The Journal of Neurophysiology Vol. 79 No. 1 January 1998, pp. 439-446
Copyright ©1998 The American Physiological Society

Changes in Intracellular Ca²⁺ Induced by GABA_A Receptor Activation and Reduction in Cl Gradient in Neonatal Rat Neocortex

Atsuo Fukuda, Kanji Muramatsu, Akihito Okabe, Yasunobu Shimano, Hideki Hida, Ichiro Fujimoto, and Hitoo Nishino

Department of Physiology, Nagoya City University Medical School, Nagoya, Aichi 467, Japan

Fukuda, Atsuo, Kanji Muramatsu, Akihito Okabe, Yasunobu Shimano, Hideki Hida, Ichiro Fujimoto, and Hitoo Nishino. Changes in intracellular Ca²⁺ induced by GABA_A receptor activation and reduction in Cl gradient in neonatal rat neocortex. J. Neurophysiol. 79: 439-446, 1998. We have studied the effects of -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 µM), 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 µM). 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 µM), 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, 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.

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