunction. Mutations in Cx47 may cause a devastating leukodystrophy called Pelizaeus-Merzbacher-like disease or a milder spastic paraplegia. In addition, CNS demyelination may be caused by defects in genes expressing astrocytic GJ proteins, which are essential for oligodendrocytes. Findings from in vitro and in vivo models of these disorders developed over the last decade indicate that most mutations cause loss of function and an inability of the mutant connexins to form functional GJs. Here we review the clinical, genetic, and neurobiological aspects of GJ disorders affecting the PNS and CNS myelinating cells.

  5. Gap-junction-mediated cell-to-cell communication.

    Science.gov (United States)

    Hervé, Jean-Claude; Derangeon, Mickaël

    2013-04-01

    Cells of multicellular organisms need to communicate with each other and have evolved various mechanisms for this purpose, the most direct and quickest of which is through channels that directly connect the cytoplasms of adjacent cells. Such intercellular channels span the two plasma membranes and the intercellular space and result from the docking of two hemichannels. These channels are densely packed into plasma-membrane spatial microdomains termed "gap junctions" and allow cells to exchange ions and small molecules directly. A hemichannel is a hexameric torus of junctional proteins around an aqueous pore. Vertebrates express two families of gap-junction proteins: the well-characterized connexins and the more recently discovered pannexins, the latter being related to invertebrate innexins ("invertebrate connexins"). Some gap-junctional hemichannels also appear to mediate cell-extracellular communication. Communicating junctions play crucial roles in the maintenance of homeostasis, morphogenesis, cell differentiation and growth control in metazoans. Gap-junctional channels are not passive conduits, as previously long regarded, but use "gating" mechanisms to open and close the central pore in response to biological stimuli (e.g. a change in the transjunctional voltage). Their permeability is finely tuned by complex mechanisms that have just begun to be identified. Given their ubiquity and diversity, gap junctions play crucial roles in a plethora of functions and their dysfunctions are involved in a wide range of diseases. However, the exact mechanisms involved remain poorly understood.

  6. Gap junctions in isolated rat aorta: evidence for contractile responses that exhibit a differential dependence on intercellular communication

    Directory of Open Access Journals (Sweden)

    Christ G.J.

    2000-01-01

    Full Text Available Connexin43 (Cx43 is a major gap junction protein present in the Fischer-344 rat aorta. Previous studies have identified conditions under which selective disruption of intercellular communication with heptanol caused a significant, readily reversible and time-dependent diminution in the magnitude of a1-adrenergic contractions in isolated rat aorta. These observations have indentified a significant role for gap junctions in modulating vascular smooth muscle tone. The goal of these steady-state studies was to utilize isolated rat aortic rings to further evaluate the contribution of intercellular junctions to contractions elicited by cellular activation in response to several other vascular spasmogens. The effects of heptanol were examined (0.2-2.0 mM on equivalent submaximal (»75% of the phenylephrine maximum aortic contractions elicited by 5-hydroxytryptamine (5-HT; 1-2 µM, prostaglandin F2a (PGF2a; 1 µM and endothelin-1 (ET-1; 20 nM. Statistical analysis revealed that 200 µM and 500 µM heptanol diminished the maximal amplitude of the steady-state contractile responses for 5-HT from a control response of 75 ± 6% (N = 26 rings to 57 ± 7% (N = 26 rings and 34.9 ± 6% (N = 13 rings, respectively (P<0.05, and for PGF2a from a control response of 75 ± 10% (N = 16 rings to 52 ± 8% (N = 19 rings and 25.9 ± 6% (N = 18 rings, respectively (P<0.05. In contrast, 200 µM and 500 µM heptanol had no detectable effect on the magnitude of ET-1-induced contractile responses, which were 76 ± 5.0% for the control response (N = 38 rings, 59 ± 6.0% in the presence of 200 µM heptanol (N = 17 rings, and 70 ± 6.0% in the presence of 500 µM heptanol (N = 23 rings (P<0.13. Increasing the heptanol concentration to 1 mM was associated with a significant decrease in the magnitude of the steady-state ET-1-induced contractile response to 32 ± 5% (21 rings; P<0.01; further increasing the heptanol concentration to 2 mM had no additional effect. In rat aorta then

  7. Close the Gap : a study on the regulation of Connexin43 gap junctional communication

    NARCIS (Netherlands)

    Zeijl, Leonie van

    2009-01-01

    Gap junctions are groups of transmembrane channels that connect the cytoplasms of adjacent cells to mediate the diffusion of small molecules, such as ions, metabolites, second messengers and small peptides. The building blocks of gap junctions are connexin proteins. The most ubiquitous and best stu

  8. Fixed-gap tunnel junction for reading DNA nucleotides.

    Science.gov (United States)

    Pang, Pei; Ashcroft, Brian Alan; Song, Weisi; Zhang, Peiming; Biswas, Sovan; Qing, Quan; Yang, Jialing; Nemanich, Robert J; Bai, Jingwei; Smith, Joshua T; Reuter, Kathleen; Balagurusamy, Venkat S K; Astier, Yann; Stolovitzky, Gustavo; Lindsay, Stuart

    2014-12-23

    Previous measurements of the electronic conductance of DNA nucleotides or amino acids have used tunnel junctions in which the gap is mechanically adjusted, such as scanning tunneling microscopes or mechanically controllable break junctions. Fixed-junction devices have, at best, detected the passage of whole DNA molecules without yielding chemical information. Here, we report on a layered tunnel junction in which the tunnel gap is defined by a dielectric layer, deposited by atomic layer deposition. Reactive ion etching is used to drill a hole through the layers so that the tunnel junction can be exposed to molecules in solution. When the metal electrodes are functionalized with recognition molecules that capture DNA nucleotides via hydrogen bonds, the identities of the individual nucleotides are revealed by characteristic features of the fluctuating tunnel current associated with single-molecule binding events.

  9. Reduction of Gap Junctional Conductance by Microinjection of Antibodies against the 27-kDa Liver Gap Junction Polypeptide

    Science.gov (United States)

    Hertzberg, E. L.; Spray, D. C.; Bennett, M. V. L.

    1985-04-01

    Antibody raised against isolated rat liver gap junctions was microinjected into coupled cells in culture to assess its influence on gap junctional conductance. A rapid inhibition of fluorescent dye transfer and electrical coupling was produced in pairs of freshly dissociated adult rat hepatocytes and myocardial cells as well as in pairs of superior cervical ganglion neurons from neonatal rats cultured under conditions in which electrotonic synapses form. The antibodies have been shown by indirect immunofluorescence to bind to punctate regions of the plasma membrane in liver. By immunoreplica analysis of rat liver homogenates, plasma membranes, and isolated gap junctions resolved on NaDodSO4/polyacrylamide gels, binding was shown to be specific for the 27-kDa major polypeptide of gap junctions. This and similar antibodies should provide a tool for further investigation of the role of cell-cell communication mediated by gap junctions and indicate that immunologically similar polypeptides comprise gap junctions in adult mammalian cells derived from all three germ layers.

  10. The gap junction proteome and its relationship to disease.

    Science.gov (United States)

    Laird, Dale W

    2010-02-01

    In recent years our understanding of connexins has advanced from viewing them simply as proteins with a surprisingly short lifespan that form gap junction channels. Connexins are now known to be multifaceted proteins at the core of many multiprotein complexes that link to structural junctional complexes and cytoskeletal elements, and also to the cellular machinery that facilitates their transport, assembly, function and internalization. Collectively, these connexin-binding proteins can be termed the 'gap junction proteome'. The mechanistic understanding of the gap junction proteome with regards to the dynamic life cycle of connexins has grown further in importance in light of the large number of human diseases attributed to connexin gene mutations and regulatory changes in connexin spatial localization and expression levels.

  11. Experimental observation of subharmonic gap structures in long Josephson junctions

    DEFF Research Database (Denmark)

    Nordahn, M.A.; Manscher, Martin; Mygind, Jesper

    1999-01-01

    The subharmonic gap structure (SGS) in long-overlap Nb-AlOx-Nb Josephson tunnel junctions has been investigated. The experimental results show peaks in the differential conductance at both odd and even integer fractions of the gap voltage, VG Furthermore, the conductance peaks at V-G/2 has been...

  12. Pallidal gap junctions-triggers of synchrony in Parkinson's disease?

    Science.gov (United States)

    Schwab, Bettina C; Heida, Tjitske; Zhao, Yan; van Gils, Stephan A; van Wezel, Richard J A

    2014-10-01

    Although increased synchrony of the neural activity in the basal ganglia may underlie the motor deficiencies exhibited in Parkinson's disease (PD), how this synchrony arises, propagates through the basal ganglia, and changes under dopamine replacement remains unknown. Gap junctions could play a major role in modifying this synchrony, because they show functional plasticity under the influence of dopamine and after neural injury. In this study, confocal imaging was used to detect connexin-36, the major neural gap junction protein, in postmortem tissues of PD patients and control subjects in the putamen, subthalamic nucleus (STN), and external and internal globus pallidus (GPe and GPi, respectively). Moreover, we quantified how gap junctions affect synchrony in an existing computational model of the basal ganglia. We detected connexin-36 in the human putamen, GPe, and GPi, but not in the STN. Furthermore, we found that the number of connexin-36 spots in PD tissues increased by 50% in the putamen, 43% in the GPe, and 109% in the GPi compared with controls. In the computational model, gap junctions in the GPe and GPi strongly influenced synchrony. The basal ganglia became especially susceptible to synchronize with input from the cortex when gap junctions were numerous and high in conductance. In conclusion, connexin-36 expression in the human GPe and GPi suggests that gap junctional coupling exists within these nuclei. In PD, neural injury and dopamine depletion could increase this coupling. Therefore, we propose that gap junctions act as a powerful modulator of synchrony in the basal ganglia. © 2014 International Parkinson and Movement Disorder Society.

  13. Gap junctions-guards of excitability

    DEFF Research Database (Denmark)

    Stroemlund, Line Waring; Jensen, Christa Funch; Qvortrup, Klaus;

    2015-01-01

    Cardiomyocytes are connected by mechanical and electrical junctions located at the intercalated discs (IDs). Although these structures have long been known, it is becoming increasingly clear that their components interact. This review describes the involvement of the ID in electrical disturbances...

  14. Molecular mechanisms of gap junction mutations in myelinating cells.

    Science.gov (United States)

    Sargiannidou, Irene; Markoullis, Kyriaki; Kleopa, Kleopas A

    2010-09-01

    There is an emerging group of neurological disorders that result from genetic mutations affecting gap junction proteins in myelinating cells. The X-linked form of Charcot Marie Tooth disease (CMT1X) is caused by numerous mutations in the GJB1 gene encoding the gap junction protein connexin32 (Cx32), which is expressed in both Schwann cells in the PNS and oligodendrocytes in the CNS. Patients with CMT1X present mainly with a progressive peripheral neuropathy, showing mixed axonal and demyelinating features. In many cases there is also clinical or subclinical involvement of the CNS with acute or chronic phenotypes of encephalopathy. Furthermore, mutations in the GJA12/GJC2 gene encoding the gap junction protein Cx47, which is expressed in oligodendrocytes, have been identified in families with progressive leukodystrophy, known as Pelizaeus-Merzbacher-like disease, as well as in patients with hereditary spastic paraplegia. Recent studies have provided insights into the pattern of gap junction protein expression and function in CNS and PNS myelinating cells. Furthermore, in vitro and in vivo disease models have clarified some of the molecular and cellular mechanisms underlying these disorders. Here we provide an overview of the clinical, genetic, and neurobiological aspects of gap junction disorders affecting the nervous system.

  15. Gap junctions in developing thalamic and neocortical neuronal networks.

    Science.gov (United States)

    Niculescu, Dragos; Lohmann, Christian

    2014-12-01

    The presence of direct, cytoplasmatic, communication between neurons in the brain of vertebrates has been demonstrated a long time ago. These gap junctions have been characterized in many brain areas in terms of subunit composition, biophysical properties, neuronal connectivity patterns, and developmental regulation. Although interesting findings emerged, showing that different subunits are specifically regulated during development, or that excitatory and inhibitory neuronal networks exhibit various electrical connectivity patterns, gap junctions did not receive much further interest. Originally, it was believed that gap junctions represent simple passageways for electrical and biochemical coordination early in development. Today, we know that gap junction connectivity is tightly regulated, following independent developmental patterns for excitatory and inhibitory networks. Electrical connections are important for many specific functions of neurons, and are, for example, required for the development of neuronal stimulus tuning in the visual system. Here, we integrate the available data on neuronal connectivity and gap junction properties, as well as the most recent findings concerning the functional implications of electrical connections in the developing thalamus and neocortex.

  16. Effects of sevoflurance preconditioning on myocardial celluar CX43 protein during ischemia/reperfusion%七氟醚预处理对缺血再灌注心肌细胞CX43蛋白表达的影响

    Institute of Scientific and Technical Information of China (English)

    章征兵; 明腾; 杨文萍; 谢学良

    2012-01-01

    目的 观察七氟醚预处理对缺血再灌注心肌细胞缝隙连接蛋白43(CX43)表达的影响,探讨七氟醚预处理对心肌保护作用的机制.方法 选取需接受手术的先天性心脏病患儿38例,随机分为七氟醚预处理组(S组)和对照组(C组),每组19例.患儿于麻醉诱导气管插管后,S组吸入七氟醚1.0 MAC 30 min,随后洗脱10 min;C组则只吸入空氧混合气体.两组麻醉诱导及维持用药相同,均按常规行体外循环(CPB).监测两组患儿年龄、体质量、主动脉阻断时间、CPB时间、机械通气时间、重症监护对间和手术后住院天数及麻醉诱导后和主动脉开放6h后血浆CK-MB、cTnI的浓度;免疫组织化学方法显示心肌CX43的分布特征及CX43含量的变化.结果 两组患儿年龄、体质量、主动脉阻断时间、CPB时间、机械通气时间、重症监护时间和手术后住院天数、术前CK-MB、cTnI浓度及转流前CX43含量和分布无统计学差异(P>0.05);S组术后6 h CK.MB、cTnI浓度低于C组(P<0.05);转流后S组CX43阳性染色的平均灰度值低于C组,阳性染色的平均面积高于C组(P<0.05),且S组cX43蛋白主要分布在闰盘上,而C组主要分布在侧连接胞膜上.结论 七氟醚预处理能减少CPB后心肌细胞CX43蛋白的降解,稳定CX43的分布,发挥心肌保护作用.%Objective To observe the effects of sevoflurance preconditioning on myocaidial celluar conexin 43 (CX43) protein during ischemia/reperfusion and explore the mechanism of myocardial protection. Methods Thirty-eight pediatric patients with congenital heart disease who need surgery were randomly allocated into sevofiurance preconditioning group (group S) and control group (group C). After induction of anesthesia, sevoflurance were inhaled for 30 minutes and followed by 10 minutes air-oxygen mixture washout in group S, group C inhale air-oxygen mixture only. Establishing car-diopulmonary bypass (CPB) as a matter of routine in two

  17. Gap junction modulation by extracellular signaling molecules: the thymus model

    Directory of Open Access Journals (Sweden)

    Alves L.A.

    2000-01-01

    Full Text Available Gap junctions are intercellular channels which connect adjacent cells and allow direct exchange of molecules of low molecular weight between them. Such a communication has been described as fundamental in many systems due to its importance in coordination, proliferation and differentiation. Recently, it has been shown that gap junctional intercellular communication (GJIC can be modulated by several extracellular soluble factors such as classical hormones, neurotransmitters, interleukins, growth factors and some paracrine substances. Herein, we discuss some aspects of the general modulation of GJIC by extracellular messenger molecules and more particularly the regulation of such communication in the thymus gland. Additionally, we discuss recent data concerning the study of different neuropeptides and hormones in the modulation of GJIC in thymic epithelial cells. We also suggest that the thymus may be viewed as a model to study the modulation of gap junction communication by different extracellular messengers involved in non-classical circuits, since this organ is under bidirectional neuroimmunoendocrine control.

  18. Microwave resonant activation in hybrid single-gap/two-gap Josephson tunnel junctions

    Science.gov (United States)

    Carabello, Steven; Lambert, Joseph G.; Mlack, Jerome; Dai, Wenqing; Li, Qi; Chen, Ke; Cunnane, Daniel; Xi, X. X.; Ramos, Roberto C.

    2016-09-01

    Microwave resonant activation is a powerful, straightforward technique to study classical and quantum systems, experimentally realized in Josephson junction devices cooled to very low temperatures. These devices typically consist of two single-gap superconductors separated by a weak link. We report the results of the first resonant activation experiments on hybrid thin film Josephson junctions consisting of a multi-gap superconductor (MgB2) and a single-gap superconductor (Pb or Sn). We can interpret the plasma frequency in terms of theories both for conventional and hybrid junctions. Using these models, we determine the junction parameters including critical current, resistance, and capacitance and find moderately high quality factors of Q0˜ 100 for these junctions.

  19. Research progress of relationship between gap junctions and movement disorders

    Directory of Open Access Journals (Sweden)

    Hai-lei WANG

    2016-12-01

    Full Text Available Gap junctions (GJ is an important way to transmit signals among body cells. Studies have confirmed that changes on structure and composition of gap junction proteins are associated with many kinds of diseases. Movement disorders is a common disease of the central nervous system (CNS, which has a certain degree of correlation with GJ, and it has become a key factor in the domestic and foreign research of pathogenesis and treatment mechanism of central nervons system diseases. This paper reviews GJ and its relationship with movement disorders. DOI: 10.3969/j.issn.1672-6731.2016.12.011

  20. Microwave dependence of subharmonic gap structure in superconducting junctions

    DEFF Research Database (Denmark)

    Sørensen, O. Hoffman; Kofoed, Bent; Pedersen, Niels Falsig

    1974-01-01

    with the superconducting energy gap itself. The location in voltage of all these structures is given by eV=(2Δ±nh ν) / m, where 2Δ is the superconducting energy gap, ν is the applied frequency, h is Planck's constant, e is the magnitude of the electronic charge, V is the dc voltage drop across the junction, and m and n...

  1. Role of connexin43-interacting proteins at gap junctions

    NARCIS (Netherlands)

    Giepmans, Ben N G

    2006-01-01

    Gap junctions are arrays of cell-to-cell channels that allow diffusion of small molecules between neighboring cells. The individual channels are formed by the four-transmembrane connexin (Cx) proteins. Recently, multiple proteins have been found to interact at the cytoplasmic site with the most abun

  2. Pallidal gap junctions-triggers of synchrony in Parkinson's disease?

    NARCIS (Netherlands)

    Schwab, B.C.; Heida, T.; Zhao, Y.; Gils, S.A. van; Wezel, Richard van

    2014-01-01

    Although increased synchrony of the neural activity in the basal ganglia may underlie the motor deficiencies exhibited in Parkinson's disease (PD), how this synchrony arises, propagates through the basal ganglia, and changes under dopamine replacement remains unknown. Gap junctions could play a majo

  3. [Gap junctions: A new therapeutic target in major depressive disorder?].

    Science.gov (United States)

    Sarrouilhe, D; Dejean, C

    2015-11-01

    Major depressive disorder is a multifactorial chronic and debilitating mood disease with high lifetime prevalence and is associated with excess mortality, especially from cardiovascular diseases and through suicide. The treatments of this disease with tricyclic antidepressants and monoamine oxidase inhibitors are poorly tolerated and those that selectively target serotonin and norepinephrine re-uptake are not effective in all patients, showing the need to find new therapeutic targets. Post-mortem studies of brains from patients with major depressive disorders described a reduced expression of the gap junction-forming membrane proteins connexin 30 and connexin 43 in the prefrontal cortex and the locus coeruleus. The use of chronic unpredictable stress, a rodent model of depression, suggests that astrocytic gap junction dysfunction contributes to the pathophysiology of major depressive disorder. Chronic treatments of rats with fluoxetine and of rat cultured cortical astrocytes with amitriptyline support the hypothesis that the upregulation of gap junctional intercellular communication between brain astrocytes could be a novel mechanism for the therapeutic effect of antidepressants. In conclusion, astrocytic gap junctions are emerging as a new potential therapeutic target for the treatment of patients with major depressive disorder.

  4. Role of connexin43-interacting proteins at gap junctions

    NARCIS (Netherlands)

    Giepmans, Ben N G

    2006-01-01

    Gap junctions are arrays of cell-to-cell channels that allow diffusion of small molecules between neighboring cells. The individual channels are formed by the four-transmembrane connexin (Cx) proteins. Recently, multiple proteins have been found to interact at the cytoplasmic site with the most

  5. Pallidal gap junctions - Triggers of synchrony in Parkinson's disease?

    NARCIS (Netherlands)

    Schwab, B.C.; Heida, Tjitske; Zhao, Yan; van Gils, Stephanus A.; van Wezel, Richard Jack Anton

    2014-01-01

    Although increased synchrony of the neural activity in the basal ganglia may underlie the motor deficiencies exhibited in Parkinson's disease (PD), how this synchrony arises, propagates through the basal ganglia, and changes under dopamine replacement remains unknown. Gap junctions could play a majo

  6. Pallidal gap junctions-triggers of synchrony in Parkinson's disease?

    NARCIS (Netherlands)

    Schwab, B.C.; Heida, T.; Zhao, Y.; Gils, S.A. van; Wezel, Richard van

    2014-01-01

    Although increased synchrony of the neural activity in the basal ganglia may underlie the motor deficiencies exhibited in Parkinson's disease (PD), how this synchrony arises, propagates through the basal ganglia, and changes under dopamine replacement remains unknown. Gap junctions could play a majo

  7. Theories of subharmonic gap structures in superconducting junctions

    DEFF Research Database (Denmark)

    Hasselberg, L.E.; Levinsen, M. T.; Samuelsen, Mogens Rugholm

    1974-01-01

    The two theories of subharmonic gap structures in superconducting junctions, multiparticle tunneling and self-coupling due to an electromagnetic field set up by the ac Josephson current, are analyzed when microwaves are applied. Both theories give the same location in voltage for the microwave......-induced satellites and the same microwave-power dependence for the subharmonic gap structure and the satellites. Therefore other properties than these are to be considered in order to distinguish between the two theories. We suggest that self-coupling is the main cause of the subharmonic gap structure....

  8. Effects of Cx43 gene modification on the proliferation and migration of the human lung squamous carcinoma cell line NCI-H226.

    Science.gov (United States)

    Zang, J-P; Wei, R

    2015-10-27

    In this study, the human lung squamous carcinoma cell line NCI-H226 was transfected with the recombinant plasmid pBudCE4.1_Cx43 to explore the role of the Cx43 gene in cell growth, cell cycle, and tumor migration. pBudCE4.1-Cx43 was transfected into human lung squamous carcinoma NCI-H226 cells using Lipofectamine TM2000. The mRNA and protein expressions of Cx43 in the transfected cells were detected by reverse transcriptase polymerase chain reaction and western blot analysis. The cell-cell communication was detected using the scratch dye tracer method and the cell cycle was detected by flow cytometry. The CCK-8 proliferation, scratch healing, and cell invasion assays were performed to evaluate the effect of the Cx43 gene transfection on the proliferation, migration, and invasive abilities of NCI-H226 cells. Cx43 mRNA and protein expressions and the fluorescence intensity in the scratch healing test were significantly higher in the experimental group than those in the control and blank groups (P migration, respectively, in the experimental group, compared to the control and blank groups (P migration of human lung squamous carcinoma cell line NCI-H226, thereby inhibiting tumor cell proliferation.

  9. Cx43基因shRNA慢病毒载体的构建及其对大鼠心肌细胞Cx43基因的作用%Construction of a lentiviral vector for RNA interference targeting rat cardiac myocytes connexin 43 gene and its effect

    Institute of Scientific and Technical Information of China (English)

    陈立; 钟国强; 涂荣会; 黎庆捷; 何艳

    2013-01-01

    目的 构建缝隙连接蛋白(Cx)43基因shRNA慢病毒载体,并检测其对大鼠心肌细胞Cx43基因的作用.方法 针对Cx43基因序列,设计RNA干扰靶点序列,合成靶序列的双链DNA,接入pGCL-GFP载体,挑选阳性克隆行PCR鉴定及测序.用pHelper1.0和pHelper2.0质粒转染293T细胞,包装产生具备感染能力的慢病毒.以293T细胞中绿色荧光蛋白的细胞数量计算病毒滴度;以最适感染复数感染大鼠心肌细胞,通过荧光显微镜观察感染效率;应用real-time PCR和Western blot法检测大鼠心肌细胞.Cx43 mRNA及Cx43蛋白表达,并评价其抑制效果.结果 经PCR鉴定和测序证实,Cx43慢病毒载体构建正确,其病毒滴度为8×108 TU/mL、转染效率为82.59%.荧光定量real-time PCR法检测Cx43 mRNA的抑制率为96.10%,Western blot法检测Cx43蛋白的抑制率为77.16%.结论 成功构建了Cx43基因shRNA慢病毒载体,其能显著抑制大鼠心肌细胞Cx43基因的表达.%Objective To construct a lentiviral RNAi vector and to detect its effect on connexin 43 (Cx43) gene in rat cardiac myocytes.Methods Short hairpin RNA (shRNA) sequence targeting rat cardiac myocytes Cx43 gene was designed.After synthesis and annealing,the double-stranded oligonucleotides (ds oligo) were connecting to pGC-LV vectors.The positive clones were selected and conducted by PCR identification and sequencing.Then,The viral particles were gen erated by cotransfection of 293T cells with the pGC-lv-Cx43 and two packaging vector (pHelper1.0,pHelper2.0),and the virus titer was determined by counting the number of GFP positive cells.After transfection of lentiviral vector into rat cardiac myocytes with multiplicity of infection (MOI),the level of Cx43 mRNA in rat cardiac myocytes was determined by realtime PCR and the level of Cx43 protein was determined by Western blot assay.The inhibitory effect was evaluated.Results The construction of Cx43 lentiviral vector was confirmed by PCR identification

  10. Role of gap junctions and hemichannels in parasitic infections.

    Science.gov (United States)

    Vega, José Luis; Subiabre, Mario; Figueroa, Felipe; Schalper, Kurt Alex; Osorio, Luis; González, Jorge; Sáez, Juan Carlos

    2013-01-01

    In vertebrates, connexins (Cxs) and pannexins (Panxs) are proteins that form gap junction channels and/or hemichannels located at cell-cell interfaces and cell surface, respectively. Similar channel types are formed by innexins in invertebrate cells. These channels serve as pathways for cellular communication that coordinate diverse physiologic processes. However, it is known that many acquired and inherited diseases deregulate Cx and/or Panx channels, condition that frequently worsens the pathological state of vertebrates. Recent evidences suggest that Cx and/or Panx hemichannels play a relevant role in bacterial and viral infections. Nonetheless, little is known about the role of Cx- and Panx-based channels in parasitic infections of vertebrates. In this review, available data on changes in Cx and gap junction channel changes induced by parasitic infections are summarized. Additionally, we describe recent findings that suggest possible roles of hemichannels in parasitic infections. Finally, the possibility of new therapeutic designs based on hemichannel blokers is presented.

  11. [Gap junction-mediated intercellular communication in astrocytes and neuroprotection].

    Science.gov (United States)

    Giaume, C; Froger, N; Koulakoff, A

    2005-06-01

    Neuroglial interaction represents a concept that is now more and more integrated in the attempts to understand who does what and how in neuronal processing and survival, in normal as well as in pathological situations. The purpose of the review is to provide an overlook about the role of glial cells, mainly astrocytes, in neuroprotection. Since a typical feature of glia is to be connected by gap junctions that allow them to be organized as a communicating network(s), we will focus this review on what is known about the contribution of astrocyte gap junctions (AGJ) in neuronal survival. As neuroglial interaction and AGJ are both affected during neurodegenerative diseases, we will also consider the above mentioned glial properties in a pathological context with a special interest in Alzheimer's disease.

  12. Symposia for a Meeting on Ion Channels and Gap Junctions

    CERN Document Server

    Sáez, Juan

    1997-01-01

    Ion channels allow us to see nature in all its magnificence, to hear a Bach suite, to smell the aroma of grandmother's cooking, and, in this regard, they put us in contact with the external world. These ion channels are protein molecules located in the cell membrane. In complex organisms, cells need to communicate in order to know about their metabolic status and to act in a coordinate manner. The latter is also accomplished by a class of ion channels able to pierce the lipid bilayer membranes of two adjacent cells. These intercellular channels are the functional subunits of gap junctions. Accordingly, the book is divided in two parts: the first part is dedicated to ion channels that look to the external world, and the second part is dedicated to gap junctions found at cell interfaces. This book is based on a series of symposia for a meeting on ion channels and gap junctions held in Santiago, Chile, on November 28-30, 1995. The book should be useful to graduate students taking the first steps in this field as...

  13. [Relationship between Helicobacter pylori infection and expression of connexin (Cx) 32 and Cx43 genes in gastric cancer and gastric precancerous lesions].

    Science.gov (United States)

    Xu, Can-Xia; Jia, Yan; Yang, Wen-Bin; Wang, Fen; Shen, Shou-Rong

    2008-06-10

    To investigate the expression of connexin (Cx)32 and Cx43 genes in gastric cancer and precancerous lesion, and to investigate the relation between the changes of expression of Cx32 and Cx43 genes and Helicobacter pylori (Hp) infection. Gastroscopy and biopsy of gastric mucosa were conducted on 33 patients with chronic superficial gastritis (CSG), 88 with precancerous lesion, and 70 with gastric cancer. Hp was detected by rapid urease test, basic fuchsin staining, and 14C-urea breath test. The CagA gene of Hp was determined by PCR. SABC immunohistochemical method was used to detect the expression of Cx32 and Cx43 genes in gastric mucosa biopsy specimens. The positive expression rates of Cx32 and Cx43 genes were 15.7% and 32.9% respectively in the gastric cancer patients, 51.1% and 54.5% in the patients with precancerous lesion, and 100.0% and 93.9% in the CSG patients. The positive Cx32 and Cx43 expression rates of the gastric cancer and precancerous lesion patients were significantly lower than those of the CSG patients (all P infection was 16.7%, not significantly different from that of the gastric cancer patients without Hp infection (13.6%). The positive Cx43 expression rate of the gastric cancer patients with Hp infection was 25%, significantly lower than that of the gastric cancer patients without Hp infection (50%, P = 0.039). The positive Cx32 and Cx43 expression rates and expression intensity of the precancerous lesion patients with Hp infection were all significantly lower than those of the precancerous lesion patients without Hp infection (all P infection was 17.9%, significantly lower than that of the CagA- Hp group (55.6%, P = 0.027), however, the positive Cx32 expression rate of the gastric cancer patients with CagA+ Hp infection was 12.8%, not significantly different from that of the gastric cancer patients with CagA- Hp infection (33.3%, P = 0.159). The positive Cx32 and Cx43 expression rates of the CSG patients with CagA+ Hp and CagA- Hp infection

  14. Mechanisms of gap junction traffic in health and disease.

    Science.gov (United States)

    Hesketh, Geoffrey G; Van Eyk, Jennifer E; Tomaselli, Gordon F

    2009-10-01

    Gap junctions (GJs) allow direct communication between cells. In the heart, GJs mediate the electrical coupling of cardiomyocytes and as such dictate the speed and direction of cardiac conduction. A prominent feature of acquired structural heart disease is remodeling of GJ protein expression and localization concomitant with increased susceptibility to lethal arrhythmias, leading many to hypothesize that the two are causally linked. Detailed understanding of the cellular mechanisms that regulate GJ localization and function within cardiomyocytes may therefore uncover potential therapeutic strategies for a significant clinical problem. This review will outline our current understanding of GJ cell biology with the intent of highlighting cellular mechanisms responsible for GJ remodeling associated with cardiac disease.

  15.  Gap junction involvement in hippocampal theta rhythm generation

    Directory of Open Access Journals (Sweden)

    Renata Bocian

    2012-10-01

    Full Text Available  Hippocampal theta rhythm is probably the best example of oscillations and synchrony phenomena occurring in neuronal networks of the central nervous system. It is well known that intraneuronal communication via chemical and electrical synapses underlies these oscillatory processes. Despite well-documented knowledge concerning the participation of chemical transmission in production of theta activity, the role of much faster gap junction communication is still not fully understood. This paper provides an overview of current research data concerning the involvement of electrical transmission in generation of the best synchronized EEG pattern recorded from the mammalian brain – theta rhythm.

  16. Role of Gap Junctions and Hemichannels in Parasitic Infections

    Directory of Open Access Journals (Sweden)

    José Luis Vega

    2013-01-01

    Full Text Available In vertebrates, connexins (Cxs and pannexins (Panxs are proteins that form gap junction channels and/or hemichannels located at cell-cell interfaces and cell surface, respectively. Similar channel types are formed by innexins in invertebrate cells. These channels serve as pathways for cellular communication that coordinate diverse physiologic processes. However, it is known that many acquired and inherited diseases deregulate Cx and/or Panx channels, condition that frequently worsens the pathological state of vertebrates. Recent evidences suggest that Cx and/or Panx hemichannels play a relevant role in bacterial and viral infections. Nonetheless, little is known about the role of Cx- and Panx-based channels in parasitic infections of vertebrates. In this review, available data on changes in Cx and gap junction channel changes induced by parasitic infections are summarized. Additionally, we describe recent findings that suggest possible roles of hemichannels in parasitic infections. Finally, the possibility of new therapeutic designs based on hemichannel blokers is presented.

  17. Mefloquine gap junction blockade and risk of pregnancy loss.

    Science.gov (United States)

    Nevin, Remington Lee

    2012-09-01

    Obstetric use of the antimalarial drug mefloquine has historically been discouraged during the first trimester and immediately before conception owing to concerns of potential fetal harm. With the rise of resistance to the antimalarial drug sulfadoxine-pyrimethamine (SP), mefloquine is now being considered as a replacement for SP for universal antenatal administration to women from malaria-endemic regions. Recent recommendations have also suggested that mefloquine may be used cautiously among pregnant travelers who cannot otherwise avoid visiting these areas. Mefloquine has been demonstrated to cause blockade of gap junction protein alpha 1 (GJA1) gap junction intercellular communication (GJIC), and recent evidence suggests that GJA1 GJIC is critical to successful embryonic implantation and early placental development. During routine use, mefloquine accumulates in organ and peripheral tissue, crosses the blood-placental barrier, and may plausibly accumulate in developing decidua and trophoblast at concentrations sufficient to interfere with GJA1 GJIC and, thus, cause deleterious effects on fetal outcomes. This conclusion is supported by epidemiological evidence that demonstrates use of the drug during early development is associated with an increased risk of miscarriage and stillbirth. Confirmatory studies are pending, but the available experimental and epidemiological evidence support renewed adherence, where feasible, to existing mefloquine package insert guidance that women avoid the drug during the periconceptional period.

  18. Modulation of gap junction channels and hemichannels by growth factors.

    Science.gov (United States)

    Schalper, Kurt A; Riquelme, Manuel A; Brañes, María C; Martínez, Agustín D; Vega, José Luis; Berthoud, Viviana M; Bennett, Michael V L; Sáez, Juan C

    2012-03-01

    Gap junction hemichannels and cell-cell channels have roles in coordinating numerous cellular processes, due to their permeability to extra and intracellular signaling molecules. Another mechanism of cellular coordination is provided by a vast array of growth factors that interact with relatively selective cell membrane receptors. These receptors can affect cellular transduction pathways, including alteration of intracellular concentration of free Ca(2+) and free radicals and activation of protein kinases or phosphatases. Connexin and pannexin based channels constitute recently described targets of growth factor signal transduction pathways, but little is known regarding the effects of growth factor signaling on pannexin based channels. The effects of growth factors on these two channel types seem to depend on the cell type, cell stage and connexin and pannexin isoform expressed. The functional state of hemichannels and gap junction channels are affected in opposite directions by FGF-1 via protein kinase-dependent mechanisms. These changes are largely explained by channels insertion in or withdrawal from the cell membrane, but changes in open probability might also occur due to changes in phosphorylation and redox state of channel subunits. The functional consequence of variation in cell-cell communication via these membrane channels is implicated in disease as well as normal cellular responses.

  19. Interstitial volume modulates the conduction velocity-gap junction relationship.

    Science.gov (United States)

    Veeraraghavan, Rengasayee; Salama, Mohamed E; Poelzing, Steven

    2012-01-01

    Cardiac conduction through gap junctions is an important determinant of arrhythmia susceptibility. Yet, the relationship between degrees of G(j) uncoupling and conduction velocity (θ) remains controversial. Conflicting results in similar experiments are normally attributed to experimental differences. We hypothesized that interstitial volume modulates conduction velocity and its dependence on G(j). Interstitial volume (V(IS)) was quantified histologically from guinea pig right ventricle. Optical mapping was used to quantify conduction velocity and anisotropy (AR(θ)). Albumin (4 g/l) decreased histologically assessed V(IS), increased transverse θ by 71 ± 10%, and lowered AR(θ). Furthermore, albumin did not change isolated cell size. Conversely, mannitol increased V(IS), decreased transverse θ by 24 ± 4%, and increased AR(θ). Mannitol also decreased cell width by 12%. Furthermore, mannitol was associated with spontaneous ventricular tachycardias in three of eight animals relative to zero of 15 during control. The θ-G(j) relationship was assessed using the G(j) uncoupler carbenoxolone (CBX). Whereas 13 μM CBX did not significantly affect θ during control, it slowed transverse θ by 38 ± 9% during mannitol (edema). These data suggest changes in V(IS) modulate θ, AR(θ), and the θ-G(j) relationship and thereby alter arrhythmia susceptibility. Therefore, V(IS) may underlie arrhythmia susceptibility, particularly in diseases associated with gap junction remodeling.

  20. Gap junction proteins are key drivers of endocrine function.

    Science.gov (United States)

    Meda, Paolo

    2017-03-08

    It has long been known that the main secretory cells of exocrine and endocrine glands are connected by gap junctions, made by a variety of connexin species that ensure their electrical and metabolic coupling. Experiments in culture systems and animal models have since provided increasing evidence that connexin signaling contributes to control the biosynthesis and release of secretory products, as well as to the life and death of secretory cells. More recently, genetic studies have further provided the first lines of evidence that connexins also control the function of human glands, which are central to the pathogenesis of major endocrine diseases. Here, we summarize the recent information gathered on connexin signaling in these systems, since the last reviews on the topic, with particular regard to the pancreatic beta cells which produce insulin, and the renal cells which produce renin. These cells are keys to the development of various forms of diabetes and hypertension, respectively, and combine to account for the exploding, worldwide prevalence of the metabolic syndrome. This article is part of a Special Issue entitled: Gap Junction Proteins edited by Jean Claude Herve. Copyright © 2017 Elsevier B.V. All rights reserved.

  1. Analysis of gap junctional intercellular communications using a dielectrophoresis-based microchip

    OpenAIRE

    Tellez-Gabriel, M.; Charrier, C.; Brounais-Le Royer, B; Mullard, M.; Brown, H K; F. Verrecchia(-ASI ASDC;); Heymann, D

    2017-01-01

    International audience; Please cite this article in press as: Tellez-Gabriel, M., et al., Analysis of gap junctional intercellular communications using a dielectrophoresis-based microchip. Gap junctions are transmembrane structures that directly connect the cytoplasm of adjacent cells, making intercellular communications possible. It has been shown that the behaviour of several tumours – such as bone tumours – is related to gap junction intercellular communications (GJIC). Several methodologi...

