Muscarinic stimulation of alpha1E Ca channels is selectively blocked by the effector antagonist function of RGS2 and phospholipase C-beta1.
Neuronal alpha1E Ca channel subunits are widely expressed in mammalian brain, where they are thought to form R-type Ca channels. Recent studies have demonstrated that R-type channels contribute to neurosecretion and dendritic Ca influx, but little is known concerning their modulation. Here we show that alpha1E channels are strongly stimulated, and only weakly inhibited, through M1 muscarinic acetylcholine receptors. Both forms of channel modulation are mediated by pertussis toxin-insensitive G-proteins. Channel stimulation is blocked by regulator of G-protein signaling 2 (RGS2) or the C-terminal region of phospholipase C-beta1 (PLCbeta1ct), which have been previously shown to function as GTPase-activating proteins for Galphaq. In contrast, RGS2 and PLCbeta1ct do not block inhibition of alpha1E through M1 receptors. Inhibition is prevented, however, by the C-terminal region of beta-adrenergic receptor kinase 1, which sequesters Gbetagamma dimers. Thus, stimulation of alpha1E is mediated by a pertussis toxin-insensitive Galpha subunit (e.g., Galphaq), whereas inhibition is mediated by Gbetagamma. The ability of RGS2 and PLCbeta1ct to selectively block stimulation indicates these proteins functioned primarily as effector antagonists. In support of this interpretation, RGS2 prevented stimulation of alpha1E with non-hydrolyzable guanosine 5'-0-(3-thiotriphosphate). We also report strong muscarinic stimulation of rbE-II, a variant alpha1E Ca channel that is insensitive to voltage-dependent inhibition. Our results predict that Galphaq-coupled receptors predominantly stimulate native R-type Ca channels. Receptor-mediated enhancement of R-type Ca currents may have important consequences for neurosecretion, dendritic excitability, gene expression, or other neuronal functions.
Melliti
K
K
Department of Biology, Utah State University, Logan, Utah 84322-5305, USA.
Meza
U
U
Adams
B
B
D000818:Animals; D001921:Brain / Q000378:metabolism; D002120:Calcium Channel Agonists / Q000494:pharmacology; D002121:Calcium Channel Blockers / Q000494:pharmacology*; D015220:Calcium Channels / Q000235:genetics / Q000378:metabolism*; D020908:Calcium Channels, R-Type; D002217:Carbachol / Q000494:pharmacology; D027682:Cation Transport Proteins; D002460:Cell Line; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000037:antagonists & inhibitors / Q000494:pharmacology; D006801:Humans; D017136:Ion Transport / Q000187:drug effects; D007527:Isoenzymes / Q000378:metabolism*; D018721:Muscarinic Agonists / Q000494:pharmacology*; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism; D009474:Neurons / Q000378:metabolism*; D018408:Patch-Clamp Techniques; D037342:Pertussis Toxin; D054799:Phospholipase C beta; D020710:RGS Proteins / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D011817:Rabbits; D051381:Rats; D043643:Receptor, Muscarinic M1; D011976:Receptors, Muscarinic / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D019311:Staurosporine / Q000494:pharmacology; D014162:Transfection; D010738:Type C Phospholipases / Q000378:metabolism*; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
C498917:Cacna1e protein, rat; D002120:Calcium Channel Agonists; D002121:Calcium Channel Blockers; D015220:Calcium Channels; D020908:Calcium Channels, R-Type; D027682:Cation Transport Proteins; D004791:Enzyme Inhibitors; D007527:Isoenzymes; D018721:Muscarinic Agonists; D009419:Nerve Tissue Proteins; D020710:RGS Proteins; C116924:RGS8 protein, human; D043643:Receptor, Muscarinic M1; D011976:Receptors, Muscarinic; C105345:Rgs2 protein, mouse; C486485:Rgs8 protein, rat; D010566:Virulence Factors, Bordetella; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D002217:Carbachol; D037342:Pertussis Toxin; D010738:Type C Phospholipases; C518033:PLCB1 protein, human; D054799:Phospholipase C beta; C518034:Plcb1 protein, mouse; C518035:Plcb1 protein, rat; D019311:Staurosporine