The distribution and the pharmacological properties of the binding of the benzodiazepine receptor antagonist [3H]-Ro 15-1788 (8-fluoro-3-carboethoxy-5,6-dihydro-5-methyl-6-oxo-4H imidazol [1,5-a] 1,4 benzodiazepine) were compared in some brain membranes of the saltwater teleost fish, Mullus surmuletus: only a single population of [3H]-Ro 15-1788 binding sites was detected. The binding was saturable and reversible with a high affinity, revealing a significant population of binding sites (Kd value of 2.1 +/- 0.2 nM and Bmax value of 1400-900 fmol mg-1 of protein, depending on fish length). The highest concentration of benzodiazepine recognition sites labelled with [3H]-Ro 15-1788 was present in the optic lobe and the olfactory bulb and the lowest concentration was found in the medulla oblongata, cerebellum and spinal cord. In order to explore behavioural selectivity as a consequence of multiple receptor subtypes, six benzodiazepine receptor ligands, flunitrazepam (5-(2-fluoro-phenyl)-1,3-dihydro-1-methyl-7-nitro-2H-1, 4-benzodiazepin-2-one), alpidem, (N,N-dipropyl-6-chloro-2-(4-chlorophenyl)imidazo [1,2-a] pyridine-3- acetamide) zolpidem {N,N,6, trimethyl-2-(4-methyl-phenyl) imidazo [1,2-a] pyridine-3-acetamide hemitartrate}, methyl beta carboline-3-carboxylate (beta CCM), Ro 15-1788 and Ro 5-4864 (4'-chlorodiazepam), were tested in vitro by binding of [3H]-Ro 15-1788 to membrane preparations from various brain areas of Mullus surmuletus, Displacement studies showed a similar rank order of efficacy of various unlabelled ligands. In all regions of the brain and in the spinal cord. GABA potentiate [3H]-flunitrazepam binding in a similar order, suggesting that the BDZ recognition sites are part of the GABAA receptor structure. These results suggest that central-type benzodiazepine receptors are present in one class of benzodiazepine binding sites in the saltwater teleost fish brain of Mullus surmuletus (type I-like). Here we report initial evidence of homogeneity of subtypes of central benzodiazepine receptors in the spinal cord of the saltwater teleost fish. Mullus surmuletus.