  2. The gap junction cellular internet: connexin hemichannels enter the signalling limelight

    National Research Council Canada - National Science Library

    Evans, W Howard; De Vuyst, Elke; Leybaert, Luc

    2006-01-01

    Cxs (connexins), the protein subunits forming gap junction intercellular communication channels, are transported to the plasma membrane after oligomerizing into hexameric assemblies called connexin hemichannels (CxHcs...

  3. Role of Cx43 gene in the process of myocardialization of proximal outflow tract septum in the mouse heart%Cx43基因对近端流出道隔心肌化过程的调控机制

    Institute of Scientific and Technical Information of China (English)

    赵晓晴; 黄国英; 谢利剑; 彭涛; 陈萍; 周国民

    2006-01-01

    目的:探讨connexin43(Cx43)基因在小鼠近端流出道隔心肌化的作用及其机制.方法:选用胚胎(ED)11.5 d至出生后1 d的Cx43基因剔除(KO)纯合型(Cx43-/-)、杂合型(Cx43+/-)及野生型(Cx43+/+)C57/BL6小鼠作为研究对象;采用PCR方法鉴定基因型;HE染色观察心脏结构,免疫组化法测定横纹肌肌动蛋白α-SCA、凋亡相关分子active caspase-3及神经嵴细胞的标志物AP-2的表达.结果:①Cx43-/-小鼠大多出生后24 h内即死亡.大体解剖见明显的右室流出道圆锥部异常膨隆,右心房扩张;组织切片HE染色示右室流出道壁大量异常小梁状组织增生突起,形成多个囊状结构,右室流出道腔明显狭窄,右室腔扩张.Cx43+/-和Cx43+/+心脏无明显异常.②Cx43-/-小鼠近端流出道隔中央区域α-SCA的表达明显滞后.③Active caspase-3组化显示Cx43+/+凋亡细胞主要出现在近端流出道隔,ED12.5至ED15.5均可见到;Cx43+/-凋亡减少,Cx43-/-则仅见很少凋亡细胞.④Cx43-/-流出道AP-2的表达增多,且表达位置异常.结论:Cx43 KO小鼠以右室流出道异常增生引起的梗阻性畸形为主要特征,该病变过程可能与胚胎期近端流出道隔心肌化迟滞有关,其近端流出道隔细胞凋亡减少和神经嵴细胞表达异常很可能参与了流出道心肌化异常的形成,表明Cx43在近端流出道隔心肌化过程中具有重要作用.

  4. Effects of doxycycline on the distribution of gap junction and susceptibility of arrhythmias in isolated ischemia-reperfusion rat hearts%多西环素对离体大鼠心肌缺血再灌注后缝隙连接分布及心律失常的影响

    Institute of Scientific and Technical Information of China (English)

    李镇; 严激; 徐健; 朱红军; 余华; 程敏; 陈康玉; 沈伟

    2011-01-01

    目的 研究多西环素对离体大鼠心肌缺血再灌注后心律失常易感性的影响,并通过分析缝隙连接的重新分布探讨其可能机制.方法 48只大鼠随机分为空白对照组、缺血再灌组、多西环素干预组.采用Langendorff灌注系统,制备离体大鼠缺血再灌注模型,用ELISA方法检测各组间大鼠心肌基质金属蛋白酶-9(MMP-9)的含量,通过Western-blot方法及免疫荧光标记方法检测心肌Cx43的分布及含量.用电生理方法观察多西环素对缺血再灌注后室性快速性心律失常发生的影响.结果 缺血再灌组及多西环素干预组较空白对照组MMP-9水平显著增高(P0.05).结论 多西环素可通过减少MMP-9的表达,减轻缺血后缝隙连接的侧化分布并减少再灌注心律失常发生.%Aim To investigate the effects and the possible mechanisms of doxycycline on the susceptibility of ventricular tachyarrhythmias in isolated ischemia-reperfusion rat hearts. Methods 48 isolated rat hearts perfused with a langendorff instrument were divided into blank group,ischemia-reperfusion group and doxycycline intervention group( n = 16 , respectively ). Matrix metalloproteinase-9( MMP-9 )were detected hy ELISA. Cx43 in the cardiomyocytes were quantified by western-blotting and localized by immunofluorescence. The risk of inducible ventricular tachyarrhythmias was evaluated by electrophysiological study. Results Doxycycline could significantly dilute MMP-9 in the ischemia-reperfusion rat hearts( P < 0. 001 ). Ventricular tachyarrhythmias tended to he reduced in doxycycline-perfused hearts( P < 0. 05 ). Redistrihution of Cx43 can he ohserved in the tissue of doxycycline-perfused hearts. Conclusion Doxycycline can prevent gap junction redistrihution hy reducing MMP-9 in the heart tissue, thereby decrease inducihle ventricular tachyarrhythmias in ischemia-reperfusion hearts.

  5. Role of gap junctions on synchronization in human neocortical networks.

    Science.gov (United States)

    Gigout, S; Deisz, R A; Dehnicke, C; Turak, B; Devaux, B; Pumain, R; Louvel, J

    2016-04-15

    Gap junctions (GJ) have been implicated in the synchronization of epileptiform activities induced by 4-aminopyrine (4AP) in slices from human epileptogenic cortex. Previous evidence implicated glial GJ to govern the frequency of these epileptiform events. The synchrony of these events (evaluated by the phase unlocking index, PUI) in adjacent areas however was attributed to neuronal GJ. In the present study, we have investigated the effects of GAP-134, a recently developed specific activator of glial GJ, on both the PUI and the frequency of the 4AP-induced epileptiform activities in human neocortical slices of temporal lobe epilepsy tissue. To delineate the impact of GJ on spatial spread of synchronous activity we evaluated the effects of carbenoxolone (CBX, a non-selective GJ blocker) on the spread in three axes 1. vertically in a given cortical column, 2. laterally within the deep cortical layers and 3. laterally within the upper cortical layers. GAP-134 slightly increased the frequency of the 4AP-induced spontaneous epileptiform activities while leaving the PUI unaffected. CBX had no effect on the PUI within a cortical column or on the PUI in the deep cortical layers. CBX increased the PUI for long interelectrodes distances in the upper cortical layers. In conclusion we provide new arguments toward the role played by glial GJ to maintain the frequency of spontaneous activities. We show that neuronal GJ control the PUI only in upper cortical layers.

  6. Effects of microtubule stabilizer on connexin 43 of isolated cardiac myocytes under hypoxia injury%微管稳定剂Taxol对缺氧心肌细胞Cx43蛋白的影响

    Institute of Scientific and Technical Information of China (English)

    王德国; 王新; 刘玉; 王星; 邢文; 李祥东; 杨胜; 王安才; 孙贤林

    2012-01-01

    Aim To observe the effects of mierotu-bule stabilizer ( Taxol) on eonnexin43 ( Cx43 ) expression and distribution of eardiae myoeyles and investi-gate its new value as an anti-arrhylhmie agent. Methods Cardiae myoevles isolated from aduk rats were exposed to hypoxia for 120 min with or without taxol ( a kind of mierotubule stabilizer ) in different. eoneentra-tions. Cellular survival was assayed by trypan blue dye-exelusion. Cx43 expression was determined by western blots. Cx43 distribution was deteeted by immu-nofluoreseenee staining and observed under laser sean-ning eonfoeal mieroseope. Results The cell viability under hypoxie eondition was redueed signifieantly whieh showed only few numbers of rod-shaped eells. Taxol promoted cell survival in a dose-dependent man- ner at the range of0. 1~10 nmol · L-1 . At the range of 100 nmol o L-1~ 10 μmol · L~1 , taxol lead toredueed cell survival. Taxol in a dose-dependent manner improved Cx43 expression even although exeessive taxol also suppressed Cx43. Cx43 was loeat ed in both ends of intaet eardiae myoeyles ( interealated disk ) and redistributed in both sides of the cell under hypoxie eonditions. Taxol dose-dependently inhibited lateraliza-tion of Cx43 in lower eoneentration and further de-ereased Cx43 in the interealated disk. Conclusions Cx43 redistribution indueed by hypoxie is proteeted by low dosage of taxol. Mierotubule stabilizer may be ae-ted as a new anti-arrhythmie agent.%目的 观察微管稳定剂Taxol对缺氧心肌细胞Cx43表达和分布的影响,探讨其作为新的抗心律失常药物的潜在价值.方法 酶解法分离的大鼠心肌细胞缺氧120 min,并以不同浓度的Taxol干预;台盼蓝排斥实验测定细胞存活率;免疫印迹检测Cx43的蛋白表达,免疫荧光染色后激光共聚焦显微镜观察分析Cx43的分布.结果 Taxol在0.1~10 nmol·L-1浓度下成剂量依赖性地提高杆状细胞数,促进细胞存活,而100 nmol·L-1~10 μmol·L-1浓度下杆状细胞数

  7. Microinjection Technique for Assessment of Gap Junction Function.

    Science.gov (United States)

    Fridman, Michael D; Liu, Jun; Sun, Yu; Hamilton, Robert M

    2016-01-01

    Gap junctions are essential for the proper function of many native mammalian tissues including neurons, cardiomyocytes, embryonic tissues, and muscle. Assessing these channels is therefore fundamental to understanding disease pathophysiology, developing therapies for a multitude of acquired and genetic conditions, and providing novel approaches to drug delivery and cellular communication. Microinjection is a robust, albeit difficult, technique, which provides considerable information that is superior to many of the simpler techniques due to its ability to isolate cells, quantify kinetics, and allow cross-comparison of multiple cell lines. Despite its user-dependent nature, the strengths of the technique are considerable and with the advent of new, automation technologies may improve further. This text describes the basic technique of microinjection and briefly discusses modern automation advances that can improve the success rates of this technique.

  8. Loss of αT-catenin alters the hybrid adhering junctions in the heart and leads to dilated cardiomyopathy and ventricular arrhythmia following acute ischemia.

    Science.gov (United States)

    Li, Jifen; Goossens, Steven; van Hengel, Jolanda; Gao, Erhe; Cheng, Lan; Tyberghein, Koen; Shang, Xiying; De Rycke, Riet; van Roy, Frans; Radice, Glenn L

    2012-02-15

    It is generally accepted that the intercalated disc (ICD) required for mechano-electrical coupling in the heart consists of three distinct junctional complexes: adherens junctions, desmosomes and gap junctions. However, recent morphological and molecular data indicate a mixing of adherens junctional and desmosomal components, resulting in a 'hybrid adhering junction' or 'area composita'. The α-catenin family member αT-catenin, part of the N-cadherin-catenin adhesion complex in the heart, is the only α-catenin that interacts with the desmosomal protein plakophilin-2 (PKP2). Thus, it has been postulated that αT-catenin might serve as a molecular integrator of the two adhesion complexes in the area composita. To investigate the role of αT-catenin in the heart, gene targeting technology was used to delete the Ctnna3 gene, encoding αT-catenin, in the mouse. The αT-catenin-null mice are viable and fertile; however, the animals exhibit progressive cardiomyopathy. Adherens junctional and desmosomal proteins were unaffected by loss of αT-catenin, with the exception of the desmosomal protein PKP2. Immunogold labeling at the ICD demonstrated in the αT-catenin-null heart a preferential reduction of PKP2 at the area composita compared with the desmosome. Furthermore, gap junction protein Cx43 was reduced at the ICD, including its colocalization with N-cadherin. Gap junction remodeling in αT-catenin-knockout hearts was associated with an increased incidence of ventricular arrhythmias after acute ischemia. This novel animal model demonstrates for the first time how perturbation in αT-catenin can affect both PKP2 and Cx43 and thereby highlights the importance of understanding the crosstalk between the junctional proteins of the ICD and its implications for arrhythmogenic cardiomyopathy.

  9. 胃癌和癌前病变患者Cx32,Cx43表达与幽门螺杆菌感染的相关性%Relationship between Helicobacter pylori infection and expression of connexin(Cx)32 and Cx43 genes in gastric cancer and gastric precancerous lesions

    Institute of Scientific and Technical Information of China (English)

    徐灿霞; 贾燕; 杨文斌; 王芬; 沈守荣

    2008-01-01

    Objective To investigate the expression of connexin (Cx) 32 and Cx43 genes in gastric cancer and precancerous lesion,and to investigate the relation between the changes of expression of Cx32 and Cx43 genes and Helicobacter pylori (Hp) infection.Methods Gastroscopy and biopsy of gastric mucosa were conducted on 33 patients with chronic superficial gastritis(CSG),88 with precancerous lesion,and 70 with gastric cancer.Hp was detected by rapid urease test,basic fuchsin staining,and 14C-urea breath test.The CagA gene of Hp Was determined by PCR.SABC immunohistochemical method was used to detect the expression of Cx32 and Cx43 genes in gastric mucosa biopsy specimens.Results The positive expression rates of Cx32 and Cx43 genes were 15.7%and 32.9%respectively in the gastric cancer patients,51.1% and 54.5%in the patients with precancerous lesion,and 100.0%and 93.9%in the CSG patients.The positive Cx32 and Cx43 expression rates of the gastric cancer and precancerous lesion patients were significantly lower than those of the CSG patients(all P<0.05).The positive Cx32 expression rate of the gastric cancer patients with Hp infection was 16.7%,not significantly different from that of the gastric cancer patients without Hp infection(13.6%).The positive Cx43 expression rate of the gastric cancer patients with Hp infection was 25%,significantly lower than that of the gastric cancer patients without Hp infection(50%,P=0.039).The positive Cx32 and Cx43 expression rates and expression intensity of the precancerous lesion patients with Hp infection were all significantly lower than those of the precancerous lesion patients without Hp infection(all P<0.05).The positive Cx43 expression rate of the gastric cancer patients with CasA+ Hp infection was 17.9%,significantly lower than that of the CasA- Hp group (55.6%,P=0.027),however,the positive Cx32 expression rate of the gastric cancer patients with CasA+Hp infection was 12.8%,not significantly different from that of the gastric cancer

  10. Regulation of Gap Junction Dynamics by UNC-44/ankyrin and UNC-33/CRMP through VAB-8 in C. elegans Neurons.

    Directory of Open Access Journals (Sweden)

    Lingfeng Meng

    2016-03-01

    Full Text Available Gap junctions are present in both vertebrates and invertebrates from nematodes to mammals. Although the importance of gap junctions has been documented in many biological processes, the molecular mechanisms underlying gap junction dynamics remain unclear. Here, using the C. elegans PLM neurons as a model, we show that UNC-44/ankyrin acts upstream of UNC-33/CRMP in regulation of a potential kinesin VAB-8 to control gap junction dynamics, and loss-of-function in the UNC-44/UNC-33/VAB-8 pathway suppresses the turnover of gap junction channels. Therefore, we first show a signal pathway including ankyrin, CRMP, and kinesin in regulating gap junctions.

  11. Connexins, E-cadherin, Claudin-7 and β-catenin transiently form junctional nexuses during the post-natal mammary gland development.

    Science.gov (United States)

    Dianati, Elham; Poiraud, Jérémy; Weber-Ouellette, Anne; Plante, Isabelle

    2016-08-01

    Gap junctions are intercellular channels made of connexins (Cxs) that allow direct communication between adjacent cells. Modulation of Cxs has been associated with abnormal development and function of the mammary gland and breast cancer. However, the mechanisms underlying their expression during normal mammary gland are not yet known. Cxs interact with components of tight and adherens junctions. Thus, we hypothesized that the expression levels of Cxs vary during mammary gland development and are regulated through stage-dependent interactions with members of the tight and adherens junctions. Our specific objectives were to: 1) determine the expression of Cxs and tight and adherens junction proteins throughout development and 2) characterize Cxs interactions with components of tight and adherens junctions. Murine mammary glands were sampled at various developmental stages (pre-pubescent to post-weaning). RT-qPCR and western-blot analyses demonstrated differential expression patterns for all gap (Cx43, Cx32, Cx26, Cx30), tight (Claudin-1, -3, -4, -7) and adherens (β-catenin, E- and P-cadherins) junctions throughout development. Interestingly, co-immunoprecipitation demonstrated interactions between these different types of junctions. Cx30 interacted with Cx26 just at the late pregnancy stage. While Cx43 showed a persistent interaction with β-catenin from virginity to post-weaning, its interactions with E-cadherin and Claudin-7 were transient. Cx32 interacted with Cx26, E-cadherin and β-catenin during lactation. Immunofluorescence results confirmed the existence of a junctional nexus that remodeled during mammary gland development. Together, our results confirm that the expression levels of Cxs vary concomitantly and that Cxs form junctional nexuses with tight and adherens junctions, suggesting the existence of common regulatory pathways.

  12. ATP- and gap junction-dependent intercellular calcium signaling in osteoblastic cells

    DEFF Research Database (Denmark)

    Jorgensen, N R; Geist, S T; Civitelli, R

    1997-01-01

    mechanically induced calcium waves in two rat osteosarcoma cell lines that differ in the gap junction proteins they express, in their ability to pass microinjected dye from cell to cell, and in their expression of P2Y2 (P2U) purinergic receptors. ROS 17/2.8 cells, which express the gap junction protein...

  13. Three-dimensional modeling and quantitative analysis of gap junction distributions in cardiac tissue.

    Science.gov (United States)

    Lackey, Daniel P; Carruth, Eric D; Lasher, Richard A; Boenisch, Jan; Sachse, Frank B; Hitchcock, Robert W

    2011-11-01

    Gap junctions play a fundamental role in intercellular communication in cardiac tissue. Various types of heart disease including hypertrophy and ischemia are associated with alterations of the spatial arrangement of gap junctions. Previous studies applied two-dimensional optical and electron-microscopy to visualize gap junction arrangements. In normal cardiomyocytes, gap junctions were primarily found at cell ends, but can be found also in more central regions. In this study, we extended these approaches toward three-dimensional reconstruction of gap junction distributions based on high-resolution scanning confocal microscopy and image processing. We developed methods for quantitative characterization of gap junction distributions based on analysis of intensity profiles along the principal axes of myocytes. The analyses characterized gap junction polarization at cell ends and higher-order statistical image moments of intensity profiles. The methodology was tested in rat ventricular myocardium. Our analysis yielded novel quantitative data on gap junction distributions. In particular, the analysis demonstrated that the distributions exhibit significant variability with respect to polarization, skewness, and kurtosis. We suggest that this methodology provides a quantitative alternative to current approaches based on visual inspection, with applications in particular in characterization of engineered and diseased myocardium. Furthermore, we propose that these data provide improved input for computational modeling of cardiac conduction.

  14. Ephaptic coupling rescues conduction failure in weakly coupled cardiac tissue with voltage-gated gap junctions

    Science.gov (United States)

    Weinberg, S. H.

    2017-09-01

    Electrical conduction in cardiac tissue is usually considered to be primarily facilitated by gap junctions, providing a pathway between the intracellular spaces of neighboring cells. However, recent studies have highlighted the role of coupling via extracellular electric fields, also known as ephaptic coupling, particularly in the setting of reduced gap junction expression. Further, in the setting of reduced gap junctional coupling, voltage-dependent gating of gap junctions, an oft-neglected biophysical property in computational studies, produces a positive feedback that promotes conduction failure. We hypothesized that ephaptic coupling can break the positive feedback loop and rescue conduction failure in weakly coupled cardiac tissue. In a computational tissue model incorporating voltage-gated gap junctions and ephaptic coupling, we demonstrate that ephaptic coupling can rescue conduction failure in weakly coupled tissue. Further, ephaptic coupling increased conduction velocity in weakly coupled tissue, and importantly, reduced the minimum gap junctional coupling necessary for conduction, most prominently at fast pacing rates. Finally, we find that, although neglecting gap junction voltage-gating results in negligible differences in well coupled tissue, more significant differences occur in weakly coupled tissue, greatly underestimating the minimal gap junctional coupling that can maintain conduction. Our study suggests that ephaptic coupling plays a conduction-preserving role, particularly at rapid heart rates.

  15. Regulation of neuronal axon specification by glia-neuron gap junctions in C. elegans

    Science.gov (United States)

    Meng, Lingfeng; Zhang, Albert; Jin, Yishi; Yan, Dong

    2016-01-01

    Axon specification is a critical step in neuronal development, and the function of glial cells in this process is not fully understood. Here, we show that C. elegans GLR glial cells regulate axon specification of their nearby GABAergic RME neurons through GLR-RME gap junctions. Disruption of GLR-RME gap junctions causes misaccumulation of axonal markers in non-axonal neurites of RME neurons and converts microtubules in those neurites to form an axon-like assembly. We further uncover that GLR-RME gap junctions regulate RME axon specification through activation of the CDK-5 pathway in a calcium-dependent manner, involving a calpain clp-4. Therefore, our study reveals the function of glia-neuron gap junctions in neuronal axon specification and shows that calcium originated from glial cells can regulate neuronal intracellular pathways through gap junctions. DOI: http://dx.doi.org/10.7554/eLife.19510.001 PMID:27767956

  16. Functional consequences of heterogeneous gap junction channel formation and its influence in health and disease.

    Science.gov (United States)

    Cottrell, G Trevor; Burt, Janis M

    2005-06-10

    The capacity of multiple connexins to hetero-oligomerize into functional heterogeneous gap junction channels has been demonstrated in vivo, in vitro, and in nonmammalian expression systems. These heterogeneous channels display gating activity, channel conductances, selectivity and regulatory behaviors that are sometimes not predicted by the behaviors of the corresponding homogeneous channels. Such observations suggest that heteromerization of gap junction proteins offers an efficient cellular strategy for finely regulating cell-to-cell communication. The available evidence strongly indicates that heterogeneous gap junction assembly is important to normal growth and differentiation, and may influence the appearance of several disease states. Definitive evidence that heterogeneous gap junction channels differentially regulate electrical conduction in excitable cells is absent. This review examines the prevalence, regulation, and implications of gap junction channel hetero-oligomerization.

  17. The role of gap junctions in the brain in health and disease.

    Science.gov (United States)

    Dere, Ekrem; Zlomuzica, Armin

    2012-01-01

    Gap junctions connect the cytosolic compartments of adjacent cells for direct electrotonic and metabolic cell-to-cell communication. Gap junctions between glial cells or neurons are ubiquitously expressed in the brain and play a role in brain development including cell differentiation, cell migration and survival, tissue homeostasis, as well as in human diseases including hearing loss, skin disease, neuropathies, epilepsy, brain trauma, and cardiovascular disease. Furthermore, gap junctions are involved in the synchronization and rhythmic oscillation of hippocampal and neocotical neuronal ensembles which might be important for memory formation and consolidation. In this review the accumulated evidence from mouse mutant and pharmacological studies using gap junction blockers is summarized and the progress made in dissecting the physiological, pathophysiological and behavioral roles of gap junction mediated intercellular communication in the brain is discussed. Copyright © 2011 Elsevier Ltd. All rights reserved.

  18. The role of gap junctions in inflammatory and neoplastic disorders (Review).

    Science.gov (United States)

    Wong, Pui; Laxton, Victoria; Srivastava, Saurabh; Chan, Yin Wah Fiona; Tse, Gary

    2017-03-01

    Gap junctions are intercellular channels made of connexin proteins, mediating both electrical and biochemical signals between cells. The ability of gap junction proteins to regulate immune responses, cell proliferation, migration, apoptosis and carcinogenesis makes them attractive therapeutic targets for treating inflammatory and neoplastic disorders in different organ systems. Alterations in gap junction profile and expression levels are observed in hyperproliferative skin disorders, lymphatic vessel diseases, inflammatory lung diseases, liver injury and neoplastic disorders. It is now recognized that the therapeutic effects mediated by traditional pharmacological agents are dependent upon gap junction communication and may even act by influencing gap junction expression or function. Novel strategies for modulating the function or expression of connexins, such as the use of synthetic mimetic peptides and siRNA technology are considered.

  19. Emergent Central Pattern Generator Behavior in Gap-Junction-Coupled Hodgkin-Huxley Style Neuron Model

    Directory of Open Access Journals (Sweden)

    Kyle G. Horn

    2012-01-01

    Full Text Available Most models of central pattern generators (CPGs involve two distinct nuclei mutually inhibiting one another via synapses. Here, we present a single-nucleus model of biologically realistic Hodgkin-Huxley neurons with random gap junction coupling. Despite no explicit division of neurons into two groups, we observe a spontaneous division of neurons into two distinct firing groups. In addition, we also demonstrate this phenomenon in a simplified version of the model, highlighting the importance of afterhyperpolarization currents ( to CPGs utilizing gap junction coupling. The properties of these CPGs also appear sensitive to gap junction conductance, probability of gap junction coupling between cells, topology of gap junction coupling, and, to a lesser extent, input current into our simulated nucleus.

  20. Analyzing the Effects of Gap Junction Blockade on Neural Synchrony via a Motoneuron Network Computational Model

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    Heraldo Memelli

    2012-01-01

    Full Text Available In specific regions of the central nervous system (CNS, gap junctions have been shown to participate in neuronal synchrony. Amongst the CNS regions identified, some populations of brainstem motoneurons are known to be coupled by gap junctions. The application of various gap junction blockers to these motoneuron populations, however, has led to mixed results regarding their synchronous firing behavior, with some studies reporting a decrease in synchrony while others surprisingly find an increase in synchrony. To address this discrepancy, we employ a neuronal network model of Hodgkin-Huxley-style motoneurons connected by gap junctions. Using this model, we implement a series of simulations and rigorously analyze their outcome, including the calculation of a measure of neuronal synchrony. Our simulations demonstrate that under specific conditions, uncoupling of gap junctions is capable of producing either a decrease or an increase in neuronal synchrony. Subsequently, these simulations provide mechanistic insight into these different outcomes.

  1. Gap-junction channels inhibit transverse propagation in cardiac muscle

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    Ramasamy Lakshminarayanan

    2005-01-01

    Full Text Available Abstract The effect of adding many gap-junctions (g-j channels between contiguous cells in a linear chain on transverse propagation between parallel chains was examined in a 5 × 5 model (5 parallel chains of 5 cells each for cardiac muscle. The action potential upstrokes were simulated using the PSpice program for circuit analysis. Either a single cell was stimulated (cell A1 or the entire chain was stimulated simultaneously (A-chain. Transverse velocity was calculated from the total propagation time (TPT from when the first AP crossed a Vm of -20 mV and the last AP crossed -20 mV. The number of g-j channels per junction was varied from zero to 100, 1,000 and 10,000 (Rgj of ∞, 100 MΩ, 10 MΩ, 1.0 MΩ, respectively. The longitudinal resistance of the interstitial fluid (ISF space between the parallel chains (Rol2 was varied between 200 KΩ (standard value and 1.0, 5.0, and 10 MΩ. The higher the Rol2 value, the tighter the packing of the chains. It was found that adding many g-j channels inhibited transverse propagation by blocking activation of all 5 chains, unless Rol2 was greatly increased above the standard value of 200 KΩ. This was true for either method of stimulation. This was explained by, when there is strong longitudinal coupling between all 5 cells of a chain awaiting excitation, there must be more transfer energy (i.e., more current to simultaneously excite all 5 cells of a chain.

  2. 维甲酸对HeLa细胞间隙连接蛋白基因cx43表达的调节作用%The regulation effect of all-trans-retinoic acid on expression of connexin gene cx43 in HeLa

    Institute of Scientific and Technical Information of China (English)

    陈必良; 马向东; 王德堂; 辛晓燕

    1999-01-01

    目的:探讨分化诱导剂维甲酸对肿瘤抑制基因--细胞间隙连接蛋白基因cx43在人子宫颈癌细胞系HeLa中表达的调节作用.方法:应用核酸原位杂交、流式细胞仪、Westen blot及Lucifer Yellow划痕标记荧光传输技术,研究维甲酸作用对HeLa细胞cx43 mRNA及其蛋白表达,以及对HeLa细胞生长和通讯功能的调节作用.结果:HeLa细胞经维甲酸处理后,原位杂交显示,HeLa细胞cx43 mRNA水平上调;流式细胞仪分析,Cx43蛋白荧光强度增强,阳性细胞计数率由1.9%上升至26.3%;Wester blot检测到分子量约43KDa的Cx43蛋白表达;HeLa细胞生长明显受抑;细胞间隙连接通讯功能有所恢复.结论:维甲酸可通过上调肿瘤抑制基因cx43及其蛋白在HeLa细胞中的表达,实现对宫颈癌细胞恶性表型的逆转作用,这可能是维甲酸肿瘤抑制作用的重要机制之一.

  3. Effects of different types of Helicobacter pylori on expressions of Cx32 and Cx43 in GES-1 cells%不同类型幽门螺杆菌对GES-1细胞Cx32、Cx43表达的影响

    Institute of Scientific and Technical Information of China (English)

    徐灿霞; 陈玉林; 邹惠芳; 陈雄

    2011-01-01

    目的 观察不同类型幽门螺杆菌(Helicobacter pylori,H.pylori)对人胃黏膜上皮细胞系GES-1细胞间隙连接蛋白(Connexin,Cx)32和43表达的影响,探讨与Cx32、Cx43表达异常有关的H.pylori菌株类型.方法 将临床培养分离的不同H.pylori菌株类型包括东亚型CagA+ H.pylori、西方型CagA+H.pylori及CagA-H.pylori与GES-1细胞共培养24 h及48 h,对照组不加H.pylori培养24 h及48 h.采用间接免疫荧光方法(IIF)及计算机图像分析技术检测GES-1细胞Cx32、Cx43表达.结果 对照组24 h和48 h及加H.pylori各组24 h GES-1细胞Cx32、Cx43表达阳性率均为100%,东亚型CagA+H.pylori组48 h Cx32、Cx43表达阳性率均低于对照组、CagA-H.pylori组和西方型CagA+H.Pylori组(P<0.05);对照组24 h和48 h Cx32、Cx43绿色荧光位于细胞膜,西方型CagA+H.pylori组和东亚型CagA+H,pylori组24 h和48 h Cx32绿色荧光大部分位于细胞膜,少部分位于细胞浆,Cx43绿色荧光大部分位于细胞浆,少部分位于细胞膜;东亚型CagA+H.pylori组和西方型Cag+H,pylori组24 h及48 hCx32、Cx43表达强度低于对照组和CagA-H.pylori组(P<0.05),且东亚型CagA+H.pylori组较西方型CagA+H.pylori组减弱更明显(P<0.05).结论 H.pylori下调GES-1细胞Cx32、Cx43表达,以CagA+H.pylori菌株特别是东亚型CagA+ H.pylori菌株作用更明显.%Objective To study the effects of different types of H. pylori on the expressions of Connexin(Cx) 32 and Cx43 in human gastric epithelial cell line GES-1 cells, and investigate the types of H. pylorirelated to the expressions of Cx32 and Cx43. Method Different types of clinical H.pylori strains were isolated and cultured, including the EasternAsian type CagA + H. pylori, the Western type CagA + H.pylori, and the CngA- H.pylori. These H.pylori strains were cocultured with GES-1 cells for 24 hours and 48 hours, respectively. The control groups were cultured without any H. pylori for 24 hours and 48 hours. The indirect immunofluorescence

  4. Treatment with the gap junction modifier rotigaptide (ZP123) reduces infarct size in rats with chronic myocardial infarction

    DEFF Research Database (Denmark)

    Haugan, Ketil; Marcussen, Niels; Kjølbye, Anne Louise;

    2006-01-01

    Treatment with non-selective drugs (eg, long-chain alcohols, halothane) that reduce gap junction intercellular communication (GJIC) is associated with reduced infarct size after myocardial infarction (MI). Therefore, it has been suggested that gap junction intercellular communication stimulating ...

  5. Local dynamics of gap-junction-coupled interneuron networks

    Science.gov (United States)

    Lau, Troy; Gage, Gregory J.; Berke, Joshua D.; Zochowski, Michal

    2010-03-01

    Interneurons coupled by both electrical gap-junctions (GJs) and chemical GABAergic synapses are major components of forebrain networks. However, their contributions to the generation of specific activity patterns, and their overall contributions to network function, remain poorly understood. Here we demonstrate, using computational methods, that the topological properties of interneuron networks can elicit a wide range of activity dynamics, and either prevent or permit local pattern formation. We systematically varied the topology of GJ and inhibitory chemical synapses within simulated networks, by changing connection types from local to random, and changing the total number of connections. As previously observed we found that randomly coupled GJs lead to globally synchronous activity. In contrast, we found that local GJ connectivity may govern the formation of highly spatially heterogeneous activity states. These states are inherently temporally unstable when the input is uniformly random, but can rapidly stabilize when the network detects correlations or asymmetries in the inputs. We show a correspondence between this feature of network activity and experimental observations of transient stabilization of striatal fast-spiking interneurons (FSIs), in electrophysiological recordings from rats performing a simple decision-making task. We suggest that local GJ coupling enables an active search-and-select function of striatal FSIs, which contributes to the overall role of cortical-basal ganglia circuits in decision-making.

  6. Gap Junctions in the Ventral Hippocampal-Medial Prefrontal Pathway Are Involved in Anxiety Regulation

    Science.gov (United States)

    Schoenfeld, Timothy J.; Kloth, Alexander D.; Hsueh, Brian; Runkle, Matthew B.; Kane, Gary A.; Wang, Samuel S.-H.

    2014-01-01

    Anxiety disorders are highly prevalent but little is known about their underlying mechanisms. Gap junctions exist in brain regions important for anxiety regulation, such as the ventral hippocampus (vHIP) and mPFC, but their functions in these areas have not been investigated. Using pharmacological blockade of neuronal gap junctions combined with electrophysiological recordings, we found that gap junctions play a role in theta rhythm in the vHIP and mPFC of adult mice. Bilateral infusion of neuronal gap junction blockers into the vHIP decreased anxiety-like behavior on the elevated plus maze and open field. Similar anxiolytic effects were observed with unilateral infusion of these drugs into the vHIP combined with contralateral infusion into the mPFC. No change in anxious behavior was observed with gap junction blockade in the unilateral vHIP alone or in the bilateral dorsal HIP. Since physical exercise is known to reduce anxiety, we examined the effects of long-term running on the expression of the neuronal gap junction protein connexin-36 among inhibitory interneurons and found a reduction in the vHIP. Despite this change, we observed no alteration in theta frequency or power in long-term runners. Collectively, these findings suggest that neuronal gap junctions in the vHIP–mPFC pathway are important for theta rhythm and anxiety regulation under sedentary conditions but that additional mechanisms are likely involved in running-induced reduction in anxiety. PMID:25411496

  7. Cisplatin-induced premature senescence with concomitant reduction of gap junctions in human fibroblasts

    Institute of Scientific and Technical Information of China (English)

    Wei ZHAO; Zhong Xiang LIN; Zhi Qian ZHANG

    2004-01-01

    To examine the role of gap junctions in cell senescence,the changes of gap junctions in cisplatin-induced premature senescence of primary cultured fibroblasts were studied and compared with the replicative senescent human fibroblasts.Dye transfer assay for gap junction function and immunofluorescent staining for connexin 43 protein distribution were done respectively. Furthermore,cytofluorimetry and DAPI fluorescence staining were performed for cell cycle and apoptosis analysis. p53 gene expression level was detected with indirect immunofluorescence. We found that cisplatin (10 mM) treatment could block cell growth cycle at G1 and induced premature senescence. The premature senescence changes included high frequency of apoptosis,elevation of p53 expression,loss of membranous gap junctions and reduction of dye-transfer capacity. These changes were comparable to the changes of replicative senescence of human fibroblasts. It was also concluded that cisplatin could induce premature senescence concomitant with inhibition of gap junctions in the fibroblasts. Loss of functional gap junctions from the cell membrane may account for the reduced intercellular communication in the premature senescent fibroblasts. The cell system we used may provide a model useful for the study of the gap junction thus promoting agents against premature senescence.

  8. Identifying connexin expression and determining gap junction intercellular communication in rainbow trout cells.

    Science.gov (United States)

    Hooper, Joshua; Poynter, Sarah J; DeWitte-Orr, Stephanie J

    2017-05-01

    Gap junctions are groups of membrane-bound channels that allow the passage of small molecules and ions between cells, permitting cell-cell communication. Because of their importance in cell homeostasis, gap junction presence and function were characterized in three commonly studied rainbow trout cell lines, namely RTgill-W1, RTgutGC, and RTG-2. Firstly, gap junction presence was determined by screening for gap junction protein alpha 7 and alpha 1 (GJA7 and GJA1) presence at the transcript level and GJA7 at the protein level. GJA7 was successfully identified at both the transcript and protein levels, and GJA1 was detected at the transcript level in all three cell lines. This is the first report of a GJA7 full-length transcript sequence in rainbow trout cells. Gap junction function, as determined by gap junction intercellular communication (GJIC), was examined using Lucifer yellow dye migration with the scrape and load technique; visualized by fluorescence microscopy. Phorbol 12-myristate 13-acetate (PMA), a gap junction inhibitor, was used to confirm the presence of functional gap junctions. Effects of serum deprivation on GJIC were also monitored; 24-h serum deprivation resulted in greater dye migration compared with 30-min serum deprivation. Both RTG-2 and RTgill-W1 showed significant dye migration that was inhibited by PMA while RTgutGC did not. Human foreskin fibroblast (HFF-1) cells were used as a positive control for gap junction presence and function. Taken together, our study shows that rainbow trout cells express connexin transcripts and proteins, and RTG-2 and, to a lesser extent, RTgill-W1 cells are able to perform GJIC.

  9. Prenatal nicotine exposure enhances Cx43 and Panx1 unopposed channel activity in brain cells of adult offspring mice fed a high-fat/cholesterol diet

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    Juan Andrés Orellana

    2014-12-01

    Full Text Available Nicotine, the most important neuroteratogen of tobacco smoke, can reproduce brain and cognitive disturbances per se when administered prenatally. However, it is still unknown if paracrine signaling among brain cells participates in prenatal nicotine-induced brain impairment of adult offspring. Paracrine signaling is partly mediated by unopposed channels formed by connexins (hemichannels and pannexins serving as aqueous pores permeable to ions and small signaling molecules, allowing exchange between the intra- and extracellular milieus. Our aim was to address whether prenatal nicotine exposure changes the activity of those channels in adult mice offspring under control conditions or subjected to a second challenge during young ages: high-fat/cholesterol (HFC diet. To induce prenatal exposure to nicotine, osmotic minipumps were implanted in CF1 pregnant mice at gestational day 5 to deliver nicotine bitartrate or saline (control solutions. After weaning, offspring of nicotine-treated or untreated pregnant mice were fed ad libitum with chow or HFC diets for 8 weeks. The functional state of Cx43 and Panx1 unopposed channels was evaluated by dye uptake experiments in hippocampal slices from 11-week-old mice. We found that prenatal nicotine increased the opening of Cx43 hemichannels in astrocytes, and Panx1 channels in microglia and neurons only if offspring mice were fed with HFC diet. Blockade of iNOS, COX2 and EP1, P2X7 and NMDA receptors, showed differential inhibition of prenatal nicotine-induced channel opening in glial cells and neurons. Importantly, inhibition of the above mentioned enzymes and receptors, or blockade of Cx43 and Panx1 unopposed channels greatly reduced ATP and glutamate release from hippocampal slices of prenatally nicotine-exposed offspring. We propose that unregulated gliotransmitter release through Cx43 and Panx1 unopposed channels may participate in brain alterations observed in offspring of mothers exposed to tobacco smoke

  10. Abundance of gap junctions at glutamatergic mixed synapses in adult Mosquitofish spinal cord neurons

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    Jose L Serrano-Velez

    2014-06-01

    Full Text Available Dye-coupling, whole-mount immunohistochemistry for gap junction channel protein connexin 35 (Cx35, and freeze-fracture replica immunogold labeling (FRIL reveal an abundance of electrical synapses/gap junctions at glutamatergic mixed synapses in the 14th spinal segment that innervates the adult male gonopodium of Western Mosquitofish, Gambusia affinis (Mosquitofish.To study gap junctions’ role in fast motor behavior, we used a minimally-invasive neural-tract-tracing technique to introduce gap junction-permeant or -impermeant dyes into deep muscles controlling the gonopodium of the adult male Mosquitofish, a teleost fish that rapidly transfers (complete in 50 of the 62 gap junctions at mixed synapses are in the 14th spinal segment.Our results support and extend studies showing gap junctions at mixed synapses in spinal cord segments involved in control of genital reflexes in rodents, and they suggest a link between mixed synapses and fast motor behavior. The findings provide a basis for studies of specific roles of spinal neurons in the generation/regulation of sex-specific behavior and for studies of gap junctions’ role in regulating fast motor behavior. Finally, the CoPA IN provides a novel candidate neuron for future studies of gap junctions and neural control of fast motor behaviors.

