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1EA 2363 Laboratoire Réponses Cellulaires et Fonctionelles à l'Hypoxie, Université Paris 13, Bobigny, France; and 2Department of Molecular and Integrative Physiology, University of Kansas Medical Center, Kansas City, Kansas
Submitted 7 June 2005 ; accepted in final form 19 September 2005
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ABSTRACT |
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 TOP ABSTRACT MATERIALS AND METHODSRESULTSDISCUSSIONGRANTSREFERENCES |
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pulmonary artery pressure; endurance training; right ventricular weight
We previously observed that rats living and training in moderate hypoxia [inspired PO2 (PIO2) of 
110 Torr for 10 wk] show a lower Pap than sedentary rats living at the same PIO2 (15). Because the measurements were obtained in normoxia and exercise training did not influence the mild polycythemic response, it appears that the lower Pap was due to attenuation of vascular remodeling by exercise training. Whether this was accompanied by effects of exercise training on hypoxia-induced pulmonary vascular smooth muscle contraction could not be determined, since the experimental design of this study did not include measurements of the Pap response to acute changes in PIO2.
In the present study, we investigated the effects of exercise training on the hypoxic pulmonary vascular response of conscious rats to acute and long-term changes in PIO2. This was done at rest and during maximal treadmill exercise. We reasoned that effects of exercise training on the pulmonary responses to short-term changes in PIO2 would largely reflect the magnitude of active vascular smooth muscle contraction; on the other hand, responses of chronically hypoxic rats would reflect modifications by exercise training of the contributions of polycythemia and pulmonary vascular remodeling to the hypertension.
The results presented here are part of a larger study on the effects of exercise training on the mechanisms of acclimatization to hypoxia. The data pertaining to systemic hemodynamics and systemic O2 transport and utilization during exercise have been published elsewhere (7).
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MATERIALS AND METHODS |
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Exercise training protocol.
Male Sprague-Dawley rats (200–225 g) were randomly assigned to sedentary or exercise-trained groups. Exercise training was carried out in normoxic conditions in an open-air, eight-lane treadmill, 5 days/wk. Work rate was started at 30 m/min for 5 min on a 10° incline and was increased 5 min/day until the rats ran for a total of 1 h. After 3 wk of training at full intensity, the animals of both groups were randomly assigned to normoxic and hypoxic subgroups, which resulted in a total of four subgroups of eight rats each: normoxic sedentary (NS), hypoxic sedentary (HS), normoxic trained (NT), and hypoxic trained (HT). The hypoxic groups, both sedentary and trained, were placed for 10 days in a hypobaric chamber where air was circulated at a barometric pressure of 380 Torr, which resulted in a PIO2 of 70 Torr. The chamber was opened five times per week for 90 min, during which both sedentary and trained rats were removed from the chamber and exposed to the normoxic environment. HT and NT rats continued to train, under normoxic conditions, at the same work rate as before. HS rats were exposed to normoxia for the same time as HT rats but did not exercise. This exercise training protocol resulted in an increase in maximal O2 uptake (
O2 max) of 20% and a two- to threefold increase in gastrocnemius muscle citrate synthase activity (7).
Maximal exercise test. At the end of the training protocol, animals were anesthetized with pentobarbital sodium (30 mg/kg ip). A polyethylene catheter (PE-50) was placed in the aortic arch via the left carotid artery, and a PE-10 catheter was advanced into the pulmonary artery via the right jugular vein with the aid of a J-shaped introducer. Adequate placement of the catheters was established by the pressure waveform and verified at autopsy. The catheters were tunnelled subcutaneously, exteriorized at the back of the neck, and flame sealed. The animals were returned to the chamber after recovery from anesthesia.
On the following day, after measurement of rectal temperature, the animals were placed on a treadmill enclosed in an airtight Lucite chamber adapted for the determination of O2 uptake (O2) and CO2 production (CO2) using the open-circuit method. The catheters were connected through sampling ports located on the top of the box enclosing the treadmill to pressure transducers. Thirty minutes after being placed on the treadmill, arterial and mixed venous blood samples were obtained via stopcocks, the blood was replaced with homologous fresh blood from the same group, and the treadmill was set at a speed of 10 m/min. This speed was maintained for 2–3 min, after which the treadmill was set at an angle of 10° and the speed increased by 4 m/min every 90–120 s until O2 max was reached. O2 max was defined as the O2 after which an increase in work rate was not associated with a further increase (±5%) in O2. Arterial and mixed venous blood samples were obtained during the last 45–60 s of exercise, while O2 and CO2 showed steady values. The box enclosing the treadmill was opened, and the rectal temperature was determined within 30 s of termination of exercise.
