In pulmonary artery smooth muscle cells (PASMCs) there is a store-operated Ca²⁺ entry (SOCE) pathway that is activated by depletion of sarcoplasmic reticulum (SR) Ca²⁺ (Ng & Gurney 2001). Nitric oxide (NO) is an important mediator of vascular contractility and from studies on cultured vascular smooth muscle cells it has been shown to inhibit SOCE (Cohen et al., 1999; Moneer et al., 2003). The aim of this study was therefore to investigate the effects of NO donors upon SOCE in freshly isolated smooth muscle cells from rat intrapulmonary arteries.Male Sprague-Dawley rats (200-300g) were killed by cervical dislocation and PASMCs isolated from intrapulmonary arteries (300-500µm outside diameter) using methods similar to those described by Drummond & Tuft (1999). PASMCs were incubated with 5µM fluo-4 AM to monitor intracellular Ca²⁺. SR store-depletion was achieved by incubation with 1µM thapsigargin in the absence of extracellular Ca²⁺.Re-addition of Ca²⁺ resulted in an increase in fluo-4 fluorescence, visualised using confocal microscopy. All experiments were carried out at room temperature. Where appropriate mean data ± S.E.M are given and n is the number of cells studied. Statistical analysis employed Student’s unpaired t test, with P < 0.05 considered to be significant. Application of Ca²⁺ to store-depleted cells resulted in a sustained increase in fluo-4 fluorescence (∆F/F₀ = 2.2 ± 0.2, n = 18). This was significantly greater than the increase in fluo-4 fluorescence observed upon re-addition of Ca²⁺ to cells deprived of extracellular Ca²⁺ for the same period in the absence of thapsigargin (∆F/F₀ = 0.5 ± 0.1, n = 16). The thapsigargin-induced increase in fluorescence was not affected by the L-type Ca²⁺ channel blocker verapamil (10µM). It was however inhibited by 45% ± 6 (n = 20) when the IP₃ receptor and SOCE inhibitor 2aminoethoxydiphenyl borate (2-APB, 75µM) was present. Consistent with previous studies, this indicates that the thapsigargin induced increase in fluorescence was due to SOCE. Neither sodium nitroprusside (10µM) nor glyceryl trinitrate (100µM) significantly affected the SOCE-dependent increase in fluo-4 fluorescence resulting from Ca²⁺ re-addition. The thapsigargin induced increase in fluorescence was reduced by only 18% ± 6 (n = 26) by sodium nitroprusside and by 14% ± 7 (n = 28) by glyceryl trinitrate. These results suggest that NO has no direct effect on the channels underlying the SOCE pathway in freshly isolated PASMCs.