Sex-specific differences in cerebral arterial myogenic tone in hypertensive and normotensive rats

Cerebral blood flow (CBF) is maintained constant despite changes in systemic blood pressure (BP) through multiple mechanisms of autoregulation such as vascular myogenic reactivity. Our aim was to determine myogenic characteristics of cannulated middle cerebral arteries (MCA) in male and female stroke-prone spontaneously hypertensive rats (SHRSP) and Wistar-Kyoto rats (WKY) at 12 wk of age under pressurised no-flow conditions. MCA pressure-diameter relationships (20-200 mmHg) were constructed in active (with calcium) and passive (without calcium) conditions, and myogenic and mechanical properties were determined. Myogenic reactivity in WKY (P < 0.05) and SHRSP (P < 0.05) males was impaired compared with their female counterparts. Comparison of SHRSP with WKY in males revealed similar myogenic reactivity, but in females SHRSP exhibited augmented myogenic reactivity (P < 0.05). In both sexes, myogenic tone yielded at lower pressure in SHRSP compared with WKY vessels (120-140 vs. 140-180 mmHg). Stress-strain relationships and elastic moduli in WKY rats showed that vessels were stiffer in females than in males. Conversely, in SHRSP, male vessels were stiffer than female vessels. Comparison of strains in males indicated that stiffness was increased in SHRSP compared with WKY vessels, whereas the converse was observed in females. These findings demonstrate that MCA myogenic and distensibility characteristics exhibit significant sex- and strain-dependent differences. Inappropriate myogenic adaptation and augmented vascular stiffness, particularly in male SHRSP, are potential limiting factors in blood flow autoregulation and may increase the predisposition for stroke-related cerebrovascular events.     

Transient middle cerebral artery occlusion causes different structural, mechanical, and myogenic alterations in normotensive and hypertensive rats

Transient focal cerebral ischemia in the rat alters vessel properties, and spontaneously hypertensive rats (SHR) show a poorer outcome after ischemia. In the present study we examined the role of hypertension on vessel properties after ischemia-reperfusion. The right middle cerebral artery (MCA) was occluded (90 min) and reperfused (24 h) in SHR (n = 12) and Wistar-Kyoto rats (WKY; n = 11). Sham-operated rats (SHR, n = 10; WKY, n = 10) were used as controls. The structural, mechanical, and myogenic properties of the MCA were assessed by pressure myography. Nuclei distribution and elastin content and organization were analyzed by confocal microscopy. Infarct volume was larger in SHR than in WKY rats. Ischemia-reperfusion induced adventitial hypertrophy associated with an increase in the total number of adventitial cells. In addition, fenestrae area and arterial distensibility increased and myogenic tone decreased in the MCA of WKY rats after ischemia-reperfusion. Hypertension per se induced hypertrophic inward remodeling. Ischemia-reperfusion decreased the cross-sectional area of the MCA in SHR, without significant changes in distensibility, despite an increase in fenestrae area. In addition, MCA myogenic properties were not altered after ischemia-reperfusion in SHR. Our results indicate that in normotensive rats, MCA develops a compensatory mechanism (i.e., enhanced distensibility and decreased myogenic tone) that counteracts the effect of ischemia-reperfusion and ensures correct cerebral irrigation. These compensatory mechanisms are lost in hypertension, thereby explaining, at least in part, the greater infarct volume observed in SHR.     

Intact female stroke-prone hypertensive rats lack responsiveness to mineralocorticoid receptor antagonists

