Novel mechanosensory role for acid sensing ion channel subtype 1a in evoking the exercise pressor reflex in rats with heart failure.

Mechanical and metabolic signals associated with skeletal muscle contraction stimulate the sensory endings of thin fibre muscle afferents, which, in turn, generates reflex increases in sympathetic nerve activity (SNA) and blood pressure (the exercise pressor reflex; EPR). EPR activation in patients and animals with heart failure with reduced ejection fraction (HF-rEF) results in exaggerated increases in SNA and promotes exercise intolerance. In the healthy decerebrate rat, a subtype of acid sensing ion channel (ASIC) on the sensory endings of thin fibre muscle afferents, namely ASIC1a, has been shown to contribute to the metabolically sensitive portion of the EPR (i.e. metaboreflex), but not the mechanically sensitive portion of the EPR (i.e. the mechanoreflex). However, the role played by ASIC1a in evoking the EPR in HF-rEF is unknown. We hypothesized that, in decerebrate, unanaesthetized HF-rEF rats, injection of the ASIC1a antagonist psalmotoxin-1 (PcTx-1; 100 ng) into the hindlimb arterial supply would reduce the reflex increase in renal SNA (RSNA) evoked via 30 s of electrically induced static hindlimb muscle contraction, but not static hindlimb muscle stretch (model of mechanoreflex activation isolated from contraction-induced metabolite-production). We found that PcTx-1 reduced the reflex increase in RSNA evoked in response to muscle contraction (n = 8; mean (SD) ∫ΔRSNA pre: 1343 (588) a.u.; post: 816 (573) a.u.; P = 0.026) and muscle stretch (n = 6; ∫ΔRSNA pre: 688 (583) a.u.; post: 304 (370) a.u.; P = 0.025). Our data suggest that, in HF-rEF rats, ASIC1a contributes to activation of the exercise pressor reflex and that contribution includes a novel role for ASIC1a in mechanosensation that is not present in healthy rats. KEY POINTS: Skeletal muscle contraction results in exaggerated reflex increases in sympathetic nerve activity in heart failure patients compared to healthy counterparts, which likely contributes to increased cardiovascular risk and impaired tolerance for even mild exercise (i.e. activities of daily living) for patients suffering with this condition. Activation of acid sensing ion channel subtype 1a (ASIC1a) on the sensory endings of thin fibre muscle afferents during skeletal muscle contraction contributes to reflex increases in sympathetic nerve activity and blood pressure, at least in healthy subjects. In this study, we demonstrate that ASIC1a on the sensory endings of thin fibre muscle afferents plays a role in both the mechanical and metabolic components of the exercise pressor reflex in male rats with heart failure. The present data identify a novel role for ASIC1a in evoking the exercise pressor reflex in heart failure and may have important clinical implications for heart failure patients.

Capillary hemodynamics and contracting skeletal muscle oxygen pressures in male rats with heart failure: Impact of soluble guanylyl cyclase activator.

In heart failure with reduced ejection fraction (HFrEF), nitric oxide-soluble guanylyl cyclase (sGC) pathway dysfunction impairs skeletal muscle arteriolar vasodilation and thus capillary hemodynamics, contributing to impaired oxygen uptake (V̇O2) kinetics. Targeting this pathway with sGC activators offers a new treatment approach to HFrEF. We tested the hypotheses that sGC activator administration would increase the O2 delivery (Q̇O2)-to-V̇O2 ratio in the skeletal muscle interstitial space (PO2is) of HFrEF rats during twitch contractions due, in part, to increases in red blood cell (RBC) flux (fRBC), velocity (VRBC), and capillary hematocrit (Hctcap). HFrEF was induced in male Sprague-Dawley rats via myocardial infarction. After 3 weeks, rats were treated with 0.3 mg/kg of the sGC activator BAY 60-2770 (HFrEF + BAY; n = 11) or solvent (HFrEF; n = 9) via gavage b.i.d for 5 days prior to phosphorescence quenching (PO2is, in contracting muscle) and intravital microscopy (resting) measurements in the spinotrapezius muscle. Intravital microscopy revealed higher fRBC (70 ± 9 vs 25 ± 8 RBC/s), VRBC (490 ± 43 vs 226 ± 35 µm/s), Hctcap (16 ± 1 vs 10 ± 1%) and a greater number of capillaries supporting flow (91 ± 3 vs 82 ± 3%) in HFrEF + BAY vs HFrEF (all P < 0.05). Additionally, PO2is was especially higher during 12-34s of contractions in HFrEF + BAY vs HFrEF (P < 0.05). Our findings suggest that sGC activators improved resting Q̇O2 via increased fRBC, VRBC, and Hctcap allowing for better Q̇O2-to-V̇O2 matching during the rest-contraction transient, supporting sGC activators as a potential therapeutic to target skeletal muscle vasomotor dysfunction in HFrEF.

Sexual dimorphism in vascular ATP-sensitive K+ channel function supporting interstitial PO2 via convective and/or diffusive O2 transport.

