Concentration dependence of neurotransmitter effects on calcium current kinetics in frog sympathetic neurones.

1. Noradrenaline (NA) slows the activation kinetics of N-type calcium channels, via G proteins. It has been suggested that the G proteins act by binding directly to the calcium channels. If the slow kinetics reflect binding and unbinding of G proteins, the rates should depend on the concentration of activated G protein. 2. We used different concentrations of NA, and increasing durations of intracellular dialysis with GTP-gamma-S, to vary the concentration of activated G protein. 3. At depolarized potentials (-20 or -10 mV), the slow activation kinetics showed no detectable concentration dependence. This analysis required correction for effects of inactivation on the measured time constants. 4. At -80 mV, reinhibition of calcium channel current was more rapid for larger responses. Thus, the effect appears to be concentration dependent at -80 mV, but not at more depolarized voltages. 5. This voltage dependence is actually expected from kinetic principles: the binding step is rate limiting when the position of equilibrium is toward the bound state (at -80 mV), but not when equilibrium favours unbinding (when the channel is open). 6. During inhibition, the channel appears to 'sense' directly the concentration of the modulator, possibly active G proteins.