Data from the Framingham Heart Study suggest that women may be more sensitive to the deleterious cardiovascular remodeling effects of aldosterone. Previous studies from our laboratory have shown that chronic treatment with spironolactone, a mineralocorticoid receptor (MR) antagonist, decreases ischemic cerebral infarct size and prevents remodeling of the middle cerebral artery (MCA) in male spontaneously hypertensive stroke-prone rats (SHRSP). Therefore, we hypothesized that MR antagonism would reduce ischemic infarct size and prevent MCA remodeling in female SHRSP. Six-week-old female SHRSP were treated for 6 wk with spironolactone (25 or 50 mg.kg(-1).day(-1)) or eplerenone (100 mg.kg(-1).day(-1)) and compared with untreated controls. At 12 wk, cerebral ischemia was induced for 18 h using the intraluminal suture occlusion technique, or the MCA was isolated for analysis of passive structure using a pressurized arteriograph. MR antagonism had no effect on infarct size or passive MCA structure in female SHRSP. To study the potential effects of estrogen, the above experiments were repeated in bilaterally ovariectomized (OVX) female SHRSP treated with spironolactone (25 mg.kg(-1).day(-1)). Infarct size and vessel structure in OVX SHRSP were not different from control SHRSP. Spironolactone had no effect on infarct size in OVX SHRSP. However, MCA lumen and outer diameters were increased in spironolactone-treated OVX SHRSP, suggesting an effect of estrogen. Cerebral artery MR expression, assessed by Western blotting, was increased in female, compared with male, SHRSP. These studies highlight an apparent sexual dimorphism of MR expression and activity in the cerebral vasculature from hypertensive rats.     

Doxycycline, a matrix metalloprotease inhibitor, reduces vascular remodeling and damage after cerebral ischemia in stroke-prone spontaneously hypertensive rats

Matrix metalloproteases (MMPs) are a family of zinc peptidases involved in extracellular matrix turnover. There is evidence that increased MMP activity is involved in remodeling of resistance vessels in chronic hypertension. Thus we hypothesized that inhibition of MMP activity with doxycycline (DOX) would attenuate vascular remodeling. Six-week-old male stroke-prone spontaneously hypertensive rats (SHRSP) were treated with DOX (50 mg·kg(-1)·day(-1) in the drinking water) for 6 wk. Untreated SHRSP were controls. Blood pressure was measured by telemetry during the last week. Middle cerebral artery (MCA) and mesenteric resistance artery (MRA) passive structures were assessed by pressure myography. MMP-2 expression in aortas was measured by Western blot. All results are means ± SE. DOX caused a small increase in mean arterial pressure (SHRSP, 154 ± 1; SHRSP + DOX, 159 ± 3 mmHg; P < 0.001). Active MMP-2 expression was reduced in aorta from SHRSP + DOX (0.21 ± 0.06 vs. 0.49 ± 0.13 arbitrary units; P < 0.05). In the MCA, at 80 mmHg, DOX treatment increased the lumen (273.2 ± 4.7 vs. 238.3 ± 6.3 μm; P < 0.05) and the outer diameter (321 ± 5.3 vs. 290 ± 7.6 μm; P < 0.05) and reduced the wall-to-lumen ratio (0.09 ± 0.002 vs. 0.11 ± 0.003; P < 0.05). Damage after transient cerebral ischemia (transient MCA occlusion) was reduced in SHRSP + DOX (20.7 ± 4 vs. 45.5 ± 5% of hemisphere infarcted; P < 0.05). In the MRA, at 90 mmHg DOX, reduced wall thickness (29 ± 1 vs. 22 ± 1 μm; P < 0.001) and wall-to-lumen ratio (0.08 ± 0.004 vs. 0.11 ± 0.008; P < 0.05) without changing lumen diameter. These results suggest that MMPs are involved in hypertensive vascular remodeling in both the peripheral and cerebral vasculature and that DOX reduced brain damage after cerebral ischemia.     

Increased sympathetic innervation in the cerebral and mesenteric arteries of hypertensive rats

