STATE-DEPENDENT INHIBITION OF NA-NEURONS AND IN A MAMMALIAN-CELL LINEEXPRESSING RAT-BRAIN TYPE IIA NA+ CHANNELS( CURRENTS BY THE NEUROPROTECTIVE AGENT 619C89 IN RAT HIPPOCAMPAL)

The compound 619C89 -piperazinyl)-5-(2,3,5-trichlorophenyl)pyrimidine] is an effective neuroprotective agent in in vivo models of cerebral i schaemia. It has been suggested to act by inhibiting voltage-gated Na channels. To test this hypothesis, the action of 619C89 on recombinan t rat brain type IIA Na+ channels expressed in Chinese hamster ovary c ells and on native Na+ channels in acutely dissociated rat hippocampal neurons has been studied using whole-cell voltage-clamp recording tec hniques. In the cell line expressing type IIA Na+ channels, 619C89 cau sed a reversible inhibition of Na+ currents in a concentration- and vo ltage-dependent manner. A half-maximal inhibitory concentration (IC50) of approximately 50 mu M was obtained at a holding potential of -90 m V whereas, with a conditioning prepulse to -60 mV for 30 s, the IC50 w as reduced to 8 mu M. Furthermore, the inhibition was markedly enhance d by a use-dependent action, which was dependent not only on the frequ ency of stimulation, but also on the duration (3.5-40 ms) of the pulse s. Trains (10-50 Hz) of up to 60 depolarizing pulses of 0.7 ms duratio n did not evoke any use-dependent inhibition in the presence of 619C89 , suggesting that this compound is not an open channel blocker. The vo ltage- and use-dependent inhibition by 619C89 was also observed on nat ive Na+ channels in hippocampal neurons. 619C89 (10 mu M) produced a s mall hyperpolarizing shift in the fast inactivation curve and a substa ntial (13 mV) hyperpolarizing shift in slow inactivation. The compound dramatically delayed the recovery from inactivation without affecting the development of inactivation. Moreover, 619C89 has no effect on th e shape of the current-voltage relationship or on the voltage activati on curve. These data indicate that 619C89 interacts selectively with t he inactivated state of the Na+ channel with an estimated affinity of 3 mu M. This primary action of 619C89 may underlie its neuroprotective effects. Copyright (C) 1966 IBRO.