Dihydropyridine enantiomers block recombinant L-type Ca2+ channels by two different mechanisms

1. The molecular basis of the state-dependent block of L-type Ca2+ channels by dihydropyridines is still poorly understood. Therefore, we studied the enantioselectivity of Ca2+ channel block by isradipine enantiomers at three holding potentials (-80, -80 and -40 mV) in Chinese hamster ovary (CHO) ce lls stably expressing the rabbit lung alpha(1C-b)-subunit. 2. The extent of enantioselectivity did not markedly change with the holdin g potential (IC50 ratios of 104-138), whereas the potency of both isradipin e enantiomers increased with depolarisation of the holding potential. 3. In addition to its block of the peak Ca2+ channel current, I-peak, (-)-i sradipine inhibited the relative current at the end of the test pulse, the so-called I-late, normalised to I-peak (I-late/I-peak). This effect was una ffected by the holding potential and revealed distinct kinetics compared to the development of conventional block of I-peak. 4. When these effects were studied using an alpha(1C-b)-mutant lacking the high-affinity dihydropyridine binding site, expressed in human embryonic ki dney (HEK 293) cells, both enantiomers blocked I-late/I-peak to a similar d egree. 5. Our data are discussed within the framework of the 'guarded receptor' an d the 'modulated receptor' hypotheses. The very different properties of the block of I-late/I-peak compared to those of the conventional high-affinity block of I-peak suggest the existence of an additional mechanism possibly mediated via a second, distinct binding site.