Ifenprodil blocks N-methyl-D-aspartate receptors by a two-component mechanism.
The inhibition of N-methyl-D-aspartate (NMDA) receptor channels by the vasodilatory and anti-ischemic agent ifenprodil was examined on cultured rat hippocampal neurons. Whole-cell and single-channel patch recordings were used. Ifenprodil inhibition of NMDA currents could be separated into two components, with IC50 values of 0.75 and 161 microM. The high and low affinity components were both voltage independent but could be separated by their kinetics and dependence on extracellular calcium and glycine. The maximal inhibition of inward current by ifenprodil (approximately 90%) was equally divided between the two components in 0.3 mM extracellular calcium and 500 nM glycine. The low affinity action of ifenprodil had rapid kinetics and appeared to result from allosteric inhibition of the glycine modulatory site on the NMDA receptor. The macroscopic kinetics of the high affinity component were slow. The rate of onset was concentration dependent, and complete recovery required 1-2 min. Unlike open-channel blockers, ifenprodil block was not use dependent, and pre-exposure to ifenprodil also reduced subsequent NMDA responses. Low concentrations of ifenprodil were less effective after calcium-dependent inactivation of whole-cell currents, but the IC50 was unaffected, suggesting that calcium and ifenprodil act on a common set of channels. On outside-out membrane patches, ifenprodil reduced the frequency of channel opening without altering the single-channel conductance. Open time histograms of the large conductance events revealed two mean open times of approximately 2 and 8 msec, but only the duration of the long openings was decreased by ifenprodil. This effect was concentration dependent and revealed a blocking rate constant of 6 x 10(7) M-1sec-1. However, the proportion of current blocked by low concentrations of ifenprodil was larger in outside-out patches than in whole-cell recordings, suggesting that intracellular factors may influence ifenprodil efficacy. These results indicate that high affinity ifenprodil binding is extracellular and does not require agonist binding or channel opening. Because low concentrations of ifenprodil only partially inhibited the current and affected only the long openings, ifenprodil may promote a modal shift in channel gating.
Legendre
P
P
Vollum Institute, Oregon Health Sciences University, Portland 97201.
Westbrook
G L
GL
D000818:Animals; D001665:Binding Sites; D002118:Calcium / Q000502:physiology; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D005998:Glycine / Q000037:antagonists & inhibitors; D007051:Ibotenic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D007473:Ion Channels / Q000187:drug effects; D016202:N-Methylaspartate / Q000494:pharmacology; D010880:Piperidines / Q000494:pharmacology*; D011073:Polyamines / Q000037:antagonists & inhibitors; D051381:Rats; D011919:Rats, Inbred Strains; D016194:Receptors, N-Methyl-D-Aspartate / Q000187:drug effects*; D014665:Vasodilator Agents / Q000494:pharmacology*; D018350:alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid
D007473:Ion Channels; D010880:Piperidines; D011073:Polyamines; D016194:Receptors, N-Methyl-D-Aspartate; D014665:Vasodilator Agents; D007051:Ibotenic Acid; D016202:N-Methylaspartate; D018350:alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; C010739:ifenprodil; D002118:Calcium; D005998:Glycine