On the role of skeletal muscle acidosis and inorganic phosphates as determinants of central and peripheral fatigue: A 31 P-MRS study.

Intramuscular hydrogen ion (H+ ) and inorganic phosphate (Pi) concentrations were dissociated during exercise to challenge their relationships with peripheral and central fatigue in vivo. Ten recreationally active, healthy men (27 ± 5 years; 180 ± 4 cm; 76 ± 10 kg) performed two consecutive intermittent isometric single-leg knee-extensor trials (60 maximal voluntary contractions; 3 s contraction, 2 s relaxation) interspersed with 5 min of rest. Phosphorus magnetic resonance spectroscopy (31 P-MRS) was used to continuously quantify intramuscular [H+ ] and [Pi] during both trials. Using electrical femoral nerve stimulation, quadriceps twitch force (Qtw ) and voluntary activation (VA) were quantified at rest and throughout both trials. Decreases in Qtw and VA from baseline were used to determine peripheral and central fatigue, respectively. Qtw was strongly related to both [H+ ] (ß coefficient: -0.9, P < 0.0001) and [Pi] (-1.1, P < 0.0001) across trials. There was an effect of trial on the relationship between Qtw and [H+ ] (-0.5, P < 0.0001), but not Qtw and [Pi] (0.0, P = 0.976). This suggests that, unlike the unaltered association with [Pi], a given level of peripheral fatigue was associated with a different [H+ ] in Trial 1 vs. Trial 2. VA was related to [H+ ] (-0.3, P < 0.0001), but not [Pi] (-0.2, P = 0.243), across trials and there was no effect of trial (-0.1, P = 0.483). Taken together, these results support intramuscular Pi as a primary cause of peripheral fatigue, and muscle acidosis, probably acting on group III/IV muscle afferents in the interstitial space, as a contributor to central fatigue during exercise. KEY POINTS: We investigated the relationship between intramuscular metabolites and neuromuscular function in humans performing two maximal, intermittent, knee-extension trials interspersed with 5 min of rest. Concomitant measurements of intramuscular hydrogen (H+ ) and inorganic phosphate (Pi) concentrations, as well as quadriceps twitch-force (Qtw ) and voluntary activation (VA), were made throughout each trial using phosphorus magnetic resonance spectroscopy (31 P-MRS) and electrical femoral nerve stimulations. Although [Pi] fully recovered prior to the onset of the second trial, [H+ ] did not. Qtw was strongly related to both [H+ ] and [Pi] across both trials. However, the relationship between Qtw and [H+ ] shifted leftward from the first to the second trial, whereas the relationship between Qtw and [Pi] remained unaltered. VA was related to [H+ ], but not [Pi], across both trials. These in vivo findings support the hypotheses of intramuscular Pi as a primary cause of peripheral fatigue, and muscle acidosis, probably acting on group III/IV muscle afferents, as a contributor to central fatigue.

Acute high-intensity exercise and skeletal muscle mitochondrial respiratory function: role of metabolic perturbation.

Recently it was documented that fatiguing, high-intensity exercise resulted in a significant attenuation in maximal skeletal muscle mitochondrial respiratory capacity, potentially due to the intramuscular metabolic perturbation elicited by such intense exercise. With the utilization of intrathecal fentanyl to attenuate afferent feedback from group III/IV muscle afferents, permitting increased muscle activation and greater intramuscular metabolic disturbance, this study aimed to better elucidate the role of metabolic perturbation on mitochondrial respiratory function. Eight young, healthy males performed high-intensity cycle exercise in control (CTRL) and fentanyl-treated (FENT) conditions. Liquid chromatography-mass spectrometry and high-resolution respirometry were used to assess metabolites and mitochondrial respiratory function, respectively, pre- and postexercise in muscle biopsies from the vastus lateralis. Compared with CTRL, FENT yielded a significantly greater exercise-induced metabolic perturbation (PCr: -67% vs. -82%, Pi: 353% vs. 534%, pH: -0.22 vs. -0.31, lactate: 820% vs. 1,160%). Somewhat surprisingly, despite this greater metabolic perturbation in FENT compared with CTRL, with the only exception of respiratory control ratio (RCR) (-3% and -36%) for which the impact of FENT was significantly greater, the degree of attenuated mitochondrial respiratory capacity postexercise was not different between CTRL and FENT, respectively, as assessed by maximal respiratory flux through complex I (-15% and -33%), complex II (-36% and -23%), complex I + II (-31% and -20%), and state 3CI+CII control ratio (-24% and -39%). Although a basement effect cannot be ruled out, this failure of an augmented metabolic perturbation to extensively further attenuate mitochondrial function questions the direct role of high-intensity exercise-induced metabolite accumulation in this postexercise response.

