Modifications of Ca2+Mobilization and Noradrenaline Release byS-Nitroso-cysteine in PC12 Cells1

The effects of nitrogen monoxide (NO)-related compounds on cytosolic free Ca2+concentrations ([Ca2+]i) and noradrenaline (NA) release in neurosecretory PC12 cells were investigated. The addition ofS-nitroso-cysteine (SNC) stimulated [Ca2+]iincreases from an intracellular Ca2+pool continuously in a concentration-dependent manner. Other NO donors, which stimulate cyclic GMP accumulation, did not cause [Ca2+]iincreases. After treatment with 0.2 mM SNC, transient increases in [Ca2+]ifrom the Ca2+pool induced by caffeine were completely abolished. The addition ofN-ethylmaleimide (NEM) caused sustained [Ca2+]iincreases from the intracellular Ca2+pool. Furthermore, caffeine did not stimulate further [Ca2+]iincreases in PC12 cells pretreated with NEM. These findings suggest that SNC and NEM predominantly interact with a caffeine-sensitive Ca2+pool. The addition of dithiothreitol (DTT) to 0.4 mM SNC-stimulated cells reduced [Ca2+]ito basal levels, and the addition of DTT to NEM-stimulated cells locked [Ca2+]iat high levels. The stimulatory effects of SNC but not NEM were not abolished by pretreatment with DTT. These findings suggest that modification of the oxidation status of the sulfhydryl groups on the caffeine-sensitive receptors by SNC or NEM regulates Ca2+channel activity in a reversible manner. SNC did not stimulate NA release by itself but did inhibit ionomycin-stimulated NA release. In contrast, NEM stimulated NA release in the absence of extracellular CaCl2and further enhanced ionomycin-stimulated NA release. Ca2+mobilization by SNC from the caffeine-sensitive pool was not a sufficient factor, and other factors stimulating NA release may be negatively regulated by SNC.