Aberrant cell-to-cell coupling in Ca2+-overloaded guinea pig ventricular muscles

2008 ◽  
Vol 294 (6) ◽  
pp. C1419-C1429 ◽  
Author(s):  
Nagomi Kurebayashi ◽  
Hiroto Nishizawa ◽  
Yuji Nakazato ◽  
Hidetake Kurihara ◽  
Satoshi Matsushita ◽  
...  
Keyword(s):  
Guinea Pig ◽  
Gap Junction ◽  
Time Lag ◽  
Propagation Delay ◽  
High Frequency Stimulation ◽  
Cell Coupling ◽  
Gap Junctional Communication ◽  
Gap Junction Channel ◽  
Junctional Communication ◽  
Cell Propagation

To investigate how intercellular coupling can be changed during Ca2+ overloading of ventricular muscle, we studied Ca2+ signals in individual cells and the histochemistry of the major gap junction channel, connexin43 (Cx43), using multicellular preparations. Papillary muscles were obtained from guinea pig ventricles and loaded with rhod-2. Sequential Ca2+ images of surface cells were obtained with a confocal microscope. In intact muscles, all cells showed simultaneous Ca2+ transients in response to field stimulation over a field of view of 0.3 × 0.3 mm2. In severely Ca2+-overloaded muscles, obtained by high-frequency stimulation in nonflowing Krebs solution, cells became less responsive to stimulation. Furthermore, nonsimultaneous but serial onsets of Ca2+ transients were often detected, suggesting a propagation delay of action potentials. The time lag of the onset between two aligned cells was sometimes as long as 100 ms. Similar lags were also observed in muscles with gap junction channels inhibited by heptanol. To investigate whether the phosphorylation state of Cx43 is affected in Ca2+-overloaded muscles, the distributions of phosphorylated and nonphosphorylated Cx43 were determined using specific antibodies. Most of the Cx43 was phosphorylated in the nonoverloaded muscles, whereas nonphosphorylated Cx43 was significantly elevated in severely Ca2+-overloaded muscles. Our results suggest that the propagation delay of action potential within a small area, a few square millimeters, can be a cause of abnormal conduction and a microreentry in Ca2+-overloaded heart. Inactivation of Na+ channels and inhibition of gap junctional communication may underlie the cell-to-cell propagation delay.

scholarly journals Connexin43 phosphorylation: structural changes and biological effects

2009 ◽  
Vol 419 (2) ◽  
pp. 261-272 ◽  
Author(s):  
Joell L. Solan ◽  
Paul D. Lampe
Keyword(s):  
Gap Junction ◽  
Structural Changes ◽  
Biological Effects ◽  
Gap Junctional Communication ◽  
Connexin Gene ◽  
Gap Junction Channel ◽  
Junctional Communication ◽  
Cell Growth And Differentiation ◽  
And Function ◽  
Gap Junctional

Vertebrate gap junctions, composed of proteins from the connexin gene family, play critical roles in embryonic development, co-ordinated contraction of excitable cells, tissue homoeostasis, normal cell growth and differentiation. Phosphorylation of connexin43, the most abundant and ubiquitously expressed connexin, has been implicated in the regulation of gap junctional communication at several stages of the connexin ‘life cycle’, including hemichannel oligomerization, export of the protein to the plasma membrane, hemichannel activity, gap junction assembly, gap junction channel gating and connexin degradation. Consistent with a short (1–5 h) protein half-life, connexin43 phosphorylation is dynamic and changes in response to activation of many different kinases. The present review assesses our current understanding of the effects of phosphorylation on connexin43 structure and function that in turn regulate gap junction biology, with an emphasis on events occurring in heart and skin.

