Article

Connexin40 and connexin43 determine gating properties of atrial gap junction channels

Department of Pharmacology, SUNY Upstate Medical University, 750 East Adams Street, Syracuse, NY 13210, USA.
Journal of Molecular and Cellular Cardiology (Impact Factor: 5.22). 05/2009; 48(1):238-45. DOI: 10.1016/j.yjmcc.2009.05.014
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ABSTRACT While ventricular gap junctions contain only Cx43, atrial gap junctions contain both Cx40 and Cx43; yet the functional consequences of this co-expression remain poorly understood. We quantitated the expression of Cx40 and Cx43 and their contributions to atrial gap junctional conductance (g(j)). Neonatal murine atrial myocytes showed similar abundances of Cx40 and Cx43 proteins, while ventricular myocytes contained at least 20 times more Cx43 than Cx40. Since Cx40 gap junction channels are blocked by 2 mM spermine while Cx43 channels are unaffected, we used spermine block as a functional dual whole cell patch clamp assay to determine Cx40 contributions to cardiac g(j). Slightly more than half of atrial g(j) and <or=20% of ventricular g(j) were inhibited. In myocytes from Cx40 null mice, the inhibition of ventricular g(j) was completely abolished, and the block of atrial g(j) was reduced to <20%. Compared to ventricular gap junctions, the transjunctional voltage (V(j))-dependent inactivation of atrial g(j) was reduced and kinetically slowed, while the V(j)-dependence of fast and slow inactivation was unchanged. We conclude that Cx40 and Cx43 are equally abundant in atrium and make similar contributions to atrial g(j). Co-expression of Cx40 accounts for most, but not all, of the differences in the V(j)-dependent gating properties between atrium and ventricle that may play a role in the genesis of slow myocardial conduction and arrhythmias.

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    • "Cell homogenates were prepared 48 h after transfection with connexin DNA as described by Gong et al. [15]. Immunoblots were performed as described earlier [5]. The protein concentrations of homogenates were determined using the method of Bradford (Bio-Rad, Richmond, CA) [16]. "
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    • "For double-labeling experiments, cells were incubated simultaneously with both mouse anti-mCherry monoclonal and rabbit anti-GFP antibodies (Abcam). Immunoblots were performed as described earlier [10] using cell homogenates prepared 48 h after transfection with connexin DNA as described by Gong et al. [11]. For immunofluorescence microscopy, cultured cells were fixed in methanol/acetone (1:1), stained as previously described [8], and imaged using a Zeiss Axioplan 2 microscope or Leica TCS SP2 laser-scanning confocal microscope. "
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    • "Cx37, Cx40, and Cx43 are abundant components of the gap junctions in various cells of the cardiovascular system, and they are coexpressed in some of these cells. In ventricular myocytes , Cx43 is the predominant connexin; however, in atrial myocytes, the abundances of Cx40 and Cx43 are approximately equal (Lin et al. 2010). In diseased myocardium, gap junctions may undergo remodeling, and the levels of Cx40 and Cx43 may be altered (reviewed by Severs et al. 2008). "
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