G-protein-independent modulation of P-type calcium channels by mu-opioids in Purkinje neurons of rat.

P-type calcium channels play a key role in the synaptic transmission between mammalian central neurons since a major part of calcium entering pre-synaptic terminals is delivered via these channels. Using conventional whole-cell patch clamp techniques we have studied the effect of mu-opioids on P-type calcium channels in acutely isolated Purkinje neurons from rat cerebellum. The selective mu-opioid agonist DAMGO (10nM) produced a small, but consistent facilitation of current through P-type calcium channels (10+/-1%, n=27, p<0.001). The effect of DAMGO was rapid (less than 10s) and fully reversible. This effect was both concentration and voltage-dependent. The EC(50) for the effect of DAMGO was 1.3+/-0.4nM and the saturating concentration was 100nM. The endogenous selective agonist of mu-opioid receptors, endomorphin-1 demonstrated similar action. Intracellular perfusion of Purkinje neurons with GTPgammaS (0.5mM) or GDPbetaS (0.5mM), as well as strong depolarizing pre-pulses (+50mV), did not eliminate facilitatory action of DAMGO on P-channels indicating that this effect is not mediated by G-proteins. Furthermore, the effect of DAMGO was preserved in the presence of a non-specific inhibitor of PKA and PKC (H7, 10microM) inside the cell. DAMGO-induced facilitation of P-current was almost completely abolished by the selective mu-opioid antagonist CTOP (100nM). These observations indicate that mu-type opioid receptors modulate P-type calcium channels in Purkinje neurons via G-protein-independent mechanism.