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J Appl Physiol 101: 348-353, 2006. First published April 20, 2006; doi:10.1152/japplphysiol.00941.2005
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HIGHLIGHTED TOPIC
Regulation of the Cerebral Circulation

Endothelium-dependent vasodilation of cerebral arteries is altered with simulated microgravity through nitric oxide synthase and EDHF mechanisms

Rhonda D. Prisby,1 M. Keith Wilkerson,3 Elke M. Sokoya,4 Robert M. Bryan, Jr.,4 Emily Wilson,5 and Michael D. Delp1,2

1Division of Exercise Physiology and 2Department of Physiology and Pharmacology, and Center for Interdisciplinary Research in Cardiovascular Sciences, West Virginia University School of Medicine, Morgantown, West Virginia; 3Department of Pharmacology, University of Vermont College of Medicine, Burlington, Vermont; 4Department of Anesthesiology, Baylor College of Medicine, Houston, Texas; and 5Department of Medical Physiology and the Cardiovascular Research Institute, Texas A&M University Health Science Center, College Station, Texas

Submitted 1 August 2005 ; accepted in final form 25 January 2006


   ABSTRACT
TOP
 ABSTRACT
MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 GRANTS
 ACKNOWLEDGMENTS
 REFERENCES
 
Cephalic elevations in arterial pressure associated with microgravity and prolonged bed rest alter cerebrovascular autoregulation in humans. Using the head-down tail-suspended (HDT) rat to chronically induce headward fluid shifts and elevate cerebral artery pressure, previous work has likewise shown cerebral perfusion to be diminished. The purpose of this study was to test the hypothesis that 2 wk of HDT reduces cerebral artery vasodilation. To test this hypothesis, dose-response relations for endothelium-dependent (2-methylthioadenosine triphosphate and bradykinin) and endothelium-independent (nitroprusside) vasodilation were determined in vitro in middle cerebral arteries (MCAs) from HDT and control rats. All in vitro measurements were done in the presence and absence of the nitric oxide synthase inhibitor NG-nitro-L-arginine methyl ester (10–5 M) and cyclooxygenase inhibitor indomethacin (10–5 M). MCA caveolin-1 protein content was measured by immunoblot analysis. Endothelium-dependent vasodilation to 2-methylthioadenosine triphosphate and bradykinin were both lower in MCAs from HDT rats. These lower vasodilator responses were abolished with NG-nitro-L-arginine methyl ester but were unaffected by indomethacin. In addition, HDT was associated with lower levels of MCA caveolin-1 protein. Endothelium-independent vasodilation was not altered by HDT. These results indicate that chronic cephalic fluid shifts diminish endothelium-dependent vasodilation through alterations in the endothelial nitric oxide synthase signaling mechanism. Such decrements in endothelium-dependent vasodilation of cerebral arteries could contribute to the elevations in cerebral vascular resistance and reductions in cerebral perfusion that occur after conditions of simulated microgravity in HDT rats.

orthostatic intolerance; cerebral blood flow; hindlimb unloading; middle cerebral artery; endothelium-dependent hyperpolarizing factor

REDUCED ORTHOSTATIC TOLERANCE often occurs in individuals after spaceflight and prolonged bed rest (1, 2, 16, 28, 35). Although many of these orthostatically intolerant individuals manifest arterial hypotension upon the assumption of an upright posture (1, 2, 16, 35), this is not always the case (2). Thus it has been hypothesized that chronic elevations in hydrostatic pressure to the head that accompany the cephalad fluid shifts associated with spaceflight and head-down-tilt bed rest may alter cerebral vasomotor responsiveness and increase cerebral vascular resistance in these individuals. Increased cerebral vascular resistance in cosmonauts has been reported after space flight (1, 16). Furthermore, cerebral blood flow velocity in bed-rested humans is attenuated across a range of arterial pressures, indicating that cerebral autoregulation of perfusion has been altered (35).

