FAST ACTIVATION AND INACTIVATION OF INOSITOL TRISPHOSPHATE-EVOKED CA2+ RELEASE IN RAT CEREBELLAR PURKINJE NEURONS

1. Calcium release from stores via inositol trisphosphate (InsP(3)) ac tivation of intracellular Ca2+ receptor-channels is thought to have a role in regulating the excitability of cerebellar Purkinje neurones. T he kinetic characteristics of InsP(3) receptor activation in Purkinje neurones are reported here. 2. InsP(3) was applied by flash photolysis of caged InsP(3) during whole-cell patch clamp. Ca2+ flux into the cy tosol was measured with a loci-affinity fluorescent Ca2+ indicator and by activation of Ca2+-dependent membrane conductance. 3. InsP(3) prod uced Ca2+ release from stores with an initial well-defined delay (mean , 85 ms at 10 mu M InsP(3)), which decreased to less than 20 ms at hig h InsP(3) concentrations. 4. The rate of rise of free [Ca2+], which pr ovides a measure of Ca2+ efflux and InsP(3) receptor activation, incre ased with increasing InsP(3) concentration in each cell and had a high absolute value of up to 1400 mu M s(-1) at 40 mu M InsP(3). The perio d of fast efflux was brief, inactivating in 25 ms at low and in 9 ms a t high InsP(3) concentration. 5. Peak free [Ca2+] was high (mean, 23 m u M with a pulse of 40 mu M InsP(3)) and increased with InsP(3) concen tration up to 80 mu M InsP(3) tested here. 6. Experiments with a flash -released, stable 5-thio-InsP(3), confirm that the low InsP(3) sensiti vity of Purkinje neurones does not result from metabolism of InsP(3). 7. The low functional affinity and fast activation by InsP(3) suggest a difference in InsP(3) receptor properties from non-neuronal cells te sted in the same way. The large Ca2+ efflux and high peak [Ca2+] proba bly result from high InsP(3) receptor-channel density. 8. Elevated cyt osolic [Ca2+] produced by Ca2+ influx through plasmalemmal Ca2+ channe ls strongly suppressed InsP(3)-evoked Ca2+ release from stores. Rapid termination of InsP(3)-evoked efflux results mainly from inhibition by high [Ca2+]. 9. The fast InsP(3) activation kinetics and rapid, stron g inactivation by Ca2+ influx suggest that interactions between InsP(3 )-mediated and membrane Ca2+ signalling could occur on a time scale co mpatible with neuronal excitation.