Abstract

Bath application of micromolar concentrations of forskolin to Xenopus oocytes that express either Torpedo electroplax or mouse muscle nicotinic acetylcholine (ACh) receptors leads to a reduction in the size of the ACh-elicited currents. This inhibition is concentration dependent and rapidly reversible, with full onset and recovery occurring within the exchange time of the recording chamber. Torpedo and mouse ACh receptors exhibit differential sensitivity to forskolin, with the Torpedo receptor showing higher affinity than the mouse receptor, with Ki values of 6.5 microM and 22 microM, respectively. The affinity for forskolin increases with ACh concentration, which rules out the possibility that forskolin acts as a competitive inhibitor. Single-channel analysis using excised patches shows that forskolin has no effect on either the single-channel amplitude or mean open time but, instead, reduces the number of channel openings per unit time, suggesting that forskolin either is a very slow channel blocker or alters receptor gating such that a fraction of the channels enter a state from which they are no longer available to open. Finally, through the use of a series of mouse-Torpedo hybrid ACh receptors, it is shown that the structural features responsible for the observed species difference in the affinity of ACh receptors for forskolin, and thus at least part of the binding site, are located on the gamma subunit.