Development of Nitric Oxide and Prostaglandin Mediation of Shear Stress–Induced Arteriolar Dilation With Aging and Hypertension
Abstract—We hypothesized that during hypertension, the impairment of mediation of shear stress–induced dilation by nitric oxide (NO) is due to the prevailing hemodynamic forces, and that mediation of this response by NO should still be present in young spontaneously hypertensive rats (SHR). Thus, responses to increases in perfusate flow eliciting increases in wall shear stress were investigated in pressurized (80 mm Hg), isolated arterioles (≈70 to 100 μm) of the left or right gracilis muscle obtained from the same WKY and SHR at 4 and 12 weeks of age. Flow-induced dilations were similar in WKY and SHR at 4 weeks (maximum, 26.5±1.8 and 24.2±2.0 μm, respectively). Also, the middle of the upward portion of the shear stress–diameter curves was similar in arterioles of the 2 strains. Inhibition of NO synthase with Nω-nitro-l-arginine (L-NNA) or inhibition of synthesis of prostaglandins (PGs) with indomethacin elicited an ≈50% reduction in flow-dependent dilation, whereas their combined administration eliminated the responses in both groups. In arterioles of 12-week-old WKY, flow-induced dilation became significantly greater (maximum, 46.1±2.3 μm) than responses of arterioles of 4-week-old WKY and 12-week-old SHR (maximum, 18.3±5.9 μm), which shifted only the shear stress–diameter curve of the 12-week-old WKY significantly to the left. Also, at 12 weeks of age, flow-dependent dilation of arterioles from SHR is mediated solely by PGs. Thus, shear stress–induced arteriolar dilation is mediated by NO and PGs in 4-week-old WKY and SHR. With aging, the release of NO and PGs increases in normotensive rats, whereas the contribution of NO to the regulation of shear stress disappears in 12-week-old SHR, which suggests that this change is probably caused by the increase in intraluminal pressure as hypertension develops.
Arteriolar endothelium plays an important role in the regulation of skeletal muscle microcirculation via the production and release of dilator factors such as nitric oxide (NO) and prostaglandins. In vivo and in vitro studies have shown that one of the most important physiological stimuli for the release of NO and prostaglandins is the presence and change in wall shear stress.1 2 3 Regulation of wall shear stress contributes to the minimization of energy loss in the circulation by adjusting vascular resistance3 ; thus, it is important to long-term regulation of blood pressure. Previous morphological studies suggest that alteration in flow during normal or pathological development can greatly influence the diameter and growth of vessels.4 5 Furthermore, shear stress–induced dilation elicited by an increase in intraluminal flow is reduced in isolated skeletal muscle arterioles of spontaneously hypertensive rats (SHR) by the age of 12 weeks.6 7 8 This alteration is likely to contribute further to the increased peripheral vascular resistance. Studies also have shown that the reason for the reduced response is an impairment of NO-mediated portion of the response, whereas the mediation by prostaglandins is retained.7 These and other findings9 10 suggest that elevated blood pressure is deleterious to the endothelium. Furthermore, recent findings have suggested that acute hypertension in the coronary circulation results in impairment of endothelium11 and that acute elevation of intraluminal pressure in isolated arterioles of normotensive rats elicits an impairment of responses mediated by NO.12
On the basis of these findings, we hypothesized that at an early age when the systemic blood pressure is still not significantly higher than normal, flow-induced dilation in skeletal muscle arterioles of normotensive and hypertensive rats should be present, similar, and mediated by both NO and prostaglandins. Furthermore, it is likely that, with aging and as hypertension develops, the increase in intraluminal pressure or other factors will affect the synthesis and release of endothelial factors that mediate shear stress–induced dilation. These questions have not been studied before. To understand the development of the regulation of wall shear stress under normotensive and hypertensive conditions, it is important to assess the role of genetic and environmental factors.
