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Cerebral blood flow and perfusion are tightly maintained through autoregulation despite changes in transmural pressure. Oxidative stress impairs cerebral blood flow, precipitating cerebrovascular events. Phosphorylation of the adaptor protein p66Shc increases mitochondrial-derived oxidative stress. The effect of p66Shc gain or loss of function in nonhypertensive rats is unclear. We hypothesized that p66Shc gain of function would impair autoregulation of cerebral microcirculation under physiological and pathological conditions. Three previously established transgenic [salt-sensitive (SS) background] p66Shc rats were used, p66-Del/SS (express p66Shc with a nine-amino acid deletion), p66Shc-knockout (KO)/SS (frameshift premature termination codon), and p66Shc signaling and knock-in substitution of Ser36Ala (p66Shc-S36A)/SS (substitution of Ser36Ala). The p66Shc-Del were also bred on Sprague-Dawley (SD) backgrounds (p66-Del/SD), and a subset was exposed to a hypertensive stimulus [ N G -nitro-l-arginine methyl ester (l-NAME)] for 4 wk. Active and passive diameters to increasing transmural pressure were measured and myogenic tone was calculated in all groups (SS and SD). Myogenic responses to increasing pressure were impaired in p66Shc-Del/SS rats relative to wild-type (WT)/SS and knock-in substitution of Ser36Ala (S36A; P < 0.05). p66-Del/SD rats did not demonstrate changes in active/passive diameters or myogenic tone relative to WT/SD but did demonstrate attenuated passive diameter responses to higher transmural pressure relative to p66-Del/SS. Four weeks of a hypertensive stimulus (l-NAME) did not alter active or passive diameter responses to increasing transmural pressure ( P = 0.86-0.99), but increased myogenic responses relative to p66-Del/SD ( P < 0.05). Collectively, we demonstrate the functional impact of p66Shc within the cerebral circulation and demonstrate that the genetic background of p66Shc rats largely drives changes in cerebrovascular function. NEW & NOTEWORTHY We demonstrate that the modulation of p66Shc signaling impairs cerebral artery myogenic tone in a low renin model of hypertension. This impairment is dependent upon the genetic background, as modulated p66Shc signaling in Sprague-Dawley rats does not impair cerebral artery myogenic tone.

Aims: Acute migraine attack in familial hemiplegic migraine type 2 (FHM2) patients is characterized by sequential hypo- and hyperperfusion. FHM2 is associated with mutations in the Na, K-ATPase α2 isoform. Heterozygous mice bearing one of these mutations (α2+/G301R mice) were shown to have elevated cerebrovascular tone and, thus, hypoperfusion that might lead to elevated concentrations of local metabolites. We hypothesize that these α2+/G301R mice also have increased cerebrovascular hyperaemic responses to these local metabolites leading to hyperperfusion in the affected part of the brain., Methods and Results: Neurovascular coupling was compared in α2+/G301R and matching wild-type (WT) mice using Laser Speckle Contrast Imaging. In brain slices, parenchymal arteriole diameter and intracellular calcium changes in neuronal tissue, astrocytic endfeet, and smooth muscle cells in response to neuronal excitation were assessed. Wall tension and smooth muscle membrane potential were measured in isolated middle cerebral arteries. Quantitative polymerase chain reaction, western blot, and immunohistochemistry were used to assess the molecular background underlying the functional changes. Whisker stimulation induced larger increase in blood perfusion, i.e. hyperaemic response, of the somatosensory cortex of α2+/G301R than WT mice. Neuronal excitation was associated with larger parenchymal arteriole dilation in brain slices from α2+/G301R than WT mice. These hyperaemic responses in vivo and ex vivo were inhibited by BaCl2, suggesting involvement of inward-rectifying K+ channels (Kir). Relaxation to elevated bath K+ was larger in arteries from α2+/G301R compared to WT mice. This difference was endothelium-dependent. Endothelial Kir2.1 channel expression was higher in arteries from α2+/G301R mice. No sex difference in functional responses and Kir2.1 expression was found., Conclusion: This study suggests that an abnormally high cerebrovascular hyperaemic response in α2+/G301R mice is a result of increased endothelial Kir2.1 channel expression. This may be initiated by vasospasm-induced accumulation of local metabolites and underlie the hyperperfusion seen in FHM2 patients during migraine attack., (Published on behalf of the European Society of Cardiology. All rights reserved. © The Author(s) 2019. For permissions, please email: journals.permissions@oup.com.)

Objectives Intracranial atherosclerotic stenosis (ICAS) is one of the most common causes of stroke worldwide. We adapted a rat model of atherosclerosis to study brain intracranial atherosclerosis, and further investigated how omega-3 fatty acids (O3FA) attenuated the development of ICAS by reducing the generation of reactive oxygen species (ROS) and the activation of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (NOX) activity. Methods Adult male Sprague-Dawley rats were divided into control normal-cholesterol or high-cholesterol diet groups with or without O3FA for up to 6 weeks. NG-nitro-L-arginine methyl ester (L-NAME, 3 mg/mL), a nitric oxide synthase inhibitor, was added to the drinking water of the high-cholesterol groups during the first 2 weeks. The rats received supplementation with O3FA (5 mg/kg/day) by gavage. At 3 and 6 weeks, we measured blood lipid levels, including low-density lipoprotein (LDL), cholesterol (CHO), triglycerides (TG), and high-density lipoprotein (HDL) as atherosclerotic blood markers. The lumen of middle cerebral artery (MCA) and the thickness of the vessel wall were assessed histologically. ROS production was measured. NOX activity and mRNA and protein expression of NOX subunits (p47 phox , gp91 phox , p22 phox , and p67 phox ) were measured. Results A high-cholesterol diet exhibited a significant increase in the classic blood markers (LDL, CHO, and TG) for atherosclerosis, as well as a decrease in HDL. These markers were found to be progressively more severe with time. Additionally, increased lumen stenosis and intimal thickening were observed in the MCA for this group. Rats given O3FA demonstrated attenuation of blood lipid levels with an absence of morphological changes.O3FA significantly reduced ROS production and NOX activity in the brain. Moreover, O3FA decreased the mRNA and protein expression of the NOX subunits p47 phox , gp91 phox , and p67 phox . Conclusions Long-term O3FA dietary supplementation prevents the development of intracranial atherosclerosis. This O3FA effect appears to be mediated by its attenuation of NOX subunit expression and NOX activity, therefore reducing ROS production. O3FA dietary supplement shows promising results in the prevention of ICAS.

