cyclic-gmp and Vasospasm--Intracranial

Soluble guanylyl cyclases (sGCs) and Ras homolog gene family, member A (rhoA)/Ras homolog gene family kinase(rho-kinase) plays a role in vascular smooth muscle relaxation in subarachnoid hemorrhage (SAH). It is of interest to examine the effect of MLB on rhoA/ROCK and sGC/cGMP/PKG expression.. A rodent SAH model was employed. Tissue samples were for sGC α 1, sGC β 1, PKG, rhoA, ROCK (Western blot), and cGMP (ELISA) measurement.. MLB morphologically improved convolution of the internal elastic lamina, distortion of endothelial wall, and necrosis of the smooth muscle in the SAH rats. Expressed cGMP, sGC α 1, sGC β 1, and PKG in the SAH groups were reduced (P < 0.01), and MLB precondition significantly induced cGMP, sGCα1, sGCβ1, and PKG. L-NAME reversed the vasodilation effect of MLB, reduced the bioexpression of PKG and cGMP (P < 0.01), and tends to reduce sGCα1 level and induce rhoA, ROCK level in MLB precondition + SAH groups.. These results demonstrate that sGC/cGMP/PKG and NO/ET pathways play pivotal roles in SAH-induced vasospasm. Through activating sGC/cGMP/PKG pathway and partially by inactivating rho-kinase in a NO-dependent mechanism, MLB shows promise to be an effective strategy for the treatment of this disease entity.

Topics: Animals; Biological Transport, Active; Camphanes; Cyclic GMP; Cyclic GMP-Dependent Protein Kinases; Disease Models, Animal; Drugs, Chinese Herbal; Guanylate Cyclase; Male; Panax notoginseng; Rats; Rats, Sprague-Dawley; Receptors, Cytoplasmic and Nuclear; Salvia miltiorrhiza; Signal Transduction; Soluble Guanylyl Cyclase; Vasospasm, Intracranial

Cerebral vasospasm is an independent predictor of poor outcome after subarachnoid hemorrhage (SAH). The nitric oxide-cyclic guanosine monophosphate (NO-cGMP) vasodilatory pathway is strongly implicated in its pathophysiology. Preliminary studies suggest that phosphodiesterase 5 (PDE5), an enzyme that degrades cGMP, may play a role because the PDE5 inhibitor sildenafil was found to reduce vasospasm after SAH. However, several questions that are critical when considering translational studies remain unanswered.. To elucidate the mechanism of action of sildenafil against vasospasm and to assess whether sildenafil attenuates SAH-induced neuronal cell death, improves functional outcome after SAH, or causes significant physiological side effects when administered at therapeutically relevant doses.. SAH was induced via endovascular perforation in male C57BL6 mice. Beginning 2 hours later, mice received sildenafil citrate (0.7, 2 or 5 mg/kg orally twice daily) or vehicle. Neurological outcome was assessed daily. Vasospasm was determined on post-SAH day 3. Brain PDE5 expression and activity, cGMP content, neuronal cell death, arterial blood pressure, and intracranial pressure were examined.. We found that PDE5 activity (but not expression) is increased after SAH, leading to decreased cGMP levels. Sildenafil attenuates this increase in PDE5 activity and restores cGMP levels after SAH. Post-SAH initiation of sildenafil was found to decrease vasospasm and neuronal cell death and markedly improve neurological outcome without causing significant physiological side effects.. Sildenafil, a US Food and Drug Administration-approved drug with a proven track record of safety in humans, is a promising new therapy for vasospasm and neurological deficits after SAH.

Topics: Animals; Blood Pressure; Cell Death; Cyclic GMP; Cyclic Nucleotide Phosphodiesterases, Type 5; Disease Models, Animal; Dose-Response Relationship, Drug; Drug Administration Schedule; Enzyme-Linked Immunosorbent Assay; Extremities; In Situ Nick-End Labeling; Intracranial Pressure; Male; Mice; Mice, Inbred C57BL; Motor Activity; Movement; Neurons; Phosphodiesterase 5 Inhibitors; Piperazines; Purines; Recovery of Function; Severity of Illness Index; Sildenafil Citrate; Subarachnoid Hemorrhage; Sulfones; Vasospasm, Intracranial

