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

1. The present study investigated the regulatory mechanism of the Na+,K+-AT Pase and the level of internal Na+ and Ca2+ in response to persistent Na+ i nflux 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 mu M) fo r 1 h resulted in a progressive rise in [Na+](i) (Delta F-Na = 64 +/- 22%) and [Ca2+](i) (Delta F-Ca = 44 +/- 14%) over 3 h. Increases in [Na+](i) and [Ca2+](i) were also observed during neuronal exposure to the Na+ ionophore monensin (50 mu M). 3. Subcellular confocal immunofluorescence quantification of alpha 3 cataly tic Na+-K+ pump subunits showed that a. veratridine-induced rise in [Na+](i ) was accompanied by a significant increase in pump density in both membran e and cytoplasmic compartments, by 39 and 54%, respectively. Similar result s were also obtained in experiments when neurones were treated with monensi n. 4. A fluorescent 9-anthroylouabain binding assay detected a 60 and 110% inc rease in phosphorylated (active) pumps after veratridine and monensin expos ure, respectively. 5. During the entire experiment, application of ouabain or veratridine alon e 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 r apid enhancement of pump synthesis, membrane redistribution and functional activity. However, these compensatory mechanisms failed to prevent persiste nt Na+ accumulation.