Key points• Inorganic nitrate (NO 3 − ) supplementation with beetroot juice (BR) in humans lowers blood pressure and the O 2 cost of exercise and may improve exercise tolerance following its reduction to nitrite (NO 2 − ) and nitric oxide (NO).• The effect of inorganic NO 3 − supplementation with BR on skeletal muscle blood flow (BF) and vascular conductance (VC) within and among locomotory muscles during exercise is unknown.• Inorganic NO 3 − supplementation with BR in rats resulted in lower exercising mean arterial pressure, lower blood [lactate], and higher total skeletal muscle hindlimb BF and VC during submaximal treadmill running.• The greater BF and VC was found in muscles and muscle parts containing primarily type IIb + d/x muscle fibres.• These data demonstrate that inorganic NO 3 − supplementation improves vascular control and elevates skeletal muscle O 2 delivery during exercise predominantly in fast-twitch type II muscles, and provide a potential mechanism by which NO 3 − supplementation improves metabolic control. Abstract Dietary nitrate (NO 3− ) supplementation, via its reduction to nitrite (NO 2 − ) and subsequent conversion to nitric oxide (NO) and other reactive nitrogen intermediates, reduces blood pressure and the O 2 cost of submaximal exercise in humans. Despite these observations, the effects of dietary NO 3 − supplementation on skeletal muscle vascular control during locomotory exercise remain unknown. We tested the hypotheses that dietary NO 3 − supplementation via beetroot juice (BR) would reduce mean arterial pressure (MAP) and increase hindlimb muscle blood flow in the exercising rat. Male Sprague-Dawley rats (3-6 months) were administered either NO 3 − (via beetroot juice; 1 mmol kg −1 day −1 , BR n = 8) or untreated (control, n = 11) tap water for 5 days. MAP and hindlimb skeletal muscle blood flow and vascular conductance (radiolabelled microsphere infusions) were measured during submaximal treadmill running (20 m min −1 , 5% grade). BR resulted in significantly lower exercising MAP (control: 137 ± 3, BR: 127 ± 4 mmHg, P < 0.05) and blood [lactate] (control: 2.6 ± 0.3, BR: 1.9 ± 0.2 mM, P < 0.05) compared to control. Total exercising hindlimb skeletal muscle blood flow (control: 108 ± 8, BR: 150 ± 11 ml min −1 (100 g) −1 , P < 0.05) and vascular conductance (control: 0.78 ± 0.05, BR: 1.16 ± 0.10 ml min −1 (100 g) −1 mmHg −1 , P < 0.05) were greater in rats that received BR compared to control. The relative differences in blood flow and vascular conductance for the 28 individual hindlimb muscles and muscle parts correlated positively with their percentage type IIb + d/x muscle fibres (blood flow: r = 0.74, vascular conductance: r = 0.71, P < 0.01 for both). These data support the hypothesis that NO 3 − supplementation improves vascular control and elevates skeletal muscle O 2 delivery during exercise predominantly in fast-twitch type II muscles, and provide a potential mechanism by which NO 3 − supplementation improves metabolic control.

The defining characteristic of chronic heart failure (CHF) is an exercise intolerance that is inextricably linked to structural and functional aberrations in the O(2) transport pathway. CHF reduces muscle O(2) supply while simultaneously increasing O(2) demands. CHF severity varies from moderate to severe and is assessed commonly in terms of the maximum O(2) uptake, which relates closely to patient morbidity and mortality in CHF and forms the basis for Weber and colleagues' (167) classifications of heart failure, speed of the O(2) uptake kinetics following exercise onset and during recovery, and the capacity to perform submaximal exercise. As the heart fails, cardiovascular regulation shifts from controlling cardiac output as a means for supplying the oxidative energetic needs of exercising skeletal muscle and other organs to preventing catastrophic swings in blood pressure. This shift is mediated by a complex array of events that include altered reflex and humoral control of the circulation, required to prevent the skeletal muscle "sleeping giant" from outstripping the pathologically limited cardiac output and secondarily impacts lung (and respiratory muscle), vascular, and locomotory muscle function. Recently, interest has also focused on the dysregulation of inflammatory mediators including tumor necrosis factor-α and interleukin-1β as well as reactive oxygen species as mediators of systemic and muscle dysfunction. This brief review focuses on skeletal muscle to address the mechanistic bases for the reduced maximum O(2) uptake, slowed O(2) uptake kinetics, and exercise intolerance in CHF. Experimental evidence in humans and animal models of CHF unveils the microvascular cause(s) and consequences of the O(2) supply (decreased)/O(2) demand (increased) imbalance emblematic of CHF. Therapeutic strategies to improve muscle microvascular and oxidative function (e.g., exercise training and anti-inflammatory, antioxidant strategies, in particular) and hence patient exercise tolerance and quality of life are presented within their appropriate context of the O(2) transport pathway.