To test our hypothesis, we characterized, in isolated arterioles of gracilis muscle of normotensive Wistar-Kyoto rats (WKY) and SHR, the magnitude and the mediation of flow-induced response at a prehypertensive age (4 weeks old) and contrasted these responses with those in vessels isolated from the same WKY and SHR at 12 weeks of age, when the systemic blood pressure is significantly elevated in SHR compared with WKY.
In the present study, male normotensive WKY (n=16), and SHR (n=13) (Charles River Laboratories, Mass) were used. The procedures followed were in agreement with institutional guidelines. Systolic blood pressure was measured by the tail-cuff method. Rats were anesthetized with intraperitoneal injections of sodium pentobarbital (Nembutal sodium, 50 mg/kg). Experiments were conducted on arterioles isolated from gracilis muscle (left and right) of the same rats at 4 and 12 weeks of age (≈70 and 100 μm in diameter), respectively. The isolation procedure of gracilis muscle arterioles has been described previously.2 6 Briefly, when rats were 4 weeks old, the left gracilis muscle of each rat was exposed by an incision of the skin. A selected portion of the gracilis muscle was then cut out and placed on a Petri dish containing cold (4°C) physiological salt solution (PSS1; pH 7.4), which was composed of (in mmol/L) 145 NaCl, 5.0 KCl, 2.0 CaCl2, 1.0 MgSO4, 1.0 NaH2PO4, 5.0 dextrose, 2.0 pyruvate, 0.02 EDTA, and 3.0 MOPS. The piece of muscle was pinned to the silicone bottom of the dish and allowed to equilibrate for ≈15 minutes. Rats were treated with an antibiotic (amoxicillin 6 mg/kg IM, BID for 3 days; Pfizer Animal Health) and an analgesic (Buprenex 0.3 mg/kg IM, BID for 3 days; Reckitt and Colman Pharmaceuticals Inc). The skin was closed with sterile suture, and rats were allowed to recover from anesthesia. At 12 weeks of age, arterioles either from the right gracilis muscle of the same rats or from another group of rats were used.
With the use of microsurgical instruments and an operating microscope (Olympus), a (1-mm-long) segment of a first-order arteriole was isolated from the gracilis muscle and surrounding tissue and transferred to the vessel chamber. The chamber contained a pair of glass micropipettes filled with physiological salt solution (PSS2) at room temperature. The PSS2 solution used for suffusion and perfusion of the vessels contained (in mmol/L) 110.0 NaCl, 5.0 KCl, 2.5 CaCl2, 1.0 MgSO4, 10.0 dextrose, 24.0 NaHCO3, and 0.02 EDTA; it was equilibrated with a mixture of 21% O2/5% CO2 balanced with N2, pH 7.4 (37°C). From a (60-mL) reservoir, the vessel chamber (15 mL) was continuously supplied with PSS2 at a rate of 40 mL/min. After the vessel was mounted on the proximal pipette and secured with sutures, the perfusion pressure was raised to 20 mm Hg to clear the debris from the lumen. Next, the other end of the vessel was mounted on the distal pipette. As described previously,2 6 both proximal (inflow) and distal (outflow) micropipettes were connected with silicone tubing to a pressure-servo syringe system (Living Systems Inc). The system was arranged to have mirror symmetry, and only pipettes with similar dimensions and equivalent resistances to flow were used. The temperature was set to 37°C (YSI temperature controller), and the vessels were allowed to equilibrate for ≈1 hour.
