Differential Respiratory Response of Isolated Cardiac Mitochondria from Young and Aged Mice towards Nitric Oxide Synthase Inhibitors

Abstract

Mitochondrial respiratory dysfunction is proposed to be an important underlying mechanism in the aging‐induced cardiac abnormalities. Identifying the mechanisms involved in the development of aging‐induced respiratory dysfunction in cardiac mitochondria could be useful to develop therapeutic strategies to reduce the risk of developing cardiovascular diseases. Role of mitochondrial nitric oxide synthase (NOS) to aging‐induced respiratory changes in cardiac mitochondria are not studied previously. Here we treated the isolated cardiac mitochondria from young (3 months) and old mice (18 months) hearts with neuronal NOS inhibitor (nNOS, ARL 17477 dihydrochloride or ARL) and endothelial NOS inhibitor (eNOS, N5‐(1‐Iminoethyl)‐L‐ornithine or NIO) and studied the respiratory characteristics using Seahorse Xfe24 analyzer.ResultsMitochondria from the old mice hearts exhibited significantly increased basal respiration (24.7%, 169.0±14.0 vs 135.5±5.1, p≤0.05) with unchanged ATP production. They also showed a significant decrease in state IIIu respiration (23%, 635.4±40.2 vs 826.4±49.2, p≤0.05) and also the membrane potential. Both ARL and NIO treatments significantly decreased state IIIu respiration (22.1%, 643.7±34.4 vs 826.4±49.2, p≤0.05 & 23.9%, 448±18.5 vs 587.1±34.8, p≤0.05 respectively) in mitochondria from young mice hearts without affecting membrane potential and oxidative stress parameters. Interestingly, ARL and NIO neither induced similar responses in the mitochondria from the old mice hearts nor they corrected the aging‐induced decline in state IIIu respiration and membrane potential. Mitochondria from the old mice hearts have increased nNOS protein levels (421%, 1.98±0.80 vs 0.38±0.20, p≤0.05) with decreased protein S‐nitrosylation (46.25%, 8.3±1.5 vs 15.44±3.2, p≤0.07). The observed differential respiratory responses of cardiac mitochondria from young and old mice following nNOS and eNOS inhibitors were associated with reduced protein S‐nitrosylation in mitochondria from young mice hearts (64.2%, 5.53±0.35 vs 15.44±3.2, p≤0.05 & 65.7%, 5.3±0.84 vs 15.44±3.2, p≤0.05 respectively) with unaltered nitrosylation in mitochondria from old mice hearts.ConclusionsThus the functional loss of mitochondrial nNOS and eNOS in the aged hearts accompanied by the reduced NO‐mediated regulation of mitochondrial proteins contributes to the aging‐induced mitochondrial dysfunction.Support or Funding InformationAmerican Heart Association (PVG: 14SDG20490359 and VNS: 16PRE31450006) and National Institute of Health: National Institute of Neurological Disorders and Stroke and National Institute of General Medical Sciences (PVK: R01NS094834).This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.