Extracellular proton-modulated pore-blocking effect of the anticonvulsant felbamate on NMDA channels

Felbamate (FBM) is a potent nonsedative anticonvulsant whose clinical effect is chiefly related to gating modi. cation (and thus use-dependent inhibition) rather than pore block of N-methyl-D-aspartate (NMDA) channels at pH 7.4. Using whole-cell recording in rat hippocampal neurons, we examined the effect of extracellular pH on FBM action. In sharp contrast to the findings at pH 7.4, the inhibitory effect of FBM on NMDA currents shows much weakened use-dependence at pH 8.4. Moreover, FBM neither accelerates the activation kinetics of the NMDA channel, nor enhances the currents elicited by very low concentrations of NMDA at pH 8.4. These differential effects of FBM between pH 7.4 and 8.4 are abolished in the mutant NMDA channels which lack proton sensitivity. Most interestingly, the inhibitory effect of FBM becomes flow-dependent and is evidently stronger in inward than in outward NMDA currents at pH 8.4. These findings indicate that FBM has a significantly more manifest pore-blocking effect on the NMDA channel at pH 8.4 than at pH 7.4. FBM therefore acts as an opportunistic pore blocker modulated by extracellular proton, suggesting that the FBM binding site is located at the junction of a widened and a narrow part of the ion conduction pathway. Also, we find that the inhibitory effect of FBM on NMDA currents is antagonized by external but not internal Na+, and that increase of external Na+ decreases the binding rate without altering the unbinding rate of FBM. These findings indicate that the FBM binding site faces the extracellular rather than the intracellular solution, and coincides with the outmost ionic (e. g., Na+) site in the NMDA channel pore. We conclude that the FBM binding site very likely is located in the external pore mouth, where extracellular proton, Na+, FBM, and NMDA channel gating have an orchestrating effect.