This study tested the effects of inhaled nitric oxide [NO; 20 parts per million (ppm)] during normoxic and hypoxic (fraction of inspired O2 = 14%) exercise on gas exchange in athletes with exercise-induced hypoxemia. Trained male cyclists (n = 7) performed two cycle tests to exhaustion to determine maximal O2 consumption (V˙o 2 max) and arterial oxyhemoglobin saturation (SaO2, Ohmeda Biox ear oximeter) under normoxic (V˙o 2 max = 4.88 ± 0.43 l/min and SaO2 = 90.2 ± 0.9, means ± SD) and hypoxic (V˙o 2 max = 4.24 ± 0.49 l/min and SaO2 = 75.5 ± 4.5) conditions. On a third occasion, subjects performed four 5-min cycle tests, each separated by 1 h at their respectiveV˙o 2 max, under randomly assigned conditions: normoxia (N), normoxia + NO (N/NO), hypoxia (H), and hypoxia + NO (H/NO). Gas exchange, heart rate, and metabolic parameters were determined during each condition. Arterial blood was drawn at rest and at each minute of the 5-min test. Arterial Po 2 (PaO2), arterial Pco 2, and SaO2 were determined, and the alveolar-arterial difference for Po 2 (A-aDo 2) was calculated. Measurements of PaO2 and SaO2 were significantly lower and A-aDo 2 was widened during exercise compared with rest for all conditions (P < 0.05). No significant differences were detected between N and N/NO or between H and H/NO for PaO2, SaO2 and A-aDo 2 (P > 0.05). We conclude that inhalation of 20 ppm NO during normoxic and hypoxic exercise has no effect on gas exchange in highly trained cyclists.
- exercise-induced hypoxemia
- pulmonary edema
- ventilation-perfusion inequality
- diffusion disequilibrium
some highly trained male endurance athletes experience decreases in arterial Po 2 (PaO2) and arterial oxyhemoglobin saturation (SaO2) and a widened alveolar-arterial difference for O2(A-aDo 2) during heavy exercise (5,13). The cause and significance of exercise-induced arterial hypoxemia (EIH) has been the topic of a considerable research effort; however, the mechanism(s) responsible remains controversial. Four potential factors have been identified: 1) venoarterial shunt, 2) relative alveolar hypoventilation, 3) ventilation-perfusion (V˙a/Q˙) inequality, and4) diffusion limitation. Venoarterial shunt has been identified as a minor contributor, and relative alveolar hypoventilation has a controversial role in the pathophysiology of EIH (6). V˙a/Q˙ relationships have been shown to worsen with exercise (10), and, during maximal exercise, ∼60% of the widened A-aDo 2 can be explained by V˙a/Q˙ mismatch (14). Pulmonary interstitial edema may explain bothV˙a/Q˙ inequality and diffusion limitations (11, 26). This would be expected to negatively influence gas exchange in the lung by lowering the compliance of the alveoli and by compressing small blood vessels, resulting in nonuniform airflow and blood flow distribution in the lungs (11). However, present techniques have failed to provide precise quantification of extravascular lung water or identification of the specific mechanisms responsible for the development of interstitial edema (6).
Inhaled nitric oxide (NO) is a potent and selective pulmonary vasodilator (8, 9). Inhalation of NO has been used effectively to treat several respiratory disorders in humans (1,15, 16) but has been shown to be detrimental in chronic obstructive pulmonary disease (1). The positive effects of NO are explained by a preferential distribution of inhaled NO to well-ventilated alveolar units, a reduction in the dispersion of ventilation distribution, lowering of pulmonary vascular pressures, and improved gas exchange. Inhalation of NO has also been utilized to exert a beneficial effect on arterial oxygenation in mountaineers with high-altitude pulmonary edema (27). Although the mechanisms of EIH remain debatable, there is some indirect evidence to support the development of transient interstitial edema as a mechanism for a widened A-aDo 2 (3, 23). We sought to test the hypothesis that inhaled NO would improve oxygenation during exercise in athletes with EIH by reducing A-aDo 2.
