Regulation of Adenylyl Cyclase Type V/VI in Smooth Muscle: Interplay of Inhibitory G Protein and Ca²⁺ Influx

Abstract

The characteristics of inhibitory regulation of adenylyl cyclase V/VI by Ca²⁺ and G proteins were examined in dispersed gastric smooth muscle cells. The mechanisms were evoked separately, sequentially, or concurrently using ligand-gated and G protein-coupled receptor agonists and receptor-independent probes (e.g, thapsigargin). During the initial phase of agonist stimulation, α,β-methylene-ATP, UTP, and ATP inhibited forskolin-stimulated cAMP formation in a concentration-dependent fashion. Inhibition by α,β-methylene-ATP, which activates ligand-gated P_2X receptors, was abolished by zero Ca²⁺, whereas inhibition by UTP, which activates P_2Y2 receptors coupled to G_q/11 and G_i3, was not affected by zero Ca²⁺ but was abolished by pertussis toxin (PTX). Inhibition by ATP, which activates both P_2X and P_2Y2 receptors, was not affected by zero Ca²⁺ alone; but after inhibition mediated by G_αi3 was blocked with PTX, inhibition by Ca²⁺influx was unmasked and was abolished by zero Ca²⁺. Inhibition by cholecystokinin-8 was observed only during the phase of capacitative Ca²⁺ influx and was blocked by zero Ca²⁺. Inhibition by UTP during this phase was not affected by zero Ca²⁺ alone; but after inhibition mediated by Gα_i3 was blocked with PTX, inhibition by Ca²⁺influx was unmasked and was abolished by zero Ca²⁺. Inhibition of adenylyl cyclase V/VI activity in smooth muscle can be mediated independently by inhibitory G proteins and Ca²⁺influx but is exclusively mediated by inhibitory G proteins when both mechanisms are triggered.