Na+ and K+ Concentrations, Extra- and Intracellular Voltages, and the Effect of TTX in Hypoxic Rat Hippocampal Slices

Department of Cell Biology, Duke University Medical Center, Durham, North Carolina 27710 Müller, Michael and George G. Somjen. Na + and K + Concentrations, Extra- and Intracellular Voltages, and the Effect of TTX in Hypoxic Rat Hippocampal Slices. J. Neurophysiol. 83: 735-745, 2000. Severe hypoxia causes rapid depolarization of CA1 neurons and glial cells that resembles spreading depression (SD). In brain slices in vitro, the SD-like depolarization and the associated irreversible loss of function can be postponed, but not prevented, by blockade of Na + currents by tetrodotoxin (TTX). To investigate the role of Na + flux, we made recordings from the CA1 region in hippocampal slices in the presence and absence of TTX. We measured membrane changes in single CA1 pyramidal neurons simultaneously with extracellular DC potential ( V o ) and either extracellular [K + ] or [Na + ]; alternatively, we simultaneously recorded [Na + ] o , [K + ] o , and V o . Confirming previous reports, early during hypoxia, before SD onset, [K + ] o began to rise, whereas [Na + ] o still remained normal and V o showed a slight, gradual, negative shift; neurons first hyperpolarized and then began to gradually depolarize. The SD-like abrupt negative V o corresponded to a near complete depolarization of pyramidal neurons and an 89% decrease in input resistance. [K + ] o increased by 47 mM and [Na + ] o dropped by 91 mM. Changes in intracellular Na + and K + concentrations, estimated on the basis of the measured extracellular ion levels and the relative volume fractions of the neuronal, glial, and extracellular compartment, were much more moderate. Because [Na + ] o dropped more than [K + ] o increased, simple exchange of Na + for K + cannot account for these ionic changes. The apparent imbalance of charge could be made up by Cl influx into neurons paralleling Na + flux and release of Mg 2+ from cells. The hypoxia-induced changes in interneurons resembled those observed in pyramidal neurons. Astrocytes responded with an initial slow depolarization as [K + ] o rose. It was followed by a rapid but incomplete depolarization as soon as SD occurred, which could be accounted for by the reduced ratio, [K + ] i /[K + ] o . TTX (1 µM) markedly postponed SD, but the SD-related changes in [K + ] o and [Na + ] o were only reduced by 23 and 12%, respectively. In TTX-treated pyramidal neurons, the delayed SD-like depolarization took off from a more positive level, but the final depolarized intracellul