Highly trained male cyclists were recruited to participate in this study (n = 8). One subject was forced to withdraw due to difficulties with placement of the arterial catheter; therefore, all data are reported for n = 7. This investigation was divided into two parts. Subjects who met the inclusion criteria inpart 1 participated in part 2. Inclusion criteria were 1) normal spirometry, that is, no history of asthma or cardiorespiratory disease, 2) maximal O2consumption (V˙o 2 max) ≥60 ml · kg−1 · min−1 and/or 5 l/min, 3) development of EIH (maximal exercise SaO2≤ 91.0%), and 4) between the ages of 18 and 40 yr. Before testing was started, subjects received a verbal description of the experiment and completed a written, informed consent form. This study was approved by the Clinical Screening Committee for Research and Other Studies Involving Human Subjects of the University of British Columbia.
Preliminary screening: part 1.
Subjects reported to the Applied Physiology Laboratory in the Allan McGavin Sports Medicine Center (Univ. British Columbia), having refrained from exhaustive exercise for 24 h, abstained from ingestion of food or fluid for 4 h, except for water, and abstained from alcohol and caffeine for 12 h. Subjects' weight and height were measured and recorded. Both spirometry and pulmonary diffusion measurements for carbon monoxide (Dl CO) were collected using the same commercial apparatus (Collins DS/PLUS II, Braintree, MA). Dl CO was determined using the single-breath method. Before Dl CO and spirometry measurements were made, subjects sat and rested for 30 min to ensure a resting heart rate and pulmonary capillary blood volume.
V˙o 2 max was determined using an incremental test to exhaustion on an electronically braked cycle ergometer (Quinton Excalibur, Lode, Groningen, The Netherlands). Subjects pedaled at a self-chosen cadence at a progressing workload, which started at 0 W and increased 30 W/min. Subjects inspired through an air flowmeter (Vacumetrics model 17150, Ventura, CA) using a two-way nonrebreathing valve (Hans-Rudolph, model 2700B, Kansas City, KS). Expired air passed into a 5-liter mixing chamber from which gas samples were analyzed at a rate of 300 ml/min for O2 and CO2 (S-3A O2 analyzer and CD-3A CO2 analyzer, Applied Electrochemistry, Pittsburgh, PA). Expired gases and minute ventilation (V˙e) were recorded using a computerized system (Rayfield, Waitsfield, VT). Gas analyzers were calibrated with gases of known concentration, and the air flowmeter was calibrated by passing 100 liters of air through the system. Heart rate was recorded every 15 s using a portable heart rate monitor (Polar Vantage XL, Kempele, Finland). SaO2 was measured by a pulse oximeter (Ohmeda Biox 3740, Louisville, CO), with values averaged and recorded every 5 s using a personal computer. Before placement of the oximeter sensor to the pinna of the ear, a topical vasodilator cream (Finalgon, Boehringer/Ingeheim, Burlington, ON) was applied to increase local perfusion. Attainment of V˙o 2 max was considered when at least three of the following four were observed:1) a plateau in O2 consumption (V˙o 2) with increasing workload,2) respiratory exchange ratio (RER) > 1.15,3) attainment of 90% of age predicted maximal heart rate, and/or 4) volitional fatigue. During part 1, cycle ergometry subjects inspired compressed air [fraction of inspired O2(Fi O2) = 20.93%]. The air was delivered from a large cylinder through a closed container of water for humidification and then into a large meteorological balloon, which acted as a reservoir for inspired air. Those who met the inclusion criteria returned on a separate day at least 72 h later to perform another maximal cycle ergometry test under hypoxic conditions (Fi O2 = 14.00%). This Fi O2 has previously been used to accentuate decreases in SaO2 in exercising trained men (17). Expired gases, heart rate, and SaO2 were determined during the hypoxic exercise session as described for the normoxic conditions.
Inhaled NO: part 2.
After completion of both V˙o 2 max tests (normoxic and hypoxic), subjects who met the inclusion criteria returned on a separate day at least 72 h later. A time line describing the experimental protocol is shown in Fig.1. Subjects were randomly assigned and blinded to each of the four following conditions: 1) normoxia (N), 2) normoxia + NO (N/NO), 3) hypoxia (H), and 4) hypoxia + NO (H/NO). Participants performed a 10- to 15-min cycling warm-up at a self-selected workload and then sat quietly on the cycle ergometer for 5 min, at which point resting data were obtained. Cycling intensity was then manually increased over 1 min to 100% of their respective maximum normoxic or hypoxic workload as determined in part 1. Subjects cycled at this intensity for 5 min, and cardiorespiratory variables were recorded at each minute in the same fashion as forpart 1. Arterial blood samples were drawn at rest and at each min of the 5-min test. After each test condition, subjects cycled easily (30–50 W) for 10 min and then rested for 50 min before commencing the next test condition. A physician was in attendance at all times and was responsible for the safety of the subjects during the study.
