Figure 4.

Kcnt2^−/− Cardiomyocyte bioenergetics and mitochondrial ultrastructure. A) Representative images of isolated cardiomyocytes from WT and Kcnt2^−/− hearts. Scale bars, 100 µm. B) OCR of isolated cardiomyocytes measured with addition of oligomycin (1 µg/ml) and either 2.5 µM BT (K_Na opener) or 500 nM FCCP (mitochondrial uncoupler). Statistics were measured using 2-way ANOVA with Bonferroni correction and post hoc t test. Bars with the same symbol are significantly different from each other (n = 4–5). Data are means ± sem. P < 0.05. C) Western blots from WT and Kcnt2^−/− heart homogenates showing levels of mitochondrial proteins (SDHA, isocitrate dehydrogenase, cyclophilin D, and electron transfer flavoprotein subunit α) and Ponceau stain loading control. D) Representative transmission electron microscopic images of fixed heart slices. Bottom left panels: insets boxes at higher magnification. Scale bar, 1 µm. Top right panels: increased magnification of single mitochondria from WT and Kcnt2^−/− mice with mitochondrial ultrastructure visible (i.e., cristae folds, outer and inner membrane contacts). Scale bar, 200 nm. E) Binned histogram of mitochondrial area or mitochondrial density, obtained from image analysis (n = 5). Data are means ± sd for each bin. F) Form-factor/aspect-ratio scatterplot. E, F) Data obtained from 1250 mitochondria, 25 fields of view, and 5 WT or Kcnt2^−/− hearts.