Mice.

The generation of floxed alleles of the genes encoding IP₃R1, IP₃R2, and IP₃R3 has been described previously (15, 37). We used two different tissue-specific Cre recombinases to delete IP₃Rs in VSMCs. First, we crossed IP₃R1 flox (IP₃R1^f/f) mice or IP₃R triple flox (IP₃R1^f/fIP₃R2^f/fIP₃R3^f/f) mice with Tg(Tagln-cre)1Her/J (Sm22α-Cre) mice (The Jackson Laboratory), respectively. IP₃R triple flox mice had been backcrossed with C57BL/6 mice for more than 7 generations. Sm22α-Cre is constitutively expressed and has been frequently used as a smooth muscle–specific Cre (38). Second, mice with inducible deletion of all 3 IP₃R genes were generated by crossing IP₃R triple flox mice with B6.FVB-Tg(Myh11-Cre/ERT2)1Soff/J (SMMHC-CreER^T2) mice (The Jackson Laboratory). Since the SMMHC-Cre gene is located on the Y chromosome in this mouse line, only male offspring carried the SMMHC-Cre gene (25). Thus, in our study, we characterized only male mice. Male IP₃R1^f/fIP₃R2^f/f IP₃R3^f/f/SMMHC-Cre⁺ mice aged 7 to 8 weeks were intraperitoneally injected with tamoxifen (50 mg/kg/d) for 5 consecutive days to delete IP₃R genes and were considered as considered as smTKO mice. Male IP₃R1^f/fIP₃R2^f/f IP₃R3^f/f/SMMHC-Cre^– mice treated with tamoxifen and male noninduced smTKO mice were used as control mice. All mice were housed under a 12-hour-day/night cycle at a temperature of 25°C.