Immunocytochemical analysis of connexin expression in the healthy and diseased cardiovascular system

Severs, Nicholas J.; Rothery, Stephen; Dupont, Emmanuel; Coppen, Steven R.; Yeh, Hung‐I; Ko, Yousang; Matsushita, T.; Kaba, Riyaz A.; Halliday, Deborah

doi:10.1002/1097-0029(20010201)52:3<301::aid-jemt1015>3.0.co;2-q

Cited by 204 publications

(146 citation statements)

References 93 publications

(102 reference statements)

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“…8,24,27 For each specimen, 10 randomly selected areas were examined. Mouse anti-rabbit immunoglobulin conjugated to CY3 (Chemicon) was used to visualize immunolabeled Cx43.…”

Section: Immunofluorescence Study and Confocal Microscopymentioning

confidence: 99%

“…Assisted by QWIN image analysis software (Leica), we measured (1) the proportion of total tissue area occupied by the Cx43 immunoreactive signal, as well as (2) the density (number/100 mm 2 ) and (3) the size of discrete clusters of immunoreactive signal, which we defined operationally as individual gap junctions. 7,24,27 …”

Section: Immunofluorescence Study and Confocal Microscopymentioning

confidence: 99%

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Downregulated myocardial connexin 43 and suppressed contractility in rabbits subjected to a cholesterol-enriched diet

Lin

Yeh

et al. 2005

Laboratory Investigation

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The effects of hypercholesterolemia on the myocardium per se include electrophysiological and mechanical alterations. Since gap junctions are essential in electromechanical coupling throughout the heart, we examined the correlation between the temporal expression of cardiac connexin 43 (Cx43), contractile function, and conduction velocity in cholesterol-fed rabbits. After a 12-week feeding period, serum cholesterol levels gradually increased (Po0.001). In contrast, expression of cardiomyocyte Cx43 protein progressively decreased (60% reduction at 12 weeks, Po0.001). Such a reduction was also demonstrated by immunoconfocal microscopy, which further showed redistribution of Cx43 gap junctions at the lateral cell membrane. The downregulation of Cx43 protein was associated with increased levels of Cx43 mRNA (3.5 -fold at 12 weeks, Po0.001) and phosphorylated c-Jun N-terminal kinase (three-fold at 12 weeks, P ¼ 0.001). Functionally, although fractional shortening of the left ventricle remained unchanged throughout the feeding protocol, the cholesterol-fed rabbits had a reduced cardiac cycle-dependent variation of integrated backscatters, a decreased mitral ring systolic velocity, and an increased modified Tei index (all Po0.001), all of which indicated impaired intrinsic myocardial contractility and attenuated ventricular systolic performance. In Langendorff-perfused hearts of cholesterol-fed rabbits, decreased conduction velocity was observed (Po0.005). Withdrawal of the cholesterol-enriched diet for 18 weeks restored the contractile parameters and Cx43 protein expression. These findings suggest that Cx43 is highly involved in the molecular mechanism of hypercholesterolemia-induced cardiac contractile dysfunction and dysrhythmias.

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“…8,24,27 For each specimen, 10 randomly selected areas were examined. Mouse anti-rabbit immunoglobulin conjugated to CY3 (Chemicon) was used to visualize immunolabeled Cx43.…”

Section: Immunofluorescence Study and Confocal Microscopymentioning

confidence: 99%

Section: Immunofluorescence Study and Confocal Microscopymentioning

confidence: 99%

Downregulated myocardial connexin 43 and suppressed contractility in rabbits subjected to a cholesterol-enriched diet

Lin

Yeh

et al. 2005

Laboratory Investigation

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“…Connexons are hexamers of four-pass membrane proteins called connexins (Cxs; Bruzzone et al, 1996;Kumar and Gilula, 1996). Once transported to the PM, GJ channels cluster into two-dimensional arrays termed plaques that can be composed of a few to many thousands of individual channels and vary from a few square nanometers to many square micrometers (Bruzzone et al, 1996;Falk, 2000a;Severs et al, 2001). GJ channels can open and close (gate) and physiological parameters, including intracellular pH, Ca 2ϩ concentration, and Cx phosphorylation, are known to modulate GJ channel gating and the extent of GJ-mediated intercellular coupling (Delmar et al, 2004;Lampe and Lau, 2004;Moreno, 2005).…”

Section: Introductionmentioning

confidence: 99%

Internalization of Large Double-Membrane Intercellular Vesicles by a Clathrin-dependent Endocytic Process