Shifted vascular optimization: the emergence of a new arteriolar behaviour with chronic metabolic disease.

NEW FINDINGS: What is the topic of this review? Altered perfusion distribution at skeletal muscle arteriolar bifurcations and how this is modified by development of chronic metabolic disease. What advances does it highlight? The outcome created is a distribution of erythrocytes in the distal microcirculation that is characterized by increased spatial heterogeneity and reduced flexibility such that mass transport/exchange within the network is impaired, with limited ability to respond to imposed challenges. This advances our understanding of how altered vascular structure and function with metabolic disease impairs perfusion to skeletal muscle at a level of resolution that would not be identified through bulk flow responses. ABSTRACT: This review is based on the presentation 'Shifted vascular optimization: the emergence of a new arteriolar behaviour with chronic metabolic disease', given at the Symposium 'Understanding Complex Behaviours in the Microcirculation: from Blood Flow to Oxygenation' during the Annual Meeting of the Physiological Society at the Aberdeen Exhibition and Conference Centre in Aberdeen, UK in July 2019. The past years of dedicated investigation on linkages between vascular (dys)function under conditions of elevated cardiovascular disease risk and tissue/organ performance have produced results and insights that frequently suffer from limited correlation and causation. Reaching out from this challenge, it was proposed that this may reflect a 'level of resolution' argument and that altered haemodynamic behaviour in vascular networks could be a stronger predictor of functional outcomes than higher resolution measures. Using this approach, we have determined that an attractor that describes the spatial and temporal shift in perfusion distribution at successive arteriolar bifurcations within the skeletal muscle is a strong predictor of functional outcomes within animals and provides novel insight into fundamental mechanistic contributors to altered patterns of intra-muscular perfusion. This article focuses on the applicability and utility of the attractor in models of cardiovascular and metabolic disease risk of increasing severity. We will also discuss the utility of the attractor in terms of understanding the effectiveness of aggressive interventions for reversing established vasculopathy and perfusion impairments.

Skeletal muscle performance in metabolic disease: Microvascular or mitochondrial limitation or both?

One of the clearly established health outcomes associated with chronic metabolic diseases (eg, type II diabetes mellitus) is that the ability of skeletal muscle to maintain contractile performance during periods of elevated metabolic demand is compromised as compared to the fatigue-resistance of muscle under normal, healthy conditions. While there has been extensive effort dedicated to determining the major factors that contribute to the compromised performance of skeletal muscle with chronic metabolic disease, the extent to which this poor outcome reflects a dysfunctional state of the microcirculation, where the delivery and distribution of metabolic substrates can be impaired, versus derangements to normal metabolic processes and mitochondrial function, versus a combination of the two, represents an area of considerable unknown. The purpose of this manuscript is to present some of the current concepts for dysfunction to both the microcirculation of skeletal muscle as well as to mitochondrial metabolism under these conditions, such that these diverse issues can be merged into an integrated framework for future investigation. Based on an interpretation of the current literature, it may be hypothesized that the primary site of dysfunction with earlier stages of metabolic disease may lie at the level of the vasculature, rather than at the level of the mitochondria.