Gap junction-mediated transfer of left-right patterning signals in the early chick blastoderm is upstream of Shh asymmetry in the node

Development ◽  
1999 ◽  
Vol 126 (21) ◽  
pp. 4703-4714 ◽  
Author(s):  
M. Levin ◽  
M. Mercola
Keyword(s):  
Gene Expression ◽  
Gap Junction ◽  
Large Scale ◽  
Chick Blastoderm ◽  
Gap Junction Channels ◽  
Gap Junctional Communication ◽  
Junctional Communication ◽  
Left And Right ◽  
The Right ◽  
Gap Junctional

Invariant patterning of left-right asymmetry during embryogenesis depends upon a cascade of inductive and repressive interactions between asymmetrically expressed genes. Different cascades of asymmetric genes distinguish the left and right sides of the embryo and are maintained by a midline barrier. As such, the left and right sides of an embryo can be viewed as distinct and autonomous fields. Here we describe a series of experiments that indicate that the initiation of these programs requires communication between the two sides of the blastoderm. When deprived of either the left or the right lateral halves of the blastoderm, embryos are incapable of patterning normal left-right gene expression at Hensen's node. Not only are both flanks required, suggesting that there is no single signaling source for LR pattern, but the blastoderm must be intact. These results are consistent with our previously proposed model in which the orientation of LR asymmetry in the frog, Xenopus laevis, depends on large-scale partitioning of LR determinants through intercellular gap junction channels (M. Levin and M. Mercola (1998) Developmental Biology 203, 90–105). Here we evaluate whether gap junctional communication is required for the LR asymmetry in the chick, where it is possible to order early events relative to the well-characterized left and right hierarchies of gene expression. Treatment of cultured chick embryos with lindane, which diminishes gap junctional communication, frequently unbiased normal LR asymmetry of Shh and Nodal gene expression, causing the normally left-sided program to be recapitulated symmetrically on the right side of the embryo. A survey of early expression of connexin mRNAs revealed that Cx43 is present throughout the blastoderm at Hamburger-Hamilton stage 2–3, prior to known asymmetric gene expression. Application of antisense oligodeoxynucleotides or blocking antibody to cultured embryos also resulted in bilateral expression of Shh and Nodal transcripts. Importantly, the node and primitive streak at these stages lack Cx43 mRNA. This result, together with the requirement for an intact blastoderm, suggests that the path of communication through gap junction channels circumvents the node and streak. We propose that left-right information is transferred unidirectionally throughout the epiblast by gap junction channels in order to pattern left-sided Shh expression at Hensen's node.

Gap-junctional communication in a communication-defective and in a communication-competent teratocarcinoma cell line

1985 ◽  
Vol 76 (1) ◽  
pp. 85-95
Author(s):  
C.W. Lo ◽  
D. Fang ◽  
M.L. Hooper
Keyword(s):  
Cell Line ◽  
Coupling Coefficient ◽  
Dye Coupling ◽  
Cell Coupling ◽  
Teratocarcinoma Cell ◽  
Gap Junctional Communication ◽  
Junctional Communication ◽  
Gap Junctional ◽  
Ionic Coupling ◽  
Cell Cell

We examined the gap-junctional communication properties of a communication-defective cell line R5/3 and its communication-competent revertant H2T12. For these studies, we carried out microelectrode impalements to monitor ionic coupling and dye coupling. Our dye-injection experiments revealed that the H2T12 cells are much more efficient in dye coupling than the R5/3 cells. This latter observation is in agreement with the previous finding that the H2T12 cells are much better metabolically coupled than the R5/3 cells. With ionic coupling measurements, however, both cell lines exhibited similar levels of cell-cell coupling. The R5/3 cells demonstrated an ionic coupling coefficient of 0.19 +/− 0.011 (S.E.M.) and H2T12 a coupling coefficient of 0.25 +/− 0.009 (S.E.M.). These results in conjunction with observations from other studies indicate that the different experimental approaches for monitoring gap-junctional communication may have different levels of sensitivity for detecting as opposed to measuring the level of cell-cell coupling.

