Timothy I. Musch

Control of microvascular oxygen pressures in rat muscles comprised of different fibre types

In response to an elevated metabolic rate , increased microvascular blood–muscle O2 flux is the product of both augmented O2 delivery and fractional O2 extraction. Whole body and exercising limb measurements demonstrate that and fractional O2 extraction increase as linear and hyperbolic functions, respectively, of . Given the presence of disparate vascular control mechanisms among different muscle fibre types, we tested the hypothesis that, in response to muscle contractions, would be lower and fractional O2 extraction (as assessed via microvascular O2 pressure, P  mvO 2) higher in fast‐ versus slow‐twitch muscles. Radiolabelled microsphere and phosphorescence quenching techniques were used to measure and P  mvO 2, respectively at rest and across the transition to 1 Hz twitch contractions at low (Lo, 2.5 V) and high intensities (Hi, 4.5 V) in rat (n= 20) soleus (Sol, slow‐twitch, type I), mixed gastrocnemius (MG, fast‐twitch, type IIa) and white gastrocnemius (WG, fast‐twitch, type IIb) muscle. At rest and for Lo and Hi (steady‐state values) transitions, P  mvO 2 was lower (all P < 0.05) in MG (mmHg: rest, 22.5 ± 1.0; Lo, 15.3 ± 1.3; Hi, 10.2 ± 1.6) and WG (mmHg: rest, 19.0 ± 1.3; Lo, 12.2 ± 1.1; Hi, 9.9 ± 1.1) than in Sol (rest, 33.1 ± 3.2 mmHg; Lo, 19.0 ± 2.3 mmHg; Hi, 18.7 ± 1.8 mmHg), despite lower and in MG and WG under each set of conditions. These data suggest that during submaximal metabolic rates, the relationship between and O2 extraction is dependent on fibre type (at least in the muscles studied herein), such that muscles comprised of fast‐twitch fibres display a greater fractional O2 extraction (i.e. lower P  mvO 2) than their slow‐twitch counterparts. These results also indicate that the greater sustained P  mvO 2 in Sol may be important for ensuring high blood–myocyte O2 flux and therefore a greater oxidative contribution to energetic requirements.

Dynamics of microvascular oxygen pressure across the rest-exercise transition in rat skeletal muscle.

There exists substantial controversy as to whether muscle oxygen (O2) delivery (QO2) or muscle mitochondrial O2 demand determines the profile of pulmonary VO2 kinetics in the rest-exercise transition. To address this issue, we adapted intravascular phosphorescence quenching techniques for measurement of rat spinotrapezius microvascular O2 pressure (PO2m). The spinotrapezius muscle intravital microscopy preparation is used extensively for investigation of muscle microcirculatory control. The phosphor palladium-meso-tetra(4-carboxyphenyl)porphyrin dendrimer (R2) at 15 mg/kg was bound to albumin within the blood of female Sprague-Dawley rats ( approximately 250 g). Spinotrapezius blood flow (radioactive microspheres) and PO2m profiles were determined in situ across the transition from rest to 1 Hz twitch contractions. Stimulation increased muscle blood flow by 240% from 16.6 +/- 3.0 to 56.2 +/- 8.3 (SE) ml/min per 100 g (P < 0.05). Muscle contractions reduced PO2m from a baseline of 31.4 +/- 1.6 to a steady-state value of 21.0 +/- 1.7 mmHg (n = 24, P < 0.01). The response profile of PO2m was well fit by a time delay of 19.2+/-2.8 sec (P < 0.05) followed by a monoexponential decline (time constant, 21.7 +/- 2.1 sec) to its steady state level. The absence of either an immediate and precipitous fall in microvascular PO2 at exercise onset or any PO2m undershoot prior to achievement of steady-state values, provides compelling evidence that O(2) delivery is not limiting under these conditions.

