MODULATION OF CA2-PROTEIN-COUPLED RECEPTORS IN SYMPATHETIC NEURONS OFMALE-RAT PELVIC GANGLIA( CURRENTS BY VARIOUS G)

The modulation of voltage-gated calcium (Ca2+) channels by various G p rotein-coupled receptor pathways was investigated in sympathetic neuro ns of the male rat major pelvic ganglion (MPG). Standard whole cell pa tch-clamp recording techniques were used to record Ca2+ currents from acutely dissociated neurons. The activation of muscarinic receptors, w hich uses a G protein pathway that was not blocked by either pertussis toxin (PTX) or cholera toxin (CTX), inhibited both N-type and L-type Ca2+ channels. The activation of alpha(2) noradrenergic receptors with the selective agonist UK14304, which used primarily a PTX-sensitive G protein pathway, inhibited only N-type Ca2+ channels. The activation of vasoactive intestinal polypeptide (VIP) receptors, which used a CTX -sensitive G protein pathway, also inhibited only N-type Ca2+ channels . UK14304 and VIP induced a bell-shaped inhibition of the Ca2+ current with a peak inhibition at around +10 mV and decreasing inhibition at more positive potentials. In contrast, the muscarine-induced Ca2+ curr ent inhibition was not bell shaped and was more prominent at more posi tive potentials. Furthermore, a large depolarization, which relieved t he current inhibition by UK14304 and VIP, did not relieve the inhibiti on by muscarine. Besides inhibiting the Ca2+ current, UK14304 and VIP also slowed the activation kinetics, an effect not seen with muscarine . Replacing external Ca2+ with Ba2+ and replacing internal ethylene gl ycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid(EGTA) with high bis-(o-aminophenoxy)-N,N,N',N'-tetraacetic acid (BAPTA) completel y blocked the inhibitory effect of muscarine. However, the inhibitory effects of both UK14304 and VIP were unaffected under these conditions . Surprisingly, the facilitation of the Ca2+ current was eliminated un der these strong calcium-buffering conditions. The activation of prote in kinase C (PKC) with phorbol 12-myristate 13-acetate (PMA) increases the amplitude of the Ca2+ current, diminishes facilitation, and reduc es the inhibition of this current by UK14304 and VIP. However, PKC act ivation did not reduce the muscarine-induced Ca2+ current inhibition. in summary, our data suggest that muscarine uses a mechanism different from UK14304 and VIP to modulate the N-type Ca2+ channels in sympathe tic neurons of the MPG. Although VIP and UK14304 use different G prote in pathways, these two different pathways most likely converge downstr eam to compete for the same target site on the N-type Ca2+ channels.