Each animal ran twice, once in normoxic conditions (PIO2 of 145 Torr) and once in hypoxia (PIO2 of 70 Torr). Both runs were carried out in the same day, with the order of hypoxic and normoxic runs being alternated in successive days. An interval of at least 4 h was allowed between runs in each rat. After the first run, the blood withdrawn in the exercise samples was replaced with fresh homologous blood, and 0.5 ml/100 g of a solution of 0.15 M NaHCO3 was administered intravenously to correct the metabolic acidosis of maximal exercise.
Gas exchange and O2 transport determinations.
The box enclosing the treadmill was airtight except for the inflow and outflow ports, which are independent of one another. PIO2 was adjusted to the desired level by mixing O2 and N2. Flow of the gas mixture entering the treadmill box was maintained constant at 20 liters ATPS/min by using a precision gas flow mixer. Inflowing and outflowing O2 and CO2 concentrations were measured continuously and simultaneously. The output of the O2 and CO2 meters was fed into a computer, and O2 and CO2 were calculated using standard gas-exchange equations (7).
Arterial and mixed venous blood samples were analyzed for pH, PO2, and PCO2 using appropriate electrodes at 38°C, and for Hb concentration and O2 saturation of Hb, and corrected for the rectal temperature by using temperature correction factors for rat blood (12, 13). Systemic arterial pressure and Pap were recorded continuously, with mean pressures obtained by electronic integration. Arterial and mixed venous O2 content were calculated from Hb concentration, PO2, and oxyhemoglobin saturation. Cardiac output (
; in ml·min–1·kg–1) was calculated as the ratio of O2 to arteriovenous O2 content. Alveolar PO2 was calculated using the alveolar gas equation assuming arterial PCO2 (PaCO2) to be equal to alveolar PCO2. Alveolar ventilation was calculated from the ratio of CO2 to PaCO2, and normalized to O2 (A/O2). Effective lung diffusing capacity for O2 (DL
) was calculated from O2, and alveolar, arterial, and mixed venous PO2 values, as described by Piiper and Scheid (24), assuming that all of the difference between arterial and alveolar PO2 is due to diffusion limitation. DL was calculated only in hypoxia because of the uncertainty introduced by the nonlinear shape of the oxyhemoglobin dissociation curve at high PO2 values.
The day after the maximal exercise bout, animals were euthanized by an overdose of barbiturate (60 mg/kg iv pentobarbital sodium), the heart was removed, and the left ventricle plus septum and the right ventricle were weighed separately.
Statistical analysis. Data are presented as means ± SE. One-way ANOVA, followed by a Bonferroni posttest for multiple comparisons, was used to determine the statistical significance between mean values. A P value of <0.05 was considered to indicate a significant difference.
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RESULTS |
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30 min of exposure to the nonresident PO2. The exercise data were obtained 10–15 min later.
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A/O2 and arterial PO2 (PaO2) and reductions in PaCO2 in all groups. This was observed both in hypoxia and in normoxia. An increase in PaO2 above resting values is a known feature of hypoxic exercise in the rat (7, 12, 13) and contrasts with the usual decrease observed in humans. The mechanisms responsible for the differences between species have been discussed before (13). The rats living in hypoxia showed the expected increase in blood Hb concentration, reflecting the polycythemia of prolonged hypoxia. Of interest is that PaO2 during hypoxia was higher in the trained rats than in their sedentary counterparts (Tables 1 and 2): NT and HT showed higher PaO2 values than NS and HS, respectively. This occurred both at rest (Table 1) and during maximal exercise (Table 2). The higher PaO2 cannot be explained by higher alveolar ventilation of the trained rats during hypoxia: there were no significant differences in A/O2 between trained rats and their sedentary counterparts, with the exception of NT vs. NS at rest, in which A/O2 was actually lower in NT (Table 1). Alveolar-arterial PO2 difference [(A-a)PO2] was lower in the trained rats (except in HS vs. HT at rest, in which there was no significant difference between groups; Table 1); DL was higher in all cases in the trained rats. The combined data suggest that the higher PaO2 is the result of a higher efficacy of pulmonary gas exchange in the trained animals.
Effects of exercise training on pulmonary hypertension.
