Impaired G protein function in gallbladder muscle from progesterone-treated guinea pigs

This study was designed to elucidate the mechanism of action of progesterone on gallbladder smooth muscle in guinea pigs. Adult male pigs were treated with either progesterone (2 mg.kg(-1).day(-1)) or saline for 7 days. Gallbladder muscle cells were isolated by enzymatic digestion with collagenase. Contractile responses td agonists were expressed as percent shortening from control cell length. [S-35]guanosine 5'-O-(3-thiotriphosphate) ([S-35]GTP gamma S)-binding properties of G proteins were assessed in crude membranes of gallbladder muscle with or without cholecystokinin octapeptide (CCK-8) stimulation. Gallbladder muscle cells from progesterone-treated guinea pigs exhibited an impaired contractile response to CCK-8, GTP gamma S, or aluminum fluoride but a normal response to potassium chloride or D-myo-inositol 1,4,5-trisphosphate compared with controls. Western blot analysis of gallbladder muscle revealed the presence of G(i1-2), G(i3), G(q/11), and G(s) proteins. The maximal contraction induced by CCK-8 was blocked by pertussis toxin and G(i) alpha(3)-specific antibodies, but not by G(i) alpha(1-2) or C(q/11)alpha antibodies. CCK-8 caused a significant increase in [S-35]GTP gamma S binding to G(i) alpha(3), but not to G(q/11)alpha or G(i) alpha(1-2) The stimulation of G(i) alpha(3) binding, however, was significantly reduced in gallbladder muscle membranes from progesterone-treated guinea pigs compared with that in control animals. In conclusion, progesterone might cause gallbladder hypomotility by downregulating G(i3) proteins.