In all protocols, only those vessels that developed spontaneous tone to pressure were used, because no vasoactive agent was added to the PSS2. After the equilibration period, flow-diameter relationships were obtained under control conditions in both strains of rats. In the arterioles, the perfusate flow was increased from 0 to 14 (for 4-week-old) or 25 (for 12-week-old) μL/min (in 2- and 5-μL/min steps, respectively).6 Flow was established at a constant intravascular pressure (80 mm Hg) by changing proximal and distal pressures to an equal degree in opposite directions to keep midpoint luminal pressure constant. The flow was measured by a ball flowmeter (Omega) that was calibrated by a Harvard perfusion pump in the range of 0 to 100 μL/min. Each flow step was maintained for ≈5 minutes to allow the vessels to reach a steady-state condition before the diameter of the arterioles was measured. After control flow-diameter curves were obtained, we subjected the vessels to Nω-nitro-l-arginine (L-NNA; 10−4 mol/L), an inhibitor of NO synthesis.2 Then, after an ≈15-minute incubation period, changes in diameter in response to step increases in perfusate flow were reassessed. The role of prostaglandins in flow-induced dilation of gracilis muscle arterioles was also assessed. To inhibit the synthesis of prostaglandins,2 indomethacin (INDO; 10−5 mol/L) was added to the PSS2 containing L-NNA. After the incubation period (≈30 minutes), the flow-diameter relationships were once more assessed. In separate experiments, INDO was given before the administration of L-NNA to exclude the effect of possible interaction between NO and cyclooxygenase.
All drugs were added to the reservoir connected to the vessel chamber, and final concentrations are reported. To assess the active tone generated by the arterioles in response to intravascular pressure, at the conclusion of each experiment, the suffusion solution was changed to a Ca2+-free PSS2 that contained sodium nitroprusside (10−4 mol/L) and EGTA (1.0 mmol/L). Vessels were incubated for 10 minutes, and then the passive diameter of arterioles at 80 mm Hg perfusion pressure was obtained.
All salts and chemicals were obtained from Sigma Chemical Co or Aldrich Co and were prepared on the day of the experiment. The diameter of vessels, under various experimental conditions, were measured with an image-shearing monitor (IPM, model 907) and recorded with an X-Y recorder (Multicorder, MC6625). The wall shear stress was calculated according to the following equation: τ=4Q η/πr3, where Q is perfusate flow, η is viscosity of the perfusate (≈0.007 poise at 37°C), and r is vessel radius. Data are presented as mean±SEM; n indicates number of rats. Statistical analyses were done by ANOVA followed by the Tukey post hoc test or regression analysis as appropriate. A value of P <0.05 was considered significant.
The systolic blood pressures of 4-week-old normotensive WKY and SHR were 122.8±3.3 and 121.5±4.7 mm Hg (n=8), respectively, whereas at 12 weeks of age, they were 133.3±2.3 and 193.5±8.1 mm Hg (n=8), respectively, which shows a significant increase in blood pressure in 12–week-old SHR.
Flow-Dependent Dilation of Arterioles From 4-Week-Old Rats
The active diameters of arterioles of 4-week-old WKY and SHR, obtained in the presence of constant intravascular pressure (80 mm Hg) and under static flow conditions, were significantly different (74.0±2.8 and 63.4±2.2 μm, respectively; P<0.01). After conclusion of the experiments and under the same conditions but in Ca2+-free solution, the passive diameter of each arteriole was also obtained (see Methods). We found that the mean passive diameters of 4-week-old WKY and SHR were also significantly different (148.8±4.2 and 122.7±2.4 μm, respectively; P<0.05), but the arteriolar tone, expressed as the percentage of passive diameter, was not different in the 2 strains of rats (49.8±1.3% and 51.7±1.5%).
Figure 1⇓, top, shows the changes in the diameter of arterioles from 4-week-old WKY and SHR in response to step increases in flow in control conditions. From 2 μL/min perfusate flow, the diameter of arterioles from 4-week-old WKY and SHR increased significantly. The increase in diameter at 14 μL/min flow was similar in arterioles of 4-week-old WKY and SHR. Also, no significant difference existed in the slope of flow-diameter curves, which indicates that arterioles of WKY and SHR exhibit similar dilations to increases in perfusate flow.
