INTRODUCTION

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HUMANS EXPOSED TO MICROGRAVITY undergo deconditioning of the cardiovascular system. Moreover, the severity of problems associated with reentry into the gravitational field increases with the duration of exposure to microgravity (51). A similar cardiovascular deconditioning is also observed with prolonged 6° head-down-tilt bed rest (HDTB; Ref. 51). Exposure to either microgravity or HDTB causes an immediate cephalad shift of body fluids and a subsequent sustained reduction in plasma volume and blood volume (51). Fluid redistribution to the upper half of the body results in a characteristic facial puffiness. This fluid redistribution is proposed to be the triggering event for subsequent adaptations of the cardiovascular system to microgravity or prolonged HDTB (51). On resumption of an upright posture after either HDTB or reentry into the gravitational field, subjects experience a number of adverse cardiovascular consequences, including resting tachycardia, decreased exercise capacity, and orthostatic intolerance characterized by either presyncopal symptoms or syncope. It has been hypothesized that these adverse effects may be related to hypovolemia, an attenuated reflex activation of sympathetic nervous system activity, reduced total peripheral resistance, and changes in peripheral vascular reactivity (2, 3, 15, 33, 38).

Hindlimb unweighting (HU) in rodents is used to simulate microgravity-induced cardiovascular deconditioning in humans because it elicits many of the cardiovascular alterations seen in humans. These include an initial cephalad fluid shift (20, 31, 52) and subsequent hypovolemia (35, 45), tachycardia (32), and reduced exercise capacity (14, 37).

A number of in vitro studies have investigated the effect of HU on vascular function. Delp and co-workers (13) found that the contraction of isolated rat aorta to various vasoconstrictor agents was reduced after 14-day HU. Using arterial tissues from 20-day HU-treated rats, Purdy and co-workers (38) also reported a significant decrease in the contraction of the carotid and femoral artery and the abdominal aorta to either norepinephrine or 68 mM K⁺. Sangha and co-workers (41) showed that this vascular hyporesponsiveness to norepinephrine in the carotid and femoral arteries, but not the abdominal aortas, was in part due to upregulation of nitric oxide-dependent vasodilator mechanisms. In the carotid artery these vasodilator mechanisms were eliminated by removal of the endothelium (42).

The effect of endothelium removal described above prompted our hypothesis that the hyporesponsiveness to norepinephrine in the HU carotid artery was associated with changes in endothelial nitric oxide function (42). Thus, in the present study, we wanted to explore whether changes in the endothelial cyclooxygenase (COX)-dependent pathway might also occur in response to HU. Endothelial cells lining the lumen of all blood vessels modulate vascular tone through the release of vasoactive agents such as nitric oxide, eicosanoids, prostacyclin, and thromboxane (50). Nitric oxide and prostacyclin are potent vasorelaxants and inhibitors of platelet aggregation that counterbalance the vasoconstrictor and platelet-aggregating properties of thromboxane (7). Nitric oxide is formed from L-arginine by nitric oxide synthase, and the constitutive form of this enzyme predominates in endothelial cells (28). Eicosanoid synthesis from arachidonic acid is regulated by the expression and activity of COX, an enzyme also reported to be expressed in endothelial cells (4). Prostacyclin, a downstream product of COX, has been shown to serve as an endothelium-derived relaxing factor in certain blood vessels (34). Alternatively, other endothelium-derived COX metabolites are suggested to oppose vasodilation. For example, Cirino et al. (4) proposed that the release of thromboxane A₂ from the endothelial cells may attenuate the profound vasodilatation observed during shock states. Iwama and co-workers (23) showed, in the aortas from spontaneously hypertensive rats, the presence of a COX-dependent vasoconstrictor metabolite that attenuated endothelium-dependent relaxations. The aim of the present study was to determine the role of endothelium-derived, COX-dependent metabolites of arachidonic acid in the reduced contractile response to norepinephrine in carotid artery rings from 20-day HU-treated rats.