Phosphorylation of connexin43 gap junction protein in uninfected and Rous sarcoma virus-transformed mammalian fibroblasts

1990 ◽  
Vol 10 (4) ◽  
pp. 1754-1763
Author(s):  
D S Crow ◽  
E C Beyer ◽  
D L Paul ◽  
S S Kobe ◽  
A F Lau
Keyword(s):  
Gap Junctions ◽  
Gap Junction ◽  
Rous Sarcoma Virus ◽  
Gap Junctional Communication ◽  
Rous Sarcoma ◽  
Junctional Communication ◽  
Junction Protein ◽  
Sarcoma Virus ◽  
Gap Junction Protein ◽  
Gap Junctional

Gap junctions are membrane channels that permit the interchange of ions and other low-molecular-weight molecules between adjacent cells. Rous sarcoma virus (RSV)-induced transformation is marked by an early and profound disruption of gap-junctional communication, suggesting that these membrane structures may serve as sites of pp60v-src action. We have begun an investigation of this possibility by identifying and characterizing putative proteins involved in junctional communication in fibroblasts, the major cell type currently used to study RSV-induced transformation. We found that uninfected mammalian fibroblasts do not appear to contain RNA or protein related to connexin32, the major rat liver gap junction protein. In contrast, vole and mouse fibroblasts contained a homologous 3.0-kilobase RNA similar in size to the heart tissue RNA encoding the gap junction protein, connexin43. Anti-connexin43 peptide antisera specifically reacted with three proteins of approximately 43, 45 and 47 kilodaltons (kDa) from communicating fibroblasts. Gap junctions of heart cells contained predominantly 45- and 47-kDa species similar to those found in fibroblasts. Uninfected fibroblast 45- and 47-kDa proteins were phosphorylated on serine residues. Phosphatase digestions of 45- and 47-kDa proteins and pulse-chase labeling studies indicated that these proteins represented phosphorylated forms of the 43-kDa protein. Phosphorylation of connexin protein appeared to occur shortly after synthesis, followed by an equally rapid dephosphorylation. In comparison with these results, connexin43 protein in RSV-transformed fibroblasts contained both phosphotyrosine and phosphoserine. Thus, the presence of phosphotyrosine in connexin43 correlates with the loss of gap-junctional communication observed in RSV-transformed fibroblasts.

Distribution, propagation, and coordination of contractile activity in lymphatics

1993 ◽  
Vol 264 (4) ◽  
pp. H1283-H1291 ◽  
Author(s):  
D. C. Zawieja ◽  
K. L. Davis ◽  
R. Schuster ◽  
W. M. Hinds ◽  
H. J. Granger
Keyword(s):  
Gap Junction ◽  
Conduction Velocity ◽  
Contractile Activity ◽  
Lymph Flow ◽  
Rat Small Intestine ◽  
Gap Junctional Communication ◽  
Junctional Communication ◽  
Gap Junctional ◽  
Flow Stream

The propagation and coordination of lymphatic contractions were studied in the mesentery of the rat small intestine using in situ microscopic observation. Indexes of lymphatic diameter were simultaneously measured at two adjacent lymphangions in spontaneously contracting lymphatics (n = 51). Diameter index, contraction frequency, and the percentage of the intersegmental contractions that were propagated and coordinated (PP) were determined at both sites. The conduction velocity of the contractile activity and the percentage of the coordinated contractions that were propagated both antegrade to the direction of lymph flow and retrograde to the flow stream were determined. The results indicate that 1) 80-90% of the lymphatic contractions in the vessels we evaluated were propagated, 2) the wave of contractile activity propagated both centrally and peripherally, and 3) the conduction velocity of the contractile activity was approximately 4-8 mm/s. We tested the hypothesis that gap junctional communication is responsible for the coordination of the contractile event. To accomplish this, we used the gap junction blockers n-heptanol and oleic acid. PP was 90 +/- 4% under normal conditions and fell to a minimum value of 55 +/- 7% during the gap junction blockade. These results indicate that gap junctional communication played an important role in the propagation and coordination of contractions that occurred in spontaneously active lymphatics.

scholarly journals Connexin Mutants Compromise the Lens Circulation and Cause Cataracts through Biomineralization

2020 ◽  
Vol 21 (16) ◽  
pp. 5822
Author(s):  
Viviana M. Berthoud ◽  
Junyuan Gao ◽  
Peter J. Minogue ◽  
Oscar Jara ◽  
Richard T. Mathias ◽  
...  
Keyword(s):  
Gap Junction ◽  
Lens Fiber ◽  
Lens Fiber Cell ◽  
Alizarin Red ◽  
General Process ◽  
Fiber Cells ◽  
Gap Junctional Communication ◽  
Junctional Communication ◽  
Gap Junctional ◽  
Junction Proteins