Figure 1⇑, bottom, shows the changes in the diameter of arterioles from 4-week-old WKY and SHR in response to step increases in wall shear stress in control conditions. From 5 dyne/cm2 of shear stress, the diameter of arterioles from 4-week-old WKY and SHR increased substantially, which delayed the increase in shear stress as flow increased. The middle portion of the shear stress–diameter curves and also the slopes of these curves were similar in arterioles of 4-week-old WKY and SHR.
Next, we investigated the endothelial mechanisms responsible for the mediation of flow-induced dilation of arterioles of WKY and SHR at 4 weeks of age. INDO, a blocker of prostaglandin synthesis, did not affect basal diameter (48.8±2.0 to 46.7±3.6 and 50.7±2.6 to 49.1±1.3 μm) but significantly reduced the dilation to increases in perfusate flow in arterioles of both strains of rats (Figure 2⇓, top and bottom). In 4-week-old WKY, the reduction of the maximum response was ≈62%, whereas in 4-week-old SHR, it was ≈45% (at maximal flow rate).
L-NNA, a NO synthase inhibitor, similarly and significantly reduced basal diameter of arterioles from the 2 strains of 4-weeks-old rats (48.8±2.5 to 42.8±2.5 and 54.0±2.6 to 46.5±3.6 μm in WKY and SHR, respectively; P<0.05). In addition, in 4-week-old WKY and SHR, L-NNA significantly reduced flow-induced arteriolar dilation (Figure 2⇑, top and bottom). For example, at 14 μL/min flow, the diameter of L-NNA–treated arterioles of 4-week-old WKY and SHR was ≈44% and ≈46% less than that of control, respectively. Also, the slopes of flow-diameter curves were significantly different between control and in the presence of L-NNA. In the presence of L-NNA, administration of INDO elicited a further significant reduction of flow-induced responses and practically eliminated the dilation to increases in perfusate flow in both 4-week-old WKY and SHR (Figure 2⇑). Figure 3⇓ summarizes the effects of INDO and L-NNA on the calculated wall shear stress–diameter curves of these arterioles. The inhibitors had similar effects on responses of arterioles, and as a result, the maintained shear stress shifted to the right in both 4-week-old WKY and SHR (Figure 3⇓). In the presence of both L-NNA and INDO, shear stress did not induce arteriolar dilation either in WKY or SHR (Figure 3⇓); thus, shear stress increased to a high level (≈250 dyne/cm2).
Flow-Dependent Dilation of Arterioles From 12-Week-Old Rats
In the presence of 80-mm Hg perfusion pressure (no flow), the active diameter of arterioles from WKY and SHR was 97.0±4.0 and 109.0±5.7 μm, respectively, whereas the passive diameter of arterioles from WKY and SHR was 177.4±7.5 and 185.9±6.7 μm, respectively. The arteriolar tone expressed as the percentage of passive diameter was not different in the two 12-week-old strains of rats (54.2±1.2% and 58.6±2.4%, respectively).
Figure 4⇓, top, shows the changes in the diameter of arterioles of 12-week-old WKY and SHR in response to step increases in flow in control conditions. From 5 μL/min perfusate flow, the diameter of arterioles of 12-week-old SHR started to deviate significantly (P<0.05) from that of 12-week-old WKY, and at 25 μL/min flow, the change in diameter of 12-week-old SHR arterioles was ≈60% less than that of 12-week-old WKY. Also, the significant difference in the slope of flow–change in diameter curves indicates that in arterioles of 12-week-old SHR, the dilation to increases in perfusate flow is markedly reduced compared with the arterioles of 12-week-old WKY. Similarly, the shear stress–diameter curves (Figure 4⇓, bottom) clearly show that the maintained shear stress is significantly lower in arterioles of 12-week-old WKY.
