Skeletal muscle mitochondrial and metabolic responses to a high-fat diet in female rats bred for high and low aerobic capacity

Abstract

Rats selected artificially to be low-capacity runners (LCR) possess a metabolic syndrome phenotype that is worsened by a high-fat diet (HFD), whereas rats selected to be high-capacity runners (HCR) are protected against HFD-induced obesity and insulin resistance. This study examined whether protection against, or susceptibility to, HFD-induced insulin resistance in the HCR-LCR strains is associated with contrasting metabolic adaptations in skeletal muscle. HCR and LCR rats (generation 20; n = 5-6; maximum running distance approximately 1800 m vs. approximately 350 m, respectively (p < 0.0001)) were divided into HFD (71.6% energy from fat) or normal chow (NC) (16.7% energy from fat) groups for 7 weeks (from 24 to 31 weeks of age). Skeletal muscle (red gastrocnemius) mitochondrial-fatty acid oxidation (FAO), mitochondrial-enzyme activity, mitochondrial-morphology, peroxisome proliferator-activated receptor gamma coactivator 1alpha (PGC-1alpha), and peroxisome proliferator-activated receptor delta (PPARdelta) expression and insulin sensitivity (intraperitoneal glucose tolerance tests) were measured. The HFD caused increased adiposity and reduced insulin sensitivity only in the LCR and not the HCR strain. Isolated mitochondria from the HCR skeletal muscle displayed a 2-fold-higher rate of FAO on NC, but both groups increased FAO following HFD. PGC-1alpha mRNA expression and superoxide dismutase activity were significantly reduced with the HFD in the LCR rats, but not in the HCR rats. PPARdelta expression did not differ between strains or dietary conditions. These results do not provide a clear connection between protection of insulin sensitivity and HFD-induced adaptive changes in mitochondrial function or transcriptional responses but do not dismiss the possibility that elevated mitochondrial FAO in the HCR may play a protective role.

Figure 1. Intraperitoneal glucose tolerance test

A glucose tolerance test was performed to assess whole body insulin sensitivity. The insulin AUC response was higher in the LCR than the HCR on both the NC, and the difference was magnified on the HFD as the LCR rats became more insulin resistant while the HCR rats did not have a significant increase in insulin AUC. No significant differences between groups or diets were found for the glucose AUC. n = 5-6/group, values are means ± SE. *Significantly higher than HCR, †significantly different from NC diet (P<0.05).

Figure 2. Total palmitate oxidation in isolated mitochondria from red gastrocnemius skeletal muscle

Total palmitate oxidation in isolated skeletal muscle mitochondria was higher in HCR than LCR on the NC diet. Although both groups had an increase in total palmitate oxidation, the increase was only significant in the LCR and resulted in no significant differences between groups after the HFD. n = 5-6/group, values are means ± SE. *Significantly higher than LCR animals, †significantly different from NC diet (P<0.05).

Figure 3. PPARδ, PGC-1α, and Cytochrome c protein and mRNA expression in red gastrocnemius skeletal muscle

On the NC diet, LCR had a trend for higher PGC-1α mRNA expression than the HCR (P=0.056), however, the expression dropped significantly in response to the HFD resulting in no differences between the groups on the HFD. When combining the HFD and NC groups the HCR had higher cytochrome c protein content than the LCR (Figure 4D). n = 5-6/group, values are means ± SE, *Significantly different between groups, † significantly different than NC diet, (P<0.05).

Figure 4. Representative images of (A) Subsarcolemmal and (B) Intermyofibrillar mitochondria

Images of SS and IMF mitochondria from the HCR and LCR on the NC and HFD.

Figure 5. Ratio of cytochrome c protein content per mitochondrial area

The ratio of cytochrome c protein per mitochondrial area was significantly higher in the HCR than the LCR on the NC diet. In response to the HFD, only the HCR had a significant increase in cytochrome c protein/mitochondrial area, resulting in the HCR group also having a larger ratio than the LCR in the HFD condition. *Significantly greater than LCR. †Significantly different than NC diet (P<0.05).

Figure 6. Superoxide dismutase activity in isolated mitochondria from red gastrocnemius muscle

The HCR had significantly lower mitochondrial SOD activity than the LCR on the NC diet. In response to the HFD, the LCR had a significant decrease in mitochondrial SOD activity while the HCR had a non-significant increase (P=0.08). n = 4-5/group, values are means ± SE, *significantly different between groups within a diet, † significantly different than NC diet, (P<0.05)