Joyner, Michael J.

The binding of heat shock protein 90 (HSP90) to endothelial nitric oxide (NO) synthase (eNOS) can enhance eNOS activation. Studies have shown that the HSP90-specific inhibitor geldanamycin (GA) can cause attenuation of NO-mediated processes. Twenty subjects participated in one of two protocols. In each protocol, one forearm of each subject was instrumented with two intradermal microdialysis probes for drug delivery. Laser Doppler flowmeters were used to measure cutaneous blood flow. Skin sites were either treated with the endothelial agonist acetylcholine or locally heated to 42°C, a maneuver that evokes NO-mediated dilation. Interventions were performed with and without GA. In the presence of GA, maximal cutaneous vascular conductance (CVC) to ACh was 20 ± 3% lower than with ACh alone ( P < 0.001). During local heating, maximal CVC in the presence of GA was 22 ± 6% lower than during heating alone ( P < 0.01). The results show that GA can attenuate NO-mediated dilation in human skin, suggesting a potential role for HSP90 in activation of eNOS in the microcirculation.

It is currently unclear whether aging alters the perfusion of active muscles during large-muscle dynamic exercise in humans. To study this issue, direct measurements of leg blood flow (femoral vein thermodilution) and systemic arterial pressure during submaximal cycle ergometry (70, 140, and 210 W) were compared between six younger (Y; 22–30 yr) and six older (O; 55–68 yr) chronically endurance-trained men. Whole body O2uptake, ventilation, and arterial and femoral venous samples for blood-gas, catecholamine, and lactate determinations were also obtained. Training duration (min/day), estimated leg muscle mass (dual-energy X-ray absorptiometry; Y, 21.5 ± 1.2 vs. O, 19.9 ± 0.9 kg), and blood hemoglobin concentration (Y, 14.9 ± 0.4 vs. O, 14.7 ± 0.2 g/dl) did not significantly differ ( P > 0.05) between groups. Leg blood flow, leg vascular conductance, and femoral venous O2 saturation were ∼20–30% lower in the older men at each work rate (all P < 0.05), despite similar levels of whole body O2 uptake. At 210 W, leg norepinephrine spillover rates and femoral venous lactate concentrations were more than twofold higher in the older men. Pulmonary ventilation was also higher in the older men at 140 (+24%) and 210 (+39%) W. These results indicate that leg blood flow and vascular conductance during cycle ergometer exercise are significantly lower in older endurance-trained men in comparison to their younger counterparts. The mechanisms responsible for this phenomenon and the extent to which they operate in other groups of older subjects deserve further attention.

Proctor, David N., Kenneth C. Beck, Peter H. Shen, Tamara J. Eickhoff, John R. Halliwill, and Michael J. Joyner. Influence of age and gender on cardiac output-V˙o2relationships during submaximal cycle ergometry. J. Appl. Physiol. 84(2): 599–605, 1998.—It is presently unclear how gender, aging, and physical activity status interact to determine the magnitude of the rise in cardiac output (Q˙c) during dynamic exercise. To clarify this issue, the present study examined theQ˙c-O2uptake (V˙o2) relationship during graded leg cycle ergometry in 30 chronically endurance-trained subjects from four groups ( n = 6–8/group): younger men (20–30 yr), older men (56–72 yr), younger women (24–31 yr), and older women (51–72 yr). Q˙c (acetylene rebreathing), stroke volume (Q˙c/heart rate), and whole bodyV˙o2were measured at rest and during submaximal exercise intensities (40, 70, and ∼90% of peakV˙o2). Baseline resting levels of Q˙c were 0.6–1.2 l/min less in the older groups. However, the slopes of theQ˙c-V˙o2relationship across submaximal levels of cycling were similar among all four groups (5.4–5.9 l/l). The absolute Q˙c associated with a given V˙o2(1.0–2.0 l/min) was also similar among groups. Resting and exercise stroke volumes (ml/beat) were lower in women than in men but did not differ among age groups. However, older men and women showed a reduced ability, relative to their younger counterparts, to maintain stroke volume at exercise intensities above 70% of peakV˙o2. This latter effect was most prominent in the oldest women. These findings suggest that neither age nor gender has a significant impact on theQ˙c-V˙o2relationships during submaximal cycle ergometry among chronically endurance-trained individuals.

This study evaluated the hypothesis that active muscle blood flow is lower during exercise at a given submaximal power output after aerobic conditioning as a result of unchanged cardiac output and blunted splanchnic vasoconstriction. Eight untrained subjects (4 men, 4 women, 23–31 yr) performed high-intensity aerobic training for 9–12 wk. Leg blood flow (femoral vein thermodilution), splanchnic blood flow (indocyanine green clearance), cardiac output (acetylene rebreathing), whole body O2 uptake (V˙o 2), and arterial-venous blood gases were measured before and after training at identical submaximal power outputs (70 and 140 W; upright 2-leg cycling). Training increased ( P < 0.05) peak V˙o 2(12–36%) but did not significantly change submaximalV˙o 2 or cardiac output. Leg blood flow during both submaximal power outputs averaged 18% lower after training ( P = 0.001; n = 7), but these reductions were not correlated with changes in splanchnic vasoconstriction. Submaximal leg V˙o 2 was also lower after training. These findings support the hypothesis that aerobic training reduces active muscle blood flow at a given submaximal power output. However, changes in leg and splanchnic blood flow resulting from high-intensity training may not be causally linked.