The values of Pap and of the ratio of Pap to (Pap/) are presented in Fig. 1. As expected, exposure of NS to hypoxia resulted in a significant increase in resting Pap and Pap/, reflecting the pulmonary vasoconstriction of acute hypoxia. Resting Pap and Pap/ of NT animals also increased during acute exposure to hypoxia, but the increase was significantly smaller than that seen in the sedentary animals. Maximal exercise had a relatively small additional effect on Pap in both NS and NT, owing to the marked decrease in Pap/ associated with exercise. A decrease in vascular resistance with exercise is a well-known feature of the pulmonary circulation. Although the intergroup differences were smaller than those seen at rest, both Pap and Pap/ of NT were significantly lower than the corresponding values of NS during hypoxic exercise, suggesting that pulmonary vascular resistance during acute hypoxic exercise is lower in NT. It is possible that this is in part due to the effect of the higher in NT than NS (Table 2)
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during hypoxia at rest, without significant differences observed between groups. During hypoxic exercise, Pap increased to a higher value in HT than in HS (Fig. 1A), possibly as a result of the higher in the former (Table 2). Nevertheless, Pap/ in hypoxic exercise was lower in HT than in HS (Fig. 1B). In contrast to the normoxic groups, however, an acute change from resident PIO2, in this case from hypoxia to normoxia, produced only modest changes in resting Pap or Pap/ in either HS or HT (Fig. 1). Although Pap/ of both hypoxic groups decreased significantly with exercise with respect to the corresponding resting values, it remained significantly higher than in the corresponding normoxic groups. As it happened at rest, transition from hypoxia to normoxia produced only minor changes in exercise Pap and Pap/ in both HT and HS rats (Fig. 1) Effects of exercise training on hypoxia-induced right ventricular hypertrophy. Table 3 shows the effects of exercise training and hypoxia on body weight and right and left ventricular weight (LV). Exercise training did not significantly change body weight; on the other hand, both hypoxic groups showed lower body weights than their normoxic counterparts. Exercise training in normoxia resulted in higher right ventricular weight (RV) and LV; both ventricles increased in weight in the same proportion, with no effect of exercise training on RV/(LV + S) (11), where S is septum weight. Hypoxia produced the expected increase in RV in the sedentary rats; this was accompanied by a lower LV, so RV/(LV + S) of HS was significantly higher than that of NS. Interestingly, RV of HT was not different from that of NT and actually lower than that of HS, so it appears as if the right ventricular hypertrophy of prolonged hypoxia is attenuated by exercise training. LV + S was smaller in both hypoxic groups than in the corresponding normoxic counterparts; however, as in normoxia, LV + S was higher in the trained rats. This resulted in the ratio RV/(LV + S) being substantially lower in HT than in HS.
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DISCUSSION |
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TOPABSTRACTMATERIALS AND METHODSRESULTSDISCUSSIONGRANTSREFERENCES |
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of acute hypoxia; 3) did not substantially influence the pulmonary hypertension of prolonged hypoxia; and 4) attenuated the right ventricular hypertrophy of prolonged hypoxia.
Experimental design.
The design of these experiments allowed us to determine the effects of exercise training on the pulmonary vascular response to short-term (30 min) and long-term (10 days) exposure to relatively severe hypoxia (PIO2 of 70 Torr, equivalent to an altitude of 5,500 m). There is abundant information in the patterns of response of rats to this level of hypoxia (5–7, 12, 13, 21) that produces clear-cut effects on the pulmonary vasculature. Exposure to hypoxia was limited to 10 days; this time was selected because previous studies in this model have shown that O2 transport variables and systemic and pulmonary hemodynamics have reached a steady state by this time (5).
Training was initiated before exposure to hypoxia; this was done to ensure that the possible effects of exercise training, which may influence hypoxia-induced pulmonary vascular responses, were already expressed at the onset of hypoxia. To prevent detraining effects, the rats continued to train while living in hypoxia. Because hypoxia reduces exercise capacity, the hypoxic animals trained in normoxia to maintain the same absolute training intensity of the normoxic animals. To control for the daily interruption of hypoxia for 90 min during training, the sedentary hypoxic rats were removed from the chamber and exposed to normoxia for the same time as the trained rats. The training regime employed here produced substantial increases in O2 max, , tissue O2 extraction, and muscle oxidative capacity (Ref. 7, and Table 1).
The pulmonary vascular response to short-term hypoxia is largely due to contraction of vascular smooth muscle, whereas the effects of prolonged hypoxia also involve polycythemia and vascular remodeling. Comparison of the effects of acute changes in resident PIO2 in rats maintained in hypoxia vs. those living in normoxia provided useful information of the relative contribution of active vasoconstriction, polycythemia, and vascular remodeling to the effects of exercise training on pulmonary hypertension.