  11. Biophysical characteristics of gap junctions in vascular wall cells: implications for vascular biology and disease

    Directory of Open Access Journals (Sweden)

    P.R. Brink

    2000-04-01

    Full Text Available The role gap junction channels play in the normal and abnormal functioning of the vascular wall is the subject of much research. The biophysical properties of gap junctions are an essential component in understanding how gap junctions function to allow coordinated relaxation and contraction of vascular smooth muscle. This study reviews the properties thus far elucidated and relates those properties to tissue function. We ask how biophysical and structural properties such as gating, permselectivity, subconductive states and channel type (heteromeric vs homotypic vs heterotypic might affect vascular smooth muscle tone.

  12. Direct Exposure to Ethanol Disrupts Junctional Cell-Cell Contact and Hippo-YAP Signaling in HL-1 Murine Atrial Cardiomyocytes.

    Directory of Open Access Journals (Sweden)

    Kanako Noritake

    Full Text Available Direct exposure of cardiomyocytes to ethanol causes cardiac damage such as cardiac arrythmias and apoptotic cell death. Cardiomyocytes are connected to each other through intercalated disks (ID, which are composed of a gap junction (GJ, adherens junction, and desmosome. Changes in the content as well as the subcellular localization of connexin43 (Cx43, the main component of the cardiac GJ, are reportedly involved in cardiac arrythmias and subsequent damage. Recently, the hippo-YAP signaling pathway, which links cellular physical status to cell proliferation, differentiation, and apoptosis, has been implicated in cardiac homeostasis under physiological as well as pathological conditions. This study was conducted to explore the possible involvement of junctional intercellular communication, mechanotransduction through cytoskeletal organization, and the hippo-YAP pathway in cardiac damage caused by direct exposure to ethanol. HL-1 murine atrial cardiac cells were used since these cells retain cardiac phenotypes through ID formation and subsequent synchronous contraction. Cells were exposed to 0.5-2% ethanol; significant apoptotic cell death was observed after exposure to 2% ethanol for 48 hours. A decrease in Cx43 levels was already observed after 3 hours exposure to 2% ethanol, suggesting a rapid degradation of this protein. Upon exposure to ethanol, Cx43 translocated into lysosomes. Cellular cytoskeletal organization was also dysregulated by ethanol, as demonstrated by the disruption of myofibrils and intermediate filaments. Coinciding with the loss of cell-cell adherence, decreased phosphorylation of YAP, a hippo pathway effector, was also observed in ethanol-treated cells. Taken together, the results provide evidence that cells exposed directly to ethanol show 1 impaired cell-cell adherence/communication, 2 decreased cellular mechanotransduction by the cytoskeleton, and 3 a suppressed hippo-YAP pathway. Suppression of hippo-YAP pathway

  13. The effects of FuFangTengLiGen preparation regulating the expressions of Cx43 and E-cad of transplantation tumor of PG cell%复方藤梨根制剂对PG荷瘤裸鼠Cx43基因表达水平及E-钙粘附素影响的研究

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    Objective: To study the anticancer effect of Chinese compound recipe FuFangTengLiGen (FFTLG) against transplantation tumor of PG cell and preliminarily explore the connection with connexin43 (Cx43) and epithelial cadherin (E-cad).Methods: Model rats of transplantation tumor of human large cell cancer PG were established. The low, medium and high dose groups of FFTLG, the blank control group were set up. After 21 days medication of FFTLG through gastrogavage, the antitumor effect of each group was compared. The gene expressions of Cx43 and E-cad in each group were quantitatively detected and compared by the application of immunohistochemistry technique. Results: FFTLG could significantly inhibit the growth of transplantation tumor. FFTLG could obviously up-regulate the expressions of Cx43 and E-cad, which in the low/medium and high dose groups were separately higher than that in the blank group. Conclusion: FFTLG has definite antitumor effect against human large cell cancer and can obviously up-regulate the expressions of Cx43 and E-cad, which may be correlated with its effect of anti-tumor.

  14. Up-Regulation Effect of 8-Br-cAMP on Expression of Connexin Gene Cx43 in Tongue Carcinoma Cell Line%8-Br-cAMP上调舌癌细胞株间隙连接蛋白基因Cx43表达的研究

    Institute of Scientific and Technical Information of China (English)

    王安训; 黄洪章; 孔庆瑜

    2001-01-01

    目的:探讨8-Br-cAMP(8-Bromo3':5'-cyclic adenosine monophosphate)对舌癌细胞株(Tca8113)细胞间隙连接蛋白基因Cx43表达的调节作用。方法:应用MTT法检测8-Br-cAMP对舌癌细胞株生长的抑制作用;应用免疫细胞化学、流式细胞仪等技术,对8-Br-cAMP诱导舌癌细胞株细胞间隙连接蛋白基因Cx43的表达进行分析。结果:舌癌细胞经10-5mol/L、10-4mol/L、10-3mol/L8-Br-cAMP处理后,细胞生长受抑制,其抑制率随浓度升高而升高;免疫细胞化学检测可见Cx43蛋白的表达明显提高;流式细胞仪分析,Cx43蛋白荧光强度增强,阳性细胞计数率由4.8%上升至26.5%,经t检验两组间存在显著性差异(P<0.01)。结论:8-Br-cAMP对舌癌细胞的抑制作用可能通过上调Cx43蛋白的表达而实现。?%Objectives:This study was designed to investigate the regulation effect of 8-Br-cAMP ( 8-Bromo 3′ :5′ -cyclic adenosine monophosphate) on the expression of connexin gene (Cx43) in human tongue carcinoma cell line (Tca8113) . Methods:The inhibitory effect of 8-Br-cAMP on growth of Tca8113 cell line was examined by methyl thiazolyl tetrazolium (MTT) assay. Cx43 gene expression in Tca8113 cells after treated with 8-Br-cAMP was analyzed by immunocytochemistry and flow cytometry . Results:After treated with 8-Br-cAMP(10-5 mol/L, 10-4 mol/L, 10-3 mol/L), Tca8113 cells was inhibited and the inhibition rate was increased with the concentration of 8-Br-cAMP; Immunocytochemistry showed Cx43 protein detectability and up-regulation in 8-Br-cAMP treated cells; The positive rate of Cx43 protein in Tca8113 ce1ls increased from 4.8% to 26.5% . There was significant difference between 8-Br-cAMP treated cells and untreated cells ( P<0.01) . Conclusion: The anti-tumor effect of 8-Br-cAMP on Tca8113 cells might be due to up-regu1ation on Cx43 gene.

  15. Modelling the effect of gap junctions on tissue-level cardiac electrophysiology

    CERN Document Server

    Bruce, Doug; Whiteley, Jonathan P

    2012-01-01

    When modelling tissue-level cardiac electrophysiology, continuum approximations to the discrete cell-level equations are used to maintain computational tractability. One of the most commonly used models is represented by the bidomain equations, the derivation of which relies on a homogenisation technique to construct a suitable approximation to the discrete model. This derivation does not explicitly account for the presence of gap junctions connecting one cell to another. It has been seen experimentally [Rohr, Cardiovasc. Res. 2004] that these gap junctions have a marked effect on the propagation of the action potential, specifically as the upstroke of the wave passes through the gap junction. In this paper we explicitly include gap junctions in a both a 2D discrete model of cardiac electrophysiology, and the corresponding continuum model, on a simplified cell geometry. Using these models we compare the results of simulations using both continuum and discrete systems. We see that the form of the action potent...

  16. Innexin gap junctions in nerve cells coordinate spontaneous contractile behavior in Hydra polyps

    Science.gov (United States)

    Takaku, Yasuharu; Hwang, Jung Shan; Wolf, Alexander; Böttger, Angelika; Shimizu, Hiroshi; David, Charles N.; Gojobori, Takashi

    2014-01-01

    Nerve cells and spontaneous coordinated behavior first appeared near the base of animal evolution in the common ancestor of cnidarians and bilaterians. Experiments on the cnidarian Hydra have demonstrated that nerve cells are essential for this behavior, although nerve cells in Hydra are organized in a diffuse network and do not form ganglia. Here we show that the gap junction protein innexin-2 is expressed in a small group of nerve cells in the lower body column of Hydra and that an anti-innexin-2 antibody binds to gap junctions in the same region. Treatment of live animals with innexin-2 antibody eliminates gap junction staining and reduces spontaneous body column contractions. We conclude that a small subset of nerve cells, connected by gap junctions and capable of synchronous firing, act as a pacemaker to coordinate the contraction of the body column in the absence of ganglia.

  17. The gap junction blocker carbenoxolone enhances propofol and sevoflurane-induced loss of consciousness

    Institute of Scientific and Technical Information of China (English)

    Zhigang Liu; Yongfang Liu; Bo Zhao; Li Du; Zhongyuan Xia; Xiangdong Chen; Tao Luo

    2012-01-01

    General anesthetics induce loss of consciousness by inhibiting ascending arousal pathways, and they interfere with gap junction electrical coupling.The present study aimed to determine whether inhibition of gap junction-mediated signaling could influence general anesthetic-induced loss of consciousness.The general anesthetics sevoflurane and propofol were used.Intracerebroventricular administration of carbenoxolone, a gap junction blocker, significantly decreased the time to loss of the righting reflex (P < 0.05), but prolonged the time to recovery of the reflex (P < 0.05).Moreover, intracerebroventricular administration of carbenoxolone increased the sensitivity to sevoflurane, with a leftward shift of the loss of righting reflex dose-response curve, and decreased the 50% effective concentration of sevoflurane.These results suggest that the gap junction blocker carbenoxolone enhances propofol and sevoflurane-mediated general anesthesia.

  18. Innexin gap junctions in nerve cells coordinate spontaneous contractile behavior in Hydra polyps

    KAUST Repository

    Takaku, Yasuharu

    2014-01-07

    Nerve cells and spontaneous coordinated behavior first appeared near the base of animal evolution in the common ancestor of cnidarians and bilaterians. Experiments on the cnidarian Hydra have demonstrated that nerve cells are essential for this behavior, although nerve cells in Hydra are organized in a diffuse network and do not form ganglia. Here we show that the gap junction protein innexin-2 is expressed in a small group of nerve cells in the lower body column of Hydra and that an anti-innexin-2 antibody binds to gap junctions in the same region. Treatment of live animals with innexin-2 antibody eliminates gap junction staining and reduces spontaneous body column contractions. We conclude that a small subset of nerve cells, connected by gap junctions and capable of synchronous firing, act as a pacemaker to coordinate the contraction of the body column in the absence of ganglia.

  19. Synopsis of the International Gap Junction Conference in Elsinore, Denmark August 5-9, 2007

    DEFF Research Database (Denmark)

    Willecke, Klaus; Nielsen, Morten Schak

    2008-01-01

    This synopsis covers the main results and conclusions from the platform presentations during the International Gap Junction Conference. More detailed information is provided in the mini reviews on controversial scientific issues, short reports of research results and conference abstracts publishe...

  20. The Microvascular Gap Junction Channel: A Route to Deliver MicroRNAs for Neurological Disease Treatment.

    Science.gov (United States)

    Thuringer, Dominique; Solary, Eric; Garrido, Carmen

    2017-01-01

    Brain microvascular endothelial cells (BMECs) separate the peripheral blood from the brain. These cells, which are surrounded by basal lamina, pericytes and glial cells, are highly interconnected through tight and gap junctions. Their permeability properties restrict the transfer of potentially useful therapeutic agents. In such a hermetic system, the gap junctional exchange of small molecules between cerebral endothelial and non-endothelial cells is crucial for maintaining tissue homeostasis. MicroRNA were shown to cross gap junction channels, thereby modulating gene expression and function of the recipient cell. It was also shown that, when altered, BMEC could be regenerated by endothelial cells derived from pluripotent stem cells. Here, we discuss the transfer of microRNA through gap junctions between BMEC, the regeneration of BMEC from induced pluripotent stem cells that could be engineered to express specific microRNA, and how such an innovative approach could benefit to the treatment of glioblastoma and other neurological diseases.

  1. The Microvascular Gap Junction Channel: A Route to Deliver MicroRNAs for Neurological Disease Treatment

    Directory of Open Access Journals (Sweden)

    Dominique Thuringer

    2017-08-01

    Full Text Available Brain microvascular endothelial cells (BMECs separate the peripheral blood from the brain. These cells, which are surrounded by basal lamina, pericytes and glial cells, are highly interconnected through tight and gap junctions. Their permeability properties restrict the transfer of potentially useful therapeutic agents. In such a hermetic system, the gap junctional exchange of small molecules between cerebral endothelial and non-endothelial cells is crucial for maintaining tissue homeostasis. MicroRNA were shown to cross gap junction channels, thereby modulating gene expression and function of the recipient cell. It was also shown that, when altered, BMEC could be regenerated by endothelial cells derived from pluripotent stem cells. Here, we discuss the transfer of microRNA through gap junctions between BMEC, the regeneration of BMEC from induced pluripotent stem cells that could be engineered to express specific microRNA, and how such an innovative approach could benefit to the treatment of glioblastoma and other neurological diseases.

  2. Structure of the gap junction channel and its implications for its biological functions.

    Science.gov (United States)

    Maeda, Shoji; Tsukihara, Tomitake

    2011-04-01

    Gap junctions consist of arrays of intercellular channels composed of integral membrane proteins called connexin in vertebrates. Gap junction channels regulate the passage of ions and biological molecules between adjacent cells and, therefore, are critically important in many biological activities, including development, differentiation, neural activity, and immune response. Mutations in connexin genes are associated with several human diseases, such as neurodegenerative disease, skin disease, deafness, and developmental abnormalities. The activity of gap junction channels is regulated by the membrane voltage, intracellular microenvironment, interaction with other proteins, and phosphorylation. Each connexin channel has its own property for conductance and molecular permeability. A number of studies have tried to reveal the molecular architecture of the channel pore that should confer the connexin-specific permeability/selectivity properties and molecular basis for the gating and regulation. In this review, we give an overview of structural studies and describe the structural and functional relationship of gap junction channels.

  3. Nanostructured thin films for multiband-gap silicon triple junction solar cells

    NARCIS (Netherlands)

    Schropp, R.E.I.; Li, H. B. T.; Franken, R.H.; Rath, J.K.; van der Werf, C.H.M.; Schuttauf, J.A.; Stolk, R.L.

    2008-01-01

    By implementing nanostructure in multiband-gap proto-Si/proto-SiGe/nc-Si:H triple junction n–i–p solar cells, a considerable improvement in performance has been achieved. The unalloyed active layers in the top and bottom cell of these triple junction cells are deposited by Hot-Wire CVD. A significan

  4. The Role of Chemical Inhibition of Gap Junctional Intercellular Communication in Toxicology.

    Science.gov (United States)

    1988-02-14

    hypothesis that chemical modulation of gap junctional intercellular communication can lead to many toxic endpoints, such as teratogenesis , tumor promotion...goal has been to test this hypothesis, namely, that chemical modulation of gap junctional intercellular communication can lead to teratogenesis ...of dium pyruvate, and 10% fetal calf serum. Under the in- low-molecular-weight radioactive labeled cubation condition with 5% CO 2 in humidified air

  5. Gap Junction Enhancer Potentiates Cytotoxicity of Cisplatin in Breast Cancer Cells

    OpenAIRE

    Ding, Ying; Nguyen, Thu Annelise

    2012-01-01

    Cisplatin is one of the most widely used anti-cancer drugs due to its ability to damage DNA and induce apoptosis. However, increasing reports of side effects and drug resistance indicate the limitation of cisplatin in cancer therapeutics. Recent studies showed that inhibition of gap junctions diminishes the cytotoxic effect and contributes to drug resistance. Therefore, identification of molecules that counteract gap junctional inhibition without decreasing the anti-cancer effect of cisplatin...

  6. Berberine potentizes apoptosis induced by X-rays irradiation probably through modulation of gap junctions

    Institute of Scientific and Technical Information of China (English)

    LIU Bing; WANG Qin; YUAN Dong-dong; HONG Xiao-ting; TAO Liang

    2011-01-01

    Background Clinical combination of some traditional Chinese medical herbs, including berberine, with irradiation is demonstrated to improve efficacy of tumor radiotherapy, yet the mechanisms for such effect remain largely unknown. The present study investigated the effect of berberine on apoptosis induced by X-rays irradiation and the relation between this effect and gap junction intercellular communication (GJIC).Methods The role of gap junctions in the modulation of X-rays irradiation-induced apoptosis was explored by manipulation of connexin (Cx) expression, and gap junction function, using oleamide, a GJIC inhibitor, and berberine.Results In transfected HeLa cells, Cx32 expression increased apoptosis induced by X-rays irradiation, while inhibition of gap junction by oleamide reduced the irradiation responses, indicating the dependence of X-rays irradiation-induced apoptosis on GJIC. Berberine, at the concentrations without cytotoxicity, enhanced apoptosis induced by irradiation only in the presence of functional gap junctions.Conclusions These results suggest that berberine potentizes cell apoptosis induced by X-rays irradiation, probably through enhancement of gap junction activity.

  7. Dissection of neuronal gap junction circuits that regulate social behavior in Caenorhabditis elegans

    Science.gov (United States)

    Jang, Heeun; Levy, Sagi; Flavell, Steven W.; Mende, Fanny; Latham, Richard; Zimmer, Manuel; Bargmann, Cornelia I.

    2017-01-01

    A hub-and-spoke circuit of neurons connected by gap junctions controls aggregation behavior and related behavioral responses to oxygen, pheromones, and food in Caenorhabditis elegans. The molecular composition of the gap junctions connecting RMG hub neurons with sensory spoke neurons is unknown. We show here that the innexin gene unc-9 is required in RMG hub neurons to drive aggregation and related behaviors, indicating that UNC-9–containing gap junctions mediate RMG signaling. To dissect the circuit in detail, we developed methods to inhibit unc-9–based gap junctions with dominant-negative unc-1 transgenes. unc-1(dn) alters a stomatin-like protein that regulates unc-9 electrical signaling; its disruptive effects can be rescued by a constitutively active UNC-9::GFP protein, demonstrating specificity. Expression of unc-1(dn) in RMG hub neurons, ADL or ASK pheromone-sensing neurons, or URX oxygen-sensing neurons disrupts specific elements of aggregation-related behaviors. In ADL, unc-1(dn) has effects opposite to those of tetanus toxin light chain, separating the roles of ADL electrical and chemical synapses. These results reveal roles of gap junctions in a complex behavior at cellular resolution and provide a tool for similar exploration of other gap junction circuits. PMID:28143932

  8. Pharmacology of gap junctions. New pharmacological targets for treatment of arrhythmia, seizure and cancer?

    Science.gov (United States)

    Salameh, Aida; Dhein, Stefan

    2005-12-20

    Intercellular communication in many organs is maintained via intercellular gap junction channels composed of connexins, a large protein family with a number of isoforms. This gap junction intercellular communication (GJIC) allows the propagation of action potentials (e.g., in brain, heart), and the transfer of small molecules which may regulate cell growth, differentiation and function. The latter has been shown to be involved in cancer growth: reduced GJIC often is associated with increased tumor growth or with de-differentiation processes. Disturbances of GJIC in the heart can cause arrhythmia, while in brain electrical activity during seizures seems to be propagated via gap junction channels. Many diseases or pathophysiological conditions seem to be associated with alterations of gap junction protein expression. Thus, depending on the target disease opening or closure of gap junctions may be of interest, or alteration of connexin expression. GJIC can be affected acutely by changing gap junction conductance or--more chronic--by altering connexin expression and membrane localisation. This review gives an overview on drugs affecting GJIC.

  9. Inter-band phase fluctuations in macroscopic quantum tunneling of multi-gap superconducting Josephson junctions

    Energy Technology Data Exchange (ETDEWEB)

    Asai, Hidehiro, E-mail: hd-asai@aist.go.jp [Electronics and Photonics Research Institute (ESPRIT), National Institute of Advanced Industrial Science and Technology (AIST), Tsukuba, Ibaraki 305-8568 (Japan); Ota, Yukihiro [CCSE, Japan Atomic Energy Agency, Kashiwa, Chiba 277-8587 (Japan); Kawabata, Shiro [Electronics and Photonics Research Institute (ESPRIT), National Institute of Advanced Industrial Science and Technology (AIST), Tsukuba, Ibaraki 305-8568 (Japan); Nori, Franco [CEMS, RIKEN, Wako-shi, Saitama 351-0198 (Japan); Physics Department, University of Michigan, Ann Arbor, MI 48109-1040 (United States)

    2014-09-15

    Highlights: • We study MQT in Josephson junctions composed of multi-gap superconductors. • We derive a formula of the MQT escape rate for multiple phase differences. • We investigate the effect of inter-band phase fluctuation on MQT. • The MQT escape rate is significantly enhanced by the inter-band phase fluctuation. - Abstract: We theoretically investigate macroscopic quantum tunneling (MQT) in a hetero Josephson junction formed by a conventional single-gap superconductor and a multi-gap superconductor. In such Josephson junctions, phase differences for each tunneling channel are defined, and the fluctuation of the relative phase differences appear which is referred to as Josephson–Leggett’s mode. We take into account the effect of the fluctuation in the tunneling process and calculate the MQT escape rate for various junction parameters. We show that the fluctuation of relative phase differences drastically enhances the escape rate.

  10. The Role of Chemical Inhibition of Gap-Junctional Intercellular Communication in Toxicology

    Science.gov (United States)

    1990-03-31

    Kalimi, "Chemical and oncoqene modulation of intercellular communication during carcinogenesis". Symposium on Molecular Cell Biology of Liver Growth...gap junct 4 onal communication during carcinoqgenesis". Molecular Cell Biology of Gap Junctions symposium, Irsee, Germany, July 18-23, 1989. 8

  11. Impact of 7,12-dimethylbenz[a]anthracene exposure on connexin gap junction proteins in cultured rat ovaries

    Energy Technology Data Exchange (ETDEWEB)

    Ganesan, Shanthi, E-mail: shanthig@iastate.edu; Keating, Aileen F., E-mail: akeating@iastate.edu

    2014-01-15

    7,12-Dimethylbenz[a]anthracene (DMBA) destroys ovarian follicles in a concentration-dependent manner. The impact of DMBA on connexin (CX) proteins that mediate communication between follicular cell types along with pro-apoptotic factors p53 and Bax were investigated. Postnatal day (PND) 4 Fisher 344 rat ovaries were cultured for 4 days in vehicle medium (1% DMSO) followed by a single exposure to vehicle control (1% DMSO) or DMBA (12.5 nM or 75 nM) and cultured for 4 or 8 days. RT-PCR was performed to quantify Cx37, Cx43, p53 and Bax mRNA level. Western blotting and immunofluorescence staining were performed to determine CX37 or CX43 level and/or localization. Cx37 mRNA and protein increased (P < 0.05) at 4 days of 12.5 nM DMBA exposure. Relative to vehicle control-treated ovaries, mRNA encoding Cx43 decreased (P < 0.05) but CX43 protein increased (P < 0.05) at 4 days by both DMBA exposures. mRNA expression of pro-apoptotic p53 was decreased (P < 0.05) but no changes in Bax expression were observed after 4 days of DMBA exposures. In contrast, after 8 days, DMBA decreased Cx37 and Cx43 mRNA and protein but increased both p53 and Bax mRNA levels. CX43 protein was located between granulosa cells, while CX37 was located at the oocyte cell surface of all follicle stages. These findings support that DMBA exposure impacts ovarian Cx37 and Cx43 mRNA and protein prior to both observed changes in pro-apoptotic p53 and Bax and follicle loss. It is possible that such interference in follicular cell communication is detrimental to follicle viability, and may play a role in DMBA-induced follicular atresia. - Highlights: • DMBA increases Cx37 and Cx43 expression prior to follicle loss. • During follicle loss both Cx37 and Cx43 expressions are reduced. • CX43 protein is absent in follicle remnants lacking an oocyte.

  12. Prenatal nicotine exposure enhances Cx43 and Panx1 unopposed channel activity in brain cells of adult offspring mice fed a high-fat/cholesterol diet.

    Science.gov (United States)

    Orellana, Juan A; Busso, Dolores; Ramírez, Gigliola; Campos, Marlys; Rigotti, Attilio; Eugenín, Jaime; von Bernhardi, Rommy

    2014-01-01

    Nicotine, the most important neuroteratogen of tobacco smoke, can reproduce brain and cognitive disturbances per se when administered prenatally. However, it is still unknown if paracrine signaling among brain cells participates in prenatal nicotine-induced brain impairment of adult offspring. Paracrine signaling is partly mediated by unopposed channels formed by connexins hemichannels (HCs) and pannexins serving as aqueous pores permeable to ions and small signaling molecules, allowing exchange between the intra- and extracellular milieus. Our aim was to address whether prenatal nicotine exposure changes the activity of those channels in adult mice offspring under control conditions or subjected to a second challenge during young ages: high-fat/cholesterol (HFC) diet. To induce prenatal exposure to nicotine, osmotic minipumps were implanted in CF1 pregnant mice at gestational day 5 to deliver nicotine bitartrate or saline (control) solutions. After weaning, offspring of nicotine-treated or untreated pregnant mice were fed ad libitum with chow or HFC diets for 8 weeks. The functional state of connexin 43 (Cx43) and pannexin 1 (Panx1) unopposed channels was evaluated by dye uptake experiments in hippocampal slices from 11-week-old mice. We found that prenatal nicotine increased the opening of Cx43 HCs in astrocytes, and Panx1 channels in microglia and neurons only if offspring mice were fed with HFC diet. Blockade of inducible nitric oxide synthase (iNOS), cyclooxygenase 2 (COX2) and prostaglandin E receptor 1 (EP1), ionotropic ATP receptor type 7 (P2X7) and NMDA receptors, showed differential inhibition of prenatal nicotine-induced channel opening in glial cells and neurons. Importantly, inhibition of the above mentioned enzymes and receptors, or blockade of Cx43 and Panx1 unopposed channels greatly reduced adenosine triphosphate (ATP) and glutamate release from hippocampal slices of prenatally nicotine-exposed offspring. We propose that unregulated gliotransmitter

  13. 不同剂量香烟烟雾提取物对阴茎海绵体NOS活性和Cx43蛋白的影响%Cigarette smoke extract reduces NOS activity and CX43 expression in the corporal cavernosum

    Institute of Scientific and Technical Information of China (English)

    刘小兵; 吴天鹏; 詹运运; 王朝阳

    2011-01-01

    目的:探讨不同剂量的香烟烟雾提取物对雄性大鼠勃起功能的影响,进一步研究吸烟导致ED的发病机制.方法:75只健康雄性SD大鼠(8周龄),随机分为5组(15只/组).A组为对照组;B组为皮下注射二甲基亚砜(DMSO)组;C组为皮下注射香烟烟雾提取物(CSE)低剂量组;D组为皮下注射CSE中剂量组;E组为皮下注射CSE高剂量组.应用SD雄性大鼠建立皮下注射CSE模型60 d后,经皮下注射阿朴吗啡(APO)后观察大鼠阴茎勃起情况,处死大鼠取阴茎海绵体组织.一部分标本采用激光共聚焦扫描显微镜方法检测缝隙连接蛋白43(Cx43)的表达;另一部分标本采用比色法测定大鼠阴茎海绵体组织中NOS活性.结果:不同剂量CSE组阴茎勃起次数、阴茎海绵体组织NOS活性和海绵体平滑肌中Cx43表达与DMSO组和对照组比较均明显减少(P0.05).结论:CSE使阴茎海绵体组织NOS活性明显降低、海绵体平滑肌中Cx43蛋白表达明显减少并且严重影响阴茎勃起功能,并且CSE剂量越大,其影响越明显.提示,Cx43蛋白表达下调、NOS活性降低可能是CSE导致ED的发病机制之一.%Objective: To study the effects of different doses of cigarette smoke extract (CSE) on the erectile function of male rats and the mechanism of smoking-induced erectile dysfunction (ED).Methods: A total of 75 healthy male SD rats were randomly divided into Groups A (control), B ( dimethyl sulphoxide [DMSO]), C ( low-dose CSE), D ( medium-dose CSE) and E ( high-dose CSE).CSE models were established in male SD rats by hypodermic injection, and 60 days later observed for penile erection following subcutaneous injection of apomorphine.Then the rats were killed and the penile cavernous body obtained for the examination of NOS activity by chromatometry and the determination of Cx43 expression by laser scanning confocal fluorescence microscopy (LCSM).Results: Compared with the control and DMSO groups, penile erection frequency, NOS

  14. Gap junctions in cells of the immune system: structure, regulation and possible functional roles

    Directory of Open Access Journals (Sweden)

    J.C. Sáez

    2000-04-01

    Full Text Available Gap junction channels are sites of cytoplasmic communication between contacting cells. In vertebrates, they consist of protein subunits denoted connexins (Cxs which are encoded by a gene family. According to their Cx composition, gap junction channels show different gating and permeability properties that define which ions and small molecules permeate them. Differences in Cx primary sequences suggest that channels composed of different Cxs are regulated differentially by intracellular pathways under specific physiological conditions. Functional roles of gap junction channels could be defined by the relative importance of permeant substances, resulting in coordination of electrical and/or metabolic cellular responses. Cells of the native and specific immune systems establish transient homo- and heterocellular contacts at various steps of the immune response. Morphological and functional studies reported during the last three decades have revealed that many intercellular contacts between cells in the immune response present gap junctions or "gap junction-like" structures. Partial characterization of the molecular composition of some of these plasma membrane structures and regulatory mechanisms that control them have been published recently. Studies designed to elucidate their physiological roles suggest that they might permit coordination of cellular events which favor the effective and timely response of the immune system.

  15. Effects of neuroinflammation on glia-glia gap junctional intercellular communication: a perspective.

    Science.gov (United States)

    Kielian, Tammy; Esen, Nilufer

    2004-01-01

    Gap junctions serve as intercellular conduits that allow for the direct transfer of small molecular weight molecules (up to 1 kDa) including ions involved in cellular excitability, metabolic precursors, and second messengers. The observation of extensive intercellular coupling and large numbers of gap junctions in the central nervous system (CNS) suggests a syncytium-like organization of glial compartments. Inflammation is a hallmark of various CNS diseases such as bacterial and viral infections, multiple sclerosis, Alzheimer's disease, and cerebral ischemia. A general consequence of brain inflammation is reactive gliosis typified by astrocyte hypertrophy and proliferation of astrocytes and microglia. Changes in gap junction intercellular communication as reflected by alterations in dye coupling and connexin expression have been associated with numerous CNS inflammatory diseases, which may have dramatic implications on the survival of neuronal and glial populations in the context of neuroinflammation. A review of the effects of inflammatory products on glia-glia gap junctional communication and glial glutamate release is presented. In addition, the hypothesis of a "syncytial switch" based upon differential regulation of gap junction expression in astrocytes and microglia during normal CNS homeostasis and neuroinflammation is proposed.

  16. Gap state charge induced spin-dependent negative differential resistance in tunnel junctions

    Science.gov (United States)

    Jiang, Jun; Zhang, X.-G.; Han, X. F.

    2016-04-01

    We propose and demonstrate through first-principles calculation a new spin-dependent negative differential resistance (NDR) mechanism in magnetic tunnel junctions (MTJ) with cubic cation disordered crystals (CCDC) AlO x or Mg1-x Al x O as barrier materials. The CCDC is a class of insulators whose band gap can be changed by cation doping. The gap becomes arched in an ultrathin layer due to the space charge formed from metal-induced gap states. With an appropriate combination of an arched gap and a bias voltage, NDR can be produced in either spin channel. This mechanism is applicable to 2D and 3D ultrathin junctions with a sufficiently small band gap that forms a large space charge. It provides a new way of controlling the spin-dependent transport in spintronic devices by an electric field. A generalized Simmons formula for tunneling current through junction with an arched gap is derived to show the general conditions under which ultrathin junctions may exhibit NDR.

  17. Blockade of Gap Junction Hemichannel Suppresses Disease Progression in Mouse Models of Amyotrophic Lateral Sclerosis and Alzheimer's Disease: e21108

    National Research Council Canada - National Science Library

    Hideyuki Takeuchi; Hiroyuki Mizoguchi; Yukiko Doi; Shijie Jin; Mariko Noda; Jianfeng Liang; Hua Li; Yan Zhou; Rarami Mori; Satoko Yasuoka; Endong Li; Bijay Parajuli; Jun Kawanokuchi; Yoshifumi Sonobe; Jun Sato; Koji Yamanaka; Gen Sobue; Tetsuya Mizuno; Akio Suzumura

    2011-01-01

    ... neurotoxic/neuroprotective roles of microglia and hardly affected disease progression. We previously demonstrated that activated microglia release a large amount of glutamate specifically through gap junction hemichannel...

  18. Spatially resolved gap closing in single Josephson junctions constructed on Bi2Te3 surface

    Science.gov (United States)

    Pang, Yuan; Wang, Junhua; Lyu, Zhaozheng; Yang, Guang; Fan, Jie; Liu, Guangtong; Ji, Zhongqing; Jing, Xiunian; Yang, Changli; Lu, Li

    2016-11-01

    Full gap closing is a prerequisite for hosting Majorana zero modes in Josephson junctions on the surface of topological insulators. Previously, we have observed direct experimental evidence of gap closing in Josephson junctions constructed on Bi2Te3 surface. In this paper we report further investigations on the position dependence of gap closing as a function of magnetic flux in single Josephson junctions constructed on Bi2Te3 surface. Project supported by the National Basic Research Program of China (Grant Nos. 2009CB929101 and 2011CB921702), the National Natural Science Foundation of China (Grant Nos. 91221203, 11174340, 11174357, 91421303, and 11527806), and the Strategic Priority Research Program B of the Chinese Academy of Sciences (Grant No. XDB07010100).

  19. Gap junctions enhancer combined with Vaughan Williams class III antiarrhythmic drugs, a promising antiarrhythmic method?

    Science.gov (United States)

    Li, Lian-dong; Zhang, Cun-tai; Ruan, Lei; Ni, Ming-ke; Quan, Xiao-qing

    2011-01-01

    Arrhythmias is one of the leading causes of death in the world. Current antiarrhythmic drugs are limited by unsatisfactory efficacy and adverse effects such as proarrhythmias. Reentry mechanism plays an important role in persistence of arrhythmias. Reentry can only continue when reentry path-length is longer than cardiac wavelength which is equal to the product of conduction velocity (CV) and effective refractory period (ERP). Gap junctions uncoupling is associated with proarrhythmic CV slowing and transmural dispersion of repolarization (TDR) increasing in many cardiac diseases. Vaughan Williams class III antiarrhythmic drugs prolong ERP with an augmented TDR which is the main mechanism of the proarrhythmic effects. Gap junctions enhancer can augment CV and diminish TDR. As a result, gap junctions enhancer combined with class III drugs may be a promising antiarrhythmic method.

  20. Regulation of gap junction channels by infectious agents and inflammation in the CNS

    Directory of Open Access Journals (Sweden)

    Paul eCastellano

    2014-05-01

    Full Text Available Gap junctions are conglomerates of intercellular channels that connect the cytoplasm of two or more cells, and facilitate the transfer of second messengers, small peptides and RNA resulting in metabolic and electrical coordination. In general, loss of gap junctional communication (GJC has been associated with cellular damage and inflammation resulting in compromise of physiological functions. Recently, it has become evident that gap junction channels also play a critical role in the pathogenesis of infectious diseases and associated inflammation. Several pathogens use the transfer of intracellular signals through GJ channels to spread infection and toxic signals that amplify inflammation to neighboring cells. Thus, identification of the mechanisms by which several infectious agents alter GJC could result in new potential therapeutic approaches to reduce inflammation and their pathogenesis.

  1. Astrocytic gap junctional networks suppress cellular damage in an in vitro model of ischemia

    Energy Technology Data Exchange (ETDEWEB)

    Shinotsuka, Takanori; Yasui, Masato; Nuriya, Mutsuo, E-mail: mnuriya@z2.keio.jp

    2014-02-07

    Highlights: • Astrocytes exhibit characteristic changes in [Ca{sup 2+}]{sub i} under OGD. • Astrocytic [Ca{sup 2+}]{sub i} increase is synchronized with a neuronal anoxic depolarization. • Gap junctional couplings protect neurons as well as astrocytes during OGD. - Abstract: Astrocytes play pivotal roles in both the physiology and the pathophysiology of the brain. They communicate with each other via extracellular messengers as well as through gap junctions, which may exacerbate or protect against pathological processes in the brain. However, their roles during the acute phase of ischemia and the underlying cellular mechanisms remain largely unknown. To address this issue, we imaged changes in the intracellular calcium concentration ([Ca{sup 2+}]{sub i}) in astrocytes in mouse cortical slices under oxygen/glucose deprivation (OGD) condition using two-photon microscopy. Under OGD, astrocytes showed [Ca{sup 2+}]{sub i} oscillations followed by larger and sustained [Ca{sup 2+}]{sub i} increases. While the pharmacological blockades of astrocytic receptors for glutamate and ATP had no effect, the inhibitions of gap junctional intercellular coupling between astrocytes significantly advanced the onset of the sustained [Ca{sup 2+}]{sub i} increase after OGD exposure. Interestingly, the simultaneous recording of the neuronal membrane potential revealed that the onset of the sustained [Ca{sup 2+}]{sub i} increase in astrocytes was synchronized with the appearance of neuronal anoxic depolarization. Furthermore, the blockade of gap junctional coupling resulted in a concurrent faster appearance of neuronal depolarizations, which remain synchronized with the sustained [Ca{sup 2+}]{sub i} increase in astrocytes. These results indicate that astrocytes delay the appearance of the pathological responses of astrocytes and neurons through their gap junction-mediated intercellular network under OGD. Thus, astrocytic gap junctional networks provide protection against tissue damage

  2. Gap junctional communication between the satellite cells of rat dorsal root ganglia.

    Science.gov (United States)

    Sakuma, E; Wang, H J; Asai, Y; Tamaki, D; Amano, K; Mabuchi, Y; Herbert, D C; Soji, T

    2001-06-01

    Many studies have described the ultrastructure of the dorsal root ganglia in various embryonic and adult animals, but in spite of the efforts of many investigators the functional role of the satellite cells in this tissue is not clearly understood. In this study, we discuss the function of this cell type based on the concept of cell-to-cell interaction through gap junctions. Five male 60 day-old Wistar strain rats were used. All animals were anesthetized with pentobarbital and perfused with glutaraldehyde fixative, then the dorsal root ganglia in levels L4, L5 and L6 were taken from each rat. After postosmication, the specimens were prepared for observation by transmission electron microscopy. All nerve cells were completely surrounded by satellite cell cytoplasmic expansions. The boundaries between adjacent nerve cells and satellite cells were complicated due to the presence of perikaryal projections of nerve cells. Gap junctions which showed the typical trilamellar structure of plasma membranes were found mainly between satellite cell processes belonging to the same nerve cell. On the other hand, some gap junctions were found between the satellite cell projections belonging to different nerve cells. The size of the gap junctions ranged from 300 to 400 nm. No gap junctions were associated with the plasma membrane of any nerve cell. In conclusion, only satellite cells can share free transcellular exchange of cytoplasmic molecules such as ions, amino acids, sugars and several second messengers including cAMP and inositol 1,4,5-triphosphate by way of gap junctions in dorsal root ganglia.