	METHODS

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Animal procedures were approved by the Institutional Animal Care and Use Committee of the University of California, Irvine. Male Wistar rats weighing 250-300 g were obtained from Simonsen Laboratories (Gilroy, CA) and housed in a temperature-controlled room (22°C) with a 12:12-h light-dark cycle. Water and rat chow were provided ad libitum. Animals were randomly assigned to control or HU groups. HU was achieved by using a tail harness to elevate the hindlimbs of the animals above the floor of the cage by a modification (38) of the method of Thomason et al. (48). Briefly, the tail was cleaned and a coat of tincture of benzoin was applied. It was then air dried until tacky. Adhesive strips (Fas-Trac of California, Van Nuys, CA) that were the width of the tail were looped through a swivel harness and pressed along the sides of the tail to form a tubular casing around the tail. Thereafter, the tail was wrapped with Elastoplast bandage (Beiersdorf, Norwalk, CT) followed by a thin layer of plaster cast material (Sammons Preston, Bolingbrook, IL). The rat was suspended by the swivel harness from a hook at the top center of the suspension cage, allowing free 360° rotation. The height of the hook was adjusted such that the front limbs were in contact with the floor and the hindlimbs, when extended, were elevated ~0.5 cm above the floor, tilting the body of the rat to an angle of 35° with the floor of the cage. The animals were HU for 20 days, after which the HU and the paired control rats were used in the following in vitro experiments.

Carotid artery ring preparation. HU and control rats were euthanized by exposure to 100% CO₂ for 60 s to induce deep anesthesia (19). The chest was opened and the heart removed. The carotid arteries were collected and placed in warm oxygenated Krebs bicarbonate solution. The tissues were cleaned under a dissecting microscope to remove extraneous fatty and connective tissue and then cut into 3-mm rings. In protocols examining responses in the absence of vascular endothelium, the endothelium was removed by using two strands of 32-gauge stainless steel wire tightly twisted to form a single wire bundle of 4-cm length. This bundle fitted easily into the lumen of the carotid artery rings and was used to remove the endothelium by gently rolling these tissues back and forth on moistened filter paper. Success of the endothelium removal in both control and HU vessels was confirmed by the loss of acetylcholine-mediated relaxation of phenylephrine-precontracted vessels.

In vitro isometric contraction experiments. Carotid artery rings were mounted in tissue baths for the measurement of isometric contraction, and they were stretched to previously determined optimal resting forces (38). In another study from the present laboratory (26), it was found that HU treatment reduced the contractile response to norepinephrine but not to serotonin. This implies that the effect of HU treatment is not mediated by possible vascular smooth muscle remodeling or vessel atrophy, and it validates our use of previously determined optimal resting forces: control carotid artery, 1.5 g; HU carotid artery, 1 g (38). Vessel rings were equilibrated for 30 min in 37°C,95% O₂-5% CO₂ gassed Krebs bicarbonate solution of the following composition (in mM): 119.2 NaCl, 4.9 KCl, 1.3 CaCl₂, 1.2 MgSO₄, 25 NaHCO₃, 11.1 glucose, 0.114 ascorbic acid, and 0.03 tetrasodium ethylenediaminetetraacetate. Subsequently, the artery rings were contracted with Krebs solution containing 100 mM K⁺ prepared by equimolar replacement of Na⁺. When the tissues had reached steady-state contraction, they were washed twice with normal Krebs solution and allowed to relax to resting levels. Contractions with 100 mM K⁺ were repeated 20-30 min later. Preliminary experiments revealed that all tissues yielded uniform magnitudes of contraction to the second and all subsequent exposures to 100 mM K⁺. Thirty to forty-five minutes after the tissues recovered from the second exposure to 100 mM K⁺, norepinephrine concentration-response curves (CRCs) were obtained by the cumulative addition of norepinephrine in 0.5-log increments. Indomethacin, ibuprofen, the thromboxane A₂ prostanoid-receptor antagonist, SQ-29548, or vehicle were added to the tissue baths 30 min before the norepinephrine CRC. The following agents were also added to the bathing medium 30 min before norepinephrine CRCs were obtained: 30 µM cocaine and 30 µM deoxycorticosterone acetate to block neuronal (17) and extraneuronal (25, 27) catecholamine uptake, respectively, and 1 µM propranolol to block -adrenoceptors (1).

View larger version (23K): [in this window] [in a new window]   Fig. 1.   Concentration-response curves for the contractile effects of norepinephrine in control (Ctl) and hindlimb unweighted (HU) carotid artery rings with and without 10 µM indomethacin. Values are means ± SE; n = 5 animals per treatment group. *HU different from control, P < 0.05. + HU different from HU in the presence of indomethacin, P < 0.05. ^ Control different from control in the presence of indomethacin, P < 0.05.