In arterioles of 12-week-old WKY and SHR (data from our previous study6 indicated by dotted line), INDO did not affect basal tone (55±1.6% versus 59±2.4% and 52.5±2.8% versus 52.9±3%, respectively) but significantly reduced thedilation to increases in perfusate flow in arterioles of both strains of rats (Figure 5⇓, top and bottom). In 12-week-old WKY, the reduction of the maximum response was ≈53% (Figure 5⇓, top), whereas in SHR (Figure 5⇓, bottom), INDO eliminated the dilation.6 In 12-week-old WKY, L-NNA significantly reduced basal tone (54.8±1.1% versus 48.7±2.8%) and flow-induced arteriolar dilation (Figure 5⇓, top). For example, at 25 μL/min flow, the change in diameter of L-NNA–treated arterioles of 12-week-old WKY was ≈42% less than that of control. Also, the slopes of flow-diameter curves were significantly different between control and in the presence of L-NNA. In contrast, in arterioles of 12-week-old SHR, L-NNA did not significantly affect basal tone (48.8±4.4% versus 45.8±4.2%) or the arteriolar dilation in response to step increases in perfusate flow (Figure 5⇓, bottom; data from our previous study are given for comparison6 ). In the presence of L-NNA, administration of INDO eliminated the dilation to increases in perfusate flow in both strains of rats (Figure 5⇓).
In Figure 6⇓, the wall shear stress–change in diameter curves in arterioles of 4- and 12-week-old WKY and SHR are depicted to compare the changes in sensitivity of arterioles to wall shear stress as a function of age. In arterioles of WKY, the upward portion (maintained shear stress) of shear stress–diameter curves shifts to the left significantly from 4 to 12 weeks of age (Figure 6⇓, top). In contrast, in arterioles of 4- and 12-week-old SHR, the shear stress–diameter curves are not different (Figure 6⇓, bottom).
The new findings of this study of isolated gracilis muscle arterioles are (1) that shear stress–induced dilations elicited by increases in perfusate flow are similar and are mediated by both NO and prostaglandins in young (4-week-old), prehypertensive SHR as well as WKY, and (2) that during development from 4 to 12 weeks of age, shear stress–dependent dilation became enhanced in WKY as a result of an increased contribution of both NO and prostaglandin, whereas in SHR, shear stress–dependent dilation is significantly attenuated because of reduced NO mediation, while the response is mediated solely by prostaglandins.
Several studies suggest that an altered function of vascular endothelial cells is intimately involved in the pathogenesis of hypertension.13 14 15 16 17 18 However, it is not clear whether alterations in the function of endothelium are primary or secondary to the development of hypertension. Also, no extant studies address the mechanisms of microvascular endothelial changes as a function of age and the development of hypertension. The changes in microvessels could be different from those of large vessels, because the myogenic response, by preventing increases in intraluminal pressure in distal segments of the peripheral circulation,19 may provide for a protection of microvascular endothelium, at least in the early phase of hypertension.
Previous studies showed that endothelium can contribute to circulatory homeostasis by shear stress–dependent regulation of vascular resistance that can be stimulated by increases in blood flow.1 3 8 Flow-dependent dilation of arterioles has not yet been investigated in young hypertensive rats, at which time systemic blood pressure is not significantly different in WKY and SHR, which prevents assessment of the possible role of adaptation or changes of this mechanism as a function of age during normal development and the development of hypertension. Therefore, we aimed to clarify whether flow-induced dilation is present in arterioles of young normotensive and genetically hypertensive rats and, if so, what endothelial factors mediate the response. Arterioles of WKY and SHR gracilis muscle were chosen for the present study, and flow-dependent responses were investigated in isolated cannulated arterioles in the presence of constant intravascular pressure. We found that at 4 weeks of age, the systemic blood pressure of WKY and SHR was not significantly different, but at 12 weeks of age, systemic blood pressure was significantly elevated in SHR. Note that the tail-cuff plethysmography used to assess systemic blood pressures of rats in the present study is not highly accurate; thus, small differences or periodic increases in systemic blood pressure of animals may not have been detected. The basal and passive diameters of 4-week-old WKY and SHR were different, for which we do not have an explanation at present. Interestingly, previous studies showed no impairment in endothelial mediation of acetylcholine-induced responses in young SHR versus WKY of the same age and blood pressure, yet structural changes such as an increase in the media/lumen ratio is already detectable.20 Nevertheless, in the present study, the myogenic tone of vessels was not different, which is important because alteration in myogenic tone may affect the magnitude of flow-induced responses.