  3. Connexin 43 ubiquitination determines the fate of gap junctions: restrict to survive.

    Science.gov (United States)

    Ribeiro-Rodrigues, Teresa M; Catarino, Steve; Pinho, Maria J; Pereira, Paulo; Girao, Henrique

    2015-06-01

    Connexins (Cxs) are transmembrane proteins that form channels which allow direct intercellular communication (IC) between neighbouring cells via gap junctions. Mechanisms that modulate the amount of channels at the plasma membrane have emerged as important regulators of IC and their de-regulation has been associated with various diseases. Although Cx-mediated IC can be modulated by different mechanisms, ubiquitination has been described as one of the major post-translational modifications involved in Cx regulation and consequently IC. In this review, we focus on the role of ubiquitin and its effect on gap junction intercellular communication.

  4. Anti-arrhythmic effect of U50,488H is mediated by preserving Cx43 protein via intracellular calcium regulation%U50,488H抗心肌缺血性心律失常与调节钙-Cx43通路有关

    Institute of Scientific and Technical Information of China (English)

    林家骥; 周贺; 裴建明

    2012-01-01

    AIM:To test the hypothesis that antiarrhythmic properties of U50,488H may be mediated by preserving Cx43 protein via regulating intracellular calcium.METHODS:Using an in vivo arrhythmia study,arrhythmia was induced by temporary occlusion of the left anterior descending (LAD) coronary artery with a silk suture for 30 min in Sprague Dawley rats weighing 250 to 350 g.U50,488H,a κ-opioid receptor (κ-OR) agonist,Nor-BNI,a κ-opioid receptor antagonist,Bay K8644,a calcium channel agonist,nifedipine,a calcium channel inhibitor and heptanol,a Cx43 inhibitor were chosen to be intravenously ( i.v.) injected into a femoral vein prior to ischemia.A sham group underwent the same surgical procedures except that the suture underneath the LAD was left untied.In an in vitro arrhythmia study,each heart was quickly removed and underwent an initial 10 min of normal baseline perfusion and was subjected to perfusion at 37℃ for 30 min (for analysis of arrhythmia) or 90 min (for Western blotting).The hearts were then randomly divided into three groups:Con (normal calcium perfusion,1.5 mmol/L),high calcium (high calcium perfusion,3.3 mmol/L) and low calcium ( low calcium perfusion,0.5 mmol/L).Before and during the ischemia period,electrocardiogram (ECG) was used to measure the incidence of arrhythmias.RESULTS:By performing ECG monitoring and immunoblotting in isolated Langendorff-perfused rat hearts,high concentrations of calcium-perfused rat hearts exhibited increased cardiac arrhythmias.Diminished expression of Cx43 protein was observed.U50,488H dose-dependently inhibited L-type calcium current in single ventricular myocytes of rats using whole-cell patch clamp techniques.These effects were blocked by nor-BNI,a selective κ-OR antagonist.Administration of U50,488H before myocardial ischemia attenuated total arrhythmia scores.This effect was blocked by nor-BNI,antagonized by Bay K8644,an L-type calcium channel agonist,and by the Cx43 uncoupler heptanol.Finally,immunoblotting data

  5. Connexin26 regulates assembly and maintenance of cochlear gap junction macromolecular complex for normal hearing

    Science.gov (United States)

    Kamiya, Kazusaku; Fukunaga, Ichiro; Hatakeyama, Kaori; Ikeda, Katsuhisa

    2015-12-01

    Hereditary deafness affects about 1 in 2000 children and GJB2 gene mutation is most frequent cause for this disease in the world. GJB2 encodes connexin26 (Cx26), a component in cochlear gap junction. Recently, we found macromolecular change of gap junction plaques with two different types of Cx26 mutation as major classification of clinical case, one is a model of dominant negative type, Cx26R75W+ and the other is conditional gene deficient mouse, Cx26f/fP0Cre as a model for insufficiency of gap junction protein [6]. Gap junction composed mainly of Cx26 and Cx30 in wild type mice formed large planar gap junction plaques (GJP). In contrast, Cx26R75W+ and Cx26f/fP0Cre showed fragmented small round GJPs around the cell border. In Cx26f/fP0Cre, some of the cells with Cx26 expression due to their cellular mosaicism showed normal large GJP with Cx26 and Cx30 only at the cell junction site between two Cx26 positive cells. These indicate that bilateral Cx26 expressions from both adjacent cells are essential for the formation of the cochlear linear GJP, and it is not compensated by other cochlear Connexins such as Connexin30. In the present study, we demonstrated a new molecular pathology in most common hereditary deafness with different types of Connexin26 mutations, and this machinery can be a new target for drag design of hereditary deafness.

  6. Variable conformation of GAP junctions linking bone cells: a transmission electron microscopic study of linear, stacked linear, curvilinear, oval, and annular junctions.

    Science.gov (United States)

    Shapiro, F

    1997-10-01

    There is a marked variability in the conformation of bone cell gap junctions in newborn murine cortical bone as defined by transmission electron microscopy (TEM). Studies were done in newborn BALB/c mouse and Sprague-Dawley rat femurs and tibias. Femoral and tibial cortices were dissected into 1 mm3 fragments and prepared in standardized fashion using modified Karnovsky fixation, 7.5% EDTA decalcification, 1% osmium tetroxide-sym collidine buffer with 1% lanthanum nitrate postfixation, Epon resin, 60 nm sections, lead citrate/uranyl acetate staining, and examination at 60 kV. Previous TEM descriptions of bone junctions have, with rare exceptions, noted only isolated linear or mildly curvilinear structures. In this study we noted gap junctional shapes on thin-section TEM preparations of osteoblasts and osteocytes to be extremely variable and complex encompassing linear, curvilinear, stacked linear, oval, and annular conformations. Multiple observations revealed linear gap junctions linking surface osteoblast cell bodies; linear, curvilinear, stacked linear, and oval junctions linking osteoblast processes in osteoid; linear and curvilinear junctions where cell processes joined with osteocyte cell bodies and each of the five conformations linking osteocyte processes within canaliculi. The annular junctions were found within osteoblast and osteocyte cytoplasm and in osteocyte cell processes within canaliculi. The annular junctions are intracellular, degenerating structures which appear as ultrastructural markers of gap junction involution. The more complex shapes reported here must be considered in (1) interpreting quantitative studies using freeze-fracture replicas, thin sections, and confocal microscopy immunolabeled junction connexin-43 components and (2) assessing gap junction biogenesis and turnover. 3-D reconstruction of bone junctions will enhance our understanding of these complex conformations.

  7. Connexins: a myriad of functions extending beyond assembly of gap junction channels

    Directory of Open Access Journals (Sweden)

    Mroue Rana M

    2009-03-01

    Full Text Available Abstract Connexins constitute a large family of trans-membrane proteins that allow intercellular communication and the transfer of ions and small signaling molecules between cells. Recent studies have revealed complex translational and post-translational mechanisms that regulate connexin synthesis, maturation, membrane transport and degradation that in turn modulate gap junction intercellular communication. With the growing myriad of connexin interacting proteins, including cytoskeletal elements, junctional proteins, and enzymes, gap junctions are now perceived, not only as channels between neighboring cells, but as signaling complexes that regulate cell function and transformation. Connexins have also been shown to form functional hemichannels and have roles altogether independent of channel functions, where they exert their effects on proliferation and other aspects of life and death of the cell through mostly-undefined mechanisms. This review provides an updated overview of current knowledge of connexins and their interacting proteins, and it describes connexin modulation in disease and tumorigenesis.

  8. Connexins: a myriad of functions extending beyond assembly of gap junction channels.

    Science.gov (United States)

    Dbouk, Hashem A; Mroue, Rana M; El-Sabban, Marwan E; Talhouk, Rabih S

    2009-03-12

    Connexins constitute a large family of trans-membrane proteins that allow intercellular communication and the transfer of ions and small signaling molecules between cells. Recent studies have revealed complex translational and post-translational mechanisms that regulate connexin synthesis, maturation, membrane transport and degradation that in turn modulate gap junction intercellular communication. With the growing myriad of connexin interacting proteins, including cytoskeletal elements, junctional proteins, and enzymes, gap junctions are now perceived, not only as channels between neighboring cells, but as signaling complexes that regulate cell function and transformation. Connexins have also been shown to form functional hemichannels and have roles altogether independent of channel functions, where they exert their effects on proliferation and other aspects of life and death of the cell through mostly-undefined mechanisms. This review provides an updated overview of current knowledge of connexins and their interacting proteins, and it describes connexin modulation in disease and tumorigenesis.

  9. Gap junctions and hemichannels composed of connexins: potential therapeutic targets for neurodegenerative diseases

    Directory of Open Access Journals (Sweden)

    Hideyuki eTakeuchi

    2014-09-01

    Full Text Available Microglia are macrophage-like resident immune cells that contribute to the maintenance of homeostasis in the central nervous system (CNS. Abnormal activation of microglia can cause damage in the CNS, and accumulation of activated microglia is a characteristic pathological observation in neurologic conditions such as trauma, stroke, inflammation, epilepsy, and neurodegenerative diseases. Activated microglia secrete high levels of glutamate, which damages CNS cells and has been implicated as a major cause of neurodegeneration in these conditions. Glutamate-receptor blockers and microglia inhibitors (e.g. minocycline have been examined as therapeutic candidates for several neurodegenerative diseases; however, these compounds exerted little therapeutic benefit because they either perturbed physiological glutamate signals or suppressed the actions of protective microglia. The ideal therapeutic approach would hamper the deleterious roles of activated microglia without diminishing their protective effects. We recently found that abnormally activated microglia secrete glutamate via gap-junction hemichannels on the cell surface. Moreover, administration of gap-junction inhibitors significantly suppressed excessive microglial glutamate release and improved disease symptoms in animal models of neurologic conditions such as stroke, multiple sclerosis, amyotrophic lateral sclerosis, and Alzheimer’s disease. Recent evidence also suggests that neuronal and glial communication via gap junctions amplifies neuroinflammation and neurodegeneration. Elucidation of the precise pathologic roles of gap junctions and hemichannels may lead to a novel therapeutic strategies that can slow and halt the progression of neurodegenerative diseases.

  10. Electrical signal transmission in a bone cell network: the influence of a discrete gap junction

    Science.gov (United States)

    Zhang, D.; Weinbaum, S.; Cowin, S. C.

    1998-01-01

    A refined electrical cable model is formulated to investigate the role of a discrete gap junction in the intracellular transmission of electrical signals in an electrically coupled system of osteocytes and osteoblasts in an osteon. The model also examines the influence of the ratio q between the membrane's electrical time constant and the characteristic time of pore fluid pressure, the circular, cylindrical geometry of the osteon, and key simplifying assumptions in our earlier continuous cable model (see Zhang, D., S. C. Cowin, and S. Weinbaum. Electrical signal transmission and gap junction regulation in a bone cell network: A cable model for an osteon. Ann. Biomed. Eng. 25:379-396, 1997). Using this refined model, it is shown that (1) the intracellular potential amplitude at the osteoblastic end of the osteonal cable retains the character of a combination of a low-pass and a high-pass filter as the corner frequency varies in the physiological range; (2) the presence of a discrete gap junction near a resting osteoblast can lead to significant modulation of the intracellular potential and current in the osteoblast for measured values of the gap junction coupling strength; and (3) the circular, cylindrical geometry of the osteon is well simulated by the beam analogy used in Zhang et al.

  11. Gap junctions and hemichannels composed of connexins: potential therapeutic targets for neurodegenerative diseases

    Science.gov (United States)

    Takeuchi, Hideyuki; Suzumura, Akio

    2014-01-01

    Microglia are macrophage-like resident immune cells that contribute to the maintenance of homeostasis in the central nervous system (CNS). Abnormal activation of microglia can cause damage in the CNS, and accumulation of activated microglia is a characteristic pathological observation in neurologic conditions such as trauma, stroke, inflammation, epilepsy, and neurodegenerative diseases. Activated microglia secrete high levels of glutamate, which damages CNS cells and has been implicated as a major cause of neurodegeneration in these conditions. Glutamate-receptor blockers and microglia inhibitors (e.g., minocycline) have been examined as therapeutic candidates for several neurodegenerative diseases; however, these compounds exerted little therapeutic benefit because they either perturbed physiological glutamate signals or suppressed the actions of protective microglia. The ideal therapeutic approach would hamper the deleterious roles of activated microglia without diminishing their protective effects. We recently found that abnormally activated microglia secrete glutamate via gap-junction hemichannels on the cell surface. Moreover, administration of gap-junction inhibitors significantly suppressed excessive microglial glutamate release and improved disease symptoms in animal models of neurologic conditions such as stroke, multiple sclerosis, amyotrophic lateral sclerosis, and Alzheimer's disease. Recent evidence also suggests that neuronal and glial communication via gap junctions amplifies neuroinflammation and neurodegeneration. Elucidation of the precise pathologic roles of gap junctions and hemichannels may lead to a novel therapeutic strategies that can slow and halt the progression of neurodegenerative diseases. PMID:25228858

  12. Increased phosphorylation of Cx36 gap junctions in the AII amacrine cells of RD retina

    Directory of Open Access Journals (Sweden)

    Elena eIvanova

    2015-10-01

    Full Text Available Retinal degeneration (RD encompasses a family of diseases that lead to photoreceptor death and visual impairment. Visual decline due to photoreceptor cell loss is further compromised by emerging spontaneous hyperactivity in inner retinal cells. This aberrant activity acts as a barrier to signals from the remaining photoreceptors, hindering therapeutic strategies to restore light sensitivity in RD. Gap junctions, particularly those expressed in AII amacrine cells, have been shown to be integral to the generation of aberrant activity. It is unclear whether gap junction expression and coupling are altered in RD. To test this, we evaluated the expression and phosphorylation state of connexin36, the gap junction subunit predominantly expressed in AII amacrine cells, in two mouse models of RD, rd10 (slow degeneration and rd1 (fast degeneration. Using Ser293-P antibody, which recognizes a phosphorylated form of connexin36, we found that phosphorylation of connexin36 in both slow and fast RD models was significantly greater than in wildtype controls. This elevated phosphorylation may underlie the increased gap junction coupling of AII amacrine cells exhibited by RD retina.

  13. Gap junctions and hemichannels composed of connexins: potential therapeutic targets for neurodegenerative diseases.

    Science.gov (United States)

    Takeuchi, Hideyuki; Suzumura, Akio

    2014-01-01

    Microglia are macrophage-like resident immune cells that contribute to the maintenance of homeostasis in the central nervous system (CNS). Abnormal activation of microglia can cause damage in the CNS, and accumulation of activated microglia is a characteristic pathological observation in neurologic conditions such as trauma, stroke, inflammation, epilepsy, and neurodegenerative diseases. Activated microglia secrete high levels of glutamate, which damages CNS cells and has been implicated as a major cause of neurodegeneration in these conditions. Glutamate-receptor blockers and microglia inhibitors (e.g., minocycline) have been examined as therapeutic candidates for several neurodegenerative diseases; however, these compounds exerted little therapeutic benefit because they either perturbed physiological glutamate signals or suppressed the actions of protective microglia. The ideal therapeutic approach would hamper the deleterious roles of activated microglia without diminishing their protective effects. We recently found that abnormally activated microglia secrete glutamate via gap-junction hemichannels on the cell surface. Moreover, administration of gap-junction inhibitors significantly suppressed excessive microglial glutamate release and improved disease symptoms in animal models of neurologic conditions such as stroke, multiple sclerosis, amyotrophic lateral sclerosis, and Alzheimer's disease. Recent evidence also suggests that neuronal and glial communication via gap junctions amplifies neuroinflammation and neurodegeneration. Elucidation of the precise pathologic roles of gap junctions and hemichannels may lead to a novel therapeutic strategies that can slow and halt the progression of neurodegenerative diseases.

  14. Remodeling of gap junctions in ischemic and nonischemic forms of heart disease.

    Science.gov (United States)

    Saffitz, Jeffrey E; Hames, Kiyomi Yamada; Kanno, Shigeto

    2007-08-01

    Electrical activation of the myocardium to produce effective pumping of blood depends on the orderly coordinated spatial and temporal transfer of current from one cell to another via gap junctions. Normal ventricular myocytes are extensively coupled by gap junctions and have the capacity to rapidly increase the amount of connexin within gap junction plaques to meet physiological demands for enhanced cell-cell communication. However, myocytes can also rapidly uncouple in response to injury or disease. In general, both acute and chronic forms of heart disease caused by diverse etiologies are associated with changes in the expression of connexins and remodeling of gap junctions. Such remodeling may have both adaptive and maladaptive consequences and contribute to major clinical processes such as heart failure and sudden cardiac death. Our laboratory has investigated mechanisms regulating cell-cell electrical coupling in the heart under physiological and pathophysiological conditions. This review is focused on selected aspects of this work pertaining to changes in coupling in response to acute and chronic ischemic heart disease and in familial cardiomyopathies caused by mutations in genes encoding desmosomal proteins.

  15. Estimation of the effective intercellular diffusion coefficient in cell monolayers coupled by gap junctions

    DEFF Research Database (Denmark)

    Olesen, Niels Erik; Hofgaard, Johannes P; von Holstein-Rathlou, Niels-Henrik;

    2012-01-01

    A recently developed dye-based assay to study gap junction permeability is analysed. The assay is based on electroporation of dye into a large number of connexin 43 expressing cells, grown to confluency on electrically conductive slides. The subsequent intercellular spread of dye to non-electroporated...

  16. Doping GaP core-shell nanowire pn-junctions

    DEFF Research Database (Denmark)

    Yazdi, Sadegh; Berg, Alexander; Borgström, Magnus T.

    2015-01-01

    The doping process in GaP core-shell nanowire pn-junctions using different precursors is evaluated by mapping the nanowires' electrostatic potential distribution by means of off-axis electron holography. Three precursors, triethyltin (TESn), ditertiarybutylselenide, and silane are investigated fo...

  17. Effect of thioridazine on gap junction intercellular communication in connexin 43-expressing cells.

    Science.gov (United States)

    Matesic, D F; Abifadel, D N; Garcia, E L; Jann, M W

    2006-07-01

    Propagation of electrical activity between myocytes in the heart requires gap junction channels, which contribute to coordinated conduction of the heartbeat. Some antipsychotic drugs, such as thioridazine and its active metabolite, mesoridazine, have known cardiac conduction side-effects, which have resulted in fatal or nearly fatal clinical consequences in patients. The physiological mechanisms responsible for these cardiac side-effects are unknown. We tested the effect of thioridazine and mesoridazine on gap junction-mediated intercellular communication between cells that express the major cardiac gap junction subtype connexin 43. Micromolar concentrations of thioridazine and mesoridazine inhibited gap junction-mediated intercellular communication between WB-F344 epithelial cells in a dose-dependent manner, as measured by fluorescent dye transfer. Kinetic analyses demonstrated that inhibition by 10 micromol/L thioridazine occurred within 5 min, achieved its maximal effect within 1 h, and was maintained for at least 24 h. Inhibition was reversible within 1 h upon removal of the drug. Western blot analysis of connexin 43 in a membrane-enriched fraction of WB-F344 cells treated with thioridazine revealed decreased amounts of unphosphorylated connexin 43, and appearance of a phosphorylated connexin 43 band that co-migrated with a "hyperphosphorylated" connexin 43 band present in TPA-inhibited cells. When tested for its effects on cardiomyocytes isolated from neonatal rats, thioridazine decreased fluorescent dye transfer between colonies of beating myocytes. Microinjection of individual cells with fluorescent dye also showed inhibition of dye transfer in thioridazine-treated cells compared to vehicle-treated cells. In addition, thioridazine, like TPA, inhibited rhythmic beating of myocytes within 15 min of application. In light of the fact that the thioridazine and mesoridazine concentrations used in these experiments are in the range of those used clinically in

  18. Sustained inhibition of rat myometrial gap junctions and contractions by lindane

    Directory of Open Access Journals (Sweden)

    Grindatti Carmen M

    2003-10-01

    Full Text Available Abstract Background Gap junctions increase in size and abundance coincident with parturition, forming an intercellular communication network that permits the uterus to develop the forceful, coordinated contractions necessary for delivery of the fetus. Lindane, a pesticide used in the human and veterinary treatment of scabies and lice as well as in agricultural applications, inhibits uterine contractions in vitro, inhibits myometrial gap junctions, and has been associated with prolonged gestation length in rats. The aim of the present study was to investigate whether brief exposures to lindane would elicit sustained inhibition of rat uterine contractile activity and myometrial gap junction intercellular communication. Methods To examine effects on uterine contraction, longitudinal uterine strips isolated from late gestation (day 20 rats were exposed to lindane in muscle baths and monitored for changes in spontaneous phasic contractions during and after exposure to lindane. Lucifer yellow dye transfer between myometrial cells in culture was used to monitor gap junction intercellular communication. Results During a 1-h exposure, 10 micro M and 100 micro M lindane decreased peak force and frequency of uterine contraction but 1 micro M lindane did not. After removal of the exposure buffer, contraction force remained significantly depressed in uterine strips exposed to 100 micro M lindane, returning to less than 50% basal levels 5 h after cessation of lindane exposure. In cultured myometrial myocytes, significant sustained inhibition of Lucifer yellow dye transfer was observed 24 h after lindane exposures as brief as 10 min and as low as 0.1 micro M lindane. Conclusion Brief in vitro exposures to lindane have long-term effects on myometrial functions that are necessary for parturition, inhibiting spontaneous phasic contractions in late gestation rat uterus and gap junction intercellular communication in myometrial cell cultures.

  19. General anesthetics have differential inhibitory effects on gap junction channels and hemichannels in astrocytes and neurons.

    Science.gov (United States)

    Liu, Xinhe; Gangoso, Ester; Yi, Chenju; Jeanson, Tiffany; Kandelman, Stanislas; Mantz, Jean; Giaume, Christian

    2016-04-01

    Astrocytes represent a major non-neuronal cell population actively involved in brain functions and pathologies. They express a large amount of gap junction proteins that allow communication between adjacent glial cells and the formation of glial networks. In addition, these membrane proteins can also operate as hemichannels, through which "gliotransmitters" are released, and thus contribute to neuroglial interaction. There are now reports demonstrating that alterations of astroglial gap junction communication and/or hemichannel activity impact neuronal and synaptic activity. Two decades ago we reported that several general anesthetics inhibited gap junctions in primary cultures of astrocytes (Mantz et al., (1993) Anesthesiology 78(5):892-901). As there are increasing studies investigating neuroglial interactions in anesthetized mice, we here updated this previous study by employing acute cortical slices and by characterizing the effects of general anesthetics on both astroglial gap junctions and hemichannels. As hemichannel activity is not detected in cortical astrocytes under basal conditions, we treated acute slices with the endotoxin LPS or proinflammatory cytokines to induce hemichannel activity in astrocytes, which in turn activated neuronal hemichannels. We studied two extensively used anesthetics, propofol and ketamine, and the more recently developed dexmedetomidine. We report that these drugs have differential inhibitory effects on gap junctional communication and hemichannel activity in astrocytes when used in their respective, clinically relevant concentrations, and that dexmedetomidine appears to be the least effective on both channel functions. In addition, the three anesthetics have similar effects on neuronal hemichannels. Altogether, our observations may contribute to optimizing the selection of anesthetics for in vivo animal studies.

  20. Connexin43 mutation causes heterogeneous gap junction loss and sudden infant death.

    Science.gov (United States)

    Van Norstrand, David W; Asimaki, Angeliki; Rubinos, Clio; Dolmatova, Elena; Srinivas, Miduturu; Tester, David J; Saffitz, Jeffrey E; Duffy, Heather S; Ackerman, Michael J

    2012-01-24

    An estimated 10% to 15% of sudden infant death syndrome (SIDS) cases may stem from channelopathy-mediated lethal arrhythmias. Loss of the GJA1-encoded gap junction channel protein connexin43 is known to underlie formation of lethal arrhythmias. GJA1 mutations have been associated with cardiac diseases, including atrial fibrillation. Therefore, GJA1 is a plausible candidate gene for premature sudden death. GJA1 open reading frame mutational analysis was performed with polymerase chain reaction, denaturing high-performance liquid chromatography, and direct DNA sequencing on DNA from 292 SIDS cases. Immunofluorescence and dual whole-cell patch-clamp studies were performed to determine the functionality of mutant gap junctions. Immunostaining for gap junction proteins was performed on SIDS-associated paraffin-embedded cardiac tissue. Two rare, novel missense mutations, E42K and S272P, were detected in 2 of 292 SIDS cases, a 2-month-old white boy and a 3-month-old white girl, respectively. Analysis of the E42K victim's parental DNA demonstrated a de novo mutation. Both mutations involved highly conserved residues and were absent in >1000 ethnically matched reference alleles. Immunofluorescence demonstrated no trafficking abnormalities for either mutation, and S272P demonstrated wild-type junctional conductance. However, junctional conductance measurements for the E42K mutation demonstrated a loss of function not rescued by wild type. Moreover, the E42K victim's cardiac tissue demonstrated a mosaic immunostaining pattern for connexin43 protein. This study provides the first molecular and functional evidence implicating a GJA1 mutation as a novel pathogenic substrate for SIDS. E42K-connexin43 demonstrated a trafficking-independent reduction in junctional coupling in vitro and a mosaic pattern of mutational DNA distribution in deceased cardiac tissue, suggesting a novel mechanism of connexin43-associated sudden death.

  1. TLR-Activated Gap Junction Channels Protect Mice against Bacterial Infection through Extracellular UDP Release.

    Science.gov (United States)

    Qin, Juliang; Zhang, Guangxu; Zhang, Xiaoyu; Tan, Binghe; Lv, Zhangsheng; Liu, Mingyao; Ren, Hua; Qian, Min; Du, Bing

    2016-02-15

    Extracellular UDP (eUDP), released as a danger signal by stressed or apoptotic cells, plays an important role in a series of physiological processes. Although the mechanism of eUDP release in apoptotic cells has been well defined, how the eUDP is released in innate immune responses remains unknown. In this study, we demonstrated that UDP was released in both Escherichia coli-infected mice and LPS- or Pam3CSK4-treated macrophages. Also, LPS-induced UDP release could be significantly blocked by selective TLR4 inhibitor Atractylenolide I and selective gap junction inhibitors carbenoxolone and flufenamic acid (FFA), suggesting the key role of TLR signaling and gap junction channels in this process. Meanwhile, eUDP protected mice from peritonitis by reducing invaded bacteria that could be rescued by MRS2578 (selective P2Y6 receptor inhibitor) and FFA. Then, connexin 43, as one of the gap junction proteins, was found to be clearly increased by LPS in a dose- and time-dependent manner. Furthermore, if we blocked LPS-induced ERK signaling by U0126, the expression of connexin 43 and UDP release was also inhibited dramatically. In addition, UDP-induced MCP-1 secretion was significantly reduced by MRS2578, FFA, and P2Y6 mutation. Accordingly, pretreating mice with U0126 and Gap26 increased invaded bacteria and aggravated mice death. Taken together, our study reveals an internal relationship between danger signals and TLR signaling in innate immune responses, which suggests a potential therapeutic significance of gap junction channel-mediated UDP release in infectious diseases. Copyright © 2016 by The American Association of Immunologists, Inc.

  2. Vascular Pericyte Impairment and Connexin43 Gap Junction Deficit Contribute to Vasomotor Decline in Diabetic Retinopathy.

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    Ivanova, Elena; Kovacs-Oller, Tamas; Sagdullaev, Botir T

    2017-08-09

    Adequate blood flow is essential to brain function, and its disruption is an early indicator in diseases, such as stroke and diabetes. However, the mechanisms contributing to this impairment remain unclear. To address this gap, in the diabetic and nondiabetic male mouse retina, we combined an unbiased longitudinal assessment of vasomotor activity along a genetically defined vascular network with pharmacological and immunohistochemical analyses of pericytes, the capillary vasomotor elements. In nondiabetic retina, focal stimulation of a pericyte produced a robust vasomotor response, which propagated along the blood vessel with increasing stimulus. In contrast, the magnitude, dynamic range, a measure of fine vascular diameter control, and propagation of vasomotor response were diminished in diabetic retinas from streptozotocin-treated mice. These functional changes were linked to several mechanisms. We found that density of pericytes and their sensitivity to stimulation were reduced in diabetes. The impaired response propagation from the stimulation site was associated with lower expression of connexin43, a major known gap junction unit in vascular cells. Indeed, selective block of gap junctions significantly reduced propagation but not initiation of vasomotor response in the nondiabetic retina. Our data establish the mechanisms for fine local regulation of capillary diameter by pericytes and a role for gap junctions in vascular network interactions. We show how disruption of this balance contributes to impaired vasomotor control in diabetes.SIGNIFICANCE STATEMENT Identification of mechanisms governing capillary blood flow in the CNS and how they are altered in disease provides novel insight into early states of neurological dysfunction. Here, we present physiological and anatomical evidence that both intact pericyte function as well as gap junction-mediated signaling across the vascular network are essential for proper capillary diameter control and vasomotor

  3. Altered detrusor gap junction communications induce storage symptoms in bladder inflammation: a mouse cyclophosphamide-induced model of cystitis.

    Science.gov (United States)

    Okinami, Takeshi; Imamura, Masaaki; Nishikawa, Nobuyuki; Negoro, Hiromitsu; Sugino, Yoshio; Yoshimura, Koji; Kanematsu, Akihiro; Hashitani, Hikaru; Ogawa, Osamu

    2014-01-01

    Lower urinary tract symptoms (LUTS) include storage, voiding and post-micturition symptoms, featuring many urological diseases. Storage symptoms are the most frequent among these and associated with overactive bladder and non-bacterial bladder inflammation such as interstitial cystitis/bladder pain syndrome (IC/BPS). Gap junction, a key regulator of hyperactive conditions in the bladder, has been reported to be involved in pathological bladder inflammation. Here we report involvement of gap junction in the etiology of storage symptoms in bladder inflammation. In this study, cyclophosphamide-induced cystitis was adapted as a model of bladder inflammation. Cyclophosphamide-treated mice showed typical storage symptoms including increased urinary frequency and reduced bladder capacity, with concurrent up-regulation of connexin 43 (GJA1), one of the major gap junction proteins in the bladder. In isometric tension study, bladder smooth muscle strips taken from the treated mice showed more pronounced spontaneous contraction than controls, which was attenuated by carbenoxolone, a gap junction inhibitor. In voiding behavior studies, the storage symptoms in the treated mice characterized by frequent voiding were alleviated by 18α-glycyrrhetinic acid, another gap junction inhibitor. These results demonstrate that cyclophosphamide-induced mouse model of cystitis shows clinical storage symptoms related with bladder inflammation and that gap junction in the bladder may be a key molecule of these storage symptoms. Therefore, gap junction in the bladder might be an alternative therapeutic target for storage symptoms in bladder inflammation.

  4. Altered detrusor gap junction communications induce storage symptoms in bladder inflammation: a mouse cyclophosphamide-induced model of cystitis.

    Directory of Open Access Journals (Sweden)

    Takeshi Okinami

    Full Text Available Lower urinary tract symptoms (LUTS include storage, voiding and post-micturition symptoms, featuring many urological diseases. Storage symptoms are the most frequent among these and associated with overactive bladder and non-bacterial bladder inflammation such as interstitial cystitis/bladder pain syndrome (IC/BPS. Gap junction, a key regulator of hyperactive conditions in the bladder, has been reported to be involved in pathological bladder inflammation. Here we report involvement of gap junction in the etiology of storage symptoms in bladder inflammation. In this study, cyclophosphamide-induced cystitis was adapted as a model of bladder inflammation. Cyclophosphamide-treated mice showed typical storage symptoms including increased urinary frequency and reduced bladder capacity, with concurrent up-regulation of connexin 43 (GJA1, one of the major gap junction proteins in the bladder. In isometric tension study, bladder smooth muscle strips taken from the treated mice showed more pronounced spontaneous contraction than controls, which was attenuated by carbenoxolone, a gap junction inhibitor. In voiding behavior studies, the storage symptoms in the treated mice characterized by frequent voiding were alleviated by 18α-glycyrrhetinic acid, another gap junction inhibitor. These results demonstrate that cyclophosphamide-induced mouse model of cystitis shows clinical storage symptoms related with bladder inflammation and that gap junction in the bladder may be a key molecule of these storage symptoms. Therefore, gap junction in the bladder might be an alternative therapeutic target for storage symptoms in bladder inflammation.

  5. Role of gap junction channel in the development of beat-to-beat action potential repolarization variability and arrhythmias.

    Science.gov (United States)

    Magyar, Janos; Banyasz, Tamas; Szentandrassy, Norbert; Kistamas, Kornel; Nanasi, Peter P; Satin, Jonathan

    2015-01-01

    The short-term beat-to-beat variability of cardiac action potential duration (SBVR) occurs as a random alteration of the ventricular repolarization duration. SBVR has been suggested to be more predictive of the development of lethal arrhythmias than the action potential prolongation or QT prolongation of ECG alone. The mechanism underlying SBVR is not completely understood but it is known that SBVR depends on stochastic ion channel gating, intracellular calcium handling and intercellular coupling. Coupling of single cardiomyocytes significantly decreases the beat-to-beat changes in action potential duration (APD) due to the electrotonic current flow between neighboring cells. The magnitude of this electrotonic current depends on the intercellular gap junction resistance. Reduced gap junction resistance causes greater electrotonic current flow between cells, and reduces SBVR. Myocardial ischaemia (MI) is known to affect gap junction channel protein expression and function. MI increases gap junction resistance that leads to slow conduction, APD and refractory period dispersion, and an increase in SBVR. Ultimately, development of reentry arrhythmias and fibrillation are associated post-MI. Antiarrhythmic drugs have proarrhythmic side effects requiring alternative approaches. A novel idea is to target gap junction channels. Specifically, the use of gap junction channel enhancers and inhibitors may help to reveal the precise role of gap junctions in the development of arrhythmias. Since cell-to-cell coupling is represented in SBVR, this parameter can be used to monitor the degree of coupling of myocardium.

  6. A rapid and sensitive assay of intercellular coupling by voltage imaging of gap junction networks.

    Science.gov (United States)

    Ceriani, Federico; Mammano, Fabio

    2013-10-21

    A variety of mechanisms that govern connexin channel gating and permeability regulate coupling in gap junction networks. Mutations in connexin genes have been linked to several pathologies, including cardiovascular anomalies, peripheral neuropathy, skin disorders, cataracts and deafness. Gap junction coupling and its patho-physiological alterations are commonly assayed by microinjection experiments with fluorescent tracers, which typically require several minutes to allow dye transfer to a limited number of cells. Comparable or longer time intervals are required by fluorescence recovery after photobleaching experiments. Paired electrophysiological recordings have excellent time resolution but provide extremely limited spatial information regarding network connectivity. Here, we developed a rapid and sensitive method to assay gap junction communication using a combination of single cell electrophysiology, large-scale optical recordings and a digital phase-sensitive detector to extract signals with a known frequency from Vf2.1.Cl, a novel fluorescent sensor of plasma membrane potential. Tests performed in HeLa cell cultures confirmed that suitably encoded Vf2.1.Cl signals remained confined within the network of cells visibly interconnected by fluorescently tagged gap junction channels. We used this method to visualize instantly intercellular connectivity over the whole field of view (hundreds of cells) in cochlear organotypic cultures from postnatal mice. A simple resistive network model reproduced accurately the spatial dependence of the electrical signals throughout the cellular network. Our data suggest that each pair of cochlear non-sensory cells of the lesser epithelial ridge is coupled by ~1500 gap junction channels, on average. Junctional conductance was reduced by 14% in cochlear cultures harboring the T5M mutation of connexin30, which induces a moderate hearing loss in connexin30T5M/T5M knock-in mice, and by 91% in cultures from connexin30-/- mice, which are

  7. Effect of sound on gap-junction-based intercellular signaling: Calcium waves under acoustic irradiation.

    Science.gov (United States)

    Deymier, P A; Swinteck, N; Runge, K; Deymier-Black, A; Hoying, J B

    2015-01-01

    We present a previously unrecognized effect of sound waves on gap-junction-based intercellular signaling such as in biological tissues composed of endothelial cells. We suggest that sound irradiation may, through temporal and spatial modulation of cell-to-cell conductance, create intercellular calcium waves with unidirectional signal propagation associated with nonconventional topologies. Nonreciprocity in calcium wave propagation induced by sound wave irradiation is demonstrated in the case of a linear and a nonlinear reaction-diffusion model. This demonstration should be applicable to other types of gap-junction-based intercellular signals, and it is thought that it should be of help in interpreting a broad range of biological phenomena associated with the beneficial therapeutic effects of sound irradiation and possibly the harmful effects of sound waves on health.

  8. Search for a proximity effect induced gap in gold/high T sub c junctions

    Energy Technology Data Exchange (ETDEWEB)

    Dessau, D.S.; Wells, B.O.; Shen, Z.; Spicer, W.E. (Stanford Electronics Laboratories, Stanford University, Stanford California 94305 (US)); Arko, A.J.; List, R.S. (Los Alamos National Laboratory, Los Alamos, New Mexico 87545 (USA)); Olson, C.G. (Ames Laboratory, Iowa State University, Ames, Iowa 50011 (USA)); Eom, C.B.; Mitzi, D.B.; Kapitulnik, A.; Geballe, T.H. (Department of Applied Physics, Stanford University, Stanford, California 94305 (USA))

    1991-03-25

    We have used high-resolution photoemission spectroscopy to search for a proximity effect induced superconducting gap in gold overlayers on {ital c}-axis single crystals of Bi{sub 2}Sr{sub 2}CaCu{sub 2}O{sub 8} and {ital a}-axis thin films of YBa{sub 2}Cu{sub 3}O{sub 7}. These two junction types give us a representative sampling of very well characterized near-ideal interfaces (gold/{ital c}-axis Bi{sub 2}Sr{sub 2}CaCu{sub 2}O{sub 8}) and junctions in which the geometry more strongly favors the existence of the proximity effect but the interfacial quality may not be as ideal (gold/{ital a}-axis YBa{sub 2}Cu{sub 3}O{sub 7}). In neither of these junction types did we observe any evidence for a proximity effect induced gap, and we place an upper limit of approximately 5 meV on its existence in the junctions that we have studied.

  9. Ultrastructure of junction areas between neurons and astrocytes in rat supraoptic nuclei

    Institute of Scientific and Technical Information of China (English)

    Li Duan; Hua Yuan; Chang-Jun Su; Ying-Ying Liu; Zhi-Ren Rao

    2004-01-01

    AIM: To determine the ultrastructure of junction areas between neurons and astrocytes of supraoptic nuclei in rats orally administered 30 g/L NaCl solution for 5 days.METHODS: The anti-connexin (CX) 43 and anti-CX32 double immunoelectromicroscopic labeled method, and anti-Fos or anti-glial fibrillary acidic protein (GFAP) immunohistochemistry were used to detect changes in the junctional area between neurons and astrocytes in supraoptic nuclei of 5 rats after 30 g/L NaCL solution was given for 5days.RESULTS: A heterotypic connexin32/connexin43 gap junction (HGJ) between neurons and astrocytes (AS) in rat supraoptic nuclei was observed, which was characterized by the thickening and dark staining of cytomembranes with a narrow cleft between them. The number of HGJs and Fos like immunoreactive (-LI) cells was significantly increased following hyperosmotic stimuli, that is, the rats were administered 30 g/L NaCl solution orally or 90 g/L NaCl solution intravenously. HGJs could be blocked with carbenoxolone (CBX), a gap junction blocker, and the number of Fos-LI neurons was significantly decreased compared with that in rats without CBX injection, while Fos-LI ASs were not affected.CONCLUSION: HGJ may be a rapid adaptive signal structure between neurons and ASs in response to stimulation.

