Efficient functional coupling of the human D3 dopamine receptor to G(o) subtype of G proteins in SH-SY5Y cells

1 The D3 dopamine receptor presumably activates G(i)/G(o) subtypes of G-pro teins, like the structurally analogous D2 receptor, but its signalling targ ets have not been clearly established due to weak functional signals from c loned receptors as heterologously expressed in mostly non-neuronal cell lin es. 2 In this study, recombinant human D3 receptors expressed in a human neurob lastoma cell line, SH-SY5Y, produced much greater signals than those expres sed in a human embryonic kidney cell line, HEK293. Quinpirole, a prototypic agonist, markedly inhibited forskolin-stimulated cyclic AMP production and Ca2+-channel (N-type) currents in SH-SY5Y cells, and enhanced GTP gamma(35 )S binding in isolated membranes, nearly ten times greater than that observ ed in HEK293 cell membranes. 3 GTP gamma(35)S-bound G alpha subunits from quinpirole-activated and solub ilized membranes were monitored upon immobilization with various G alpha-sp ecific antibodies. G alpha(o) subunits (not G alpha(i)) were highly labelle d with GTP gamma(35)S in SH-SY5Y, but not in HEK293 cell membranes, despite their abundance in the both cell types, as shown with reverse transcriptio n-polymerase chain reaction and Western blots. N-type Ca2+ channels and ade nylyl cyclase V (D3-specific effector), on the other hand, exist only in SH -SY5Y cells. 4 More efficient coupling of the D3 receptor to G(o) subtypes in SH-SY5Y th an HEK293 cells may be attributed, at least in part, to the two D3 neuronal effecters only present in SH-SY5Y cells (N-type Ca2+-channels and adenylyl cyclase V). The abundance of G(o) subtypes in the both cell lines seems to indicate their availability not a limiting factor.