Abstract 2699: Mechanisms Underlying Increased Ca2+ Influx in SERCA2 Kockout Cardiomyocytes
In normal cardiomyocytes, contraction is predominantly dependent on Ca2+ release from the sarcoplasmic reticulum (SR), and reduced activity of the SR Ca2+ ATPase (SERCA) has been linked to heart failure. We have recently observed that mice with cardiomyocyte-specific excision of SERCA (KO) compensate for markedly reduced SR function by increasing trans-sarcolemmal Ca2+ flux. Here we investigated the mechanisms underlying the greater Ca2+ entry. At seven weeks following gene deletion, SR Ca2+ release was not detectable in KO cardiomyocytes. Expression of the L-type Ca2+ channel was increased in KO as protein levels of the pore forming subunit,α1C, and regulatory subunit, α2/δ1, were 178% and 147% of control values, respectively. Peak L-type Ca2+ currents were 49% larger in KO than control, and slower Ca2+ current decay kinetics resulted in integrated currents which were 220% of control values. The action potential (AP) was also prolonged in KO. Treatment with 20 μM nifedipine or 0 mM Ca2+ in the external solution abolished this difference, indicating that the longer AP resulted from increased Ca2+ current. Indeed, measurements of transient outward K+ current were similar in KO and control. Switching the voltage stimulus from the control AP to the KO AP increased the magnitude of the Ca2+ transient by 67% in KO myocytes. This augmentation predominantly resulted from increased integrated Ca2+ current. Thus, overall L-type Ca2+ entry in KO was increased from control values by 3–3.5 fold when AP alterations were considered. AP prolongation in control cells also increased Ca2+ influx, but augmentation of the Ca2+ transient was less marked (19%) due to reduced efficiency of Ca2+-induced Ca2+ release. Finally, we observed that treatment with 5 μM KB-R7943 markedly reduced Ca2+ transients in KO cells but not in controls, indicating a greater role of reverse-mode Na+/Ca2+ exchange in KO. Thus, increased expression of the Ca2+ channel, absence of Ca2+-dependent inactivation, action potential prolongation, and augmented reverse-mode Na+/Ca2+ exchange can together greatly enhance Ca2+ influx when SR function is reduced. These observations suggest that reduced SR function in human heart failure may be treated by strategies which augment sarcolemmal Ca2+ fluxes.