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Inhibition of bioenergetics provides novel insights into recruitment of PINK 1‐dependent neuronal mitophagy
Author(s) -
Shin Yea Seul,
Ryall James G.,
Britto Joanne M.,
Lau Chew L.,
Devenish Rodney J.,
Nagley Phillip,
Beart Philip M.
Publication year - 2019
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1111/jnc.14667
Subject(s) - mitophagy , mitochondrion , depolarization , microbiology and biotechnology , antimycin a , rotenone , biology , respiratory chain , pink1 , biochemistry , membrane potential , chemistry , biophysics , autophagy , apoptosis
Contributions of damaged mitochondria to neuropathologies have stimulated interest in mitophagy. We investigated triggers of neuronal mitophagy by disruption of mitochondrial energy metabolism in primary neurons. Mitophagy was examined in cultured murine cerebellar granule cells after inhibition of mitochondrial respiratory chain by drugs rotenone, 3‐nitropropionic acid, antimycin A, and potassium cyanide, targeting complexes I, II , III , and IV , respectively. Inhibitor concentrations producing slow cellular demise were determined from analyses of cellular viability, morphology of neuritic damage, plasma membrane permeability, and oxidative phosphorylation. Live cell imaging of dissipation of mitochondrial membrane potential (ΔΨ m ) by drugs targeting mitochondrial complexes was referenced to complete depolarization by carbonyl cyanide m‐chlorophenyl hydrazone. While inhibition of complexes I, III and IV effected rapid dissipation of ΔΨ m , inhibition of complex II using 3‐nitropropionic acid led to minimal depolarization of mitochondria. Nonetheless, all respiratory chain inhibitors triggered mitophagy as indicated by increased aggregation of mitochondrially localized PINK 1. Mitophagy was further analyzed using a dual fluorescent protein biosensor reporting mitochondrial relocation to acidic lysosomal environment. Significant acidification of mitochondria was observed in neurons treated with rotenone or 3‐nitropropionic acid, revealing mitophagy at distal processes. Neurons treated with antimycin A or cyanide failed to show mitochondrial acidification. Minor dissipation of ΔΨ m by 3‐nitropropionic acid coupled with vigorous triggering of mitophagy suggested depolarization of mitochondria is not a necessary condition to trigger mitophagy. Moreover, weak elicitation of mitophagy by antimycin A, subsequent to loss of ΔΨ m , suggested that mitochondrial depolarization is not a sufficient condition for triggering robust neuronal mitophagy. Our findings provide new insight into complexities of mitophagic clearance of neuronal mitochondria.