Modulation of the GABAA receptor by barbiturates and pregnane steroids: differential effects of the influence of assay temperature.
The effect of temperature on the modulation of the GABAA receptor by barbiturates and steroids has been investigated in-vitro using a radioreceptor binding assay. Displaceable [3H]muscimol binding to a crude membrane preparation from rat cerebral cortex was enhanced by the endogenous steroid metabolite, 5 beta-pregnan-3 alpha-ol-20-one, by the synthetic steroid, alphaxalone, and by pentobarbitone in a dose-dependent manner. Hydrocortisone and corticosterone had no significant effect on [3H]muscimol binding. Analysis of binding data using a curve-fitting program ('Ligand') showed that both pentobarbitone (1 mM) and 5 beta-pregnan-3 alpha-ol-20-one (10 microM) increased the apparent number of high affinity binding sites in the membrane but had no effect on the affinity of [3H]muscimol binding (Kd approx. 11 nM). Increasing the assay temperature from 0 degrees C to 35 degrees C decreased [3H]muscimol binding and decreased the enhancement of binding by pentobarbitone but had no effect on 5 beta-pregnan-3 alpha-ol-20-one enhancement of binding. 5 alpha-Pregnan-3 alpha-ol-20-one increased the apparent rate of association of [3H]muscimol binding to its receptor whereas pentobarbitone had no effect. These different effects on the apparent association rate and the different responses to temperature, suggest that the barbiturate and steroid may interact with the GABAA receptor through different binding sites.
Yau
J L
JL
Department of Pharmacology and Clinical Pharmacology, Ninewells Hospital, University of Dundee, UK.
Balfour
D J
DJ
Stevenson
I H
IH
D000818:Animals; D001463:Barbiturates / Q000494:pharmacology*; D066298:In Vitro Techniques; D008297:Male; D009118:Muscimol / Q000493:pharmacokinetics; D011278:Pregnanes / Q000494:pharmacology*; D011869:Radioligand Assay; D051381:Rats; D011919:Rats, Inbred Strains; D011963:Receptors, GABA-A / Q000187:drug effects / Q000378:metabolism*; D013570:Synaptic Membranes / Q000187:drug effects / Q000378:metabolism; D013696:Temperature
D001463:Barbiturates; D011278:Pregnanes; D011963:Receptors, GABA-A; D009118:Muscimol