MIBEFRADIL (RO-40-5967) BLOCKS MULTIPLE TYPES OF VOLTAGE-GATED CALCIUM CHANNELS IN CULTURED RAT SPINAL MOTONEURONS

The actions of the novel calcium (Ca2+) channel antagonist mibefradil (Ro 40-5967), a selective T-type channel blocker in myocardium, were i nvestigated in embryonic rat spinal motoneurones maintained in culture . Whole-cell currents were recorded with the patch-clamp technique. Mo toneurones displayed transient, low-voltage-activated (LVA) and, more sustained, high-voltage-activated (HVA) Ca2+ currents. The LVA current s were small and preferentially blocked by amiloride and low doses of nickel. Most of the HVA Ca2+ current flowed through N-type Ca2+ channe ls, while L-, and P/Q-type channels represented a smaller fraction. Mi befradil caused a rapid and reversible dose-dependent block of inward Ca2+ channel currents. Inhibition was nearly complete at 10 mu M, sugg esting mibefradil blockade of all subclasses of Ca2+ channels. The IC5 0 was approximately 1.4 mu M on currents measured at 0 mV, from a hold ing potential of -90 mV. Inhibition of LVA Ca2+ current occurred over the same concentration range. Slow tail currents induced by the dihydr opyridine agonist Bay K 8644 were also blocked by mibefradil, although with a slightly lower potency (IC50 = 3.4 mu M). These broad inhibito ry effects of mibefradil on Ca2+ influx were also supported by the str ong inhibition of depolarization-induced intracellular calcium transie nts, measured from Indo-1 loaded motoneurones imaged with confocal mic roscopy. We conclude that mibefradil has potent blocking effects on Ca 2+ channels in mammalian motoneurones. We hypothesize that therapeutic and pharmacological effects of mibefradil may involve actions on Ca2 channels other than type T.