Measurement of GABA(A) receptor function in rat cultured cerebellar granule cells by the Cytosensor microphysiometer

1 gamma-Aminobutyric acid (GABA), acting via the GABA(A) receptor, increased the extracellular acidification rate of rat primary cultured cerebellar granule cells, measured by the Cytosensor microphysiometer. 2 The optimal conditions for the measurement of GABA(A) receptor function in cerebellar granule cells by microphysiometry were: cells seeded at 9-12x10(5) cells/transwell cup an maintained in vitro for 8 days, GABA stimulation performed at 25 degrees C, with a stimulation time of 33 s. 3 GABA stimulated a concentration-dependent increase in the extracellular acidification rate with an EC50 of 2.0+/-0.2 mu M (means+/-s.e.mean, n=7 experiments) and maximal increase (E-max) over basal response of 15.4+/-1.2%. 4 The sub-maximal GABA-stimulated increase in acidification rate could be potentiated by the 1,4-benzodiazepine, flunitrazepam (100 nM). The 10 nM GABA response showed the maximal benzodiazepine facilitation (GABA alone, 1.4 mu V s(-1), GABA+flunitrazepam, 3.8 mu V s(-1), mean increment over basal, n=7). 5 The GABA-stimulated increase in acidification rate was inhibited by the GABA(A) antagonist, bicuculline (100 mu M) (90% inhibition at 1 mM GABA). 6 The results of this study show that activation of GABA(A) receptors in rat cerebellar granule cells caused an increase in the extracellular acidification rate; an effect which was potentiated by benzodiazepines and inhibited by a GABA(A) receptor antagonist. This paper defines the conditions and confirms the feasibility of using microphysiometry to investigate GABA(A) receptor function in primary cultured CNS neurones. The microphysiometer provides a rapid and sensitive technique to investigate the regulation of the GABA(A) receptor in populations of neurones.