Flow-Induced Dilation at 4 Weeks of Age
At 4 weeks of age, in response to increases in perfusate flow, arterioles of WKY and SHR exhibited similar dilations, as indicated by the slope of the flow-diameter curves of SHR and WKY arterioles (Figure 1⇑). In both normotensive and hypertensive rats, inhibition of either NO or prostaglandin synthesis alone significantly reduced the dilation to flow. Combined application of these 2 inhibitors nearly completely eliminated flow-induced dilation of WKY and SHR arterioles. These findings demonstrate that at a prehypertensive age in arterioles of rat gracilis muscle, both NO and prostaglandins are involved in the endothelial mediation of dilation after increases in perfusate flow. This proportion seems to be ≈41% and ≈48%, respectively; these 2 endothelium-derived factors are responsible for the full mediation of the response. These findings also indicate that flow-induced NO and prostaglandin release are present in arterioles of young SHR. Others also have found NO-dependent vasodilation in young SHR.18 In agreement with these functional data, previous studies already demonstrated that the level of endothelial NO synthase (eNOS) protein was similar in the aorta of 4-week-old SHR and WKY.21
Flow-Induced Dilation at 12 Weeks of Age
In arterioles of 12-week-old WKY, we found that flow-dependent dilation is significantly enhanced compared with that in 4-week-old WKY and that this dilation is mediated by endothelium-derived NO and prostaglandins. In contrast, in 12-week-old hypertensive rats, the magnitude of flow-induced dilation is similar to what was observed at 4 weeks of age. Therefore, the dilation to increases in flow is reduced compared with 12-week-old normotensive rats. Inhibition of NO synthase by L-NNA did not reduce the response, whereas INDO treatment nearly completely eliminated the impaired flow-induced dilation6 in 12-week-old SHR. The findings suggest that increases in perfusate flow do not elicit an NO-mediated dilation of these arterioles but do stimulate the synthesis of prostaglandins that are responsible for the dilation in response to increases in flow in arterioles of hypertensive rats.
Thus, in normotensive rats, the synthesis of NO and prostaglandins increases with age and elicits augmented flow-induced dilation in older (12-week-old) rats. Previous findings showed that expression of eNOS is markedly increased in proliferating cultured bovine aortic endothelial cells, which suggests that similar events take place in normotensive WKY during development.22 The underlying reason for the enhanced appearance of eNOS could be the continuous presence of wall shear stress, which may also increase with the increase in blood pressure in WKY. In SHR, the greater increase in blood pressure (and, hence, wall shear stress) leads to the impairment of NO mediation, whereas the synthesis of prostaglandins is not affected. Indeed, during the development of hypertension, a decline in the activity and expression of eNOS has been shown in the rat aorta.21 In addition, a reduced release of NO was accompanied by depressed eNOS activity in thoracic aorta.23 Other studies showed that the decline of eNOS protein is accompanied by an increase of inducible NOS expression in Wistar rats and an increased plasma concentration of nitrate and nitrite.24 Contrary to these findings, the plasma concentration of serum nitrate/nitrite is reduced in individuals with essential hypertension,25 whereas Bonnardeaux et al26 showed no association of the eNOS gene with human essential hypertension. However, at present, we do not know whether alterations in wall shear stress are linked to NO release by inducible NOS and what the cellular origin of NO in the plasma in this condition may be.