The pathogenesis of cerebral vasospasm is likely to be multifactorial. Strong evidence has indicated that decreasing levels of NO after SAH seem to be important. A PDE-V inhibitor, tadalafil, theoretically increases NO levels. Our study investigated the vasodilatory efficacy of tadalafil on the cerebral arteries with measurement of basilar artery diameters on angiography.. We used 42 male Wistar-Albino rats to test our hypothesis. They were assigned randomly into the following seven groups: group 1: control (only saline), group 2: SAH only (killed on day 2), group 3: SAH + tadalafil (killed on day 2), group 4: SAH only (killed on day 4), group 5: SAH + tadalafil (killed on day 4), group 6: saline + tadalafil (killed on day 2) and group 7: saline + tadalafil (killed on day 4). The three different parts of basilar artery diameters were measured angiographically.. There were statistically significant differences between the SAH and SAH groups treated with tadalafil at days 2 and 4. Comparison between control and tadalafil groups showed no significant differences. This result indicated that tadalafil has a vasodilatory effect on vasoconstricted arteries, but no effect on normal basilar arteries.. Our study results showed that tadalafil has a vasodilatory effect on both acute and chronic periods of cerebral vasospasm. We also concluded that cerebral angiography can be used safely for investigation of cerebral vasospasm in animal studies.

Topics: Animals; Basilar Artery; Carbolines; Cerebral Angiography; Cerebral Arteries; Cerebrovascular Circulation; Cyclic GMP; Disease Models, Animal; Drug Administration Schedule; Male; Nitric Oxide; Phosphodiesterase Inhibitors; Rats; Rats, Wistar; Subarachnoid Hemorrhage; Tadalafil; Time Factors; Treatment Outcome; Vasodilation; Vasodilator Agents; Vasospasm, Intracranial

Tetramethylpyrazine (TMP), an ingredient of Chinese herbal Szechwan lovage rhizome, shows vasorelaxant effect. Cerebral vasospasm (CVS) after subarachnoid hemorrhage (SAH) is associated with high mortality and morbidity. Here, we evaluated the effect of TMP in a model of CVS and sought to identify the underlying mechanisms of action. A rabbit SAH model was established by injection of the autoblood via cisterna magna. Cerebral blood flow and arterial diameter were measured by Transcranial Doppler (TCD) and Computed Tomography Angiography (CTA). Expression of eNOS and PDE-V in basilar artery (BA) was assessed by western blots. Levels of nitric oxide (NO) in plasma and cerebral spinal fluid, and of intra-endothelium Ca(2+) were measured. Significantly reduced diameter and accelerated blood flow velocity were detected in BAs of SAH animals (P<0.05 vs. sham group). Expression of eNOS and NO was increased, and PDE-V expression was reduced by TMP.TMP ameliorated cerebral vasospasm (P<0.05 vs. SAH group), and L-NAME (a NOS inhibitor) partly abrogated the effects of TMP. TMP induced a dose-dependent increase of intra-endothelium Ca(2+). The current results demonstrated that the vasorelaxant effect of TMP was at least in part via regulation of NO/cGMP signaling.

Topics: Animals; Basilar Artery; Blotting, Western; Calcium Signaling; Cerebral Angiography; Cyclic GMP; Cyclic Nucleotide Phosphodiesterases, Type 5; Disease Models, Animal; Endothelial Cells; Male; Microscopy, Confocal; Nitric Oxide; Nitric Oxide Synthase Type III; Pyrazines; Rabbits; Signal Transduction; Subarachnoid Hemorrhage; Tomography, X-Ray Computed; Ultrasonography, Doppler, Transcranial; Vasodilator Agents; Vasospasm, Intracranial

Myosin light chain phosphatase plays a critical role in modulating smooth muscle contraction in response to a variety of physiologic stimuli. A downstream target of the RhoA/Rho-kinase and nitric oxide (NO)/cGMP/cyclic GMP-dependent kinase (cGKI) pathways, myosin light chain phosphatase activity reflects the sum of both calcium sensitization and desensitization pathways through phosphorylation and dephosphorylation of the myosin phosphatase targeting subunit (MYPT1). As cerebral blood flow is highly spatio-temporally modulated under normal physiologic conditions, severe perturbations in normal cerebral blood flow, such as in cerebral vasospasm, can induce neurological deficits. In nonpermeabilized cerebral vessels stimulated with U-46619, a stable mimetic of endogenous thromboxane A2 implicated in the etiology of cerebral vasospasm, we observed significant increases in contractile force, RhoA activation, regulatory light chain phosphorylation, as well as phosphorylation of MYPT1 at Thr-696, Thr-853, and surprisingly Ser-695. Inhibition of nitric oxide signaling completely abrogated basal MYPT1 Ser-695 phosphorylation and significantly increased and potentiated U-46619-induced MYPT1 Thr-853 phosphorylation and contractile force, indicating that NO/cGMP/cGKI signaling maintains basal vascular tone through active inhibition of calcium sensitization. Surprisingly, a fall in Ser-695 phosphorylation did not result in an increase in phosphorylation of the Thr-696 site. Although activation of cGKI with exogenous cyclic nucleotides inhibited thromboxane A2-induced MYPT1 membrane association, RhoA activation, contractile force, and regulatory light chain phosphorylation, the anticipated decreases in MYPT1 phosphorylation at Thr-696/Thr-853 were not observed, indicating that the vasorelaxant effects of cGKI are not through dephosphorylation of MYPT1. Thus, thromboxane A2 signaling within the intact cerebral vasculature induces "buffered" vasoconstrictions, in which both the RhoA/Rho-kinase calcium-sensitizing and the NO/cGMP/cGKI calcium-desensitizing pathways are activated.

