CHANGES IN INTRACELLULAR CA2- GRADIENT IN NEONATAL RAT NEOCORTEX( INDUCED BY GABA(A) RECEPTOR ACTIVATION AND REDUCTION IN CL)

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 pyram idal neurons of rat somatosensory cortex. The Cl-gradient was reduced either with furosemide or by oxygen glucose deprivation. Immature slic es 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 th e ratio of fluorescence intensities (R-F340/F380) The magnitude of the [Ca2+](i) increases induced by oxygen-glucose deprivation was signifi cantly reduced (by 44%) by bicuculline (10 mu M), a GABA(A) receptor a ntagonist. 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). Continu ous 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 bicuculli ne(10 mu M). The initial but not the sustained phase was abolished by Ni2+. In the presence of Ni2+, the remaining sustained response was in hibited by the addition of 2-amino-5-phosphonopentanoic acid (AP5, 20 mu M), a selective N-methyl-D-aspartate (NMDA) receptor antagonist. Th us the initial peak and the sustained phase of the GABA-evoked [Ca2+]( i) increase were mediated by Ca2+ influx through VDCCs and NMDA recept or channels, respectively, and both phases were initiated via the GABA (A) receptor. These results indicate that, in neocortical pyramidal ne urons, a reduction in the Cl-gradient converts the GABA(A) receptor-me diated action from nothing or virtually nothing to a large and sustain ed accumulation of cellular Ca2+. This accumulation is the result of C a2+ influx mainly through the NMDA receptor channel. Thus GABA(A) norm ally an inhibitory transmitter, may play an aggravating role in excito toxicity if a shift in the Cl-equilibrium potential occurs, as reporte d previously, during cerebral ischemia.