The density of catecholamine-containing nerve fibers was studied in the cerebral and mesenteric arteries from normotensive Wistar-Kyoto rats (WKY), spontaneously hypertensive rats (SHR), and stroke-prone SHR (SHRSP) in the growing (SHR, WKY) and adult (SHR, SHRSP, WKY) animals. Cerebral arteries from SHR showed an increased adrenergic innervation from day 1. The nerve plexuses reached an adult pattern earlier in SHR than in WKY. The arteries from adult SHR and SHRSP (22 weeks old) showed a markedly higher nerve density than WKY. There was a positive linear correlation between blood pressure and nerve density for four cerebral arteries. The mesenteric arteries were not innervated at birth. However, hyperinnervation of these arteries in the SHR was already present at 10 days of age as compared with WKY. Sympathectomy with anti-nerve growth factor and guanethidine caused a complete disappearance of fluorescent fibers in the mesenteric arteries from SHR and WKY, and in the cerebral arteries of WKY. The same procedure caused only partial denervation of the cerebral arteries from hypertensive animals. We postulate that the increase in nerve density in the cerebral arteries from the hypertensive rats may contribute to the development of arterial hypertrophy in chronic hypertension through the trophic effect of the sympathetic innervation on vascular structure.     

Cerebral artery responses to pressure and flow in uremic hypertensive and spontaneously hypertensive rats

Impaired cerebral blood flow autoregulation is seen in uremic hypertension, whereas in nonuremic hypertension autoregulation is shifted toward higher perfusion pressure. The cerebral artery constricts in response to a rise in either lumen pressure or flow; we examined these responses in isolated middle cerebral artery segments from uremic Wistar-Kyoto rats (WKYU), normotensive control rats (WKYC), and spontaneously hypertensive rats (SHR). Pressure-induced (myogenic) constriction developed at 100 mmHg; lumen flow was then increased in steps from 0 to 98 microl/min. Some vessels were studied after endothelium ablation. Myogenic constriction was significantly lower in WKYU (28 +/- 2.9%) compared with both WKYC (39 +/- 2.5%, P = 0.035) and SHR (40 +/- 3.1%, P = 0.018). Flow caused constriction of arteries from all groups in an endothelium-independent manner. The response to flow was similar in WKYU and WKYC, whereas SHR displayed increased constriction compared with WKYU (P < 0.001) and WKYC (P < 0.001). We conclude that cerebral myogenic constriction is decreased in WKYU, whereas flow-induced constriction is enhanced in SHR.     

Impact of Short-Term Treatment with Telmisartan on Cerebral Arterial Remodeling in SHR

Background and Purpose

Chronic hypertension decreases internal diameter of cerebral arteries and arterioles. We recently showed that short-term treatment with the angiotensin II receptor blocker telmisartan restored baseline internal diameter of small cerebral arterioles in spontaneously hypertensive rats (SHR), via reversal of structural remodeling and inhibition of the angiotensin II vasoconstrictor response. As larger arteries also participate in the regulation of cerebral circulation, we evaluated whether similar short-term treatment affects middle cerebral arteries of SHR.

Methods

Baseline internal diameters of pressurised middle cerebral arteries from SHR and their respective controls, Wistar Kyoto rats (WKY) and responses to angiotensin II were studied in a small vessel arteriograph. Pressure myogenic curves and passive internal diameters were measured following EDTA deactivation, and elastic modulus from stress-strain relationships.

Results

Active baseline internal diameter was 23% lower in SHR compared to WKY, passive internal diameter (EDTA) 28% lower and elastic modulus unchanged. Pressure myogenic curves were shifted to higher pressure values in SHR. Telmisartan lowered blood pressure but had no effect on baseline internal diameter nor on structural remodeling (passive internal diameter and elastic modulus remained unchanged compared to SHR). Telmisartan shifted the pressure myogenic curve to lower pressure values than SHR.

Conclusion

In the middle cerebral arteries of SHR, short-term treatment with telmisartan had no effect on structural remodeling and did not restore baseline internal diameter, but allowed myogenic tone to adapt towards lower pressure values.     