To study the effects of chronic headward fluid shifts on cerebrovascular control mechanisms, investigators have used the head-down tail-suspended (HDT) rat as a model (11, 29–31, 34). Previous work has shown that after a 2-wk period of elevated fluid pressure to the head of HDT rats, cerebral blood flow is lower and vascular resistance is higher in numerous regions of the brain during normal standing and head-up tilt (30). This elevation in cerebral vascular resistance was associated with enhanced basal tone of middle cerebral arteries (MCAs) from HDT rats through the downregulation of an endothelium-dependent nitric oxide (NO) synthase (NOS) mechanism (30). Furthermore, it was found that the levels of endothelial NOS (eNOS) protein were markedly decreased in MCAs from HDT rats relative to control animals. The purpose of this investigation was to determine whether HDT adversely affects stimulated vasodilator responsiveness of isolated MCAs, and, in particular, endothelium-dependent vasodilation. It was hypothesized that endothelium-dependent vasodilation is diminished in MCAs from HDT rats through the NOS signaling mechanism. In addition, we investigated whether caveolin-1 protein, an endogenous inhibitor of eNOS and a structural protein involved in organization of plasma membrane caveolae, was elevated in HDT rats.


   MATERIALS AND METHODS
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 ABSTRACT
 MATERIALS AND METHODS
RESULTS
 DISCUSSION
 GRANTS
 ACKNOWLEDGMENTS
 REFERENCES
 
The procedures employed in this study were approved by the Institutional Animal Care and Use Committee and conform with the Guide for the Care and Use of Laboratory Animals published by the US National Institutes of Health (NIH Publication no. 85-23, revised 1996). Six- to 8-mo-old male Sprague-Dawley rats were randomly assigned to either cage control or 2-wk HDT groups. The 2-wk HDT treatment has been shown to induce cerebral and peripheral cardiovascular alterations (8, 21, 29–31).

After the 2-wk experimental period, the animals were anesthetized with pentobarbital sodium (35 mg/kg ip) and decapitated, and the brain was rapidly removed and placed in a 4°C physiological saline-albumin buffer solution (PSS). MCAs were isolated for in vitro experimentation or determination of endothelial caveolin-1 protein content.

In Vitro Studies

Isolated segments of the MCA were placed in vessel chambers containing PSS at 37°C as previously described (2, 3, 13, 25, 30, 33). A micropipette filled with PSS was inserted into one end of the vessel and secured with 11-0 nylon ophthalmic suture. The other end of the vessel was cannulated with a second resistance-matched micropipette and secured with suture. After cannulation, each isolated vessel in the tissue chamber was transferred to the stage of an inverted microscope coupled to a video camera, video micrometer, videotape recorder, and data-acquisition system. To maintain a constant intraluminal pressure of 75 mmHg (10), the micropipettes cannulating the MCAs were connected to two independent hydrostatic fluid reservoirs; pressure was measured through side arms of the reservoir lines with low-volume-displacement strain gauge transducers. Leaks were detected by pressurizing the vessel and then closing the valves to the reservoirs and verifying that intraluminal pressure remained constant. Vessels free of leaks equilibrated for at least 1 h at 37°C to develop basal tone; the bathing solution was replaced every 15 min during the equilibration period.