Cerebrovascular CO 2 reactivity is affected by nitric oxide (NO). We tested the hypothesis that sildenafil selectively potentiates NO-cGMP signaling, which affects CO 2 reactivity. Fourteen healthy males (34 ± 2 yr) were enrolled in the study. Blood pressure (BP), ECG, velocity of cerebral blood flow (CBF; measured by transcranial Doppler), and end-tidal CO 2 (EtCO 2 ) were assessed at baseline (CO 2 ~39 mmHg), during hyperventilation (CO 2 ~24 mmHg), during hypercapnia (CO 2 ~46 mmHg), during boluses of phenylephrine (25-200 µg), and during graded head-up tilting (HUT). Measurements were repeated 1 h after 100 mg sildenafil were taken. Results showed that sildenafil did not affect resting BP, heart rate, CBF peak and mean velocities, estimated regional cerebrovascular resistance (eCVR; mean BP/mean CBF), breath/min, and EtCO 2 : 117 ± 2/67 ± 3 mmHg, 69 ± 3 beats/min, 84 ± 5 and 57 ± 4 cm/s, 1.56 ± 0.1 mmHg·cm -1 ·s -1 , 14 ± 0.5 breaths/min, and 39 ± 0.9 mmHg, respectively. Sildenafil increased and decreased the hypercapnia induced in CBF and eCVR, respectively. Sildenafil also attenuated the decrease in peak velocity of CBF, 25 ± 2 vs. 20 ± 2% ( P < 0.05) and increased the eCVR, 2.5 ± 0.2 vs. 2 ± 0.2% ( P < 0.03) during hyperventilation. Sildenafil did not affect CBF despite significant increases in the eCVRs that were elicited by phenylephrine and HUT. This investigation suggests that sildenafil, which potentiates the NO-cGMP signaling, seems to affect the cerebrovascular CO 2 reactivity without affecting the static and dynamic pressure-dependent mechanisms of cerebrovascular autoregulation., (Copyright © 2017 the American Physiological Society.)

Marfan syndrome (MFS) is a connective tissue disorder that is often associated with the fibrillin-1 (Fbn1) gene mutation and characterized by cardiovascular alterations, predominantly ascending aortic aneurysms. Although neurovascular complications are uncommon in MFS, the improvement in Marfan patients' life expectancy is revealing other secondary alterations, potentially including neurovascular disorders. However, little is known about small-vessel pathophysiology in MFS. MFS is associated with hyperactivated transforming growth factor (TGF)-β signaling, which among numerous other downstream effectors, induces the NADPH oxidase 4 (Nox4) isoform of NADPH oxidase, a strong enzymatic source of H2O2 We hypothesized that MFS induces middle cerebral artery (MCA) alterations and that Nox4 contributes to them. MCA properties from 3-, 6-, or 9-mo-old Marfan (Fbn1(C1039G/+)) mice were compared with those from age/sex-matched wild-type littermates. At 6 mo, Marfan compared with wild-type mice developed higher MCA wall/lumen (wild-type: 0.081 ± 0.004; Marfan: 0.093 ± 0.002; 60 mmHg; P < 0.05), coupled with increased reactive oxygen species production, TGF-β, and Nox4 expression. However, wall stiffness and myogenic autoregulation did not change. To investigate the influence of Nox4 on cerebrovascular properties, we generated Marfan mice with Nox4 deficiency (Nox4(-/-)). Strikingly, Nox4 deletion in Marfan mice aggravated MCA wall thickening (cross-sectional area; Marfan: 6,660 ± 363 μm(2); Marfan Nox4(-/-): 8,795 ± 824 μm(2); 60 mmHg; P < 0.05), accompanied by decreased TGF-β expression and increased collagen deposition and Nox1 expression. These findings provide the first evidence that Nox4 mitigates cerebral artery structural changes in a murine model of MFS., (Copyright © 2016 the American Physiological Society.)

Background/aim: H2S is a novel vasoactivator. To verify the hypothesis that H2S may act as an endothelium-derived hyperpolarizing factor (EDHF) in the rat cerebrovasculature, the role of H2S in endothelium-derived relaxing factor (EDRF)-mediated responses was investigated., Methods: Cystathionine-γ-lyase (CSE) was knocked down with an siRNA technique. Artery diameter, hyperpolarization and Ca2+-activated K+ (KCa) current were measured., Results: CSE knockdown was indicated by a decrease in protein and mRNA expression in the rat middle cerebral artery (MCA) and cerebral basilar artery (CBA). Acetylcholine (ACh) induced significant hyperpolarization and vasodilation in endothelium-intact MCA and CBA. Removal of the endothelium abolished these responses. The nitric oxide (NO) synthase inhibitor L-NAME, but not the PGI2 production inhibitor indomethacin, significantly inhibited ACh-induced hyperpolarization and vasodilation in the CBA. In the presence of L-NAME and indomethacin, ACh-induced hyperpolarization and vasodilation in the MCA and CBA were attenuated. The non-NO/PGI2-mediated responses were abolished by the KCa channel blockers charybdotoxin and apamin. In the cerebral arteries from the CSE knockdown rat, non-NO/PGI2-mediated responses were significantly attenuated, and the remaining responses were abolished by charybdotoxin and apamin or the CSE inhibitor propargylglycine. CSE knockdown did not affect L-NAME-sensitive responses in the CBA. Sodium hydrosulfide (NaHS) augmented the KCa current in CBA vascular smooth muscle cells., Conclusion: EDHF-mediated responses in rat cerebral arteries were due to H2S activating the KCa channel., (© 2016 S. Karger AG, Basel.)

Objective: To observe the effect of electroacupuncture (EA) intervention on expression and content of protein kinase C (PKC) in the middle cerebral artery in acute cerebral infarction (ACI) rats so as to explore its mechanism underlying improvement of ACI., Methods: Wistar rats were randomly divided into normal control (n = 6), sham operation (n = 30), ACI model (n = 30), and EA (n = 30) groups, and the latter three groups were further divided into 0. 5 h, 1 h, 3 h, 6 h and 12 h subgroups (n = 6 in each subgroup). The ACI model was established by occlusion of the middle cerebral artery (MCAO). EA (15 Hz, 1 mA) was applied to "Shuigou" (GV 26) for 20 min. The PKC expression levels and activity in the vascular smooth muscle of the middle cerebral artery were detected using immunohistochemistry and ELISA, respectively., Results: In comparison with the control group, the immunoactivity and activities of PKC in the middle cerebral artery tissue at 0. 5 h, 1 h, 3 h, 6 h and 12 h were significantly increased in the model group (P<0. 05). After EA intervention, the expression levels and activities of PKC at the 5 time-points were markedly down-regulated in comparison with the model group at the same corresponding time-point (P<0. 05). No significant changes of PKC expression and activity were found in the sham operation group (P>0. 05)., Conclusion: EA intervention can up-regulate the immunoactivity and activity of PKC in the vascular smooth muscle of the middle cerebral artery in ACI rats, which may contribute to its effect in improving ACI by relieving arterial spasm.

