5-HT1A RECEPTOR ACTIVATES NA+ H+ EXCHANGE IN CHO-K1 CELLS THROUGH G(I-ALPHA-2) AND G(I-ALPHA-3)/

5-HT1A receptors couple to many signaling pathwaysin CHO-K1 cells thro ugh pertussis toxin-sensitive G proteins, The purpose of this study wa s to determine which members of the G(i/o/z) family mediate 5-HT1A rec eptor-activated Na+/H+ exchange as measured by microphysiometry of cel l monolayers. The method was extensively validated, showing that proto n efflux was sodium-dependent, inhibited by amiloride analogs, and act ivated by growth factors, phorbol ester, calcium ionophore, and hypert onic stress. 5-HT and the specific agonist (+/-)-8-hydroxy-2-(di-N-pro pylamino)tetralin hydrobromide rapidly stimulated proton efflux that w as blocked by a specific receptor antagonist, amiloride analogs or per tussis toxin, The activation by 5-HT depended upon extracellular sodiu m and could be demonstrated under conditions of imposed intracellular acid load, as well as in the presence and absence of glycolytic substr ate, Acceleration of proton efflux was not inhibited by sequestration of G protein beta gamma-subunits, a maneuver that blocked 5-HT1A recep tor activation of mitogen-activated protein kinase, Transfection of G( z alpha) and pertussis toxin-resistant mutants of G(o alpha) and G(i a lpha 1) did not reverse the blockade induced by pertussis toxin, In co ntrast, pertussis toxin-resistant mutants of G(i alpha 2) and G(i alph a 3) ''rescued'' the ability of 5-HT to increase proton efflux after p ertussis toxin treatment, These experiments demonstrate clearly that G (i alpha 2) and G(i alpha 3) can specifically mediate rapid agonist-in duced acceleration of Na+/H+ exchange.