Impact of dietary nitrate supplementation via beetroot juice on exercising muscle vascular control in rats

•  Inorganic nitrate (NO3−) supplementation with beetroot juice (BR) in humans lowers blood pressure and the O2 cost of exercise and may improve exercise tolerance following its reduction to nitrite (NO2−) and nitric oxide (NO). •  The effect of inorganic NO3− supplementation with BR on skeletal muscle blood flow (BF) and vascular conductance (VC) within and among locomotory muscles during exercise is unknown. •  Inorganic NO3− 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 NO3− supplementation improves vascular control and elevates skeletal muscle O2 delivery during exercise predominantly in fast‐twitch type II muscles, and provide a potential mechanism by which NO3− supplementation improves metabolic control.

Oxygen exchange profile in rat muscles of contrasting fibre types.

To determine whether fibre type affects the O2 exchange characteristics of skeletal muscle at the microcirculatory level we tested the hypothesis that, following the onset of contractions, muscle comprising predominately type I fibres (soleus, Sol, 86 % type I) would, based on demonstrated blood flow responses, exhibit a blunted microvascular PO2 (PO2,m, which is determined by the O2 delivery (Q̇O2) to O2 uptake (V̇O2) ratio) profile (assessed via phosphorescence quenching) compared to muscle of primarily type II fibres (peroneal, Per, 84 % type II). PO2,m was measured at rest, and following the rest‐contractions (twitch, 1 Hz, 2–4 V for 120 s) transition in Sol (n= 6) and Per (n= 6) muscles of Sprague‐Dawley rats. Both muscles exhibited a delay followed by a mono‐exponential decrease in PO2,m to the steady state. However, compared with Sol, Per demonstrated (1) a larger change in baseline minus steady state contracting PO2,m (ΔPO2,m) (Per, 13.4 ± 1.7 mmHg; Sol, 8.6 ± 0.9 mmHg, P < 0.05); (2) a faster mean response time (i.e. time delay (TD) plus time constant (τ); Per, 23.8 ± 1.5 s; Sol, 39.6 ± 4.3 s, P < 0.05); and therefore (3) a greater rate of PO2,m decline (ΔPO2,m/τ; Per, 0.92 ± 0.08 mmHg s−1; Sol, 0.42 ± 0.05 mmHg s−1, P < 0.05). These data demonstrate an increased microvascular pressure head of O2 at any given point after the initial time delay for Sol versus Per following the onset of contractions that is probably due to faster Q̇O2 dynamics relative to those of V̇O2.

Dynamics of oxygen uptake following exercise onset in rat skeletal muscle

Technical limitations have precluded measurement of the V(O(2)) profile within contracting muscle (mV(O(2))) and hence it is not known to what extent V(O(2)) dynamics measured across limbs in humans or muscles in the dog are influenced by transit delays between the muscle microvasculature and venous effluent. Measurements of capillary red blood cell flux and microvascular P(O(2)) (P(O(2)m)) were combined to resolve the time course of mV(O(2)) across the rest-stimulation transient (1 Hz, twitch contractions). mV(O(2)) began to rise at the onset of contractions in a close to monoexponential fashion (time constant, J = 23.2 +/- 1.0 sec) and reached its steady-state value at 4.5-fold above baseline. Using computer simulation in healthy and disease conditions (diabetes and chronic heart failure), our findings suggest that: (1) mV(O(2)) increases essentially immediately (< 2 sec) following exercise onset; (2) within healthy muscle the J blood flow (thus O(2) delivery, J Q(O(2)m)) is faster than JmV(O(2)) such that oxygen delivery is not limiting, and 3) a faster P(O(2)m) fall to a P(O(2)m) value below steady-state values within muscle from diseased animals is consistent with a relatively sluggish Q(O(2)m) response compared to that of mV(O(2)).

Dynamics of microvascular oxygen partial pressure in contracting skeletal muscle of rats with chronic heart failure.