We have reported that a reduction in renal production of 20-HETE contributes to development of hypertension in Dahl salt-sensitive (SS) rats. The present study examined whether 20-HETE production is also reduced in the cerebral vasculature of SS rats and whether this impairs the myogenic response and autoregulation of cerebral blood flow (CBF). The production of 20-HETE, the myogenic response of middle cerebral arteries (MCA), and autoregulation of CBF were compared in SS, SS-5(BN) rats and a newly generated CYP4A1 transgenic rat. 20-HETE production was 6-fold higher in cerebral arteries of CYP4A1 and SS-5(BN) than in SS rats. The diameter of the MCA decreased to 70 ± 3% to 65 ± 6% in CYP4A1 and SS-5(BN) rats when pressure was increased from 40 to 140 mmHg. In contrast, the myogenic response of MCA isolated from SS rats did not constrict. Administration of a 20-HETE synthesis inhibitor, HET0016, abolished the myogenic response of MCA in CYP4A1 and SS-5(BN) rats but had no effect in SS rats. Autoregulation of CBF was impaired in SS rats compared with CYP4A1 and SS-5(BN) rats. Blood-brain barrier leakage was 5-fold higher in the brain of SS rats than in SS-5(BN) and SS.CYP4A1 rats. These findings indicate that a genetic deficiency in the formation of 20-HETE contributes to an impaired myogenic response in MCA and autoregulation of CBF in SS rats and this may contribute to vascular remodeling and cerebral injury following the onset of hypertension., (Copyright © 2015 the American Physiological Society.)

Myogenic responses by resistance vessels are a key component of autoregulation in brain, thus playing a crucial role in regulating cerebral blood flow and protecting the blood-brain barrier against potentially detrimental elevations in blood pressure. Although cerebrovascular disease is often accompanied by alterations in myogenic responses, mechanisms that control these changes are poorly understood. Peroxisome proliferator-activated receptor γ has emerged as a regulator of vascular tone. We hypothesized that interference with peroxisome proliferator-activated receptor γ in smooth muscle would augment myogenic responses in cerebral arteries. We studied transgenic mice expressing a dominant-negative mutation in peroxisome proliferator-activated receptor γ selectively in smooth muscle (S-P467L) and nontransgenic littermates. Myogenic tone in middle cerebral arteries from S-P467L was elevated 3-fold when compared with nontransgenic littermates. Rho kinase is thought to play a major role in cerebrovascular disease. The Rho kinase inhibitor, Y-27632, abolished augmented myogenic tone in middle cerebral arteries from S-P467L mice. CN-03, which modifies RhoA making it constitutively active, elevated myogenic tone to ≈60% in both strains, via a Y-27632-dependent mechanism. Large conductance Ca(2+)-activated K(+) channels (BKCa) modulate myogenic tone. Inhibitors of BKCa caused greater constriction in middle cerebral arteries from nontransgenic littermates when compared with S-P467L. Expression of RhoA or Rho kinase-I/II protein was similar in cerebral arteries from S-P467L mice. Overall, the data suggest that peroxisome proliferator-activated receptor γ in smooth muscle normally inhibits Rho kinase and promotes BKCa function, thus influencing myogenic tone in resistance arteries in brain. These findings have implications for mechanisms that underlie large- and small-vessel disease in brain, as well as regulation of cerebral blood flow., (© 2014 American Heart Association, Inc.)

Vasospasm is known to contribute to delayed cerebral ischemia following subarachnoid hemorrhage (SAH). We hypothesized that vasospasm initiates structural changes within the vessel wall, possibly aggravating ischemia and leading to resistance to vasodilator treatment. We therefore investigated the effect of blood on cerebral arteries with respect to contractile activation and vascular remodeling. In vitro experiments on rodent basilar and middle cerebral arteries showed a gradual contraction in response to overnight exposure to blood. After incubation with blood, a clear inward remodeling was found, reducing the caliber of the passive vessel. The transglutaminase inhibitor L682.777 fully prevented this remodeling. Translation of the in vitro findings to an in vivo SAH model was attempted in rats, using both a single prechiasmatic blood injection model and a double cisterna magna injection model, and in mice, using a single prechiasmatic blood injection. However, we found no substantial changes in active or passive biomechanical properties in vivo. We conclude that extravascular blood can induce matrix remodeling in cerebral arteries, which reduces vascular caliber. This remodeling depends on transglutaminase activity. However, the current rodent SAH models do not permit in vivo confirmation of this mechanism., (© 2015 S. Karger AG, Basel.)

Endothelial dysfunction occurs in conduit and cerebral resistance arteries with advancing age. Lifelong caloric restriction (CR) can prevent the onset of age-related dysfunction in many tissues, but its effects on cerebral resistance artery function, as compared with conduit artery function, have not been determined. We measured endothelium-dependent dilation (EDD) in the carotid artery and middle cerebral artery (MCA) from young (5-7 months), old ad libitum fed (AL, 29-32 months), and old lifelong CR (CR, 40 % CR, 29-32 months) B6D2F1 mice. Compared with young, EDD for old AL was 24 % lower in the carotid and 47 % lower in the MCA (p < 0.05). For old CR, EDD was not different from young in the carotid artery (p > 0.05), but was 25 % lower than young in the MCA (p < 0.05). EDD was not different between groups after NO synthase inhibition with N(ω)-nitro-L-arginine methyl ester in the carotid artery or MCA. Superoxide production by the carotid artery and MCA was greater in old AL compared with young and old CR (p < 0.05). In the carotid, incubation with the superoxide scavenger TEMPOL improved EDD for old AL (p > 0.05), with no effect in young or old CR (p > 0.05). In the MCA, incubation with TEMPOL or the NADPH oxidase inhibitor apocynin augmented EDD in old AL (p < 0.05), but reduced EDD in young and old CR (p < 0.05). Thus, age-related endothelial dysfunction is prevented by lifelong CR completely in conduit arteries, but only partially in cerebral resistance arteries. These benefits of lifelong CR on EDD result from lower oxidative stress and greater NO bioavailability.

Hydrogen sulfide (H(2)S) is an endogenous mediator with peripheral vasorelaxant effects; however, the mechanism of H(2)S-induced vasorelaxation in cerebral blood vessels has not been extensively studied. Vasorelaxation studies were performed on middle cerebral arteries from male Sprague Dawley rats using wire myography. Immunofluorescence staining was used to detect the presence of the H(2)S-producing enzyme cystathionine-γ-lyase (CSE). CSE was present in the endothelium and smooth muscle of middle cerebral arteries. The CSE substrate, L-cysteine, induced vasorelaxation that was sensitive to the CSE inhibitor DL-propargylglycine. This relaxation was independent of endothelium, suggesting that H(2)S was produced in the vascular smooth muscle. The H(2)S donor, sodium hydrogen sulfide (NaHS; 0.1-3.0 mM) produced concentration-dependent relaxation, which was unaffected by endothelium removal. Nifedipine (3 μM) significantly reduced the maximum relaxation elicited by NaHS. Inhibiting potassium (K(+)) conductance with 50 mM K(+) significantly attenuated NaHS-induced relaxation, however, selective blockers of ATP sensitive (K(ATP)), calcium sensitive (K(Ca)), voltage dependent (K(V)), or inward rectifier (K(ir)) channels alone or in combination did not affect the response to NaHS. 4,4-diisothiocyanatostilbene-2,2-disulfonic acid (DIDS; 300 μM) caused a significant rightward shift of the NaHS concentration-response curve, but this effect could not be explained by inhibition of Cl(-) channels or Cl(-)/HCO (3)(-) exchange, as selective blockade of these mechanisms had no effect. These findings suggest endogenous H(2)S can regulate cerebral vascular function. The H(2)S-mediated relaxation of middle cerebral arteries is DIDS sensitive and partly mediated by inhibition of L-type calcium channels, with an additional contribution by K channels but not K(ATP), K(Ca), K(V), or K(ir) subtypes.