During N, subjects rested for 5 min and cycled while breathing normoxic gas. Condition N/NO consisted of normoxic gas with 20 parts per million (ppm) NO delivered to the inspiratory tubing. During condition H, subjects rested for 5 min and cycled while inhaling hypoxic gas (Fi O2 = 14.00%); condition H/NO consisted of the same hypoxic gas with 20 ppm NO. During all conditions, the inspired air was delivered from a large cylinder through water for humidification and then into a large meteorological balloon, which acted as a reservoir before being inspired by the subject. NO was delivered at the distal end of the tubing while inspired concentrations of O2, NO, and nitrogen dioxide were monitored continuously during each test condition 5 cm from the subject's mouth using a commercial apparatus (PulmoNOx II, Pulmonox, Tofield, AB). The NO-delivery system was calibrated before each experiment as per the manufacturer's specifications. The concentration of NO used in the present study (20 ppm) has previously been shown to improve V˙a/Q˙ distributions, PaO2, and pulmonary vascular resistance in pigs (24), reverse hypoxic pulmonary vasoconstriction (HPV), and redistribute blood flow to better ventilated areas of the lung in sheep (21).
Arterial blood sampling.
A 20-gauge arterial catheter was inserted in the radial artery of the nondominant hand by percutaneous cannulation using 1% local anesthesia (lidocaine) and sterile technique and was then secured to the skin. Adequate collateral circulation via the ulnar artery (Allen's test) was estimated before the cannula was inserted. A minimum volume extension tube, connected in series with two three-way stopcocks arranged at right angles, was flushed with a saline-heparin solution. A rapid response (<0.01 s) thermistor (18T, Physitemp Instruments, Clifton, NJ) used to measure peak arterial blood temperature was inserted through a Touhy-Borsch heparin lock (Abbott Hospitals, North Chicago, IL). Catheter patency was maintained with a continuous heparin infusion (1 ml 1:1,000 units in 500 ml normal saline at 3 ml/h). At the onset of sampling, 12 ml of blood was withdrawn, and the final 3 ml was collected in preheparinized plastic syringes. The remaining 9 ml were then slowly reinfused. Samples were withdrawn at rest and at 1-min intervals for the duration of each test (4 conditions × 6 samples per condition = 24 samples/subject). Blood samples were placed on ice until analyzed for H+concentration, Po 2, Pco 2, base excess, and HCO3 −(CIBA-Corning 278 blood-gas system, CIBA-Corning Diagnostics, Medfield, MA). PaO2 was corrected for temperature and H+ concentration. Temperature increased 0.9 ± 0.1°C (mean ± SD) from rest to 5 min of exercise across all trials. SaO2 levels were calculated based on corrected PaO2. The alveolar gas equation was used to calculated alveolar Po 2 and A-aDo 2 (20).
Mean values and measures of variability were determined for descriptive, anthropometric, and lung function variables obtained during preliminary screening. Maximal cycle ergometry data frompart 1 were compared using t-tests for dependent samples (normoxia vs. hypoxia). Experimental data were analyzed using a four (condition) by six (time) two-way factorial ANOVA with repeated measures on both factors. When sphericity was not assumed, Greenhouse-Geisser P values were utilized. When significantF ratios were observed, Scheffé's test was applied post hoc to determine where the differences occurred. The level of significance was set at P < 0.05. Statistical power calculations were performed a priori to estimate an appropriate minimum sample size of five. A sample size of seven was utilized to ensure sufficient statistical power (1 − β = 0.8).
Physical and maximal exercise data.
Descriptive data and resting pulmonary function data are presented in Table 1. Lung parameters were within normal values predicted for men of similar age, height, and weight except for Dl CO, which was significantly elevated. Data from normoxic and hypoxic maximal cycle ergometry tests are presented in Table 2. Significant differences were observed between normoxic and hypoxic conditions for V˙o 2 max, RER, maximal heart rate, and power output, whereas no significant differences were detected for V˙e. From rest to maximal exercise, mean values for SaO2 dropped significantly under both normoxic (97.7 to 90.2) and hypoxic (97.0 to 75.5) conditions.