OBJECTIVE This investigation tested the hypothesis that the dynamics of muscle microvascular O(2) pressure (PO(2)m, which reflects the ratio of O(2) utilization [V*O(2)] to O(2) delivery [Q*O(2)]) following the onset of contractions would be altered in chronic heart failure (CHF). METHODS Female Sprague-Dawley rats were subjected to a myocardial infarction (MI) or a sham operation (Sham). Six to 10 weeks post Sham (n=6) or MI (n=17), phosphorescence quenching techniques were utilized to determine PO(2)m dynamics at the onset of spinotrapezius muscle contractions (1 Hz). RESULTS MI rats were separated into groups with Moderate (n=10) and Severe (n=7) CHF based upon the degree of left ventricular (LV) dysfunction as indicated by structural abnormalities (increased right ventricle weight and lung weight normalized to body weight). LV end-diastolic pressure was elevated significantly in both CHF groups compared with Sham (Sham, 3+/-1; Moderate CHF, 9+/-2; Severe CHF, 27+/-4 mmHg, P<0.05). The PO(2)m response was modeled using time delay and exponential components to fit the PO(2)m response to the steady-state. Compared with Shams, the time constant (tau) of the primary PO(2)m response was significantly speeded in Moderate CHF (tau, Sham, 19.0+/-1.5; Moderate CHF, 13.2+/-1.9 s, P<0.05) and slowed in Severe CHF (tau, 28.2+/-3.4 s, P<0.05). Within the Severe CHF group, tau increased linearly with the product of right ventricular and lung weight (r=0.83, P<0.05). CONCLUSIONS These results suggest that CHF alters the dynamic matching of muscle V*O(2)-to-Q*O(2) across the transition from rest to contractions and that the nature of that perturbation is dependent upon the severity of cardiac dysfunction.

Effects of aging on microvascular oxygen pressures in rat skeletal muscle

Aging alters skeletal muscle vascular geometry and control such that the dynamics of muscular blood flow (Q) and O2 delivery (Q(O2)) may be impaired across the rest-exercise transition. If, at the onset of muscle contractions, Q dynamics are slowed disproportionately to those of muscle O2 uptake (V(O2), microvascular PO2 (PO2m) would be reduced and blood-tissue O2 transfer compromised. This investigation determined the effects of aging on PO2m (a direct reflection of the Q(O2)-to-V(O2) ratio), at rest and across the rest-contractions transition in the spinotrapezius of young (approximately 6 months, n = 9) and old (>24 months, n = 10) male Fisher 344/Brown Norway hybrid rats. Phosphorescence quenching techniques were used to quantify PO2m, and test the hypothesis that, across the rest-contractions (twitch, 1 Hz; 4-6 V, 240 s) transition, aging would transiently reduce the Q(O2)-to-V(O2) ratio causing a biphasic profile in which PO2m fell below steady-state contracting values. Old rats had a lower pre-contraction baseline PO2m than young (27.1+/-1.9 versus 33.8+/-1.6 mmHg, P<0.05, respectively). In addition, in old rats PO2m demonstrated a pronounced difference between the absolute nadir and end-contracting values (2.5+/-0.9 mmHg), which was absent in young rats. In conclusion, unlike their young counterparts, old rats exhibited a transiently reduced PO2m across the rest-contractions transition that may impair blood-tissue O2 exchange and elevate the O2 deficit, thereby contributing to premature fatigue.

Effects of prior contractions on muscle microvascular oxygen pressure at onset of subsequent contractions

In humans, pulmonary oxygen uptake (V̇O2) kinetics may be speeded by prior exercise in the heavy domain. This ‘speeding’ arises potentially as the result of an increased muscle O2 delivery (Q̇O2) and/or a more rapid elevation of oxidative phosphorylation. We adapted phosphorescence quenching techniques to determine the QO2‐to‐O2 utilization (Q̇O2/V̇O2) characteristics via microvascular O2 pressure (PO2,m) measurements across sequential bouts of contractions in rat spinotrapezius muscle. Spinotrapezius muscles from female Sprague‐Dawley rats (n= 6) were electrically stimulated (1 Hz twitch, 3–5 V) for two 3 min bouts (ST1 and ST2) separated by 10 min rest. PO2,m responses were analysed using an exponential + time delay (TD) model. There was no significant difference in baseline and ΔPO2,m between ST1 and ST2 (28.5 ± 2.6 vs. 27.9 ± 2.4 mmHg, and 13.9 ± 1.8 vs. 14.1 ± 1.3 mmHg, respectively). The TD was reduced significantly in the second contraction bout (ST1, 12.2 ± 1.9; ST2, 5.7 ± 2.2 s, P < 0.05), whereas the time constant of the exponential PO2,m decrease was unchanged (ST1, 16.3 ± 2.6; ST2, 17.6 ± 2.7 s, P > 0.1). The shortened TD found in ST2 led to a reduced time to reach 63 % of the final response of ST2 compared to ST1 (ST1, 28.3 ± 3.0; ST2, 20.2 ± 1.8 s, P < 0.05). The speeding of the overall response in the absence of an elevated PO2,m baseline (which had it occurred would indicate an elevated QO2/V̇O2) or muscle blood flow suggests that some intracellular process(es) (e.g. more rapid increase in oxidative phosphorylation) may be responsible for the increased speed of PO2,m kinetics after prior contractions under these conditions.

