Journal zyxwvutsrqponm of Neurochemislry zyxwvutsrqponm Raven Press, zyxwvutsrqponmlk Ltd., New York zyxwvutsrqponm 0 1989 International Society for Neurochemistry Distribution and Pharmacological Properties of the GABAA/ Benzodiazepine/Chloride Ionophore Receptor Complex in the Brain of the Fish zyxw AnguiZZa anguiZZa M. G. Corda, B. Longoni, *A. Cau, *S. Paci, *S. Salvadori, *U. Laudani, and G. Biggio Department of Experimental Biolom, Chair of Pharmacology; and *Institute of Zoology, University of Cagliari, Cagliari, Italy Abstract: In the present study, we characterized the distri- bution and the pharmacological properties of the different components of the GABAA receptor complex in the brain of the eel (Anguilla anguilla). Benzodiazepine recognition sites labeled “in vitro” with [3H]flunitrazepam zyxwvuts ( [3H]FNT) were present in highest concentration in the optic lobe and in lowest concentration in the medulla oblongata and spinal cord. A similar distribution was observed in the density of zyxwvu y- [3H]aminobutyric acid ([3H]GABA) binding sites. GABA in- creased the binding of [3H]FNT in a concentration-dependent manner, with a maximal enhancement of zyxwvut 45% above the control value, and, vice versa, diazepam stimulated the bind- ing of [3H]GABAto eel brain membrane preparations. The density of benzodiazepine and GABA recognition sites and their reciprocal regulation were similar to those observed in the rat brain. In contrast, the binding of the specific ligand for the C1- ionophore, t-[35S]butylbicyclophosphorothionate ([35S]TBPS), to eel brain membranes was lower than that found in the rat brain. In addition, [35S]TBPS binding in eel brain was less sensitive to the inhibitory effects of GABA and muscimol and much more sensitive to the stimulatory effect of bicuculline, when compared with [35S]TBPS binding in the rat brain. Moreover, the uptake of 36Cl- into eel brain membrane vesicles was only marginally stimulated by con- centrations of GABA or muscimol that significantly enhanced the 36Cl- uptake into rat brain membrane vesicles. Finally, intravenous administration of the P-carboline inverse agonist 6,7-dimethoxy-4-ethyl-~-carboline-3-carboxylic acid methyl ester (20 mg/kg) and of the chloride channel blocker pen- tylenetetrazole (80 mg/kg) produced convulsions in eels that were antagonized by diazepam at doses five to 20 times higher than those required to produce similar effects in rats. The results may indicate a different functional activity of the GABA-coupled chloride ionophore in the fish brain as com- pared with the mammalian brain. Key Words: GABAA re- ceptor complex-Brain-Eel (Anguilla anguil1a)-Benzo- diazepine recognition sites-Flunitrazepam-y-Aminobu- tyric acid-t-Butylbicyclophosphorothionate-Diazepam- Chloride ionophore. Corda M. G. et al. Distribution and pharmacological properties of the GABAA/benzodiazepine/ chloride ionophore receptor complex in the brain of the fish Anguilla anguilla. J. Neurochem. 52, 1025-1034 (1989). y-Aminobutyric acid (GABA) is the major inhibitory neurotransmitter in brain (Kmjevic, 1974; Enna and Gallagher, 1983). In neuronal membranes, GABA in- teracts with specific recognition sites that are believed to be part of a macromolecular protein complex that also contains a recognition site for benzodiazepines, a recognition site for convulsant [picrotoxin, pentylene- tetrazole (PTZ), and t-butylbicyclophosphorothionate (TBPS)] and anticonvulsant (barbiturates) drugs, and a chloride ion channel (for review, see Olsen and Ven- ter, 1986). Whereas the biochemical, physiological, and pharmacological properties of the GABAA receptor ~ ~~~ Received March 17, 1988; revised manuscript received July 26, 1988; accepted September 12, 1988. Address correspondence and reprint requests to Dr. M. G. Corda at Department of Experimental Biology, Chair of Pharmacology, University of Cagliari, Via Palabanda, 12,09123 Cagliari, Italy. complex have been extensively studied in the mam- malian brain, the characterization of this receptor complex in fishes has received little attention. In fact, only a few binding studies have measured the distri- bution of benzodiazepine, GABA, and convulsant/ barbiturate sites in the brain of higher bony fishes (Nielsen et al., 1978; Fernholm et al., 1979; Mann and Enna, 1980; Wilkinson et al., 1983; Cole et al., 1984; Mathis and Tunnicliff, 1984). Moreover, there appears to be a paucity of data on the biochemical and phar- macological properties of the chloride ionophore cou- pled to the GABAA receptor in fish brain. Abbreviations used: P-CCE, ethyl-~-carboline-3-carboxylate; DMCM, 6,7-dimethoxy-4-ethyl-~-carboline-3-carboxylic acid methyl ester; FNT, flunitrazepam; GABA, y-aminobutyric acid; kD, kilodalton(s); PTZ, pentylenetetrazole; TBPS, t-butylbicyclophosphorothionate. 1025