Differential regulation of synaptic GABAA receptors by cAMP-dependent protein kinase in mouse cerebellar and olfactory bulb neurones.

Abstract

1. It has been demonstrated that the regulation of recombinant GABAA receptors by phosphorylation depends on the subunit composition. Here we studied the regulation of synaptic GABAA receptor function by cAMP-dependent protein kinase (PKA) in neurones expressing distinct receptor subtypes. 2. Light microscopic immunocytochemistry revealed that granule cells of the olfactory bulb express only the beta3 as the beta subunit variant, whereas cerebellar stellate and basket cells express only the beta2 as the beta subunit. 3. In cerebellar interneurones, intracellular application of 20 microM microcystin, a protein phosphatase 1/2A inhibitor, prolonged (63 +/- 14 %; mean +/- s.e.m.) the decay time course of miniature IPSCs (mIPSCs) without significantly affecting their amplitude, rise time and frequency. The effect of microcystin could be blocked by co-applying PKA inhibitory peptide (PKA-I, 1 microM). 4. No significant changes in any of the mIPSC parameters could be detected after intracellular application of PKA-I alone or following the inhibition of calcineurin with FK506 (50 nM). 5. In granule cells of the olfactory bulb expressing the beta3 subunit fast and slowly rising mIPSCs were detected, resulting in a bimodal distribution of the 10-90 % rise times, suggesting two distinct populations of events. Fast rising mIPSCs (mIPSCFR) had a 10-90 % rise time of 410 +/- 50 micros, an amplitude of 68 +/- 6 pA, and a weighted decay time constant (tauw) of 15.8 +/- 2.9 ms. In contrast, slowly rising mIPSCs (mIPSCSR) displayed an approximately threefold slower rise time (1.15 +/- 0.12 ms), 57 % smaller amplitude (29 +/- 1.7 pA), but had a tauw (16.8 +/- 3.0 ms) similar to that of the fast events. 6. mIPSCs in olfactory granule cells were not affected by the intracellular perfusion of microcystin. In spite of this, intracellular administration of constitutively active PKA caused a small, gradual, but significant increase (18 +/- 5 %) in the amplitude of the events without changing their time course. 7. These findings demonstrate a cell-type-dependent regulation of synaptic inhibition by protein phosphorylation. Furthermore, our results show that the effect of PKA-mediated phosphorylation on synaptic inhibition depends upon the subunit composition of postsynaptic GABAA receptors.