A high-potassium diet reduces infarct size and improves vascular structure in hypertensive rats

High-potassium diets can improve vascular function, yet the effects of potassium supplementation on ischemic stroke have not been studied. We hypothesized that dietary potassium supplementation would reduce ischemic cerebral infarct size by reversing cerebral artery hypertrophy. Six-week-old male stroke-prone spontaneously hypertensive rats (SHRSP) were fed diets containing 0.79% potassium (LK) or 2.11% potassium (HK) for 6 wk; Wistar-Kyoto (WKY) rats were fed the LK diet. The HK diet did not reduce blood pressure, as measured by telemetry, in the SHRSP. Cerebral ischemia was induced by middle cerebral artery (MCA) occlusion. The resultant infarct was smaller in the HK-SHRSP than in the LK-SHRSP: 55.1 +/- 6.3 vs. 71.4 +/- 2.4% of the hemisphere infarcted (P < 0.05). Infarcts were smaller in WKY rats (33.5 +/- 4.8%) than in LK-SHRSP or HK-SHRSP. The vessel wall of MCAs from LK-SHRSP was hypertrophied compared with WKY rats; this was reversed in HK-SHRSP. RT-PCR analysis of the cerebral vessels showed that expression of platelet-derived growth factor receptors-alpha and -beta, epidermal growth factor receptor, and collagen I and III was increased in the vessels from LK-SHRSP compared with WKY rats and reduced in HK-SHRSP. These results suggest that potassium supplementation provides neuroprotection in a model of ischemic stroke independent of blood pressure and possibly through changes in vascular structure.     

Relative contribution of Rho kinase and protein kinase C to myogenic tone in rat cerebral arteries in hypertension

Arterial smooth muscle constriction in response to pressure, i.e., myogenic tone, may involve calcium-dependent and calcium-sensitization mechanisms. Calcium sensitization in vascular smooth muscle is regulated by kinases such as PKC and Rho kinase, and activity of these kinases is known to be altered in cardiovascular disorders. In the present study, we evaluated the relative contribution of PKC and Rho kinase to myogenic tone in cerebral arteries in hypertension. Myogenic tone and arterial wall calcium in Wistar-Kyoto (WKY) rats and spontaneously hypertensive rats (SHR) were measured simultaneously, and the effect of PKC and Rho kinase inhibitors on myogenic tone was evaluated. SHR arteries showed significantly greater myogenic tone than WKY arteries. Pressure/wall tension-arterial wall calcium curves showed a hyperbolic relation in WKY rats, but the curves for SHR arteries were parabolic. Myogenic tone was decreased by the Rho kinase inhibitors Y-27632 and HA-1077, with a significantly greater effect in SHR than in WKY arteries. Reduction in myogenic tone produced by the PKC inhibitor bisindolylmaleimide I in WKY and SHR arteries was significantly less than that produced by Rho kinase inhibition. The pressure-dependent increase in myogenic tone was significantly decreased by Y-27632, and the decrease was markedly greater than that produced by bisindolylmaleimide I in SHR arteries. In WKY arteries, the pressure-dependent increase in myogenic tone was decreased to a similar extent by Y-27632 and bisindolylmaleimide I. These results suggest greater myogenic tone with increased calcium sensitization in SHR arteries, largely because of Rho kinase activation, with a minor contribution of PKC activation.     

Elevated production of 20-HETE in the cerebral vasculature contributes to severity of ischemic stroke and oxidative stress in spontaneously hypertensive rats