Phosphodiesterase 5 (PDE5) is associated with migraine pathophysiology, stroke recovery and vasospasm treatment. The potential vascular interplay of PDE5 inhibitors sildenafil, tadalafil and UK-114,542 was studied by intra- versus extra-luminal administration in rat middle cerebral arteries in vitro and on middle meningeal arteries in vivo. By Western blot PDE5 was detected in both cerebral and meningeal arteries, though with minor variations in band intensity between vascular beds. Rat middle cerebral artery diameter was investigated using pressurised arteriography, applying UK-114,542, sildenafil, and tadalafil intra- or extra-luminally. Effects on the dural middle meningeal artery were studied in the in vivo closed cranial window model. At high concentrations, abluminal sildenafil and UK-114,542, but not tadalafil, induced dilatation of the middle cerebral artery. Luminal application elicited a contraction of 4% (sildenafil, P=0.03) and 10% (tadalafil, P=0.02). In vivo, sildenafil, but not tadalafil, dose-dependently dilated middle meningeal artery concomitant to blood pressure reduction (1-3mg/kg);1mg/kg sildenafil inducing 60 ± 14% (P=0.04) and vehicle (DMSO) 13 ± 6% dilatation. In conclusion, PDE5 inhibitors applied luminally had minor contractile effect, whereas abluminal sildenafil induced middle cerebral artery dilatation above therapeutic levels. In vivo, sildenafil dilated middle meningeal artery concomitant with a reduction in blood pressure. Tadalafil had no dilatory effects. PDE5 inhibitors show differential vascular activity in cerebral arteries from healthy animals; arterial dilatation is seen primarily above therapeutic levels. Such findings support clinical studies showing no vasodilator effects of sildenafil on cerebral arteries in healthy subjects., (Copyright © 2011 Elsevier B.V. All rights reserved.)

A major cause of stroke is cerebral ischemia in regions supplied by the middle cerebral artery (MCA). In this study, we hypothesized that compromised cardiovascular function during static exercise may involve altered expression of inducible NOS (iNOS) protein within the rostral ventrolateral medulla (RVLM) and caudal ventrolateral medulla (CVLM). We compared cardiovascular responses and iNOS protein expression within the left and right sides of both RVLM and CVLM in sham-operated rats and in rats with a 90 min left-sided MCA occlusion (MCAO) followed by 24 h of reperfusion. Increases in blood pressure during a static muscle contraction were attenuated in MCAO rats compared with sham-operated rats. Also, iNOS expression within the left RVLM was augmented compared with the right RVLM in MCAO rats and compared with both RVLM quadrants in sham-operated rats. In contrast, compared with sham-operated rats and the right CVLM of MCAO rats, iNOS expression was attenuated in the left CVLM in left-sided MCAO rats. These data suggest that the attenuation of pressor responses during static exercise in MCAO rats involves overexpression of iNOS within the ipsilateral RVLM and attenuation in iNOS within the ipsilateral CVLM. Differential expression of iNOS within the medulla plays a role in mediating cardiovascular responses during static exercise following stroke.

Although the use of antenatal glucocorticoids has resulted in decreased neonatal morbidity/mortality, recent animal studies have raised concerns regarding adverse effects of these medications on postnatal cardiovascular function. We hypothesized that antenatal betamethasone (Beta) exposure alters cerebral vascular reactivity in adult female sheep. We observed that K-induced constriction was comparable in middle cerebral artery (MCA) from Beta-exposed animals and age-matched controls. Pressure-induced constriction was significantly attenuated in MCA from Beta-exposed compared with control sheep. Inhibition of NOS significantly augmented pressure-induced constriction in MCA from both Beta-exposed and control sheep, whereas cyclooxygenase (COX) inhibition augmented pressure-induced constriction only in MCA from Beta-exposed sheep. Furthermore, NOS and COX inhibition significantly attenuated bradykinin (BK)-induced dilation in MCA from both Beta-exposed and control sheep. However, there seemed to be a greater contribution of both NOS and COX to BK-induced dilation in Beta-exposed compared with control MCA. Our findings demonstrate that fetal exposure to a clinically relevant course of Beta alters cerebral vascular tone and reactivity in adult female sheep.

Blocking or impairment of the sphenopalatine ganglia (SPG) is an effective therapy of cluster headache and other pain syndromes. Contrarily, unilateral SPG-stimulation reduces infarction size in the rat permanent suture model. Well, what are the effects of the SPG damage on the following brain ischemia? This study was aimed to investigate the effects of resection of the nerves bundle from the SPG of rat on the brain lesions following middle cerebral artery occlusion (MCAO), and evaluated the roles of the nitric oxygen synthase (NOS) immunoreactive perivascular nerves of cerebral arteries in MCAO. We found that 7 days after bilateral resections of the nerves bundle from the SPG, the NOS activity perivascular nerves in the middle cerebral arteries disappeared, and the infarction volume and the TUNEL positive cells increased significantly after 24 h MCAO, which implicated that the NOS contained nerves from the SPG maybe have an important role in the MCAO.

Purpose: Oxidized low-density lipoprotein receptor 1 (LOX1) is a critical factor for atherosclerosis in a variety of vascular diseases; however, its major role in cerebral arterial dissecting aneurysm is unclear., Clinical Presentation: We present a case of remarkable contrast of LOX1 expression in ruptured and unruptured multiple middle cerebral artery dissections and discuss the correlation of LOX1 with matrix metalloproteinases (MMPs). A 59-year-old woman presented with subarachnoid hemorrhage associated with left temporal subcortical hematoma. Emergent cerebral angiography demonstrated aneurysmal dilatation at the origin of the left anterior temporal artery (ATA) and occlusion on the distal side of ATA. Infectious aneurysm was excluded. Intraoperative findings showed ruptured dissection of the left ATA and unruptured aneurysmal dilatation of another temporal branch of the left M1 portion. Both lesions were trapped by clips and resected. Histopathological examination confirmed that both ruptured and unruptured aneurysmal dilatations were diagnosed as arterial dissections. Immunohistochemical examination demonstrated remarkable expressions of LOX1, MMP-2, and MMP-9 in hypertrophic media outside the intima in ruptured dissection, on the other hand, those expressions in the intima and inside hypertrophic media in the unruptured dissection., Conclusions: This is the first report to reveal immunohistochemical findings of LOX1 and MMPs in multiple dissections of MCA. The contrast localization of LOX1 and MMPs might contribute to the fragility of the arterial wall layer of ruptured/unruptured arterial dissections.