KEY POINTS: Inhibition of pancreatic ATP-sensitive K+ (KATP ) channels is the intended effect of oral sulphonylureas to increase insulin release in diabetes. However, pertinent to off-target effects of sulphonylurea medication, sex differences in cardiac KATP channel function exist, whereas potential sex differences in vascular KATP channel function remain unknown. In the present study, we assessed vascular KATP channel function (topical glibenclamide superfused onto fast-twitch oxidative skeletal muscle) supporting blood flow and interstitial O2 delivery-utilization matching ( PO2 is) during twitch contractions in male, female during pro-oestrus and ovariectomized female (F+OVX) rats. Glibenclamide decreased blood flow (convective O2 transport) and interstitial PO2 in male and female, but not F+OVX, rats. Compared to males, females also demonstrated impaired diffusive O2 transport and a faster fall in interstitial PO2 . Our demonstration, in rats, that sex differences in vascular KATP channel function exist support the tentative hypothesis that oral sulphonylureas may exacerbate exercise intolerance and morbidity, especially in premenopausal females. ABSTRACT: Vascular ATP-sensitive K+ (KATP ) channels support skeletal muscle blood flow ( Q̇m ), interstitial O2 delivery ( Q̇O2 )-utilization ( V̇O2 ) matching (i.e. interstitial-myocyte O2 flux driving pressure; PO2 is) and exercise tolerance. Potential sex differences in skeletal muscle vascular KATP channel function remain largely unexplored. We hypothesized that local skeletal muscle KATP channel inhibition via glibenclamide superfusion (5 mg kg-1 GLI; sulphonylurea diabetes medication) in anaesthetized female Sprague-Dawley rats, compared to males, would demonstrate greater reductions in contracting (1 Hz, 7 V, 180 s) fast-twitch oxidative mixed gastrocnemius (97% type IIA+IID/X+IIB) Q̇m (15 µm microspheres) and PO2 is (phosphorescence quenching), resulting from more compromised convective ( Q̇O2 ) and diffusive ( DO2  ) O2 conductances. Furthermore, these GLI-induced reductions in ovary-intact females measured during pro-oestrus would be diminished following ovariectomy (F+OVX). GLI similarly impaired mixed gastrocnemius V̇O2 in both males (↓28%) and females (↓33%, both P < 0.032) via reduced Q̇m (male: ↓31%, female: ↓35%, both P < 0.020), Q̇O2 (male: 5.6 ± 0.5 vs. 4.0 ± 0.5, female: 6.4 ± 1.1 vs. 4.2 ± 0.6 mL O2  min-1 100 g tissue-1 , P < 0.022) and the resulting PO2 is, with females also demonstrating a reduced DO2  (0.40 ± 0.07 vs. 0.30 ± 0.04 mL O2  min-1 100 g tissue-1 , P < 0.042) and a greater GLI-induced speeding of PO2 is fall (mean response time: Sex × Drug interaction, P = 0.026). Conversely, GLI did not impair the mixed gastrocnemius of F+OVX rats. Therefore, in patients taking sulphonylureas, these results support the potential for impaired vascular KATP channel function to compromise muscle Q̇m and therefore exercise tolerance. Such an effect, if present, would likely contribute to adverse cardiovascular events in premenopausal females more than males.

Vascular ATP-sensitive K+ channels support maximal aerobic capacity and critical speed via convective and diffusive O2 transport.

KEY POINTS: Oral sulphonylureas, widely prescribed for diabetes, inhibit pancreatic ATP-sensitive K+ (KATP ) channels to increase insulin release. However, KATP channels are also located within vascular (endothelium and smooth muscle) and muscle (cardiac and skeletal) tissue. We evaluated left ventricular function at rest, maximal aerobic capacity ( V̇ O2 max) and submaximal exercise tolerance (i.e. speed-duration relationship) during treadmill running in rats, before and after systemic KATP channel inhibition via glibenclamide. Glibenclamide impaired critical speed proportionally more than V̇ O2 max but did not alter resting cardiac output. Vascular KATP channel function (topical glibenclamide superfused onto hindlimb skeletal muscle) resolved a decreased blood flow and interstitial PO2 during twitch contractions reflecting impaired O2 delivery-to-utilization matching. Our findings demonstrate that systemic KATP channel inhibition reduces V̇ O2 max and critical speed during treadmill running in rats due, in part, to impaired convective and diffusive O2 delivery, and thus V̇ O2 , especially within fast-twitch oxidative skeletal muscle. ABSTRACT: Vascular ATP-sensitive K+ (KATP ) channels support skeletal muscle blood flow and microvascular oxygen delivery-to-utilization matching during exercise. However, oral sulphonylurea treatment for diabetes inhibits pancreatic KATP channels to enhance insulin release. Herein we tested the hypotheses that: i) systemic KATP channel inhibition via glibenclamide (GLI; 10 mg kg-1 i.p.) would decrease cardiac output at rest (echocardiography), maximal aerobic capacity ( V̇ O2 max) and the speed-duration relationship (i.e. lower critical speed (CS)) during treadmill running; and ii) local KATP channel inhibition (5 mg kg-1 GLI superfusion) would decrease blood flow (15 µm microspheres), interstitial space oxygen pressures (PO2 is; phosphorescence quenching) and convective and diffusive O2 transport ( Q̇ O2 and DO2 , respectively; Fick Principle and Law of Diffusion) in contracting fast-twitch oxidative mixed gastrocnemius muscle (MG: 9% type I+IIa fibres). At rest, GLI slowed left ventricular relaxation (2.11 ± 0.59 vs. 1.70 ± 0.23 cm s-1 ) and decreased heart rate (321 ± 23 vs. 304 ± 22 bpm, both P < 0.05) while cardiac output remained unaltered (219 ± 64 vs. 197 ± 39 ml min-1 , P > 0.05). During exercise, GLI reduced V̇ O2 max (71.5 ± 3.1 vs. 67.9 ± 4.8 ml kg-1 min-1 ) and CS (35.9 ± 2.4 vs. 31.9 ± 3.1 m min-1 , both P < 0.05). Local KATP channel inhibition decreased MG blood flow (52 ± 25 vs. 34 ± 13 ml min-1 100 g tissue-1 ) and PO2 isnadir (5.9 ± 0.9 vs. 4.7 ± 1.1 mmHg) during twitch contractions. Furthermore, MG V̇ O2 was reduced via impaired Q̇ O2 and DO2 (P < 0.05 for each). Collectively, these data support that vascular KATP channels help sustain submaximal exercise tolerance in healthy rats. For patients taking sulfonylureas, KATP channel inhibition may exacerbate exercise intolerance.