Transient receptor potential vanilloid 4 channels are important regulators of parenchymal arteriole dilation and cognitive function.

OBJECTIVE: Hypertension-associated PA dysfunction reduces cerebral perfusion and impairs cognition. This is associated with impaired TRPV4-mediated PA dilation; therefore, we tested the hypothesis that TRPV4 channels are important regulators of cerebral perfusion, PA structure and dilation, and cognition. METHODS: Ten- to twelve-month-old male TRPV4 knockout (WKY-Trpv4em4Mcwi ) and age-matched control WKY rats were studied. Cerebral perfusion was measured by MRI with arterial spin labeling. PA structure and function were assessed using pressure myography and cognitive function using the novel object recognition test. RESULTS: Cerebral perfusion was reduced in the WKY-Trpv4em4Mcwi rats. This was not a result of PA remodeling because TRPV4 deletion did not change PA structure. TRPV4 deletion did not change PA myogenic tone development, but PAs from the WKY-Trpv4em4Mcwi rats had severely blunted endothelium-dependent dilation. The WKY-Trpv4em4Mcwi rats had impaired cognitive function and exhibited depressive-like behavior. The WKY-Trpv4em4Mcwi rats also had increased microglia activation, and increased mRNA expression of GFAP and tumor necrosis factor alpha suggesting increased inflammation. CONCLUSION: Our data indicate that TRPV4 channels play a critical role in cerebral perfusion, PA dilation, cognition, and inflammation. Impaired TRPV4 function in diseases such as hypertension may increase the risk of the development of vascular dementia.

Skeletal muscle energetics are compromised only during high-intensity contractions in the Goto-Kakizaki rat model of type 2 diabetes.

Type 2 diabetes (T2D) presents with hyperglycemia and insulin resistance, affecting over 30 million people in the United States alone. Previous work has hypothesized that mitochondria are dysfunctional in T2D and results in both reduced ATP production and glucose disposal. However, a direct link between mitochondrial function and T2D has not been determined. In the current study, the Goto-Kakizaki (GK) rat model of T2D was used to quantify mitochondrial function in vitro and in vivo over a broad range of contraction-induced metabolic workloads. During high-frequency sciatic nerve stimulation, hindlimb muscle contractions at 2- and 4-Hz intensities, the GK rat failed to maintain similar bioenergetic steady states to Wistar control (WC) rats measured by phosphorus magnetic resonance spectroscopy, despite similar force production. Differences were not due to changes in mitochondrial content in red (RG) or white gastrocnemius (WG) muscles (cytochrome c oxidase, RG: 22.2 ± 1.6 vs. 23.3 ± 1.7 U/g wet wt; WG: 10.8 ± 1.1 vs. 12.1 ± 0.9 U/g wet wt; GK vs. WC, respectively). Mitochondria isolated from muscles of GK and WC rats also showed no difference in mitochondrial ATP production capacity in vitro, measured by high-resolution respirometry. At lower intensities (0.25-1 Hz) there were no detectable differences between GK and WC rats in sustained energy balance. There were similar phosphocreatine concentrations during steady-state contraction and postcontractile recovery (τ = 72 ± 6 s GK versus 71 ± 2 s WC). Taken together, these results suggest that deficiencies in skeletal muscle energetics seen at higher intensities are not due to mitochondrial dysfunction in the GK rat.

Quantification of Mitochondrial Oxidative Phosphorylation in Metabolic Disease: Application to Type 2 Diabetes.