Measurement of left atrial pressure to calculate pulmonary vascular resistance in conscious, closed-chest rats is impractical. Accordingly, data on Pap/ were used as an indication of changes in pulmonary vascular resistance. In all cases, the changes in Pap/ produced by the various interventions paralleled the changes in Pap, indicating that the latter are not the result of changes in . Maximal exercise resulted in modest increases in Pap and marked decreases in Pap/, reflecting the well know decrease in pulmonary vascular resistance associated with increases in . These factors suggest that, in the present experiments, changes in Pap/ are an adequate reflection of changes in pulmonary vascular resistance.
Effects of exercise training on pulmonary gas exchange.
The trained rats showed higher PaO2 during hypoxia than their sedentary counterparts. This effect cannot be explained by higher ventilatory responses to hypoxia in the trained animals (Tables 1 and 2), suggesting that efficacy of pulmonary gas exchange is increased by exercise training. (A-a)PO2 and the apparent pulmonary DL, were used as global indexes of efficacy of pulmonary gas exchange. DL was used in addition to (A-a)PO2 since it takes into account O2 and mixed venous PO2, changes that could influence (A-a)PO2 in the absence of changes in pulmonary function. Both indexes are influenced by pulmonary diffusing capacity, A/O2 mismatch, and venoarterial shunt (24). From the present data, it is not possible to determine which of these factors is responsible for the effects of exercise training on pulmonary gas exchange. Hypoxia in humans and rats widens A/O2 distribution; this is exaggerated by exercise (20, 29). Although, in theory, exercise training could improve efficacy of gas exchange by reducing the spread of A/O2 values, this may not be a very effective mechanism to increase PaO2 in hypoxia, since the contribution of A/O2 heterogeneity to the (A-a)PO2 is sharply reduced with decreasing PIO2 (29).
Increased pulmonary diffusing capacity could also contribute to the higher PaO2 of the trained animals. Exercise training does not modify the structural determinants of diffusing capacity, i.e., the lung does not grow in response to exercise training (28). However, O2 diffusion in the lung may be influenced by nonstructural features such as capillary blood volume, pulmonary capillary Hb concentration, , and transcapillary fluid exchange. Exercise training could theoretically act through one or more of these factors.
Although the underlying mechanism remains unidentified, the attenuation of the decrease in PaO2 in hypoxia brought about by exercise training could be a valuable adaptive strategy. The major compensatory mechanism to acute hypoxia is the increase in ventilation, which serves to attenuate the drop in PaO2. Although this is a highly useful mechanism, its energy costs can be substantial, particularly when energy demands are elevated such as in exercise. An increased efficacy of pulmonary gas exchange would permit further attenuation of hypoxemia for a given level of alveolar ventilation.
Effects of exercise training on acute hypoxic pulmonary hypertension.
The increases in Pap and in Pap/ in response to acute hypoxia were significantly smaller in the NT rats than in their sedentary counterparts. Because the main factor responsible for the acute pulmonary hypertension is active vasoconstriction, the most likely explanation is that exercise training attenuates the hypoxia-induced contraction of pulmonary vascular smooth muscle. The mechanism underlying this effect is not apparent from the present data; however, in view of the improvement in pulmonary gas exchange produced by exercise training, a possible explanation may be that the attenuated pressor response of the NT rats is due to a less severe hypoxic stimulus in these animals.
Other factors, in addition to the PaO2 levels, could tend to attenuate the increase in pulmonary vascular resistance during hypoxia produced by exercise training. A decrease in the pulmonary vasoconstrictive response to endothelin (18) has been demonstrated in trained rats. Pulmonary arterial rings obtained from exercise-trained pigs show increased endothelial-dependent vasodilation compared with rings from untrained controls (19). Exercise training-induced increases in endothelial NO synthase have been reported in systemic vascular beds (16). Exercise-induced elevated NO levels in exhaled air of trained individuals suggest that the increased NO availability of exercise training observed in the systemic circulation could extend to the pulmonary circulation (4).
The attenuation of the acute pulmonary pressor response to hypoxia by exercise training should provide a beneficial effect by lowering the right ventricular afterload in trained subjects. The smaller increase in cardiac work would tend to maintain myocardial tissue PO2 during conditions such as exercise in acute hypoxia in which myocardial O2 demands are increased in the presence of limited O2 supply.
Effects of exercise training on pulmonary hypertension of prolonged hypoxia.
