Preconditioning increases S-nitrosylation of the L-type Ca2+ channel and SERCA2a

Abstract

Nitric oxide (NO) has been shown to be an important signaling messenger in ischemic preconditioning (IPC). Accordingly, we investigated whether protein S-nitrosylation occurs in IPC hearts and whether an increase of NO production by S-nitrosoglutathione (GSNO) elicits similar effects on S-nitrosylation and protection. C57BL/6J mice (12–15 weeks) hearts were perfused in the Langendorff mode. Hearts were subjected to: (1) control perfusion; (2) ischemic preconditioning (IPC); or (3) GSNO preconditioning (GSNO-PC), followed by 20 min of no-flow ischemia and 40 min of reperfusion. Recovery of post-ischemic left ventricular developed pressure (LVDP) was measured at the end of reperfusion. Compared with control, IPC and GSNO-PC significantly improved the post-ischemic recovery of LVDP (62.1±6.7% and 54.8±7.9% vs. 36.8±4.8% in control). IPC and GSNO-PC both significantly increased the S-nitrosothiol (SNO) contents in heart membrane fractions (9.9±0.4 in IPC and 9.2±0.5 in GSNO-PC vs. 3.4±0.6 pmol/mg protein in control). IPC and GSNO-PS also both significantly increased S-nitrosylation of the L-type Ca2+ channel α1 subunit. Using a combination of biotin switch method, streptavidin chromatography, and mass spectrometry proteomic analysis, the cardiac sarcoplasmic reticulum Ca2+ ATPase (SERCA2a) was also identified as an S-nitrosylated protein in both IPC and GSNO-PC hearts. Thus, the similarity of protection and protein S-nitrosylation modification in IPC and GSNO-PC hearts suggest that protein S-nitrosylation may play an important cardioprotective role in preconditioning heart.