Ageing-induced decrease in cardiac mitochondrial function in healthy rats

Abstract

It is widely recognized that mitochondrial dysfunction is a key component of the multifactorial process of ageing. The effects of age on individual components of mitochondrial function vary across species and strains. In this study we investigated the oxygen consumption, the mitochondrial membrane potential (Δψ), the sensitivity of mitochondrial permeability transition pore (mPTP) to calcium overload, and the production of reactive oxygen species (ROS) in heart mitochondria isolated from old compared with adult healthy Sprague-Dawley rats. Respirometry studies and Δψ measurements were performed with an Oxygraph-2k equipped with a tetraphenylphosphonium electrode. ROS production and calcium retention capacity were measured spectrofluorimetrically. Our results show an important decline for all bioenergetic parameters for both complex I and complex II supported-respiration, a decreased Δψ in mitochondria energized with complex I substrates, and an increased mitochondrial ROS production in the old compared with the adult group. Mitochondrial sensitivity to Ca²⁺-induced mPTP opening was also increased in the old compared with the adult animals. Moreover, the protective effect of cyclosporine A on mPTP opening was significantly reduced in the old group. We conclude that healthy ageing is associated with a decrease in heart mitochondria function in Sprague-Dawley rats.

Radical Oxygen Species, Exercise and Aging: An Update

“…This is thought to be due to slowed electron transport and prolongation of ubisemiquinone radical occupancy in the complex [57]. However, most studies with animal models have reported a decrease in Dw with aging [58,59]. Therefore the Dw factor cannot explain the rise in ROS production in older subjects.…”

Mitochondria and aging: A role for the mitochondrial transition pore?

“…Indeed, pharmacological mPTP inhibitors failed to produce significant effects in either normal or stressed conditions. For instance, cyclosporin A (CsA) was unable to inhibit carboxyatractyloside‐induced permeability transition in aged mitochondria (García, Zazueta, Martínez‐Abundis, Pavón, & Chávez, 2009) and to prolong the time necessary to induce mPTP opening in isolated mitochondria (Duicu et al., 2013) and in cardiomyocytes isolated from old rats (Liu, Zhu, Brink, Glass, & Rebecchi, 2011). Similarly, the ability of sevoflurane and isoflurane conditioning (Li et al., 2013; Zhu et al., 2010) and of the GSK‐3β inhibitor SB‐216763 (Zhu, Rebecchi, Glass, Brink, & Liu, 2011) to protect against myocardial ischemia–reperfusion injury, which is mediated by inhibition of mPTP opening in young rats, is abrogated in senescent animals.…”

Modulation of mitochondrial respiratory function and ROS production by novel benzopyran analogues

“…Ventricular tissue was manually triturated with subtilisin (5 mg/g wet tissue), a non-specific protease, to release the interfibrillar mitochondria. The suspension was homogenised with a tissue homogenizer (Glas-Col 099C K5424 CE) for maximum 30 s. The final tissue homogenate obtained was processed at 4°C using the differential centrifugation technique as previously described (Duicu et al 2013). Mitochondrial protein concentration was determined using biuret method (Gornall et al 1949).…”

“…The substrate-uncoupler-inhibitor-titration (SUIT) protocol, GM State2 + ADP OXPHOS + c + Omy State4 + FCCP ETS + (Ama) ROX , was previously described (Duicu et al 2013) and comprised the following steps: (i) addition of complex I respiratory substrates, 10 mmol/L glutamate (G) and 2 mmol/L malate (M), State 2 (basal respiration); (ii) addition of 5 mmol/L adenosine diphosphate (ADP) to assess the maximal oxidative phosphorylation capacity (OXPHOS), State 3 (active respiration); (iii) addition of 10 mol/L cytochrome c (c) to check for the integrity of mitochondrial outer membrane; (iv) addition of 2 g/mL oligomycin (Omy), an F 0 F 1 -ATP synthase inhibitor, to inhibit State 3 at the level of State 4; (v) successive titrations (0.5 mol/L steps) with FCCP (carbonyl cyanide p-(trifluoro-methoxy) phenyl-hydrazone) to obtain the uncoupled respiration and to measure the electron transport system capacity, ETS; (vi) inhibition of respiration with 2.5 mol/L antimycin A (Ama) to measure the residual oxygen consumption, ROX state. Mitochondrial respiration was corrected for oxygen flux due to instrumental background and ROX.…”

“…Mitochondrial H 2 O 2 release was measured using the Amplex Red (10 mol/L) fluorescent marker (wavelengths: excitation at 530 nm and emission at 590 nm, Hitachi F-7000 spectrofluorimeter) as previously described (Duicu et al 2013). RHM (0.25 mg protein/mL) were incubated in 2 mL incubation buffer (250 mmol/L sucrose, 1 mmol/L EGTA, 1 mmol/L EDTA, 20 mmol/L Tris/HCl, and 1.5 mg/mL defatted BSA, pH 7.4, 37°C), supplemented with CI-dependent substrates: G (5 mmol/L) and M (5 mmol/L).…”