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J Physiol Volume 525, Number 2, 343-353, June 1, 2000
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The Journal of Physiology (2000), 525.2, pp. 343-353
© Copyright 2000 The Physiological Society

Regulation of Na+,K+-ATPase by persistent sodium accumulation in adult rat thalamic neurones

Vladimir V. Senatorov, Peter K. Stys and Bin Hu

Loeb Health Research Institute, Ottawa Hospital, University of Ottawa, Ottawa, Ontario, Canada K1Y 4E9

  1. The present study investigated the regulatory mechanism of the Na+,K+-ATPase and the level of internal Na+ and Ca2+ in response to persistent Na+ influx in acutely dissociated rat thalamic neurones.

  2. Whole-cell patch-clamp recordings and Na+ imaging revealed a stable [Na+]i and low background pump activity. Exposure to veratridine (50 µM) for 1 h resulted in a progressive rise in [Na+]i (DeltaFNa = 64 ± 22 %) and [Ca2+]i (DeltaFCa = 44 ± 14 %) over 3 h. Increases in [Na+]i and [Ca2+]i were also observed during neuronal exposure to the Na+ ionophore monensin (50 µM).

  3. Subcellular confocal immunofluorescence quantification of alpha3 catalytic Na+-K+ pump subunits showed that a veratridine-induced rise in [Na+]i was accompanied by a significant increase in pump density in both membrane and cytoplasmic compartments, by 39 and 54 %, respectively. Similar results were also obtained in experiments when neurones were treated with monensin.

  4. A fluorescent 9-anthroylouabain binding assay detected a 60 and 110 % increase in phosphorylated (active) pumps after veratridine and monensin exposure, respectively.

  5. During the entire experiment, application of ouabain or veratridine alone induced little cell swelling and death, but pump inhibition in cells pre-loaded with Na+ led to rapid cell swelling and necrosis.

  6. The above results indicate that a persistent influx of Na+ may trigger rapid enhancement of pump synthesis, membrane redistribution and functional activity. However, these compensatory mechanisms failed to prevent persistent Na+ accumulation.



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