Enantioselective modulation of GABAergic synaptic transmission by steroids and benz[e]indenes in hippocampal microcultures.

The effects of enantiomers of the neurosteroid analogues, 3alpha-hydroxy-5alpha-pregnan-20-one (DHP) and 3alpha-hydroxy-5alpha-androstane-17beta-carbonitrile (ACN), and the benz[e]indene, BI-1, on synaptic currents were examined in microcultures of rat hippocampal neurons. Over the range of 0.1-10 microM, the (+)-enantiomers were more potent and effective than their (-)-enantiomeric counterparts in enhancing gamma-aminobutyric acid (GABA)A receptor-mediated evoked synaptic currents. The (+)-enantiomers had small effects on peak currents, but slowed the decay of inhibitory synaptic currents, resulting in 2-3-fold increases in charge transfer during inhibitory synaptic events at 10 microM. Similar prolongations of spontaneous miniature inhibitory postsynaptic currents (IPSCs) and responses to brief GABA pulses to outside-out patches suggest that the prolongations of evoked synaptic currents result primarily from postsynaptic effects. In contrast, the (-)-enantiomers had little effect on evoked IPSCs at concentrations < or = 1 microM, but enhanced inhibitory transmission at 10 microM. At concentrations < or = 1 microM, neither the (+)- nor (-)-enantiomers altered glutamate-mediated excitatory synaptic currents. At 10 microM, (+)-DHP and (+)-ACN depressed excitatory responses in a bicuculline-sensitive fashion, suggesting that direct chloride channel gating by the steroids contributed to the depression. These data indicate that certain steroids and benz[e]indenes augment inhibitory synaptic transmission enantioselectively and provide strong support for the hypothesis that steroids act at specific sites on synaptic GABA(A) receptors rather than via alteration of membrane lipids.

Pharmacological and physiological properties of a putative ganglionic nicotinic receptor, alpha 3 beta 4, expressed in transfected eucaryotic cells.

Neuronal nicotinic acetylcholine receptor subunits alpha 3 (PCA48E) and beta 4S (ZPC13) were expressed in human embryonic kidney (HEK)-293 cells by calcium phosphate transfection. In the presence of atropine, acetylcholine (ACh) induced fast activating currents which exhibited desensitization and inward rectification. The EC50 for ACh was 202 +/- 32 microM with a Hill coefficient of 1.9 +/- 0.4. The rank order of nicotinic agonist potency was 1,1-dimethyl-4-phenylpiperozinium (DMPP) > cytisine = nicotine approximately equal to ACh. The maximal response elicited by DMPP was substantially less than that elicited by other agonists, suggesting that DMPP is a partial agonist. ACh (500 microM) responses were very effectively blocked by equimolar concentrations (100 microM) of the ganglionic antagonists d-tubocurarine, mecamylamine and hexamethonium. Equal concentrations of the potent muscle receptor antagonist decamethonium and the competitive antagonist dihydro-beta-erythroidine were much less effective. alpha bungaro-toxin (1 microM) had little effect on ACh-induced responses. This physiological and pharmacological profile is consistent with a ganglionic nicotinic response.