Hypertension is a major risk factor for stroke, but the factors that contribute to the increased incidence and severity of ischemic stroke in hypertension remain to be determined. 20-hydroxyeicosatetraenoic acid (20-HETE) has been reported to be a potent constrictor of cerebral arteries, and inhibitors of 20-HETE formation reduce infarct size following cerebral ischemia. The present study examined whether elevated production of 20-HETE in the cerebral vasculature could contribute to the larger infarct size previously reported after transient middle cerebral artery occlusion (MCAO) in hypertensive strains of rat [spontaneously hypertensive rat (SHR) and spontaneously hypertensive stroke-prone rat (SHRSP)]. The synthesis of 20-HETE in the cerebral vasculature of SHRSP measured by liquid chromatography-tandem mass spectrometry was about twice that seen in Wistar-Kyoto (WKY) rats. This was associated with the elevated expression of cytochrome P-450 (CYP)4A protein and CYP4A1 and CYP4A8 mRNA. Infarct volume after transient MCAO was greater in SHRSP (36+/-4% of hemisphere volume) than in SHR (19+/-5%) or WKY rats (5+/-2%). This was associated with a significantly greater reduction in regional cerebral blood flow (rCBF) in SHR and SHRSP than in WKY rats during the ischemic period (78% vs. 62%). In WKY rats, rCBF returned to 75% of control following reperfusion. In contrast, SHR and SHRSP exhibited a large (166+/-18% of baseline) and sustained (1 h) postischemic hyperperfusion. Acute blockade of the synthesis of 20-HETE with N-hydroxy-N'-(4-butyl-2-methylphenyl)-formamidine (HET0016; 1 mg/kg) reduced infarct size by 59% in SHR and 87% in SHRSP. HET0016 had no effect on the fall in rCBF during MCAO but eliminated the hyperemic response. HET0016 also attenuated vascular O2*- formation and restored endothelium-dependent dilation in cerebral arteries of SHRSP. These results indicate the production of 20-HETE is elevated in the cerebral vasculature of SHRSP and contributes to oxidative stress, endothelial dysfunction, and the enhanced sensitivity to ischemic stroke in this hypertensive model.     

Nicardipine normalizes elevated levels of antioxidant activity in response to xanthine oxidase-induced oxidative stress in hypertensive rat heart

It has been reported that the production of oxygen radicals mediated by xanthine oxidase (XO) is stimulated in hypertensive cardiovascular endothelium, suggesting involvement of oxidative stress in pathogenesis of hypertension. In this study we estimated the effect of nicardipine, a calcium blocker, on the oxidative stress and antioxidant activities in left ventricles from spontaneously hypertensive rat (SHR) and stroke-prone SHR (SHRSP). The activity of XO increased 3.5-fold in SHR and 6.2-fold in SHRSP compared to that in normal controls (WKY). Interestingly, the levels of glutathione (GSH) and the activity of its synthesizing enzyme (γ-glutamylcysteine synthetase, γ-GCS) elevated concomitantly in SHR and SHRSP: the level of GSH increased 1.2-fold in SHR and 1.3-fold in SHRSP. The activity of γ-GCS was elevated 1.5-fold in SHR and 2.4-fold in SHRSP, accompanying an increase in the expression of its mRNA. Treatment of these rats with nicardipine, for 4 weeks improved blood pressure, from 176 ± 10 to 140 ± 8 mmHg in SHR, and from 201 ± 11 to 167 ± 5 mmHg in SHRSP, respectively, and decreased wet weight of heart, levels of GSH, and the activities of XO and γ-GCS. Nicardipine reduced the expression of γ-GCS mRNA. Collectively, these results suggest that reactive oxygen species produced by XO in hypertensive rat heart cause induction of the expression of γ-GCS and nicardipine plays a role in reducing the oxidative stress in hypertensive heart.     

Effects of ischemia and myogenic activity on active and passive mechanical properties of rat cerebral arteries

Passive (papaverine induced) and active (spontaneous pressure induced) biomechanical properties of ischemic and nonischemic rat middle cerebral arteries (MCAs) were studied under pressurized conditions in vitro. Ischemic (1 h of occlusion), contralateral, and sham-operated control MCAs were isolated from male Wistar rats (n = 22) and pressurized using an arteriograph system that allowed control of transmural pressure (TMP) and measurement of lumen diameter and wall thickness. Three mechanical stiffness parameters were computed: overall passive stiffness (beta), pressure-dependent modulus changes (E(inc,p)), and smooth muscle cell (SMC) activity-dependent changes (E(inc,a)). The beta-value for ischemic vessels was increased compared with sham vessels (13.9 +/- 1.7 vs. 9.1 +/- 1.4, P < 0.05), indicating possible short-term remodeling due to ischemia. E(inc,p) increased with pressure in the passive vessels (P < 0.05) but remained relatively constant in the active vessels for all vessel types, indicating that pressure-induced SMC contractile activity (i.e., myogenic reactivity) in cerebral arteries leads to the maintenance of a constant elastic modulus within the autoregulatory pressure range. E(inc,a) increased with pressure for all conditions, signifying that changes in stiffness are influenced by SMC activity and vascular tone.     