  10. Gap junctions are essential for murine primordial follicle assembly immediately before birth.

    Science.gov (United States)

    Teng, Zhen; Wang, Chao; Wang, Yijing; Huang, Kun; Xiang, Xi; Niu, Wanbao; Feng, Lizhao; Zhao, Lihua; Yan, Hao; Zhang, Hua

    2016-02-01

    The reserve of primordial follicles determines the reproductive ability of the female mammal over its reproductive life. The primordial follicle is composed of two types of cells: oocytes and surrounding pre-granulosa cells. However, the underlying mechanism regulating primordial follicle assembly is largely undefined. In this study, we found that gap junction communication (GJC) established between the ovarian cells in the perinatal mouse ovary may be involved in the process. First, gap junction structures between the oocyte and surrounding pre-granulosa cells appear at about 19.0 dpc (days post coitum). As many as 12 gap junction-related genes are upregulated at birth, implying that a complex communication may exist between ovarian cells, because specifically silencing the genes of individual gap junction proteins, such as Gja1, Gja4 or both, has no influence on primordial follicle assembly. On the other hand, non-specific blockers of GJC, such as carbenoxolone (CBX) and 18α-glycyrrhetinic acid (AGA), significantly inhibit mouse primordial follicle assembly. We proved that the temporal window for establishment of GJC in the fetal ovary is from 19.5 dpc to 1 dpp (days postpartum). In addition, the expression of ovarian somatic cell (OSC)-specific genes, such as Notch2, Foxl2 and Irx3, was negatively affected by GJC blockers, whereas oocyte-related genes, such as Ybx2, Nobox and Sohlh1, were hardly affected, implying that the establishment of GJC during this period may be more important to OSCs than to oocytes. In summary, our results indicated that GJC involves in the mouse primordial follicle assembly process at a specific temporal window that needs Notch signaling cross-talking. © 2016 Society for Reproduction and Fertility.

  11. SLO BK Potassium Channels Couple Gap Junctions to Inhibition of Calcium Signaling in Olfactory Neuron Diversification.

    Science.gov (United States)

    Alqadah, Amel; Hsieh, Yi-Wen; Schumacher, Jennifer A; Wang, Xiaohong; Merrill, Sean A; Millington, Grethel; Bayne, Brittany; Jorgensen, Erik M; Chuang, Chiou-Fen

    2016-01-01

    The C. elegans AWC olfactory neuron pair communicates to specify asymmetric subtypes AWCOFF and AWCON in a stochastic manner. Intercellular communication between AWC and other neurons in a transient NSY-5 gap junction network antagonizes voltage-activated calcium channels, UNC-2 (CaV2) and EGL-19 (CaV1), in the AWCON cell, but how calcium signaling is downregulated by NSY-5 is only partly understood. Here, we show that voltage- and calcium-activated SLO BK potassium channels mediate gap junction signaling to inhibit calcium pathways for asymmetric AWC differentiation. Activation of vertebrate SLO-1 channels causes transient membrane hyperpolarization, which makes it an important negative feedback system for calcium entry through voltage-activated calcium channels. Consistent with the physiological roles of SLO-1, our genetic results suggest that slo-1 BK channels act downstream of NSY-5 gap junctions to inhibit calcium channel-mediated signaling in the specification of AWCON. We also show for the first time that slo-2 BK channels are important for AWC asymmetry and act redundantly with slo-1 to inhibit calcium signaling. In addition, nsy-5-dependent asymmetric expression of slo-1 and slo-2 in the AWCON neuron is necessary and sufficient for AWC asymmetry. SLO-1 and SLO-2 localize close to UNC-2 and EGL-19 in AWC, suggesting a role of possible functional coupling between SLO BK channels and voltage-activated calcium channels in AWC asymmetry. Furthermore, slo-1 and slo-2 regulate the localization of synaptic markers, UNC-2 and RAB-3, in AWC neurons to control AWC asymmetry. We also identify the requirement of bkip-1, which encodes a previously identified auxiliary subunit of SLO-1, for slo-1 and slo-2 function in AWC asymmetry. Together, these results provide an unprecedented molecular link between gap junctions and calcium pathways for terminal differentiation of olfactory neurons.

  12. Endothelin uncouples gap junctions in sustentacular cells and olfactory ensheathing cells of the olfactory mucosa.

    Science.gov (United States)

    Le Bourhis, Mikaël; Rimbaud, Stéphanie; Grebert, Denise; Congar, Patrice; Meunier, Nicolas

    2014-09-01

    Several factors modulate the first step of odour detection in the rat olfactory mucosa (OM). Among others, vasoactive peptides such as endothelin might play multifaceted roles in the different OM cells. Like their counterparts in the central nervous system, the olfactory sensory neurons are encompassed by different glial-like non-neuronal OM cells; sustentacular cells (SCs) surround their cell bodies, whereas olfactory ensheathing cells (OECs) wrap their axons. Whereas SCs maintain both the structural and ionic integrity of the OM, OECs assure protection, local blood flow control and guiding of olfactory sensory neuron axons toward the olfactory bulb. We previously showed that these non-neuronal OM cells are particularly responsive to endothelin in vitro. Here, we confirmed that the endothelin system is strongly expressed in the OM using in situ hybridization. We then further explored the effects of endothelin on SCs and OECs using electrophysiological recordings and calcium imaging approaches on both in vitro and ex vivo OM preparations. Endothelin induced both robust calcium signals and gap junction uncoupling in both types of cells. This latter effect was mimicked by carbenoxolone, a known gap junction uncoupling agent. However, although endothelin is known for its antiapoptotic effect in the OM, the uncoupling of gap junctions by carbenoxolone was not sufficient to limit the cellular death induced by serum deprivation in OM primary culture. The functional consequence of the endothelin 1-induced reduction of the gap junctional communication between OM non-neuronal cells thus remains to be elucidated. © 2014 Federation of European Neuroscience Societies and John Wiley & Sons Ltd.

  13. Pharmacological blockade of gap junctions induces repetitive surging of extracellular potassium within the locust CNS.

    Science.gov (United States)

    Spong, Kristin E; Robertson, R Meldrum

    2013-10-01

    The maintenance of cellular ion homeostasis is crucial for optimal neural function and thus it is of great importance to understand its regulation. Glial cells are extensively coupled by gap junctions forming a network that is suggested to serve as a spatial buffer for potassium (K(+)) ions. We have investigated the role of glial spatial buffering in the regulation of extracellular K(+) concentration ([K(+)]o) within the locust metathoracic ganglion by pharmacologically inhibiting gap junctions. Using K(+)-sensitive microelectrodes, we measured [K(+)]o near the ventilatory neuropile while simultaneously recording the ventilatory rhythm as a model of neural circuit function. We found that blockade of gap junctions with either carbenoxolone (CBX), 18β-glycyrrhetinic acid (18β-GA) or meclofenamic acid (MFA) reliably induced repetitive [K(+)]o surges and caused a progressive impairment in the ability to maintain baseline [K(+)]o levels throughout the treatment period. We also show that a low dose of CBX that did not induce surging activity increased the vulnerability of locust neural tissue to spreading depression (SD) induced by Na(+)/K(+)-ATPase inhibition with ouabain. CBX pre-treatment increased the number of SD events induced by ouabain and hindered the recovery of [K(+)]o back to baseline levels between events. Our results suggest that glial spatial buffering through gap junctions plays an essential role in the regulation of [K(+)]o under normal conditions and also contributes to a component of [K(+)]o clearance following physiologically elevated levels of [K(+)]o.

  14. Connexin 30 expression and frequency of connexin heterogeneity in astrocyte gap junction plaques increase with age in the rat retina.

    Science.gov (United States)

    Mansour, Hussein; McColm, Janet R; Cole, Louise; Weible, Michael; Korlimbinis, Anastasia; Chan-Ling, Tailoi

    2013-01-01

    We investigated age-associated changes in retinal astrocyte connexins (Cx) by assaying Cx numbers, plaque sizes, protein expression levels and heterogeneity of gap junctions utilizing six-marker immunohistochemistry (IHC). We compared Wistar rat retinal wholemounts in animals aged 3 (young adult), 9 (middle-aged) and 22 months (aged). We determined that retinal astrocytes have gap junctions composed of Cx26, -30, -43 and -45. Cx30 was consistently elevated at 22 months compared to younger ages both when associated with parenchymal astrocytes and vascular-associated astrocytes. Not only was the absolute number of Cx30 plaques significantly higher (Pgap junctions was demonstrated by the significant increase in the number of Cx26/Cx45 gap junctions with age. We also found gap junctions comprised of 1, 2, 3 or 4 Cx proteins suggesting that retinal astrocytes use various connexin protein combinations in their gap junctions during development and aging. These data provides new insight into the dynamic and extensive Cx network utilized by retinal astrocytes for communication within both the parenchyma and vasculature for the maintenance of normal retinal physiology with age. This characterisation of the changes in astrocytic gap junctional communication with age in the CNS is crucial to the understanding of physiological aging and age-related neurodegenerative diseases.

  15. Critical role of gap junction coupled KATP channel activity for regulated insulin secretion.

    Directory of Open Access Journals (Sweden)

    Jonathan V Rocheleau

    2006-02-01

    Full Text Available Pancreatic beta-cells secrete insulin in response to closure of ATP-sensitive K+ (KATP channels, which causes membrane depolarization and a concomitant rise in intracellular Ca2+ (Cai. In intact islets, beta-cells are coupled by gap junctions, which are proposed to synchronize electrical activity and Cai oscillations after exposure to stimulatory glucose (>7 mM. To determine the significance of this coupling in regulating insulin secretion, we examined islets and beta-cells from transgenic mice that express zero functional KATP channels in approximately 70% of their beta-cells, but normal KATP channel density in the remainder. We found that KATP channel activity from approximately 30% of the beta-cells is sufficient to maintain strong glucose dependence of metabolism, Cai, membrane potential, and insulin secretion from intact islets, but that glucose dependence is lost in isolated transgenic cells. Further, inhibition of gap junctions caused loss of glucose sensitivity specifically in transgenic islets. These data demonstrate a critical role of gap junctional coupling of KATP channel activity in control of membrane potential across the islet. Control via coupling lessens the effects of cell-cell variation and provides resistance to defects in excitability that would otherwise lead to a profound diabetic state, such as occurs in persistent neonatal diabetes mellitus.

  16. Critical role of gap junction coupled KATP channel activity for regulated insulin secretion.

    Science.gov (United States)

    Rocheleau, Jonathan V; Remedi, Maria S; Granada, Butch; Head, W Steven; Koster, Joseph C; Nichols, Colin G; Piston, David W

    2006-02-01

    Pancreatic beta-cells secrete insulin in response to closure of ATP-sensitive K+ (KATP) channels, which causes membrane depolarization and a concomitant rise in intracellular Ca2+ (Cai). In intact islets, beta-cells are coupled by gap junctions, which are proposed to synchronize electrical activity and Cai oscillations after exposure to stimulatory glucose (>7 mM). To determine the significance of this coupling in regulating insulin secretion, we examined islets and beta-cells from transgenic mice that express zero functional KATP channels in approximately 70% of their beta-cells, but normal KATP channel density in the remainder. We found that KATP channel activity from approximately 30% of the beta-cells is sufficient to maintain strong glucose dependence of metabolism, Cai, membrane potential, and insulin secretion from intact islets, but that glucose dependence is lost in isolated transgenic cells. Further, inhibition of gap junctions caused loss of glucose sensitivity specifically in transgenic islets. These data demonstrate a critical role of gap junctional coupling of KATP channel activity in control of membrane potential across the islet. Control via coupling lessens the effects of cell-cell variation and provides resistance to defects in excitability that would otherwise lead to a profound diabetic state, such as occurs in persistent neonatal diabetes mellitus.

  17. Gap junctions linking the dendritic network of GABAergic interneurons in the hippocampus.

    Science.gov (United States)

    Fukuda, T; Kosaka, T

    2000-02-15

    The network of GABAergic interneurons connected by chemical synapses is a candidate for the generator of synchronized oscillations in the hippocampus. We present evidence that parvalbumin (PV)-containing GABAergic neurons in the rat hippocampal CA1 region, known to form a network by mutual synaptic contacts, also form another network connected by dendrodendritic gap junctions. Distal dendrites of PV neurons run parallel to the alveus (hippocampal white matter) and establish multiple contacts with one another at the border between the stratum oriens and the alveus. In electron microscopic serial section analysis, gap junctions could be identified clearly at 24% of these contact sites. A dendrodendritic chemical synapse and a mixed synapse also were found between PV-immunoreactive dendrites. Three-dimensional reconstruction of the dendritic arborization revealed that both PV neurons of the well known vertical type (presumptive basket cells and axoaxonic cells) and those of another horizontal type constitute the dendritic network at the light microscopic level. The extent of dendritic fields of single PV neurons in the lateral direction was 538 +/- 201 micrometer (n = 5) in the vertical type and 838 +/- 159 micrometer (n = 6) in the horizontal type. Our previous and present observations indicate that PV-containing GABAergic neurons in the hippocampus form the dual networks connected by chemical and electrical synapses located at axosomatic and dendrodendritic contact sites, respectively. Gap junctions linking the dendritic network may mediate coherent synaptic inputs to distant interneurons and thereby facilitate the synchronization of oscillatory activities generated in the interneuron network.

  18. Zebrafish Cx35: cloning and characterization of a gap junction gene highly expressed in the retina.

    Science.gov (United States)

    McLachlan, Elizabeth; White, Thomas W; Ugonabo, Chioma; Olson, Carl; Nagy, James I; Valdimarsson, Gunnar

    2003-09-15

    The vertebrate connexin gene family encodes protein subunits of gap junction channels, which provide a route for direct intercellular communication. Consequently, gap junctions play a vital role in many developmental and homeostatic processes. Aberrant functioning of gap junctions is implicated in many human diseases. Zebrafish are an ideal vertebrate model to study development of the visual system as they produce transparent embryos that develop rapidly, thereby facilitating morphological and behavioral testing. In this study, zebrafish connexin35 has been cloned from a P1 artificial chromosome (PAC) library. Sequence analysis shows a high degree of similarity to the Cx35/36 orthologous group, which are expressed primarily in nervous tissue, including the retina. The gene encodes a 304-amino acid protein with a predicted molecular weight of approximately 35 kDa. Injection of zebrafish Cx35 RNA into paired Xenopus oocytes elicited intercellular electrical coupling with weak voltage sensitivity. In development, Cx35 is first detectable by Northern analysis and RT-PCR, at 2 days post-fertilization (2 dpf), and in the adult it is expressed in the brain and retina. Immunohistochemical analysis revealed that the Cx35 protein is expressed in two sublaminae of the inner plexiform layer of the adult retina. A similar pattern was seen in the 4 and 5 dpf retina, but no labeling was detected in the retina of earlier embryos.

  19. A unified framework for spiking and gap-junction interactions in distributed neuronal network simulations

    Directory of Open Access Journals (Sweden)

    Jan eHahne

    2015-09-01

    Full Text Available Contemporary simulators for networks of point and few-compartment model neurons come with a plethora of ready-to-use neuron and synapse models and support complex network topologies. Recent technological advancements have broadened the spectrum of application further to the efficient simulation of brain-scale networks on supercomputers. In distributed network simulations the amount of spike data that accrues per millisecond and process is typically low, such that a common optimization strategy is to communicate spikes at relatively long intervals, where the upper limit is given by the shortest synaptic transmission delay in the network. This approach is well-suited for simulations that employ only chemical synapses but it has so far impeded the incorporation of gap-junction models, which require instantaneous neuronal interactions. Here, we present a numerical algorithm based on a waveform-relaxation technique which allows for network simulations with gap junctions in a way that is compatible with the delayed communication strategy. Using a reference implementation in the NEST simulator, we demonstrate that the algorithm and the required data structures can be smoothly integrated with existing code such that they complement the infrastructure for spiking connections. To show that the unified framework for gap-junction and spiking interactions achieves high performance and delivers high accuracy...

  20. Immunohistochemical localization of a gap junction protein (connexin43) in the muscularis externa of murine, canine, and human intestine

    DEFF Research Database (Denmark)

    Mikkelsen, H B; Huizinga, J D; Thuneberg, L

    1993-01-01

    Electron-microscopic studies have revealed a heterogeneous distribution of gap junctions in the muscularis externa of mammalian intestines. This heterogeneity is observed at four different levels: among species; between small and large intestines; between longitudinal and circular muscle layers; ...

  1. ZP123 increases gap junctional conductance and prevents reentrant ventricular tachycardia during myocardial ischemia in open chest dogs

    DEFF Research Database (Denmark)

    Xing, Dezhi; Kjølbye, Anne Louise; Nielsen, Morten S

    2003-01-01

    INTRODUCTION: The aim of this study was to determine if the stable antiarrhythmic peptide (AAP) analogue ZP123 increases gap junctional intercellular conductance and prevents reentrant ventricular tachycardia (VT) during coronary artery occlusion. METHODS AND RESULTS: Voltage clamp experiments...

  2. Gap junctional regulation of pressure, fluid force, and electrical fields in the epigenetics of cardiac morphogenesis and remodeling.

    Science.gov (United States)

    Seki, Akiko; Nishii, Kiyomasa; Hagiwara, Nobuhisa

    2015-05-15

    Epigenetic factors of pressure load, fluid force, and electrical fields that occur during cardiac contraction affect cardiac development, morphology, function, and pathogenesis. These factors are orchestrated by intercellular communication mediated by gap junctions, which synchronize action potentials and second messengers. Misregulation of the gap junction protein connexin (Cx) alters cardiogenesis, and can be a pathogenic factor causing cardiac conduction disturbance, fatal arrhythmia, and cardiac remodeling in disease states such as hypertension and ischemia. Changes in Cx expression can occur even when the DNA sequence of the Cx gene itself is unaltered. Posttranslational modifications might reduce arrhythmogenic substrates, improve cardiac function, and promote remodeling in a diseased heart. In this review, we discuss the epigenetic features of gap junctions that regulate cardiac morphology and remodeling. We further discuss potential clinical applications of current knowledge of the structure and function of gap junctions. Copyright © 2014 Elsevier Inc. All rights reserved.

  3. Inhibition of dye-coupling in Patella (mollusca) embryos by microinjection of antiserum against Nephrops (arthropoda) gap junctions

    NARCIS (Netherlands)

    Serras, F.; Buultjens, T.E.J.; Finbow, M.E.

    1988-01-01

    Antiserum raised against Nephrops gap junctions was injected into single cells of the 2-, 4-, 8-, 16-, and 32-cell stage of the Patella vulgata embryos. The pattern of junctional communication by iontophoresis of Lucifer Yellow CH was tested at the 32-cell stage. The results show that the normal pat

  4. Inhibition of dye-coupling in Patella (mollusca) embryos by microinjection of antiserum against Nephrops (arthropoda) gap junctions

    NARCIS (Netherlands)

    Serras, F.; Buultjens, T.E.J.; Finbow, M.E.

    Antiserum raised against Nephrops gap junctions was injected into single cells of the 2-, 4-, 8-, 16-, and 32-cell stage of the Patella vulgata embryos. The pattern of junctional communication by iontophoresis of Lucifer Yellow CH was tested at the 32-cell stage. The results show that the normal

  5. The Carboxyl Tail of Connexin32 Regulates Gap Junction Assembly in Human Prostate and Pancreatic Cancer Cells*

    Science.gov (United States)

    Katoch, Parul; Mitra, Shalini; Ray, Anuttoma; Kelsey, Linda; Roberts, Brett J.; Wahl, James K.; Johnson, Keith R.; Mehta, Parmender P.

    2015-01-01

    Connexins, the constituent proteins of gap junctions, are transmembrane proteins. A connexin (Cx) traverses the membrane four times and has one intracellular and two extracellular loops with the amino and carboxyl termini facing the cytoplasm. The transmembrane and the extracellular loop domains are highly conserved among different Cxs, whereas the carboxyl termini, often called the cytoplasmic tails, are highly divergent. We have explored the role of the cytoplasmic tail of Cx32, a Cx expressed in polarized and differentiated cells, in regulating gap junction assembly. Our results demonstrate that compared with the full-length Cx32, the cytoplasmic tail-deleted Cx32 is assembled into small gap junctions in human pancreatic and prostatic cancer cells. Our results further document that the expression of the full-length Cx32 in cells, which express the tail-deleted Cx32, increases the size of gap junctions, whereas the expression of the tail-deleted Cx32 in cells, which express the full-length Cx32, has the opposite effect. Moreover, we show that the tail is required for the clustering of cell-cell channels and that in cells expressing the tail-deleted Cx32, the expression of cell surface-targeted cytoplasmic tail alone is sufficient to enhance the size of gap junctions. Our live-cell imaging data further demonstrate that gap junctions formed of the tail-deleted Cx32 are highly mobile compared with those formed of full-length Cx32. Our results suggest that the cytoplasmic tail of Cx32 is not required to initiate the assembly of gap junctions but for their subsequent growth and stability. Our findings suggest that the cytoplasmic tail of Cx32 may be involved in regulating the permeability of gap junctions by regulating their size. PMID:25548281

  6. A mechanism of gap junction docking revealed by functional rescue of a human-disease-linked connexin mutant.

    Science.gov (United States)

    Gong, Xiang-Qun; Nakagawa, So; Tsukihara, Tomitake; Bai, Donglin

    2013-07-15

    Gap junctions are unique intercellular channels formed by the proper docking of two hemichannels from adjacent cells. Each hemichannel is a hexamer of connexins (Cxs) - the gap junction subunits, which are encoded by 21 homologous genes in the human genome. The docking of two hemichannels to form a functional gap junction channel is only possible between compatible Cxs, but the underlying molecular mechanism is unclear. On the basis of the crystal structure of the Cx26 gap junction, we developed homology models for homotypic and heterotypic channels from Cx32 and/or Cx26; these models predict six hydrogen bonds at the docking interface of each pair of the second extracellular domain (E2). A Cx32 mutation N175H and a human-disease-linked mutant N175D were predicted to lose the majority of the hydrogen bonds at the E2 docking-interface; experimentally both mutations failed to form morphological and functional gap junctions. To restore the lost hydrogen bonds, two complementary Cx26 mutants - K168V and K168A were designed to pair with the Cx32 mutants. When docked with Cx26K168V or K168A, the Cx32N175H mutant was successfully rescued morphologically and functionally in forming gap junction channels, but not Cx32 mutant N175Y. By testing more homotypic and heterotypic Cx32 and/or Cx26 mutant combinations, it is revealed that a minimum of four hydrogen bonds at each E2-docking interface are required for proper docking and functional channel formation between Cx26 and Cx32 hemichannels. Interestingly, the disease-linked Cx32N175D could be rescued by Cx26D179N, which restored five hydrogen bonds at the E2-docking interface. Our findings not only provide a mechanism for gap junction docking for Cx26 and Cx32 hemichannels, but also a potential therapeutic strategy for gap junction channelopathies.

  7. Irsogladine maleate regulates gap junctional intercellular communication-dependent epithelial barrier in human nasal epithelial cells.

    Science.gov (United States)

    Miyata, Ryo; Nomura, Kazuaki; Kakuki, Takuya; Takano, Ken-Ichi; Kohno, Takayuki; Konno, Takumi; Sawada, Norimasa; Himi, Tetsuo; Kojima, Takashi

    2015-04-01

    The airway epithelium of the human nasal mucosa acts as the first physical barrier that protects against inhaled substances and pathogens. Irsogladine maleate (IM) is an enhancer of gastric mucosal protective factors via upregulation of gap junctional intercellular communication (GJIC). GJIC is thought to participate in the formation of functional tight junctions. However, the effects of IM on GJIC and the epithelial barrier in human nasal epithelial cells (HNECs) remain unknown. To investigate the effects of IM on GJIC and the tight junctional barrier in HNECs, primary cultures of HNECs transfected with human telomerase reverse transcriptase (hTERT-HNECs) were treated with IM and the GJIC inhibitors oleamide and 18β-GA. Some cells were pretreated with IM before treatment with TLR3 ligand poly(I:C) to examine whether IM prevented the changes via TLR3-mediated signal pathways. In hTERT-HNECs, GJIC blockers reduced the expression of tight junction molecules claudin-1, -4, -7, occludin, tricellulin, and JAM-A. IM induced GJIC activity and enhanced the expression of claudin-1, -4, and JAM-A at the protein and mRNA levels with an increase of barrier function. GJIC blockers prevented the increase of the tight junction proteins induced by IM. Furthermore, IM prevented the reduction of JAM-A but not induction of IL-8 and TNF-α induced by poly(I:C). In conclusion, IM can maintain the GJIC-dependent tight junctional barrier via regulation of GJIC in upper airway nasal epithelium. Therefore, it is possible that IM may be useful as a nasal spray to prevent the disruption of the epithelial barrier by viral infections and exposure to allergens in human nasal mucosa.

  8. Gap Junction in the Teleost Fish Lineage: Duplicated Connexins May Contribute to Skin Pattern Formation and Body Shape Determination.

    Science.gov (United States)

    Watanabe, Masakatsu

    2017-01-01

    Gap junctions are intercellular channels that allow passage of ions and small molecules between adjacent cells. Gap junctions in vertebrates are composed of connexons, which are an assembly of six proteins, connexins. Docking of two connexons on the opposite cell surfaces forms a gap junction between the cytoplasm of two neighboring cells. Connexins compose a family of structurally related four-pass transmembrane proteins. In mammals, there are ~20 connexins, each of which contributes to unique permeability of gap junctions, and mutations of some connexin-encoding genes are associated with human diseases. Zebrafish has been predicted to contain 39 connexin-encoding genes; the high number can be attributed to gene duplication during fish evolution, which resulted in diversified functions of gap junctions in teleosts. The determination of body shapes and skin patterns in animal species is an intriguing question. Mathematical models suggest principle mechanisms explaining the diversification of animal morphology. Recent studies have revealed the involvement of gap junctions in fish morphological diversity, including skin pattern formation and body shape determination. This review focuses on connexins in teleosts, which are integrated in the mathematical models explaining morphological diversity of animal skin patterns and body shapes.

  9. 异氟烷对心肌动作电位及缝隙连接蛋白Cx43的作用%Effects of Isoflurane on Action Potential of Heart and Expression of Myocardial Connexin 43

    Institute of Scientific and Technical Information of China (English)

    谷宇; 高鸿; 杨烨

    2012-01-01

    目的:观察异氟烷对心肌动作电位及缝隙连接蛋白Cx43表达的影响.方法:健康家兔40只,采用Langendorff离体心脏平衡灌注15min后,随机分为5组:I0.65组,用0.65MAC异氟烷的K-H液灌注;I0,65H用0.65MAC异氟烷加0.5mmol/L庚醇的K-H液灌注;I1.3组,用1.3MAC异氟烷的K-H液灌注;H0.5组,用庚醇0.5mmol/L的K-H液灌注;S组,用K-H液灌注.观察记录灌流15min时(基础值)和给药15min时的心率(HR)及心内膜心肌(Endo)、中层心肌(M)、心外膜心肌(Epi)的动作电位时程(APD)和振幅(APA),灌注完毕取左心室心肌组织,用免疫组织化学检测心肌Cx43蛋白表达.结果:I0.65组心肌动作电位无明显影响,I1.3组APD显著延长(P<0.05),H0.5组HR显著减慢(P<0.05),APD显著延长(P<0.05),k0.65H组给药后有发生心律失常甚至心电活动停止现象;Cx43蛋白表达I0.65H组较H0.5组表达少(P<0.05),H0.5组较I0.65、I1.3组表达少(P<0.05),I1.3组较I0.65组、S组表达少(P<0.05).结论:异氟烷与庚醇对心脏动作电位和缝隙连接蛋白Cx43可能有着相似的作用,延长动作单位时程,减少Cx43蛋白表达,改变Cx43蛋白分布;异氟烷可能通过阻滞缝隙连接,对心脏动作电位产生作用.%Objective: To observe the effects of isoflurane on action potential ( AP) of heart and the expression of myocardial connexin43 (Cx43). Methods: Forty isolated rabbit hearts were perfused for 15 minutes with modified Krebs-Hensleit buffer using Langendorff device. Then, all the hearts were randomly divided into five groups (re =8 in each) : group I0.65, group I0.65H , group 1.3, group t0. 5 and group S, in which hearts were perfused with Kreb-Hensleit buffer containing 0. 65 MAC isoflurane, 0.65 MAC isoflurane plus 0.5 mmol/L heptanol, 1.3 MAC isoflurane, 0.5 mmol/L heptanol and Kreb-Hensleit buffer only respectively. Heart rates ( HR) , AP course ( APD) and AP amplitude ( APA) of endocardium ( Endo) , mid-layer myocardium ( M ) and

  10. Alteration of cadherin isoform expression and inhibition of gap junctions in stomach carcinoma cells

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    To explore cell malignant phenotype correlated changes of cell surface adhesion molecules and cell-cell communication in carcinogenesis, human stomach transformed and cancer cell lines were investigated. Expressions of E-cadherin, N-cadherin, ?-catenin, ?-catenin as well as gap junction (GJ) protein Cx32 were studied by utilization of immunoblotting, immunocytochemical and fluorescent dye transfer methods. Mammalian normal stomach mucosal cells expressed E-cadherin but not N-cadherin. E-cadherin immunofluorescence was detected at cell membranous adherens junctions (AJ) where colocalization with immunofluorescent staining of inner surface adhesion plaque proteins ?- and ?-catenins was observed. The existence of E-cadherin/ catenin (?-, ?-) protein complexes as AJ was suggested. In transformed and stomach cancer cells E-cadherin was inhibited, instead, N-cadherin was expressed and localized at membranous AJ where co-staining with ?- and ?-catenin fluorescence was observed. Formation of N-cadherin/catenin (?-, ?-) protein complex at AJs of transformed and cancer cells was suggested. The above observations were further supported by immunoblotting results. Normal stomach muscosal and transformed cells expressed Cx32 at membranous GJ and were competent of gap junction communication (GJIC). In stomach cancer cells, Cx32 was inhibited and GJIC was defective. The results suggested that changes of signal pathways mediated by both cell adhesion and cell communication systems are associated intracellular events of stomach carcinogenesis. The alteration of cadherin isoform from E- to N-cadherin in transformed and stomach cancer cells is the first report.

  11. Expression of gap junction protein connexin 43 in bovine urinary bladder tumours.

    Science.gov (United States)

    Corteggio, A; Florio, J; Roperto, F; Borzacchiello, G

    2011-01-01

    The aetiopathogenesis of urinary bladder tumours in cattle involves prolonged ingestion of bracken fern and infection by bovine papillomavirus types 1 or 2 (BPV-1/2). The oncogenic activity of BPV is largely associated with the major oncoprotein E5. Gap junctions are the only communicating junctions found in animal tissues and are composed of proteins known as connexins. Alterations in connexin expression have been associated with oncogenesis. The present study investigated biochemically and immunohistochemically the expression of connexin 43 in samples of normal (n=2), dysplastic (n=3) and neoplastic (n=23) bovine urothelium. The tumours included 10 carcinomas in situ, five papillary urothelial carcinomas and eight invasive urothelial carcinomas. Normal and dysplastic urothelium had membrane expression of connexin 43, but this was reduced in samples of carcinoma in situ. Papillary urothelial carcinomas showed moderate cytoplasmic and membrane labelling, while invasive carcinoma showed loss of connexin 43 expression. Copyright © 2010 Elsevier Ltd. All rights reserved.

  12. Band-gaps in long Josephson junctions with periodic phase-shifts

    Science.gov (United States)

    Ahmad, Saeed; Susanto, Hadi; Wattis, Jonathan A. D.

    2017-04-01

    We investigate analytically and numerically a long Josephson junction on an infinite domain, having arbitrary periodic phase shift of κ, that is, the so-called 0-κ long Josephson junction. The system is described by a one-dimensional sine-Gordon equation and has relatively recently been proposed as artificial atom lattices. We discuss the existence of periodic solutions of the system and investigate their stability both in the absence and presence of an applied bias current. We find critical values of the phase-discontinuity and the applied bias current beyond which static periodic solutions cease to exist. Due to the periodic discontinuity in the phase, the system admits regions of allowed and forbidden bands. We perturbatively investigate the Arnold tongues that separate the region of allowed and forbidden bands, and discuss the effect of an applied bias current on the band-gap structure. We present numerical simulations to support our analytical results.

  13. Entrainment, retention, and transport of freely swimming fish in junction gaps between commercial barges operating on the Illinois Waterway

    Science.gov (United States)

    Davis, Jeremiah J.; Jackson, Patrick; Engel, Frank; LeRoy, Jessica Z.; Neeley, Rebecca N.; Finney, Samuel T.; Murphy, Elizabeth A.

    2016-01-01

    Large Electric Dispersal Barriers were constructed in the Chicago Sanitary and Ship Canal (CSSC) to prevent the transfer of invasive fish species between the Mississippi River Basin and the Great Lakes Basin while simultaneously allowing the passage of commercial barge traffic. We investigated the potential for entrainment, retention, and transport of freely swimming fish within large gaps (> 50 m3) created at junction points between barges. Modified mark and capture trials were employed to assess fish entrainment, retention, and transport by barge tows. A multi-beam sonar system enabled estimation of fish abundance within barge junction gaps. Barges were also instrumented with acoustic Doppler velocity meters to map the velocity distribution in the water surrounding the barge and in the gap formed at the junction of two barges. Results indicate that the water inside the gap can move upstream with a barge tow at speeds near the barge tow travel speed. Water within 1 m to the side of the barge junction gaps was observed to move upstream with the barge tow. Observed transverse and vertical water velocities suggest pathways by which fish may potentially be entrained into barge junction gaps. Results of mark and capture trials provide direct evidence that small fish can become entrained by barges, retained within junction gaps, and transported over distances of at least 15.5 km. Fish entrained within the barge junction gap were retained in that space as the barge tow transited through locks and the Electric Dispersal Barriers, which would be expected to impede fish movement upstream.

  14. Osteoclastogenesis is influenced by modulation of gap junctional communication with antiarrhythmic peptides.

    Science.gov (United States)

    Kylmäoja, Elina; Kokkonen, Hanna; Kauppinen, Kyösti; Hussar, Piret; Sato, Tetsuji; Haugan, Ketil; Larsen, Bjarne Due; Tuukkanen, Juha

    2013-03-01

    Osteoclasts are formed by the fusion of mononuclear precursor cells of the monocyte-macrophage lineage. Among several putative mechanisms, gap-junctional intercellular communication (GJC) has been proposed to have a role in osteoclast fusion and bone resorption. We examined the role of GJC in osteoclastogenesis and in vitro bone resorption with mouse bone marrow hematopoietic stem cells and RAW 264.7 cells. Blocking of gap junctions with 18-α-glycyrrhetinic acid (18GA) led to inhibition of osteoclastogenesis and in vitro bone resorption. Similarly, the GJC inhibitor GAP27 inhibited osteoclast formation. GJC modulation with the antiarrhythmic peptides (AAPs) led to increased amounts of multinuclear RAW 264.7 osteoclasts as well as increased number of nuclei per multinuclear cell. In the culture of bone marrow hematopoietic stem cells in the presence of bone marrow stromal cells AAP reduced the number of osteoclasts, and coculture of MC3T3-E1 preosteoblasts with RAW 264.7 macrophages prevented the action of AAPs to promote osteoclastogenesis. The present data indicate that AAPs modulate the fusion of the pure culture of cells of the monocyte-macrophage lineage. However, the fusion is influenced by GJC in cells of the osteoblast lineage.

  15. Massive Dirac fermion transport in a gapped graphene-based magnetic tunnel junction

    Science.gov (United States)

    Soodchomshom, Bumned; Tang, I.-Ming; Hoonsawat, Rassmidara

    2009-08-01

    The spin transport in a graphene-based magnetic (NG/ferromagnetic barrier (FB)/NG) tunnel junction with the graphene sheet being grown on a SiC substrate is investigated. Zhou et al. [Nat. Mater. 6 (2007) 770] has shown that in these epitaxial grown graphene sheets, the electrons behave like massive relativistic particles with an energy gap of 2 Δ∼260 meV opening up in the energy spectrum of the massive relativistic electron. Basing on assumption that gap in graphene can occur under the influence of the magnetic field, we find that in the case of thick ferromagnetic graphene barriers, the electronic gap causes the barrier to behave as a strong insulator when the gate potential is in the range 400-130 meVswitched from a 100% spin up current to a 100% spin down current by small variation of V G from V G E f , the features of a perfect spin filtering electronic junction.

  16. Intracerebroventricular injection of lipopolysaccharide increases gene expression of connexin32 gap junction in rat hippocampus.

    Science.gov (United States)

    Abbasian, Mohammad; Sayyah, Mohammad; Babapour, Vahab; Mahdian, Reza

    2013-01-01

    Gap junctions are intercellular membrane channels that provide direct cytoplasmic continuity between adjacent cells. This communication can be affected by changes in expression of gap junctional subunits called Connexins (Cx). Changes in the expression and function of connexins are associated with number of brain neurodegenerative diseases. Neuroinflammation is a hallmark of various central nervous system (CNS) diseases, like multiple sclerosis, Alzheimer's disease and epilepsy. Neuroinflammation causes change in Connexins expression. Hippocampus, one of the main brain regions with a wide network of Gap junctions between different neural cell types, has particular vulnerability to damage and consequent inflammation. Cx32 - among Connexins- is expressed in hippocampal Olygodandrocytes and some neural subpopulations. Although multiple lines of evidence indicate that there is an association between neuroinflammation and the expression of connexin, the direct effect of neuroinflammation on the expression of connexins has not been well studied. In the present study, the effect of neuroinflammation induced by the Lipopolysaccharide (LPS) on Cx32 gene and protein expressions in rat hippocampus is evaluated. LPS (2.5µg/rat) was infused into the rat cerebral ventricles for 14 days. Cx32 mRNA and protein levels were measured by Real Time PCR and Western Blot after 1st, 7th and 14th injection of LPS in the hippocampus. Significant increase in Cx32 mRNA expression was observed after 7th injection of LPS (P < 0.001). However, no significant change was observed in Cx32 protein level. LPS seems to modify Cx32 GJ communication in the hippocampus at transcription level but not at translation or post-translation level. In order to have a full view concerning modification of Cx32 GJ communication, effect of LPS on Cx32 channel gating should also be determined.

