gamma-Aminobutyric acid receptor-regulated 36Cl- flux in mouse cortical slices.

An improved neurochemical assay for gamma-aminobutyric acid (GABA) function has been developed using tracer-radioactive chloride efflux in mouse cortical slices. Careful maintenance of the brain slice viability resulted in a 3-fold stimulation of 36Cl- efflux rate by the GABA agonist muscimol (EC50 = 3 microM), comparable to electrophysiologic and other chloride flux preparations. The shape of the muscimol dose-response curve was shallow, suggestive of negative cooperativity or heterogeneous receptors, but tissue uptake of agonist, possible diffusion barriers and apparent functional desensitization complicated these results. The response to muscimol was inhibited by GABAA receptor antagonists such as RU5135 and was enhanced by barbiturates and benzodiazepines. As observed previously, barbiturates stimulated 36Cl- efflux rate on their own and potentiated the response (potency and maximal effect) to muscimol in a stereospecific and picrotoxin-sensitive manner. Benzodiazepine receptor ligands alone did not alter 36Cl- flux, but agonists such as flunitrazepam enhanced the response to muscimol, an effect sensitive to the antagonist Ro15-1788. The inverse agonist methyl 6,7-dimethoxy-4-ethyl-beta-carboline-3-carboxylate did not inhibit muscimol-activated 36Cl- flux. The anthelminthic-insecticide avermectin B1a stimulated 36Cl- flux by itself, and this response was apparently additive with that of muscimol. This brain slice chloride flux assay is therefore suitable for the assessment of activity including dose-response curves for GABAA agonists, antagonists and modulators including benzodiazepines.