The capillary bed constitutes a vast surface facilitating exchange of O2, substrates and metabolites between blood and organs. In contracting skeletal muscle capillary blood flow and O2 diffusing capacity as well as O2 flux may increase two orders of magnitude above resting. Chronic diseases such as heart failure, diabetes and also sepsis impair these processes leading to compromised energetic, metabolic and ultimately contractile function. Among researchers seeking to understand blood-myocyte exchange in health and the bases for dysfunction in disease there is a fundamental disconnect between microcirculation specialists and many physiologists and physiologist clinicians. Whereas the former observe capillaries and capillary function directly (muscle intravital microscopy) the latter generally use indirect methodologies (e.g., post-mortem tissue analysis, 1-methyl xanthine, contrast enhanced ultrasound, permeability surface area product) and interpret their findings based upon August Krogh’s observations made nearly a century ago. “Kroghian” theory holds that only a small fraction of capillaries support red blood cell (RBC) flux in resting muscle leaving the vast majority to be “recruited” (i.e., initiate RBC flux) during contractions which would constitute the basis for increasing capillary exchange surface area and reducing capillary-mitochondrial diffusion distances. Experimental techniques each have their strengths and weaknesses and often the correct or complete answer to a problem emerges from integration across multiple technologies. Today Krogh’s entrenched “capillary recruitment” hypothesis is challenged by direct observations of capillaries in contracting muscle; something that he and his colleagues could not do. Moreover, in the peer-reviewed scientific literature, application of a range of contemporary physiological technologies including intravital microscopy of contracting muscle, magnetic resonance and near infrared spectroscopy and phosphorescence quenching combined with elegant in situ and in vivo models suggest that the role of the capillary bed, at least in contracting muscle, is subserved without the necessity for de novo capillary recruitment of previously non-flowing capillaries. When viewed within the context of the capillary recruitment hypothesis, this evidence casts serious doubt on the interpretation of those data that are based upon Kroghian theory and indirect methodologies. Thus today a wealth of evidence calls for a radical revision of blood-muscle exchange theory to one in which most capillaries support RBC flux at rest and, during contractions, capillary surface area is “recruited” along the length of previously flowing capillaries. This occurs, in part, by elevating capillary hematocrit and extending the length of the capillary available for blood-myocyte exchange (i.e., longitudinal recruitment). Our understanding of blood-myocyte O2 and substrate/metabolite exchange in health and the mechanistic bases for dysfunction in disease demands no less.

The O2 requirements of contracting skeletal muscle may increase 100-fold above rest. In 1919 August Krogh’s brilliant insights recognized the capillary as the principal site for this increased blood-myocyte O2 flux. Based on the premise that most capillaries did not sustain RBC flux at rest Krogh proposed that capillary recruitment (i.e., initiation of red blood cell (RBC) flux in previously non-flowing capillaries) increased the capillary surface area available for O2 flux and reduced mean capillary-to-mitochondrial diffusion distances. More modern experimental approaches reveal that most muscle capillaries may support RBC flux at rest. Thus, rather than contraction-induced capillary recruitment per se, increased RBC flux and hematocrit within already-flowing capillaries likely elevate perfusive and diffusive O2 conductances and hence blood-myocyte O2 flux. Additional surface area for O2 exchange is recruited but, crucially, this may occur along the length of already-flowing capillaries (i.e. longitudinal recruitment). Today, the capillary is still considered the principal site for O2 and substrate delivery to contracting skeletal muscle. Indeed, the presence of very low intramyocyte O2 partial pressures (PO2’s) and the absence of PO2 gradients, whilst refuting the relevance of diffusion distances, place an even greater importance on capillary hemodynamics. This emergent picture calls for a paradigm-shift in our understanding of the function of capillaries by de-emphasizing de novo ‘capillary recruitment.’ Diseases such as heart failure impair blood-myocyte O2 flux, in part, by decreasing the proportion of RBC-flowing capillaries. Knowledge of capillary function in healthy muscle is requisite for identification of pathology and efficient design of therapeutic treatments.

Critical speed (CS) constitutes an important metabolic and performance demarcator. However, active skeletal muscle blood flow distribution specifically surrounding CS remains unknown. We tested the hypotheses that CS could be accurately determined in the running rat and that measurement of hindlimb inter-and intramuscular blood flow below and above CS would support that the greatest muscle fibre recruitment above, relative to below, CS occurs in the predominantly glycolytic muscles. Seven male Sprague-Dawley rats performed five constant-speed tests to exhaustion at speeds between 95 and 115% of the speed that elicitedV O 2 max to determine CS. Subsequent constant-speed tests were performed at speeds incrementally surrounding CS to determine time to exhaustion,V O 2 , and hindlimb muscle blood flow distribution. Speed and time to exhaustion conformed to a hyperbolic relationship (r 2 = 0.92 ± 0.03) which corresponded to a linear 1/time function (r 2 = 0.93 ± 0.02) with a CS of 48.6 ± 1.0 m min −1 . Time to exhaustion below CS was ∼5× greater (P < 0.01) than that above. Below CSV O 2 stabilized at a submaximal value (58.5 ± 2.5 ml kg −1 min −1 ) whereas above CSV O 2 (81.7 ± 2.5 ml kg −1 min −1 ) increased toV O 2 max (84.0 ± 1.8 ml kg −1 min −1 , P > 0.05 vs. above CS). The 11 individual muscles or muscle parts that evidenced the greatest blood flow increases above, relative to below, CS were composed of ≥69% Type IIb/d/x muscle fibres. Moreover, there was a significant correlation (P < 0.05, r = 0.42) between the increased blood flow above expressed relative to below CS and the percentage Type IIb/d/x fibres found in the individual muscles or muscle parts. These data validate the powerful CS construct in the rat and identify that running above CS, relative to below CS, incurs disproportionate blood flow increases (indicative of recruitment) in predominantly highly glycolytic muscle fibres.