  17. Simvastatin Sodium Salt and Fluvastatin Interact with Human Gap Junction Gamma-3 Protein.

    Science.gov (United States)

    Marsh, Andrew; Casey-Green, Katherine; Probert, Fay; Withall, David; Mitchell, Daniel A; Dilly, Suzanne J; James, Sean; Dimitri, Wade; Ladwa, Sweta R; Taylor, Paul C; Singer, Donald R J

    2016-01-01

    Finding pleiomorphic targets for drugs allows new indications or warnings for treatment to be identified. As test of concept, we applied a new chemical genomics approach to uncover additional targets for the widely prescribed lipid-lowering pro-drug simvastatin. We used mRNA extracted from internal mammary artery from patients undergoing coronary artery surgery to prepare a viral cardiovascular protein library, using T7 bacteriophage. We then studied interactions of clones of the bacteriophage, each expressing a different cardiovascular polypeptide, with surface-bound simvastatin in 96-well plates. To maximise likelihood of identifying meaningful interactions between simvastatin and vascular peptides, we used a validated photo-immobilisation method to apply a series of different chemical linkers to bind simvastatin so as to present multiple orientations of its constituent components to potential targets. Three rounds of biopanning identified consistent interaction with the clone expressing part of the gene GJC3, which maps to Homo sapiens chromosome 7, and codes for gap junction gamma-3 protein, also known as connexin 30.2/31.3 (mouse connexin Cx29). Further analysis indicated the binding site to be for the N-terminal domain putatively 'regulating' connexin hemichannel and gap junction pores. Using immunohistochemistry we found connexin 30.2/31.3 to be present in samples of artery similar to those used to prepare the bacteriophage library. Surface plasmon resonance revealed that a 25 amino acid synthetic peptide representing the discovered N-terminus did not interact with simvastatin lactone, but did bind to the hydrolysed HMG CoA inhibitor, simvastatin acid. This interaction was also seen for fluvastatin. The gap junction blockers carbenoxolone and flufenamic acid also interacted with the same peptide providing insight into potential site of binding. These findings raise key questions about the functional significance of GJC3 transcripts in the vasculature and

  18. Simvastatin Sodium Salt and Fluvastatin Interact with Human Gap Junction Gamma-3 Protein.

    Directory of Open Access Journals (Sweden)

    Andrew Marsh

    Full Text Available Finding pleiomorphic targets for drugs allows new indications or warnings for treatment to be identified. As test of concept, we applied a new chemical genomics approach to uncover additional targets for the widely prescribed lipid-lowering pro-drug simvastatin. We used mRNA extracted from internal mammary artery from patients undergoing coronary artery surgery to prepare a viral cardiovascular protein library, using T7 bacteriophage. We then studied interactions of clones of the bacteriophage, each expressing a different cardiovascular polypeptide, with surface-bound simvastatin in 96-well plates. To maximise likelihood of identifying meaningful interactions between simvastatin and vascular peptides, we used a validated photo-immobilisation method to apply a series of different chemical linkers to bind simvastatin so as to present multiple orientations of its constituent components to potential targets. Three rounds of biopanning identified consistent interaction with the clone expressing part of the gene GJC3, which maps to Homo sapiens chromosome 7, and codes for gap junction gamma-3 protein, also known as connexin 30.2/31.3 (mouse connexin Cx29. Further analysis indicated the binding site to be for the N-terminal domain putatively 'regulating' connexin hemichannel and gap junction pores. Using immunohistochemistry we found connexin 30.2/31.3 to be present in samples of artery similar to those used to prepare the bacteriophage library. Surface plasmon resonance revealed that a 25 amino acid synthetic peptide representing the discovered N-terminus did not interact with simvastatin lactone, but did bind to the hydrolysed HMG CoA inhibitor, simvastatin acid. This interaction was also seen for fluvastatin. The gap junction blockers carbenoxolone and flufenamic acid also interacted with the same peptide providing insight into potential site of binding. These findings raise key questions about the functional significance of GJC3 transcripts in the

  19. Connexin50 couples axon terminals of mouse horizontal cells by homotypic gap junctions.

    Science.gov (United States)

    Dorgau, Birthe; Herrling, Regina; Schultz, Konrad; Greb, Helena; Segelken, Jasmin; Ströh, Sebastian; Bolte, Petra; Weiler, Reto; Dedek, Karin; Janssen-Bienhold, Ulrike

    2015-10-01

    Horizontal cells in the mouse retina are of the axon-bearing B-type and contribute to the gain control of photoreceptors and to the center-surround organization of bipolar cells by providing feedback and feedforward signals to photoreceptors and bipolar cells, respectively. Horizontal cells form two independent networks, coupled by dendro-dendritic and axo-axonal gap junctions composed of connexin57 (Cx57). In Cx57-deficient mice, occasionally the residual tracer coupling of horizontal cell somata was observed. Also, negative feedback from horizontal cells to photoreceptors, potentially mediated by connexin hemichannels, appeared unaffected. These results point to the expression of a second connexin in mouse horizontal cells. We investigated the expression of Cx50, which was recently identified in axonless A-type horizontal cells of the rabbit retina. In the mouse retina, Cx50-immunoreactive puncta were predominantly localized on large axon terminals of horizontal cells. Electron microscopy did not reveal any Cx50-immunolabeling at the membrane of horizontal cell tips invaginating photoreceptor terminals, ruling out the involvement of Cx50 in negative feedback. Moreover, Cx50 colocalized only rarely with Cx57 on horizontal cell processes, indicating that both connexins form homotypic rather than heterotypic or heteromeric gap junctions. To check whether the expression of Cx50 is changed when Cx57 is lacking, we compared the Cx50 expression in wildtype and Cx57-deficient mice. However, Cx50 expression was unaffected in Cx57-deficient mice. In summary, our results indicate that horizontal cell axon terminals form two independent sets of homotypic gap junctions, a feature which might be important for light adaptation in the retina. © 2015 Wiley Periodicals, Inc.

  20. The spike timing precision of FitzHugh-Nagumo neuron network coupled by gap junctions

    Institute of Scientific and Technical Information of China (English)

    Zhang Su-Hua; Zhan Yong; Yu Hui; An Hai-Long; Zhao Tong-Jun

    2006-01-01

    It has been proved recently that the spike timing can play an important role in information transmission, so in this paper we develop a network with N-unit FitzHugh-Nagumo neurons coupled by gap junctions and discuss the dependence of the spike timing precision on synaptic coupling strength, the noise intensity and the size of the neuron ensemble. The calculated results show that the spike timing precision decreases as the noise intensity increases; and the ensemble spike timing precision increases with coupling strength increasing. The electric synapse coupling has a more important effect on the spike timing precision than the chemical synapse coupling.

  1. The study of the expression of the Cx43 in the oral squamous cell carcinoma and Tca8113%间隙连接蛋白43在口腔颌面部鳞癌组织及舌癌细胞株(Tca8113)中表达的研究

    Institute of Scientific and Technical Information of China (English)

    宋娟; 张汉东

    2012-01-01

    目的:探讨间隙连接蛋白43(Cx43)在口腔鳞癌组织(oscc)和舌鳞癌细胞株(Tca8113)中的表达及其生物学行为的关系:细胞分化诱导剂全反式维甲酸(ATRA)对人舌鳞癌细胞株(Tca8113)中Cx43表达的影响.方法:应用免疫组织、细胞化学、免疫荧光及结合免疫印迹技术,观察OSCC组织中以及ATRA对Tca8113细胞株进行培养及分化诱导后Cx43在蛋白水平的变化.结果:Cx43在正常口腔鳞状上皮主要表达于相邻细胞间相互接触的细胞膜上,而在OSCC组织中,Cx43细胞膜表达与组织的分化程度、转移之间有显著性差异(Cx43:x2=7.42、12.43,P<0.05、P<0.01).免疫细胞化学检测可见舌癌细胞中Cx43异常定位于细胞浆中和/或细胞膜不与邻近细胞接触的游离缘上.Tca8113经ATRA诱导后,Cx43蛋白表达增加.结论:Cx43表达的改变与OSCC的发生和发展有关.ATRA可以通过上调Cx43表达,实现对肿瘤生长的抑制%Objective: To investigate the expression of connexin 43 and its relationship with the biological behavior in oral squamous cell carcinoma (OSCC) and Tca8113;to investigate the effects of connexin 43 in tongue carcinoma cell line (Tca8113) by all-trans-retinoic acid (ATRA). Method: Immunohistochemical and immunofluorescence was used to detect the expressions of Cx43 in oral squamous carcinoma. Western blot was adopted to examine the expressions of Cx43 in tongue carcinoma cell line (Tca8113) by all-trans-retinoic acid (ATRA). Result: In normal oral squamous epithelium the expression of Cx43 was typically shown in the membrane at intercellular contact,but in OSCC they were aberrantly expressed in cytoplasm.The significant difference was observed between low Cx43 expression and differentiation,metastasis (Cx43:X2= 7.42,12.43,P <0.05,P <0.01).In Tca8113 cell line Cx43 was detected mainly either intracytoplas-mically or in plasma membrane free from contact with other cells by immunocytochemistry and immunofluorescence

  2. Potassium ion recycling pathway via gap junction systems in the mammalian cochlea and its interruption in hereditary nonsyndromic deafness.

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    Kikuchi, T; Adams, J C; Miyabe, Y; So, E; Kobayashi, T

    2000-01-01

    In the mammalian cochlea, there are two independent gap junction systems, the epithelial cell gap junction system and the connective tissue cell gap junction system. Thus far, four different connexin molecules, including connexin 26, 30, 31, and 43, have been reported in the cochlea. The two networks of gap junctions form the route by which K+ ions that pass through the sensory cells during mechanosensory transduction can be recycled back to the endolymphatic space, from which they reenter the sensory cells. Activation of hair cells by acoustic stimuli induces influx of K+ ions from the endolymph to sensory hair cells. These K+ ions are released basolaterally to the extracellular space of the organ of Corti, from which they enter the cochlear supporting cells. Once inside the supporting cells they move via the epithelial cell gap junction system laterally to the lower part of the spiral ligament. The K+ ions are released into the extracellular space of the spiral ligament by root cells and taken up by type II fibrocytes. This uptake incorporates K+ into the connective tissue gap junction system. Within this system, the K+ ions pass through the tight junctional barrier of the stria vascularis and are released within the intrastrial extracellular space. The marginal cells of the stria vascularis then take up K+ and return it to the endolymphatic space, where it can be used again in sensory transduction. It is highly probable that mutations of connexin genes that result in human nonsyndromic deafness cause dysfunction of cochlear gap junctions and thereby interrupt K+ ion recirculation pathways. In addition to connexin mutations, other conditions may disrupt gap junctions within the ear. For example, mice with a functionally significant mutation of Brain-4, which is expressed in the connective tissue cells within the cochlea, show marked depression of the endolymphatic potential and profound sensorineural hearing loss. It seems likely that disruption of connective

  3. Blockade of gap junction hemichannel suppresses disease progression in mouse models of amyotrophic lateral sclerosis and Alzheimer's disease.

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    Takeuchi, Hideyuki; Mizoguchi, Hiroyuki; Doi, Yukiko; Jin, Shijie; Noda, Mariko; Liang, Jianfeng; Li, Hua; Zhou, Yan; Mori, Rarami; Yasuoka, Satoko; Li, Endong; Parajuli, Bijay; Kawanokuchi, Jun; Sonobe, Yoshifumi; Sato, Jun; Yamanaka, Koji; Sobue, Gen; Mizuno, Tetsuya; Suzumura, Akio

    2011-01-01

    Glutamate released by activated microglia induces excitotoxic neuronal death, which likely contributes to non-cell autonomous neuronal death in neurodegenerative diseases, including amyotrophic lateral sclerosis and Alzheimer's disease. Although both blockade of glutamate receptors and inhibition of microglial activation are the therapeutic candidates for these neurodegenerative diseases, glutamate receptor blockers also perturbed physiological and essential glutamate signals, and inhibitors of microglial activation suppressed both neurotoxic/neuroprotective roles of microglia and hardly affected disease progression. We previously demonstrated that activated microglia release a large amount of glutamate specifically through gap junction hemichannel. Hence, blockade of gap junction hemichannel may be potentially beneficial in treatment of neurodegenerative diseases. In this study, we generated a novel blood-brain barrier permeable gap junction hemichannel blocker based on glycyrrhetinic acid. We found that pharmacologic blockade of gap junction hemichannel inhibited excessive glutamate release from activated microglia in vitro and in vivo without producing notable toxicity. Blocking gap junction hemichannel significantly suppressed neuronal loss of the spinal cord and extended survival in transgenic mice carrying human superoxide dismutase 1 with G93A or G37R mutation as an amyotrophic lateral sclerosis mouse model. Moreover, blockade of gap junction hemichannel also significantly improved memory impairments without altering amyloid β deposition in double transgenic mice expressing human amyloid precursor protein with K595N and M596L mutations and presenilin 1 with A264E mutation as an Alzheimer's disease mouse model. Our results suggest that gap junction hemichannel blockers may represent a new therapeutic strategy to target neurotoxic microglia specifically and prevent microglia-mediated neuronal death in various neurodegenerative diseases.

  4. Blockade of gap junction hemichannel suppresses disease progression in mouse models of amyotrophic lateral sclerosis and Alzheimer's disease.

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    Hideyuki Takeuchi

    Full Text Available BACKGROUND: Glutamate released by activated microglia induces excitotoxic neuronal death, which likely contributes to non-cell autonomous neuronal death in neurodegenerative diseases, including amyotrophic lateral sclerosis and Alzheimer's disease. Although both blockade of glutamate receptors and inhibition of microglial activation are the therapeutic candidates for these neurodegenerative diseases, glutamate receptor blockers also perturbed physiological and essential glutamate signals, and inhibitors of microglial activation suppressed both neurotoxic/neuroprotective roles of microglia and hardly affected disease progression. We previously demonstrated that activated microglia release a large amount of glutamate specifically through gap junction hemichannel. Hence, blockade of gap junction hemichannel may be potentially beneficial in treatment of neurodegenerative diseases. METHODS AND FINDINGS: In this study, we generated a novel blood-brain barrier permeable gap junction hemichannel blocker based on glycyrrhetinic acid. We found that pharmacologic blockade of gap junction hemichannel inhibited excessive glutamate release from activated microglia in vitro and in vivo without producing notable toxicity. Blocking gap junction hemichannel significantly suppressed neuronal loss of the spinal cord and extended survival in transgenic mice carrying human superoxide dismutase 1 with G93A or G37R mutation as an amyotrophic lateral sclerosis mouse model. Moreover, blockade of gap junction hemichannel also significantly improved memory impairments without altering amyloid β deposition in double transgenic mice expressing human amyloid precursor protein with K595N and M596L mutations and presenilin 1 with A264E mutation as an Alzheimer's disease mouse model. CONCLUSIONS: Our results suggest that gap junction hemichannel blockers may represent a new therapeutic strategy to target neurotoxic microglia specifically and prevent microglia-mediated neuronal

  5. Role of gap junctions in acetylcholine-induced vasodilation of proximal and distal arteries of the rat mesentery.

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    Hill, C E; Hickey, H; Sandow, S L

    2000-07-01

    We have previously shown that myoendothelial gap junctions are more prevalent in distal than in proximal arteries of the rat mesentery. In the present study we have investigated the role of gap junctions in the mechanism of action of endothelium-derived hyperpolarizing factor (EDHF) in these same vessels following relaxation with acetylcholine. Arteries were pre-constricted with phenylephrine and concentration response curves to acetylcholine were constructed in the presence of N(G)-nitro-L-arginine methyl ester (L-NAME; 10(-5) M) and indomethacin (10(-5) M) to prevent effects due to the release of nitric oxide and prostacyclins. Nitric oxide was found to have only a small role in the relaxation of the proximal vessels and was not involved in the relaxations of the distal vessels. 18 alpha-Glycyrrhetinic acid (10(-5) M), a putative gap junction uncoupler, significantly reduced acetylcholine-induced relaxations by 50% in both proximal and distal vessels. Potassium channel antagonists, tetraethylammonium chloride (TEA; 10(-3) M) and barium chloride (10(-4) M), together abolished the dilatory response in the proximal mesenteric arteries, but did not completely block responses in the distal arteries. The data suggest that gap junctions contribute significantly to the acetylcholine-induced relaxation in both proximal and distal arteries of the rat mesentery. We hypothesize that the absence of a correlation between the role of gap junctions and the incidence of myoendothelial gap junctions in these same vessels is due to significant effects of the inhibitors on gap junctions located in the smooth muscle layers of the larger vessels.

  6. A computational approach to detect gap junction plaques and associate them with cells in fluorescent images.

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    Goldberg, Joshua S; Vadakkan, Tegy J; Hirschi, Karen K; Dickinson, Mary E

    2013-04-01

    Intercellular signaling is a fundamental requirement for complex biological system function and survival. Communication between adjoining cells is largely achieved via gap junction channels made up of multiple subunits of connexin proteins, each with unique selectivity and regulatory properties. Intercellular communication via gap junction channels facilitates transmission of an array of cellular signals, including ions, macromolecules, and metabolites that coordinate physiological processes throughout tissues and entire organisms. Although current methods used to quantify connexin expression rely on number or area density measurements in a field of view, they lack cellular assignment, distance measurement capabilities (both within the cell and to extracellular structures), and complete automation. We devised an automated computational approach built on a contour expansion algorithm platform that allows connexin protein detection and assignment to specific cells within complex tissues. In addition, parallel implementation of the contour expansion algorithm allows for high-throughput analysis as the complexity of the biological sample increases. This method does not depend specifically on connexin identification and can be applied more widely to the analysis of numerous immunocytochemical markers as well as to identify particles within tissues such as nanoparticles, gene delivery vehicles, or even cellular fragments such as exosomes or microparticles.

  7. Gap junctions are essential for generating the correlated spike activity of neighboring retinal ganglion cells.

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    Béla Völgyi

    Full Text Available Neurons throughout the brain show spike activity that is temporally correlated to that expressed by their neighbors, yet the generating mechanism(s remains unclear. In the retina, ganglion cells (GCs show robust, concerted spiking that shapes the information transmitted to central targets. Here we report the synaptic circuits responsible for generating the different types of concerted spiking of GC neighbors in the mouse retina. The most precise concerted spiking was generated by reciprocal electrical coupling of GC neighbors via gap junctions, whereas indirect electrical coupling to a common cohort of amacrine cells generated the correlated activity with medium precision. In contrast, the correlated spiking with the lowest temporal precision was produced by shared synaptic inputs carrying photoreceptor noise. Overall, our results demonstrate that different synaptic circuits generate the discrete types of GC correlated activity. Moreover, our findings expand our understanding of the roles of gap junctions in the retina, showing that they are essential for generating all forms of concerted GC activity transmitted to central brain targets.

  8. Self-organized synchronous oscillations in a network of excitable cells coupled by gap junctions.

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    Lewis, T J; Rinzel, J

    2000-11-01

    Recent evidence suggests that electrical coupling plays a role in generating oscillatory behaviour in networks of neurons; however, the underlying mechanisms have not been identified. Using a cellular automata model proposed by Traub et al (Traub R D, Schmitz D, Jefferys J G and Draguhn A 1999 High-frequency population oscillations are predicted to occur in hippocampal pyramidal neural networks interconnected by axo-axonal gap junctions Neuroscience 92 407-26), we describe a novel mechanism for self-organized oscillations in networks that have strong, sparse random electrical coupling via gap junctions. The network activity is generated by random spontaneous activity that is moulded into regular population oscillations by the propagation of activity through the network. We explain how this activity gives rise to particular dependences of mean oscillation frequency on network connectivity parameters and on the rate of spontaneous activity, and we derive analytical expressions to approximate the mean frequency and variance of the oscillations. In doing so, we provide insight into possible mechanisms for frequency control and modulation in networks of neurons.

  9. Treatment with the gap junction modifier rotigaptide (ZP123) reduces infarct size in rats with chronic myocardial infarction.

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    Haugan, Ketil; Marcussen, Niels; Kjølbye, Anne Louise; Nielsen, Morten Schak; Hennan, James K; Petersen, Jørgen Søberg

    2006-02-01

    Treatment with non-selective drugs (eg, long-chain alcohols, halothane) that reduce gap junction intercellular communication (GJIC) is associated with reduced infarct size after myocardial infarction (MI). Therefore, it has been suggested that gap junction intercellular communication stimulating compounds may increase infarct size. The antiarrhythmic peptide analogue rotigaptide (ZP123) increases cardiac gap junction intercellular communication and the purpose of the present study was to examine the effects of rotigaptide treatment on infarct size. Myocardial infarction was induced in male rats by ligation of the left anterior descending artery (LAD). Rats (n = 156) were treated with rotigaptide at three dose levels or vehicle from the onset of ischemia and for 3 weeks following LAD occlusion. Infarct size was determined using histomorphometry after 3 weeks treatment. Rotigaptide treatment producing steady state plasma levels of 0.8 +/- 0.1, 5.5 +/- 0.5, and 86 +/- 8 nmol/L had no effect on mortality, but reduced infarct size to 90 +/- 10% (P = 0.41), 67 +/- 7% (P = 0.005), and 82 +/- 7% (P = 0.13), respectively relative to vehicle-treated myocardial infarction rats (100 +/- 12%). In contrast to what was predicted, our data demonstrates that rotigaptide treatment was associated with a significant infarct size reduction. We conclude that whereas treatment with non-selective inhibitors of gap junction intercellular communication cause a reduction in infarct size, this information cannot be extrapolated to the effects of compounds that selectively increase gap junction intercellular communication.

  10. Connexin36 Gap Junction Blockade Is Ineffective at Reducing Seizure-Like Event Activity in Neocortical Mouse Slices

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    Logan J. Voss

    2010-01-01

    Full Text Available Despite much research, there remains controversy over the role of gap junctions in seizure processes. Many studies report anticonvulsant effects of gap junction blockade, but contradictory results have also been reported. The aim of this study was to clarify the role of connexin36 (Cx36 gap junctions in neocortical seizures. We used the mouse neocortical slice preparation to investigate the effect of pharmacological (mefloquine and genetic (Cx36 knockout mice (Cx36KO manipulation of Cx36 gap junctions on two seizure models: low-magnesium artificial cerebrospinal fluid (ACSF and aconitine perfusion in low-magnesium ACSF. Low-magnesium- (nominally zero and aconitine- (230 nM induced seizure-like event (SLE population activity was recorded extracellularly. The results were consistent in showing that neither mefloquine (25 μM nor genetic knockdown of Cx36 expression had anticonvulsant effects on SLE activity generated by either method. These findings call into question the widely held idea that open Cx36 gap junctions promote seizure activity.

  11. Perfect GMR effect in gapped graphene-based ferromagnetic normal ferromagnetic junctions

    Institute of Scientific and Technical Information of China (English)

    Hossein Karbaschi; Gholam Reza Rashedi

    2015-01-01

    We investigate the quantum transport property in gapped graphene-based ferromagnetic/normal/ferromagnetic (FG/NG/FG) junctions by using the Dirac–Bogoliubov–de Gennes equation. The graphene is fabricated on SiC and BN substrates separately, so carriers in FG/NG/FG structures are considered as massive relativistic particles. Transmission prob-ability, charge, and spin conductances are studied as a function of exchange energy of ferromagnets (h), size of graphene gap, and thickness of normal graphene region (L) respectively. Using the experimental values of Fermi energy in the normal graphene part (EFN∼400 meV) and energy gap in graphene (260 meV for SiC and 50 meV for BN substrate), it is shown that this structure can be used for both spin-up and spin-down polarized current. The latter case has different behavior of gapped FG/NG/FG from that of gapless FG/NG/FG structures. Also perfect charge giant magnetoresistance is observed in a range of EFN−mv2F

  12. Connexin43 hemichannel-mediated regulation of connexin43.

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    Kai Li

    Full Text Available BACKGROUND: Many signaling molecules and pathways that regulate gap junctions (GJs protein expression and function are, in fact, also controlled by GJs. We, therefore, speculated an existence of the GJ channel-mediated self-regulation of GJs. Using a cell culture model in which nonjunctional connexin43 (Cx43 hemichannels were activated by cadmium (Cd(2+, we tested this hypothesis. PRINCIPAL FINDINGS: Incubation of Cx43-transfected LLC-PK1 cells with Cd(2+ led to an increased expression of Cx43. This effect of Cd(2+ was tightly associated with JNK activation. Inhibition of JNK abolished the elevation of Cx43. Further analysis revealed that the changes of JNK and Cx43 were controlled by GSH. Supplement of a membrane-permeable GSH analogue GSH ethyl ester or GSH precursor N-acetyl-cystein abrogated the effects of Cd(2+ on JNK activation and Cx43 expression. Indeed, Cd(2+ induced extracellular release of GSH. Blockade of Cx43 hemichannels with heptanol or Cx43 mimetic peptide Gap26 to prevent the efflux of GSH significantly attenuated the Cx43-elevating effects of Cd(2+. CONCLUSIONS: Collectively, our results thus indicate that Cd(2+-induced upregulation of Cx43 is through activation of nonjunctional Cx43 hemichannels. Our findings thus support the existence of a hemichannel-mediated self-regulation of Cx43 and provide novel insights into the molecular mechanisms of Cx43 expression and function.

  13. Connexin-47 and connexin-32 in gap junctions of oligodendrocyte somata, myelin sheaths, paranodal loops and Schmidt-Lanterman incisures: implications for ionic homeostasis and potassium siphoning.

    Science.gov (United States)

    Kamasawa, N; Sik, A; Morita, M; Yasumura, T; Davidson, K G V; Nagy, J I; Rash, J E

    2005-01-01

    The subcellular distributions and co-associations of the gap junction-forming proteins connexin 47 and connexin 32 were investigated in oligodendrocytes of adult mouse and rat CNS. By confocal immunofluorescence light microscopy, abundant connexin 47 was co-localized with astrocytic connexin 43 on oligodendrocyte somata, and along myelinated fibers, whereas connexin 32 without connexin 47 was co-localized with contactin-associated protein (caspr) in paranodes. By thin-section transmission electron microscopy, connexin 47 immunolabeling was on the oligodendrocyte side of gap junctions between oligodendrocyte somata and astrocytes. By freeze-fracture replica immunogold labeling, large gap junctions between oligodendrocyte somata and astrocyte processes contained much more connexin 47 than connexin 32. Along surfaces of internodal myelin, connexin 47 was several times as abundant as connexin 32, and in the smallest gap junctions, often occurred without connexin 32. In contrast, connexin 32 was localized without connexin 47 in newly-described autologous gap junctions in Schmidt-Lanterman incisures and between paranodal loops bordering nodes of Ranvier. Thus, connexin 47 in adult rodent CNS is the most abundant connexin in most heterologous oligodendrocyte-to-astrocyte gap junctions, whereas connexin 32 is the predominant if not sole connexin in autologous ("reflexive") oligodendrocyte gap junctions. These results clarify the locations and connexin compositions of heterologous and autologous oligodendrocyte gap junctions, identify autologous gap junctions at paranodes as potential sites for modulating paranodal electrical properties, and reveal connexin 47-containing and connexin 32-containing gap junctions as conduits for long-distance intracellular and intercellular movement of ions and associated osmotic water. The autologous gap junctions may regulate paranodal electrical properties during saltatory conduction. Acting in series and in parallel, autologous and

  14. Functional assessment of gap junctions in monolayer and three-dimensional cultures of human tendon cells using fluorescence recovery after photobleaching

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    Kuzma-Kuzniarska, Maria; Yapp, Clarence; Pearson-Jones, Thomas W.; Jones, Andrew K.; Hulley, Philippa A.

    2014-01-01

    Abstract. Gap junction-mediated intercellular communication influences a variety of cellular activities. In tendons, gap junctions modulate collagen production, are involved in strain-induced cell death, and are involved in the response to mechanical stimulation. The aim of the present study was to investigate gap junction-mediated intercellular communication in healthy human tendon-derived cells using fluorescence recovery after photobleaching (FRAP). The FRAP is a noninvasive technique that allows quantitative measurement of gap junction function in living cells. It is based on diffusion-dependent redistribution of a gap junction-permeable fluorescent dye. Using FRAP, we showed that human tenocytes form functional gap junctions in monolayer and three-dimensional (3-D) collagen I culture. Fluorescently labeled tenocytes following photobleaching rapidly reacquired the fluorescent dye from neighboring cells, while HeLa cells, which do not communicate by gap junctions, remained bleached. Furthermore, both 18 β-glycyrrhetinic acid and carbenoxolone, standard inhibitors of gap junction activity, impaired fluorescence recovery in tendon cells. In both monolayer and 3-D cultures, intercellular communication in isolated cells was significantly decreased when compared with cells forming many cell-to-cell contacts. In this study, we used FRAP as a tool to quantify and experimentally manipulate the function of gap junctions in human tenocytes in both two-dimensional (2-D) and 3-D cultures. PMID:24390370

  15. A Gap Junction Protein, Inx2, Modulates Calcium Flux to Specify Border Cell Fate during Drosophila oogenesis

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    Ghosh, Ritabrata; Deshpande, Girish

    2017-01-01

    Intercellular communication mediated by gap junction (GJ) proteins is indispensable during embryogenesis, tissue regeneration and wound healing. Here we report functional analysis of a gap junction protein, Innexin 2 (Inx2), in cell type specification during Drosophila oogenesis. Our data reveal a novel involvement of Inx2 in the specification of Border Cells (BCs), a migratory cell type, whose identity is determined by the cell autonomous STAT activity. We show that Inx2 influences BC fate specification by modulating STAT activity via Domeless receptor endocytosis. Furthermore, detailed experimental analysis has uncovered that Inx2 also regulates a calcium flux that transmits across the follicle cells. We propose that Inx2 mediated calcium flux in the follicle cells stimulates endocytosis by altering Dynamin (Shibire) distribution which is in turn critical for careful calibration of STAT activation and, thus for BC specification. Together our data provide unprecedented molecular insights into how gap junction proteins can regulate cell-type specification. PMID:28114410

  16. Gap junction and hemichannel-independent actions of connexins on cell and tissue functions--an update.

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    Zhou, Jade Z; Jiang, Jean X

    2014-04-17

    Connexins, a family of transmembrane proteins, are components of both gap junction channels and hemichannels, which mediate the exchange of ions and small molecules between adjacent cells, and between the inside and outside of the cell, respectively. Substantial advancements have been made in the comprehension of the role of gap junctions and hemichannels in coordinating cellular events. In recent years, a plethora of studies demonstrate a role of connexin proteins in the regulation of tissue homeostasis that occurs independently of their channel activities. This is shown in the context of cell growth, adhesion, migration, apoptosis, and signaling. The major mechanisms of these channel-independent activities still remain to be discovered. In this review, we provide an updated overview on the current knowledge of gap junction- and hemichannel-independent functions of connexins, in particular, their effects on tumorigenesis, neurogenesis and disease development. Copyright © 2014 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

  17. Vitamin D3 regulates the formation and degradation of gap junctions in androgen-responsive human prostate cancer cells.

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    Linda Kelsey

    Full Text Available 1α-25(OH2 vitamin D3 (1-25D, an active hormonal form of Vitamin D3, is a well-known chemopreventive and pro-differentiating agent. It has been shown to inhibit the growth of several prostate cancer cell lines. Gap junctions, formed of proteins called connexins (Cx, are ensembles of cell-cell channels, which permit the exchange of small growth regulatory molecules between adjoining cells. Cell-cell communication mediated by gap junctional channels is an important homeostatic control mechanism for regulating cell growth and differentiation. We have investigated the effect of 1-25D on the formation and degradation of gap junctions in an androgen-responsive prostate cancer cell line, LNCaP, which expresses retrovirally-introduced Cx32. Connexin32 is expressed by the luminal and well-differentiated cells of normal prostate and prostate tumors. Our results document that 1-25D enhances the expression of Cx32 and its subsequent assembly into gap junctions. Our results further show that 1-25D prevents androgen-regulated degradation of Cx32, post-translationally, independent of androgen receptor (AR-mediated signaling. Finally, our findings document that formation of gap junctions sensitizes Cx32-expressing LNCaP cells to the growth inhibitory effects of 1-25D and alters their morphology. These findings suggest that the growth-inhibitory effects of 1-25D in LNCaP cells may be related to its ability to modulate the assembly of Cx32 into gap junctions.

  18. Involvement of connexin 43 in acupuncture analgesia

    Institute of Scientific and Technical Information of China (English)

    HUANG Guang-ying; ZHENG Cui-hong; YU Wei-chang; TIAN Dai-shi; WANG Wei

    2009-01-01

    Background Connexin 43 (Cx43) is one of the major components of human keratinocyte gap junctions. To study whether gap junctional intercellular communication participates in the transfer of acupoint signals and acupuncture analgesia, the expression of Cx43 was studied in Zusanli (ST36) acupoints compared with control non-acupoint regions in rats after acupuncture. In addition, Cx43 heterozygous gene knockout mice were used to further explore the relationship between Cx43 and acupuncture analgesia. Methods The expression of Cx43 was detected by immunohistochemistry, immunoblotting, and RT-PCR for the Cx43 protein and mRNA. The influence of the Cx43 gene knockout on acupuncture analgesia was measured by a hot plate and observing the writhing response on Cx43 heterozygous gene knockout mice. Results Immunohistochemistry showed abundant Cx43 expression in some cells in the skin and subcutaneous tissue of rat ST36 acupoints. The mRNA and protein levels of Cx43 in acupoints were significantly higher than those in the control points in the non-acupuncture group, and even more so after acupuncture. The hot plate and writhing response experiments showed that partial knockout of the Cx43 gene decreased acupuncture analgesia. Conclusion Cx43 expression and acupuncture analgesia showed a positive correlation.

  19. Stochastic Model of Gap Junctions Exhibiting Rectification and Multiple Closed States of Slow Gates.

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    Snipas, Mindaugas; Kraujalis, Tadas; Paulauskas, Nerijus; Maciunas, Kestutis; Bukauskas, Feliksas F

    2016-03-29

    Gap-junction (GJ) channels formed from connexin (Cx) proteins provide direct pathways for electrical and metabolic cell-cell communication. Earlier, we developed a stochastic 16-state model (S16SM) of voltage gating of the GJ channel containing two pairs of fast and slow gates, each operating between open (o) and closed (c) states. However, experimental data suggest that gates may in fact contain two or more closed states. We developed a model in which the slow gate operates according to a linear reaction scheme, o↔c1↔c2, where c1 and c2 are initial-closed and deep-closed states that both close the channel fully, whereas the fast gate operates between the open state and the closed state and exhibits a residual conductance. Thus, we developed a stochastic 36-state model (S36SM) of GJ channel gating that is sensitive to transjunctional voltage (Vj). To accelerate simulation and eliminate noise in simulated junctional conductance (gj) records, we transformed an S36SM into a Markov chain 36-state model (MC36SM) of GJ channel gating. This model provides an explanation for well-established experimental data, such as delayed gj recovery after Vj gating, hysteresis of gj-Vj dependence, and the low ratio of functional channels to the total number of GJ channels clustered in junctional plaques, and it has the potential to describe chemically mediated gating, which cannot be reflected using an S16SM. The MC36SM, when combined with global optimization algorithms, can be used for automated estimation of gating parameters including probabilities of c1↔c2 transitions from experimental gj-time and gj-Vj dependencies.

  20. Managing the complexity of communication: regulation of gap junctions by post-translational modification

    DEFF Research Database (Denmark)

    Axelsen, Lene Nygaard; Callø, Kirstine; von Holstein-Rathlou, Niels-Henrik

    2013-01-01

    expression by transcription and translation is of great importance, the trafficking, channel activity and degradation are also under tight control. The function of connexins can be regulated by several post translational modifications, which affect numerous parameters; including number of channels, open......Gap junctions are comprised of connexins that form cell-to-cell channels which couple neighboring cells to accommodate the exchange of information. The need for communication does, however, change over time and therefore must be tightly controlled. Although the regulation of connexin protein...... probability, single channel conductance or selectivity. The most extensively investigated post translational modifications are phosphorylations, which have been documented in all mammalian connexins. Besides phosphorylations, some connexins are known to be ubiquitinated, SUMOylated, nitrosylated, hydroxylated...

  1. The NA+/K+-ATPase controls gap junctions via membrane microdomain interactions in rat smooth muscles.

    DEFF Research Database (Denmark)

    Matchkov, Vladimir; Nilsson, Holger; Aalkjær, Christian

    in regulation of the intercellular communication. We have here shown that gap junctions between SMCs are regulated through an interaction between the Na+/K+-ATPase and the Na+/Ca2+-exchanger leading to an increase in [Ca2+]i in discrete areas near the plasma membrane. We have also suggested that this Na......The Na+/K+-ATPase is known to interact with many membrane and cytosolic proteins by organizing various signaling complexes. These interactions were suggested to be important in regulation of various cellular responses. Pumping activity of the Na+/K+-ATPase is suggested to be essential for some...... in rat mesenteric small arteries. Paired cultured rat smooth muscle cells (A7r5) were used as a model for electrical coupling of SMC by measuring membrane capacitance (Cm). PCR, Western blotting and immunohistochemistry were used to identify the membrane transporters. SMCs were uncoupled (evaluated...

  2. Conduction abnormalities and ventricular arrhythmogenesis: The roles of sodium channels and gap junctions

    Directory of Open Access Journals (Sweden)

    Gary Tse

    2015-12-01

    Full Text Available Ventricular arrhythmias arise from disruptions in the normal orderly sequence of electrical activation and recovery of the heart. They can be categorized into disorders affecting predominantly cellular depolarization or repolarization, or those involving action potential (AP conduction. This article briefly discusses the factors causing conduction abnormalities in the form of unidirectional conduction block and reduced conduction velocity (CV. It then examines the roles that sodium channels and gap junctions play in AP conduction. Finally, it synthesizes experimental results to illustrate molecular mechanisms of how abnormalities in these proteins contribute to such conduction abnormalities and hence ventricular arrhythmogenesis, in acquired pathologies such as acute ischaemia and heart failure, as well as inherited arrhythmic syndromes.

  3. Interrupting the inflammatory cycle in chronic diseases--do gap junctions provide the answer?

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    Green, Colin R; Nicholson, Louise F B

    2008-12-01

    A number of chronic diseases, including neurodegenerative, cardiovascular and metabolic disorders, are associated with genetic susceptibility. Some may originate on exposure to an environmental stimulus. Regardless of genetic predisposition or external stimulus, these chronic diseases, once triggered, share an inflammatory component making them effectively persistent "wounds". There is also increasing evidence that the presence of one disease can cause activation of another apparently unrelated disease, leading to multiple disorders via activation of an immune response that 'fast forwards' disease progression. Here we review common aspects of a number of chronic disease conditions, and put forward the proposal that gap junction modulation may provide an opportunity to break the inflammatory cycle that sustains and links these disorders.

  4. Targeting neuronal gap junctions in mouse retina offers neuroprotection in glaucoma.

    Science.gov (United States)

    Akopian, Abram; Kumar, Sandeep; Ramakrishnan, Hariharasubramanian; Roy, Kaushambi; Viswanathan, Suresh; Bloomfield, Stewart A

    2017-06-30

    The progressive death of retinal ganglion cells and resulting visual deficits are hallmarks of glaucoma, but the underlying mechanisms remain unclear. In many neurodegenerative diseases, cell death induced by primary insult is followed by a wave of secondary loss. Gap junctions (GJs), intercellular channels composed of subunit connexins, can play a major role in secondary cell death by forming conduits through which toxic molecules from dying cells pass to and injure coupled neighbors. Here we have shown that pharmacological blockade of GJs or genetic ablation of connexin 36 (Cx36) subunits, which are highly expressed by retinal neurons, markedly reduced loss of neurons and optic nerve axons in a mouse model of glaucoma. Further, functional parameters that are negatively affected in glaucoma, including the electroretinogram, visual evoked potential, visual spatial acuity, and contrast sensitivity, were maintained at control levels when Cx36 was ablated. Neuronal GJs may thus represent potential therapeutic targets to prevent the progressive neurodegeneration and visual impairment associated with glaucoma.