Piehl

Lehmann

Gumpert

et al. 2007

MBoC

156

228

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Beyond its well-documented role in vesicle endocytosis, clathrin has also been implicated in the internalization of large particles such as viruses, pathogenic bacteria, and even latex beads. We have discovered an additional clathrin-dependent endocytic process that results in the internalization of large, double-membrane vesicles at lateral membranes of cells that are coupled by gap junctions (GJs). GJ channels bridge apposing cell membranes to mediate the direct transfer of electrical currents and signaling molecules from cell to cell. Here, we report that entire GJ plaques, clusters of GJ channels, can be internalized to form large, double-membrane vesicles previously termed annular gap junctions (AGJs). These internalized AGJ vesicles subdivide into smaller vesicles that are degraded by endo/lysosomal pathways. Mechanistic analyses revealed that clathrin-dependent endocytosis machinery-components, including clathrin itself, the alternative clathrin-adaptor Dab2, dynamin, myosin-VI, and actin are involved in the internalization, inward movement, and degradation of these large, intercellular double-membrane vesicles. These findings contribute to the understanding of clathrin's numerous emerging functions. INTRODUCTIONThe role of clathrin in endocytosis is well documented. This protein forms a typical curved lattice around endocytic vesicles that are internalized at the plasma membrane (PM). In addition, the involvement of clathrin in several uncharacteristic endocytic processes has been reported, including the internalization of viruses, pathogenic bacteria, and large latex beads (Aggeler and Werb, 1982;Ehrlich et al., 2004;Rust et al., 2004;Veiga and Cossart, 2005). Here, we describe another function of the clathrin-dependent endocytic machinery that results in the internalization of large, doublemembrane vesicles at lateral PMs of cells that are coupled by gap junctions (GJs).GJs are ubiquitously distributed channels that connect the cytoplasms of two apposing cells each participating in this connection via a half channel termed a connexon to provide direct cell-to-cell communication. Connexons are hexamers of four-pass membrane proteins called connexins (Cxs;Bruzzone et al., 1996;Kumar and Gilula, 1996). Once transported to the PM, GJ channels cluster into two-dimensional arrays termed plaques that can be composed of a few to many thousands of individual channels and vary from a few square nanometers to many square micrometers (Bruzzone et al., 1996; Falk, 2000a;Severs et al., 2001). GJ channels can open and close (gate) and physiological parameters, including intracellular pH, Ca 2ϩ concentration, and Cx phosphorylation, are known to modulate GJ channel gating and the extent of GJ-mediated intercellular coupling (Delmar et al., 2004;Lampe and Lau, 2004;Moreno, 2005). However, the extent of intercellular coupling could also be regulated through altering the number of GJ channels in the PM.Cxs have a surprisingly short half-life of only 1-5 h, leading to a rapid GJ and Cx protein turnover (Fallon and Goodeno...

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“…In the heart, at least the connexins Cx30.2, Cx37, Cx40, Cx43, and Cx45 are expressed, each with a different contribution pattern and abundance in the different cells of the heart (Severs et al 2001;Duffy et al 2006). Thus Cx43 and Cx40 are the most abundant cardiac connexins.…”

Section: Introductionmentioning

confidence: 99%

Neonatal Rat Cardiomyocytes Show Characteristics of Nonhomotypic Gap Junction Channels

Schulte

Scheffler

Rojas-Gómez

et al. 2008

Cell Communication & Adhesion

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Neonatal rat cardiomyocytes mainly coexpress the connexins Cx40, Cx43, and to a small amount Cx45, leading to potential formation of mixed (heteromeric/heterotypic) gap junction channels. Using the dual-voltage clamp technique with switching clamp circuits, the authors investigated voltage sensitivity of gap junction channels between cell pairs of Cx40, Cx43, and Cx45 stably transfected HeLa cells and compared those data to data obtained from cell pairs of cultured neonatal rat cardiomyocytes. In accordance to previously published data, the relationship between normalized conductance and transjunctional voltage (g/V j ) was quasisymmetrical for the transfected HeLa cells, indicating homotypic gap junction channels. Boltzmann curves fitted to data obtained from neonatal rat cardiomyocyte pairs expressing both Cx40 and Cx43 showed an asymmetrical inactivation pattern, which cannot be explained by the presence of pure populations of homotypic gap junction channels of either isoform. In conclusion the authors assume the additional presence of heterotypic and possibly even heteromeric gap junction channels in neonatal rat cardiomyocytes.

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Immunocytochemical analysis of connexin expression in the healthy and diseased cardiovascular system

Cited by 204 publications

References 93 publications

Downregulated myocardial connexin 43 and suppressed contractility in rabbits subjected to a cholesterol-enriched diet

Downregulated myocardial connexin 43 and suppressed contractility in rabbits subjected to a cholesterol-enriched diet

Internalization of Large Double-Membrane Intercellular Vesicles by a Clathrin-dependent Endocytic Process

Neonatal Rat Cardiomyocytes Show Characteristics of Nonhomotypic Gap Junction Channels

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