Voltage-dependent acceleration of Ca(v)1.2 channel current decay by (+)- and (-)-isradipine.
Inhibition of Ca(v)1.2 by antagonist 1,4 dihydropyridines (DHPs) is associated with a drug-induced acceleration of the calcium (Ca(2+)) channel current decay. This feature is contradictorily interpreted as open channel block or as drug-induced inactivation. To elucidate the underlying molecular mechanism we investigated the effects of (+)- and (-)-isradipine on Ca(v)1.2 inactivation gating at different membrane potentials. alpha(1)1.2 Constructs were expressed together with alpha(2)-delta- and beta(1a)- subunits in Xenopus oocytes and drug-induced changes in barium current (I(Ba)) kinetics analysed with the two microelectrode voltage clamp technique. To study isradipine effects on I(Ba) decay without contamination by intrinsic inactivation we expressed a mutant (V1504A) lacking fast voltage-dependent inactivation. At a subthreshold potential of -30 mV a 200-times higher concentration of (-)-isradipine was required to induce a comparable amount of inactivation as by (+)-isradipine. At +20 mV the two enantiomers were equally efficient in accelerating the I(Ba) decay. Faster recovery from (-)- than from (+)-isradipine-induced inactivation at -80 mV in a Ca(v)1.2 construct (tau((-)-isr.(Cav1.2))=0.74 s<tau((+)-isr.(Cav1.2))=2.85 s) and even more rapid recovery of V1504A (tau((-)-isr.(V1504A))=0.39 s<tau((+)-isr.(V1504A))=1.98 s) indicated that drug-induced determinants and determinants of intrinsic inactivation (V1504) stabilize the DHP-induced channel conformation in an additive manner. In the voltage range between -25 and 20 mV where the channels inactivate predominantly from the open state the (+)- and (-)-isradipine-induced acceleration of the I(Ba) decay in V1504A displayed similar voltage-dependence as intrinsic fast inactivation of Ca(v)1.2. Our data suggest that the isradipine-induced acceleration of the Ca(v)1.2 current decay reflects enhanced fast voltage-dependent inactivation and not open channel block.
Berjukow
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Institut für Biochemische Pharmakologie, Peter-Mayr-Strasse 1, A-6020 Innsbruck, Austria.
Hering
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D019943:Amino Acid Substitution; D000818:Animals; D002121:Calcium Channel Blockers / Q000494:pharmacology*; D020746:Calcium Channels, L-Type / Q000235:genetics / Q000502:physiology*; D004305:Dose-Response Relationship, Drug; D004558:Electric Stimulation; D017275:Isradipine / Q000737:chemistry / Q000494:pharmacology*; D008564:Membrane Potentials / Q000187:drug effects*; D009154:Mutation; D009865:Oocytes; D021122:Protein Subunits; D011817:Rabbits; D013237:Stereoisomerism; D013997:Time Factors; D014981:Xenopus
D002121:Calcium Channel Blockers; D020746:Calcium Channels, L-Type; D021122:Protein Subunits; D017275:Isradipine