Effect of (+)-sparteine on nicotinic acetylcholine receptors in the neurons of rat superior cervical ganglion.

The effects of (+)-sparteine, a ganglionic blocking agent, on acetylcholine (ACh)-induced membrane currents and on fast excitatory postsynaptic currents (EPSCs) were studied in the neurons of rat isolated superior cervical ganglion, with the whole-cell patch-clamp recording method and the two-electrode voltage-clamp method, respectively. (+)-Sparteine (2 microM) reduced the ACh-induced current caused by activation of nicotinic ACh receptors (AChRs) in a voltage-independent manner at membrane potentials of -50 mV to +30 mV, whereas its blocking effect increased at more negative membrane potentials. The dose-response relationship for ACh was modified by 2 microM (+)-sparteine at -50 mV and at -90 mV in a fashion typical for competitive rather than noncompetitive antagonists. The apparent mean open time of the AChR channel, as estimated from the power density spectrum of the ACh-induced current fluctuations at -90 mV, was not decreased by 2 microM (+)-sparteine, in contrast to what was observed with hexamethonium, the well known open-channel blocker for ganglionic AChRs. At higher concentrations, i.e., 5 microM and 10 microM (lower concentrations were not effective), (+)-sparteine reduced the amplitude of the EPSC and the time constant of the EPSC decay. The former effect was voltage independent, whereas the latter effect was voltage independent at membrane potentials of -70 mV and more positive and increased at membrane potentials of -90 and -110 mV. These results suggest that (+)-sparteine produces in ganglionic AChRs a competitive blocking effect and, in addition, an open-channel blockade. The latter component probably provides a smaller contribution than does the former to the blockade by (+)-sparteine of the ACh-induced current. Conformational analysis of the (+)-sparteine molecule was performed, and the dimensions of the molecule were measured. Minimum dimensions of the space-filling profile for two conformers, high and low populated, were found to be 7.3 x 7.9 A and 6.8 x 7.5 A, respectively. Both profiles are larger than the channel profile at which the open-channel blockers have been suggested to bind, which may explain comparatively low open-channel-blocking activity of (+)-sparteine.