Phosphorylation-dependent regulation of nonselective cation channels in guinea pig chromaffin cells

In guinea pig chromaffin cells, acetylcholine induces a nonselective cation current by activating a muscarinic receptor, probably m4. We investigated activation and deactivation processes of this current. The calmodulin antagonists trifluoperazine and calmidazolium reversibly suppressed the muscarinic activation of the nonselective cation current in a dose-dependent manner; the former was about six times less potent than the latter. H-7, a kinase inhibitor, also reversibly inhibited the muscarinic current with a concentration eliciting 50% of maximal inhibition of 277 microM. Internal application of vanadate, a nonspecific phosphatase inhibitor, enhanced the muscarinic current and markedly slowed the time course of its deactivation, but the phosphatase 1 and 2A inhibitors okadaic acid and calyculin A had no effects. Lowering Mg2+ concentration in the patch solution mimicked the effects of vanadate on the muscarinic current. Internal dialysis with vanadate or low-Mg2+ solution gradually led to development of an inward current, and the related voltage dependence was similar to that seen with the muscarinic cation current. These results suggest that kinase and Mg(2+)-dependent phosphatase are responsible for activation and deactivation of nonselective cation channels. The channels seem to be under the influence of both enzymes, even in the absence of external signals.