Topics: 15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; Animals; Blood Flow Velocity; Cerebral Arteries; Cyclic GMP; Cyclic GMP-Dependent Protein Kinases; Enzyme Activation; Male; Muscle Contraction; Myosin-Light-Chain Phosphatase; Nitric Oxide; Phosphorylation; Protein Phosphatase 1; Rabbits; Rats; rho-Associated Kinases; rhoA GTP-Binding Protein; Signal Transduction; Thromboxane A2; Vasoconstriction; Vasoconstrictor Agents; Vasospasm, Intracranial

The aim of this study was to investigate the ability of a SERM, RLX, to prevent vasospasm in a rabbit model of SAH.. Thirty-four New Zealand white rabbits were allocated into 3 groups randomly. Subarachnoid hemorrhage was induced by injecting autologous blood into the cisterna magna. The treatment groups were as follows: (1) sham operated (no SAH [n = 12]), (2) SAH only (n = 12), and (3) SAH plus RLX (n = 10). Basilar artery lumen areas and arterial wall thickness were measured to assess vasospams in all groups.. There was a statistically significant difference between the mean basilar artery cross-sectional areas and the mean arterial wall thickness measurements of the control and SAH-only groups (P < .05). The difference between the mean basilar artery cross-sectional areas and the mean arterial wall thickness measurements in the RLX-treated group was statistically significant (P < .05). The difference between the SAH group and the SAH + RLX group was also statistically significant (P < .05).. These findings demonstrate that RLX has marked vasodilatatory effect in an experimental model of SAH in rabbits. This observation may have clinical implications suggesting that this SERM drug could be used as possible anti-vasospastic agent in patients without major adverse effects.

Topics: Animals; Basilar Artery; Calcium Channels, L-Type; Cerebral Arteries; Cyclic GMP; Disease Models, Animal; Endothelial Cells; Estrogen Receptor alpha; Male; Muscle, Smooth, Vascular; Myosin Light Chains; Rabbits; Raloxifene Hydrochloride; Selective Estrogen Receptor Modulators; Subarachnoid Hemorrhage; Treatment Outcome; Vasodilation; Vasodilator Agents; Vasospasm, Intracranial

Cerebral vasospasm remains a major cause of morbidity and mortality in patients with subarachnoid hemorrhage. Heme oxygenase-1 (HO-1) is an oxidative stress-inducible enzyme with multiple protective functions against vascular and neurological diseases, including delayed cerebral vasospasm. In the present study, intravenous administration (i.v.) of nicaraven (1 mg/kg/min, for 2 days after subarachnoid hemorrhage) ameliorated delayed cerebral vasospasm in rat subarachnoid hemorrhage models, marked synergistic induction of HO-1 protein (> 2.5-fold than 'subarachnoid hemorrhage with saline i.v.'), and elicited a rapid increase of cGMP accumulation in the basilar arteries. In the sham-operated rats, nicaraven could not induce HO-1 expression. Antisense HO-1 oligodeoxynucleotides abrogated this HO-1 induction and the antivasospastic effect of nicaraven. In vitro study using Hela cells, nicaraven enhanced the human HO-1 promoter (-4.5 kbp) activity, which was pre-activated with the blood component oxyhemoglobin to mimic the ability of subarachnoid hemorrhage. These results suggest that this enhanced HO-1 expression through a combination of pathological state and pharmacological agent could be an effective strategy to improve the prognosis of heme- and oxidative stress-induced diseases, such as delayed cerebral vasospasm.