Type 2 diabetes (T2D) is a growing health concern with nearly 400 million affected worldwide as of 2014. T2D presents with hyperglycemia and insulin resistance resulting in increased risk for blindness, renal failure, nerve damage, and premature death. Skeletal muscle is a major site for insulin resistance and is responsible for up to 80% of glucose uptake during euglycemic hyperglycemic clamps. Glucose uptake in skeletal muscle is driven by mitochondrial oxidative phosphorylation and for this reason mitochondrial dysfunction has been implicated in T2D. In this review we integrate mitochondrial function with physiologic function to present a broader understanding of mitochondrial functional status in T2D utilizing studies from both human and rodent models. Quantification of mitochondrial function is explained both in vitro and in vivo highlighting the use of proper controls and the complications imposed by obesity and sedentary lifestyle. This review suggests that skeletal muscle mitochondria are not necessarily dysfunctional but limited oxygen supply to working muscle creates this misperception. Finally, we propose changes in experimental design to address this question unequivocally. If mitochondrial function is not impaired it suggests that therapeutic interventions and drug development must move away from the organelle and toward the cardiovascular system.

Chronic atorvastatin and exercise can partially reverse established skeletal muscle microvasculopathy in metabolic syndrome.

It has long been known that chronic metabolic disease is associated with a parallel increase in the risk for developing peripheral vascular disease. Although more clinically relevant, our understanding about reversing established vasculopathy is limited compared with our understanding of the mechanisms and development of impaired vascular structure/function under these conditions. Using the 13-wk-old obese Zucker rat (OZR) model of metabolic syndrome, where microvascular dysfunction is sufficiently established to contribute to impaired skeletal muscle function, we imposed a 7-wk intervention of chronic atorvastatin treatment, chronic treadmill exercise, or both. By 20 wk of age, untreated OZRs manifested a diverse vasculopathy that was a central contributor to poor muscle performance, perfusion, and impaired O2 exchange. Atorvastatin or exercise, with the combination being most effective, improved skeletal muscle vascular metabolite profiles (i.e., nitric oxide, PGI2, and thromboxane A2 bioavailability), reactivity, and perfusion distribution at both individual bifurcations and within the entire microvascular network versus responses in untreated OZRs. However, improvements to microvascular structure (i.e., wall mechanics and microvascular density) were less robust. The combination of the above improvements to vascular function with interventions resulted in an improved muscle performance and O2 transport and exchange versus untreated OZRs, especially at moderate metabolic rates (3-Hz twitch contraction). These results suggest that specific interventions can improve specific indexes of function from established vasculopathy, but either this process was incomplete after 7-wk duration or measures of vascular structure are either resistant to reversal or require better-targeted interventions. NEW & NOTEWORTHY We used atorvastatin and/or chronic exercise to reverse established microvasculopathy in skeletal muscle of rats with metabolic syndrome. With established vasculopathy, atorvastatin and exercise had moderate abilities to reverse dysfunction, and the combined application of both was more effective at restoring function. However, increased vascular wall stiffness and reduced microvessel density were more resistant to reversal. Listen to this article's corresponding podcast at https://ajpheart.podbean.com/e/reversal-of-microvascular-dysfunction/ .

Measuring Mitochondrial Function: From Organelle to Organism.

Mitochondrial energy production is crucial for normal daily activities and maintenance of life. Herein, the logic and execution of two main classes of measurements are outlined to delineate mitochondrial function: ATP production and oxygen consumption. Aerobic ATP production is quantified by phosphorus magnetic resonance spectroscopy (31PMRS) in vivo in both human subjects and animal models using the same protocols and maintaining the same primary assumptions. Mitochondrial oxygen consumption is quantified by oxygen polarography and applied in isolated mitochondria, cultured cells, and permeabilized fibers derived from human or animal tissue biopsies. Traditionally, mitochondrial functional measures focus on maximal oxidative capacity-a flux rate that is rarely, if ever, observed outside of experimental conditions. Perhaps more physiologically relevant, both measurement classes herein focus on one principal design paradigm; submaximal mitochondrial fluxes generated by graded levels of ADP to map the function for ADP sensitivity. We propose this function defines the bioenergetic role that mitochondria fill within the myoplasm to sense and match ATP demands. Any deficit in this vital role for ATP homeostasis leads to symptoms often seen in cardiovascular and cardiopulmonary diseases, diabetes, and metabolic syndrome.