Background: Gamma-glutamyl carboxylation, a reaction essential for the biosynthesis of vitamin K-dependent coagulation factors, requires the participation of the gamma-glutamyl carboxylase (GGCX), vitamin K epoxide reductase (VKORC1), and NAD(P)H:quinone oxidoreductase (NQO1). We evaluated the role of these genotype polymorphisms in patients with large-artery atherosclerotic stroke., Methods: In this hospital-based case-control study, 117 patients who were categorized as having large-artery atherosclerotic stroke and 115 age- and gender-matched controls were recruited. Genotyping determination for the GGCX1 (Gln325Arg), NQO1 (Pro187Ser), and VKORC1 (rs9923231) polymorphisms was performed. The associations of genotype with ischemic stroke (IS) risk were examined., Results: A higher genotypic frequency of NQO1 C609T was found in the controls than in the patients, manifesting a 0.47-fold risk reduction in IS (95% CI=0.25-0.87). A tendency toward a reduced IS risk was statistically significant in those subjects who carried a greater number of the NQO1, GGCX, and VKORC1 polymorphisms (aOR=0.58, P(trend)=0.005). The synergistic effect of multiple genes on risk reduction was more significant in a subset of patients who were not alcoholics and who were non-smokers (P<0.05)., Conclusions: Compartmentation of coagulation factor metabolism may account for the preferential role of NQO1, GGCX, and VKORC1 polymorphisms to lower the risk for large-artery atherosclerotic stroke., (Copyright 2010 Elsevier B.V. All rights reserved.)

Aims: The modulation of myogenic function and cerebral blood flow (CBF) by nitric oxide (NO) synthases (NOS) was assessed in the middle cerebral arteries (MCAs) of Kyoto Wistar stroke prone hypertensive rats (SHRsp) in relation to haemorrhagic stroke development., Methods and Results: MCAs were studied with a pressure myograph. CBF in MCA perfusion domain was measured using laser Doppler techniques. NOS isozymes were identified using immunohistochemistry. MCAs expressed endothelial, neuronal, and inducible NOS (eNOS, nNOS, and iNOS, respectively) in the endothelium, nNOS and traces of iNOS in smooth muscle and adventitial cells. Before stroke, MCA pressure-dependent constriction (PDC) was superimposed over basal non-pressure-dependent tone (BNPDT). Endothelial NO generation and non-endothelial nNOS but not iNOS reduced BNPDT and increased the lumen diameter at which PDC initiated without altering the amplitude of PDC. NOS inhibition decreased CBF and increased the upper blood pressure limit of autoregulation. PDC, CBF autoregulation, and NOS dilatory influence were lost, and BNPDT was increased in MCAs from SHRsp with stroke. The expression of NOS isozymes and MCA reactivity to NO donors was not altered. NOS activity was not recovered by in vitro l-arginine or tetrahydrobiopterin supplementation, l-arginase inhibition or superoxide scavengers., Conclusion: The loss of PDC and CBF autoregulation during hypertension may facilitate over-perfusion and cerebral haemorrhage formation in SHRsp. NOS dysfunction in MCAs preceded stroke and involved the inactivation of eNOS and nNOS in areas not subjected to hyper-distension. The elevation in BNPDT due to NOS inactivation may oppose over-perfusion in the absence of CBF autoregulation.

Background/aims: In cerebral arteries, nitric oxide (NO) release plays a key role in suppressing vasomotion. Our aim was to establish the pathways affected by NO in rat middle cerebral arteries., Methods: In isolated segments of artery, isometric tension and simultaneous measurements of either smooth muscle membrane potential or intracellular [Ca(2+)] ([Ca(2+)](SMC)) changes were recorded., Results: In the absence of L-NAME, asynchronous propagating Ca(2+) waves were recorded that were sensitive to block with ryanodine, but not nifedipine. L-NAME stimulated pronounced vasomotion and synchronous Ca(2+) oscillations with close temporal coupling between membrane potential, tone and [Ca(2+)](SMC). If nifedipine was applied together with L-NAME, [Ca(2+)](SMC) decreased and synchronous Ca(2+) oscillations were lost, but asynchronous propagating Ca(2+) waves persisted. Vasomotion was similarly evoked by either iberiotoxin, or by ryanodine, and to a lesser extent by ODQ. Exogenous application of NONOate stimulated endothelium-independent hyperpolarization and relaxation of either L-NAME-induced or spontaneous arterial tone. NO-evoked hyperpolarization involved activation of BK(Ca) channels via ryanodine receptors (RYRs), with little involvement of sGC. Further, in whole cell mode, NO inhibited current through L-type voltage-gated Ca(2+) channels (VGCC), which was independent of both voltage and sGC., Conclusion: NO exerts sGC-independent actions at RYRs and at VGCC, both of which normally suppress cerebral artery myogenic tone., (Copyright 2009 S. Karger AG, Basel.)

Ca(2+) sensitization has been postulated to contribute to the myogenic contraction of resistance arteries evoked by elevation of transmural pressure. However, the biochemical evidence of pressure-induced increases in phosphorylated myosin light chain phosphatase (MLCP) targeting subunit 1 (MYPT1) and/or 17 kDa protein kinase C (PKC)-potentiated protein phosphatase 1 inhibitor protein (CPI-17) required to sustain this view is not currently available. Here, we determined whether Ca(2+) sensitization pathways involving Rho kinase (ROK)- and PKC-dependent phosphorylation of MYPT1 and CPI-17, respectively, contribute to the myogenic response of rat middle cerebral arteries. ROK inhibitors (Y27632, 0.03-10 micromol l(-1); H1152, 0.001-0.3 micromol l(-1)) and PKC inhibitors (GF109203X, 3 micromol l(-1); Gö6976; 10 micromol l(-1)) suppressed myogenic vasoconstriction between 40 and 120 mmHg. An improved, highly sensitive 3-step Western blot method was developed for detection and quantification of MYPT1 and CPI-17 phosphorylation. Increasing pressure from 10 to 60 or 100 mmHg significantly increased phosphorylation of MYPT1 at threonine-855 (T855) and myosin light chain (LC(20)). Phosphorylation of MYPT1 at threonine-697 (T697) and CPI-17 were not affected by pressure. Pressure-evoked elevations in MYPT1-T855 and LC(20) phosphorylation were reduced by H1152, but MYPT1-T697 phosphorylation was unaffected. Inhibition of PKC with GF109203X did not affect MYPT1 or LC(20) phosphorylation at 100 mmHg. Our findings provide the first direct, biochemical evidence that a Ca(2+) sensitization pathway involving ROK-dependent phosphorylation of MYPT1 at T855 (but not T697) and subsequent augmentation of LC(20) phosphorylation contributes to myogenic control of arterial diameter in the cerebral vasculature. In contrast, suppression of the myogenic response by PKC inhibitors cannot be attributed to block of Ca(2+) sensitization mediated by CPI-17 or MYPT1 phosphorylation.

