D-2 dopamine receptors reduce N-type Ca2+ currents in rat neostriatal cholinergic interneurons through a membrane-delimited, protein-kinase-C-insensitive pathway

Dopamine has long been known to regulate the activity of striatal cholinergic interneurons and the release of acetylcholine. Yet, the cellular mechanisms by which this regulation occurs have not been elucidated. One way in which dopamine might act is by modulating voltage-dependent Ca2+ channels. To test this hypothesis, the impact of dopaminergic agonists on Ca2+ channels in neostriatal cholinergic interneurons was studied by combined whole cell voltage-clamp recording and single-cell reverse transcription-polymerase chain reactions. Cholinergic interneurons were identified by the presence of choline acetyltransferase mRNA. Nearly all interneurons tested (90%, n = 17) coexpressed D-2 (short and long isoforms) and D-1b (D-5) dopamine receptor mRNAs. D-1a receptor mRNA was found in only a small subset (20%) of the sample and D-3 and D-4 receptor mRNAs were undetectable. D-2 receptor agonists rapidly and reversibly reduced N-type Ca2+ currents. D-1b/D-1a receptor activation had little or no effect on Ca2+ currents. The D-2 receptor antagonist sulpiride blocked the effect of D-2 agonists. Dialysis with guanosine-5'-O-(2-thiodiphosphate) or brief exposure to the G protein (G(i/o)) alkylating agent N-ethylmaleimide also blocked the D-2 modulation. The reduction in N-type currents was neither accompanied by kinetic slowing nor significantly reversed by depolarizing prepulses. The D-2 receptor effects were mediated by a membrane-delimited pathway, because the modulation was not seen in cell-attached patches when agonist was applied to the bath and was not disrupted by perturbations in cytosolic signaling pathways known to be linked to D-2 receptors. Activation of M2 muscarinic receptors occluded the D-2 modulation, suggesting a shared signaling element. However, activation of protein kinase C attenuated the M2 modulation without significantly affecting the D-2 modulation. Taken together, our results suggest that activation of D-2 dopamine receptors in cholinergic interneurons reduces N-type Ca2+ currents via a membrane-delimited, Gi,, class G protein pathway that is not regulated by protein kinase C. This signaling pathway may underlie the ability of D-2 receptors to reduce striatal acetylcholine release.