Impact of presymptomatic COVID-19 on vascular and skeletal muscle function: a case study.

The impact of COVID-19 has been largely described after symptom development. Although the SARS-CoV-2 virus elevates heart rate (HR) prior to symptom onset, whether this virus evokes other presymptomatic alterations is unknown. This case study details the presymptomatic impact of COVID-19 on vascular and skeletal muscle function in a young woman [24 yr, 173.5 cm, 89 kg, body mass index (BMI): 29.6 kg·m-2]. Vascular and skeletal muscle function were assessed as part of a separate study with the first and second visits separated by 2 wk. On the evening following the second visit, the participant developed a fever and a rapid antigen test confirmed a positive COVID-19 diagnosis. Compared with the first visit, the participant presented with a markedly elevated HR (∼30 beats/min) and a lower mean blood pressure (∼8 mmHg) at the second visit. Vascular function measured by brachial artery flow-mediated dilation, reactive hyperemia, and passive leg movement were all noticeably attenuated (25%-65%) as was leg blood flow during knee extension exercise. Muscle strength was diminished as was ADP-stimulated respiration (30%), assessed in vitro, whereas there was a 25% increase in the apparent Km. Lastly, an elevation in IL-10 was observed prior to symptom onset. Notably, 2.5 mo after diagnosis symptoms of fatigue and cough were still present. Together, these findings provide unique insight into the physiological responses immediately prior to onset of COVID-19 symptoms; they suggest that SARS-CoV-2 negatively impacts vascular and skeletal muscle function prior to the onset of common symptoms and may set the stage for the widespread sequelae observed following COVID-19 diagnosis.NEW & NOTEWORTHY This unique case study details the impact of SARS-CoV-2 infection on vascular and skeletal muscle function in a young predominantly presymptomatic woman. Prior to COVID-19 diagnosis, substantial reductions in vascular, skeletal muscle, and mitochondrial function were observed along with an elevation in IL-10. This integrative case study indicates that the presymptomatic impact of COVID-19 is widespread and may help elucidate the acute and long-term sequelae of this disease.

The hypertension advantage and natural selection: Since type 2 diabetes associates with co-morbidities and premature death, why have the genetic variants remained in the human genome?

Type 2 diabetes is a major public health crisis around the world. It is estimated that more than 300 million people worldwide have type 2 diabetes. Furthermore, the World Health Organization estimates that deaths from the complications of diabetes will increase by two thirds between 2008 and 2030. Since type 2 diabetes is a major public health crisis, why have the genetic variants for diabetes not been removed from the genome by natural selection? We hypothesize that insulin resistance, a predisposition to type 2 diabetes, and the associated elevation in sympathetic nervous system activity and arterial blood pressure provided an advantage to humans who lived as hunter-gatherers. Specifically, sympathetic hyperactivity stimulates the renin-angiotensin aldosterone system, promotes sodium reabsorption, and increases blood volume, heart rate, stroke volume and peripheral vascular resistance, thus inducing hypertension. The hypertension in turn provides a hemodynamic advantage for hunter-gatherers. Specifically, sympathetic hyperactivity and increased blood pressure increases blood flow delivery to working muscles by increasing cardiac output and shunting blood from non-active tissue. This natural selection for hypertension occurred during the time in human evolutionary history when the lifespan of most individuals was probably 30-40 years, and morbidity and mortality from cardiovascular disorders was limited. Thus, the selection pressure for elevation in sympathetic nervous system activity and blood pressure provided an advantage for hunting and gathering that would be greater than the selection pressure exerted by the manifestations of cardiovascular disease in aged individuals.

Obesity and inactivity, not hyperglycemia, cause exercise intolerance in individuals with type 2 diabetes: Solving the obesity and inactivity versus hyperglycemia causality dilemma.

