Pulses of extracellular K+ produce two cytosolic Ca2+ transients that display different temperature dependence and carbonyl cyanide m-chlorophenyl sensitivity in SH-SY5Y cells.

In SH-SY5Y cells we have shown that stimulation with high extracellular K+ ([K+]e) evokes a transient increase in cytoplasmic Ca2+ ([Ca2+]cyt) (K+on) that is triggered by the opening of voltage-dependent Ca2+ channels and followed by Ca2+ -induced Ca2+ release from the endoplasmic reticulum (Xu, F., Zhang, J., Recio-Pinto, E. and Blanck, T.J., Halothane and isoflurane augment depolarization-induced cytosolic CA2+ transients and attenuate carbachol-stimulated CA2+ transients, Anesthesiology, 92 (2000) 1746-56). The removal of high-[K+]e results in a second transient increase in [Ca2+]cyt (K+off) that is independent of extracellular Ca2+ (Corrales, A., Montoya, G.J., Sutachan, J.J., Cornillez-Ty, G., Garavito-Aguilar, Z., Xu, F., Blanck, T.J. and Recio-Pinto, E., Transient increases in extracellular K+ produce two pharmacological distinct cytosolic Ca2+ transients, Brain Res., 1031 (2005) 174-184). In this study we further characterize the properties of K+off. We found that K+off was detectable at near physiological temperatures (34-36 degrees C) but, depending on the level of [K+]e, it was undetectable or highly diminished at room temperature. In contrast, K+on was increased by lowering the temperature. Extracellular Na+ -replacement with K+ did not affect K+off, indicating that K+off was not generated by osmolarity changes. Replacement of extracellular Na+ with choline+ did not affect K+off, indicating that K+off did not result from activity changes of the plasma membrane Na+/Ca2+ exchanger. Caffeine decreased K+on but not K+off. CCCP (carbonyl cyanide m-chlorophenyl), a protonophore uncoupler that decreases mitochondrial Ca2+ uptake, decreased K+on but not K+off. CGP37157, an inhibitor of the mitochondria Na+/Ca2+ exchanger, decreased K+off when added alone but not when added simultaneously with CCCP. Clonazepam had similar effects as CGP37157. These findings indicate that the generation of K+off is strongly temperature-dependent and its pharmacology is distinct from the [Ca2+]cyt changes observed previously at room temperature.