Aims: Cigarette smoking is one of the strongest risk factors for stroke. However, the underlying molecular mechanisms that smoke leads to the pathogenesis of stroke are incompletely understood., Methods: Dimethyl sulfoxide (DMSO)-soluble (lipid-soluble) cigarette smoking particles (DSP) were extracted from cigarette smoke (0.8 mg nicotine per cigarette; Marlboro). Rat cerebral arteries were isolated and organ cultured in the presence of DSP (0.2 microl/ml, equivalent to the plasma level in smokers) for 24 h. The expression of matrix metalloproteinase 9 and 13 (MMP9 and MMP13), angiotensin receptor 1 and 2 (AT(1) and AT(2)), interleukin 6 and inducible nitric oxide synthase (iNOS) were investigated at mRNA level by real-time PCR and/or at protein level by immunohistochemistry. In addition, the activity of three mitogen-activated protein kinases (p38, ERK 1/2 and SAPK/JNK) and their downstream transcription factors (ATF-2, Elk-1 and c-Jun) were examined., Results: We observed that compared with control (DMSO-treated cerebral arteries), the cerebral arteries treated by DSP exhibited enhanced expression of MMP13 and AT(1) receptors, but not of AT(2) receptors, at both mRNA and protein levels, suggesting that a transcriptional mechanism is most likely involved in the DSP effects. This is further supported by the findings that DSP induced phosphorylation of p38 mitogen-activated protein kinases inflammatory signal protein in parallel with activation of its downstream transcription factor ATF-2 and Elk-1. However, ERK 1/2 and SAPK/JNK activities were markedly expressed in the control (organ culture per se with DMSO), and DSP failed to further enhance the activation of ERK 1/2 and SAPK/JNK in the cerebral arteries., Conclusions: DSP induces cerebral vessel inflammation with activation of p38 MAPK inflammatory signal and the downstream transcriptional factors (ATF-2 and Elk-1) in parallel with enhanced extracellular-matrix-related gene transcription and increased AT(1) receptor expression in the cerebral arteries, which are key events in stroke pathogenesis.

Objective: We investigated the effect of ischemia and reperfusion on the vasoactive function of penetrating brain parenchymal arterioles under pressurized conditions., Methods: Parenchymal arterioles (< 50 microm in diameter) from within the middle cerebral artery territory were dissected from male Wistar rats that were either nonischemic control (n = 16) or ischemic for one hour and reperfused for 24 hours (n = 16) by temporary filament occlusion of the middle cerebral artery. Arterioles were mounted on glass cannulas within an arteriograph chamber that allowed for the measurement of lumen diameter and control over intravascular pressure., Results: After one hour of equilibration at 10 mmHg, spontaneous myogenic tone developed in both groups of animals, constricting control arterioles from 69 +/- 9 to 49 +/- 11 microm (29.5 +/- 10.2%) and ischemic arterioles from 66 +/- 9 to 45 +/- 11 microm (33.1 +/- 14.1%); p > 0.05. Contraction to the nitric oxide synthase inhibitor nitro-L-arginine (10(-4)M) was significantly diminished in ischemic arterioles, constricting only 3.2 +/- 3.3 vs. 15.6 +/- 12.5% in control arterioles (p = 0.017). Both groups dilated to nifedipine; however, the response was significantly diminished after ischemia. The EC50 for nifedipine in control arterioles was 3.54 +/- 0.11 vs. 9.90 +/- 0.71 nM for ischemic arterioles (p = 0.024)., Conclusions: These findings demonstrate that functional changes occur in brain parenchymal arterioles after ischemia and reperfusion, a result that may significantly influence stroke outcome by altering blood flow to an ischemic region.

The fetal cardiovascular responses to hypoxia include decreased peripheral blood flow and increased cerebral, cardiac, and adrenal blood flow. Prostanoids, metabolites of cyclooxygenase enzyme activity, have potent effects on vascular tone in both the adult and the fetus. To examine the role of prostanoids in the regulation of fetal cerebral blood flow (CBF) during acute hypoxic stress, eight near term fetal sheep were studied after infusing vehicle or diclofenac, a cyclooxygenase inhibitor, followed by a 30-min period of hypoxia (arterial Po(2) 12 Torr). In the control experiments, CBF, measured continuously with laser Doppler flowmetry, increased to 148% of baseline values (p < 0.01) and cerebral vascular resistance decreased to 70% of baseline values after 30 min of hypoxic stress. During diclofenac infusion, hypoxia resulted in a CBF increase to only 129% of baseline, a significant attenuation (p < 0.05), accompanied by decreased plasma prostanoid concentrations. Increases in mean arterial blood pressure during hypoxia were also attenuated by diclofenac infusion. Flow and pressure responses were not accompanied by changes in cerebral vascular resistance. These results indicate that prostanoids indirectly modulate fetal CBF responses to hypoxia, but that their effects are mediated through modulation of systemic rather than cerebral vascular tone.

Reactive oxygen species play a role in neuronal damage following cerebral ischemia-reperfusion. We tested whether activity of the superoxide-generating enzyme, NADPH-oxidase, is enhanced in cerebral arteries within, adjacent and distant from the ischemic core. The right middle cerebral artery (MCA) of conscious rats was temporarily occluded by perivascular injection of endothelin-1 to induce stroke (ET-1; n=19). Control rats were injected with saline (n=9). At 24 h or 72 h post-administration of ET-1, the MCA and its branches within the ipsilateral penumbra and infarcted core, corresponding arteries in the contralateral hemisphere, and basilar artery were excised. Anatomically similar arteries were excised from saline-injected rats. At 24 h after stroke, NADPH-stimulated superoxide production by arteries from the infarcted core did not differ from levels generated by arteries from control rats, whereas levels were significantly lower 72 h after stroke. However, at both time points after stroke, superoxide production by arteries from the ischemic penumbra was 8-fold greater than levels generated by arteries from control rats. Surprisingly, even in the non-ischemic arteries from the contralateral hemisphere and in the basilar artery, superoxide production was increased approximately 4- to 6-fold at 24 h, but had returned to normal 72 h after stroke. The NADPH-oxidase inhibitor, diphenyleneiodonium, virtually abolished superoxide production by all arteries. Thus, the activity of NADPH-oxidase is enhanced in cerebral arteries from the ischemic penumbra at 24 h and 72 h following cerebral ischemia. Additionally, NADPH-oxidase activity is temporarily enhanced after cerebral ischemia within arteries from non-ischemic parts of the brain.

