The interaction of general anaesthetics with recombinant GABA A and glycine receptors expressed in Xenopus laevis oocytes: a comparative study

1 The effects of five structurally dissimilar general anaesthetics were examined in voltage‐clamp recordings of agonist‐evoked currents mediated by recombinant γ‐aminobutyric acid (GABA) A receptors composed of human α 1 β 1 and γ 2L subunits expressed in Xenopus laevis oocytes. A quantitative comparison of the effects of these agents was made upon recombinant glycine receptors expressed as a homo‐oligomer of human α 1 subunits, or as a hetero‐oligomer of human α 1 and rat β subunits. 2 Complementary RNA‐injected oocytes expressing GABA A receptors responded to bath applied GABA with an EC 50 of 158±34 μ m . Oocytes expressing α 1 and α 1 β glycine receptors subsequent to cDNA injection displayed EC 50 values of 76±2 μ m and 66±2 μ m , respectively, in response to bath applied glycine. 3 Picrotoxin antagonized responses mediated by homo‐oligomeric α 1 glycine receptors with an IC 50 of 4.2±0.8 μ m . Hetero‐oligomeric α1β glycine receptors were at least 100‐fold less sensitive to blockade by picrotoxin. 4 With the appropriate agonist EC 10 , propofol enhanced GABA and glycine‐evoked currents to approximately the maximal response produced by a saturating concentration of either agonist (i.e. I max ). The calculated EC 50 values were 2.3±0.2 μ m , 16±3 μ m and 27±2 μ m , for GABA A α 1 β 1 γ 2L , glycine α 1 and α 1 β receptors, respectively. At relatively high concentrations, propofol was observed to activate directly both GABA A and glycine receptors. 5 Pentobarbitone potentiated GABA‐evoked currents to 117±8.5% of I max with an EC 50 of 65±3 μ m . The barbiturate also produced a substantial enhancement of the glycine‐evoked currents, I max and EC 50 values being 71±2% and 845±66 μ m and 51±10% and 757±30 μ m for homomeric α1 and heteromeric α1β glycine receptors respectively. At high concentrations, pentobarbitone directly activated GABA A , but not glycine, receptors. 6 The potentiation by propofol or pentobarbitone of currents mediated by α 1 homo‐oligomeric glycine receptors was in both cases associated with a parallel sinistral shift of the glycine concentration‐effect curve. The effects of binary combinations of pentobarbitone and propofol at maximally effective concentrations were mutually occlusive suggesting a common site, or mechanism, of action. 7 GABA‐evoked currents were maximally potentiated by etomidate to 79±2% of I max (EC 50 of 8.1±0.9 μ m ). By contrast, glycine‐induced currents mediated by α 1 and α 1 β glycine receptor isoforms were enhanced only to 29±4% and 28±3% of I max . Limited solubility precluded the calculation of EC 50 values for the effect of etomidate at glycine receptors. None of the receptor isoforms examined were directly activated by etomidate. 8 The neurosteroid 5α‐pregnan‐3α‐ol‐20‐one potentiated GABA‐evoked currents to 69±4% of I max , with an EC 50 value of 89±6 n m . In contrast, both α 1 homo‐oligomeric and α 1 β heter‐oligomeric glycine receptors were insensitive to the action of this steroid. A direct agonist action of the steroid was discernible at GABA A , but not glycine, receptors. 9 Trichloroethanol, the active metabolite of the general anaesthetic chloral hydrate, enhanced glycine‐evoked currents to 77±10% and 94±4% of I max on α 1 and α 1 β glycine receptors, with EC 50 values of 3.5±0.1 m m and 5.9±0.3 m m respectively. On GABA A receptors, trichloroethanol had a lower maximum enhancement (52±5% of I max ), but a slightly higher potency (EC 50 1.0±0.1 m m ). Trichloroethanol activated neither GABA A , nor glycine, receptors. 10 The data demonstrate a variety of intravenous general anaesthetic agents, at clinically relevant concentrations, to augment preferentially GABA‐evoked currents mediated by the α 1 β 1 γ 2L receptor subunit combination as compared to their effects on both α 1 and α 1 β glycine receptors. However, the presence on glycine receptors of lower affinity modulatory binding sites for pentobarbitone, propofol and trichloroethanol may aid in the identification of the molecular determinants of the CNS actions of these anaesthetics.