Phospholamban is required for CaMKII-dependent recovery of Ca transients and SR Ca reuptake during acidosis in cardiac myocytes

Initially during acidosis, Ca transient amplitude (Δ[Ca] _i) and the rate constant of [Ca]_i decline (k _Ca) are decreased, but later during acidosis Δ[Ca]_i and k_Ca partially recover. This recovery in rat myocytes could be inhibited by KN-93 suggesting that CaMKII-dependent protein phosphorylation (and enhanced SR Ca uptake) may be responsible. To test whether phospholamban (PLB) is required for the Δ[Ca]_i and k_Ca recovery during acidosis, we used isolated myocytes from PLB knockout (PLB-KO) vs. wild-type (WT) mice. [Ca]_i was measured using fluo-3. During the initial phase of acidosis (1-4 min), Δ[Ca]_i decreased in WT myocytes (n = 8) from 1.75 ± 0.19 to 1.10 ± 0.13 ΔF/F ₀ (P < 0.05) and k_Ca decreased from 3.20 ± 0.22 to 2.38 ± 0.18 s^-1 (P < 0.05). Later during acidosis (6-12 min), Δ[Ca]_i partially recovered to 1.41 ± 0.18 ΔF/F₀ and k_Ca to 2.78 ± 0.22 s^-1 (i.e. both recovered by ∼50%). CaMKII inhibition using KN-93 completely prevented this recovery of Δ[Ca]_i and k_Ca during late acidosis in WT myocytes. In PLB-KO myocytes (n = 11) Δ[Ca] _i decreased during early acidosis from 2.92 ± 0.31 to 1.33 ± 0.17 ΔF/F₀ (P < 0.05) and k_Ca decreased from 10.45 ± 0.56 to 7.58 ± 0.68 s^-1 (P < 0.05). However, Δ[Ca]_i did not recover during late acidosis and k _Ca decreased even more (6.59 ± 0.65 s^-1). Parallel results were seen for contractile parameters. We conclude that PLB is crucial to the recovery of Δ[Ca]_i and k_Ca during acidosis. Moreover, PLB phosphorylation by CaMKII plays an important role in limiting the decline in Ca transients (and contraction) during acidosis.