Dynamic heterogeneity of exercising muscle blood flow and O2 utilization.

Resolving the bases for different physiological functioning or exercise performance within a population is dependent on our understanding of control mechanisms. For example, when most young healthy individuals run or cycle at moderate intensities, oxygen uptake (VO2) kinetics are rapid and the amplitude of the VO2 response is not constrained by O2 delivery. For this to occur, muscle O2 delivery (i.e., blood flow × arterial O2 concentration) must be coordinated superbly with muscle O2 requirements (VO2), the efficacy of which may differ among muscles and distinct fiber types. When the O2 transport system succumbs to the predations of aging or disease (emphysema, heart failure, and type 2 diabetes), muscle O2 delivery and O2 delivery-VO2 matching and, therefore, muscle contractile function become impaired. This forces greater influence of the upstream O2 transport pathway on muscle aerobic energy production, and the O2 delivery-VO2 relationship(s) assumes increased importance. This review is the first of its kind to bring a broad range of available techniques, mostly state of the art, including computer modeling, radiolabeled microspheres, positron emission tomography, magnetic resonance imaging, near-infrared spectroscopy, and phosphorescence quenching to resolve the O2 delivery-VO2 relationships and inherent heterogeneities at the whole body, interorgan, muscular, intramuscular, and microvascular/myocyte levels. Emphasis is placed on the following: 1) intact humans and animals as these provide the platform essential for framing and interpreting subsequent investigations, 2) contemporary findings using novel technological approaches to elucidate O2 delivery-VO2 heterogeneities in humans, and 3) future directions for investigating how normal physiological responses can be explained by O2 delivery-VO2 heterogeneities and the impact of aging/disease on these processes.

Blood flow and O2 extraction as a function of O2 uptake in muscles composed of different fiber types.

We examined how the greater vasodilatory capacity of slow--(ST) versus fast-twitch (FT) muscles impacts the relationship between blood flow (Q ) and O2 uptake (VO2) and, consequently, the O2 extraction (a-vO2 diff.)-to-VO2 relationship. Q was measured with radiolabelled microspheres, while VO2 was calculated by the Fick principle using measurements of microvascular O2 pressure (phosphorescence quenching) at rest, low--(2.5 V) and high-intensity contractions (4.5 V) for soleus (Sol; ST, n=5), mixed-gastrocnemius (MG; FT, n=7) and white-gastrocnemius (WG; FT, n=7). The slope of the Q-to-VO2 relationship (delta Q/delta VO2] ) was not different among muscles (Sol = 5.5 +/- 0.2, MG = 6.0 +/- 0.11 and WG = 5.8 +/- 0.06; P > 0.05). In contrast, the intercept was greater (P < 0.05) for Sol (16.3 +/- 2.7 ml min(-1) 100 g(-1)) versus MG and WG (in ml min(-1) 100 g(-1): 1.39 +/- 0.26 and 1.45 +/- 0.23, respectively; MG and WG, P > 0.05). In addition, the a-vO2 diff.-to-VO2] relationship for Sol was shifted rightward compared to MG and WG. These data suggest that the increase in Q for a given change in VO2 is similar for slow- and fast-twitch muscles, at least for the range of metabolic rates and muscles studied herein and that a-vO2 diff. differences result from the lower resting Q in FT muscles.