Cu2+, Co2+, and Mn2+ modify the gating kinetics of high-voltage-activated Ca2+ channels in rat palaeocortical neurons

The effects of three divalent metal cations (Mn2+, Co2+, and Cu2+) on high-voltage-activated (HVA) Ca2+ currents were studied in acutely dissociated pyramidal neurons of rat piriform cortex using the patch-clamp technique. Cu2+, Mn2+, and Co2+ blocked HVA currents conducted by Ba2+ ( IBa) with IC50 of approximately 920 nM, approximately 58 micro M, and approximately 65 micro M, respectively. Additionally, after application of non-saturating concentrations of the three cations, residual currents activated with substantially slower kinetics than control IBa. As a consequence, the current fraction abolished by the blocking cations typically displayed, in its early phase, an unusually fast-decaying transient. The latter phenomenon turned out to be a subtraction artifact, since none of the pharmacological components (L-, N-, P/Q-, and R-type) that constitute the total HVA currents under study showed a similarly fast early decay: hence, the slow activation kinetics of residual currents was not due to the preferential inhibition of a fast-activating/inactivating component, but rather to a true slowing effect of the blocker cations. The percent IBa-amplitude inhibition caused by Mn2+, Co2+, and Cu2+ was voltage-independent over the whole potential range explored (up to +30 mV), hence the slowing of IBa activation kinetics was not due to a mechanism of voltage- and time-dependent relief from block. Moreover, Mn2+, Co2+, and Cu2+ significantly reduced I(Ba) deactivation speed upon repolarization, which also is not compatible with a depolarization-dependent unblocking mechanism. The above results show that 1) Cu2+ is a particularly potent HVA Ca2+-channel blocker in rat palaeocortical neurons; and 2) Mn2+, Co2+, and Cu2+, besides exerting a blocking action on HVA Ca2+-channels, also modify Ca2+-current activation and deactivation kinetics, most probably by directly interfering with channel-state transitions.