Experimental design.   A series of in vitro experiments was performed to investigate the effects of HDT on endothelium-dependent and endothelium-independent vasodilation. The first series determined endothelium-dependent vasodilator responsiveness to the P2Y1-purinoceptor agonist 2-methylthioadenosine triphosphate (2-MeS-ATP; 10–9 to 10–6 M) alone and in the presence of the NOS inhibitor NG-nitro-L-arginine methyl ester (L-NAME; 10–5 M), the cyclooxygenase inhibitor indomethacin (Indo; 10–5 M), both L-NAME (10–5 M) and Indo (10–5 M), and during blockade of NOS (L-NAME; 10–5 M) and Ca2+-activated potassium channels (KCa; 10–7 M charybdotoxin and 3 x 10–7 M apamin) as previously described (14, 25, 30). A second series of studies determined the endothelium-dependent vasodilator responsiveness of MCAs to the B2-receptor agonist bradykinin (BK; 10–10 to 10–6 M) alone and in the presence of L-NAME (10–5 M) or Indo (10–5 M). To elicit endothelium-dependent vasodilation, MeS-ATP, BK, and the various inhibitors were added to the luminal perfusate in the fluid reservoirs connected to the pipettes cannulating the MCA. The height of the reservoirs was then adjusted to achieve an intraluminal flow of 20 µl/min, a rate of which does not induce a vascular response that differs between MCAs from control or HDT rats (30). In a final series of studies, concentration-response relations to the abluminal application of the exogenous NO donor sodium nitroprusside (SNP; 10–10 to 10–5 M) were determined alone and in the presence of L-NAME (10–5 M). To avoid possible tachyphylaxis, only one concentration-response determination was made per vessel segment. At the conclusion of each experiment, arteries were incubated in Ca2+-free PSS for 1 h to determine maximal diameter at an intraluminal pressure of 75 mmHg.

Drugs and reagents.   SNP, BK, L-NAME, Indo, charybdotoxin, and apamin were purchased from Sigma, and 2-MeS-ATP from ICN Pharmaceuticals PSS buffer contained (in mM) 145 NaCl, 4.7 KCl, 1.2 NaH2PO4, 1.17 MgSO4, 2.0 CaCl2, 5.0 glucose, 2.0 pyruvate, 0.02 EDTA, and 3.0 MOPS (Sigma), plus bovine serum albumin (USB Chemicals, 10 mg/ml), with a pH of 7.4. Ca2+-free PSS buffer contained 2 mM EDTA and CaCl2 was replaced with 2.0 mM NaCl.

Evaluation of Caveolin-1 Expression

MCAs were snap frozen and stored at –80°C in 0.5-ml microcentrifuge tubes. Differences in caveolin-1 protein expression in MCAs were assessed by immunoblot analysis as previously described (27, 30). Briefly, MCA proteins (10 µg total protein per sample) were subjected to SDS-PAGE (8–16% gradient gel) and transferred to nitrocellulose membrane. Membranes were blocked and incubated with primary antibody overnight at 4°C. Primary antibody dilutions were as follows: caveolin-1, 1:1,250, and GAPDH, 1:1,000 in blocking buffer. After washing, membranes were incubated with the appropriate horseradish peroxidase-conjugated species-specific anti-IgG (1:50,000 to 1:100,000 depending on primary antibody) for 2 h at 25°C. Peroxidase activity was detected by use of West Dura Extended (Pierce). Normalization for loading differences and blot-to-blot variability in density was accomplished by using ratios of the densitometry signals for caveolin-1 vs. GAPDH protein. GAPDH was chosen as a normalization control after empirical verification that no quantitative differences in expression was observed in vessels from control and HDT samples when equal protein amounts were loaded.

Statistical Analysis

To control for possible variations in vessel size between groups (30, 31), vasodilator responses were recorded as actual diameters and subsequently expressed as a percentage of maximal vasodilation according to the following formula:

Formula

where Dm is the maximal diameter recorded at 75 mmHg in calcium-free PSS, Ds is the steady-state diameter recorded after each dose of 2-MeS-ATP, BK, or SNP, and Db is the initial baseline diameter recorded immediately before the application of the first dose of vasodilator substance as previously described (25, 27, 33). Concentration-diameter relations were evaluated by repeated-measures ANOVA to detect differences within (dose) and between (experimental groups) factors. Pairwise comparisons between specific levels were made through post hoc analysis (Student-Newman-Keuls) when a significant main effect was found. The development of basal tone was expressed as the percent constriction relative to maximal diameter and was calculated as:

Formula

A one-way ANOVA was used to determine differences between groups for body weight, muscle weights, basal tone, and maximal diameter. All values are presented as means ± SE. A value of P < 0.05 was required for significance.