Neuronal nitric oxide synthase regulation of skeletal muscle functional hyperemia: exercise training and moderate compensated heart failure.

Nitric oxide (NO) modulates oxygen delivery-utilization matching in resting and contracting skeletal muscle. Recent reports indicate that neuronal NO synthase (nNOS)-mediated vasoregulation during contractions is enhanced with exercise training and impaired with chronic heart failure (HF). Consequently, we tested the hypothesis that selective nNOS inhibition (S-methyl-l-thiocitrulline; SMTC, 2.1 µmol/kg) would produce attenuated reductions in muscle blood flow during moderate/heavy submaximal exercise in sedentary HF rats compared to their healthy counterparts. In addition, SMTC was expected to evoke greater reductions in exercising muscle blood flow in trained compared to sedentary healthy and HF rats. Blood flow during submaximal treadmill running (20 min/m, 5% grade) was determined via radiolabeled microspheres pre- and post-SMTC administration in healthy sedentary (Healthy + Sed, n = 8), healthy exercise trained (Healthy + ExT, n = 8), HF sedentary (HF + Sed, left ventricular end-diastolic pressure (LVEDP) = 12 ± 1 mmHg, n = 8), and HF exercise trained (HF + ExT, LVEDP = 16 ± 2 mmHg, n = 7) rats. nNOS contribution to exercising total hindlimb blood flow (ml/min/100 g) was not increased by training in either healthy or HF groups (Healthy + Sed: 105 ± 11 vs. 108 ± 16; Healthy + ExT: 96 ± 9 vs. 91 ± 7; HF + Sed: 124 ± 6 vs. 110 ± 12; HF + ExT: 107 ± 13 vs. 101 ± 8; control vs. SMTC, respectively; p > .05 for all). Similarly, SMTC did not reduce exercising blood flow in the majority of individual hindlimb muscles in any group (p > .05 for all, except for the semitendinosus and adductor longus in HF + Sed and the adductor longus in HF + ExT; p < .05). Contrary to our hypothesis, we find no support for either upregulation of nNOS function contributing to exercise hyperemia after training or its dysregulation with chronic HF.

Effects of comorbid type II diabetes mellitus and heart failure on rat hindlimb and respiratory muscle blood flow during treadmill exercise.

In rats with type II diabetes mellitus (T2DM) compared with nondiabetic healthy controls, muscle blood flow (Q̇m) to primarily glycolytic hindlimb muscles and the diaphragm muscle are elevated during submaximal treadmill running consequent to lower skeletal muscle mass, a finding that held even when muscle mass was normalized to body mass. In rats with heart failure with reduced ejection fraction (HF-rEF) compared with healthy controls, hindlimb Q̇m was lower, whereas diaphragm Q̇m is elevated during submaximal treadmill running. Importantly, T2DM is the most common comorbidity present in patients with HF-rEF, but the effect of concurrent T2DM and HF-rEF on limb and respiratory Q̇m during exercise is unknown. We hypothesized that during treadmill running (20 m·min-1; 10% incline), hindlimb and diaphragm Q̇m would be higher in T2DM Goto-Kakizaki rats with HF-rEF (i.e., HF-rEF + T2DM) compared with nondiabetic Wistar rats with HF-rEF. Ejection fractions were not different between groups (HF-rEF: 30 ± 5; HF-rEF + T2DM: 28 ± 8%; P = 0.617), whereas blood glucose was higher in HF-rEF + T2DM (209 ± 150 mg/dL) compared with HF-rEF rats (113 ± 28 mg/dL; P = 0.040). Hindlimb muscle mass normalized to body mass was lower in rats with HF-rEF + T2DM (36.3 ± 1.6 mg/g) than in nondiabetic HF-rEF counterparts (40.3 ± 2.7 mg/g; P < 0.001). During exercise, Q̇m was elevated in rats with HF-rEF + T2DM compared with nondiabetic counterparts to the hindlimb (HF-rEF: 100 ± 28; HF-rEF + T2DM: 139 ± 23 mL·min-1·100 g-1; P < 0.001) and diaphragm (HF-rEF: 177 ± 66; HF-rEF + T2DM: 215 ± 93 mL·min-1·100g-1; P = 0.035). These data suggest that the pathophysiological consequences of T2DM on hindlimb and diaphragm Q̇m during treadmill running in the GK rat persist even in the presence of HF-rEF.NEW & NOTEWORTHY Herein, we demonstrate that rats comorbid with heart failure with reduced ejection fraction (HF-rEF) and type II diabetes mellitus (T2DM) have a higher hindlimb and respiratory muscle blood flow during submaximal treadmill running (20 m·min-1; 10% incline) compared with nondiabetic HF-rEF counterparts. These data may carry important clinical implications for roughly half of all patients with HF-rEF who present with T2DM.