To explore the hypothesis that lower body muscle mass correlates with orthostatic tolerance, 18 healthy volunteers (age 18–48 yr; 10 men, 8 women) underwent a graded lower body negative pressure (LBNP) protocol consisting of six, 5-min stages of suction up to 60 mmHg in 10-mmHg increments. Forearm blood flow, heart rate, and blood pressure were measured, and forearm vascular resistance was calculated. Leg muscle mass was assessed by dual-energy X-ray absorptiometry. All subjects received standard intravenous hydration for at least 8 h before the study. Six men and four women completed all stages of LBNP. Four men and four women developed presyncopal symptoms, including marked bradycardia and/or hypotension, at LBNP levels of 30 mmHg ( n = 2; 1 man, 1 woman), 40 mmHg ( n = 2; 1 man, 1 woman), and 50 mmHg ( n = 4; 2 men, 2 women). The presyncopal subjects had leg muscle masses ranging from 19.5 to 25.2 kg in men and from 11.7 to 16.6 kg in women. In subjects who completed all stages of LBNP, leg muscle mass ranged from 17.5 to 24.1 kg in men and from 10.4 to 18.0 kg in women. Leg muscle mass did not differ between presyncopal subjects and those who completed the protocol. Furthermore, there were no differences in the hemodynamic responses to LBNP between subjects with low vs. high leg mass. These data suggest that leg muscle mass is not a critical determinant of LBNP tolerance in otherwise healthy men and women.

We conducted a series of studies to develop and test a rapid, noninvasive method to measure limb venous compliance in humans. First, we measured forearm volume (mercury-in-Silastic strain gauges) and antecubital intravenous pressure during inflation of a venous collecting cuff around the upper arm. Intravenous pressure fit the regression line, −0.3 ± 0.7 + 0.95 ± 0.02 ⋅ cuff pressure ( r = 0.99 ± 0.00), indicating cuff pressure is a good index of intravenous pressure. In subsequent studies, we measured forearm and calf venous compliance by inflating the venous collecting cuff to 60 mmHg for 4 min, then decreasing cuff pressure at 1 mmHg/s (over 1 min) to 0 mmHg, using cuff pressure as an estimate of venous pressure. This method produced pressure-volume curves fitting the quadratic regression (Δlimb volume) = β0 + β1 ⋅ (cuff pressure) + β2 ⋅ (cuff pressure)2, where Δ is change. Curves generated with this method were reproducible from day to day (coefficient of variation: 4.9%). In 11 subjects we measured venous compliance via this method under two conditions: with and without (in random order) superimposed sympathetic activation (ischemic handgrip exercise to fatigue followed by postexercise ischemia). Calf and forearm compliance did not differ between control and sympathetic activation ( P > 0.05); however, the data suggest that unstressed volume was reduced by the maneuver. These studies demonstrate that venous pressure-volume curves can be generated both rapidly and noninvasively with this technique. Furthermore, the results suggest that although whole-limb venous compliance is under negligible sympathetic control in humans, unstressed volume can be affected by the sympathetic nervous system.

Dyke, Christopher K., Niki M. Dietz, Robert L. Lennon, David O. Warner, and Michael J. Joyner. Forearm blood flow responses to handgripping after local neuromuscular blockade. J. Appl. Physiol. 84(2): 754–758, 1998.—To test the hypothesis that acetylcholine “spillover” from motor nerves contributes significantly to skeletal muscle vasodilation during exercise, we measured the forearm blood flow responses during attempted handgripping after local paralysis of the forearm with the neuromuscular-blocking drug pipecuronium. This compound blocks postsynaptic nicotinic receptors but has no impact on acetylcholine release from motor nerves. The drug was administered selectively to one forearm by using regional intravenous drug administration techniques in five subjects. Pipecuronium reduced maximum forearm grip strength from 40.0 ± 3.2 kg before treatment to 0.0 kg after treatment. By contrast, drug administration had no effect on maximum voluntary contraction in the untreated forearm (41.3 ± 3.3 vs. 41.4 ± 2.7 kg). During 2 min of attempted maximal contraction of the paralyzed forearm, the forearm blood flow increased from only 3.4 ± 0.8 to 4.8 ± 1.2 ml ⋅ 100 ml−1 ⋅ min−1( P < 0.05). Heart rate increased from 63 ± 3 to 73 ± 8 beats/min ( P > 0.05) during attempted contraction, and only three of five subjects showed obvious increases in heart rate. Mean arterial pressure increased significantly ( P < 0.05) from 102 ± 6 to 109 ± 9 mmHg during attempted contractions. When these increases in flow are considered in the context of the marked (10-fold or greater) increases in flow seen in contracting forearm skeletal muscle, it appears that acetylcholine spillover from motor nerves has, at most, a minimal impact on the hyperemic responses to contraction in humans.