   RESULTS
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DISCUSSION
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Maximal MCA intraluminal diameter was similar between groups (control, 239 ± 4 µm; HDT, 225 ± 4 µm), although MCA wall thickness was greater in HDT rats (control, 11 ± 1 µm; HDT, 17 ± 1 µm). Development of basal tone was greater in MCAs from HDT (34 ± 3%) than control (24 ± 2%) rats, and, correspondingly, baseline diameter was less in HDT cerebral arteries (control, 185 ± 9 µm; HDT, 147 ± 6 µm).

Vasodilator Responses

Endothelium-dependent vasodilation induced by 2-MeS-ATP was less in MCAs from HDT rats (Fig. 1). L-NAME completely abolished vasodilator responses of control arteries to 2-MeS-ATP, whereas the NOS inhibitor only partially blocked the vasodilation of MCAs from HDT animals. The cyclooxygenase inhibitor Indo had no effect on the vasodilator responses of either control or HDT MCAs (Fig. 1). The addition of both L-NAME and Indo completely blocked vasodilation in control MCAs, but only partially blocked the vasodilation in HDT MCAs (Fig. 2), similar to that of L-NAME alone. The combined NOS and KCa blockade obliterated all responses to 2-MeS-ATP in MCAs from both groups (Fig. 2). Endothelium-dependent vasodilation induced by BK was less in MCAs from HDT rats (Fig. 3). L-NAME inhibited the vasodilator responses in both control and HDT arteries to BK and abolished differences in BK-mediated vasodilation between groups. In contrast, Indo had no effect on BK-mediated vasodilation in MCA from control or HDT rats (data not shown). The endothelium-independent vasodilation induced by the NO-donor SNP did not differ between groups either in the presence or absence of L-NAME (Fig. 4).


Figure 1
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  		Fig. 1. Concentration-response relation of middle cerebral arteries from Control and head-down tail-suspended (HDT) rats to 2-methylthioadenosine triphosphate (2-MeS-ATP) alone and in the presence of the nitric oxide synthase (NOS) inhibitor NG-nitro-L-arginine methyl ester (L-NAME; 10–5 M) and the cyclooxygenase inhibitor indomethacin (Indo; 10–5 M). Values are means ± SE. *HDT vasodilator response significantly different from respective control response (P ≤ 0.05).

 

Figure 2
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  		Fig. 2. Concentration-response relation of middle cerebral arteries from Control and HDT rats to 2-MeS-ATP alone and in the presence of the combined NOS (L-NAME; 10–5 M) and cyclooxygenase (Indo; 10–5 M) inhibitors, and the combined NOS (L-NAME; 10–5 M) and Ca2+-activated potassium channels [10–7 M charybdotoxin (ChTX) and 3 x 10–7 M apamin (Apa)] inhibitors. Values are means ± SE. *HDT vasodilator response significantly different from respective control response (P ≤ 0.05).

 

Figure 3
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  		Fig. 3. Concentration-response relation of middle cerebral arteries from Control and HDT rats to bradykinin alone and in the presence of L-NAME (10–5 M). Values are means ± SE. *HDT vasodilator response significantly different from respective control response (P ≤ 0.05).

 

Figure 4
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  		Fig. 4. Concentration-response relation of middle cerebral arteries from Control and HDT rats to the nitric oxide donor sodium nitroprusside alone and in the presence of L-NAME (10–5 M). Values are means ± SE. Responses did not differ among groups (P > 0.05).

 
Caveolin-1 Expression

HDT resulted in a lower in caveolin-1 protein content in MCAs (Fig. 5).


Figure 5
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  		Fig. 5. Relative changes in caveolin-1 protein expression of middle cerebral arteries from Control and HDT rats. Expression was quantified by Western blot analysis and normalized to the expression of GAPDH. Values are means ± SE. *HDT caveolin-1 protein content significantly less than that in Control arteries (P ≤ 0.05).