Angiotensin converting enzyme inhibition improves cerebrovascular control during exercise in male rats with heart failure.

We investigated the effects of chronic (∼7 weeks) treatment with the angiotensin converting enzyme (ACE) inhibitor Captopril in rats with heart failure with reduced ejection fraction (HF-rEF) on brain blood flow (BF; radiolabeled microspheres) at rest and during submaximal exercise. We hypothesized that middle cerebral, posterior cerebral, and cerebellar BF during submaximal exercise (20 m/min, 5% incline) would be reduced in rats with HF-rEF (n = 10) compared to healthy (SHAM, n = 10) controls and HF-rEF rats chronically treated with Captopril (HF-rEF + Cap., n = 20). During submaximal exercise middle cerebral (HF-rEF + Cap.: 274 ± 12; HF-rEF: 234 ± 23; SHAM: 248 ± 24 ml/min/100 g) and cerebellar (HF-rEF + Cap.: 222 ± 14; HF-rEF: 243 ± 22; SHAM: 214 ± 23 ml/min/100 g) BF increased from rest in all groups with no difference among groups (P > 0.24). Posterior cerebral BF increased from rest in all groups but was lower than SHAM (394 ± 46 ml/min/100 g; P = 0.03) in HF-rEF (298 ± 19 ml/min/100 g) but not HF-rEF + Cap. (356 ± 18 ml/min/100 g; P = 0.14), supporting the concept that ACE inhibition in HF-rEF elevates brain BF increases, at least to the posterior cerebral region, during moderate intensity exercise/physical activity.

Exaggerated sympathetic and cardiovascular responses to dynamic mechanoreflex activation in rats with heart failure: Role of endoperoxide 4 and thromboxane A2 receptors.

The primary purpose of this investigation was to determine the role played by endoperoxide 4 receptors (EP4-R) and thromboxane A2 receptors (TxA2-R) during isolated dynamic muscle mechanoreflex activation in rats with heart failure with reduced ejection fraction (HF-rEF) and sham-operated healthy controls. We found that injection of the EP4-R antagonist L-161,982 (1 µg) into the arterial supply of the hindlimb had no effect on the peak pressor response to dynamic hindlimb muscle stretch in HF-rEF (n = 6, peak ∆MAP pre: 27 ± 7; post: 27 ± 4 mm Hg; P = 0.99) or sham (n = 6, peak ∆MAP pre: 15 ± 3; post: 13 ± 3 mm Hg; P = 0.67) rats. In contrast, injection of the TxA2-R antagonist daltroban (80 µg) into the arterial supply of the hindlimb reduced the pressor response to dynamic hindlimb muscle stretch in HF-rEF (n = 11, peak ∆MAP pre: 28 ± 4; post: 16 ± 2 mm Hg; P = 0.02) but not sham (n = 8, peak ∆MAP pre: 17 ± 3; post: 16 ± 3; P = 0.84) rats. Our data suggest that TxA2-Rs on thin fibre muscle afferents contribute to the exaggerated mechanoreflex in HF-rEF.

Exercise training decreases intercostal and transversus abdominis muscle blood flows in heart failure rats during submaximal exercise.

Diaphragm muscle blood flow (BF) and vascular conductance (VC) are elevated with chronic heart failure (HF) during exercise. Exercise training (ExT) elicits beneficial respiratory muscle and pulmonary system adaptations in HF. We hypothesized that diaphragm BF and VC would be lower in HF rats following ExT than their sedentary counterparts (Sed). Respiratory muscle BFs and mean arterial pressure were measured via radiolabeled microspheres and carotid artery catheter, respectively, during submaximal treadmill exercise (20 m/min, 5 % grade). During exercise, no differences were present between HF + ExT and HF + Sed in diaphragm BFs (201 ± 36 vs. 227 ± 44 mL/min/100 g) or VCs (both, p > 0.05). HF + ExT compared to HF + Sed had lower intercostal BF (27 ± 3 vs. 41 ± 5 mL/min/100 g) and VC (0.21 ± 0.02 vs. 0.31 ± 0.04 mL/min/mmHg/100 g) during exercise (both, p < 0.05). Further, HF + ExT compared to HF + Sed had lower transversus abdominis BF (20 ± 1 vs. 35 ± 6 mL/min/100 g) and VC (0.14 ± 0.02 vs. 0.27 ± 0.05 mL/min/mmHg/100 g) during exercise (both, p < 0.05). These data suggest that exercise training lowers the intercostal and transversus abdominis BF responses in HF rats during submaximal treadmill exercise.