Halliwill, John R., Lori A. Lawler, Tamara J. Eickhoff, Michael J. Joyner, and Sharon L. Mulvagh. Reflex responses to regional venous pooling during lower body negative pressure in humans. J. Appl. Physiol. 84(2): 454–458, 1998.—Lower body negative pressure is frequently used to simulate orthostasis. Prior data suggest that venous pooling in abdominal or pelvic regions may have major hemodynamic consequences. Therefore, we developed a simple paradigm for assessing regional contributions to venous pooling during lower body negative pressure. Sixteen healthy men and women underwent graded lower body negative pressure protocols to 60 mmHg while wearing medical antishock trousers to prevent venous pooling under three randomized conditions: 1) no trouser inflation (control), 2) only the trouser legs inflated, and 3) the trouser legs and abdominopelvic region inflated. Without trouser inflation, heart rate increased 28 ± 4 beats/min, mean arterial pressure fell −3 ± 2 mmHg, and forearm vascular resistance increased 51 ± 9 units at 60 mmHg lower body negative pressure. With inflation of either the trouser legs or the trouser legs and abdominopelvic region, heart rate and mean arterial pressure did not change during lower body negative pressure. By contrast, although the forearm vasoconstrictor response to lower body negative pressure was attenuated by inflation of the trouser legs (Δforearm vascular resistance 33 ± 10 units, P < 0.05 vs. control), attenuation was greater with the inflation of the trouser legs and abdominopelvic region (Δforearm vascular resistance 16 ± 5 units, P < 0.05 vs. control and trouser legs-only inflation). Thus the hemodynamic consequences of pooling in the abdominal and pelvic regions during lower body negative pressure appear to be less than in the legs in healthy individuals.

We tested the hypothesis that ACh or nitric oxide (NO) might be involved in the vasodilation that accompanies a single contraction of the forearm. Eight adults (3 women and 5 men) completed single 1-s-duration contractions of the forearm to raise and lower a weight equivalent to ∼20% maximal voluntary contraction through a distance of 5 cm. In a second protocol, each subject had a cuff, placed completely about the forearm, inflated to 120 mmHg for a 1-s period, then released as a simulation of the mechanical effect of muscle contraction. Three conditions were studied, always in this order: 1) control, with intra-arterial infusion of saline; 2) after muscarinic blockade with atropine; and 3) after NO synthase inhibition with N G-monomethyl-l-arginine (l-NMMA) plus atropine. Forearm blood flow (FBF), measured by combined pulsed and echo Doppler ultrasound, was reduced at rest with l-NMMA-atropine compared with the other two conditions. After the single contraction, there were no effects of atropine, butl-NMMA reduced the peak FBF and the total postcontraction hyperemia. After the single cuff inflation, atropine had no effects, whereasl-NMMA caused changes similar to those seen after contraction, reducing the peak FBF and the total hyperemia. The observation thatl-NMMA reduced FBF in response to both cuff inflation and a brief contraction indicates that NO from the vascular endothelium might modulate the basal level of vascular tone and the mechanical component of the hyperemia with exercise. It is unlikely that ACh and NO from the endothelium are involved in the dilator response to a single muscle contraction.

Joyner, Michael J., and Niki M. Dietz.Nitric oxide and vasodilation in human limbs. J. Appl. Physiol. 83(6): 1785–1796, 1997.—Both the skeletal muscle and skin of humans possess remarkable abilities to vasodilate. Marked vasodilation can be seen in these vascular beds in response to a variety of common physiological stimuli. These stimuli include reactive hyperemia (skin and muscle), exercise hyperemia (muscle), mental stress (muscle), and whole body heating (skin). The physiological mechanisms that cause vasodilation in response to these stimuli are poorly understood, and the substance(s) responsible for it remain unclear. In this context, recent attention has been focused on the possible contribution of nitric oxide (NO) to the regulation of hyperemic responses in human skin and skeletal muscle. The emerging picture is that NO is not an essential component of the dilator response seen during reactive hyperemia. However, it does appear that NO may play a modest role in exercise hyperemia. NO appears to play a major role in the skeletal muscle vasodilation seen in response to mental stress in humans. Preliminary evidence also indicates that NO is not essential for the normal dilator responses observed in the cutaneous circulation during body heating in humans, but this issue needs further study. There are a number of possible mechanisms that might mediate NO release in humans, and the role of these mechanisms in the various hyperemic responses is also poorly understood. The role of altered NO-mediated vasodilation in some disease states is also discussed. Whereas NO is a potent vasodilating substance, the actions of NO alone do not explain a variety of poorly understood vasodilator mechanisms in conscious humans. Much work remains for those interested in the role of NO in the regulation of blood flow to the skin and skeletal muscle of humans.