FIGURE 3.

Diurnal variations in myocardial triglyceride levels (A and B), net triglyceride synthesis rate (C), lipolysis rate (D), and triglyceride synthesis rate (E) in WT hearts were altered in CCM hearts. Myocardial triglyceride levels (A) were measured in WT and CCM hearts isolated at ZT 0, 6, 12, and 18. Oil red O staining (B) was performed on frozen sections from WT and CCM hearts isolated at ZT0. The black bar denotes 100 μm. Net incorporation of [³H]oleate into triglyceride (C) was measured in ex vivo perfused WT and CCM hearts isolated at ZT0, 6, 12, and 18; after the 40-min perfusion with buffer containing [9,10-³H]oleate (0.067 mCi/liter), the hearts were freeze-clamped and stored until analysis. Myocardial lipolysis (D) and triglyceride synthesis (E) were determined in ex vivo perfused WT and CCM hearts isolated at ZT6 and ZT18 using a pulse-chase method. The hearts were initially pulsed with 1.2 mm oleate and tracer amounts of [1-¹⁴C]oleate (0.12 mCi/liter) for 1 h followed by a 15-min wash with nonradioactive buffer and subsequently chased with buffer containing 0.4 mm oleate and tracer amounts of [9,10-³H]oleate (0.067 mCi/liter) for an additional 45 min. Data are represented as the mean ± S.E. Statistical analysis was performed using a two-way ANOVA followed by a post hoc comparison with Bonferroni correction. * denotes p < 0.05 time effect, and # denotes p < 0.05 genotype effect (n = 6 for myocardial triglyceride, n = 3–4 for Oil red O staining, n = 6–7 for net triglyceride synthesis, and n = 2–4 for lipolysis and triglyceride synthesis measurements).