Regulation of autophagic and mitophagic flux during chronic contractile activity-induced muscle adaptations.

Abstract

Autophagy and mitophagy are important for training-inducible muscle adaptations, yet it remains unclear how these systems are regulated throughout the adaptation process. Here, we studied autophagic and mitophagic flux in the skeletal muscles of Sprague-Dawley rats (300-500g) exposed to chronic contractile activity (CCA; 3h/day, 9V, 10Hz continuous, 0.1ms pulse duration) for 1, 2, 5, and 7days (N = 6-8/group). In order to determine the flux rates, colchicine (COL; 0.4mg/ml/kg) was injected 48h before tissue collection, and we evaluated differences of autophagosomal protein abundances (LC3-II and p62) between colchicine- and saline-injected animals. We confirmed that CCA resulted in mitochondrial adaptations, including improved state 3 respiration as early as day 1 in permeabilized muscle fibers, as well significant increases in mitochondrial respiratory capacity and marker proteins in IMF mitochondria by day 7. Mitophagic and autophagic flux (LC3-II and p62) were significantly decreased in skeletal muscle following 7days of CCA. Notably, the mitophagic system seemed to be downregulated prior (day 3-5) to changes in autophagic flux (day 7), suggesting enhanced sensitivity of mitophagy compared to autophagy with chronic muscle contraction. Although we detected no significant change in the nuclear translocation of TFEB, a regulator of lysosomal biogenesis, CCA increased total TFEB protein, as well as LAMP1, in skeletal muscle. Thus, chronic muscle activity reduces mitophagy in parallel with improved mitochondrial function, and this is supported by enhanced lysosomal degradation capacity.