Adenoviral SERCA1a gene transfer to adult rat ventricular myocytes induces physiological changes in calcium handling.

Objective: We examined the functional consequences of expressing adult rabbit fast skeletal sarcoplasmic reticulum (SR) Ca2+-ATPase (SERCA1a) in isolated adult rat ventricular myocytes. Methods: Myocytes were infected with a recombinant adenovirus harboring SERCA1a. Then 2 days after myocyte infection, protein expression was estimated using Western blot and SDS–PAGE analysis. We also measured the ATP-dependent oxalate-facilitated Ca2+ uptake of myocyte homogenates and monitored Ca2+ transient in myocytes loaded with the Ca2+ dye, indo-1. Results: SERCA1a gene expression resulted in a 36% increase in the total SERCA protein level in infected myocytes compared to controls ( P <0.01), while SERCA2 and phospholamban levels did not change. This increase was associated with a 42% rise in SR Ca2+ uptake ( P <0.01), while τ (the time constant of Ca2+ transient decay), and the time to peak fell by 32% ( P <0.01) and 38% ( P <0.001), respectively. Increasing the frequency of stimulation from 0.2 to 2 Hz decreased τ in both cell types ( P <0.01). However, the decrease was much smaller in infected ( P <0.01) than in uninfected cells ( P <0.001). Isoproterenol (1 μM) further decreased τ in infected myocytes by 23% ( P <0.05). In these cells, the diastolic [Ca2+]i decreased by 50% ( P <0.05) while the systolic [Ca2+]i increased by 19% ( P <0.05). No difference was found in the speed of SR Ca2+ reloading after caffeine washout between the two cell types. Conclusion: Adenovirus-mediated SERCA1a gene transfer to adult rat ventricular myocytes enhances SR Ca2+ handling to a degree similar to that observed following physiological stimulation.