Metabolic and power output during 5-min cycling.
Arterial blood variables during 5-min cycling.
All data are reported in Tables 5and 6 and Figs.2 and 3. Across all time points, there were no significant differences for PaO2 between N and N/NO or between H and H/NO (see Fig. 2). Both hypoxic conditions were significantly lower than both normoxic conditions at all measurement periods. PaO2 values were significantly lower at 1, 2, 3, 4, and 5 min of exercise compared with rest for all inspired gas conditions. Similar results were observed for SaO2, except that values at 1 and 2 min were not significantly different from rest under conditions of N and N/NO (see Fig. 3). A-aDo 2was significantly different at all time periods compared with rest for all inspired gas mixtures, and significant differences were detected between N/NO and H at rest and during all exercise measurements. Arterial Pco 2(PaCO2) was not significantly different between gas conditions but was lower compared with rest throughout all exercise for H and H/NO and at minutes 3, 4, and 5 for both N and N/NO. No statistically significant differences were detected between N vs. N/NO or between H vs. H/NO for pH, HCO3 −, or base excess (Tables 5 and 6).
The present study is the first to systematically examine the effects of inhaled NO during normoxic and hypoxic exercise in athletes with EIH. Unique to this investigation was the observation that inhalation of 20 ppm NO during normoxic and hypoxic high-intensity, short-duration exercise did not significantly affect gas exchange,V˙o 2, or cycling power in highly trained athletes with EIH.
The mechanism(s) of EIH remains controversial. However,V˙a/Q˙ inequality and diffusion limitations may be causative (6, 11, 30). The mechanisms explaining the increases in V˙a/Q˙ inequality and diffusion limitation are unknown but may be related to the development of transient pulmonary edema (6). Some authors have suggested that, in elite athletes, the integrity of the pulmonary blood-gas barrier is altered at extreme levels of exercise (29). However, identification of pulmonary edema and pulmonary stress failure has been difficult to achieve in exercising humans and remains controversial. Although the specific mechanism forV˙a/Q˙ inequality and diffusion limitations is debatable, the end result is a widened A-aDo 2. Inhaled NO, a selective pulmonary vasodilator (8, 9), is used in the treatment of diseases characterized by pulmonary hypertension and hypoxemia (1, 15, 16). The rationale is based on the fact that NO, given by inhalation, only dilates those pulmonary vessels that are well ventilated. As a result, pulmonary gas exchange is improved. We hypothesized that inhaled NO would improved gas exchange in athletes with EIH.
Inhaled NO during normoxia.
The present investigation demonstrated no differences in gas exchange,V˙o 2, power, or heart rate between N and N/NO at rest or during exercise. These findings do not support our original hypothesis but do agree with observations in resting sheep (17) and the fact that PaO2 was not altered in normal individuals who inhaled NO at rest (8). In addition, our findings are consistent with data that reported no effects of inhaled NO on Q˙ or Dl CO in humans (2) or PaO2 andV˙a/Q˙ in dogs (12) under normoxic conditions.
Studies that have sought to examine the effects of inhaled NO during exercise in humans have been few. To date, only one group (7) has investigated the effects of NO inhalation on pulmonary gas exchange during exercise in highly trained athletes. In agreement with the present study, they observed no differences for ventilatory and performance parameters. However, they found that inhalation of NO caused PaO2 to decrease at rest and during exercise compared with breathing room air. This is in contrast to the present study and the resting data of Frostell et al. (8). It is important to note that arterial blood-gas measurements were corrected for temperature in the present study, whereas those of Durand et al. (7) were not. This would likely overestimate the drop in PaO2 and SaO2 they observed during exercise and impact on the interpretation of their results.
Although pulmonary pressures were not measured in this study, we can speculate on two possible explanations for our normoxic results:1) pulmonary pressures were not altered or 2) pulmonary pressures were reduced with no effect on gas exchange. We believe it is more probable that the first scenario occurred, although we cannot exclude the second possibility. The pulmonary capillary bed in these athletes may have been already maximally dilated, and inhalation of a vasodilator would have no further effect. Therefore, no effect on pulmonary pressure would be expected nor would any alteration in gas exchange occur. Our results are therefore consistent with the hypothesis of Dempsey (4), who postulated that pulmonary capillary blood volume reaches its maximal morphological limit and further dilation is not anatomically possible. However, in light of our lack of hemodynamic measures, we emphasize the speculative nature of this explanation of our negative findings.
