The Panx1 inhibitor spironolactone attenuates HPV. Representative tracing (A) of pulmonary artery pressure (PAP) in isolated perfused mouse lungs and quantitative group data analysis (B) show attenuation of the vasoconstrictive response to hypoxia (1% O₂)—measured as increase in PAP (ΔPAP)—by spironolactone (20 µMol/L in (A); 0–40 µMol/L in (B)) in a dose-dependent manner (n = 5 per group). Time-resolved group data (C) and quantification of area-under-the-curve (AUC) (D) show attenuated increase in intracellular Ca²⁺ concentration in response to hypoxia—as measured by ratiometric imaging of Fura 2 at 340 and 380 nm (F340/F380) in hPASMC—by spironolactone as compared to vehicle (n = 4 each). Time-resolved group data (E) and quantification of area under the curve (AUC) (F) show attenuated increase in intracellular Ca²⁺ concentration in response to hypoxia in hPASMC by the Panx1 specific inhibitory peptide ¹⁰Panx1 (200 µMol/L, n = 9) as compared to scrambled peptide (n = 7). (G–J) Knockdown of Panx1 decreased both RNA as well as protein levels and led to a marked attenuation of the [Ca²⁺]_i response to hypoxia. (K–M) In hPASMC, knockdown of NR3C2 by siRNA (siNR3C2, n = 7) reduced NR3C2 mRNA expression as well as NR3C2 protein levels compared to control cells treated with non-targeting scrambled siRNA (siScr, n = 4–6) but (N) did not affect the [Ca²⁺]_i increase in response to hypoxia. Data are mean ± SEM; Data were analysed using Kruskal–Wallis test (B) or Mann–Whitney U-test (D, F, G, I, J, K, M, N); *P < 0.05; ns, not significant.