Desmethoxyverapamil-sensitive calcium channels in rat portal vein smooth muscle

Abstract

The whole-cell patch clamp technique was used to analyze the properties of the phenylalkylamine-sensitive calcium channels in smooth muscle cells isolated from the portal vein. (−)-D888 dose dependently inhibited the calcium current elicited from a holding potential of −40 mV (IC 50 = 1.3 nM) in a frequency-dependent manner. No voltage dependence of the inhibition was noted. Independent high- and low-affinity binding sites for (−)-[ 3H]D888 were identified. Calcium entry blockers such as (−)-D888, d-cis-diltiazem and nicardipine completely or partially antagonized the (−)-[ 3H]D888 binding at both types of sites. The properties of this cross-inhibition suggest that phenylalkylamines and d-cis-diltiazem bind at common sites in vascular smooth muscles whereas dihydropyridines bind at distinct sites which are allosterically coupled to the phenylalkylamine sites. As the IC 50 for (−)-D888 found from electrophysiological experiments is not identical to the equilibrium dissociation constants for the high- and low-affinity sites found from binding data (0.47 and 50 nM, respectively), it is suggested that binding of (−)-D888 to both high- and low-affinity sites may be involved in the inhibitory effect of (−)-D888 on calcium channels. Furthermore, these two different binding sites may correspond to two different subtypes of phenylalkylamine-sensitive calcium channels in smooth muscle cells.