Inhaled NO during hypoxia.
Unique to this investigation was the delivery of NO to individuals with EIH during hypoxic exercise. As expected, during H and H/NO, PaO2 and SaO2 were significantly lower than N and N/NO at all time points (Figs. 2 and 3). The drop in oxygenation during hypoxic exercise was consistent with that observed in previous EIH studies (22). Similar to the normoxic trials, inhaled NO did not alter gas exchange,V˙o 2, or cycling power during hypoxia. Pison et al. (21) showed that addition of 20 ppm NO to a hypoxic (Fi O2 = 12%) gas mixture returned pulmonary gas-exchange measures to baseline values in mechanically ventilated sheep. These are in contrast to our results in which no statistical difference was observed for gas-exchange variables between H and H/NO. This was an unexpected result; we expected that HPV would be induced by using a hypoxic inspiratory gas mixture at rest and during exercise.
We hypothesized that inhaled NO would have reversed the transient HPV and improved pulmonary gas exchange, as demonstrated in healthy humans breathing hypoxic gas (21). Why did we observe no effect of NO on gas exchange during the resting hypoxic condition? Potentially, the HPV at rest in the present study was of a small enough magnitude and duration to have had a minimal effect on gas exchange. However, this seems unlikely given that the time course for HPV has a distinct initial rapid constriction that reaches a peak within minutes and shows profound increases in pulmonary vascular resistance within the time frame used in the present study (28). An alternate explanation is that the timing of resting samples impacted on arterial blood-gas values. Resting samples were taken after a warm-up period that was preceded by a 5-min resting period. Possibly, the warm-up influenced the V˙a/Q˙ relationship and may explain the lack of effect of NO on gas exchange. Furthermore, subjects may have been slightly hyperventilating at the time of the resting sample (PaCO2 of 36–38 Torr), which could have affected the influence of NO.
The lack of effect of NO during hypoxic exercise is consistent with the observations of Koizumi et al. (17). Inhalation of NO during exercise in sheep had no effect on PaO2. In addition, inhaled NO did not change the time course or magnitude of changes in pulmonary pressures in the exercising sheep.
Limitations of the study.
A debatable point is how much of the inhaled NO actually reached the lower airways? Although it was not possible to measure the amount of inhaled NO that reaches the alveoli, we assume that subjects in the present study did indeed inhale 20 ppm NO. We are confident that NO was delivered to the respiratory system, as it was measured 5 cm from the point of inspiration. The experimental approach and NO concentration employed in our study were similar to those employed by others (7). It is possible that the concentration of NO used in this study was not sufficient to induce vasodilation; however, this seems unlikely given that similar concentrations have been used previously to show significant alterations in gas exchange and pulmonary pressures (19, 21, 25). Our choice of 20 ppm was based on the above-mentioned studies and other clinical investigations.
It is possible that we failed to detect a statistical difference between conditions due to our small sample size and insufficient statistical power. However, this seems unlikely. Post hoc statistical power (1 − β) calculations were performed utilizing a computer statistical program for gas-exchange variables across normoxic and hypoxic conditions. Power values ranged from 0.72 to 0.84, indicating sufficient protection against type II errors.
Determination of pulmonary artery pressure and cardiac output in the present study, together with V˙a/Q˙ measures, would have allowed for more definitive conclusions on the effect of inhaled NO during exercise. Because the use of NO was hypothesized to act as a selective pulmonary vasodilator, the lack of the measurement of pulmonary artery and wedge pressures represents a major limitation of this study.
In summary, we have demonstrated that inhalation of 20 ppm NO during normoxic and hypoxic high-intensity, short-duration cycle exercise did not significantly affect gas exchange in athletes with EIH. Cardiorespiratory variables and cycling power output were also unaffected by NO inhalation during normoxia and hypoxia. We conclude that inhalation of 20 ppm NO during normoxic and hypoxic exercise has no effect on gas exchange in highly trained cyclists.
Address for reprint requests and other correspondence: D. C. McKenzie, Allan McGavin Sports Medicine Center, 3055 Wesbrook Mall, Vancouver, BC, Canada V6T 1Z3 (E-mail:).
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- Copyright © 2001 the American Physiological Society