The molecular mechanisms which underlie the rapid and dramatic central depressant actions of intravenous general anaesthetics are not known. Intravenous agents such as propofol, etomidate, thiopentone and alphaxalone are structurally diverse and hence it is difficult to conceive that they might act through by common mechanism. However, surprisingly a number of studies have now demonstrated that many clinical and experimental intravenous anaesthetics, at clinically relevant concentrations, share the property of enhancing the actions of GABA acting at the GABA_A receptor. Although such observations do not exclude alternative receptors or ion channels as anaesthetic targets, the enhancement of neuronal inhibition by an action on the major inhibitory receptor in the central nervous system has a logical appeal.

Utilizing the Xenopus laevis oocyte expression system we have compared the actions of propofol, etomidate, pentobarbitone and alphaxalone on GABA_A , glycine, neuronal nicotinic, 5-HT₃, NMDA and AMPA receptors. These studies demonstrate etomidate, propofol and alphaxalone to be selective for the GABA_A receptor. Similarly pentobarbitone also enhances the actions of GABA acting at the GABA_A receptor although it additionally has actions at other receptors (eg. neuronal nicotinic). These studies reinforce other work highlighting the GABA_A receptor as an important molecular target for these agents.

The GABA_A receptor exists as a pentameric arrangement of subunits drawn from five families from a_1-6, b_1-3, g_1-3, d, e and f .These subunits exhibit distinct expression patterns within the central nervous system. The functional properties of the receptors that these different subunits form are unlikely to be homogeneous, but may be brain region ,or indeed neurone specific. Hence, we have investigated whether general anaesthetics interact identically with different GABA_A receptor isoforms. Of particular interest was the specificity of etomidate which produced a large GABA modulatory effect when acting at receptors composed of a_6b2g2 or _a6b3g2 subunits. By contrast, this anaesthetic had little effect on a_6b1g2 receptors. By creating a series of chimaeric b_1-2 and b_2-1 constructs, together with site-directed mutagenesis we demonstrated that this selectivity for b₂- or b₃- cf. _b1- subunit containing receptors resides within a single amino acid residue (asparagine [N] for b₂ and b₃ and serine [S] for b₁) which is located in the transmembrane region 2 (TM2) of the subunit, a part of the protein that is thought to contribute to the lining of the chloride ion channel. We are currently performing experiments to determine whether or not this amino acid forms part of a general anaesthetic binding pocket within the b subunit, or alternatively is essential for permitting the effects of the binding of etomidate to be transduced.

Irrespective of whether or not this key amino acid influences anaesthetic binding or transduction, these observations suggest future experiments in animals genetically engineered to express etomidate insensitive b subunits. Such experiments should clarify whether or not the GABA_A receptor is an important target for mediating the behavioural effects of general anaesthetics. A similar approach is currently being successfully utilized to determine which GABA_A receptor isoforms mediate the anxiolytic and sedative effects of diazepam.

References:

Belelli, D. et al. (1999). General anaesthetic action at transmitter-gated inhibitory amino acid receptors. Trends Pharmacol. Sci. 20, 496-502.

McKernan R.M. et al. (2000). Sedative but not anxiolytic properties of benzodiazepines are mediated by the GABA_A receptor a₁ subtype. Nature Neuroscience 3, 587-592.

Low K. et al. (2000). Molecular and neuronal substrate for the selective attenuation of anxiety. Science 290, 131-134.