Characterization of the binding of [ 3H]SR 33805 to the slow Ca 2+ channel in rat heart sarcolemmal membrane

Abstract

SR 33805 is a representative of a new class of compounds (indole sulfone) that inhibits L-type Ca 2+ channels. [ 3H]SR 33805 has been shown to bind with a high affinity ( K d ∼ 20 pM calculated from saturation isotherms and association/dissociation kinetics) to a single site in a purified preparation of rat cardiac sarcolemmal membranes. The binding was found to be saturable and reversible. The maximal binding capacity was in approximately 1:1 stoichiometry with that of other Ca 2+ channel antagonists. Various cations (Na +, Ca 2+, Cd 2+, and La 3+) were shown to inhibit specific [ 3H]SR 33805 binding, with La 3+ being the most potent. Using a range of receptor or channel ligands (including ω-conotoxin and Na + and K + channel modulators), only the L-type Ca 2+ channel antagonists were found to displace [ 3H]SR 33805. However, dihydropyridines, phenylalkylamines, benzothiazepines, and diphenyl-butylpiperidines were found to inhibit [ 3H]SR 33805 in a non-competitive manner as demonstrated by displacement experiments in addition to dissociation kinetics. In contrast, the interaction of SR 33805 with fantofarone has been found to be competitive. Binding of [ 3H]SR 33805 (and [ 3H]fantofarone) is entropy driven as opposed to that of the [ 3H]nitrendipine which is enthalpy driven. From these results we suggest that SR 33805 binds with a high affinity to a unique site on the L-type Ca 2+ channel found in rat cardiac sarcolemmal membranes. This site is equivalent to that of fantofarone and in allosteric interaction with that of the dihydropyridines, phenylalkylamines and benzothiazepines.