Hindlimb Denervation Alters the Regulation of Autophagy and Mitophagy

Abstract

During chronic muscle disuse, fibre atrophy occurs concomitantly with reductions in mitochondrial content and function. The reduction in organelle content is due, in part, to depressed mitochondrial biogenesis and elevations in intracellular degradation processes. The regulation of mitochondrial biogenesis has been studied extensively. However, the processes that underlie the degradation of mitochondria have yet to be fully elucidated. The breakdown of the organelle is due to a selective form of autophagy, termed mitophagy. When a mitochondrion becomes dysfunctional it dissociates from the mitochondrial network within the muscle and is subsequently degraded by lysosomes. The objective of this work is to better understand the processes of autophagy and mitophagy in the context of muscle disuse. Accordingly, we employed a hindlimb denervation protocol in which we unilaterally sectioned the peroneal nerve of one hindlimb, using the contralateral limb as a control in Sprague‐Dawley rats. Protein measurements of autophagy and mitochondrial content markers were made at 1, 3 and 7 days post‐denervation. We observed significant 25–30% (p<0.05) reductions in extensor digitorum longus (EDL) and tibialis anterior (TA) muscle mass by 7 days post‐denervation. Significant elevations in the autophagy proteins Tfeb, Beclin1 and ATG‐7 were also measured 7 days post denervation, while mitochondrial protein COX subunit‐IV and COX enzyme activity were reduced by 25% and 40%, respectively. However, SS and IMF mitochondrial respiratory capacity was diminished, accompanied by enhanced reactive oxygen species production, as early as 1–3 days following denervation. To investigate the early changes in autophagy and mitophagy flux that may explain the alterations in mitochondrial content and function, a subgroup of animals was treated with colchicine (4mg/kg/day) for 2 days to inhibit autophagic breakdown in response to 1 and 3 days of denervation. LC3‐II autophagy flux was increased as early as 1 and 3 days post‐denervation, but diminished by 7 days, in part as a result of a denervation‐induced increase in LC3‐II expression. Mitophagy flux was increased in parallel in SS mitochondria, and by 3 days in the IMF subfraction. These changes appeared to coincide with the declines in mitochondrial function, suggesting that although mitophagy is elevated, it is not sufficient to eliminate all of the dysfunctional organelles. Furthermore, the immediate elevations in autophagy/mitophagy flux at 1 and 3 days occurred prior to the upregulation of autophagy and lysosomal markers at 7 days post‐denervation, suggesting that the intrinsic activity of the autophagosomal breakdown pathway is sufficient in the early time course, but is subsequently upregulated to meet the demands of prolonged denervation.Support or Funding InformationSupported by NSERC, CanadaThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.