Influence of GABAA receptor gamma2 splice variants on receptor kinetics and isoflurane modulation

Gamma-aminobutyric acid type A (GABAA) receptors, the major inhibitory receptors in the brain, are important targets of many drugs, including general anesthetics. These compounds exert multiple effects on GABAA receptors, including direct activation, prolongation of deactivation kinetics, and reduction of inhibitory postsynaptic current amplitudes. However, the degree to which these actions occur differs for different agents and synapses, possibly because of subunit-specific effects on postsynaptic receptors. In contrast to benzodiazepines and intravenous anesthetics, there is little information available about the subunit dependency of actions of volatile anesthetics. Therefore, the authors studied in detail the effects of isoflurane on recombinant GABAA receptors composed of several different subunit combinations.

Human embryonic kidney 293 cells were transiently transfected with rat complementary DNAs of alpha1beta2, alpha1beta2gamma2L, alpha1beta2gamma2S, alpha5beta3, or alpha5beta3gamma2S subunits. Using rapid application and whole cell patch clamp techniques, cells were exposed to 10- and 2,000-ms pulses of gamma-aminobutyric acid (1 mm) in the presence or absence of isoflurane (0.25, 0.5, 1.0 mm). Anesthetic effects on decay kinetics, peak amplitude, net charge transfer and rise time were measured. Statistical significance was assessed using the Student t test or one-way analysis of variance followed by the Tukey post hoc test.

Under control conditions, incorporation of a gamma2 subunit conferred faster deactivation kinetics and reduced desensitization. Isoflurane slowed deactivation, enhanced desensitization, and reduced peak current amplitude in alphabeta receptors. Coexpression with a gamma2 subunit caused these effects of isoflurane to be substantially reduced or abolished. Although the two gamma2 splice variants imparted qualitatively similar macroscopic kinetic properties, there were significant quantitative differences between effects of isoflurane on deactivation and peak current amplitude in gamma2S- versus gamma2L-containing receptors. The net charge transfer resulting from brief pulses of gamma-aminobutyric acid was decreased by isoflurane in alphabeta but increased in alphabetagamma receptors.

The results indicate that subunit composition does substantially influence modulation of GABAA receptors by isoflurane. Specifically, the presence of a gamma2 subunit and the identity of its splice variant are important factors in determining physiologic and pharmacologic properties. These results may have functional implications in understanding how anesthetic effects on specific types of GABAA receptors in the brain contribute to changes in brain function and behavior.