  5. Effects of extremely low frequency magnetic fields on gap junctional intercellular communication and its mechanism

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    The study on biological effect of electromagnetic fields has been paid close attention in recent years. Gap junctional intercellular communication (GJIC) plays an important role in the maintenance of cell proliferation and differentiation, and in the multistage process of carcinogenesis. A series of researches showed that xtremely low frequency (ELF) magnetic fields not only enhance the inhibition of GJIC induced by 12-O-tetradecanoylphorbol-13-acetate, but also inhibit GJIC directly when the intensity is equal to or more than 0.4  mT, and that the mechanisms of GJIC inhibition by ELF magnetic fields are due to hyperphosphorylation of connexin 43, which is mediated by protein kinase C-activated signal transduction, and the internalization of connexin 43 from plasma membrane to cytoplasm.

  6. Gap junction proteins: master regulators of the planarian stem cell response to tissue maintenance and injury.

    Science.gov (United States)

    Peiris, T Harshani; Oviedo, Néstor J

    2013-01-01

    Gap junction (GJ) proteins are crucial mediators of cell-cell communication during embryogenesis, tissue regeneration and disease. GJ proteins form plasma membrane channels that facilitate passage of small molecules across cells and modulate signaling pathways and cellular behavior in different tissues. These properties have been conserved throughout evolution, and in most invertebrates GJ proteins are known as innexins. Despite their critical relevance for physiology and disease, the mechanisms by which GJ proteins modulate cell behavior are poorly understood. This review summarizes findings from recent work that uses planarian flatworms as a paradigm to analyze GJ proteins in the complexity of the whole organism. The planarian model allows access to a large pool of adult somatic stem cells (known as neoblasts) that support physiological cell turnover and tissue regeneration. Innexin proteins are present in planarians and play a fundamental role in controlling neoblast behavior. We discuss the possibility that GJ proteins participate as cellular sensors that inform neoblasts about local and systemic physiological demands. We believe that functional analyses of GJ proteins will bring a complementary perspective to studies that focus on the temporal expression of genes. Finally, integrating functional studies along with molecular genetics and epigenetic approaches would expand our understanding of cellular regulation in vivo and greatly enhance the possibilities for rationally modulating stem cell behavior in their natural environment. This article is part of a Special Issue entitled: The communicating junctions, roles and dysfunctions. Copyright © 2012 Elsevier B.V. All rights reserved.

  7. Influence of drugs on gap junctions in glioma cell lines and primary astrocytes in vitro

    Directory of Open Access Journals (Sweden)

    Zahra eMoinfar

    2014-05-01

    Full Text Available Gap junctions (GJs are hemichannels on cell membrane. Once they are intercellulary connected to the neighboring cells, they build a functional syncytium which allows rapid transfer of ions and molecules between cells. This characteristic makes GJs a potential modulator in proliferation, migration and development of the cells. So far, several types of GJs are recognized on different brain cells as well as in glioma. Astrocytes, as one of the major cells that maintain neuronal homeostasis, express different types of GJs that let them communicate with neurons, oligodendrocytes and endothelial cells of the blood brain barrier; however, the main GJ in astrocytes is connexin 43. There are different cerebral diseases in which astrocyte GJs might play a role. Several drugs have been reported to modulate gap junctional communication in the brain which can consequently have beneficial or detrimental effects on the course of treatment in certain diseases. However, the exact cellular mechanism behind those pharmaceutical efficacies on GJs is not well-understood. Accordingly, how specific drugs would affect GJs and what some consequent specific brain diseases would be are the interests of the authors of this chapter. We would focus on pharmaceutical effects on GJs on astrocytes in specific diseases where GJs could possibly play a role including: 1 migraine and a novel therapy for migraine with aura, 2 neuroautoimmune diseases and immunomodulatory drugs in the treatment of demyelinating diseases of the central nervous system such as multiple sclerosis, 3 glioma and antineoplastic and anti-inflammatory agents that are used in treating brain tumors and 4 epilepsy and anticonvulsants that are widely used for seizures therapy. All of the above-mentioned therapeutic categories can possibly affect GJs expression of astrocytes and the role is discussed in the upcoming chapter.

  8. Intercellular signaling via cyclic GMP diffusion through gap junctions restarts meiosis in mouse ovarian follicles.

    Science.gov (United States)

    Shuhaibar, Leia C; Egbert, Jeremy R; Norris, Rachael P; Lampe, Paul D; Nikolaev, Viacheslav O; Thunemann, Martin; Wen, Lai; Feil, Robert; Jaffe, Laurinda A

    2015-04-28

    Meiosis in mammalian oocytes is paused until luteinizing hormone (LH) activates receptors in the mural granulosa cells of the ovarian follicle. Prior work has established the central role of cyclic GMP (cGMP) from the granulosa cells in maintaining meiotic arrest, but it is not clear how binding of LH to receptors that are located up to 10 cell layers away from the oocyte lowers oocyte cGMP and restarts meiosis. Here, by visualizing intercellular trafficking of cGMP in real-time in live follicles from mice expressing a FRET sensor, we show that diffusion of cGMP through gap junctions is responsible not only for maintaining meiotic arrest, but also for rapid transmission of the signal that reinitiates meiosis from the follicle surface to the oocyte. Before LH exposure, the cGMP concentration throughout the follicle is at a uniformly high level of ∼2-4 μM. Then, within 1 min of LH application, cGMP begins to decrease in the peripheral granulosa cells. As a consequence, cGMP from the oocyte diffuses into the sink provided by the large granulosa cell volume, such that by 20 min the cGMP concentration in the follicle is uniformly low, ∼100 nM. The decrease in cGMP in the oocyte relieves the inhibition of the meiotic cell cycle. This direct demonstration that a physiological signal initiated by a stimulus in one region of an intact tissue can travel across many layers of cells via cyclic nucleotide diffusion through gap junctions could provide a general mechanism for diverse cellular processes.

  9. Intercellular signaling via cyclic GMP diffusion through gap junctions restarts meiosis in mouse ovarian follicles

    Science.gov (United States)

    Shuhaibar, Leia C.; Egbert, Jeremy R.; Norris, Rachael P.; Lampe, Paul D.; Nikolaev, Viacheslav O.; Thunemann, Martin; Wen, Lai; Feil, Robert; Jaffe, Laurinda A.

    2015-01-01

    Meiosis in mammalian oocytes is paused until luteinizing hormone (LH) activates receptors in the mural granulosa cells of the ovarian follicle. Prior work has established the central role of cyclic GMP (cGMP) from the granulosa cells in maintaining meiotic arrest, but it is not clear how binding of LH to receptors that are located up to 10 cell layers away from the oocyte lowers oocyte cGMP and restarts meiosis. Here, by visualizing intercellular trafficking of cGMP in real-time in live follicles from mice expressing a FRET sensor, we show that diffusion of cGMP through gap junctions is responsible not only for maintaining meiotic arrest, but also for rapid transmission of the signal that reinitiates meiosis from the follicle surface to the oocyte. Before LH exposure, the cGMP concentration throughout the follicle is at a uniformly high level of ∼2–4 μM. Then, within 1 min of LH application, cGMP begins to decrease in the peripheral granulosa cells. As a consequence, cGMP from the oocyte diffuses into the sink provided by the large granulosa cell volume, such that by 20 min the cGMP concentration in the follicle is uniformly low, ∼100 nM. The decrease in cGMP in the oocyte relieves the inhibition of the meiotic cell cycle. This direct demonstration that a physiological signal initiated by a stimulus in one region of an intact tissue can travel across many layers of cells via cyclic nucleotide diffusion through gap junctions could provide a general mechanism for diverse cellular processes. PMID:25775542

  10. Caveolin-1 modulates cardiac gap junction homeostasis and arrhythmogenecity by regulating cSrc tyrosine kinase.

    Science.gov (United States)

    Yang, Kai-Chien; Rutledge, Cody A; Mao, Mao; Bakhshi, Farnaz R; Xie, An; Liu, Hong; Bonini, Marcelo G; Patel, Hemal H; Minshall, Richard D; Dudley, Samuel C

    2014-08-01

    Genome-wide association studies have revealed significant association of caveolin-1 (Cav1) gene variants with increased risk of cardiac arrhythmias. Nevertheless, the mechanism for this linkage is unclear. Using adult Cav1(-/-) mice, we revealed a marked reduction in the left ventricular conduction velocity in the absence of myocardial Cav1, which is accompanied with increased inducibility of ventricular arrhythmias. Further studies demonstrated that loss of Cav1 leads to the activation of cSrc tyrosine kinase, resulting in the downregulation of connexin 43 and subsequent electric abnormalities. Pharmacological inhibition of cSrc mitigates connexin 43 downregulation, slowed conduction, and arrhythmia inducibility in Cav1(-/-) animals. Using a transgenic mouse model with cardiac-specific overexpression of angiotensin-converting enzyme (ACE8/8), we demonstrated that, on enhanced cardiac renin-angiotensin system activity, Cav1 dissociated from cSrc because of increased Cav1 S-nitrosation at Cys(156), leading to cSrc activation, connexin 43 reduction, impaired gap junction function, and subsequent increase in the propensity for ventricular arrhythmias and sudden cardiac death. Renin-angiotensin system-induced Cav1 S-nitrosation was associated with increased Cav1-endothelial nitric oxide synthase binding in response to increased mitochondrial reactive oxidative species generation. The present studies reveal the critical role of Cav1 in modulating cSrc activation, gap junction remodeling, and ventricular arrhythmias. These data provide a mechanistic explanation for the observed genetic link between Cav1 and cardiac arrhythmias in humans and suggest that targeted regulation of Cav1 may reduce arrhythmic risk in cardiac diseases associated with renin-angiotensin system activation. © 2014 American Heart Association, Inc.

  11. Involvement of gap junction channels in the pathophysiology of migraine with aura

    Directory of Open Access Journals (Sweden)

    Denis eSarrouilhe

    2014-02-01

    Full Text Available Migraine is a common, recurrent and disabling primary headache disorder with a genetic component which affects up to 20 % of the population. One third of all patients with migraine experiences aura, a focal neurological disturbance that manifests itself as visual, sensitive or motor symptoms preceding the headache. In the pathophysiology of migraine with aura, activation of the trigeminovascular system from the meningeal vessels mediates migraine pain via the brainstem and projections ascend to the thalamus and cortex. Cortical spreading depression (CSD was proposed to trigger migraine aura and to activate perivascular trigeminal nerves in the cortex. Quinine, quinidine and the derivative mefloquine are able to inhibit CSD suggesting an involvement of neuronal connexin36 channels in CSD propagation. More recently, CSD was shown to induce headache by activating the trigeminovascular system through the opening of stressed neuronal Pannexin1 channels. A novel benzopyran compound, tonabersat, was selected for clinical trial on the basis of its inhibitory activity on CSD and neurogenic inflammation in animal models of migraine. Interestingly, in the time course of animal model trials, tonabersat was shown to inhibit trigeminal ganglion neuronal-glial cell gap junctions, suggesting that this compound could prevent peripheral sensitization within the ganglion. Three clinical trials aimed at investigating the effectiveness of tonabersat as a preventive drug were negative, and conflicting results were obtained in other trials concerning its ability to relieve attacks. In contrast, in another clinical trial, tonabersat showed a preventive effect on attacks of migraine with aura but had no efficacy on non-aura attacks. Gap junction channels seem to be involved in several ways in the pathophysiology of migraine with aura and emerge as a new promising putative target in treatment of this disorder.

  12. Involvement of gap junction channels in the pathophysiology of migraine with aura

    Science.gov (United States)

    Sarrouilhe, Denis; Dejean, Catherine; Mesnil, Marc

    2014-01-01

    Migraine is a common, recurrent, and disabling primary headache disorder with a genetic component which affects up to 20% of the population. One third of all patients with migraine experiences aura, a focal neurological disturbance that manifests itself as visual, sensitive or motor symptoms preceding the headache. In the pathophysiology of migraine with aura, activation of the trigeminovascular system from the meningeal vessels mediates migraine pain via the brainstem and projections ascend to the thalamus and cortex. Cortical spreading depression (CSD) was proposed to trigger migraine aura and to activate perivascular trigeminal nerves in the cortex. Quinine, quinidine and the derivative mefloquine are able to inhibit CSD suggesting an involvement of neuronal connexin36 channels in CSD propagation. More recently, CSD was shown to induce headache by activating the trigeminovascular system through the opening of stressed neuronal Pannexin1 channels. A novel benzopyran compound, tonabersat, was selected for clinical trial on the basis of its inhibitory activity on CSD and neurogenic inflammation in animal models of migraine. Interestingly, in the time course of animal model trials, tonabersat was shown to inhibit trigeminal ganglion (TGG) neuronal-glial cell gap junctions, suggesting that this compound could prevent peripheral sensitization within the ganglion. Three clinical trials aimed at investigating the effectiveness of tonabersat as a preventive drug were negative, and conflicting results were obtained in other trials concerning its ability to relieve attacks. In contrast, in another clinical trial, tonabersat showed a preventive effect on attacks of migraine with aura but had no efficacy on non-aura attacks. Gap junction channels seem to be involved in several ways in the pathophysiology of migraine with aura and emerge as a new promising putative target in treatment of this disorder. PMID:24611055

  13. Dysfunction of mitochondria and deformed gap junctions in the heart of IL-18-deficient mice.

    Science.gov (United States)

    Li, Wen; Jin, Denan; Hata, Masaki; Takai, Shinji; Yamanishi, Kyosuke; Shen, Weili; El-Darawish, Yosif; Yamanishi, Hiromichi; Okamura, Haruki

    2016-08-01

    Interleukin-18 (IL-18) was discovered as an interferon-γ-inducing factor and has been regarded as a proinflammatory cytokine. However, IL-18 is ubiquitously expressed both in immune/inflammatory cells and in nonimmune cells, and its biological roles have not been sufficiently elucidated. Here, we demonstrate that IL-18-deficient [IL-18 knockout (KO)] mice have heart abnormalities that may be related to impaired autophagy. In endurance running tests, IL-18KO mice ran significantly shorter distances compared with wild-type (WT) mice. Echocardiographs indicated disability in the systolic and diastolic functions of the IL-18KO mouse heart. Immunostaining of connexin 43 showed heterogeneous localization of gap junctions in the lateral membranes of the IL-18KO cardiac myocytes. Western blotting analysis revealed decreased phosphorylated connexin 43 in the IL-18KO heart. Electron microscopy revealed unusual localization of intercalated disks, swollen or damaged mitochondria, and broad, indistinct Z-lines in the IL-18KO heart. In accordance with the morphological observation, mitochondrial respiratory function, including that of complexes I and IV, was impaired, and production of reactive oxygen species was augmented in IL-18KO hearts. Notably, levels of LC3-II were markedly lower in the IL-18KO hearts than in WT hearts. In the culture of cardiac myocytes of IL-18KO neonates, exogenous IL-18 upregulated LC3-II and increased the number of intact mitochondria with high mitochondrial membrane potential. These results indicated that IL-18 has roles apart from those as a proinflammatory cytokine in cardiac myocytes and suggested that IL-18 contributes to the homeostatic maintenance of mitochondrial function and gap-junction turnover in cardiac myocytes, possibly by upregulating autophagy.

  14. Connexin domains relevant to the chemical gating of gap junction channels

    Directory of Open Access Journals (Sweden)

    C. Peracchia

    1997-05-01

    Full Text Available Most cells exchange ions and small metabolites via gap junction channels. These channels are made of two hemichannels (connexons, each formed by the radial arrangement of six connexin (Cx proteins. Connexins span the bilayer four times (M1-M4 and have both amino- and carboxy-termini (NT, CT at the cytoplasmic side of the membrane, forming two extracellular loops (E1, E2 and one inner (IL loop. The channels are regulated by gates that close with cytosolic acidification (e.g., CO2 treatment or increased calcium concentration, possibly via calmodulin activation. Although gap junction regulation is still unclear, connexin domains involved in gating are being defined. We have recently focused on the CO2 gating sensitivity of Cx32, Cx38 and various mutants and chimeras expressed in Xenopus oocytes and studied by double voltage clamp. Cx32 is weakly sensitive to CO2, whereas Cx38 is highly sensitive. A Cx32 chimera containing the second half of the inner loop (IL2 of Cx38 was as sensitive to CO2 as Cx38, indicating that this domain plays an important role. Deletion of CT by 84% did not affect CO2 sensitivity, but replacement of 5 arginines (R with sparagines (N at the beginning of CT (C1 greatly enhanced the CO2 sensitivity of Cx32. This suggests that whereas most of CT is irrelevant, positive charges of C1 maintain the CO2 sensitivity of Cx32 low. As a hypothesis we have proposed a model that involves charge interaction between negative residues of the beginning of IL1 and positive residues of either C1 or IL2. Open and closed channels would result from IL1-C1 and IL1-IL2 interactions, respectively

  15. First-principles spin-transfer torque in CuMnAs |GaP |CuMnAs junctions

    Science.gov (United States)

    Stamenova, Maria; Mohebbi, Razie; Seyed-Yazdi, Jamileh; Rungger, Ivan; Sanvito, Stefano

    2017-02-01

    We demonstrate that an all-antiferromagnetic tunnel junction with current perpendicular to the plane geometry can be used as an efficient spintronic device with potential high-frequency operation. By using state-of-the-art density functional theory combined with quantum transport, we show that the Néel vector of the electrodes can be manipulated by spin-transfer torque. This is staggered over the two different magnetic sublattices and can generate dynamics and switching. At the same time the different magnetization states of the junction can be read by standard tunneling magnetoresistance. Calculations are performed for CuMnAs |GaP |CuMnAs junctions with different surface terminations between the antiferromagnetic CuMnAs electrodes and the insulating GaP spacer. We find that the torque remains staggered regardless of the termination, while the magnetoresistance depends on the microscopic details of the interface.

  16. Relative Roles of Gap Junction Channels and Cytoplasm in Cell-to-Cell Diffusion of Fluorescent Tracers

    Science.gov (United States)

    Safranyos, Richard G. A.; Caveney, Stanley; Miller, James G.; Petersen, Nils O.

    1987-04-01

    Intercellular (tissue) diffusion of molecules requires cytoplasmic diffusion and diffusion through gap junctional (or cell-to-cell) channels. The rates of tissue and cytoplasmic diffusion of fluorescent tracers, expressed as an effective diffusion coefficient, De, and a cytoplasmic diffusion coefficient, Dcyt, have been measured among the developing epidermal cells of a larval beetle, Tenebrio molitor L., to determine the contribution of the junctional channels to intercellular diffusion. Tracer diffusion was measured by injecting fluorescent tracers into cells and quantitating the rate of subsequent spread into adjacent cells. Cytoplasmic diffusion was determined by fluorescence photobleaching. These experiments show that gap junctional channels constitute approximately 70-80% of the total cell-to-cell resistance to the diffusion of organic tracers at high concentrations in this tissue. At low concentrations, however, the binding of tracer to cytoplasm slows down the cytoplasmic diffusion, which may limit intercellular diffusion.

  17. Gap-junction coupling and ATP-sensitive potassium channels in human β -cell clusters: Effects on emergent dynamics

    Science.gov (United States)

    Loppini, A.; Pedersen, M. G.; Braun, M.; Filippi, S.

    2017-09-01

    The importance of gap-junction coupling between β cells in pancreatic islets is well established in mouse. Such ultrastructural connections synchronize cellular activity, confine biological heterogeneity, and enhance insulin pulsatility. Dysfunction of coupling has been associated with diabetes and altered β -cell function. However, the role of gap junctions between human β cells is still largely unexplored. By using patch-clamp recordings of β cells from human donors, we previously estimated electrical properties of these channels by mathematical modeling of pairs of human β cells. In this work we revise our estimate by modeling triplet configurations and larger heterogeneous clusters. We find that a coupling conductance in the range 0.005 -0.020 nS/pF can reproduce experiments in almost all the simulated arrangements. We finally explore the consequence of gap-junction coupling of this magnitude between β cells with mutant variants of the ATP-sensitive potassium channels involved in some metabolic disorders and diabetic conditions, translating studies performed on rodents to the human case. Our results are finally discussed from the perspective of therapeutic strategies. In summary, modeling of more realistic clusters with more than two β cells slightly lowers our previous estimate of gap-junction conductance and gives rise to patterns that more closely resemble experimental traces.

  18. Effect of Mefloquine, a Gap Junction Blocker, on Circadian Period2 Gene Oscillation in the Mouse Suprachiasmatic Nucleus

    Directory of Open Access Journals (Sweden)

    Jinmi Koo

    2015-09-01

    Full Text Available BackgroundIn mammals, the master circadian pacemaker is localized in an area of the ventral hypothalamus known as the suprachiasmatic nucleus (SCN. Previous studies have shown that pacemaker neurons in the SCN are highly coupled to one another, and this coupling is crucial for intrinsic self-sustainability of the SCN central clock, which is distinguished from peripheral oscillators. One plausible mechanism underlying the intercellular communication may involve direct electrical connections mediated by gap junctions.MethodsWe examined the effect of mefloquine, a neuronal gap junction blocker, on circadian Period 2 (Per2 gene oscillation in SCN slice cultures prepared from Per2::luciferase (PER2::LUC knock-in mice using a real-time bioluminescence measurement system.ResultsAdministration of mefloquine causes instability in the pulse period and a slight reduction of amplitude in cyclic PER2::LUC expression. Blockade of gap junctions uncouples PER2::LUC-expressing cells, in terms of phase transition, which weakens synchrony among individual cellular rhythms.ConclusionThese findings suggest that neuronal gap junctions play an important role in synchronizing the central pacemaker neurons and contribute to the distinct self-sustainability of the SCN master clock.

  19. A variant in the carboxyl-terminus of connexin 40 alters GAP junctions and increases risk for tetralogy of Fallot

    NARCIS (Netherlands)

    Guida, V.; Ferese, R.; Rocchetti, M.; Bonetti, M.; Sarkozy, A.; Cecchetti, S.; Gelmetti, V.; Lepri, F.; Copetti, M.; Lamorte, G.; Cristina Digilio, M.; Marino, B.; Zaza, A.; den Hertog, J.; Dallapiccola, B.; De Luca, A.

    2013-01-01

    GJA5 gene (MIM no. 121013), localized at 1q21.1, encodes for the cardiac gap junction protein connexin 40. In humans, copy number variants of chromosome 1q21.1 have been associated with variable phenotypes comprising congenital heart disease (CHD), including isolated TOF. In mice, the deletion of Gj

  20. Connexin composition in apposed gap junction hemiplaques revealed by matched double-replica freeze-fracture replica immunogold labeling.

    Science.gov (United States)

    Rash, John E; Kamasawa, Naomi; Davidson, Kimberly G V; Yasumura, Thomas; Pereda, Alberto E; Nagy, James I

    2012-06-01

    Despite the combination of light-microscopic immunocytochemistry, histochemical mRNA detection techniques and protein reporter systems, progress in identifying the protein composition of neuronal versus glial gap junctions, determination of the differential localization of their constituent connexin proteins in two apposing membranes and understanding human neurological diseases caused by connexin mutations has been problematic due to ambiguities introduced in the cellular and subcellular assignment of connexins. Misassignments occurred primarily because membranes and their constituent proteins are below the limit of resolution of light microscopic imaging techniques. Currently, only serial thin-section transmission electron microscopy and freeze-fracture replica immunogold labeling have sufficient resolution to assign connexin proteins to either or both sides of gap junction plaques. However, freeze-fracture replica immunogold labeling has been limited because conventional freeze fracturing allows retrieval of only one of the two membrane fracture faces within a gap junction, making it difficult to identify connexin coupling partners in hemiplaques removed by fracturing. We now summarize progress in ascertaining the connexin composition of two coupled hemiplaques using matched double-replicas that are labeled simultaneously for multiple connexins. This approach allows unambiguous identification of connexins and determination of the membrane "sidedness" and the identities of connexin coupling partners in homotypic and heterotypic gap junctions of vertebrate neurons.

  1. Analysis of the Gap Junction-dependent Transfer of miRNA with 3D-FRAP Microscopy.

    Science.gov (United States)

    Lemcke, Heiko; Voronina, Natalia; Steinhoff, Gustav; David, Robert

    2017-06-19

    Small antisense RNAs, like miRNA and siRNA, play an important role in cellular physiology and pathology and, moreover, can be used as therapeutic agents in the treatment of several diseases. The development of new, innovative strategies for miRNA/siRNA therapy is based on an extensive knowledge of the underlying mechanisms. Recent data suggest that small RNAs are exchanged between cells in a gap junction-dependent manner, thereby inducing gene regulatory effects in the recipient cell. Molecular biological techniques and flow cytometric analysis are commonly used to study the intercellular exchange of miRNA. However, these methods do not provide high temporal resolution, which is necessary when studying the gap junctional flux of molecules. Therefore, to investigate the impact of miRNA/siRNA as intercellular signaling molecules, novel tools are needed that will allow for the analysis of these small RNAs at the cellular level. The present protocol describes the application of three-dimensional fluorescence recovery after photobleaching (3D-FRAP) microscopy to elucidating the gap junction-dependent exchange of miRNA molecules between cardiac cells. Importantly, this straightforward and non-invasive live-cell imaging approach allows for the visualization and quantification of the gap junctional shuttling of fluorescently labeled small RNAs in real time, with high spatio-temporal resolution. The data obtained by 3D-FRAP confirm a novel pathway of intercellular gene regulation, where small RNAs act as signaling molecules within the intercellular network.

  2. Dynamic trafficking and delivery of connexons to the plasma membrane and accretion to gap junctions in living cells

    NARCIS (Netherlands)

    Lauf, Undine; Giepmans, Ben N G; Lopez, Patricia; Braconnot, Sebastien; Chen, Shu-Chih; Falk, Matthias M

    2002-01-01

    Certain membrane channels including acetylcholine receptors, gap junction (GJ) channels, and aquaporins arrange into large clusters in the plasma membrane (PM). However, how these channels are recruited to the clusters is unknown. To address this question, we have investigated delivery of GJ channel

  3. Down-regulation of membrana granulosa cell gap junctions is correlated with irreversible commitment to resume meiosis in golden Syrian hamster oocytes.

    Science.gov (United States)

    Racowsky, C; Baldwin, K V; Larabell, C A; DeMarais, A A; Kazilek, C J

    1989-08-01

    One of the currently popular hypotheses for the regulation of meiotic resumption in mammalian oocytes proposes that the preovulatory surge of luteinizing hormone causes down-regulation of follicular gap junctions, which in turn disrupts transfer of a meiotic arrester from the somatic cells into the oocyte. The present study has investigated this hypothesis by examining the integrity of membrana granulosa cell gap junctions during the period of irreversible commitment to maturation of golden Syrian hamster oocytes in vivo. Our results have revealed a significant progressive decrease in the fractional area of cell surface occupied by gap junction membrane with increasing percentage of oocytes irreversibly committed to mature (1.946% and 0.921% fractional gap junction area at 0% and 100% oocytes irreversibly committed to mature, respectively, P less than 0.05). This net loss of membrana granulosa cell gap junctions from the cell surface was accompanied by a significant decrease in density of gap junction particles, whether they were arranged in rectilinear or non-rectilinear packing patterns. Furthermore, the number of gap junction particles per unit area of surface membrane scanned also underwent a significant progressive decrease with increasing percentage of oocytes irreversibly committed to mature. These data with the hamster are consistent with the hypothesis that down-regulation of membrana granulosa cell gap junctions may be of central importance in the regulation of gonadotropic stimulation of meiotic resumption in mammalian oocytes.

  4. Gap junctional communication modulates gene transcription by altering the recruitment of Sp1 and Sp3 to connexin-response elements in osteoblast promoters

    Science.gov (United States)

    Stains, Joseph P.; Lecanda, Fernando; Screen, Joanne; Towler, Dwight A.; Civitelli, Roberto

    2003-01-01

    Loss-of-function mutations of gap junction proteins, connexins, represent a mechanism of disease in a variety of tissues. We have shown that recessive (gene deletion) or dominant (connexin45 overexpression) disruption of connexin43 function results in osteoblast dysfunction and abnormal expression of osteoblast genes, including down-regulation of osteocalcin transcription. To elucidate the molecular mechanisms of gap junction-sensitive transcriptional regulation, we systematically analyzed the rat osteocalcin promoter for sensitivity to gap junctional intercellular communication. We identified an Sp1/Sp3 containing complex that assembles on a minimal element in the -70 to -57 region of the osteocalcin promoter in a gap junction-dependent manner. This CT-rich connexin-response element is necessary and sufficient to confer gap junction sensitivity to the osteocalcin proximal promoter. Repression of osteocalcin transcription occurs as a result of displacement of the stimulatory Sp1 by the inhibitory Sp3 on the promoter when gap junctional communication is perturbed. Modulation of Sp1/Sp3 recruitment also occurs on the collagen Ialpha1 promoter and translates into gap junction-sensitive transcriptional control of collagen Ialpha1 gene expression. Thus, regulation of Sp1/Sp3 recruitment to the promoter may represent a potential general mechanism for transcriptional control of target genes by signals passing through gap junctions.

  5. Homotypic gap junctional communication associated with metastasis increases suppression increases with PKA kinase activity and is unaffected by P13K inhibition

    Science.gov (United States)

    Loss of gap junctional intercellular communication (GJIC) between cancer cells is a common characteristic of malignant transformation. This communication is mediated by connexin proteins that make up the functional units of gap junctions. Connexins are highly regulated at the protein level and phosp...

  6. The antiarrhythmic peptide analog rotigaptide (ZP123) stimulates gap junction intercellular communication in human osteoblasts and prevents decrease in femoral trabecular bone strength in ovariectomized rats

    DEFF Research Database (Denmark)

    Jørgensen, Niklas Rye; Teilmann, Stefan Cuoni; Henriksen, Zanne

    2005-01-01

    and strength in vivo. Cell coupling and calcium signaling were assessed in vitro on human, primary, osteoblastic cells. In vivo effects of rotigaptide on bone strength and density were determined 4 wk after ovariectomy in rats treated with either vehicle, sc injection twice daily (300 nmol per kilogram body......Gap junctions play an important role in bone development and function, but the lack of pharmacological tools has hampered the gap junction research. The antiarrhythmic peptides stimulate gap junction communication between cardiomyocytes, but effects in noncardiac tissue are unknown. The purpose...... of this study was to examine whether antiarrhythmic peptides, which are small peptides increasing gap junctional conductivity, show specific binding to osteoblasts and investigate the effect of the stable analog rotigaptide (ZP123) on gap junctional intercellular communication in vitro and on bone mass...

  7. The effect of a gap-junction blocker, carbenoxolone, on ischemic brain injury and cortical spreading depression.

    Science.gov (United States)

    Tamura, K; Alessandri, B; Heimann, A; Kempski, O

    2011-10-27

    Cortical spreading depression (CSD) has been shown to cause secondary cell loss in experimental models of brain injury and in patients, and blocking of CSD is a potential neuroprotective strategy. Here we tested the hypothesis that gap junctions affect CSD under physiological conditions as well as infarct development in a rat two-vein occlusion model suited to study pathophysiology of the penumbra (n = 71). We applied the gap junction blocker carbenoxolone (CBX) or saline intra-ventricularly. Interestingly, CBX temporarily increased systemic blood pressure and cortical blood flow (41% and 53%, 15 min after 250 μg CBX). We induced CSD with cortical microinjection of potassium chloride (KCl), counted how many spontaneous CSDs after CSD induction were elicited and measured the propagation velocity. After 250 μg CBX administration, significant 37.5 ± 6.5 additional CSDs were seen. CSD velocity increased significantly after 50 μg and 250 μg CBX. Occlusion of two adjacent cortical veins using Rose Bengal dye and fiberoptic illumination followed by 250 μg CBX or saline showed a significant more than doubling of infarct volumes 7 days after CBX. The current experiments provide evidence that CBX can accelerate the initiation and propagation of CSD suggesting opening of gap junctions is not required for CSD propagation. Blocking gap junctions worsens outcome from focal cerebral ischemia. Hence, measures intended to improve spatial buffering via astroglial gap junctions could have therapeutic potential in disease processes involving CSD. Copyright © 2011 IBRO. Published by Elsevier Ltd. All rights reserved.

  8. Protein preconcentration using nanofractures generated by nanoparticle-assisted electric breakdown at junction gaps.

    Directory of Open Access Journals (Sweden)

    Chun-Ping Jen

    Full Text Available Sample preconcentration is an important step that increases the accuracy of subsequent detection, especially for samples with extremely low concentrations. Due to the overlapping of electrical double layers in the nanofluidic channel, the concentration polarization effect can be generated by applying an electric field. Therefore, a nonlinear electrokinetic flow is induced, which results in the fast accumulation of proteins in front of the induced ionic depletion zone, the so-called exclusion-enrichment effect. Nanofractures were created in this work to preconcentrate proteins via the exclusion-enrichment effect. The protein sample was driven by electroosmotic flow and accumulated at a specific location. The preconcentration chip for proteins was fabricated using simple standard soft lithography with a polydimethylsiloxane replica. Nanofractures were formed by utilizing nanoparticle-assisted electric breakdown. The proposed method for nanofracture formation that utilizes nanoparticle deposition at the junction gap between microchannels greatly decreases the required electric breakdown voltage. The experimental results indicate that a protein sample with an extremely low concentration of 1 nM was concentrated to 1.5×10(4-fold in 60 min using the proposed chip.

  9. Connexin expression and gap-junctional intercellular communication in ES cells and iPS cells.

    Science.gov (United States)

    Oyamada, Masahito; Takebe, Kumiko; Endo, Aya; Hara, Sachiko; Oyamada, Yumiko

    2013-01-01

    Pluripotent stem cells, i.e., embryonic stem (ES) and induced pluripotent stem (iPS) cells, can indefinitely proliferate without commitment and differentiate into all cell lineages. ES cells are derived from the inner cell mass of the preimplantation blastocyst, whereas iPS cells are generated from somatic cells by overexpression of a few transcription factors. Many studies have demonstrated that mouse and human iPS cells are highly similar but not identical to their respective ES cell counterparts. The potential to generate basically any differentiated cell types from these cells offers the possibility to establish new models of mammalian development and to create new sources of cells for regenerative medicine. ES cells and iPS cells also provide useful models to study connexin expression and gap-junctional intercellular communication (GJIC) during cell differentiation and reprogramming. In 1996, we reported connexin expression and GJIC in mouse ES cells. Because a substantial number of papers on these subjects have been published since our report, this Mini Review summarizes currently available data on connexin expression and GJIC in ES cells and iPS cells during undifferentiated state, differentiation, and reprogramming.

  10. Connexin expression and gap-junctional intercellular communication in ES cells and iPS cells

    Directory of Open Access Journals (Sweden)

    Masahito eOyamada

    2013-07-01

    Full Text Available Pluripotent stem cells, i.e., embryonic stem (ES and induced pluripotent stem (iPS cells, can indefinitely proliferate without commitment and differentiate into all cell lineages. ES cells are derived from the inner cell mass of the preimplantation blastocyst, whereas iPS cells are generated from somatic cells by overexpression of a few transcription factors. Many studies have demonstrated that mouse and human iPS cells are highly similar but not identical to their respective ES cell counterparts. The potential to generate basically any differentiated cell types from these cells offers the possibility to establish new models of mammalian development and to create new sources of cells for regenerative medicine. ES cells and iPS cells also provide useful models to study connexin expression and gap-junctional intercellular communication (GJIC during cell differentiation and reprogramming. In 1996, we reported connexin expression and GJIC in mouse ES cells. Because a substantial number of papers on these subjects have been published since our report, this Mini Review summarizes currently available data on connexin expression and GJIC in ES cells and iPS cells during undifferentiated state, differentiation, and reprogramming.

  11. Formation of antiwaves in gap-junction-coupled chains of neurons

    Science.gov (United States)

    Urban, Alexander; Ermentrout, Bard

    2012-07-01

    Using network models consisting of gap-junction-coupled Wang-Buszaki neurons, we demonstrate that it is possible to obtain not only synchronous activity between neurons but also a variety of constant phase shifts between 0 and π. We call these phase shifts intermediate stable phase-locked states. These phase shifts can produce a large variety of wavelike activity patterns in one-dimensional chains and two-dimensional arrays of neurons, which can be studied by reducing the system of equations to a phase model. The 2π periodic coupling functions of these models are characterized by prominent higher order terms in their Fourier expansion, which can be varied by changing model parameters. We study how the relative contribution of the odd and even terms affects what solutions are possible, the basin of attraction of those solutions, and their stability. These models may be applicable to the spinal central pattern generators of the dogfish and also to the developing neocortex of the neonatal rat.

  12. Extracellular domains play different roles in gap junction formation and docking compatibility.

    Science.gov (United States)

    Bai, Donglin; Wang, Ao Hong

    2014-02-15

    GJ (gap junction) channels mediate direct intercellular communication and play an important role in many physiological processes. Six connexins oligomerize to form a hemichannel and two hemichannels dock together end-to-end to form a GJ channel. Connexin extracellular domains (E1 and E2) have been shown to be important for the docking, but the molecular mechanisms behind the docking and formation of GJ channels are not clear. Recent developments in atomic GJ structure and functional studies on a series of connexin mutants revealed that E1 and E2 are likely to play different roles in the docking. Non-covalent interactions at the docking interface, including hydrogen bonds, are predicted to form between interdocked extracellular domains. Protein sequence alignment analysis on the docking compatible/incompatible connexins indicate that the E1 domain is important for the formation of the GJ channel and the E2 domain is important in the docking compatibility in heterotypic channels. Interestingly, the hydrogen-bond forming or equivalent residues in both E1 and E2 domains are mutational hot spots for connexin-linked human diseases. Understanding the molecular mechanisms of GJ docking can assist us to develop novel strategies in rescuing the disease-linked connexin mutants.

  13. Inhibition of gap junction communication at ectopic Eph/ephrin boundaries underlies craniofrontonasal syndrome.

    Directory of Open Access Journals (Sweden)

    Alice Davy

    2006-10-01

    Full Text Available Mutations in X-linked ephrin-B1 in humans cause craniofrontonasal syndrome (CFNS, a disease that affects female patients more severely than males. Sorting of ephrin-B1-positive and -negative cells following X-inactivation has been observed in ephrin-B1(+/- mice; however, the mechanisms by which mosaic ephrin-B1 expression leads to cell sorting and phenotypic defects remain unknown. Here we show that ephrin-B1(+/- mice exhibit calvarial defects, a phenotype autonomous to neural crest cells that correlates with cell sorting. We have traced the causes of calvarial defects to impaired differentiation of osteogenic precursors. We show that gap junction communication (GJC is inhibited at ectopic ephrin boundaries and that ephrin-B1 interacts with connexin43 and regulates its distribution. Moreover, we provide genetic evidence that GJC is implicated in the calvarial defects observed in ephrin-B1(+/- embryos. Our results uncover a novel role for Eph/ephrins in regulating GJC in vivo and suggest that the pleiotropic defects seen in CFNS patients are due to improper regulation of GJC in affected tissues.

  14. Phenotypic variability in gap junction syndromic skin disorders: experience from KID and Clouston syndromes' clinical diagnostics.