Gap junction-mediated intercellular communication facilitates the circulation of ions, small molecules, and metabolites in the avascular eye lens. Mutants of the lens fiber cell gap junction proteins, connexin46 (Cx46) and connexin50 (Cx50), cause cataracts in people and in mice. Studies in mouse models have begun to elucidate the mechanisms by which these mutants lead to cataracts. The expression of the dominant mutants causes severe decreases in connexin levels, reducing the gap junctional communication between lens fiber cells and compromising the lens circulation. The impairment of the lens circulation results in several changes, including the accumulation of Ca2+ in central lens regions, leading to the formation of precipitates that stain with Alizarin red. The cataract morphology and the distribution of Alizarin red-stained material are similar, suggesting that the cataracts result from biomineralization within the organ. In this review, we suggest that this may be a general process for the formation of cataracts of different etiologies.

scholarly journals The Sleep-inducing Lipid Oleamide Deconvolutes Gap Junction Communication and Calcium Wave Transmission in Glial Cells

1997 ◽  
Vol 139 (7) ◽  
pp. 1785-1792 ◽  
Author(s):  
Xiaojun Guan ◽  
Benjamin F. Cravatt ◽  
George R. Ehring ◽  
James E. Hall ◽  
Dale L. Boger ◽  
...  
Keyword(s):  
Gap Junction ◽  
Glial Cells ◽  
Wave Transmission ◽  
Calcium Wave ◽  
Mediated Communication ◽  
Gap Junctional Communication ◽  
Gap Junction Communication ◽  
Junctional Communication ◽  
Nervous System Function ◽  
Gap Junctional

Oleamide is a sleep-inducing lipid originally isolated from the cerebrospinal fluid of sleep-deprived cats. Oleamide was found to potently and selectively inactivate gap junction–mediated communication between rat glial cells. In contrast, oleamide had no effect on mechanically stimulated calcium wave transmission in this same cell type. Other chemical compounds traditionally used as inhibitors of gap junctional communication, like heptanol and 18β-glycyrrhetinic acid, blocked not only gap junctional communication but also intercellular calcium signaling. Given the central role for intercellular small molecule and electrical signaling in central nervous system function, oleamide- induced inactivation of glial cell gap junction channels may serve to regulate communication between brain cells, and in doing so, may influence higher order neuronal events like sleep induction.

scholarly journals Differential phosphorylation of the gap junction protein connexin43 in junctional communication-competent and -deficient cell lines.

1990 ◽  
Vol 111 (5) ◽  
pp. 2077-2088 ◽  
Author(s):  
L S Musil ◽  
B A Cunningham ◽  
G M Edelman ◽  
D A Goodenough
Keyword(s):  
Cell Adhesion ◽  
Gap Junctions ◽  
Gap Junction ◽  
Cell Lines ◽  
Gap Junctional Communication ◽  
Competent Cells ◽  
Deficient Cell ◽  
Junctional Communication ◽  
Gap Junctional ◽  
Cell Cell

Connexin43 is a member of the highly homologous connexin family of gap junction proteins. We have studied how connexin monomers are assembled into functional gap junction plaques by examining the biosynthesis of connexin43 in cell types that differ greatly in their ability to form functional gap junctions. Using a combination of metabolic radiolabeling and immunoprecipitation, we have shown that connexin43 is synthesized in gap junctional communication-competent cells as a 42-kD protein that is efficiently converted to a approximately 46-kD species (connexin43-P2) by the posttranslational addition of phosphate. Surprisingly, certain cell lines severely deficient in gap junctional communication and known cell-cell adhesion molecules (S180 and L929 cells) also expressed 42-kD connexin43. Connexin43 in these communication-deficient cell lines was not, however, phosphorylated to the P2 form. Conversion of S180 cells to a communication-competent phenotype by transfection with a cDNA encoding the cell-cell adhesion molecule L-CAM induced phosphorylation of connexin43 to the P2 form; conversely, blocking junctional communication in ordinarily communication-competent cells inhibited connexin43-P2 formation. Immunohistochemical localization studies indicated that only communication-competent cells accumulated connexin43 in visible gap junction plaques. Together, these results establish a strong correlation between the ability of cells to process connexin43 to the P2 form and to produce functional gap junctions. Connexin43 phosphorylation may therefore play a functional role in gap junction assembly and/or activity.

