Regulation of cardiac gap junction channel permeability and conductance by several phosphorylating conditions
Abstract Short term (15 min) effects of activators of protein kinase A (PKA), PKC and PKG on cardiac macroscopic (g j ) and single channel (&ggr; j ) gap junctional conductances were studied in pairs of neonatal rat cardiomyocytes. Under dual whole-cell voltage-clamp, PKC activation by 100 nM TPA increased g j by 16 ± 2% (mean ± S.E.M, n=9), 1.5 mM of the PKG activator 8-bromo-cGMP (8Br-cGMP) decreased g j by 26 ± 2% (n=4), whereas 1.5 mM of the PKA activator 8Br-cAMP did not affect g j (1 ± 5%, n=11). Single cardiac gap junction channel events, resolved in the presence of heptanol, indicated two &ggr; j sizes of 20 pS and 40–45 pS. Under control conditions, the larger events were most frequently observed. Whereas 8Br-cAMP did not change this distribution, TPA or 8Br-cGMP shifted the &ggr; j distribution to the lower sizes. Diffusion of 6-carboxyfluorescein (6-CF), a gap junction permeant tracer, from the injected cell to neighboring cells was studied on small clusters of neonatal rat cardiomyocytes. Under control conditions, 6-CF labeled 8.4 ± 0.4 cells (mean ± S.E.M, n=31). Whereas 8Br-cAMP did not change the extent of dye transfer (8.1 ± 0.5 cells, n=10), TPA restricted the diffusion of 6-CF to 2.2 ± 0.2 cells (n=30) and 8Br-cGMP to 3.5 ± 0.3 cells (n=10). This suggests that permeability and single channel conductance of Cx43 gap junction channels are parallel related. Altogether, these results point to the differential modulation of electrical and metabolic coupling of cardiac cells by various phosphorylating conditions.
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