Figure 5. Akt→SR adenoviral expression improves SR Ca²⁺_ihandling, release and re-uptake during acute LT toxicity.

A, NRVM were infected with the indicated adenoviral constructs (SERCA 2a, DN-PLB, Akt→SR or HA-B56α) for 36 h prior to either vehicle (distilled water) or 2 h LT (0.05 ng/mL PA + 0.025 ng/mL LF) treatment. Ratiometric Ca²⁺_i determination by Fluo-4/Fura-Red fluorescence was normalized to baseline fluorescence (dye only). Lysates harvested from NRVM infected with an empty expression vector were used as the virus control. B, Representative immunoblots of DN-PLB, SERCA 2a. Akt→SR or empty vector adenoviral expression in the presence or absence of LT treatment reveals that both SERCA 2a and Akt→SR expression induces phosphorylation of PLB-T¹⁷ . C, Representative Ca²⁺ transients monitored by Fura-2 ratiometic (360/380 nm) fluorescence imaging for CMs field-stimulated at 3.0 Hz are depicted for adenovirus-treated (ad.) CMs in the absence of LT (left) and in the presence of LT 2 h treatment (right). Expression of empty vector (EV) adenovirus serves as virus control (top) for CA-MEK7 (middle) and Akt→SR (bottom). D, Group data for Ca²⁺ transient peak height [ΔF_(360/380)], tau, velocities of Ca²⁺ release and re-uptake for EV (virus control), CA-MEK7 and Akt→SR after 2 h LT-treatment of CMs field-stimulated at 1, 2 and 3 Hz. Data are averaged from n=5 dispersions, averaging 4 groups of CMs and 5-10 transients per group. *P<0.05 and ^§P<0.001 versus virus control.