Tension-velocity relationships in hypertensive mesenteric resistance arteries

Increased total peripheral resistance is the cardinal haemodynamic disorder in essential hypertension. This could be secondary to alterations in the mechanical properties of vascular smooth muscle. Adequate study has not been made of the tension-velocity (T-V) relationship in hypertensive resistance arterial smooth muscle. Increased narrowing in such arteries would result in increased resistance. The objectives of this investigation were to determine whether there is (i) increased narrowing capacity (-delta C/C omicron, where C stands for arterial internal circumference and C omicron is the optimal arterial internal circumference for maximum tension development); (ii) an increased maximum velocity of isobaric narrowing (Vmax) measured in C omicron per second; (iii) an increased wall thickness (h); and (iv) an increased active stress development (Tmax) in the spontaneously hypertensive rat (SHR; n = 5) compared with the normotensive Wistar Kyoto (WKY; n = 5) and MK-421 (an angiotensin I converting enzyme inhibitor) treated spontaneously hypertensive rat (MK-421 trt. SHR; n = 5) mesenteric resistance (diameter, less than 300 micron) arteries. Analysis of the data for arteries constricting isobarically against a range of pressures revealed that (a) the SHR -deltaC/C omicron values at pressures ranging from 20 to 120 mmHg (1 mmHg = 133.322 Pa) showed significantly increased narrowing compared with the MK-421 trt. SHR and WKY -deltaC/C omicron values in this same pressure range (p less than 0.01), and (b) the SHR derived Vmax of 0.83 +/- 0.08 C omicron/s was significantly faster than either the MK-421 trt.(ABSTRACT TRUNCATED AT 250 WORDS)     

Respective contribution of age, mean arterial pressure, and body weight on central arterial distensibility in SHR

In spontaneously hypertensive rats (SHR), carotid and aortic distensibilities measured at operational blood pressure (BP) are reduced. Increased body weight and mean arterial pressure (MAP) are both known to reduce distensibility independently. However, whether, after adjustment to body weight and mean BP, distensibility remains reduced in SHR has never been investigated. Carotid and abdominal aorta distensibilities were measured under anesthesia in SHR at 5, 12, 52, and 78 wk of age, and measurements were compared with age-matched normotensive Wistar rats. Each age group was composed of 9 or 10 animals. We determined distensibility using echo-tracking techniques of high resolution. Compared with Wistar rats, carotid and aortic distensibilities measured at operational MAP are reduced in SHR. This reduction is accentuated with age, particularly for the carotid artery. After adjustment to body weight and MAP, carotid and aortic distensibilities become identical in Wistar and SHR (or even slightly increased in SHR) but continue to be reduced with age, mainly for the carotid artery. In conclusion, in SHR, age and high BP do not have a parallel and similar influence on the reduction of arterial distensibility. Aging constantly reduces arterial distensibility, whereas MAP levels contribute to maintenance of arterial function.     

Myogenic response of rat femoral small arteries in relation to wall structure and [Ca(2+)](i)

The present study investigated the influence of media thickness on myogenic tone and intracellular calcium concentration ([Ca(2+)](i)) in rat skeletal muscle small arteries. A ligature was loosely tied around one external iliac artery of 5-wk-old spontaneously hypertensive rats. At 18 wk of age, femoral artery blood pressure was 102 +/- 11 mmHg (n = 15) on the ligated side and 164 +/- 6 mmHg (n = 15) on the contralateral side. Small arteries feeding the gracilis muscle had a reduced media cross-sectional area and a reduced media-to-lumen ratio on the ligated side, where also the range of myogenic constriction was shifted to lower pressures. However, when expressed as a function of wall stress, diameter responses were nearly identical. [Ca(2+)](i) was higher in vessels from the ligated hindlimb at pressures above 10 mmHg, but vasoconstriction was not accompanied by changes in [Ca(2+)](i). Thus the myogenic constriction here seems due primarily to changes in intracellular calcium sensitivity, which are determined mainly by the force per cross-sectional area of the wall and therefore altered by changes in vascular structure.     

