Fig. 5

Effect of TRPV4 agonist on calpain 1-NLRP3/cas-1-GSDMD pathway in the hippocampus. A. The TRPV4 agonist GSK1016790A decreased inactive/total calpain 1 ratio (independent-samples t test, t = 12.72, P < 0.01) but did not change inactive/total calpain 2 ratio (independent-samples t test, t = 0.19, P = 0.86) in the hippocampus. B and C. GSK1016790A increased NLRP3 (B, Mann–Whitney U test, P < 0.01), c-cas-1 (B, Mann–Whitney U test, P < 0.01), IL-1β (B, Mann–Whitney U test, P < 0.01), and N-GSDMD (B, Mann–Whitney U test, P < 0.01) protein levels, and GSDMD⁺ cells (red arrow) number (C, independent-samples t test, t = − 20.96, P < 0.01) in the hippocampus. D. GSDMD⁺/NeuN⁺ (D-i, white arrow), GSDMD⁺/GFAP⁺ (D-ii, white star) and GSDMD⁺/Iba-1⁺ (D-iii, white triangle) cells in the hippocampus of control and GSK1016790A-injected mice. E and F. The calpain inhibitor MDL-28170 increased inactive/total calpain 1 ratio (E, Mann–Whitney U test, P < 0.01) and decreased c-cas-1 (F, independent-samples t test, t = 8.73, P < 0.01), IL-1β (F, independent-samples t test, t = 9.35, P < 0.01), and N-GSDMD protein levels (F, independent-samples t test, t = 13.52, P < 0.01) but did not affect NLRP3 protein levels (F, independent-samples t test, t = − 0.20, P = 0.85) in the hippocampus of GSK1016790A-injected mice. G and H. The NLRP3 inhibitor MCC950 (G) and cas-1 inhibitor Ac-YVAD-cmk (H) decreased c-cas-1 (G, independent-samples t test, t = 8.17, P < 0.01; H, Mann–Whitney U test, P < 0.01), IL-1β (G, independent-samples t test, t = 11.62, P < 0.01; H, Mann–Whitney U test, P < 0.01) and N-GSDMD protein levels (G, Mann–Whitney U test, P < 0.01; H, independent-samples t test, t = 13.38, P < 0.01) in the hippocampus in GSK1016790A-injected mice. Scale = 50 μm. ×4 objective for the top row in D-i, D-ii and D-iii; ×10 objective for C-DAPI, C-GSDMD and C-Merge; ×20 objective for C-enlarge, the down row in D-i, D-ii and D-iii. ^<<P < 0.01 vs. Control (icv.), ^$$P < 0.01 vs. GSK1016790A + vehicle, ^%%P < 0.01 vs. GSK1016790A + vehicle