Thromboxane A2 receptors contribute to the exaggerated exercise pressor reflex in male rats with heart failure.

Mechanical and metabolic signals associated with skeletal muscle contraction stimulate the sensory endings of thin fiber muscle afferents and produce reflex increases in sympathetic nerve activity and blood pressure during exercise (i.e., the exercise pressor reflex; EPR). The EPR is exaggerated in patients and animals with heart failure with reduced ejection fraction (HF-rEF) and its activation contributes to reduced exercise capacity within this patient population. Accumulating evidence suggests that the exaggerated EPR in HF-rEF is partially attributable to a sensitization of mechanically activated channels produced by thromboxane A2 receptors (TxA2 -Rs) on those sensory endings; however, this has not been investigated. Accordingly, the purpose of this investigation was to determine the role played by TxA2 -Rs on the sensory endings of thin fiber muscle afferents in the exaggerated EPR in rats with HF-rEF induced by coronary artery ligation. In decerebrate, unanesthetized rats, we found that injection of the TxA2 -R antagonist daltroban (80 µg) into the arterial supply of the hindlimb reduced the pressor response to 30 s of electrically induced 1 Hz dynamic hindlimb muscle contraction in HF-rEF (n = 8, peak ∆MAP pre: 22 ± 3; post: 14 ± 2 mmHg; p = 0.01) but not sham (n = 10, peak ∆MAP pre: 13 ± 3; post: 11 ± 2 mmHg; p = 0.68) rats. In a separate group of HF-rEF rats (n = 4), we found that the systemic (intravenous) injection of daltroban had no effect on the EPR (peak ΔMAP pre: 26 ± 7; post: 25 ± 7 mmHg; p = 0.50). Our data suggest that TxA2 -Rs on thin fiber muscle afferents contribute to the exaggerated EPR evoked in response to dynamic muscle contraction in HF-rEF.

Nitric oxide synthase inhibition during treadmill exercise reveals fiber-type specific vascular control in the rat hindlimb.

The control of vascular tone during exercise is highly complex and integrated. Specifically, in regards to the contribution of nitric oxide (NO), the observed magnitude and muscle fiber-type dependency of the NO contribution to exercise hyperemia may differ depending on the timing of NO synthase (NOS) inhibition with respect to the exercise bout (i.e., administration prior to vs. during exercise). We tested the hypothesis that, in the presence of prior cyclooxygenase inhibition (indomethacin, 5 mg/kg(-1)), NOS inhibition (N(G)-nitro-L-arginine methyl ester, L-NAME; 10 mg/kg) administered during submaximal treadmill exercise would blunt blood flow and vascular conductance (VC) in the hindlimb muscle(s) of the rat with the greatest reductions in blood flow and VC occurring in the predominantly oxidative muscles. Adult female Wistar rats (n = 10, age: 3-4 mo) ran on a motor-driven treadmill (20 m/min, 10% grade). Total and regional hindlimb muscle blood flow and VC were determined via radiolabeled microspheres before (control) and after L-NAME administration during exercise. L-NAME reduced (P < 0.05) total hindlimb muscle blood flow (control: 123 + or - 10, L-NAME: 103 + or - 7 ml x min(-1) x 100g(-1)) and VC (control: 0.95 + or - 0.09, L-NAME: 0.63 + or - 0.05 ml x min(-1) x 100g(-1) x mmHg(-1)). There was a significant correlation (r = 0.51, P < 0.05) between the absolute reductions in VC after L-NAME and the percent sum of type I and IIa fibers in the individual muscles and muscle parts; however, there was no correlation (P = 0.62) when expressed as blood flow. Surprisingly, the highly oxidative muscles demonstrated a marked ability to maintain oxygen delivery, which differs substantially from previous reports of L-NAME infusion prior to exercise in these muscles. The demonstration that NO is an important regulator of blood flow and VC in the rat hindlimb during treadmill exercise, but that the fiber-type dependency of NO is altered markedly when NOS inhibition is performed during, vs. prior to, exercise, lends important insights into the integrated nature of vascular control during exercise.

Dietary nitrate supplementation opposes the elevated diaphragm blood flow in chronic heart failure during submaximal exercise.

Chronic heart failure (CHF) results in a greater cost of breathing and necessitates an elevated diaphragm blood flow (BF). Dietary nitrate (NO3‾) supplementation lowers the cost of exercise. We hypothesized that dietary NO3‾ supplementation would attenuate the CHF-induced greater cost of breathing and thus the heightened diaphragm BF during exercise. CHF rats received either 5days of NO3‾-rich beetroot (BR) juice (CHF+BR, n=10) or a placebo (CHF, n=10). Respiratory muscle BFs (radiolabeled microspheres) were measured at rest and during submaximal exercise (20m/min, 5% grade). Infarcted left ventricular area and normalized lung weight were not significantly different between groups. During submaximal exercise, diaphragm BF was markedly lower for CHF+BR than CHF (CHF+BR: 195±28; CHF: 309±71mL/min/100g, p=0.04). The change in diaphragm BF from rest to exercise was less (p=0.047) for CHF+BR than CHF. These findings demonstrate that dietary NO3‾ supplementation reduces the elevated diaphragm BF during exercise in CHF rats thus providing additional support for this therapeutic intervention in CHF.