Obesity, a sedentary lifestyle and type 2 diabetes are intricately linked conditions contributing to reduced exercise tolerance, significant morbidity, and premature deaths. It is unknown whether the reported exercise intolerance associated with type 2 diabetes is a direct result of the hyperglycemia, the impact of a relatively sedentary lifestyle, or increased adiposity. We hypothesize that obesity and inactivity, not hyperglycemia, cause exercise intolerance in individuals with type 2 diabetes. An analysis of the literature and results from the Goto-Kakizaki (GK) rat model of type 2 diabetes strongly support this hypothesis. GK rats were not sedentary or obese when compared with Wistar control rats and did not have exercise intolerance. Specifically, despite being hyperglycemic, GK rats demonstrated a longer treadmill run time to exhaustion (150.6 ±â€¯9.0 vs. 77.2 ±â€¯12.9 min), further distance run (1506 ±â€¯90 vs. 772 ±â€¯129 m), more work performed per gram muscle (44.0 ±â€¯2.8 vs. 21.9 ±â€¯3.8 kg*m/g) and a small increase in total vertical work performed when accounting for body mass (116.8 ±â€¯6.3 versus 98.9 ±â€¯15.2 kg*m). These results document preserved exercise tolerance in the non-obese, non-sedentary GK rat supporting the hypothesis that the reported exercise intolerance in models of type 2 diabetes is dependent on obesity and inactivity. Solving the obesity and inactivity versus hyperglycemia causality dilemma is important in understanding the development of type 2 diabetes and implications for future pharmacological and life style interventions.

Type 2 diabetes mellitus in the Goto-Kakizaki rat impairs microvascular function and contributes to premature skeletal muscle fatigue.

Despite extensive investigation into the impact of metabolic disease on vascular function and, by extension, tissue perfusion and organ function, interpreting results for specific risk factors can be complicated by the additional risks present in most models. To specifically determine the impact of type 2 diabetes without obesity on skeletal muscle microvascular structure/function and on active hyperemia with elevated metabolic demand, we used 17-wk-old Goto-Kakizaki (GK) rats to study microvascular function at multiple levels of resolution. Gracilis muscle arterioles demonstrated blunted dilation to acetylcholine (both ex vivo proximal and in situ distal arterioles) and elevated shear (distal arterioles only). All other alterations to reactivity appeared to reflect compromised endothelial function associated with increased thromboxane (Tx)A2 production and oxidant stress/inflammation rather than alterations to vascular smooth muscle function. Structural changes to the microcirculation of GK rats were confined to reduced microvessel density of ~12%, with no evidence for altered vascular wall mechanics. Active hyperemia with either field stimulation of in situ cremaster muscle or electrical stimulation via the sciatic nerve for in situ gastrocnemius muscle was blunted in GK rats, primarily because of blunted functional dilation of skeletal muscle arterioles. The blunted active hyperemia was associated with impaired oxygen uptake (V̇o2) across the muscle and accelerated muscle fatigue. Acute interventions to reduce oxidant stress (TEMPOL) and TxA2 action (SQ-29548) or production (dazmegrel) improved muscle perfusion, V̇o2, and muscle performance. These results suggest that type 2 diabetes mellitus in GK rats impairs skeletal muscle arteriolar function apparently early in the progression of the disease and potentially via an increased reactive oxygen species/inflammation-induced TxA2 production/action on network function as a major contributing mechanism. NEW & NOTEWORTHY The impact of type 2 diabetes mellitus on vascular structure/function remains an area lacking clarity. Using diabetic Goto-Kakizaki rats before the development of other risk factors, we determined alterations to vascular structure/function and skeletal muscle active hyperemia. Type 2 diabetes mellitus reduced arteriolar endothelium-dependent dilation associated with increased thromboxane A2 generation. Although modest microvascular rarefaction was evident, there were no other alterations to vascular structure/function. Skeletal muscle active hyperemia was blunted, although it improved after antioxidant or anti-thromboxane A2 treatment.