Other than the fact that the cystic fibrosis transmembrane conductance regulator (CFTR) Cl- channel can be activated by cAMP dependent kinase (PKA), little is known about the signal transduction pathways regulating CFTR. Since G-proteins play a principal role in signal transduction regulating several ion channels [4, 5, 9], we sought to test whether G-proteins control CFTR Cl- conductance (CFTR G(Cl)) in the native sweat duct (SD). We permeabilized the basolateral membrane with alpha-toxin so as to manipulate cytosolic nucleotides. We activated G-proteins and monitored CFTR G(Cl) activity as described earlier [20, 23, 25]. We now show that activating G-proteins with GTP-gamma-S (100 microm) also activates CFTR G(Cl) in the presence of 5 mm ATP alone (without exogenous cAMP). GTP-gamma-S increased CFTR G(Cl) by 44 +/- 20 mS/cm(2) (mean +/- se; n = 7). GDP (10 mm) inhibited G-protein activation of CFTR G(Cl) even in the presence of GTP-gamma-S. The heterotrimeric G-protein activator (AlF(4-) in the cytoplasmic bath activated CFTR G(Cl) (increased by 51.5 +/- 9.4 mS/cm(2) in the presence of 5 mm ATP without cAMP, n = 6), the magnitude of which was similar to that induced by GTP-gamma-S. Employing immunocytochemical-labeling techniques, we localized Galphas, Galphai, Galphaq, and Gbeta at the apical membranes of the sweat duct. Further, we showed that the mutant CFTR G(Cl) in ducts from cystic fibrosis (CF) subjects could be partially activated by G-proteins. The magnitude of mutant CFTR G(Cl) activation by G-proteins was smaller as compared to non-CF ducts but comparable to that induced by cAMP in CF ducts. We conclude that heterotrimeric G-proteins are present in the apical membrane of the native human sweat duct which may help regulate salt absorption by controlling CFTR G(Cl) activity.