MECHANISMS OF ANOXIA-INDUCED DEPOLARIZATION IN BRAIN-STEM NEURONS - IN-VITRO CURRENT AND VOLTAGE-CLAMP STUDIES IN THE ADULT-RAT

To determine the mechanisms underlying the depolarization induced by a noxia in brainstem neurons, we studied single neurons in brainstem sli ces using conventional micro-electrodes and freshly dissociated hypogl ossal and vagal cells using patch clamp techniques (whole-cell configu ration). Since glutamate concentration increases in the extracellular space during 02 deprivation, we first tested whether N-methyl-D-aspara te (NMDA) and non-NMDA receptors are involved in this anoxia-induced d epolarization. APV, MK-801, CNQX and KYN (NMDA and non-NMDA blockers), which bathed slices after control anoxia runs, did not affect the dep olarization trajectory. Decreasing extracellular Na+ (Na(o)+) from 150 mM to 5 mM attenuated markedly and significantly the depolarization o bserved during anoxia (15-20% of control). The relation between intrac ellular adenosine triphosphate (ATP) and the anoxia-induced depolariza tion was also investigated in the slice and in dissociated single brai nstem neurons. In the slice, iontophoresis of ATP did not give consist ent results. Since we could not ascertain that ATP was actually iontop horesed through high resistance (50-80 MOMEGA) microelectrodes, we pat ched single neurons and studied the effect of clamping intracellular A TP levels on the hyperpolarizing holding current (I(H)) in the voltage clamp mode. The increase in I(H) with anoxia (or cyanide) was markedl y attenuated in cells patched with pipettes containing ATP. We conclud e that in brainstem neurons, the anoxia-induced depolarization: (a) is not a function of an increase in extracellular glutamate concentratio n; and (b) depends on Na+ and ATP-mediated processes.