RyR metabolic environment influences Ca²⁺ spark release in PMI mice. (A; top) Representative line-scan images of Ca²⁺ sparks obtained in permeabilized myocytes from sham (left) or from PMI (right) mice, with the same cell first in control (C.Sol.; top) and then “failing” (F.Sol.; bottom) solutions. (Bottom) Bar graph showing the average data of Ca²⁺ spark frequency in permeabilized cells from sham (n = 6) and PMI mice (n = 8) in control (Ct.Sol.; gray and dark red bars for sham and PMI, respectively) and failing solutions (F.Sol.; dark green for sham and dark cyan bar for PMI). (B; top) Line-scan examples of caffeine-evoked Ca²⁺ release in permeabilized myocytes in failing solution from sham (left) or from PMI (right) mice. (Bottom) Averaged caffeine-evoked Ca²⁺ transient peak (F/F₀) of permeabilized myocytes from sham (dark green bar; n = 6 cells from two animals) and PMI (blue bar; n = 8 cells from two animals) mice. (C–E) Ca²⁺ spark characteristics, Ca²⁺ spark amplitude (peak F/F₀; C), Ca²⁺ spark duration at half-peak amplitude (FDHM; D), and Ca²⁺ spark width at half-peak amplitude (FWHM; E) of permeabilized myocytes from sham (dark green bar; n = 1,263 Ca²⁺ sparks, from six cells, from two hearts) and PMI (dark cyan bar; n = 1,391 sparks, from eight cells, from two hearts) mice in failing solutions. *, P < 0.05 versus myocytes from sham mice; ***, P < 0.001 versus Sham cells in control solution; ^###, P < 0.001 versus the same group of myocytes in control solution. Error bars represent the SEM.