Role of cyclic AMP and protein kinase A in K+ channel activation by calcitonin gene-related peptide (CGRP) in the guinea-pig ureter.

1. The aim of this study was to assess whether agents that interfere with the intracellular actions of cAMP and activation of protein kinase A (PKA) prevent the inhibitory action of human alpha-calcitonin gene-related peptide (CGRP) in the guinea-pig ureter smooth muscle. The action of CGRP was compared to that of the K+ channel opener, cromakalim, and the adenylyl cyclase activator, forskolin, toward electrical field stimulation- (EFS) induced myogenic twitch contractions of the ureter. To further verify the role of cAMP in the action of CGRP, we also studied the effect of stable cAMP analogues and of the phosphodiesterase inhibitor, 3-isobutyl-1-methylxanthine (IBMX). 2. Maximally effective concentrations of CGRP (0.1 microM) or forskolin (10 microM) produced a transient suppression of twitches. Cromakalim (3 microM) likewise produced a prompt suppression of twitches that in most cases exceeded 15 min. The early suppressant effect of CGRP or forskolin was inhibited by 1 or 10 microM glibenclamide; about 30% of the effect of CGRP was glibenclamide-resistant. The effect of cromakalim was totally suppressed by glibenclamide. 3. The inhibitory effect of CGRP was concentration-dependently reduced by low concentrations of barium chloride (IC50 63 microM), which blocked with similar potency the inhibitory action of cromakalim (IC50 60 microM). Glibenclamide (10 nM-10 microM) concentration-dependently inhibited the effect of CGRP and cromakalim with IC50S of 0.13 and 0.72 microM, respectively. 4. The cAMP analogues dibutyrye-cAMP (1-3 mM), 8-(4-chlorophenylthio)cAMP (0.3-1 mM) and Sp-cAMP monophosphothioate (0.1-0.3 mM) were either ineffective or poorly effective in inhibiting twitches. The cGMP analog, 8Br-cGMP (100-300 microM) produced a slowly developing, glibenclamide (1 microM)-resistant partial inhibition (25-30%) of twitches. 5. IBMX (1-300 microM) produced a concentration-dependent inhibition of twitches (EC50 16 microM). IBMX (100 microM) produced a large (peak 91%) and transient inhibition: glibenclamide (1 microM) blocked the early peak of the inhibitory action of IBMX, similar to the effect observed toward CGRP and forskolin.