 

   DISCUSSION
TOP
 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
GRANTS
 ACKNOWLEDGMENTS
 REFERENCES
 
The purpose of this study was to test the hypothesis that HDT diminishes endothelium-dependent vasodilation in MCAs through a NO signaling mechanism. The results demonstrate that endothelium-dependent vasodilation mediated through 2-MeS-ATP and BK is attenuated in MCAs from HDT rats. The vasodilator responsiveness of MCAs from control and HDT rats was not affected by cyclooxygenase inhibition. In contrast, inhibition of NOS activity with L-NAME abolished the greater vasodilation of MCAs from control animals, indicating the deficit in endothelium-dependent vasodilation of MCAs from HDT rats occurs through the NOS signaling mechanism. In addition, we anticipated that the previously reported decrease in eNOS protein expression (30) and the diminished endothelium-dependent vasodilation (present study) would be coupled to an increase in caveolin-1 expression in MCAs from HDT rats. However, contrary to our hypothesis, there was a decreased expression of caveolin-1 (Fig. 5), suggesting a more complex relation among these and possibly other variables associated with the adaptation of cerebral arteries to simulated microgravity. Vasodilation in response to sodium nitroprusside, a NO donor that mediates an endothelium-independent vasodilation, did not differ between groups, indicating that smooth muscle cell relaxation to NO remains unaltered in MCAs from HDT rats. These results suggest that reductions in endothelium-mediated vasodilation could contribute to the diminished perfusion and lower vascular conductance that have been reported to occur in key regulatory areas of the brain after chronic HDT (30).

The mechanism of the reduced endothelium-dependent vasodilation appears to result from a downregulation of the endothelial NO signaling pathway. It has been widely reported that eNOS expression is sensitive to chronic changes in intravascular blood flow and shear stress (6, 22). Indeed, we have previously shown that cerebral blood flow during HDT is reduced by ~25% (29) and that eNOS protein content is diminished in MCAs from HDT rats (30). On the basis of these studies we hypothesized that proteins that are involved in the regulation of NOS activity might also be altered in response to HDT. One such protein was caveolin-1, an inhibitor of eNOS activity, which was expected to be elevated in accordance with the observed decrease in NO-mediated vasodilation. Contrary to expectations, levels of caveolin-1 protein were diminished in MCAs from HDT rats (Fig. 5).

Caveolin-1 is a scaffolding protein that is considered to be a primary marker and structural protein of plasma membrane caveolae. It is expressed in a number of cell types, including vascular endothelial and smooth muscle cells. In addition to its structural role in the organization of caveolae, caveolin-1 binds to eNOS and maintains it in an inactive state until it is competed off by calmodulin (5). A role for caveolin-1 in modulating eNOS activity is corroborated by increased NO production in the caveolin knockout mice (24). However, decreased caveolin-1 may also contribute to the altered vasodilator responses because it is implicated in modulation of smooth muscle contractile events (17). Additionally, decreased caveolin-1 levels are associated with increased smooth muscle proliferation (15, 23, 24), and thus the diminished levels of caveolin-1 content may contribute to the structural changes in the wall of cerebral arteries from HDT rats, i.e., the increase in medial wall thickness (30, 31, present study). Decreased vascular levels of caveolin-1 expression are also associated with other pathologies, such as aging (26) and hypertension (E. Wilson, unpublished observations), where endothelium-dependent vasodilation is diminished.

