Electrophysiological characterization of potent agonists and antagonists at pre‐ and postsynaptic GABAB receptors on neurones in rat brain slices

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Abstract
1 Intracellular recordings were made from neurones in striatum (caudate-putamen) and substantia nigra pars compacta in rat brain slices. Three GABAB agonists, baclofen, 3-aminopropylphosphinic acid (3-APPA) and 3-aminopropyl(methyl)phosphinic acid (SK&F 97541), depressed excitatory postsynaptic potentials (e.p.s.ps) mediated by glutamate in the striatum, and hyperpolarized neurones in the substantia nigra. The ability of 3-aminopropyl(diethoxymethyl)phosphinic acid (CGP 35348), 3-aminopropyl (hexyl)phosphinic acid (3-APHPA) and phaclofen to antagonize these responses was assessed. 2 Striatal e.p.s.ps, studied in the presence of bicuculline (30 μm), were reduced in amplitude by 92% with 6,7-dinitroquinoxaline-2,3-dione (DNQX; 30 μm). These e.p.s.ps were depressed by up to 95% by SK&F 97541 and baclofen with EC50s of 0.092 μm and 1.25 μm respectively. The maximal effect of 3-APPA was 67% with an EC50 of 0.83 μm. Agonist concentration-effect data fitted a single-site logistic model. GABAB agonists were without effect on striatal neurone membrane potential, input resistance or depolarizations induced by applied glutamate. 3 The depression of striatal e.p.s.ps by SK&F 97541 was reversibly antagonized by CGP 35348, 3-APHPA and phaclofen with estimated equilibrium dissociation constants (KB) of 11.2 ± 1.7 μm (n = 4), 13.3 ± 0.4 μm (n = 3), and 405 ± 43 μm (n = 3) respectively. CGP 35348 and 3-APHPA appeared to act competitively (Schild plot slopes of 0.99 and 1.01 respectively). 4 Nigral neurones were hyperpolarized by up to 25 mV by SK&F 97541 and baclofen with EC50s of 0.15 μm and 3.6 μm respectively. The maximum hyperpolarization by 3-APPA was only 84% that of the other agonists, with an EC50 of 9.0 μm. Agonist concentration-effect data fitted a single-site logistic model. 5 The SK&F 97541-induced hyperpolarization was reversibly antagonized by CGP 35348, 3-APHPA and phaclofen with estimated KBs of 17.6 ± 4.4 (n = 3), 14.0 ± 1.5 (n = 4), and >400 μm (n = 1) respectively. CGP 35348 appeared competitive (Schild plot slope of 0.99). Antagonists were also tested with baclofen as agonist, yielding similar KB estimates as for SK&F 97541. 6 It is concluded that at both the presynaptic and postsynaptic sites examined, SK&F 97541 was about 10 fold more potent than baclofen or 3-APPA. The antagonists CGP 35348 and 3-APHPA (KB 10–20 μm) were about 20 fold more potent than phaclofen. The similarities in relative agonist potency and estimated antagonist affinity between these two functionally distinct GABAB receptors renders them pharmacologically indistinguishable at present.