Mechanisms for the coordination of intercellular calcium signaling in insulin-secreting cells

1997 ◽  
Vol 110 (4) ◽  
pp. 497-504 ◽  
Author(s):  
D. Cao ◽  
G. Lin ◽  
E.M. Westphale ◽  
E.C. Beyer ◽  
T.H. Steinberg
Keyword(s):  
Gap Junction ◽  
Islet Cells ◽  
Cytosolic Calcium ◽  
Calcium Waves ◽  
Gap Junctional Communication ◽  
Junctional Communication ◽  
Junction Protein ◽  
Insulin Secreting Cells ◽  
Gap Junction Protein ◽  
Gap Junctional

Insulin-mediated increases in cytosolic calcium are synchronized among the cells in a pancreatic islet, and result in pulsatile secretion of insulin. Pancreatic beta cells express the gap junction protein connexin43 and are functionally coupled, making gap junctional communication a likely mechanism for the synchronization of calcium transients among islet cells. To define the mechanism by which pancreatic islet cells coordinate calcium responses, we studied mechanically-induced intercellular calcium waves in the communication-deficient rat insulinoma cell line RINm5f, and in RINm5f cells transfected with the gap junction protein connexin43. Both RINm5f and RINm5f cells transfected with connexin43 propagated calcium waves that required release of calcium from intracellular stores, did not involve gap junctional communication, and appeared to be mediated by autocrine activity of secreted ATP acting on P2U purinergic receptors. Connexin43 transfectants also propagated calcium waves that required gap junctional communication and influx of extracellular calcium through voltage-gated calcium channels. Gap junction-dependent intercellular calcium waves were inhibited by preventing plasma membrane depolarization. These studies demonstrate two distinct pathways by which insulin-secreting cells can coordinate cytosolic calcium rises, and show that it is by ionic traffic that gap junctions synchronize calcium-dependent events in these cells.

scholarly journals Differential regulation of distinct types of gap junction channels by similar phosphorylating conditions.

1995 ◽  
Vol 6 (12) ◽  
pp. 1707-1719 ◽  
Author(s):  
B R Kwak ◽  
M M Hermans ◽  
H R De Jonge ◽  
S M Lohmann ◽  
H J Jongsma ◽  
...  
Keyword(s):  
Gap Junction ◽  
Single Channel ◽  
Cell Communication ◽  
Differential Regulation ◽  
Cell Coupling ◽  
Gap Junction Channels ◽  
Physiological Importance ◽  
Gap Junction Channel ◽  
Conductance States ◽  
Channel Properties

Studies on physiological modulation of intercellular communication mediated by protein kinases are often complicated by the fact that cells express multiple gap junction proteins (connexins; Cx). Changes in cell coupling can be masked by simultaneous opposite regulation of the gap junction channel types expressed. We have examined the effects of activators and inhibitors of protein kinase A (PKA), PKC, and PKG on permeability and single channel conductance of gap junction channels composed of Cx45, Cx43, or Cx26 subunits. To allow direct comparison between these Cx, SKHep1 cells, which endogenously express Cx45, were stably transfected with cDNAs coding for Cx43 or Cx26. Under control conditions, the distinct types of gap junction channels could be distinguished on the basis of their permeability and single channel properties. Under various phosphorylating conditions, these channels behaved differently. Whereas agonists/antagonist of PKA did not affect permeability and conductance of all gap junction channels, variable changes were observed under PKC stimulation. Cx45 channels exhibited an additional conductance state, the detection of the smaller conductance states of Cx43 channels was favored, and Cx26 channels were less often observed. In contrast to the other kinases, agonists/antagonist of PKG affected permeability and conductance of Cx43 gap junction channels only. Taken together, these results show that distinct types of gap junction channels are differentially regulated by similar phosphorylating conditions. This differential regulation may be of physiological importance during modulation of cell-to-cell communication of more complex cell systems.

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