Diazepam-potentiated [3H]phenytoin binding is associated with peripheral-type benzodiazepine receptors and not with voltage-dependent sodium channels.
In the presence of diazepam, [3H]phenytoin binds with high affinity to brain membranes. The present experiments examined whether this high affinity [3H]phenytoin-binding site co-localized with the standard [3H]phenytoin-binding site on the voltage-dependent sodium channel (VDSC). Veratridine, a pharmacological activator of the voltage-dependent sodium channel, that inhibits standard [3H]phenytoin binding, failed to affect the high affinity diazepam-potentiated [3H]phenytoin binding in brain membranes, suggesting that the potentiated binding interaction resides at a site distinct from the voltage-dependent sodium channel. This possibility was confirmed by anion exchange chromatography of digitonin-solubilized rat brain membranes, as diazepam-potentiated high affinity [3H]phenytoin binding eluted in column fractions that were distinct from [3H]saxitoxin-defined voltage-dependent sodium channels. To examine whether diazepam-potentiated [3H]phenytoin binding might be associated with other 'classic' benzodiazepine receptor sites, we tested whether specific ligands for benzodiazepine receptors would either produce or block potentiated [3H]phenytoin binding. Neither agonists, nor antagonists, of the high affinity central-type benzodiazepine receptor affected potentiated [3H]phenytoin binding, suggesting that the high affinity potentiated binding site is not likely associated with central benzodiazepine receptors. Peripheral-type benzodiazepine receptor agonists, however, did potentiate [3H]phenytoin binding, and a specific receptor antagonist (PK11195) attenuated the potentiation seen with diazepam. Overall, these data illustrate that [3H]phenytoin interacts with a novel site in brain membranes that is distinct from the voltage-dependent sodium channel and is allosterically revealed by peripheral-type, but not central-type, benzodiazepine receptor agonists.
Francis
J
J
Division of Cell and Molecular Biology, Toronto Western Research Institute, Toronto Western Hospital, 399 Bathurst Street, M5T 2S8, Toronto, Ontario, Canada.
Eubanks
J H
JH
McIntyre Burnham
W
W
D000818:Animals; D001921:Brain / Q000378:metabolism; D002852:Chromatography, Ion Exchange; D003975:Diazepam / Q000494:pharmacology*; D004072:Digitonin / Q000494:pharmacology; D004357:Drug Synergism; D004594:Electrophysiology; D008024:Ligands; D008297:Male; D008566:Membranes / Q000378:metabolism; D010672:Phenytoin / Q000378:metabolism*; D051381:Rats; D020318:Rats, Long-Evans; D011963:Receptors, GABA-A / Q000378:metabolism*; D015222:Sodium Channels / Q000378:metabolism / Q000502:physiology*; D012995:Solubility; D014316:Tritium; D014701:Veratridine / Q000494:pharmacology
D008024:Ligands; D011963:Receptors, GABA-A; D015222:Sodium Channels; D014316:Tritium; D010672:Phenytoin; D014701:Veratridine; D004072:Digitonin; D003975:Diazepam