Despite the fact that the NOS mechanism of endothelium-dependent vasodilation is down, it appears that an endothelium-derived hyperpolarizing factor (EDHF) is concomitantly upregulated with HDT during vasodilator responses induced by 2-Me-ATP (Fig. 1), the more potent of the two endothelium-dependent vasodilator substances studied. EDHF does not normally play any role in endothelium-dependent relaxation induced by 2-Me-ATP (12, 20). This is evident in the present investigation, in which inhibition of NOS activity with L-NAME completely abolished 2-MeS-ATP-mediated vasodilation in MCAs from control rats (Fig. 1). However, in MCAs from HDT rats L-NAME only partially abolished the vasodilator response to 2-Me-ATP. The residual vasodilation was, however, eliminated with the additional KCa blockade, indicating that the reduction in NOS-mediated vasodilation was partially compensated for by an upregulation of an EDHF-mediated response. This type of adaptive response of the cerebral vasculature is not unique to the HDT model. Several pathological conditions, including ischemia-reperfusion injury (20) and traumatic brain injury (12), have shown attenuated endothelial NO-mediated vasodilation with a simultaneous potentiation of EDHF-mediated vasodilation. Thus this appears to represent an adaptive response of cerebral arteries under certain conditions, such as simulated microgravity, to enhance endothelium-dependent vasodilation mediated by some (e.g., 2-MeS-ATP), but not all (e.g., BK) agonists.

Although the present study and previous works (11, 30) implicate a downregulation of the NOS signaling pathway for diminished endothelium-mediated vasodilation and enhanced vasoconstriction of cerebral arteries, not all studies have come to similar conclusions. For example, Ma et al. (19) reported that eNOS and inducible NOS protein content of a collection of cerebral arteries were not different between control and HDT rats. Furthermore, nitrate and nitrite content, stable metabolites of NO, were elevated in these cerebral arteries from HDT rats, indicating that NO bioavailability in cerebral arteries is actually enhanced by simulated microgravity. In another study, Zhang et al. (34) have examined the effects of HDT on vasodilator responses through endothelium-dependent and -independent mechanisms in basilar arteries. Similar to the results of the present study, Zhang et al. found that HDT did not alter vasodilator responses to the endothelium-independent agonists SNP and adenosine. However, unlike those responses of the present study, they also reported that endothelium-dependent vasodilator responses to acetylcholine and thrombin were unaltered by HDT. Several factors could contribute to the apparent disparity in results from these studies, including the cerebral arteries studied (MCA vs. basilar artery), the methodology employed (pressurized artery measuring changes in vessel diameter vs. wire myograph measuring changes in isometric tension), and the agonists used (2-MeS-ATP and BK vs. acetylcholine and thrombin). Another variable confounding comparison between the present study and that of Zhang et al. is the duration of the HDT treatment (2 wk vs. 4 wk). Because Zhang et al. used a 4-wk HDT treatment period, it is possible that a compensatory upregulation of the EDHF mechanism of vasodilation (e.g., Fig. 2) could have normalized the deficit in NO-mediated vasodilation over the more prolonged period of HDT. Thus the possibility remains that compensatory mechanisms may act to normalize endothelium-dependent vasodilation through different signaling pathways with prolonged exposure to simulated microgravity.

One possible stimulus for the attenuation of endothelium-dependent vasodilation and the downregulation of the NOS signaling pathway is a decrement in physical activity imposed by the HDT treatment. HDT does not, however, elicit a generalized impairment of endothelial function. For example, endothelium-dependent vasodilation of the aorta from HDT rats is normal (9). In skeletal muscle, endothelium-dependent vasodilation and eNOS protein expression are depressed in arterioles from the soleus muscle (8, 32), a muscle that undergoes significant atrophy and reductions in activity during the HDT treatment. In contrast, other muscles such as the superficial portion of gastrocnemius muscle that do not undergo such dramatic changes in activity and mass have normal endothelium-dependent vasodilation and eNOS expression (8, 32). Thus reductions in endothelium-dependent vasodilation are not generalized among muscles or throughout the body but appear to be evident only in tissue undergoing significant decrements in metabolic activity. If physical inactivity is linked with the impairment of endothelial function in the cerebral circulation, it may be related specifically to altered patterns of cerebral perfusion associated with exercise or inactivity (7).

