To study effects of mitochondrial complex I (CI, NADH:ubiquinone oxidoreductase) deficiency, we inactivated the Ndufs4 gene, which encodes an 18 kDa subunit of the 45-protein CI complex. Although small, Ndufs4 knockout (KO) mice appeared healthy until approximately 5 weeks of age, when ataxic signs began, progressing to death at approximately 7 weeks. KO mice manifested encephalomyopathy including a retarded growth rate, lethargy, loss of motor skill, blindness, and elevated serum lactate. CI activity in submitochondrial particles from KO mice was undetectable by spectrophotometric assays. However, CI-driven oxygen consumption by intact tissue was about half that of controls. Native gel electrophoresis revealed reduced levels of intact CI. These data suggest that CI fails to assemble properly or is unstable without NDUFS4. KO muscle has normal morphology but low NADH dehydrogenase activity and subsarcolemmal aggregates of mitochondria. Nonetheless, total oxygen consumption and muscle ATP and phosphocreatine concentrations measured in vivo were within normal parameters.

To explore the lethal, ataxic phenotype of complex I deficiency in Ndufs4 knockout (KO) mice, we inactivated Ndufs4 selectively in neurons and glia (NesKO mice). NesKO mice manifested the same symptoms as KO mice including retarded growth, loss of motor ability, breathing abnormalities, and death by ∼7 wk. Progressive neuronal deterioration and gliosis in specific brain areas corresponded to behavioral changes as the disease advanced, with early involvement of the olfactory bulb, cerebellum, and vestibular nuclei. Neurons, particularly in these brain regions, had aberrant mitochondrial morphology. Activation of caspase 8, but not caspase 9, in affected brain regions implicate the initiation of the extrinsic apoptotic pathway. Limited caspase 3 activation and the predominance of ultrastructural features of necrotic cell death suggest a switch from apoptosis to necrosis in affected neurons. These data suggest that dysfunctional complex I in specific brain regions results in progressive glial activation that promotes neuronal death that ultimately results in mortality.cerebellum | mitochondria | gliosis | neurodegeneration | vestibular nucleus

Summary Mitochondrial dysfunction plays a key pathogenic role in aging skeletal muscle resulting in significant healthcare costs in the developed world. However, there is no pharmacologic treatment to rapidly reverse mitochondrial deficits in the elderly. Here we demonstrate that a single treatment with the mitochondrial targeted peptide SS-31 restores in vivo mitochondrial energetics to young levels in aged mice after only one hour. Young (5 month old) and old (27 month old) mice were injected intraperitoneally with either saline or 3 mg/kg of SS-31. Skeletal muscle mitochondrial energetics were measured in vivo one hour after injection using a unique combination of optical and 31P magnetic resonance spectroscopy. Age related declines in resting and maximal mitochondrial ATP production, coupling of oxidative phosphorylation (P/O), and cell energy state (PCr/ATP) were rapidly reversed after SS-31 treatment, while SS-31 had no observable effect on young muscle. These effects of SS-31 on mitochondrial energetics in aged muscle were also associated with a more reduced glutathione redox status and lower mitochondrial H2O2 emission. Skeletal muscle of aged mice was more fatigue resistant in situ one hour after SS-31 treatment and eight days of SS-31 treatment led to increased whole animal endurance capacity. These data demonstrate that SS-31 represents a new strategy for reversing age-related deficits in skeletal muscle with potential for translation into human use.