Mechanism of acetylcholine action on pacemaker current (i(f)) in canine Purkinje fibers.

We have recently reported in canine Purkinje fibers that acetylcholine (ACh) can reverse the positive voltage shift of the pacemaker current (i(f)) induced by beta-adrenergic stimulation while having no direct action of its own. We have now investigated this effect of ACh on the cyclic adenosine monophosphate (cAMP) cascade in more detail. We find that addition of a membrane permeable analogue of cAMP (8-chlorophenylthio cAMP), 0.5-1 mM, increased the amplitude of i(f). This action was not reversed by 1 microM ACh, implying that ACh acts at a step prior to cAMP action. We then looked at the steps controlling intracellular concentration of cAMP. Inhibiting the phosphodiesterase with 100 microM isobutyl-1-methylxanthine (IBMX) increased i(f). This action, however, was reversed by ACh. Finally we investigated whether the action of forskolin, a direct activator of adenylyl cyclase, could be reversed by ACh. Forskolin (10-20 microM) increased i(f), and ACh at 1 microM partially reversed this action of forskolin. These results suggest that, in canine Purkinje fibers, ACh reverses the positive action of beta-adrenergic agents on i(f) via a decrease in cAMP production.