Carbohydrates and lipids provide the majority of substrates to fuel mitochondrial oxidative phosphorylation. Metabolic inflexibility, defined as an impaired ability to switch between these fuels, is implicated in a number of metabolic diseases. Here, we explore the mechanism by which physical inactivity promotes metabolic inflexibility in skeletal muscle. We developed a mouse model of sedentariness, small mouse cage (SMC), that, unlike other classic models of disuse in mice, faithfully recapitulated metabolic responses that occur in humans. Bioenergetic phenotyping of skeletal muscle mitochondria displayed metabolic inflexibility induced by physical inactivity, demonstrated by a reduction in pyruvate-stimulated respiration ( J O ₂ ) in the absence of a change in palmitate-stimulated J O ₂ . Pyruvate resistance in these mitochondria was likely driven by a decrease in phosphatidylethanolamine (PE) abundance in the mitochondrial membrane. Reduction in mitochondrial PE by heterozygous deletion of phosphatidylserine decarboxylase (PSD) was sufficient to induce metabolic inflexibility measured at the whole-body level, as well as at the level of skeletal muscle mitochondria. Low mitochondrial PE in C2C12 myotubes was sufficient to increase glucose flux toward lactate. We further implicate that resistance to pyruvate metabolism is due to attenuated mitochondrial entry via mitochondrial pyruvate carrier (MPC). These findings suggest a mechanism by which mitochondrial PE directly regulates MPC activity to modulate metabolic flexibility in mice.

Diabetic cardiomyopathy, an increasingly global epidemic and a major cause of heart failure with preserved ejection fraction (HFpEF), is associated with hyperglycemia, insulin resistance, and intracardiomyocyte calcium mishandling. Here we identify that, in db/db mice with type 2 diabetes–induced HFpEF, abnormal remodeling of cardiomyocyte transverse-tubule microdomains occurs with downregulation of the membrane scaffolding protein cardiac bridging integrator 1 (cBIN1). Transduction of cBIN1 by AAV9 gene therapy can restore transverse-tubule microdomains to normalize intracellular distribution of calcium-handling proteins and, surprisingly, glucose transporter 4 (GLUT4). Cardiac proteomics revealed that AAV9-cBIN1 normalized components of calcium handling and GLUT4 translocation machineries. Functional studies further identified that AAV9-cBIN1 normalized insulin-dependent glucose uptake in diabetic cardiomyocytes. Phenotypically, AAV9-cBIN1 rescued cardiac lusitropy, improved exercise intolerance, and ameliorated hyperglycemia in diabetic mice. Restoration of transverse-tubule microdomains can improve cardiac function in the setting of diabetic cardiomyopathy and can also improve systemic glycemic control.

The development of high‐resolution respirometry (HRR) has greatly expanded the analytical scope to study mitochondrial respiratory control relative to specific tissue/cell types across various metabolic states. Specifically, the Oroboros Oxygraph 2000 (O2k) is a common tool for measuring rates of mitochondrial respiration and is the focus of this perspective. The O2k platform is amenable for answering numerous bioenergetic questions. However, inherent variability with HRR‐derived data, both within and amongst users, can impede progress in bioenergetics research. Therefore, we advocate for several vital considerations when planning and conducting O2k experiments to ultimately enhance transparency and reproducibility across laboratories. In this perspective, we offer guidance for best practices of mitochondrial preparation, protocol selection, and measures to increase reproducibility. The goal of this perspective is to propagate the use of the O2k, enhance reliability and validity for both new and experienced O2k users, and provide a reference for peer reviewers.

Older age is the primary risk factor for most chronic diseases, including Alzheimer’s disease (AD). Current preclinical models to study brain aging and AD are mainly transgenic and harbor mutations intended to mirror brain pathologies associated with human brain aging/AD (eg, by increasing production of the amyloid precursor protein, amyloid beta [Aβ], and/or phosphorylated tau, all of which are key pathological mediators of AD). Although these models may provide insight on pathophysiological processes in AD, none completely recapitulate the disease and its strong age-dependence, and there has been limited success in translating preclinical results and treatments to humans. Here, we describe 2 nontransgenic guinea pig (GP) models, a standard PigmEnTed (PET) strain, and lesser-studied Dunkin-Hartley (DH) strain, that may naturally mimic key features of brain aging and AD in humans. We show that brain aging in PET GP is transcriptomically similar to human brain aging, whereas older DH brains are transcriptomically more similar to human AD. Both strains/models also exhibit increased neurofilament light chain (NFL, a marker of neuronal damage) with aging, and DH animals display greater S100 calcium-binding protein B (S100β), ionized calcium-binding adapter molecule 1 (Iba1), and Aβ and phosphorylated tau—which are all important markers of neuroinflammation-associated AD. Collectively, our results suggest that both the PET and DH GP may be useful, nontransgenic models to study brain aging and AD, respectively.

Impaired mitochondrial function and disrupted proteostasis contribute to musculoskeletal dysfunction. However, few interventions simultaneously target these two drivers to prevent musculoskeletal decline. Nuclear factor erythroid 2-related factor 2 (Nrf2) activates a transcriptional programme promoting cytoprotection, metabolism, and proteostasis. We hypothesized daily treatment with a purported Nrf2 activator, PB125, in Hartley guinea pigs, a model of musculoskeletal decline, would attenuate the progression of skeletal muscle mitochondrial dysfunction and impaired proteostasis and preserve musculoskeletal function. We treated 2- and 5-month-old male and female Hartley guinea pigs for 3 and 10 months, respectively, with the phytochemical compound PB125. Longitudinal assessments of voluntary mobility were measured using Any-Maze (TM) open-field enclosure monitoring. Cumulative skeletal muscle protein synthesis rates were measured using deuterium oxide over the final 30 days of treatment. Mitochondrial oxygen consumption in soleus muscles was measured using high resolution respirometry. In both sexes, PB125 (1) increased electron transfer system capacity; (2) attenuated the disease/age-related decline in coupled and uncoupled mitochondrial respiration; and (3) attenuated declines in protein synthesis in the myofibrillar, mitochondrial and cytosolic subfractions of the soleus. These effects were not associated with statistically significant prolonged maintenance of voluntary mobility in guinea pigs. Collectively, treatment with PB125 contributed to maintenance of skeletal muscle mitochondrial respiration and proteostasis in a pre-clinical model of musculoskeletal decline. Further investigation is necessary to determine if these documented effects of PB125 are also accompanied by slowed progression of other aspects of musculoskeletal dysfunction.