Strong cAMP Response to a GPCR Agonist Challenge Despite Apparent Inactivation

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A large variety of neurotransmitters, hormones, paracrine agents, and odorants exert their effects through G protein coupled receptors (GPCRs). These receptors activate intracellular G proteins that in turn modulate the activity of different effector proteins. Based largely on the study of the light-activated receptor rhodopsin and the beta-adrenergic receptor, the vast majority of GPCRs are assumed to undergo a process of inactivation or silencing involving receptor phosphorylation and capping by one of the arrestins. The advent of improved cAMP sensors in living cells and enhanced molecular information about phosphodiesterases has led to an increased appreciation of the role that these enzymes play in the shaping and termination of cAMP signals. We report here a high-resolution measurement of vasoactive intestinal peptide (VIP)-triggered cAMP signals near the surface membrane in pituitary GH4C1 cells, using modified cyclic nucleotide-gated ion channels. The signals are transient and the falling phase is due solely to an increase in phosphodiesterase activity. There is no detectable receptor inactivation on this timescale. Surprisingly, this system is highly responsive to subsequent increases in VIP levels. We show that such responsiveness can arise in a system in which receptors remain active and phosphodiesterase activity is just slightly higher than adenylyl cyclase activity, but not in a system that exhibits classical receptor inactivation. The upregulation of phosphodiesterase activity represents a type of inactivation or desensitization that causes a decline in the response to the initial agonist dose and limits the intracellular spread of cAMP. However, this mechanism has the unique feature of allowing the cell to respond to subsequent agonist challenges.