Effects of Type II diabetes on muscle microvascular oxygen pressures.

We tested the hypothesis that muscle microvascular O2 pressure (PmvO2; reflecting the O2 delivery (QO2) to O2 uptake (VO2) ratio) would be lowered in the spinotrapezius muscle of Goto-Kakizaki (GK) Type II diabetic rats (n=7) at rest and during twitch contractions when compared to control (CON; n=5) rats. At rest, PmvO2 was lower in GK versus CON rats (CON: 29+/-2; GK: 18+/-2Torr; P<0.05). At the onset of contractions, GK rats evidenced a faster change in PmvO2 than CON (i.e., time constant (tau); CON: 16+/-4; GK: 6+/-2s; P<0.05). In contrast to the monoexponential fall in PmvO2 to the steady-state level seen in CON, GK rats exhibited a biphasic PmvO2 response that included a blunted (or non-existent) PmvO2 decrease followed by recovery to a steady-state PmvO2 that was at, or slightly above, resting values. Compared with CON, this decreased PmvO2 across the transition to a higher metabolic rate in Type II diabetes would be expected to impair blood-muscle O2 exchange and contractile function.

Effect of chronic heart failure in older rats on respiratory muscle and hindlimb blood flow during submaximal exercise.

Submaximal exercise diaphragm blood flow (BF) is elevated in young chronic heart failure (CHF) rats, while it is unknown if this occurs in older animals. Respiratory and hindlimb muscle BFs (radiolabeled microspheres) were measured at rest and during submaximal exercise (20m/min, 5% grade) in older healthy (n=7) and CHF (n=6) Fischer 344X Brown Norway rats (27-29 mo old). Older CHF, compared to healthy, rats had greater (p<0.01) left ventricular end-diastolic pressure and right ventricle and lung weight (normalized to body weight). During submaximal exercise, respiratory and hindlimb muscle BFs increased (p<0.02) in both groups, while diaphragm BF was higher (CHF: 257±32; healthy: 121±9mL/min/100g, p<0.01) and hindlimb BF lower (CHF: 111±10; healthy: 133±12mL/min/100g, p=0.04) in older CHF compared to healthy rats. Submaximal exercise hindlimb BF was negatively related (r=-0.93; p=0.03) to diaphragm BF in older CHF rats. During submaximal exercise, diaphragm BF is elevated in older CHF compared to healthy rats in proportion to the compromised hindlimb BF.

Respiratory muscle blood flow during exercise: Effects of sex and ovarian cycle.

Sex and ovarian cycle have been speculated to modify respiratory muscle blood flow control during exercise, but the findings are inconclusive. We tested the hypotheses that females would have higher respiratory muscle blood flow and vascular conductance (VC) compared with males during exercise and that this difference would be accentuated in proestrus vs. ovariectomized (OVA) females. Mean arterial pressure (carotid artery catheter) and respiratory muscle blood flow (radiolabeled microspheres) were measured during moderate-intensity (24 m/min, 10% grade) exercise in male (n = 9), female (n = 9), and OVA female (n = 7) rats and near-maximal (60 m/min, 5% grade) exercise in male (n = 5) and female (n = 7) rats. At rest, diaphragm, intercostal, and transversus abdominis blood flow were not different (P = 0.33) among groups. During moderate-intensity exercise, diaphragm (M: 124 ± 16; F: 140 ± 14; OVA: 140 ± 20 ml·min-1·100 g-1), intercostal (M: 33 ± 5; F: 34 ± 5; OVA: 30 ± 5 ml·min-1·100 g-1), and transversus abdominis blood flow (M: 24 ± 4; F: 35 ± 7; OVA: 35 ± 9 ml·min-1·100 g-1) significantly increased in all groups compared with rest but were not different (P = 0.12) among groups. From rest to moderate-intensity exercise, diaphragm (P < 0.03) and transversus abdominis (P < 0.04) VC increased in all groups, whereas intercostal VC increased only for males and females (P = 0.01). No differences (P > 0.13) existed in VC among groups. During near-maximal exercise, diaphragm (M: 304 ± 62; F: 283 ± 17 ml·min-1·100 g-1), intercostal (M: 29 ± 8; F: 40 ± 6 ml·min-1·100 g-1), and transversus abdominis (M: 85 ± 14; F: 86 ± 9 ml·min-1·100 g-1) blood flow and VC were not different (P > 0.27) between males and females. These data demonstrate that respiratory muscle blood flow and vascular conductance at rest and during exercise are not affected by sex or ovarian cycle in rats.NEW & NOTEWORTHY It has been proposed that sex and ovarian cycle modulate respiratory muscle blood flow control during exercise. We demonstrate herein that neither sex nor ovarian cycle influences respiratory muscle blood flow or vascular conductance at rest or during exercise in rats.

Impact of aging on muscle blood flow in chronic heart failure.