Topics: Animals; Basilar Artery; Cerebral Angiography; Cyclic GMP; Enzyme Induction; Gene Expression Regulation, Enzymologic; Heme Oxygenase-1; Humans; Male; Niacinamide; Oligodeoxyribonucleotides, Antisense; Oxyhemoglobins; Promoter Regions, Genetic; Rats; Rats, Sprague-Dawley; Subarachnoid Hemorrhage; Time Factors; Vasospasm, Intracranial

Intracranial aneurysmal rupture is the common cause of spontaneous subarachnoid haemorrhage (SAH). This haemorrhage is typically diffuse and located in extracerebral subarachnoid space in which main cerebral arterial branches are situated. The intimate and long-term contact of arterial wall and blood products in the closed space causes the cerebral vasospasm as a serious and frequent complication of SAH. It is connected with significant morbidity and mortality due to developing of focal cerebral ischaemia and subsequently cerebral infarction. The aim of our experimental research was to create the animal model of vasospasm using the femoral artery due to examination of reduced basic dilator activity cause in arterial wall after SAH. The important characteristic of major cerebral arteries is their localization in the closed subarachnoid space which enables their to have long-term contact with blood products after haemorrhage. Thirty six femoral arteries (FA) of eighteen female rats weighing about 300 g were used. In vivo, femoral arteries are microsurgically prepared in both inguinal regions in all rats. Eighteen arteries were encompassed by polytetrafluoroethylene (PTFE) material forming closed tube and autologous blood was injected in the tube around the arterial wall. Additional eighteen arteries, as a control group, were also put in PTFE tube but without exposing to the blood. All rats are left to live for eight days. Afterwards, rats were sacrificed and their arteries were in vitro examined including an isometric tension measurement and histological changes analysis. The tension was measured during application of vasoconstrictors and vasodilatators (nitric oxide, NO). FA exposed to periadventitial blood exhibit hyper reactivity to constrictors (KCl, phenylephrine, acetylcholine) compared to control group. It was also found that NO donor (sodium nitroprusside) diminished arterial spasm induced by blood and vasoconstrictors. In conclusion, FA can be used as a model for vasospasm correlating with cerebral vasospasm after SAH and therefore this model can be utilized in future experiments assessing cerebral vasospasm. The reduced basic dilator activity of spastic femoral artery is caused by an absence of gaseous messenger NO next to the arteries but not by diminished response vasculature to NO. Absence of NO after SAH probably causes the reduced basic dilator activity of cerebral arteries as well. The guanylate-cyclase level in the arterial wall is conseque

Topics: Adenomatous Polyposis Coli; Animals; Cyclic GMP; Femoral Artery; Hemorrhage; Loss of Heterozygosity; Microsatellite Repeats; Models, Animal; Nitric Oxide; Polytetrafluoroethylene; Rats; Subarachnoid Hemorrhage; Vasodilator Agents; Vasospasm, Intracranial

Vasospasm after subarachnoid hemorrhage (SAH) may result from hemoglobin-mediated removal of nitric oxide (NO) from the arterial wall. We tested the ability of the long-acting, water-soluble, NO donor (Z)-1-[N-(2-aminoethyl)-N-(2-ammonioethyl)amino]diazen-1-1,2-diolate (DETA/NO), delivered via continuous intracisternal infusion, to prevent vasospasm in a nonhuman primate model of SAH.. First, vasorelaxation in response to DETA/NO was characterized in vitro by using monkey basilar artery rings under isometric tension. Next, monkeys were randomized to undergo angiography, unilateral SAH, and no treatment (SAH only, n = 4) or treatment with DETA/NO (1 mmol/L, 12 ml/d, n = 4) or decomposed DETA/NO (at the same dose, n = 4). Vasospasm was assessed by angiography, which was performed on Day 0 and Day 7. Levels of cyclic adenosine monophosphate and cyclic guanosine monophosphate (cGMP) were measured in cerebral arteries on Day 7.. DETA/NO produced significant relaxation of monkey arteries in vitro, which reached a maximum at concentrations of 10(-5) mol/L. In monkeys, angiography demonstrated significant vasospasm of the right intradural cerebral arteries in all three groups, with no significant difference in vasospasm among the groups (P > 0.05, analysis of variance). The ratios of cGMP or cyclic adenosine monophosphate levels in the right and left middle cerebral arteries were not different among the groups (P > 0.05, analysis of variance). There was no significant correlation between arterial cGMP contents and the severity of vasospasm.. DETA/NO did not prevent vasospasm. There was no correlation between the severity of vasospasm and cyclic adenosine monophosphate and cGMP levels in the cerebral arteries. These results suggest that events downstream of cyclic nucleotides may be abnormal during vasospasm.

