Secondary P2X1 receptor activation is resistant to nitric oxide but is substantially attenuated by increased ectonucleotidase activity. [Ca²⁺]_i responses (A-E) and ATP secretion (F) induced by a range of agonists (U46619 [U4], thrombin [Thr], collagen [Coll], Pam₃CSK₄ [Pam], and α,βmeATP) were studied under control conditions and after exposure to either spermine NONOate (sNO; 100μM) or increased apyrase (3.2 U mL⁻¹). In addition, the contribution of P2X1 receptors was assessed by desensitization before addition of external Ca²⁺ using 0.6μM α,βmeATP. Asterisks indicate the significance level compared with control samples in the absence of sNO, and hash symbols indicate the significance level compared with samples in the presence of sNO (E) or compared with thrombin in the presence of sNO (F).