Modulation of p66Shc impairs cerebrovascular myogenic tone in low renin but not low nitric oxide models of systemic hypertension.

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 [NG-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.

Abnormal neurovascular coupling as a cause of excess cerebral vasodilation in familial migraine.

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.

Omega-3 fatty acid supplement reduces activation of NADPH oxidase in intracranial atherosclerosis stenosis.

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 (p47phox, gp91phox, p22phox, and p67phox) 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 p47phox, gp91phox, and p67phox. 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.

Nitric oxide-sensitive guanylyl cyclase signaling affects CO2-dependent but not pressure-dependent regulation of cerebral blood flow.

Cerebrovascular CO2 reactivity is affected by nitric oxide (NO). We tested the hypothesis that sildenafil selectively potentiates NO-cGMP signaling, which affects CO2 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 CO2 (EtCO2) were assessed at baseline (CO2 ~39 mmHg), during hyperventilation (CO2 ~24 mmHg), during hypercapnia (CO2 ~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 EtCO2: 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 CO2 reactivity without affecting the static and dynamic pressure-dependent mechanisms of cerebrovascular autoregulation.

Involvement of Hydrogen Sulfide in Endothelium-Derived Relaxing Factor-Mediated Responses in Rat Cerebral Arteries.

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.

NADPH oxidase 4 attenuates cerebral artery changes during the progression of Marfan syndrome.

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)-ß 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-ß, 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-ß 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.

[Effect of Electroacupuncture at "Shuigou" (GV 26) on Immunoactivity and Content of Protein Kinase C in the Middle Cerebral Artery in Acute Cerebral Infarction Rats].

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.

Cerebral Artery Remodeling in Rodent Models of Subarachnoid Hemorrhage.

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.

Genetic interference with peroxisome proliferator-activated receptor γ in smooth muscle enhances myogenic tone in the cerebrovasculature via A Rho kinase-dependent mechanism.

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.

Impaired myogenic response and autoregulation of cerebral blood flow is rescued in CYP4A1 transgenic Dahl salt-sensitive rat.

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.

Beneficial effects of lifelong caloric restriction on endothelial function are greater in conduit arteries compared to cerebral resistance arteries.

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.

An investigation of the mechanisms of hydrogen sulfide-induced vasorelaxation in rat middle cerebral arteries.

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.

Differential vasoactive effects of sildenafil and tadalafil on cerebral arteries.

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.

Transient middle cerebral artery occlusion and reperfusion alters inducible NOS expression within the ventrolateral medulla and modulates cardiovascular function during static exercise.

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.

Antenatal betamethasone alters vascular reactivity in adult female ovine cerebral arteries.

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.

Resection of the nerves bundle from the sphenopalatine ganglia tend to increase the infarction volume following middle cerebral artery occlusion.

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.

Genotype polymorphisms of GGCX, NQO1, and VKORC1 genes associated with risk susceptibility in patients with large-artery atherosclerotic stroke.

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.

Lectin-like oxidized low-density lipoprotein receptor 1 and matrix metalloproteinase expression in ruptured and unruptured multiple dissections of distal middle cerebral artery: case report.

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.

Nitric oxide suppresses cerebral vasomotion by sGC-independent effects on ryanodine receptors and voltage-gated calcium channels.

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.

Alterations in the modulation of cerebrovascular tone and blood flow by nitric oxide synthases in SHRsp with stroke.

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.