Topics: Animals; Cerebral Angiography; Cyclic AMP; Cyclic GMP; Disease Models, Animal; Macaca fascicularis; Middle Cerebral Artery; Nitric Oxide Donors; Severity of Illness Index; Subarachnoid Hemorrhage; Triazenes; Vasospasm, Intracranial

Hemoglobin contributes to vasospasm after subarachnoid hemorrhage. One mechanism may involve binding of nitric oxide, destruction of nitric oxide, or both. Support for this mechanism would be evidence that nitric oxide donors prevent vasospasm. This study attempted to provide such evidence.. A randomized, blinded study was conducted in which 13 monkeys underwent cerebral angiography and creation of a right subarachnoid hemorrhage. Subcutaneous osmotic pumps were implanted to deliver sodium nitroprusside (n = 7) or vehicle (n = 6) via catheters into the right basal cisterns. Seven days later, angiography was repeated, and the animals were humanely killed. Levels of cyclic nucleotides, hemoglobins, and thiocyanate were measured.. Significant vasospasm of the right middle cerebral artery was present in animals treated with sodium nitroprusside (35 +/- 22% reduction in diameter, P < 0.05, paired t test) and placebo (28 +/- 20% reduction, P < 0.05, not significantly different from nitroprusside group by unpaired t test). Adequate delivery of sodium nitroprusside was supported by the finding of a significant increase in cyclic guanosine monophosphate levels in the cerebral arteries of treated animals compared with placebo (P < 0.05, unpaired t test). Thiocyanate was not present in significantly increased amounts in animals treated with nitroprusside, although this group did display elevated concentrations of nitrosyl hemoglobin (measured by electron paramagnetic resonance spectroscopy) and cyanomethemoglobin (measured by spectrophotometry) in the cerebrospinal fluid on Day 7.. The lack of effect of sodium nitroprusside was not the result of inadequate drug delivery because cyclic guanosine monophosphate levels were significantly increased in vasospastic arteries. Vasospasm may not have been prevented because of a toxic effect of sodium nitroprusside metabolites, involvement of smooth muscle relaxation or contraction processes downstream of cyclic guanosine monophosphate, or both.

Topics: Animals; Brain; Cerebral Angiography; Cerebral Arteries; Cisterna Magna; Cyclic GMP; Hemoglobins; Injections; Macaca fascicularis; Nitric Oxide Donors; Nitroprusside; Treatment Failure; Vasospasm, Intracranial

Cyclic GMP (cGMP) mediates smooth muscle relaxation in the central nervous system. In subarachnoid hemorrhage (SAH), decreases in intrinsic nitric oxide (NO) cause cerebral vasospasms due to the regulation of cGMP formation by NO-mediated pathways. As phosphodiesterase type V (PDE V) selectively hydrolyzes cGMP, we hypothesized that PDE V may function in the initiation of vasospasm. This study sought to identify the altered PDE V expression and activity in the vasospastic artery in a canine SAH model. We also used this system to examine possible therapeutic strategies to prevent vasospasm. Using a canine model of SAH, we induced cerebral vasospasm in the basilar artery (BA). Following angiographic confirmation of vasospasm on day 7, PDE V expression was immunohistochemically identified in smooth muscle cells of the vasospastic BA but not in cells of a control artery. The isolation of PDE enzymes using a sepharose column confirmed increased PDE V activity in the vasospastic artery only through both inhibition studies, using the highly selective PDE V inhibitor, sildenafil citrate, and Western blotting. Preliminary in vivo experiment using an oral PDE V inhibitor at 0.83 mg kg(-1) demonstrated partial relaxation of the spastic BA. PDE V activity was increased from control levels within the BA seven days after SAH. PDE V expression was most prominent in smooth muscle cells following SAH. These results suggest that clinical administration of a PDE V inhibitor may be a useful therapeutic tool in the prevention of vasospasm following SAH.

Topics: 3',5'-Cyclic-GMP Phosphodiesterases; Animals; Blotting, Western; Cerebral Angiography; Cerebral Arteries; Cyclic GMP; Cyclic Nucleotide Phosphodiesterases, Type 5; Disease Models, Animal; Dogs; Immunohistochemistry; Male; Myocytes, Smooth Muscle; Phosphodiesterase Inhibitors; Phosphoric Diester Hydrolases; Piperazines; Purines; Sildenafil Citrate; Subarachnoid Hemorrhage; Sulfones; Time Factors; Vasospasm, Intracranial