Fig 1: Activation of RyR1 but not of IP3R is necessary to increase the mitochondrial O2 consumption after membrane depolarization. Adult FDB muscle fibers maintained in Seahorse medium with BTS (10 μM) were pre-incubated for 1 h with xestospongin B (10 μM) or dantrolene (50 μM). The fibers were depolarized 5 min before the measurement. (A,D) Representative kinetics of OCR in fibers pre-incubated with xestospongin B or dantrolene respectively. (B,E) Quantification of basal and ATP-linked OCR in muscle fibers pre-incubated with xestospongin B or dantrolene respectively. (C,F) Percent distribution of the basal, ATP-linked, reserve capacity, proton leak and non-mitochondrial OCR were calculated. OCR after FCCP administration was considered as 100% (maximal OCR). N = 5 independents experiments. *p < 0.05; **p < 0.01 compared with the control condition.
Fig 2: Activation of IP3R and RyR1 participate in the mitochondrial Ca2+ increase after membrane depolarization. Adult FDB muscle was electroporated with plasmids encoding the molecular Ca2+ sensor CEPIA3mt. Fibers were maintained in Krebs Ringer buffer with BTS (10 μM) and pre-incubated for 1 h with xestospongin B (10 μM), dantrolene (50 μM) or both. The representative kinetics and maximal fluorescence was calculated in each condition (A,B). sh-IP3R1-RFP partially reduced the calcium increases in response to high K+ (C,D), scale bar is 15 μm. (E) Muscle fibers electroporated with plasmids encoding CEPIA3mt and pre-incubated with caged IP3. The photolysis increased the mitochondrial calcium level. Color bar represents the relative change in fluorescence. (F) Percent of response to different number of UV flashes is shown. N = 6 experiments were performed and 25 fibers were evaluated each time. †Means difference vs. control stimulated with K+. *Difference vs. resting; *p < 0.05; **/††p < 0.01.
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