Quantitative analysis of tetrapentylammonium-induced blockade of open N-methyl-d-aspartate channels

The blockade of open N-methyl-d-aspartate (NMDA) channels by tetrapentylamm onium (TPentA) in acutely isolated rat hippocampal neurons was studied usin g whole-cell patch-clamp techniques. TPentA prevented the closure of the NM DA channel following what is known as the foot-in-the-door mechanism. Hooke d tail currents appearing after termination of the agonist (aspartate) and TPentA coapplication were analyzed quantitatively according to the correspo nding sequential kinetic model. Studies of the hooked tail current amplitud e and the degree of the stationary current inhibition dependence on the blo cker concentration led to a new method for estimation of fast foot-in-the-d oor blocker binding/unbinding rate constants. The application of this metho d to the NMDA channel blockade by TPentA allowed finding the values of its binding (1.48 mu M-1 s(-1)) and unbinding (14 s(-1)) rate constants. An ana lysis of the dependence of the electric charge carried during the hooked ta il current on the blocker concentration led to a new method for estimation of the maximum NMDA channel open probability, P-o. The value of P-o found i n experiments with TPentA was 0.04.