Ion Channel Regulation By Signal Transduction Pathways

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We have focused on three specific ion channels: CaV1.2, BKca, GIRK. CaV1.2 channels show modal gating that is regulated by dihydropyridine, protein phosphorylatiion, and calcium. We have obtained new evidence from single channel recordings that dihydropyridines modulate channel activity by altering their susceptibility to phosphorylation. Furthermore, calcium inhibits channel activity through dephosphorylation of the channel protein. The BKca potassium channels, which are encoded by the KCNMA1 gene (slo) and regulate action potential duration in endocrine cells and nerve terminals. Genetic studies of ethanol action on locomotor coordination in invertebrate model organisms have identified BKca channels as a prominent target of ethanol action. Other studies have identified the Gs-cAMP-PKA signaling system as a target of ethanol action. Therefore, we have studied the mechanism of BKca regulation by ethanol in a model genetic organism, C. elegans. We identified the PKA site on ceBKca; when it was mutated the worms were impervious to ethanol up t o 500 mM ! Inhibition of the Ser/Thr protein phosphatase PP1 by targeted expression of a mammalian inhibitor protein in ce neurons also prevented ethanol from disrupting locomotion. We have confirmed this dependence of ethanol action on PP1 activity in rat hippocampal slices, in which ethanol inhibits LTP on CA1 pyramidal dendrites. In collaboration with Dr. Birnbaumer's group, TRPC channels have been implicated in calcium entry in response to signaling through Gq coupled receptors. GIRK channels regulate excitability of neurons, endocrine cells and cardiac myocytes. We have investigated GIRK channel regulation by G proteins, phospholipids, and protein phosphorylation using concatenated recombinant channels at the single channel level to avoid confusion by changes in expression. In particular we have used somatostatin stimulation of GIRK channel activity to investigate the effector for Go.