View larger version (23K): [in this window] [in a new window]   Fig. 2.   Concentration-response curves for the contractile effects of norepinephrine in Ctl and HU carotid artery rings with and without 100 µM ibuprofen. Values are means ± SE; n = 5 animals per treatment group. *HU different from control, P < 0.05. + HU different from HU in the presence of ibuprofen. P < 0.05.

View larger version (26K): [in this window] [in a new window]   Fig. 3.   Concentration-response curves for the contractile effects of norepinephrine in Ctl and HU carotid artery rings with and without 0.1 µM SQ-29548. Values are means ± SE; n = 5 animals per treatment group. *HU different from control, P < 0.05. + HU different from HU in the presence of SQ-29548, P < 0.05.

View larger version (25K): [in this window] [in a new window]   Fig. 4.   Concentration-response curves for the contractile effects of norepinephrine in endothelium-denuded Ctl and HU carotid artery rings with and without 10 µM indomethacin (Indo). Values are means ± SE; n = 8 animals per treatment group.

View larger version (25K): [in this window] [in a new window]   Fig. 5.   Concentration-response curves for the contractile effects of norepinephrine in Ctl and HU carotid artery rings in the presence of 10 µM Indo. HU vessels are with and without the nitric oxide inhibitor NG-nitro-L-arginine methyl ester (L-NAME; 10 µM). Values are means ± SE; n = 5 animals per treatment group. *HU different from control, P < 0.05. + HU different from HU in the presence of L-NAME. P < 0.05.

View larger version (23K): [in this window] [in a new window]   Fig. 6.   Concentration-response curves for the contractile effects of norepinephrine in Ctl and HU carotid artery (Cart) rings with and without 10 µM L-NAME. Values are mean ± SE; n = 5 animals per treatment group. *HU different from HU+L-NAME, P < 0.05; + HU different from control, P < 0.05.

	RESULTS

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An earlier study from the present laboratory indicated that HU treatment resulted in hyporesponsiveness to norepinephrine in arterial tissues (38). Removal of the endothelium restored the observed hyporesponsiveness to norepinephrine toward control in the carotid artery rings from HU-treated rats (41). The purpose of the present study was to assess the possible contribution of endothelium-derived, COX-dependent arachidonic acid metabolites to the vascular hyporesponsiveness to norepinephrine in the carotid artery rings from HU-treated rats. The first approach was to expose the carotid artery rings from HU and control rats to 10 µM indomethacin, a COX inhibitor, before norepinephrine CRCs were obtained. Addition of either indomethacin or ibuprofen (see below) had no effect on resting force. As observed in our laboratory's earlier studies (38), HU treatment resulted in a significantly reduced response to norepinephrine in the carotid artery (Fig. 1). Indomethacin did not affect the response to norepinephrine in control arteries, except at the highest norepinephrine concentration used (300 µM). In this case, indomethacin increased the contraction. In contrast, indomethacin caused a further depression in the contractile response to norepinephrine in the HU-treated carotid artery rings. In six of the eight experiments shown in Fig. 1, HU tissues contracted weakly to norepinephrine in the presence of indomethacin. However, norepinephrine elicited no response in two experiments, resulting in a nonsignificant effect of norepinephrine in these tissues. To confirm these results, the effects of a structurally different COX inhibitor, 100 µM ibuprofen, were assessed. Ibuprofen had no effect on control carotid artery rings but significantly depressed the contractile response to norepinephrine in the carotid artery rings from HU-treated rats (Fig. 2).

Taken together, these results indicated that HU treatment upregulated the activity of a COX-dependent vasoconstrictor substance. PGH₂ and thromboxane A₂ are known vasoconstrictor agents that exert their effects through the thromboxane A₂ prostanoid receptor (6). Thus experiments were carried out to assess the consequences of selective thromboxane A₂ prostanoid-receptor blockade using SQ-29548 (36). As shown in Fig. 3, 0.1 µM SQ-29548 had no effect on control carotid artery rings but further depressed the contraction to norepinephrine in HU vessel rings. The similar effects of COX and thromboxane A₂ prostanoid-receptor inhibition suggested that the contractile response to norepinephrine in HU carotid artery rings was mediated, in part, by a product of the COX pathway that acted on the thromboxane A₂ prostanoid receptor.

To determine whether the COX-dependent vasoconstrictor metabolite was derived from the endothelium, norepinephrine CRCs were obtained in endothelium-denuded carotid artery rings in the presence and absence of indomethacin. As shown in Fig. 4, indomethacin had no effect on the norepinephrine CRC in HU carotid artery. This result demonstrates that the COX-dependent vasoconstrictor metabolite was derived from the endothelium. In addition, indomethacin had no effect on the norepinephrine CRC in control carotid artery.

