Changes in intracellular Ca2+ induced by GABAA receptor activation and reduction in Cl- gradient in neonatal rat neocortex

Changes in intracellular Ca2+ induced by GABA, receptor activation and reduction in Cl- gradient in neonatal rat neocortex. J. Neurophysiol. 79: 439-446, 1998. We have studied the effects of gamma-aminobutyric acid (GABA) and of reducing the Cl- gradient on the [Ca2+](i) in pyramidal neurons of rat somatosensory cortex. The Cl-gradient was reduced either with furosemide or by oxygen glucose deprivation. Immature slices taken at postnatal day (P) 7-14 were labeled with fura-2, and [Ca2+](i) was monitored in identified pyramidal cells in layer II/III as the ratio of fluorescence intensities (R-F340/F380) The magnitude of the [Ca2+](i) increases induced by oxygen-glucose deprivation was significantly reduced (by 44%) by bicuculline (10 mu M), a GABA(A) receptor antagonist. Under normal conditions, GABA generally did not raise [Ca2+](i), although in some neurons a small and transient [Ca2+]i increase was observed. These transient [Ca2+](i) increases were blocked by Ni2+ (1 mM), a blocker of voltage-dependent Ca2+ channels (VDCCs). Continuous perfusion with GABA did not cause a sustained elevation of [Ca2+](i) but bicuculline caused [Ca2+](i) oscillations. After inhibition of Cl-extrusion with furosemide (1.5 mM), GABA induced a large [Ca2+](i) increase consisting of an initial peak followed by a sustained phase. Both the initial and the sustained phases were eliminated by bicuculline(10 mu M). The initial but not the sustained phase was abolished by Ni2+. In the presence of Ni2+, the remaining sustained response was inhibited by the addition of 2-amino-5-phosphonopentanoic acid (AP5, 20 mu M), a selective N-methyl-D-aspartate (NMDA) receptor antagonist. Thus the initial peak and the sustained phase of the GABA-evoked [Ca2+](i) increase were mediated by Ca2+ influx through VDCCs and NMDA receptor channels, respectively, and both phases were initiated via the GABA(A) receptor. These results indicate that, in neocortical pyramidal neurons, a reduction in the Cl-gradient converts the GABA(A) receptor-mediated action from nothing or virtually nothing to a large and sustained accumulation of cellular Ca2+. This accumulation is the result of Ca2+ influx mainly through the NMDA receptor channel. Thus GABA(A) normally an inhibitory transmitter, may play an aggravating role in excitotoxicity if a shift in the Cl-equilibrium potential occurs, as reported previously, during cerebral ischemia.