cyclic-gmp and Subarachnoid-Hemorrhage

Subarachnoid hemorrhage (SAH) usually results from ruptured aneurysm, but how leaked hemoglobin regulates the microcirculation in the pathophysiology of early brain injury after SAH is still unclear. In the present study, we sought to investigate the role and possible mechanism of hemoglobin induced pericyte phenotype transformation in the regulation of microcirculation after SAH. Endovascular perforation SAH rat model, brain slices and cultured pericytes were used, and intervened with endothelial nitric oxide synthase (eNOS) antagonist L-NNA and its agonist scutellarin, hemoglobin, DETA/NO (nitric oxide(NO) donor), PITO (NO scavenger), 8-Br-cGMP (cGMP analog). We found modulating eNOS regulated pericyte α-SMA phenotype transformation, microcirculation, and neurological function in SAH rats. Modulating eNOS also affected eNOS expression, eNOS activity and NO availability after SAH. In addition, we showed hemoglobins penetrated into brain parenchyma after SAH. And hemoglobins significantly reduced the microvessel diameters at pericyte sites, due to the effects of hemoglobin inducing α-SMA expressions in cultured pericytes and brain slices via inhibiting NO/cGMP pathway. In conclusion, pericyte α-SMA phenotype mediates acute microvessel constriction after SAH possibly by hemoglobin suppressing NO/cGMP signaling pathway. Therefore, by targeting the eNOS and pericyte α-SMA phenotype, our present data may shed new light on the management of SAH patients.

Topics: Actins; Animals; Cyclic GMP; Disease Models, Animal; Enzyme Activation; Gene Expression; Hemoglobins; Male; Microcirculation; Nitric Oxide; Nitric Oxide Synthase Type III; Pericytes; Phenotype; Rats; Signal Transduction; Subarachnoid Hemorrhage

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

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

Cerebrovascular dysfunction after subarachnoid hemorrhage (SAH) may contribute to ischemia, but little is known about the contribution of intracerebral arterioles. In this study, the authors tested the hypothesis that SAH inhibits the vascular reactivity of intracerebral arterioles and documented the time course of this dysfunction.. Subarachnoid hemorrhage was induced using an endovascular filament model in halothane-anesthetized male Sprague-Dawley rats. Penetrating intracerebral arterioles were harvested 2, 4, 7, or 14 days postinsult, cannulated using a micropipette system that allowed luminal perfusion and control of luminal pressure, and evaluated for reactivity to vasodilator agents.. Spontaneous tone developed in all pressurized (60 mm Hg) intracerebral arterioles harvested in this study (from 66 rats), with similar results in the sham and SAH groups. Subarachnoid hemorrhage did not affect dilation responses to acidic pH (6.8) but led to a persistent impairment of endothelium-dependent dilation responses to adenosine triphosphate (p < 0.01), as well as a transient attenuation (p < 0.05) of vascular smooth muscle-dependent dilation responses to adenosine, sodium nitroprusside, and 8-Br-cyclic guanosine monophosphate (cGMP). Impairment of NO-mediated dilation was more sustained than adenosine- and 8-Br-cGMP-induced responses (up to 7 days postinsult compared with 2 days). All smooth muscle-dependent responses returned to sham levels by 14 days after SAH.. Subarachnoid hemorrhage led to a persistent impairment of endothelium-dependent dilation and a transient attenuation of vascular smooth muscle-dependent dilation responses to adenosine. Impairment of NO-mediated dilation occurred when the response to cGMP was intact, suggesting a change in cGMP levels rather than an alteration in intracellular mechanisms downstream from cGMP.

Topics: Adenosine; Adenosine Triphosphate; Animals; Arterioles; Blood Pressure; Carbon Dioxide; Cerebrovascular Circulation; Cyclic GMP; Endothelium, Vascular; Hydrogen-Ion Concentration; Male; Muscle Tonus; Muscle, Smooth, Vascular; Nitric Oxide; Nitroprusside; Oxygen; Rats; Rats, Sprague-Dawley; Subarachnoid Hemorrhage; Vasodilation; Vasodilator Agents

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

We designed the present experiments to investigate the involvement of endogenous nitric oxide synthase (NOS) inhibitors, dimethylarginine dimethylaminohydrolase (DDAH) as a hydrolyzing enzyme of the NOS inhibitors, NOS, arginase which shares l-arginine as a common substrate with NOS, and endothelin-1 (ET-1) in the pulmonary dysfunction after induction of experimental subarachnoid hemorrhage (SAH) in the rabbit. SAH was induced by injecting autologous blood into the cisterna magna, and controls were injected with saline. On day 2, pulmonary arteries were isolated for determinations. A significant impairment of the endothelium-dependent relaxation (EDR) caused by acetylcholine was found in 20 cases (43.5%) out of 46 SAH animals, and the same animals exhibited accompanying the significantly impaired cyclic GMP production, accumulated endogenous NOS inhibitors, attenuated DDAH activity, enhanced arginase activity and accumulated ET-1 within the vessel wall. Meanwhile, there were no differences in endothelial NOS activity per se and sodium nitroprusside-induced relaxation between the animals with an impaired EDR and those without such a change. ET-1 content within aortic wall was increased with concomitant decrease in cyclic GMP production after the intraperitoneal application of authentic monomethylarginine as a NOS inhibitor in the rat. The current results suggest that accumulated endogenous NOS inhibitors and enhanced arginase activity possibly bring about the impaired NO production, thereby attenuating the EDR and contributing to the accumulation of ET-1 within the vessel wall. The accumulated endogenous NOS inhibitors at least partly result from the decreased DDAH activity. These alterations may be relevant to the pulmonary dysfunction after induction of SAH.

