Endogenous regulator of G-protein signaling proteins modify N-type calcium channel modulation in rat sympathetic neurons.
Experiments using heterologous overexpression indicate that regulator of G-protein signaling (RGS) proteins play important roles in Gbetagamma-mediated ion channel modulation. However, the roles subserved by endogenous RGS proteins have not been extensively examined because tools for functionally inhibiting natively expressed RGS proteins are lacking. To address this void, we used a strategy in which Galpha(oA) was rendered insensitive to pertussis toxin (PTX) and RGS proteins by site-directed mutagenesis. Either PTX-insensitive (PTX-i) or both PTX- and RGS-insensitive (PTX/RGS-i) mutants of Galpha(oA) were expressed along with Gbeta(1) and Ggamma(2) subunits in rat sympathetic neurons. After overnight treatment with PTX to suppress natively expressed Galpha subunits, voltage-dependent Ca(2+) current inhibition by norepinephrine (NE) (10 microm) was reconstituted in neurons expressing either PTX-i or PTX/RGS-i Galpha(oA). When compared with neurons expressing PTX-i Galpha(oA), the steady-state concentration-response relationships for NE-induced Ca(2+) current inhibition were shifted to lower concentrations in neurons expressing PTX/RGS-i Galpha(oA). In addition to an increase in agonist potency, the expression of PTX/RGS-i Galpha(oA) dramatically retarded the current recovery after agonist removal. Interestingly, the alteration in current recovery was accompanied by a slowing in the onset of current inhibition. Together, our data suggest that endogenous RGS proteins contribute to membrane-delimited Ca(2+) channel modulation by regulating agonist potency and kinetics of G-protein-mediated signaling in neuronal cells.
Jeong
S W
SW
Laboratory of Molecular Physiology, Guthrie Research Institute, Sayre, Pennsylvania 18840, USA.
Ikeda
S R
SR
D000818:Animals; D020864:Calcium Channels, N-Type / Q000187:drug effects / Q000502:physiology*; D002478:Cells, Cultured; D020962:Heterotrimeric GTP-Binding Proteins / Q000235:genetics / Q000502:physiology*; D008297:Male; D009474:Neurons / Q000187:drug effects / Q000502:physiology*; D009638:Norepinephrine / Q000494:pharmacology*; D037342:Pertussis Toxin; D020710:RGS Proteins / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D015398:Signal Transduction; D017783:Superior Cervical Ganglion / Q000166:cytology / Q000502:physiology*; D010566:Virulence Factors, Bordetella / Q000494:pharmacology*
D020864:Calcium Channels, N-Type; D020710:RGS Proteins; D010566:Virulence Factors, Bordetella; D037342:Pertussis Toxin; D020962:Heterotrimeric GTP-Binding Proteins; D009638:Norepinephrine