Atrial natriuretic polypeptide (ANP) in the development of spontaneously hypertensive rats (SHR) and stroke-prone SHR (SHRSP)

In order to investigate the pathophysiological role of atrial natriuretic polypeptide (ANP) in genetic hypertensive rats, the atrial content and plasma concentration of ANP were measured by a sensitive radioimmunoassay (RIA) for rat ANP in 5-, 10- and 20-week-old spontaneously hypertensive rats (SHR) and stroke-prone SHR (SHRSP) and compared to age-matched Wistar Kyoto rats (WKY). Atrial content of immunoreactive ANP (ir-ANP) tended to be higher in SHR and was already significantly higher in SHRSP than in WKY at 5 weeks of age. Atrial content in the hypertensive strains became significantly higher than in WKY when hypertension developed at 10 and 20 weeks. On the other hand, plasma ir-ANP in SHR was significantly lower than in WKY at 5 weeks, however, it became significantly higher in both SHR and SHRSP than in WKY at 10 and 20 weeks. These findings suggest that ANP release may increase to compensate for the elevation of blood pressure in SHR and SHRSP and that biosynthesis of ANP may be concomitantly stimulated, resulting in an increase in atrial ANP.     

Mechanics and Composition of Middle Cerebral Arteries from Simulated Microgravity Rats with and without 1-h/d –Gx Gravitation

Background

To elucidate further from the biomechanical aspect whether microgravity-induced cerebral vascular mal-adaptation might be a contributing factor to postflight orthostatic intolerance and the underlying mechanism accounting for the potential effectiveness of intermittent artificial gravity (IAG) in preventing this adverse effect.

Methodology/Principal Findings

Middle cerebral arteries (MCAs) were isolated from 28-day SUS (tail-suspended, head-down tilt rats to simulate microgravity effect), S+D (SUS plus 1-h/d −G_x gravitation by normal standing to simulate IAG), and CON (control) rats. Vascular myogenic reactivity and circumferential stress-strain and axial force-pressure relationships and overall stiffness were examined using pressure arteriography and calculated. Acellular matrix components were quantified by electron microscopy. The results demonstrate that myogenic reactivity is susceptible to previous pressure-induced, serial constrictions. During the first-run of pressure increments, active MCAs from SUS rats can strongly stiffen their wall and maintain the vessels at very low strains, which can be prevented by the simulated IAG countermeasure. The strains are 0.03 and 0.14 respectively for SUS and S+D, while circumferential stress being kept at 0.5 (10⁶ dyn/cm²). During the second-run pressure steps, both the myogenic reactivity and active stiffness of the three groups declined. The distensibility of passive MCAs from S+D is significantly higher than CON and SUS, which may help to attenuate the vasodilatation impairment at low levels of pressure. Collagen and elastin percentages were increased and decreased, respectively, in MCAs from SUS and S+D as compared with CON; however, elastin was higher in S+D than SUS rats.

Conclusions

Susceptibility to previous myogenic constrictions seems to be a self-limiting protective mechanism in cerebral small resistance arteries to prevent undue cerebral vasoconstriction during orthostasis at 1-G environment. Alleviating of active stiffening and increasing of distensibility of cerebral resistance arteries may underlie the countermeasure effectiveness of IAG.     

Free and bound forms of atrial natriuretic peptide (ANP) in rat plasma: preferential increase of free ANP in spontaneously hypertensive rats (SHR) and stroke-prone SHR (SHRSP)

