Restricted distribution of connexin40, a gap junctional protein, in mammalian heart.

Abstract

Connexin40 (Cx40) is a member of the connexin family of gap junction proteins. Its mRNA, abundant in lung, is also present in mammalian heart, although in lower amount. Rabbit antipeptide antibodies directed to the COOH terminus (residues 335 to 356) of rat Cx40 were characterized to investigate the distribution of Cx40 in rat and guinea pig cardiac tissues. The affinity-purified antibodies detect specifically a major protein (Mr, 40000) in immunoblots of total extracts from rat lung and rat and guinea pig heart. In sections of guinea pig atrial tissue treated for immunofluorescence, a strong labeling associated with myocytes was seen with a distribution consistent with that of intercalated disks. The results of immunoelectron microscopy carried out with guinea pig atrial tissue showed that epitopes recognized by these antibodies were exclusively associated with gap junctions. These results, added to those of control experiments, demonstrate that antibodies 335-356 are specific for Cx40. Doublelabeling experiments carried out with lung sections using anti-factor VIII and anti-Cx40 antibodies suggest that Cx40 is expressed in blood vessel endothelial cells. In guinea pig and rat heart sections, investigated using both immunofluorescence and immunoperoxidase techniques, a signal was also found to be associated with vascular walls. In guinea pig heart, only atrial myocytes are Cx4O-positive. No labeling was detected in ventricular myocytes, including those of the His bundle and the bundle branches, which otherwise do express connexin43 (Cx43). In rat heart Cx4O -expressing myocytes are localized in the conduction system, ie, the His bundle, the bundle branches, and the Purkinje fibers. Cx43 is not detected either in the His bundle or in the proximal parts of the bundle branches, and consequently, Cx4O is the first connexin demonstrated in this region of the rat conduction system. Cx40 was not detected in the working ventricular myocytes. Doublelabeling experiments carried out with hen anti-Cx43 antibodies and rabbit anti-Cx4O antibodies demonstrated that, in tissues expressing both Cx43 and Cx4O, these two connexins were localized in the same immunoreactive sites. A few sites, however, appear to contain only one or the other of these two connexins. (Circ Res. 1994;74:839-851

Developmental regulation of connexin 40 gene expression in mouse heart correlates with the differentiation of the conduction system

“…In adult mouse and rat hearts CX40 was shown to be expressed in the His bundle and the bundle branches (Gros et al, 1994. The present study demonstrates that CX40 is already expressed at 14 dpc in the early differentiating bundle branches on both sides of the growing interventricular septum.…”

“…In addition, in the adult rat and mouse, CX43 is also absent from the ventricular conduction tissue, i.e., the His bundle and the proximal part of the bundle branches (Van Kempen et al, 1991;Gourdie et al, 1992). Interestingly, CX40 distribution studies (Gourdie et al, 1993;Gros et al, 1994Gros et al, , 1995Van Kempen et al, 1995) showed that CX40 has a more restricted distribution than CX43 in adult mammalian hearts. These authors demonstrated that (2x40 is expressed not only in atria (depending on the species) but also in ventricular conduction system myocytes.…”

Incompatibility of connexin 40 and 43 Hemichannels in gap junctions between mammalian cells is determined by intracellular domains.

“…The difference in yi(main state) between Cx4O and Cx4O*43El,2 channels was surprising, since both connexins presumably possessed the same four transmembrane domains Ml-M4. In the case of heterotypic (Bruzzone et al, 1993) and human HeLa cells (Elfgang et al, 1995 surrounding working myocardium expresses Cx43 (Bastide et al, 1993;Gros et al, 1994 (Elfgang et al, 1995 Figure 3C). Previously, it had been shown that each immunofluorescent signal on contacting membranes of primary hepatocytes coincided with a gap junctional plaque (Dermietzel et al, 1987).…”

“…This functional incompatibility is likely to contribute to the preferential propagation of action potentials along the bundles of murine conductive myocardiocytes (expressing Cx4O) which are surrounded by the working myocardium (expressing Cx43; Bastide et al, 1993;Gros et al, 1994). As yet, the molecular determinants causing the incompatibility between Cx4O and Cx43 hemichannels were not known.…”

Chemistry, Physiology, and Pharmacology of β.-Adrenergic Mechanisms in the Heart. Why are .β-Blocker Antiarrhythmics Superior?

“…Cx40, Cx43 and Cx45 are the most abundant connexin isoforms of the working mammalian myocardial cells and the conduction system. Cx40 is mainly expressed in the atrium and in the conduction system [44][45][46][47][48][49], while Cx43 can be found both in atrial and ventricular cells but not in sinoatrial and atrioventricular nodes [46,[48][49][50][51][52][53][54][55]. Cx45 is preferentially expressed in sinoatrial and atrioventricular nodes, His-bundle and bundle branches [56][57][58].…”