The flower buds of Buddleja officinalis MAXIM (Loganiaceae) are used to treat headache and inflammatory diseases in traditional Korean medicine. In the present study, the neuroprotective effects of the methanolic extract of B. officinalis (BOME) and of its hexane fraction (BOHF) were investigated in a middle cerebral artery occlusion (MCAo, 120 min occlusion, 24 h reperfusion) Sprague-Dawley rat model. BOME or BOHF (100 mg/kg, p.o.) was twice administered 30 min before the onset of MCAo and 2 h after reperfusion. BOME and BOHF treated groups showed infarct volumes reduced by 33.9% and 68.2%, respectively, at 2 h occlusion. In BOHF treated animals, cyclooxygenase-2 and iNOS inductions were inhibited in ischemic hemispheres at both the mRNA and protein levels. Furthermore, in vitro studies showed that BOME and BOHF both inhibited LPS-induced nitric oxide production in BV-2 mouse microglial cells. These results suggest that the anti-inflammatory and the microglial activation inhibitory effects of B. officinalis extract may contribute to its neuroprotective effects in brain ischemia.

The purpose of this study was to determine whether chronic blockade of the angiotensin II (ANG II) AT1 receptor under normal physiological conditions impairs vascular relaxation mechanisms in isolated middle cerebral arteries (MCA). Male Sprague-Dawley rats on a standard diet were given losartan (1 mg/mL) in the drinking water or normal water ad libitum for 7 days. Vessel diameters were measured by television microscopy before and during exposure to various vasodilator agonists and reductions in PO2 from 140 mm Hg to 35-45 mm Hg. Dilations to acetylcholine (1 microM), the stable prostacyclin analogue iloprost (10 pg/mL), and the Gs protein activator cholera toxin (1 ng/mL) were completely eliminated in vessels from losartan-treated animals. However, middle cerebral arteries from control and losartan-treated rats still demonstrated significant dilations in response to reduced PO2. Hypoxic dilation of middle cerebral arteries from control rats was eliminated by indomethacin (1 microM) and unaffected by the NOS inhibitor L-NAME (100 microM) whereas dilation in response to reduced PO2 in middle cerebral arteries from losartan-treated rats was eliminated by L-NAME and unaffected by indomethacin. Middle cerebral arteries from control and losartan-treated animals exhibited similar dilations in response to the NO-donor sodium nitroprusside (1 microM). These data suggest that AT1 receptor activation is important in maintaining normal vascular relaxation mechanisms in cerebral resistance arteries during normal physiological conditions, and that AT1 receptor blockade causes a shift in the mechanisms of hypoxic dilation of middle cerebral arteries from cyclooxygenase metabolites to NO.

We previously showed that testosterone, administered in vivo, increases the tone of cerebral arteries. A possible underlying mechanism is increased vasoconstriction through the thromboxane A2 (TxA2) pathway. Therefore, we investigated the effect of chronic testosterone treatment (4 wk) on TxA2 synthase levels and the contribution of TxA2 to vascular tone in rat middle cerebral arteries (MCAs). Using immunofluorescence and confocal microscopy, we demonstrated that TxA2 synthase is present in MCA segments in both smooth muscle and endothelial layers. Using Western blot analysis, we found that TxA2 synthase protein levels are higher in cerebral vessel homogenates from testosterone-treated orchiectomized (ORX + T) rats compared with orchiectomized (ORX) control animals. Functional consequences of changes in cerebrovascular TxA2 synthase were determined using cannulated, pressurized MCA segments in vitro. Constrictor responses to the TxA2 mimetic U-46619 were not different between the ORX + T and ORX groups. However, dilator responses to either the selective TxA2 synthase inhibitor furegrelate or the TxA2-endoperoxide receptor (TP) antagonist SQ-29548 were greater in the ORX + T compared with ORX group. In endothelium-denuded arteries, the dilation to furegrelate was attenuated in both the ORX and ORX + T groups, and the difference between the groups was abolished. These data suggest that chronic testosterone treatment enhances TxA2-mediated tone in rat cerebral arteries by increasing endothelial TxA2 synthesis without altering the TP receptors mediating constriction. The effect of in vivo testosterone on cerebrovascular TxA2 synthase, observed here after chronic hormone administration, may contribute to the risk of vasospasm and thrombosis related to cerebrovascular disease.

Brain aromatase has been shown to be increased in expression after neurotoxic damage and to exert neuroprotection via generation of local oestrogens. The present study investigates the topography and time course of brain aromatase expression after experimental stroke (middle cerebral artery occlusion (MCAO)). Ovariectomised stroke prone spontaneously hypertensive rats underwent distal MCAO by electrocoagulation. Immunohistochemistry revealed increased brain aromatase expression at 24h and 8 days in the cortical penumbra/peri-infarct zones with no increase evident at 2h or 30 days post-MCAO. Double label studies indicate that some of the increased aromatase expression is associated with astrocytic processes. Thus, this is the first evidence that aromatase protein is increased after MCAO and the location (peri-infarct), time course (within 24h) and cellular localisation (astrocytic) indicate the potential for aromatase to promote the survival of cells in the penumbra after experimental stroke by local synthesis of oestrogens.

In rats of Krushinsky-Molodkina strain (KMR), the audiogenic stress induced epileptiform seizure and development of acute disturbances of cerebral circulation of hemorrhagic nature. The inhibitor of NO-synthase (L-NNA) increased the severity of clinical symptoms, mortality, and the intensity of intracranial hemorrhages. In contrast, L-arginine elevated the resistance of KMRs to acoustic stress. The intensity of nitrergic innervation was analyzed in preparations of the middle cerebral artery with the use of histochemical NADPH-diaphorase staining. In preparations of control KMRs, a net of NADPH-positive perivascular nerve fibers was found, while in experimental KMRs, in an hour after sound stimulation, such fibers practically were not revealed. Preliminary exposure of KMRs in hypoxic conditions (1 hour in hypobaric chamber at simulated altitude of 5000 m above the sea level) decreased the development of stress lesions. The protective effect of hypoxic training disappeared after the administration of NO-synthase inhibitor (L-NNA). The study demonstrated participation of nitric oxide (NO) in adaptive reactions of cerebral hemodynamics linked with the significant increase of cerebral blood flow.