In a previous study, it was observed that endothelium removal increased the maximal contraction of HU carotid artery to norepinephrine (42). As a consequence, that contraction moved closer to the maximal norepinephrine-induced contraction in the control carotid artery. Inspection of Fig. 4 revealed a similar result. The maximal response to norepinephrine in HU was closer to that in control carotid artery in endothelium-denuded (34%; Fig. 4), compared with endothelium-intact (46%; Fig. 1), vessels. However, the CRCs in the control and HU denuded carotid arteries were clearly not superimposed. Statistical analysis found no significant differences between the two CRCs. However, power calculations revealed that, for contractions to 1-10 µM norepinephrine, there was insufficient statistical power to establish a lack of significant difference between these two CRCs.

The effect of the nitric oxide synthase inhibition with 10 µM L-NAME was assessed in endothelium-intact control and HU carotid artery rings pretreated with indomethacin to eliminate the contribution of a vasoconstrictor prostaglandin. Addition of L-NAME had no effect on resting force. As shown in Fig. 5, HU treatment markedly depressed the contraction to norepinephrine. However, the contractile response of the HU carotid artery rings in the presence of L-NAME increased and the norepinephrine CRC became superimposed on that of the control tissues in the absence of L-NAME. L-NAME also markedly increased the contractile response to norepinephrine in the control tissues. Thus the HU-induced reduction in contractile response was retained in the presence of indomethacin plus L-NAME.

Additional experiments were carried out to determine the effect of L-NAME alone, i.e., in the absence of indomethacin. As shown in Fig. 6, the norepinephrine CRC in the HU carotid artery was depressed, compared with control. However, in the presence of L-NAME, the HU CRC was elevated and became superimposed on the control CRC in the absence of L-NAME. L-NAME had no significant effect in control carotid arteries but tended to cause an increase. L-NAME treatment also increased the variability of vessel contraction, as indicated by larger standard errors of the mean.

	DISCUSSION

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HU treatment causes vascular hyporesponsiveness to norepinephrine in several arteries of the rat (10, 11, 13, 38), including the abdominal aorta and the femoral and carotid arteries. Results from a recent study (42) provided evidence that nitric oxide derived from the endothelium contributed to the HU-induced hyporesponsiveness of the carotid artery. Also, the present finding (Fig. 4) that endothelium removal reduced the HU effect is consistent with such a role for nitric oxide. The present study was undertaken to explore whether COX-dependent metabolites of arachidonic acid might also contribute to the effect of HU treatment.

HU treatment depressed the contractile response of the carotid artery to norepinephrine, as reported previously (38). Pretreatment with either of two different COX inhibitors caused a further depression of the contractile response to norepinephrine in HU carotid artery rings but had no effect in control artery rings. Importantly, the thromboxane A₂ prostanoid-receptor antagonist, SQ-29548, mimicked these effects of COX inhibition. Taken together, these results suggested that a COX-dependent metabolite of arachidonic acid acted on thromboxane A₂ prostanoid receptors to mediate a large component of the contractile response to norepinephrine in HU carotid artery. Finally, endothelium removal eliminated the blocking effect of indomethacin in the HU carotid artery (see Fig. 4). This demonstrated that the COX-dependent metabolite was derived from the endothelium.

Endothelium removal was shown previously (42) and in the present study (Fig. 4) to increase the contraction of HU carotid artery to norepinephrine. In our laboratory's earlier study (42), this was the first evidence to suggest that HU treatment might upregulate endothelial vasodilator function. This suggestion was strengthened by observations that HU treatment caused 1) a 10-fold increase in the sensitivity of carotid artery to acetylcholine-induced vasodilation (42) and 2) a significant increase in endothelial nitric oxide synthase expression in this vessel (41).

In endothelium-denuded vessels, the norepinephrine CRC in HU carotid artery approached, but did not become superimposed on, the CRC in the control vessel (Fig. 4). If HU-induced hyporesponsiveness to norepinephrine were due to an endothelium-dependent mechanism exclusively, endothelium removal should have caused the CRCs in control and HU carotid arteries to become superimposed. Because they were not superimposed, we suggest that HU, in addition to its endothelial effect, impaired vascular smooth muscle function, thereby contributing to the depression of contractility in the carotid artery. Power calculations revealed that the eight experiments represented in Fig. 4 of the present study were insufficient to detect a significant HU effect in denuded carotid artery. Additional research is required to address this issue.

