Nitric oxide activates ca2+ entry via a mechanism distinct from store-operated ca2+ channels

Nitric oxide (NO) has been implicated as a mediator of Ca²⁺ entry in response to Ca²⁺ store depletion (store-operated channels; SOCs), possibly by increasing cGMP. The present studies investigated the action or NO donors and cGMP on cytosolic Ca²⁺ and revealed rapid and profound actions of NO on possible direct activation of Ca²⁺ entry channels. The potent NO donor, GEA3162, a 4-aryl-substituted oxatriazol. rapidly activated (within 1 min) a transient increase in intracellular free Ca ²⁺ in DDTMF-2 cells. The effect was due to activation of Ca ²⁺ influx since no effect of GEA3162 was observed in Ca ²⁺-free medium. After GEA3162-treatment of cells in the absence of external Ca1' reapplication of Ca²⁺ resulted in an immediate and large increase in cytosolic Ca²⁺ indicating that the entry mechanism remained in an activated state but was rapidly deactivated. To determine whether this influx was related to SOC activity, experiments investigated the effects of the imidazole derivative, SKF96365, a known SOC inhibitor. After thapsigarain treatment, SOC-induced Ca²⁺ entry activated by depletion of external Ca²⁺ was blocked by 200 μM SKF96365; in contrast, GEA3162-induced Ca²⁺ influx was unaffected by SKF96365. Moreover, the action of GEA3162 was not inhibited by the guanylate cyclase inhibitor, LY83583, suggesting that NO-induced Ca²⁺ entry is not mediated via changes in cGMP levels. No change in the level of cGMP could be detected in response to GEA3162. Permeability to other divalent cation was assessed. Ca²⁺ influx was totally blocked by La³⁺. GEA3162 induced Ni²⁺ entry but only in the presence of Ca²⁺; this indicates that Ca ²⁺ is required for activation of the NO-induced channel. The results reveal that NO can induce a rapid and profound influx of Ca²⁺ via a mechanism independent of store-filling and distinct from SOC-medlated Ca ²⁺ entry channels. The action of NO does not appear to be mediated by cGMP, and could reflect a more direct interaction perhaps via S-nitrosylation of an entry channel or regulatory proteins thereof.