1. Inflammatory changes in peripheral arteries have been reported in animal models of hypertension. Whether they occur in cerebral arteries (CA) with hypertension induced by deprivation of endogenous nitric oxide (NO) remains unknown. 2. In the present study, we compared the arteriolar injury score (AIS) and perivascular inflammation in CA between hypertensive and normotensive rats following NO deprivation with the NO synthase inhibitor N(omega)-nitro-L-arginine methyl ester (L-NAME). Five-week-old male spontaneously hypertensive rats (SHR) and Wistar -Kyoto (WKY) rats were fed with L-NAME (1 mg/mL) for 4 weeks. 3. Nitric oxide deprivation resulted in time-dependent elevations in tail-cuff pressure (representing systolic blood pressure (SBP)) in both SHR and WKY rats. The magnitude of increase in SBP was larger in SHR (+81.0 +/- 3.2 vs+25.0 +/- 2.2 mmHg; P < 0.01). Arteriolar hyalinosis and AIS in various segments of the CA were assessed with periodic acid-Schiff staining and inflammatory cells were immunostained with the antibody against macrophage/monocyte marker (ED1). The ED1+ cells appeared in the middle CA of L-NAME-treated SHR as early as 2 weeks after treatment. These cells were not observed in L-NAME-treated WKY rats and untreated SHR. More ED1+ cells were found in L-NAME-treated SHR than L-NAME-treated WKY rats after 4 weeks treatment. 4. The AIS and number of ED1+ cells around the perivascular area of the internal carotid artery were significantly higher in L-NAME-treated compared with untreated rats (AIS: 137 +/- 28 vs 46 +/- 10 for WKY rats, respectively; 169 +/- 18 vs 53 +/- 6 for SHR, respectively (P < 0.01); ED1+ cells: 7.9 +/- 0.6 vs 1.3 +/- 0.9 for WKY rats, respectively; 13.6 +/- 2.7 vs 2.1 +/- 0.9 for SHR, respectively (P < 0.01)), although SBP was higher in untreated SHR than in L-NAME-treated WKY rats (170 +/- 4 vs 137 +/- 4 mmHg, respectively; P < 0.05). 5. These findings suggest that ED1+ cells appeared in the middle CA of L-NAME-SHR as early as 2 weeks after treatment. Chronic inhibition of NO accelerates hypertension and induces perivascular inflammation.

Although vascular dysregulation has been documented in patients with extracranial vascular disease, transient ischemic attacks, and stroke, the pathomechanisms are poorly understood. To model thromboembolic stroke in rats, photochemically induced nonocclusive common carotid artery thrombosis (CCAT) was used to generate a platelet thrombus in the carotid artery of anesthetized rats. After CCAT, platelet aggregates break off the thrombus, travel to the distal cerebral vasculature, damage blood vessels, and cause small infarctions. The authors hypothesized that deficits in the endothelial nitric oxide synthase (eNOS) pathway may be responsible for vascular dysfunction after embolic stroke. To examine the functional status of the eNOS system, they measured eNOS-dependent dilation after CCAT by applying acetylcholine through a cranial window over the middle cerebral artery. The authors also measured eNOS mRNA and protein in the middle cerebral artery to determine whether functional changes were caused by alterations in expression. eNOS-dependent dilation was reduced at 6 hours, elevated at 24 hours, and returned to baseline 72 hours after CCAT. Endothelial nitric oxide synthase mRNA increased at 2 hours and was followed by a rise in protein 24 hours after CCAT. Changes in the eNOS system may account for some of the observed vascular deficits in patients with cerebrovascular disease.

Hypertension-associated alterations of the nitric oxide (NO) pathway were analyzed in middle cerebral arteries (MCA) from normotensive (WKY) and hypertensive (SHR) rats. The vasoconstrictor response to prostaglandin F2alpha (PGF(2 alpha), 30 and 100 microM) was smaller in MCA from SHR than from WKY. Endothelium-dependent relaxations to bradykinin (1 nM-10 microM) or acetylcholine (10 microM) were similar in MCA from both strains, whereas the endothelium-independent response to sodium nitroprusside (1 nM-0.1 mM) was smaller in MCA from SHR. L-arginine (L-Arg, 10 microM) similarly inhibited the vasoconstrictor responses in both strains; however, the inhibitory effect of 100 microM of L-Arg was greater in MCA from SHR. N(omega)-nitro-L-arginine methyl ester (L-NAME, 100 microM), but not aminoguanidine (100 microM) or 7-nitroindazole (10 microM), increased basal tone, potentiated the PGF(2 alpha)-induced vasoconstrictor responses and reduced the bradykinin-elicited relaxation in a similar way in MCA from WKY and SHR. N(omega)-nitro-L-arginine methyl ester also antagonized the inhibitory effect of 10 microM of L-Arg. Incubation for 5 h with lipopolysaccharide (10 microg/ml) similarly reduced the response to PGF(2 alpha) in MCA from WKY and SHR; this reduction was antagonized by dexamethasone (1 microM). Cerebral arteries expressed endothelial (eNOS) and neuronal (nNOS) NO synthase similarly in both strains, but inducible NOS (iNOS) expression was more evident in SHR. Lipopolysaccharide increased iNOS expression in both strains to a similar level. The basal constitutive NOS (cNOS) and iNOS activities were similar in arteries from WKY and SHR. Lipopolysaccharide increased iNOS activity only in arteries from SHR. These results indicate that hypertension did not impair endothelial NO production by NOS activation but induced an up-regulation of basal iNOS expression.

Background and Purpose: Instability of the vascular tone (vasomotion) develops in several cerebrovascular diseases associated with endothelial dysfunction. The aim of the present study was to characterize cerebral vasomotion induced by diminished NO production with quantitative evaluation and chaos analysis. We tested the hypothesis that activation of thromboxane receptors mediates chaotic vasomotion after NO synthase (NOS) inhibition., Methods: Measurements of vascular tension were carried out in isolated rat middle cerebral arteries. The extent of vasomotion was characterized by tension instability, whereas vasomotion complexity was assessed by chaos analysis., Results: Blocking the basal NO release by N(omega)-nitro-L-arginine (L-NA) induced vasomotion, which was further enhanced and became irregular after UTP administration. The NO donor sodium nitroprusside was able to reverse this effect, and stable steady-state conditions reappeared. The guanylyl cyclase inhibitor 1H-(1,2,4)oxadiazolo[4,3-a]quinoxaline-1-one (ODQ) or coapplication of ODQ and L-NA had an effect identical to that of L-NA alone. Vasoconstriction by K(+) failed to induce vasomotion in intact vessels or in the presence of L-NA or ODQ. The thromboxane receptor antagonist ICI 192605 dose-dependently attenuated the vasomotion induced by L-NA and UTP, and the thromboxane-receptor agonist U-46619 induced significant vasomotion in intact vessels., Conclusions: The lack of NO in cerebral vessels provokes vulnerability to chaotic vasomotion, which can be triggered by the administration of UTP, whereas excess NO reverses it to stable conditions. The vasomotion after blockade of the NO-cGMP pathway is mediated by activation of thromboxane receptors.

The role of the neuronal isoform of the nitric oxide (NO) synthase (nNOS) in the regulation of the cerebrovascular tone was studied in vitro. Selective inhibition of nNOS by 7-nitro indazole monosodium salt (7-NINA) failed to alter the resting tension and the relaxant effect of bradykinin in isolated rat middle cerebral arteries. These results indicate that 1./ 7-NINA is selective for nNOS and 2./ cerebrovascular nNOS is involved neither in the resting NO production nor in the mediation of the relaxant effect of bradykinin. Therefore, nNOS-derived NO that contributes to the maintenance of the resting cerebral blood flow in vivo appears to be released from neurons and/or glial cells.