Chronic heart failure (CHF) is manifested principally in the elderly population. Therefore, to understand the causes of exercise intolerance in CHF patients, it is imperative to resolve the effects of aging on muscle blood flow (BF) in CHF. To address this issue, we determined the muscle BF response to submaximal treadmill exercise (20 m/min, 5% grade) in young (Y(CHF): 6-8 mo, 412 +/- 11 g, n = 11) and old (O(CHF): 27-29 mo, 494 +/- 10 g, n = 8) Fischer 344 x Brown Norway rats with similar degrees of myocardial infarction-induced left ventricular (LV) dysfunction [resting LV end-diastolic pressure: Y(CHF) = 24 +/- 2, O(CHF) = 22 +/- 2 mmHg; derivative of LV pressure over time: Y(CHF) = 5,168 +/- 285; O(CHF) = 5,050 +/- 165 mmHg/s; lung weight normalized to body weight: Y(CHF) = 9.14 +/- 0.72; O(CHF) = 8.21 +/- 0.29 mg/g (all P > 0.05)]. The exercising heart rate response was blunted in O(CHF) compared with Y(CHF) rats (Y(CHF) = 454 +/- 8, O(CHF) = 395 +/- 9 beats/min; P < 0.05). BF (radiolabeled microspheres) to the total hindlimb musculature and to each of the 28 individual muscles examined was similar between Y(CHF) and O(CHF) rats under resting conditions. During exercise, BF to five of the hindlimb muscles that normally possess a majority of slow-twitch oxidative and fast-twitch oxidative glycolytic muscle fibers increased significantly less (-25 to -42%) for O(CHF) compared with Y(CHF) rats. In contrast, BF to 14 of the hindlimb muscles that normally possess a majority of fast-twitch glycolytic muscle fibers was increased (+22 to +337%) for O(CHF) vs. Y(CHF) rats, which contributed to a greater mass-specific total hindlimb BF response in O(CHF) rats (Y(CHF) = 78 +/- 5, O(CHF) = 100 +/- 11 ml.min(-1).100 g(-1); P < 0.05) and coincided with greater reductions in BF to the kidneys and splanchnic organs during exercise in O(CHF) vs. Y(CHF). In conclusion, there appears to be a profound age-related redistribution of BF from the highly oxidative to the highly glycolytic muscles of the hindlimb during exercise in O(CHF) compared with Y(CHF) rats. This phenomenon is qualitatively similar to that reported previously for healthy young and old rats.

Effects of eccentric exercise on microcirculation and microvascular oxygen pressures in rat spinotrapezius muscle.

A single bout of eccentric exercise results in muscle damage, but it is not known whether this is correlated with microcirculatory dysfunction. We tested the following hypotheses in the spinotrapezius muscle of rats either 1 (DH-1; n = 6) or 3 (DH-3; n = 6) days after a downhill run to exhaustion (90-120 min; -14 degrees grade): 1) in resting muscle, capillary hemodynamics would be impaired, and 2) at the onset of subsequent acute concentric contractions, the decrease of microvascular O(2) pressure (Pmv(o(2))), which reflects the dynamic balance between O(2) delivery and O(2) utilization, would be accelerated compared with control (Con, n = 6) rats. In contrast to Con muscles, intravital microscopy observations revealed the presence of sarcomere disruptions in DH-1 and DH-3 and increased capillary diameter in DH-3 (Con: 5.2 +/- 0.1; DH-1: 5.1 +/- 0.1; DH-3: 5.6 +/- 0.1 mum; both P < 0.05 vs. DH-3). At rest, there was a significant reduction in the percentage of capillaries that sustained continuous red blood cell (RBC) flux in both DH running groups (Con: 90.0 +/- 2.1; DH-1: 66.4 +/- 5.2; DH-3: 72.9 +/- 4.1%, both P < 0.05 vs. Con). Capillary tube hematocrit was elevated in DH-1 but reduced in DH-3 (Con: 22 +/- 2; DH-1: 28 +/- 1; DH-3: 16 +/- 1%; all P < 0.05). Although capillary RBC flux did not differ between groups (P > 0.05), RBC velocity was lower in DH-1 compared with Con (Con: 324 +/- 43; DH-1: 212 +/- 30; DH-3: 266 +/- 45 mum/s; P < 0.05 DH-1 vs. Con). Baseline Pmv(O(2)) before contractions was not different between groups (P > 0.05), but the time constant of the exponential fall to contracting Pmv(O(2)) values was accelerated in the DH running groups (Con: 14.7 +/- 1.4; DH-1: 8.9 +/- 1.4; DH-3: 8.7 +/- 1.4 s, both P < 0.05 vs. Con). These findings are consistent with the presence of substantial microvascular dysfunction after downhill eccentric running, which slows the exercise hyperemic response at the onset of contractions and reduces the Pmv(O(2)) available to drive blood-muscle O(2) delivery.

Skeletal muscle ouabain binding sites are reduced in rats with chronic heart failure.