    Science.gov (United States)

    Kutkowska-Kaźmierczak, Anna; Niepokój, Katarzyna; Wertheim-Tysarowska, Katarzyna; Giza, Aleksandra; Mordasewicz-Goliszewska, Maria; Bal, Jerzy; Obersztyn, Ewa

    2015-08-01

    Connexins belong to the family of gap junction proteins which enable direct cell-to-cell communication by forming channels in adjacent cells. Mutations in connexin genes cause a variety of human diseases and, in a few cases, result in skin disorders. There are significant differences in the clinical picture of two rare autosomal dominant syndromes: keratitis-ichthyosis-deafness (KID) syndrome and hidrotic ectodermal dysplasia (Clouston syndrome), which are caused by GJB2 and GJB6 mutations, respectively. This is despite the fact that, in both cases, malfunctioning of the same family proteins and some overlapping clinical features (nail dystrophy, hair loss, and palmoplantar keratoderma) is observed. KID syndrome is characterized by progressive vascularizing keratitis, ichthyosiform erythrokeratoderma, and neurosensory hearing loss, whereas Clouston syndrome is characterized by nail dystrophy, hypotrichosis, and palmoplantar keratoderma. The present paper presents a Polish patient with sporadic KID syndrome caused by the mutation of p.Asp50Asn in GJB2. The patient encountered difficulties in obtaining a correct diagnosis. The other case presented is that of a family with Clouston syndrome (caused by p.Gly11Arg mutation in GJB6), who are the first reported patients of Polish origin suffering from this disorder. Phenotype diversity among patients with the same genotypes reported to date is also summarized. The conclusion is that proper diagnosis of these syndromes is still challenging and should always be followed by molecular verification.

  15. Role of hippocampal CA1 area gap junction channels on morphine state-dependent learning.

    Science.gov (United States)

    Beheshti, Siamak; Hosseini, Seyyed Akbar Mir Seyyed; Noorbakhshnia, Maryam; Eivani, Mehdi

    2014-12-15

    Morphine produces a state dependent learning. The hippocampus is involved in this kind of learning. Gap junctions (GJs) are involved in some of the effects of morphine and exist in different areas of the hippocampus. We investigated the effects of blocking GJ channels of the hippocampal CA1 area, by means of pre-test bilateral injection of carbenoxolone (CBX), on morphine state dependent learning, using a passive avoidance task. Post-training subcutaneous administrations of morphine (0.5, 2.5, 5 and 7.5 mg/kg) dose-dependently impaired memory retrieval. Pre-test administration of morphine (0.5, 2.5, 5 and 7.5 mg/kg) induced a state-dependent retrieval of the memory acquired under post-training morphine influence. Pre-test injections of CBX (25, 75 and 150 nM) dose dependently prevented memory retrieval by post-training (7.5 mg/kg) and pre-test (0.5, 2.5, 5, 7.5 mg/kg) injections of morphine. The results suggest that intercellular coupling via GJ channels of the hippocampal CA1 area modulates morphine state dependent learning.

  16. Dimethylarsenic acid damages cellular DNA and inhibits gap junctional intercellular communication between human skin fibroblast cells

    Institute of Scientific and Technical Information of China (English)

    GuoXB; DengFR

    2002-01-01

    Although arsenic is identified as a human carcinogen,there is currently no accepted mechanism for its action or an established animal model for evaluating the carcinogenic activity of arsenic.To elucidate the mechanism of arsenic arcinogenesis,we investigated the effect of dimethylarsenic acid(DMAA),the main metabolite of inorganic arsenic in humans,on the cellular DNA and gap junctional intercellular communication (GJIC) between human skin fibroblast cells.Single-cell gel electrophoresis (SCGE) assay was used to detect the DNA damage in human skin fibroblast cells exposed to DMAA,and the GJIC between cells was detected by the scrape loading/dye transfer assay.DMAA at concentrations of 0.01-1.0 mmol·L-1 induced DNA damage in a dose-dependent manner,and GJIC between human skin fibroblast cells was significantly inhibited by DMAA at 1.0 mmol·L-1.Our results suggest that both genotoxic and nongenotoxic mechanism are involved in the mechanism of DMAA-induced cellular toxicity.

  17. Extract from the zooxanthellate jellyfish Cotylorhiza tuberculata modulates gap junction intercellular communication in human cell cultures.

    Science.gov (United States)

    Leone, Antonella; Lecci, Raffaella Marina; Durante, Miriana; Piraino, Stefano

    2013-05-22

    On a global scale, jellyfish populations in coastal marine ecosystems exhibit increasing trends of abundance. High-density outbreaks may directly or indirectly affect human economical and recreational activities, as well as public health. As the interest in biology of marine jellyfish grows, a number of jellyfish metabolites with healthy potential, such as anticancer or antioxidant activities, is increasingly reported. In this study, the Mediterranean "fried egg jellyfish" Cotylorhiza tuberculata (Macri, 1778) has been targeted in the search forputative valuable bioactive compounds. A medusa extract was obtained, fractionated, characterized by HPLC, GC-MS and SDS-PAGE and assayed for its biological activity on breast cancer cells (MCF-7) and human epidermal keratinocytes (HEKa). The composition of the jellyfish extract included photosynthetic pigments, valuable ω-3 and ω-6 fatty acids, and polypeptides derived either from jellyfish tissues and their algal symbionts. Extract fractions showed antioxidant activity and the ability to affect cell viability and intercellular communication mediated by gap junctions (GJIC) differentially in MCF-7 and HEKa cells. A significantly higher cytotoxicity and GJIC enhancement in MCF-7 compared to HEKa cells was recorded. A putative action mechanism for the anticancer bioactivity through the modulation of GJIC has been hypothesized and its nutraceutical and pharmaceutical potential was discussed.

  18. Cloning, mapping and mutation analysis of human gene GJB5 encoding gap junction protein b-5

    Institute of Scientific and Technical Information of China (English)

    XIA; Jiahui; (夏家辉); ZHENG; Duo; (郑多),; TANG; Dongsheng; (唐冬生); DAI; Heping; (戴和平); PAN; Qian; (潘乾); LONG; Zhigao; (龙志高); LIAO; Xiaodong; (廖晓东)

    2001-01-01

    By homologous EST searching and nested PCR a new human gene GJB5 encoding gap junction protein b-5 was identified. GJB5 was genetically mapped to human chromosome 1p33-p35 by FISH. RT-PCR revealed that it was expressed in skin, placenta and fetal skin. DNA sequencing of GJB5 was carried out in 142 patients with sensorineural hearing impairment and probands of 36 families with genetic diseases, including erythrokeratodermia (5 families), Charcot-Marie-Tooth disease (13), ptosis (4), and retinitis pigmentosa and deafness (14). Two missense mutations (686A→G, H229R; 25C→T, L9F) were detected in two sensorineural hearing impairment families. A heterologous deletion of 18 bp within intron was found in 3 families with heredity hearing impairment, and in one of the 3 families, a missense mutation (R265P) was identified also. But the deletion and missense mutation seemed not segregating with hearing impairment in the family. No abnormal mRNA or mRNA expression was detected in deletion carriers by RT-PCR analysis in skin tissue. Mutation analysis in 199 unaffected individuals revealed that two of them were carriers with the same 18 bp deletion.

  19. Cloning, mapping and mutation analysis of human gene GJB5 encoding gap junction protein b-5

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    By homologous EST searching and nested PCR a new human gene GJB5encoding gap junction protein b-5 was identified. GJB5 was genetically mapped to human chromosome 1p33-p35 by FISH. RT-PCR revealed that it was expressed in skin, placenta and fetal skin. DNA sequencing of GJB5 was carried out in 142 patients with sensorineural hearing impairment and probands of 36 families with genetic diseases, including erythrokeratodermia (5 families), Charcot-Marie-Tooth disease (13), ptosis (4), and retinitis pigmentosa and deafness (14). Two missense mutations (686A→G, H229R; 25C→T, L9F) were detected in two sensorineural hearing impairment families. A heterologous deletion of 18 bp within intron was found in 3 families with heredity hearing impairment, and in one of the 3 families, a missense mutation (R265P) was identified also. But the deletion and missense mutation seemed not segregating with hearing impairment in the family. No abnormal mRNA or mRNA expression was detected in deletion carriers by RT-PCR analysis in skin tissue. Mutation analysis in 199 unaffected individuals revealed that two of them were carriers with the same 18 bp deletion.

  20. ``Hybrid'' multi-gap/single-gap Josephson junctions: Evidence of macroscopic quantum tunneling in superconducting-to-normal switching experiments on MgB2/I/Pb and MgB2/I/Sn junctions

    Science.gov (United States)

    Carabello, Steve; Lambert, Joseph; Dai, Wenqing; Li, Qi; Chen, Ke; Cunnane, Daniel; Xi, X. X.; Ramos, Roberto

    We report results of superconducting-to-normal switching experiments on MgB2/I/Pb and MgB2/I/Sn junctions, with and without microwaves. These results suggest that the switching behavior is dominated by quantum tunneling through the washboard potential barrier, rather than thermal excitations or electronic noise. Evidence includes a leveling in the standard deviation of the switching current distribution below a crossover temperature, a Lorentzian shape of the escape rate enhancement peak upon excitation by microwaves, and a narrowing in the histogram of escape counts in the presence of resonant microwave excitation relative to that in the absence of microwaves. These are the first such results reported in ``hybrid'' Josephson tunnel junctions, consisting of multi-gap and single-gap superconducting electrodes.

  1. Dipyridamole increases gap junction coupling in bovine GM-7373 aortic endothelial cells by a cAMP-protein kinase A dependent pathway.

    Science.gov (United States)

    Begandt, D; Bintig, W; Oberheide, K; Schlie, S; Ngezahayo, A

    2010-02-01

    The scrape-loading/dye transfer technique was applied on the bovine aortic endothelial cell line GM-7373 to analyze the effects of the antithrombolytic drug dipyridamole on gap junction coupling in endothelial cells. We found that a cell treatment for 24 h with dipyridamole in therapeutically relevant concentrations (1-100 microM) increased gap junction coupling in a dose dependent manner. Similar to dipyridamole, forskolin as well as 8-Br-cAMP increased the gap junction coupling, while dibutyryl-cGMP (db-cGMP) did not affect the gap junction coupling of the GM-7373 endothelial cells. In parallel, a pharmacological inhibition of protein kinase A (PKA) with N-[2-(p-bromocinnamylamino)ethyl]-5-isoquinolinesulfonamide dihydrochloride (H-89), antagonised the action of dipyridamole on gap junction coupling. We propose that the observed dipyridamole induced increase in gap junction coupling in endothelial cells is related to a cAMP-PKA dependent phosphorylation pathway. The report shows that gap junction coupling in endothelial cells is a suitable therapeutic target for treatment of cardiovascular diseases.

  2. Effects of Taxol on the expression and distribution of connexin 43 in cultured cardiac myocytes under hypoxia%Taxol对缺氧培养乳鼠心肌细胞Cx43蛋白表达及分布的影响

    Institute of Scientific and Technical Information of China (English)

    王德国; 王新; 王安才; 朱红军; 孙贤林

    2013-01-01

    目的:观察微管稳定剂Taxol对缺氧导致的培养乳鼠心肌细胞Cx43表达和分布的影响.方法:差速贴壁梯度离心法分离培养乳大鼠心肌细胞,取培养4d细胞缺氧120 min,分别以不同浓度的Taxol干预;免疫印迹法检测聚合态微管蛋白含量、心肌细胞Cx43的蛋白表达,免疫荧光染色后激光共聚焦显微镜观察Cx43的分布.结果:正常培养心肌细胞Cx43分布在核膜和细胞的闰盘处,缺氧120 min可导致心肌细胞微管解聚,Cx43蛋白表达降低,在心肌细胞间连接处分布规律散失,核膜Cx43分布减弱或消失,而均匀分布在细胞膜上;在低剂量的Taxol作用下,心肌1细胞微管解聚状态缓解,Cx43蛋白表达和分布异常明显改善;随着Taxol剂量增加,这种改善作用更明显,呈现剂量依赖性.结论:缺氧引起乳鼠心肌细胞微管解聚,Cx43蛋白表达降低分布紊乱,微管稳定剂Taxol可以显著地保护缺氧导致的心肌Cx43异常,具有潜在的抗缺血性心律失常价值.

  3. Gap junction proteins in the blood-brain barrier control nutrient-dependent reactivation of Drosophila neural stem cells.

    Science.gov (United States)

    Spéder, Pauline; Brand, Andrea H

    2014-08-11

    Neural stem cells in the adult brain exist primarily in a quiescent state but are reactivated in response to changing physiological conditions. How do stem cells sense and respond to metabolic changes? In the Drosophila CNS, quiescent neural stem cells are reactivated synchronously in response to a nutritional stimulus. Feeding triggers insulin production by blood-brain barrier glial cells, activating the insulin/insulin-like growth factor pathway in underlying neural stem cells and stimulating their growth and proliferation. Here we show that gap junctions in the blood-brain barrier glia mediate the influence of metabolic changes on stem cell behavior, enabling glia to respond to nutritional signals and reactivate quiescent stem cells. We propose that gap junctions in the blood-brain barrier are required to translate metabolic signals into synchronized calcium pulses and insulin secretion.

  4. Pharmacological and Genetic Evidence for Gap Junctions as Potential New Insecticide Targets in the Yellow Fever Mosquito, Aedes aegypti.

    Science.gov (United States)

    Calkins, Travis L; Piermarini, Peter M

    2015-01-01

    The yellow fever mosquito Aedes aegypti is an important vector of viral diseases that impact global health. Insecticides are typically used to manage mosquito populations, but the evolution of insecticide resistance is limiting their effectiveness. Thus, identifying new molecular and physiological targets in mosquitoes is needed to facilitate insecticide discovery and development. Here we test the hypothesis that gap junctions are valid molecular and physiological targets for new insecticides. Gap junctions are intercellular channels that mediate direct communication between neighboring cells and consist of evolutionarily distinct proteins in vertebrate (connexins) and invertebrate (innexins) animals. We show that the injection of pharmacological inhibitors of gap junctions (i.e., carbenoxolone, meclofenamic acid, or mefloquine) into the hemolymph of adult female mosquitoes elicits dose-dependent toxic effects, with mefloquine showing the greatest potency. In contrast, when applied topically to the cuticle, carbenoxolone was the only inhibitor to exhibit full efficacy. In vivo urine excretion assays demonstrate that both carbenoxolone and mefloquine inhibit the diuretic output of adult female mosquitoes, suggesting inhibition of excretory functions as part of their mechanism of action. When added to the rearing water of 1st instar larvae, carbenoxolone and meclofenamic acid both elicit dose-dependent toxic effects, with meclofenamic acid showing the greatest potency. Injecting a double-stranded RNA cocktail against innexins into the hemolymph of adult female mosquitoes knock down whole-animal innexin mRNA expression and decreases survival of the mosquitoes. Taken together these data indicate that gap junctions may provide novel molecular and physiological targets for the development of insecticides.

  5. Mechanistical studies on gap junctions and their role in tumour promotion : Structure-activity relationships of polychlorinated biphelyls

    OpenAIRE

    Bager, Yvonne

    1997-01-01

    Chemically induced carcinogenesis is a multi-stage process that can be divided into at least three major stages, initiation, promotion and progression. The promotion stage involves clonal expansion of genetically altered cells (i.e. initiated cells). The final stage, progression, includes a karyotypic instability of cells that may result in malignancy. Several lines of evidence support the theory that inhibition of gap junctional intercellular communication plays a sig...

  6. Pharmacological and Genetic Evidence for Gap Junctions as Potential New Insecticide Targets in the Yellow Fever Mosquito, Aedes aegypti.

    Directory of Open Access Journals (Sweden)

    Travis L Calkins

    Full Text Available The yellow fever mosquito Aedes aegypti is an important vector of viral diseases that impact global health. Insecticides are typically used to manage mosquito populations, but the evolution of insecticide resistance is limiting their effectiveness. Thus, identifying new molecular and physiological targets in mosquitoes is needed to facilitate insecticide discovery and development. Here we test the hypothesis that gap junctions are valid molecular and physiological targets for new insecticides. Gap junctions are intercellular channels that mediate direct communication between neighboring cells and consist of evolutionarily distinct proteins in vertebrate (connexins and invertebrate (innexins animals. We show that the injection of pharmacological inhibitors of gap junctions (i.e., carbenoxolone, meclofenamic acid, or mefloquine into the hemolymph of adult female mosquitoes elicits dose-dependent toxic effects, with mefloquine showing the greatest potency. In contrast, when applied topically to the cuticle, carbenoxolone was the only inhibitor to exhibit full efficacy. In vivo urine excretion assays demonstrate that both carbenoxolone and mefloquine inhibit the diuretic output of adult female mosquitoes, suggesting inhibition of excretory functions as part of their mechanism of action. When added to the rearing water of 1st instar larvae, carbenoxolone and meclofenamic acid both elicit dose-dependent toxic effects, with meclofenamic acid showing the greatest potency. Injecting a double-stranded RNA cocktail against innexins into the hemolymph of adult female mosquitoes knock down whole-animal innexin mRNA expression and decreases survival of the mosquitoes. Taken together these data indicate that gap junctions may provide novel molecular and physiological targets for the development of insecticides.

  7. Critical role of gap junction communication, calcium and nitric oxide signaling in bystander responses to focal photodynamic injury

    OpenAIRE

    Calì, Bianca; Ceolin, Stefano; Ceriani, Federico; Bortolozzi, Mario; Agnellini, Andrielly H.R.; Zorzi, Veronica; Predonzani, Andrea; Bronte, Vincenzo; Molon, Barbara; Mammano, Fabio

    2015-01-01

    Ionizing and nonionizing radiation affect not only directly targeted cells but also surrounding “bystander” cells. The underlying mechanisms and therapeutic role of bystander responses remain incompletely defined. Here we show that photosentizer activation in a single cell triggers apoptosis in bystander cancer cells, which are electrically coupled by gap junction channels and support the propagation of a Ca2+ wave initiated in the irradiated cell. The latter also acts as source of nitric oxi...

  8. Critical role of gap junction communication, calcium and nitric oxide signaling in bystander responses to focal photodynamic injury

    OpenAIRE

    Cali, B.; Ceolin, S.; Ceriani, F.; Bortolozzi, M.; Agnellini, A.H.R.; Zorzi, V.; Predonzani, A.; BRONTE, V; Molon, B.; Mammano, F.

    2015-01-01

    Ionizing and nonionizing radiation affect not only directly targeted cells but also\\ud surrounding “bystander” cells. The underlying mechanisms and therapeutic role of\\ud bystander responses remain incompletely deined. Here we show that photosentizer\\ud activation in a single cell triggers apoptosis in bystander cancer cells, which are\\ud electrically coupled by gap junction channels and support the propagation of a Ca2+\\ud wave initiated in the irradiated cell. The latter also acts as source...

  9. Gap junctional communication between vascular cells. Induction of connexin43 messenger RNA in macrophage foam cells of atherosclerotic lesions.

    OpenAIRE

    Polacek, D.; Lal, R; Volin, M. V.; Davies, P F

    1993-01-01

    The structure and function of blood vessels depend on the ability of vascular cells to receive and transduce signals and to communicate with each other. One means by which vascular cells have been shown to communicate is via gap junctions, specifically connexin43. In atherosclerosis, the normal physical patterns of communication are disrupted by the subendothelial infiltration and accumulation of blood monocytes, which in turn can differentiate into resident foam cells. In this paper we repor...

  10. Analytically determined topological phase diagram of the proximity-induced gap in diffusive n-terminal Josephson junctions

    Science.gov (United States)

    Amundsen, Morten; Ouassou, Jabir Ali; Linder, Jacob

    2017-01-01

    Multiterminal Josephson junctions have recently been proposed as a route to artificially mimic topological matter with the distinct advantage that its properties can be controlled via the superconducting phase difference, giving rise to Weyl points in 4-terminal geometries. A key goal is to accurately determine when the system makes a transition from a gapped to non-gapped state as a function of the phase differences in the system, the latter effectively playing the role of quasiparticle momenta in conventional topological matter. We here determine the proximity gap phase diagram of diffusive n-terminal Josephson junctions (), both numerically and analytically, by identifying a class of solutions to the Usadel equation at zero energy in the full proximity effect regime. We present an analytical equation which provides the phase diagram for an arbitrary number of terminals n. After briefly demonstrating the validity of the analytical approach in the previously studied 2- and 3-terminal cases, we focus on the 4-terminal case and map out the regimes where the electronic excitations in the system are gapped and non-gapped, respectively, demonstrating also in this case full agreement between the analytical and numerical approach.

  11. Implanted neural progenitor cells regulate glial reaction to brain injury and establish gap junctions with host glial cells.

    Science.gov (United States)

    Talaverón, Rocío; Matarredona, Esperanza R; de la Cruz, Rosa R; Macías, David; Gálvez, Victoria; Pastor, Angel M

    2014-04-01

    Transplantation of neural stem/progenitor cells (NPCs) in the lesioned brain is able to restore morphological and physiological alterations induced by different injuries. The local microenvironment created at the site of grafting and the communication between grafted and host cells are crucial in the beneficial effects attributed to the NPC implants. We have previously described that NPC transplantation in an animal model of central axotomy restores firing properties and synaptic coverage of lesioned neurons and modulates their trophic factor content. In this study, we aim to explore anatomical relationships between implanted NPCs and host glia that might account for the implant-induced neuroprotective effects. Postnatal rat subventricular zone NPCs were isolated and grafted in adult rats after transection of the medial longitudinal fascicle. Brains were removed and analyzed eight weeks later. Immunohistochemistry for different glial markers revealed that NPC-grafted animals displayed significantly greater microglial activation than animals that received only vehicle injections. Implanted NPCs were located in close apposition to activated microglia and reactive astrocytes. The gap junction protein connexin43 was present in NPCs and glial cells at the lesion site and was often found interposed within adjacent implanted and glial cells. Gap junctions were identified between implanted NPCs and host astrocytes and less frequently between NPCs and microglia. Our results show that implanted NPCs modulate the glial reaction to lesion and establish the possibility of communication through gap junctions between grafted and host glial cells which might be involved in the restorative effects of NPC implants.

  12. Cataract-causing mutation of human connexin 46 impairs gap junction, but increases hemichannel function and cell death.

    Directory of Open Access Journals (Sweden)

    Qian Ren

    Full Text Available Connexin channels play a critical role in maintaining metabolic homeostasis and transparency of the lens. Mutations in connexin genes are linked to congenital cataracts in humans. The G143R missense mutation on connexin (Cx 46 was recently reported to be associated with congenital Coppock cataracts. Here, we showed that the G143R mutation decreased Cx46 gap junctional coupling in a dominant negative manner; however, it significantly increased gap junctional plaques. The G143R mutant also increased hemichannel activity, inversely correlated with the level of Cx46 protein on the cell surface. The interaction between cytoplasmic loop domain and C-terminus has been shown to be involved in gating of connexin channels. Interestingly, the G143R mutation enhanced the interaction between intracellular loop and Cx46. Furthermore, this mutation decreased cell viability and the resistance of the cells to oxidative stress, primarily due to the increased hemichannel function. Together, these results suggest that mutation of this highly conserved residue on the cytoplasmic loop domain of Cx46 enhances its interaction with the C-terminus, resulting in a reduction of gap junction channel function, but increased hemichannel function. This combination leads to the development of human congenital cataracts.

  13. MicroRNA Intercellular Transfer and Bioelectrical Regulation of Model Multicellular Ensembles by the Gap Junction Connectivity.

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    Cervera, Javier; Meseguer, Salvador; Mafe, Salvador

    2017-08-17

    We have studied theoretically the microRNA (miRNA) intercellular transfer through voltage-gated gap junctions in terms of a biophysically grounded system of coupled differential equations. Instead of modeling a specific system, we use a general approach describing the interplay between the genetic mechanisms and the single-cell electric potentials. The dynamics of the multicellular ensemble are simulated under different conditions including spatially inhomogeneous transcription rates and local intercellular transfer of miRNAs. These processes result in spatiotemporal changes of miRNA, mRNA, and ion channel protein concentrations that eventually modify the bioelectrical states of small multicellular domains because of the ensemble average nature of the electrical potential. The simulations allow a qualitative understanding of the context-dependent nature of the effects observed when specific signaling molecules are transferred through gap junctions. The results suggest that an efficient miRNA intercellular transfer could permit the spatiotemporal control of small cellular domains by the conversion of single-cell genetic and bioelectric states into multicellular states regulated by the gap junction interconnectivity.

  14. Blockade of gap junction hemichannel protects secondary spinal cord injury from activated microglia-mediated glutamate exitoneurotoxicity.

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    Umebayashi, Daisuke; Natsume, Atsushi; Takeuchi, Hideyuki; Hara, Masahito; Nishimura, Yusuke; Fukuyama, Ryuichi; Sumiyoshi, Naoyuki; Wakabayashi, Toshihiko

    2014-12-15

    We previously demonstrated that activated microglia release excessive glutamate through gap junction hemichannels and identified a novel gap junction hemichannel blocker, INI-0602, that was proven to penetrate the blood-brain barrier and be an effective treatment in mouse models of amyotrophic lateral sclerosis and Alzheimer disease. Spinal cord injury causes tissue damage in two successive waves. The initial injury is mechanical and directly causes primary tissue damage, which induces subsequent ischemia, inflammation, and neurotoxic factor release resulting in the secondary tissue damage. These lead to activation of glial cells. Activated glial cells such as microglia and astrocytes are common pathological observations in the damaged lesion. Activated microglia release glutamate, the major neurotoxic factor released into the extracellular space after neural injury, which causes neuronal death at high concentration. In the present study, we demonstrate that reduction of glutamate-mediated exitotoxicity via intraperitoneal administration of INI-0602 in the microenvironment of the injured spinal cord elicited neurobehavioral recovery and extensive suppression of glial scar formation by reducing secondary tissue damage. Further, this intervention stimulated anti-inflammatory cytokines, and subsequently elevated brain-derived neurotrophic factor. Thus, preventing microglial activation by a gap junction hemichannel blocker, INI-0602, may be a promising therapeutic strategy in spinal cord injury.

  15. Microfluidic application-specific integrated device for monitoring direct cell-cell communication via gap junctions between individual cell pairs

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    Lee, Philip J.; Hung, Paul J.; Shaw, Robin; Jan, Lily; Lee, Luke P.

    2005-05-01

    Direct cell-cell communication between adjacent cells is vital for the development and regulation of functional tissues. However, current biological techniques are difficult to scale up for high-throughput screening of cell-cell communication in an array format. In order to provide an effective biophysical tool for the analysis of molecular mechanisms of gap junctions that underlie intercellular communication, we have developed a microfluidic device for selective trapping of cell-pairs and simultaneous optical characterizations. Two different cell populations can be brought into membrane contact using an array of trapping channels with a 2μm by 2μm cross section. Device operation was verified by observation of dye transfer between mouse fibroblasts (NIH3T3) placed in membrane contact. Integration with lab-on-a-chip technologies offers promising applications for cell-based analytical tools such as drug screening, clinical diagnostics, and soft-state biophysical devices for the study of gap junction protein channels in cellular communications. Understanding electrical transport mechanisms via gap junctions in soft membranes will impact quantitative biomedical sciences as well as clinical applications.

  16. Cochlear implantation effect on deaf children with gap junction protein beta 2 gene mutation

    Institute of Scientific and Technical Information of China (English)

    KONG Ying; LIU Sha; WANG Su-ju; Li Shu-jing; LIANG Shuang

    2013-01-01

    Background The popularization and promotion of gene diagnosis technology makes it possible to detect deafness genes for children with congenital hearing impairment,and the proportion of gap junction protein beta 2 (GJB2) gene mutations in cochlear implant patients is 26.5% We did follow-up evaluation on auditory rehabilitation effect for all 31 deaf children with GJB2 gene mutation after cochlear implantation to provide a reference for such patients.Methods Application of “the genetic deafness gene chip detection kit” and “gene complete sequence analysis” were applied to conduct detection on common genetic deafness gene mutation hotspots of the hearing impaired children with cochlear implantation.To conduct auditory rehabilitation effect evaluation on all 31 cases of patients with GJB2 genetic deafness after 3,6 and 12 months of the operation respectively.The single factor repeated measure analysis of variance (ANOVA) was applied to analysis whether there were significant difference among the results of initial consonant of a Chinese syllable recognition at 3 different stages after the operation,the results of vowel of a Chinese syllable recognition at 3 different stages after the operation,and the results of two-syllable recognition at 3 different stages after the operation.Results The 235delC is the high-incidence mutational site in 31 cases of patients with GJB2 genetic deafness,and the total detection rate is up to 90.3% (28/31).There were significant differences in the initial consonant and the vowel of a Chinese syllable recognition rate,and the two-syllable recognition rates at 3,6,and 12 months after the operation (P<0.01).Conclusion Cochlear implantation is a safe and effective measure for auditory reconstruction,enabling patients with GJB2 hereditary severe sensorineural deafness to achieve auditory speech recognition effectively.

  17. Thalamic gap junctions control local neuronal synchrony and influence macroscopic oscillation amplitude during EEG alpha rhythms

    Directory of Open Access Journals (Sweden)

    Stuart eHughes

    2011-08-01

    Full Text Available Although EEG alpha ( (8-13 Hz rhythms are often considered to reflect an ‘idling’ brain state, numerous studies indicate that they are also related to many aspects of perception. Recently, we outlined a potential cellular substrate by which such aspects of perception might be linked to basic  rhythm mechanisms. This scheme relies on a specialized subset of rhythmically bursting thalamocortical (TC neurons (high-threshold bursting cells in the lateral geniculate nucleus (LGN which are interconnected by gap junctions (GJs. By engaging GABAergic interneurons, that in turn inhibit conventional relay-mode TC neurons, these cells can lead to an effective temporal framing of thalamic relay-mode output. Although the role of GJs is pivotal in this scheme, evidence for their involvement in thalamic  rhythms has thus far mainly derived from experiments in in vitro slice preparations. In addition, direct anatomical evidence of neuronal GJs in the LGN is currently lacking. To address the first of these issues we tested the effects of the GJ inhibitors, carbenoxolone (CBX and 18-glycyrrhetinic acid (18-GA, given directly to the LGN via reverse microdialysis, on spontaneous LGN and EEG  rhythms in behaving cats. We also examined the effect of CBX on  rhythm-related LGN unit activity. Indicative of a role for thalamic GJs in these activities, 18-GA and CBX reversibly suppressed both LGN and EEG  rhythms, with CBX also decreasing neuronal synchrony. To address the second point, we used electron microscopy to obtain definitive ultrastructural evidence for the presence of GJs between neurons in the cat LGN. As interneurons show no phenotypic evidence of GJ coupling (i.e. dye-coupling and spikelets we conclude that these GJs must belong to TC neurons. The potential significance of these findings for relating macroscopic changes in  rhythms to basic cellular processes is discussed.

  18. Characterization of a gap-junctional intercellular communication (GJIC) assay using cigarette smoke.

    Science.gov (United States)

    Roemer, Ewald; Lammerich, Hans-Peter; Conroy, Lynda L; Weisensee, Dirk

    2013-06-07

    Inhibition of gap-junctional intercellular communication (GJIC) via exposure to various toxic substances has been implicated in tumor promotion. In the present study, cigarette smoke total particulate matter (TPM), a known inhibitor of GJIC, were used to characterize a new GJIC screening assay in three independent experiments. The main features of this assay were automated fluorescence microscopy combined with non-invasive parachute technique. Rat liver epithelial cells (WB-F344) were stained with the fluorescent dye Calcein AM (acetoxymethyl) and exposed to TPM from the Kentucky Reference Cigarette 2R4F (a blend of Bright and Burley tobaccos) and from two single-tobacco cigarettes (Bright and Burley) for 3h. Phorbol-12-myristate-13-acetate (TPA) was used as positive control and 0.5% dimethyl sulfoxide (DMSO) as solvent control. The transfer of dye to adjacent cells (percentage of stained cells) was used as a measure of cellular communication. A clear and reproducible dose-response of GJIC inhibition following TPM exposure was seen. Reproducibility and repeatability measurements for the 2R4F cigarette were 3.7% and 6.9%, respectively. The half-maximal effective concentration values were 0.34ng/ml for TPA, 0.050mg/ml for the 2R4F, 0.044mg/ml for the Bright cigarette, and 0.060mg/ml for the Burley cigarette. The assay was able to discriminate between the two single-tobacco cigarettes (P<0.0001), and between the single-tobacco cigarettes and the 2R4F (P=0.0008, 2R4F vs. Burley and P<0.0001, 2R4F vs. Bright). Thus, this assay can be used to determine the activity of complex mixtures such as cigarette smoke with high throughput and high precision. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

  19. INHIBITION OF GAP JUNCTIONAL INTERCELLULAR COMMUNICATION BY PERFLUORINATED COMPOUNDS IN RAT LIVER AND DOLPHIN KIDNEY EPITHELIAL CELL LINES IN VITRO AND SPRAGUE-DAWLEY RATS IN VIVO

    Science.gov (United States)

    Abstract Gap Junctional Intercellular Communication (GJIC) is the major pathway of intercellular signal transduction, and is, thus, important for normal cell growth and function. Recent studies have revealed a global distribution of some perfluorinated organic compounds e...

  20. Connexin mutation that causes dominant congenital cataracts inhibits gap junctions, but not hemichannels, in a dominant negative manner.

    Science.gov (United States)

    Banks, Eric A; Toloue, Masoud M; Shi, Qian; Zhou, Zifei Jade; Liu, Jialu; Nicholson, Bruce J; Jiang, Jean X

    2009-02-01

    The connexin (Cx) 50, E48K, mutation is associated with a human dominant congenital cataract; however, the underlying molecular mechanism has not been characterized. The glutamate (E) residue at position 48 is highly conserved across animal species and types of connexins. When expressed in paired Xenopus oocytes, human (h) and chicken (ch) Cx50 E48K mutants showed no electrical coupling. In addition, this mutation acts in a dominant negative manner when paired hetero-typically or hetero-merically with wild-type Cx50, but has no such effect on Cx46, the other lens fiber connexin. A similar loss-of-function and dominant negative effect was observed using dye transfer assays in the same system. By using two different dye transfer methods, with two different tracer dyes, we found chCx50 E48K expressed in chicken lens embryonic fibroblast cells by retroviral infection similarly failed to induce dye coupling, and prevented wild-type chCx50 from forming functional gap junctions. In contrast to its effect on gap junctions, the E48K mutation has no effect on hemichannel activity when assayed using electrical conductance in oocytes, and mechanically induced dye uptake in cells. Cx50 is functionally involved in cell differentiation and lens development, and the E48K mutant promotes primary lens cell differentiation indistinguishable from wild-type chCx50, despite its lack of junctional channel function. Together the data show that mutations affecting gap junctions but not hemichannel function of Cx50 can lead to dominant congenital cataracts in humans. This clearly supports the model of intercellular coupling of fiber cells creating a microcirculation of nutrients and metabolites required for lens transparency.

  1. Detecting and estimating rectification of gap junction conductance based on simulations of dual-cell recordings from a pair and a network of coupled cells.

    Science.gov (United States)

    Fortier, Pierre A

    2010-07-21

    Gap junctions can exhibit rectification of conductance. Some reports use inequality of coupling coefficients as the first sign of the possible existence of rectification (Devor and Yarom, 2002; Fan et al., 2005; Levavi-Sivan et al., 2005; Mann-Metzer and Yarom, 1999; Nolan et al., 1999; Szabadics et al., 2001). However, mathematical modeling and simulations of electrotonic coupling between an isolated pair of neurons showed conditions where the coupling coefficients were unreliable indicators of rectification. On the other hand, the transfer resistances were found to be reliable indicators of junctional rectification. The existing mathematical model of cell coupling (Bennett, 1966; Devor and Yarom, 2002; Verselis and Veenstra, 2000) was extended in order to measure rectification of the junctional conductances directly between dual-recorded neurons whether isolated or surrounded by a simulated 3-dimensional network of heterogeneous cells whose gap junctions offered parallel paths for current flow between the recorded neurons. The results showed that the transfer resistances could still detect rectification of the gap junction linking the dual-recorded neurons when embedded in a coupled cell network and that a mathematical model could estimate the conductances in both directions through this gap junction using only data that would be available from real dual-intracellular penetrations which allow electrophysiological recordings and intracellular staining. Rectification of gap junctions in unrecorded cells of a biologically realistic coupled cell network had negligible effects on the voltage responses of the dual-recorded neurons because of minimal current passing through these surrounding cells.

  2. Depression of Intraocular Pressure Following Inactivation of Connexin43 in the Nonpigmented Epithelium of the Ciliary Body

    Science.gov (United States)

    Calera, Mónica R.; Wang, Zhao; Sanchez-Olea, Roberto; Paul, David L.; Civan, Mortimer M.; Goodenough, Daniel A.

    2010-01-01

    Purpose Conditional inactivation of connexin43 (Cx43) in the pigmented epithelium of the mouse eye results in a reduction in aqueous humor production and complete loss of the vitreous chamber. It was proposed that gap junctions between pigmented and nonpigmented epithelia of the ciliary body are critical for the production of the aqueous humor. To form such junctions, Cx43 in the pigmented epithelium must interact with connexin(s) present in the adjacent cells of the nonpigmented epithelium. The importance of Cx43 expression in the nonpigmented epithelium for the establishment of gap junctions and the regulation of intraocular pressure was tested. Methods To inactivate Cx43 in the nonpigmented epithelium of the mouse eye, a mouse line was crossed with a floxed Cx43 locus (Cx43flox/flox) and a transgenic mouse line expressing cre recombinase under the control of the Pax6α promoter. General eye structure was evaluated by light microscopy, gap junctions were analyzed by electron microscopy, and intraocular pressure was directly assessed with micropipettes. Results In Pax6α-cre/Cx43flox/flox mice, Cx43 was partially inactivated in the nonpigmented epithelium of the ciliary body and iris. Animals developed dilatations between the pigmented and nonpigmented epithelia and displayed a significant reduction in intraocular pressure. However, gap junctions between the ciliary epithelial layers were decreased but not eliminated. Conclusions Cx43 expression in the nonpigmented epithelium of the ciliary body contributes to the formation of gap junctions with the cells of the pigmented epithelium. These gap junctions play a critical role in maintaining the physical integrity of the ciliary body epithelium. Although the partial loss of Cx43 from the nonpigmented epithelium was correlated with a measurable drop in intraocular pressure, possible changes in Cx43 in the aqueous outflow pathway may provide an additional contribution to the observed phenotype. PMID:19168903

  3. Cardioprotective effect of connexin 43 expression regulated by PI3K/Akt on hydrogen sulfide postconditioning in isolated ischemic and reperfused rat hearts%PI3K/Akt调节线粒体Cx43蛋白表达在H2S后处理离体大鼠心肌中的保护作用

    Institute of Scientific and Technical Information of China (English)

    张可璇; 毛洪雅; 孟雪; 惠夏; 张邓新; 季永

    2013-01-01

    Aim To investigate whether the PBK/Akt signaling pathway regulates connexin 43 expression to protect isolated rat hearts against ischemia/reperfusion ( I/R ) injury in hydrogen sulfide postconditioning. Methods Hearts of 56 male SD rats were isolated and linked to the Langendorff apparatus. They were randomly divided into 4 groups( n = 14): ischemia reper-fusion group ( I/R ), PBK/Akt signaling pathway inhibitor LY294002 group ( LY ), hydrogen sulfide post-conditioning group ( NP ), and hydrogen sulfide with to all the nuclei counted was used as apoptotic index ( AI ). The expression of total Cx43 and phosphoryla-tion Cx43 in mitochondria were determined with Western blot analysis at the end of reperfusion. Results No differences in baseline hemodynamics were observed a-mong the experimental groups ( P > 0. 05 ). After reperfusion, compared with I/R group, NP group had better hemodynamics; the myocardial infarct size and cardiocyte apoptotic index were much lower ( P < 0. 05 ); the expression of tCx43 ( total connexin 43 , t