In addition to the diminished endothelium-dependent vasodilation that occurs in cerebral arteries after HDT, other factors likely contribute to diminished cerebral perfusion after HDT (30). Increased vasoconstrictor responses to KCl, arginine vasopressin, and 5-hydroxytryptamine have been reported after 4 wk of HDT in basilar arteries (34). In MCAs from HDT rats, Geary et al. (11) reported greater myogenic vasoconstriction, and Wilkerson et al. (30) also reported enhanced basal tone and greater vasoconstrictor responses to transmural pressure, shear stress, and KCl. Collectively, these studies suggest that multiple mechanisms involving diminished endothelium-dependent vasodilation and greater vasoconstriction of cerebral arteries contribute to the lower brain blood flow and vascular conductance apparent in rats exposed to conditions of simulated microgravity.

Altered autoregulation of cerebral perfusion has been reported to occur in humans subsequent to prolonged headward fluid shifts, such as those induced by microgravity and head-down bed rest (1, 2, 16, 35). Previous work in animals has shown that inhibition of the NOS signaling pathway results in a downward shift in the cerebral autoregulatory curve (i.e., lower cerebral blood flow for a given arterial pressure) (18). Therefore, results of the present study, as well as previous reports demonstrating a HDT-induced enhancement of cerebral artery vasoconstriction through the downregulation of the NOS signaling pathway (11, 30), may underlie the reductions in cerebral blood flow that occur in rats during and after exposure to simulated microgravity (29, 30). Likewise, diminished endothelium-mediated vasodilation and enhanced vasoconstrictor responsiveness of the cerebral resistance vasculature provide a plausible explanation for the mechanism underlying the downward shift in cerebral autoregulation reported to occur in humans after periods of reduced gravitational stress and bed rest. Such a change in cerebral vascular responsiveness could induce orthostatic intolerance in normotensive individuals by limiting cerebral blood flow, or it could exacerbate difficulties in maintaining cerebral perfusion in individuals experiencing orthostatic hypotension (4).

In conclusion, the present study demonstrates that 2-MeS-ATP- and BK-elicited vasodilation is diminished in MCAs from HDT rats and that this attenuated endothelium-dependent vasodilation occurs through alterations in the NOS signaling pathway (Figs. 1 and 3). Furthermore, the diminished endothelium-dependent vasodilation is associated with reduced NOS (30) and caveolin-1 (present study) protein expression. Concomitant with the attenuated NOS component of the 2-MeS-ATP-mediated vasodilation is an enhancement of an EDHF-mediated vasodilator component in MCAs from HDT rats. Although the upregulation of an EDHF signaling mechanisms appears to partially compensate for the diminished NOS signaling pathway, the net effect is that endothelium-dependent vasodilation in HDT rats remains lower than that in cerebral arteries from control animals. The functional consequence of these and other vascular alterations (e.g., 11, 30, 31, 34) with HDT appear to be greater cerebral vascular resistance and lower cerebral perfusion (30). When applied to the human condition, these results suggest that reported alterations in cerebral autoregulation associated with microgravity or prolonged bed rest may be the result of diminished levels of cerebral artery endothelial NO. Although diminished NO bioavailability may be an appropriate adaptation of the cerebral vasculature to the elevations in arterial pressure associated with chronic headward fluid shifts, this adaptive response may limit the ability of cerebral vasculature to respond to abrupt alterations in pressure, such as that which occurs with orthostatic stress (1, 2, 16, 35).


   GRANTS
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 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 GRANTS
ACKNOWLEDGMENTS
 REFERENCES
 
This work was supported by National Aeronautics and Space Administration Grants NNA04CC66G and NCC2-1166 and by National Space Biomedical Research Institute Grant NCC9-58-42 CA00209.


   ACKNOWLEDGMENTS
TOP
 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 GRANTS
 ACKNOWLEDGMENTS
REFERENCES
 
The authors gratefully acknowledge Jan L. Patterson for technical contributions to this work.


   FOOTNOTES
 

Address for reprint requests and other correspondence: M. D. Delp, Div. Exercise Physiology, West Virginia Univ. School of Medicine, Morgantown, WV 26506 (e-mail: mdelp{at}hsc.wvu.edu)

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.


   REFERENCES
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 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 GRANTS
 ACKNOWLEDGMENTS
 REFERENCES
 

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