Experiments were carried out with indomethacin to block COX and with L-NAME to block nitric oxide synthase. It is important to note that these agents were added to the bathing medium after the artery rings were equilibrated at their optimum resting forces. Neither indomethacin nor L-NAME had any effect on resting force. This has several implications. First, we suggest that there was little or no basal synthesis and release of a vasoconstrictor prostaglandin. If there had been, this prostaglandin would have produced baseline tone and the addition of indomethacin would have caused the vessel to relax. This argument also supports the view that the indomethacin-sensitive component of the contraction to norepinephrine was not due to the spontaneous release of the vasoconstrictor prostaglandin. Rather, it appears that the production of the prostaglandin was dependent on the activation of -adrenoceptors. It is possible that, in some presently unknown way, HU treatment coupled vasoconstrictor prostaglandin production as a second messenger to the -adrenoceptor. Alternatively, HU treatment could simply have increased the expression of the synthesis pathway for the prostaglandin. In this case, synthesis and release could have been triggered by cellular shape changes associated with the norepinephrine-induced contraction. Further research is required to differentiate between these possibilities.

Second, the lack of effect of L-NAME on resting tone has two possible interpretations. It may indicate that there is no spontaneous release of nitric oxide. This interpretation presupposes that the carotid artery must possess myogenic tone. If true, that tone could be suppressed by spontaneously released nitric oxide. In that case, L-NAME would have blocked nitric oxide synthesis and release and unmasked the myogenic tone. If this scenario were true, the lack of tone development would suggest that there is no spontaneous release of nitric oxide. Alternatively, the carotid artery may not possess myogenic tone. In this case, blockade of nitric oxide release by L-NAME would not have an effect under any circumstances. These two possibilities cannot be distinguished by the present results.

Throughout most of the norepinephrine CRC, indomethacin had no effect on the contraction of control carotid arteries. This implies that activation of -adrenoceptors in control arteries was not associated with prostaglandin. In contrast, indomethacin (and also ibuprofen and SQ-29548) further suppressed an already decreased contraction to norepinephrine in HU carotid arteries. This effect of indomethacin on HU carotid artery contractions was eliminated in endothelium-denuded vessels. These observations support the major conclusion of this study that HU treatment induces the expression of the synthetic machinery for a vasoconstrictor prostaglandin within the endothelium. Moreover, synthesis and release of such a prostaglandin contributed to the contraction of HU carotid artery to norepinephrine.

These results and those of our laboratory's earlier studies suggest that HU treatment upregulated both a vasodilator, nitric oxide (42), and a vasoconstrictor, prostaglandin (present study), in the endothelium of carotid artery. To explore this further, it was hypothesized that blockade of both nitric oxide synthase and COX would have an effect equivalent to endothelium removal. Namely, the norepinephrine-induced contraction in HU arteries would more closely approximate that in the control arteries in the presence of L-NAME and indomethacin. However, the results obtained were far more complex, at least with respect to control carotid arteries, than this hypothesis suggests. First, it should be noted that L-NAME increased the contraction of HU arteries to norepinephrine, both in the presence (Fig. 5) and absence (Fig. 6) of indomethacin. This is consistent with an enhanced role for nitric oxide in HU carotid artery. What was unexpected was the dramatic increase in the contraction of control carotid arteries to norepinephrine in the presence of both L-NAME and indomethacin (Fig. 5). We offer the following speculation. In Fig. 1, it can be seen that indomethacin had no effect on the contraction of control carotid artery except at the highest concentration of norepinephrine, 300 µM. The fact that indomethacin enhanced the contraction to this concentration of norepinephrine suggests the involvement of a vasodilator prostaglandin. In Fig. 6, it can be seen that the nitric oxide synthase inhibitor, L-NAME, caused a modest, nonsignificant elevation of the norepinephrine CRC in control carotid artery. Thus it is possible that control arteries possess two potential vasodilator pathways, a prostaglandin and nitric oxide. Blocking either of these pathways has little effect because they are both relatively weak and/or each can compensate for the other. However, blocking both pathways has a synergistic effect that manifests as a marked enhancement of contraction.

