Benzodiazepine Receptors in Rat Brain: Action of Triton X-100 and Localization in Relation to the Synaptic Region

Abstract

Membranes were isolated from the cerebral cortices of rat brain and submitted to binding with ³H-flunitrazepam (³H-FNZP). Specific binding gave a hyperbolic saturation curve reaching a maximum between 10–12 nM suggesting a single population of sites. By Scatchard analysis a K_D = 4 nM and a B_max = 407 fmol/mg protein was obtained; the Hill number was 0.97. The effect of Triton X-100 was analyzed between concentrations of 6.4 × 10^-8 and 5 × 10^-1%. With 6.4 × 10^-6 and 6.4 × 10^-3% there is an activation of the binding without loss of protein. This activation is not reversed by washing. At 6.4 × 10^-2% or higher concentrations there is an inhibition that is partially reversed and also solubilization of some receptors. The activation with low detergent is due to an increase in B_max without change in K_D. With 2 × 10^-1% and 5 × 10^-1% Triton X-100 there is considerable decrease in B_max and in the latter case, an increase in affinity, as well. The results obtained through the action of Triton X-100 suggest that a proportion of benzodiazepine receptors are localized pre-synaptically. These findings are discussed in relation to previous studies from this laboratory on the localization of other central receptors with reference to the synaptic region.