Notably, studies measuring NO synthase and gene expression were done on endothelial cells either in tissue culture or in those isolated from large conduit vessels. These results may not be applicable to microvascular endothelial cells because they are likely to undergo a different process of adaptation. Our recent studies show that an acute increase in intraluminal pressure from 80 to 160 mm Hg (for 30 minutes) in isolated arterioles attenuates flow-dependent dilation as a result of enhanced production of superoxide12 27 interfering with NO. We can safely assume that in these vessels eNOS was present, because before pressure treatment, NO mediation was intact. Furthermore, application of superoxide dismutase and catalase, scavengers of reactive oxygen species, prevented the impairment, which suggests a primary role for hemodynamic forces in the impairment of the endothelial l-arginine pathway.
To reconcile some of the divergent results, one has to take into account the fact that the final physiological response, namely the dilation, depends not only on the presence of the specific gene, message, or protein, but also on a host of other factors, such as substrate availability, cofactors (tetrahydrobiopterin, Ca2+, and calmodulin), and levels of superoxide dismutase and superoxide.28 29 Nevertheless, it remains an intriguing question as to why in hypertension an impairment occurs in the signal transduction that links alterations in shear stress to NO release but not to prostaglandin release. The pathological consequences of inappropriate regulation of wall shear stress is that arterioles tend to promote increases in peripheral resistance and elevation of blood pressure.29 Higher blood pressure increases blood flow velocity, which in the presence of reduced vascular diameter further increases wall shear stress; this then can set up a pathological positive feedback mechanism. Regulation of wall shear stress at higher values not only would impose an extra burden on cardiac function but could further impair endothelial function.
In conclusion, the new findings of the present study are that in young normotensive and genetically hypertensive rats, shear stress–induced dilation of arterioles are present and are not different. This dilation increases with age in normotensive rats because of the increased release of NO and prostaglandins, whereas this dilation reduces with age in hypertensive rats as a result of an absence of NO mediation. Furthermore, the present findings suggest that in hypertension, elevated hemodynamic forces rather than genetic factors are primarily responsible for the development of impaired endothelial regulation of wall shear stress in skeletal muscle arterioles.
This work was supported by National Institutes of Health grants HL-46813 and HL-43023, AHA grant 9930244N, and AHA NY state affiliate grant 9830015T. We appreciate the excellent secretarial assistance of Mary Browne and Miriam Nunez.
- Received April 28, 1999.
- Revision received May 26, 1999.
- Accepted July 13, 1999.
Koller A, Kaley G. Endothelium regulates skeletal muscle microcirculation by a blood flow velocity sensing mechanism. Am J Physiol. 1990;258(pt 2):H916–H920.
Koller A, Dong S, Huang A, Kaley G. Corelease of nitric oxide and prostaglandins mediates flow-dependent dilation of rat gracilis muscle arterioles. Am J Physiol. 1994;267(pt 2):H326–H332.
Koller A, Kaley G. Endothelial regulation of wall shear stress and blood flow in skeletal muscle microcirculation. Am J Physiol. 1991;260(pt 2):H862–H868.
Hutchins PM, Darnell AE. Observation of a decreased number of small arterioles in spontaneously hypertensive rats. Circ Res. 1974;34:35(suppl I):I-161–I-165.
Prewitt RL, Chen II, Dowell R. Development of microvascular rarefaction in the spontaneously hypertensive rat. Am J Physiol. 1982;243(pt 2):H243–H251.
Koller A, Huang A. Impaired nitric oxide-mediated flow-induced dilation in arterioles of spontaneously hypertensive rats. Circ Res. 1994;74:416–421.
Koller A, Huang A. Shear stress-induced dilation is attenuated in skeletal muscle arterioles of hypertensive rats. Hypertension. 1995;25(pt 2):758–763.
