Effect of G protein heterotrimer composition on coupling of neurotransmitter receptors to N-type Ca~2+ channel modulation in sympathetic neurons

摘要

Voltage-dependent (VD) inhibition of N-type Ca~2+ channels is mediated primarily by neurotransmitter receptors that couple to pertussis toxin (PTX)-sensitive G proteins (such as G_o. and G_i). To date, however, the composition of heterotrimeric complexes, i.e.. specific Gαβγ combinations, capable of coupling receptors to N-type Ca~2+ channels has not been defined. We addressed this question by heterologously expressing identified Gαβγ combina- tions in PTX-treated rat sympathetic neurons and testing for reconstitution of agonist-mediated VD inhibition. The heterolo- gously expressed Gα subunits were rendered PTX-insensitive by mutating the codon specifying the ADP ribosylation site. The following results were obtained from this approach. (I) Expression of Gα_oA, Gα_oB, and Gα_i2 (aIong with Gβ_1γ2) reconstituted VD inhibition mediated by α2-adrenergic, adenosine, somatostatin, and prostaglandin E2 receptors. Conversely, expression of Gα_i1 and Go1s was ineffective at restoring coupling. (ii) Coupling efficiency, as determined from the magnitude of reconstituted Ca~2+ Current inhibition. depended on both the receptor and Gcr subtype. The following rank order of coupling efficiency was observed: Gα_oA = G_αoB > Gαiz for α2-adrenergic receptor: Gαi2 > GαoA = GαoB for adenosine and prostaglandin E2 receptors; and GαoB = Gαi2 > GαoA for the somatostatin receptor. (iii} In general. varying the Gpy composition of G羙A-containing heterotrimers had little effect on the coupling of α2-adrenergic receptors to the VD pathway. Taken together, these results suggest that multiple, diverse Gopy com- binations are capable of coupling neurotransmitter receptors to VD inhibition of N-type Ca~2+ channels. Thus. if exquisite Gcrpwcou- pling specificity exists in situ. it cannot arise solely from the inherent inability of other Gopy combinations to form functional signaling complexes.