Inhibitory guanosine triphosphatebinding protein-mediated regulation of vasopressin action in isolated single medullary tubules of mouse kidney

Vasopressin (AVP) plays a key role in maximal urine concen-tration by stimulating NaCi reabsorption in the medullary thick ascending limbs of Henle (MAL) and by increasing water permeability in the medullary collecting tubules (MCT). These effects of AVP in MAL and MCT are mediated by cAMP. Alpha2-adrenergic stimulation in MCI, and high ambient Ca2+ and PGE2 in MAL inhibit AVP-dependent cAMP production and thereby modulate urine concentration. The present study was undertaken to clarify the mechanisms underlying the inhi-bition of AVP-dependent cAMP production by these agents using microdissected mouse MAL and MCT. Preincubation of MCI and MAL with 1 gg/ml pertussis toxin for 3 and 6 h, respectively, resulted in ADP-ribosylation of an- 41-kD pro-tein, which was presumably an alpha subunit of the inhibitory GTP-binding protein G.. Epinephrine, 10-6 M, via alpha2-adrenergic stimulation, inhibited AVP-dependent cAMP pro-duction in MCT. Preincubation of MCT for 3 h with pertussis toxin abolished the inhibition of AVP-dependent cAMP pro-duction by epinephrine. High ambient Ca2 ' and PGE2 both inhibited AVP-dependent cAMP production in MAL. Prein-cubation of MAL for 6 h with pertussis toxin abolished the inhibition by high ambient Ca2 ' and attenuated the inhibition by PGE2. Preincubation ofMCT or MAL with pertussis toxin for 1 h was ineffective in ADP-ribosylation and did not modify the inhibition of AVP-dependent cAMP production by these agents in both nephron segments. Our data suggest that the inhibition of AVP-dependent cAMP production by alpha2-adrenergic stimulation in MCI, and by high ambient Ca2 ' and PGE2 in MAL, is mediated, at least in part, through activation of GQ