Intrinsic skeletal muscle abnormalities decrease muscular endurance in chronic heart failure (CHF). In CHF patients, the number of skeletal muscle Na(+)-K(+) pumps that have a high affinity for ouabain (i.e., the concentration of [(3)H]ouabain binding sites) is reduced, and this reduction is correlated with peak oxygen uptake. The present investigation determined whether the concentration of skeletal muscle [(3)H]ouabain binding sites found during CHF is related to 1) severity of the disease state, 2) muscle fiber type composition, and/or 3) endurance capacity. Four muscles were chosen that represented slow-twitch oxidative (SO), fast-twitch oxidative glycolytic (FOG), fast-twitch glycolytic (FG), and mixed fiber types. Measurements were obtained 8-10 wk postsurgery in 23 myocardial infarcted (MI) and 18 sham-operated control (sham) rats. Eighteen rats had moderate left ventricular (LV) dysfunction [LV end-diastolic pressure (LVEDP) < 20 mmHg], and five had severe LV dysfunction (LVEDP > 20 mmHg). Rats with severe LV dysfunction had significant pulmonary congestion and were likely in a chronic state of compensated congestive failure as indicated by an approximately twofold increase in both lung and right ventricle weight. Run time to fatigue and maximal oxygen uptake (VO(2 max)) were significantly reduced ( downward arrow39 and downward arrow28%, respectively) in the rats with severe LV dysfunction and correlated with the magnitude of LV dysfunction as indicated by LVEDP (run time: r = 0.60, n = 21, P < 0.01 and VO(2 max): r = 0.93, n = 13, P < 0.01). In addition, run time to fatigue was significantly correlated with VO(2 max) (r = 0.87, n = 15, P < 0.01). The concentration of [(3)H]ouabain binding sites (B(max)) was significantly reduced (21-28%) in the three muscles comprised primarily of oxidative fibers [soleus: 259 +/- 14 vs. 188 +/- 17; plantaris: 295 +/- 17 vs. 229 +/- 18; red portion of gastrocnemius: 326 +/- 17 vs. 260 +/- 14 pmol/g wet tissue wt]. In addition, B(max) was significantly correlated with VO(2 max) (soleus: r = 0.54, n = 15, P < 0.05; plantaris: r = 0.59, n = 15, P < 0.05; red portion of gastrocnemius: r = 0.65, n = 15, P < 0.01). These results suggest that downregulation of Na(+)-K(+) pumps that possess a high affinity for ouabain in oxidative skeletal muscle may play an important role in the exercise intolerance that attends severe LV dysfunction in CHF.

Downhill running: a model of exercise hyperemia in the rat spinotrapezius muscle.

To utilize the rat spinotrapezius muscle as a model to investigate the microcirculatory consequences of exercise training, it is necessary to design an exercise protocol that recruits this muscle. There is evidence that the spinotrapezius is derecruited during standard treadmill exercise protocols performed on the uphill treadmill (i.e., 6 degrees incline). This investigation tested the hypothesis that downhill running would effectively recruit the spinotrapezius muscle as assessed by the presence of an exercise hyperemia response. We used radioactive 15-microm microspheres to determine blood flows in the spinotrapezius and selected hindlimb muscles of female Sprague-Dawley rats at rest and during downhill (i.e., -14 degrees incline; 331 +/- 5 g body wt, n = 7) and level (i.e., 0 degrees incline; 320 +/- 11 g body wt, n = 5) running at 30 m/min. Both level and downhill exercise increased blood flow to all hindlimb muscles (P < 0.01). However, in marked contrast to the absence of a hyperemic response to level running, blood flow to the spinotrapezius muscle increased from 26 +/- 6 ml.min(-1).100 g(-1) at rest to 69 +/- 8 ml.min(-1).100 g(-1) during downhill running (P < 0.01). These findings indicate that downhill running represents an exercise paradigm that recruits the spinotrapezius muscle and thereby constitutes a tenable physiological model for investigating the adaptations induced by exercise training (i.e., the mechanisms of altered microcirculatory control by transmission light microscopy).

Altered regional blood flow responses to submaximal exercise in older rats.

Maximal aerobic capacity and the ability to sustain submaximal exercise (Ex) declines with advancing age. Whether altered muscle blood flow (BF) plays a mechanistic role in these effects remains to be resolved. The present investigation determined the effects of aging on the hemodynamic and regional BF response to submaximal Ex in rats. Heart rate (HR), mean arterial pressure (MAP), and BF to different organs (kidneys, splanchnic organs, and 28 hindlimb muscles) were determined at rest and during submaximal treadmill Ex (20 m/min, 5% grade) with radiolabeled microspheres in young (Y; 6-8 mo old, 339 +/- 8 g, n = 9) and old (O; 27-29 mo old, 504 +/- 18 g, n = 7) Fischer 344 x Brown Norway rats. Results demonstrated that HR, MAP, and BF to the pancreas, small and large intestine, and total hindlimb musculature were similar between Y and O rats at rest. BF to the kidneys, spleen, and stomach were 33, 60, and 43% lower, respectively, in O compared with Y rats. BF to the total hindlimb musculature increased (P < 0.05) during Ex and was similar for both Y and O rats (Y: 16 +/- 3 to 124 +/- 7 vs. O: 20 +/- 3 to 137 +/- 12 ml.min-1.100 g-1). However, in O vs. Y rats, BF was reduced in 6 (highly oxidative) and elevated in 8 (highly glycolytic) of the 28 individual hindquarter muscles or muscle parts examined (P < 0.05). During Ex, BF to the spleen and stomach decreased (P < 0.05) from rest in Y rats, whereas BF decreased in the kidneys, pancreas, spleen, stomach, as well as the small and large intestines of O rats. In conclusion, these data demonstrate that, despite similar increases in total hindlimb BF in Y and O rats during submaximal Ex, there is a profound BF redistribution from highly oxidative to highly glycolytic muscles.