Although exercise training moderated the acute pressor response to hypoxia in the rats living in normoxia, it had much smaller effects on the pulmonary hypertension of prolonged hypoxia. Increased vascular smooth muscle tone, vascular remodeling, and polycythemia are thought to be the major factors responsible for the pulmonary hypertension of chronic hypoxia (1, 10, 17). The relatively small effect of acute return to normoxia on Pap and Pap/ of chronically hypoxic rats suggests that hypoxia-induced increased vascular smooth muscle tone played a minor role in the hypertension of prolonged hypoxia. It is also clear that exercise training did not influence polycythemia, since Hb concentrations of HS and HT were essentially identical (Tables 1 and 2). Accordingly, it would appear that exercise training did not affect the remaining contributing factor of chronic hypoxic pulmonary hypertension either, namely pulmonary vascular remodeling. This contrasts with observations from our own laboratories (15), which demonstrated an attenuation of pulmonary hypertension in rats exposed for 10 wk at PIO2 of 110 Torr, a level of hypoxia more moderate than the one employed in the present study. Because the measurements were carried out under normoxic conditions, the effect cannot be explained on the basis of an attenuation of hypoxia-induced vascular smooth muscle contraction by exercise training (15). Furthermore, both trained and sedentary rats living in moderate hypoxia showed similar polycythemic responses. Accordingly, the effect of exercise training on the pulmonary hypertension of moderate hypoxia appears to be due to attenuation of pulmonary vascular remodeling. The present study shows that a similar effect of exercise training does not occur under conditions of more severe hypoxia. The reason for this apparent discrepancy may be due to the different severity of hypoxia as well as duration of the training regime between both studies: in the present study, hypoxia was more severe and training shorter than in our earlier study. It is clear that the possible effect of exercise training on remodeling and vasoactive properties of the pulmonary circulation during prolonged hypoxia should be the subject of further research.
Effects of exercise training on hypoxia-induced right ventricular hypertrophy. As expected (5, 6, 21, 25), acclimatization to hypoxia induced right ventricular hypertrophy accompanying the sustained pulmonary hypertension. However, RV was significantly lower in HT than in HS, despite comparable right ventricular afterload. The mechanism for the smaller increase in RV is not clear. A possible explanation is that the attenuation of the pulmonary hypertension in the early stages of hypoxia in the trained rats may have delayed the development of right ventricular hypertrophy. Pap of sedentary rats increases immediately on exposure to hypoxia (5), whereas this increase is attenuated in trained rats (Fig. 1). Although Pap eventually increases to similar levels in sedentary and trained animals (Fig. 1), the time during which the right ventricular afterload is increased may be shorter in the trained rats, and this may influence the development of right ventricular hypertrophy. Alternatively, factors in addition to mechanical loads could modulate the development of right ventricular hypertrophy. Steudel et al. (26) have shown that inhalation of NO prevented right ventricular hypertrophy in mice acclimatized to hypoxia despite pulmonary hypertension and polycythemia. If exercise training increases NO availability in the pulmonary circulation, right ventricular hypertrophy could be moderated.
In summary, the present study shows that exercise training improves effectiveness of pulmonary gas exchange and attenuates the decrease in PaO2 secondary to acute hypoxic exposure. This is accompanied by smaller increases in Pap and Pap/ on acute exposure to hypoxia, indicating that exercise training attenuates the contraction of pulmonary vascular smooth muscle that follows hypoxia. It is possible that this attenuated vasoconstriction is due to the higher PaO2 observed in trained rats. Despite the smaller initial increase in Pap, exercise training does not substantially influence the development of chronic hypoxic pulmonary hypertension, indicating that the polycythemia and vascular remodeling that follows prolonged exposure to PIO2 of 70 Torr are not affected by exercise training. Despite comparable right ventricular afterload levels at the end of the protocol, right ventricular hypertrophy was moderated in the HT rats. This could be the result of a delay in the development of pulmonary hypertension. Although the underlying mechanisms are not known, improvement of pulmonary gas exchange and reduction in the initial increase in Pap represent useful responses to acute hypoxia.
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GRANTS |
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FOOTNOTES |
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The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
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REFERENCES |
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TOPABSTRACTMATERIALS AND METHODSRESULTSDISCUSSIONGRANTSREFERENCES |
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O2 max more than living and training in normoxia. J Appl Physiol 90: 2057–2062, 2001.This article has been cited by other articles:
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  C. Yilmaz, D. M. Dane, and C. C. W. Hsia Assessing recruitment of lung diffusing capacity in exercising guinea pigs with a rebreathing technique J Appl Physiol, July 1, 2008; 105(1): 316 - 321. [Abstract] [Full Text] [PDF]
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