Research Article: In vivo imaging reveals mitophagy independence in the maintenance of axonal mitochondria during normal aging

Date Published: August 07, 2017

Publisher: John Wiley and Sons Inc.

Author(s): Xu Cao, Haiqiong Wang, Zhao Wang, Qingyao Wang, Shuang Zhang, Yuanping Deng, Yanshan Fang.


Mitophagy is thought to be a critical mitochondrial quality control mechanism in neurons and has been extensively studied in neurological disorders such as Parkinson’s disease. However, little is known about how mitochondria are maintained in the lengthy neuronal axons in the context of physiological aging. Here, we utilized the unique Drosophila wing nerve model and in vivo imaging to rigorously profile changes in axonal mitochondria during aging. We revealed that mitochondria became fragmented and accumulated in aged axons. However, lack of Pink1 or Parkin did not lead to the accumulation of axonal mitochondria or axonal degeneration. Further, unlike in in vitro cultured neurons, we found that mitophagy rarely occurred in intact axons in vivo, even in aged animals. Furthermore, blocking overall mitophagy by knockdown of the core autophagy genes Atg12 or Atg17 had little effect on the turnover of axonal mitochondria or axonal integrity, suggesting that mitophagy is not required for axonal maintenance; this is regardless of whether the mitophagy is PINK1‐Parkin dependent or independent. In contrast, downregulation of mitochondrial fission–fusion genes caused age‐dependent axonal degeneration. Moreover, Opa1 expression in the fly head was significantly decreased with age, which may underlie the accumulation of fragmented mitochondria in aged axons. Finally, we showed that adult‐onset, neuronal downregulation of the fission–fusion, but not mitophagy genes, dramatically accelerated features of aging. We propose that axonal mitochondria are maintained independently of mitophagy and that mitophagy‐independent mechanisms such as fission–fusion may be central to the maintenance of axonal mitochondria and neural integrity during normal aging.

Partial Text

Healthy mitochondria are critical for maintaining normal bioenergetically demanding activities of neurons. Such energy demand in neuronal axons is likely to be especially extreme due to the activities such as synaptic transmission, generating and propagating action potentials, and transporting biomaterials over a long distance. Deleterious mitochondrial changes such as a decrease in mitochondrial integrity and function are associated with aging and neurodegenerative diseases (Bratic & Larsson, 2013; López‐Otín et al., 2013). Mitophagy is an important mitochondrial quality control mechanism that selectively eliminates damaged mitochondria by autophagy (Wang and Klionsky, 2014; Wei et al., 2015). The process is regulated by the PTEN‐induced putative kinase 1 (PINK1) and the E3 ubiquitin ligase Parkin (Pickrell & Youle, 2015), whose mutations can cause familial forms of Parkinson’s disease (PD).

Neuronal aging is known to be associated with deleterious changes in mitochondria including a decrease in mitochondrial biogenesis, the respiratory chain efficacy and ATP generation, an increase in the production of reactive oxygen species, accumulation of mitochondrial DNA mutations, and reduction in mitochondrial transport and turnover (Green et al., 2011; López‐Otín et al., 2013; Bratic & Larsson, 2013). The development of the in vivo imaging paradigm of the Drosophila wing nerve has enabled the systematic characterization of mitochondrial morphology in axons during aging in this study. Moreover, taking the advantage of vast genetic tools in Drosophila, we have manipulated mitochondrial quality control genes in a spatially and temporally controlled manner, and directly visualized the consequence on mitochondria and axonal integrity in vivo. The imaging data show a clear change in mitochondrial heterogeneity from long, tubular mitochondria in young axons toward short, round mitochondria in aged axons. This change is concurrent with an increased number of axonal mitochondria. Thus, aging is associated with an accumulation of fragmented mitochondria in axons. Our finding is consistent with a recent study showing that the axonal mitochondria size decreased in aged C. elegans (Morsci et al., 2016). This is in addition to the reported decline of axonal mitochondrial transport during aging in nematode neurons (Morsci et al., 2016) and mouse retinal ganglion cells (Takihara et al., 2015).

This study is supported by the National Key R&D Program of China (No. 2016YFA0501902), the National Natural Science Foundation of China (No. 31471017 and No. 81671254), and the State High‐Tech Development Plan of China (the ‘863 Program’, No. 2014A020526) to Y.F.

X.C. planned and performed experiments, analyzed data, and wrote the manuscript. H.W. and Q.W. performed experiments. X.C., H.W., and Z.W. contributed critical reagents. S.Z. and Y.D. provided technical assistance. Y.F. designed and supervised research and wrote the manuscript.

The authors declare no conflict of interest.




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