There are precedents in other animal models for an interaction between nitric oxide and a COX-dependent vasoconstrictor substance, both released from the endothelium. Koga et al. (29) and Iwama et al. (23) assessed the maximal acetylcholine-induced relaxation of precontracted aorta rings from spontaneously hypertensive and normotensive rats. They found that this relaxation was attenuated by both age and hypertension. This relaxation was blocked by L-NAME, indicating that it was mediated by nitric oxide. Importantly, the attenuation of this relaxation was reversed by either COX inhibition (29) or thromboxane A₂ prostanoid-receptor blockade (23). Thus both studies (23, 29) provided evidence that a COX-dependent vasoconstrictor substance had been released from the endothelium and that this substance accounted for the age and hypertension-induced attenuation of relaxation to acetylcholine.

The mechanisms responsible for the HU-mediated induction of an endothelially derived, COX-dependent vasoconstrictor substance in carotid artery are unknown. However, there is evidence to support the hypothesis that the upregulated nitric oxide itself may have stimulated the production of the COX metabolite. A direct interaction of the nitric oxide and COX pathways has been reported for various cell types and tissues (16, 40, 47). Davidge et al. (7) observed an activation of COX by nitric oxide in cultured microvascular endothelial cells. This interaction could have occurred via intermediary pathways as follows. Under conditions of enhanced oxidative stress, nitric oxide reacts with superoxide anions to form peroxynitrite (22). This could lead to an increase in lipid peroxidation that would enhance COX activity (21). Using Western immunoblots of the thoracic aorta, Davidge et al. (8) reported an increased COX expression in older rats and suggested that a natural age-related increase in endogenous lipid peroxidation may have increased COX expression in these rats. Whether the HU-induced increase in endothelially derived nitric oxide in carotid artery was responsible for induction of the COX-dependent metabolite observed in the present study is unknown. Future experiments will be required to elucidate this issue. However, endothelial nitric oxide does not contribute to the hyporesponsiveness to norepinephrine in either abdominal aorta (13, 42) or femoral artery from HU rats (42). Moreover, in contrast to the carotid artery, COX inhibition has no effect on the contraction of these vessels to norepinephrine (26; Sangha and Purdy, unpublished observations). This lends further support to the possibility that a causal relationship exists between elevated endothelial nitric oxide activity and the induction of an endothelium-derived vasoconstrictor prostaglandin.

The mechanisms underlying the HU-induced upregulation of endothelial nitric oxide function (41, 42) in carotid artery are unknown. However, endothelial nitric oxide vasodilator activity is regulated by changes in blood flow and associated shear stress (44). We speculate that HU increased shear stress in the carotid artery by causing a cephalad fluid shift (20, 31, 52). In turn, this could have increased endothelial nitric oxide function.

An increase in endothelial nitric oxide function is not a universal feature of blood vessels from the HU rat. Jasperse et al. (24) found that endothelial nitric oxide synthase protein expression was reduced in rat soleus muscle feed arteries after 2 wk of HU. In addition, these authors and Delp et al. (12) found that the vasodilation of soleus feed arteries to acetylcholine was reduced. These observations may also reflect HU-induced changes in shear stress. Delp et al. found that shear stress was reduced in soleus feed arteries after 2 wk of HU.

Geary et al. (18) obtained functional evidence, in rat cerebral artery, that HU reduced nitric oxide activity. Because this was responsible, in part, for an increase in myogenic tone in this vessel, these authors suggested that the reduced nitric oxide activity was an appropriate adaptation. They suggested that elevated cerebrovascular myogenic tone and consequent vasoconstriction could act to protect the brain from overperfusion. In view of this argument, the hyporesponsiveness of the carotid artery seems inappropriate with respect to proper maintenance of brain blood flow. We suggest that the main effect of carotid artery hyporesponsiveness is to contribute to orthostatic hypotension, a known consequence of both simulated (30) and real (51) microgravity.

In summary, the present study confirms our laboratory's earlier observation (42) that HU treatment increases endothelium-dependent, nitric oxide-mediated vascular mechanisms in the carotid artery. In addition, we have uncovered an endothelium-dependent vasoconstrictor component of the response of HU treated carotid arteries to norepinephrine. This component appears to compensate partially for the HU-induced vascular hyporesponsiveness in this blood vessel. It is unknown whether exposure of humans in microgravity produces similar endothelium-dependent vasodilator and vasoconstrictor changes. The present finding of an apparent COX-dependent vasoconstrictor component raises the caution that the use of COX inhibitors by astronauts may have the potential to exacerbate postural intolerance by further depressing the ability of arteries to constrict to the endogenous neurotransmitter, norepinephrine.