Regulation of CPT I activity in intermyofibrillar and subsarcolemmal mitochondria from human and rat skeletal muscle

Carnitine palmitoyltransferase I (CPT I) is considered the rate-limiting enzyme in the transfer of long-chain fatty acids (LCFA) into the mitochondria and is reversibly inhibited by malonyl-CoA (M-CoA) in vitro. In rat skeletal muscle, M-CoA levels decrease during exercise, releasing the inhibition of CPT I and increasing LCFA oxidation. However, in human skeletal muscle, M-CoA levels do not change during moderate-intensity exercise despite large increases in fat oxidation, suggesting that M-CoA is not the sole regulator of increased CPT I activity during exercise. In the present study, we measured CPT I activity in intermyofibrillar ( IMF) and subsarcolemmal ( SS) mitochondria isolated from human vastus lateralis (VL), rat soleus ( Sol), and red gastrocnemius (RG) muscles. We tested whether exercise-related levels ( similar to 65% maximal O-2 uptake) of calcium and adenylate charge metabolites ( free AMP, ADP, and Pi) could override the M-CoA-induced inhibition of CPT I activity and explain the increased CPT I flux during exercise. Protein content was similar to 25 - 40% higher in IMF than in SS mitochondria in all muscles. Maximal CPT I activity was similar in IMF and SS mitochondria in all muscles ( VL: 282 +/- 46 vs. 280 +/- 51; Sol: 390 +/- 81 vs. 368 +/- 82; RG: 252 +/- 71 vs. 278 +/- 44 nmol . min(-1) . mg protein(-1)). Sensitivity to M-CoA did not differ between IMF and SS mitochondria in all muscles ( 25 - 31% inhibition in VL, 52 - 70% in Sol and RG). Calcium and adenylate charge metabolites did not override the M-CoA-induced inhibition of CPT I activity in mitochondria isolated from VL, Sol, and RG muscles. Decreasing pH from 7.1 to 6.8 reduced CPT I activity by similar to 34 - 40% in both VL mitochondrial fractions. In summary, this study reports no differences in CPT I activity or sensitivity to M-CoA between IMF and SS mitochondria isolated from human and rat skeletal muscles. Exercise-induced increases in calcium and adenylate charge metabolites do not appear responsible for upregulating CPT I activity in human or rat skeletal muscle during moderate aerobic exercise.