Free and bound forms of atrial natriuretic peptide (ANP) in rat plasma were analysed by gel permeation chromatography combined with a radioimmunoassay (RIA) for rat ANP (rANP). Gel permeation chromatography showed two immunoreactive peaks in rat plasma, one corresponding to alpha-rANP, rANP(99-126), and the other eluted at a high molecular weight, clearly different from gamma-rANP, rANP(1-126). The chromatographic profile of rat plasma after incubation with synthetic alpha-rANP demonstrated that the high molecular immunoreactivity had ANP-binding capacity. This bound form of ANP was almost totally excluded following extraction procedure, therefore, the immunoreactive ANP (ir-ANP) measured with the extraction assay was mainly free ANP. On the other hand, direct RIA may detect not only the free but also the bound form of ANP. Using both direct RIA and the extraction method, bound forms of plasma ANP in spontaneously hypertensive rats (SHR) and stroke-prone SHR (SHRSP) were compared to normotensive Wistar Kyoto rats (WKY). Bound forms of plasma ANP in 20-week-old SHR and SHRSP were significantly higher than that in age-matched WKY. The ratio of free/bound form of plasma ANP in SHR and SHRSP also significantly increased compared to WKY, indicating a preferential increase in free ANP in the plasma of these hypertensive rats. These findings suggest that a bound form of ANP may be present in rat plasma and that it may play some pathophysiological role in the hypertension of SHR and SHRSP. Increased free ANP in plasma may indicate a compensatory increase in ANP release in these hypertensive rats.     

Increased medial smooth muscle cell length is responsible for vascular hypertrophy in young hypertensive rats

Large mesenteric arteries from 3- to 4-wk-old spontaneously hypertensive rats (SHR) showed medial hypertrophy and an increased contractile response to various agonists before significant blood pressure increase. Here we determined the cellular nature of this vascular hypertrophy. Large mesenteric arteries from SHR and Wistar-Kyoto (WKY) rats were fixed at maximal relaxation either with an in situ perfusion fixation or an in vitro fixation method. With the use of morphometric protocols and confocal microscopy, the volume of the medial wall and lumen, numerical density of smooth muscle cell nuclei in the medial layer, and smooth muscle cell and nuclear length were measured. Both methods of fixation yielded similar results, showing significant medial volume expansion in SHR than WKY without lumen change. Numerical density of medial smooth muscle cells was significantly less in SHR than WKY, and their total number per 100 microm length were similar between the strains. Average smooth muscle nuclear and cell length from SHR was significantly longer than that of WKY. Regression analysis showed that the increase in smooth muscle cell length explained 80% of the medial volume increase. We concluded that increased smooth muscle cell length in prehypertensive SHR is responsible for increased medial volume in the mesenteric arteries.     

Aortic stiffness in vivo in hypertensive rat via echo-tracking: analysis of the pulsatile distension waveform

Large-artery stiffening is a major risk factor in aging and hypertension. Elevated blood pressure (BP) and vascular wall properties participate in arterial stiffening; we aimed to evaluate their respective role by combining echo-tracking and the spontaneously hypertensive rats (SHR) treated with low doses of a nitric oxide synthase inhibitor, shown to have arterial stiffening. Normotensive [Wistar-Kyoto (WKY)], SHR, and SHR treated for 2 wk with N(G)-nitro-L-arginine methyl ester (SHRLN) were anesthetized; BP and distension (pulsatile displacement) of the aortic walls with the ArtLab echo-tracking device were measured. Stiffness index increased in SHRLN vs. SHR; compliance, distensibility, and the slopes and area of the distension-pressure loop curve decreased. The pulsatile distension and pressure waveforms were strongly altered in SHRLN. Maximal values were decreased and increased, respectively, and the waveform kinetics also differed. Thus the area under the curve adjusted to heart rate (AUC/ms) was calculated. Acute BP reductions were induced by diltiazem in SHR and SHRLN, to levels similar to those of WKY. In SHR, compliance, distensibility, stiffness index, and the ascending slope of the distension-pressure loop reached the values of WKY, whereas they were only partially improved in SHRLN. Aortic distension (maximal value and AUC/ms) and the area of the distension-pressure loop were improved in SHR, but not in SHRLN. These data confirm the aortic stiffening induced by nitric oxide reduction in SHR. They show that the ArtLab system analyzes aortic stiffness in rats, and that the aortic pulsatile distension waveform is a parameter strongly dependent on the vascular wall properties.