Matrougui K, Jacques M, Lévy BI, Henrion D. Impaired nitric oxide- and prostaglandin-mediated responses to flow in resistance arteries of hypertensive rats. Hypertension. 1997;30:942–947.
Rizzoni D, Porteri E, Castellano M, Bettoni G, Muiesan ML, Tiberio G, Giulini SM, Rossi G, Bernini G, Agabiti-Rosei E. Endothelial dysfunction in hypertension is independent from the etiology and from vascular structure. Hypertension. 1998;31(pt 2):335—341.
Bund SJ, West KP, Heagerty AM. Effects of protection from pressure on resistance artery morphology and reactivity in spontaneously hypertensive and WKY rats. Circ Res. 1991;68:1230–1240.
Huang A, Sun D, Kaley G, Koller A. Superoxide released to high intra-arteriolar pressure reduces nitric oxide-mediated shear stress- and agonist-induced dilations. Circ Res. 1998;83:960–965.
Lockette WG, Otsuha Y, Carretero OA. The loss of endothelium-dependent vascular relaxation in hypertension. Hypertension. 1986;8(suppl II):61—66.
Tesfamariam B, Halpern W. Endothelium-dependent and endothelium-independent vasodilation in resistance arteries from hypertensive rats. Hypertension. 1988;11:440–444.
Boegehold MA. Reduced influence of nitric oxide on arteriolar tone in hypertensive Dahl rats. Hypertension. 1992;19:290–295.
Cardillo C, Kilcoyne CM, Quyyumi AA, Cannon RO, Panza JA. Selective defect in nitric oxide synthesis may explain the impaired endothelium-dependent vasodilation in patients with essential hypertension. Circulation. 1998;97:851–856.
Radaelli A, Mircoli L, Mori I, Mancia G, Ferrari AU. Nitric oxide-dependent vasodilation in young spontaneously hypertensive rats. Hypertension. 1998;32:735–739.
Sun D, Kaley G, Koller A. Characteristics and origin of the myogenic response in isolated gracilis muscle arterioles. Am J Physiol. 1994;266(pt 2):H1177–H1183.
Kong JQ, Taylor DA, Fleming WW. Mesenteric vascular responses of young spontaneously hypertensive rats. J Pharmacol Exp Ther. 1991;1:258:13–17.
Chou Tz-C, Yen M-H, Li C-Y, Ding Y-A. Alterations of nitric oxide synthase expression with aging and hypertension in rats. Hypertension. 1998;31:643–648.
Arnal J-F, Yamin J, Dockery S, Harrison DG. Regulation of endothelial nitric oxide synthase mRNA, protein, and activity during cell growth. Am J Physiol. 1994;267(pt 1)):C1381–C1388.
Cernadas MR, de Miguel LS, García-Duran M, González-Fernández F, Millás I, Montón M, Rodrigo J, Rico L, Fernández P, de Frutos T, Rodríguez-Feo JA, Guerra J, Caramelo C, Casado S, López-Farré A. Expression of constitutive and inducible nitric oxide synthases in the vascular wall of young and aging rats. Circ Res. 1998;83:279–286.
Node K, Kitakaze M, Yoshikawa H, Kosaka H, Hori M. Reduced plasma concentrations of nitrogen oxide in individuals with essential hypertension. Hypertension. 1997;30(pt1):405–408.
Bonnardeaux A, Nadaud S, Charru A, Jeunemaitre X, Corvol P, Soubrier F. Lack of evidence for linkage of the endothelial cell nitric oxide synthase gene to essential hypertension. Circulation. 1995;91:96–102.
Nakazono K, Watanabe N, Matsuno K, Sasaki J, Sato T, Inoue M. Does superoxide underlie the pathogenesis of hypertension? Proc Natl Acad Sci U S A. 1991;88:10045–10048.
Cosentino F, Katusic ZS. Tetrahydrobiopterin and dysfunction of endothelial nitric oxide synthase coronary arteries. Circulation. 1994;91:139–144.