Topics: Acetylcholine; Amidohydrolases; Animals; Arginase; Cyclic GMP; Cyclooxygenase Inhibitors; Dose-Response Relationship, Drug; Endothelin-1; Indomethacin; Lung Diseases; Male; Models, Biological; Nitric Oxide Donors; Nitric Oxide Synthase; Nitroarginine; Nitroprusside; omega-N-Methylarginine; Oxadiazoles; Pulmonary Artery; Quinoxalines; Rabbits; Rats; Rats, Sprague-Dawley; Subarachnoid Hemorrhage; Vasoconstriction

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

This study was aimed to evaluate the influence of an antagonist of heme oxygenase, zinc protoporphyrin IX (ZnPPIX), on the production of endogenous carbon monoxide (CO) and the secondary cerebral injury after subarachnoid hemorrhage (SAH). Wistar rats were divided into non-SAH, SAH, and ZnPPIX groups. Autologus arterial hemolysate was injected into rat cisterna magna to induce SAH. CO and cyclic guanosine monophosphate (cGMP) levels in the brain, and lactate dehydrogenase (LDH) activity in serum were determined 24 hours and 72 hours after cisternal injection. It was found that 24 hours and 72 hours after SAH, the CO contents in SAH group were increased by 20.76% and 37.36%, respectively. CO content in ZnPPIX group was statistically lower than that in SAH group. No obvious change of cGMP content in SAH group was found. However, cGMP content in ZnPPIX group was lower than that in SAH group. Serum LDH activity increased significantly after induction of SAH. LDH activity in ZnPPIX group increased to a greater extent. It was concluded that ZnPPIX aggravates the cerebral injury secondary to experimental SAH by inhibiting the production of endogenous CO. The activation of HO/CO pathway is an intrinsic protective mechanism against cerebral ischemic injury after SAH.

Topics: Animals; Brain Ischemia; Carbon Monoxide; Cyclic GMP; Heme Oxygenase (Decyclizing); L-Lactate Dehydrogenase; Protoporphyrins; Rats; Rats, Wistar; Subarachnoid Hemorrhage; Time Factors

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

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

This study was designed to determine the effect of subarachnoid hemorrhage (SAH) on potassium (K+) channels involved in relaxations of cerebral arteries to nitrovasodilators. The effects of K+ channel inhibitors on relaxations to 3-morpholinosydnonimine (SIN-1) and sodium nitroprusside (SNP) were studied in rings of basilar arteries obtained from untreated dogs and dogs exposed to SAH. The levels of cyclic GMP were measured by radioimmunoassay. In rings without endothelium, concentration-dependent relaxations to SIN-1 (10(-9)-10(-4) mol/L) and SNP (10(-9)-10(-4) mol/L) were not affected by SAH, whereas increase in cyclic GMP production stimulated by SIN-1 (10(-6) mol/L) was significantly suppressed after SAH. The relaxations to SIN-1 and SNP were reduced by charybdotoxin (CTX: 10(-7) mol/L), a selective Ca(2+)-activated K+ channel inhibitor, in both normal and SAH arteries; however, the reduction of relaxations by CTX was significantly greater in SAH arteries. By contrast, the relaxations to these nitrovasodilators were not affected by glyburide (10(-5) mol/L), an ATP-sensitive K+ channel inhibitor, in both normal and SAH arteries. These findings suggest that in cerebral arteries exposed to SAH, CA(2+)-activated K+ channels may play a compensatory role in mediation of relaxations to nitric oxide. This may help to explain mechanisms of relaxations to nitrovasodilators in arteries with impaired production of cyclic GMP.

Topics: 4-Aminopyridine; Animals; Basilar Artery; Charybdotoxin; Cyclic GMP; Dogs; Female; Humans; Male; Molsidomine; Nitroprusside; Potassium Channels; Subarachnoid Hemorrhage; Uridine Triphosphate; Vasodilation; Vasodilator Agents

The authors investigated the changes and the potential of cyclic nucleotide-dependent signal transduction, which induces smooth muscle relaxation, in the basilar artery with severe vasospasm in dogs with double experimental subarachnoid hemorrhage (SAH) to explore at which biochemical level the arterial dilative capability was impaired. The amount of cyclic adenosine and guanosine monophosphates (cAMP and cGMP) decreased significantly in the basilar artery after SAH. The activities of adenylate and guanylate cyclases also were decreased significantly in the smooth muscle cells of the basilar artery 4 days after SAH. In addition to the failure of the pathways to produce cyclic nucleotides, the activities of cAMP- and cGMP-dependent protein kinases, which are representative actual enzymes that amplify the signal for vascular dilation, also significantly decreased together with the almost total loss of activation by cyclic nucleotides in the same basilar artery after SAH. It was revealed that the system for smooth muscle relaxation was impaired severely in the cerebral arteries with severe vasospasm after SAH, on the biochemical basis of significantly less vasodilative capability and in several of the steps to produce the cyclic nucleotides of intracellular signal transduction.

Topics: Adenylyl Cyclases; Animals; Basilar Artery; Cyclic AMP; Cyclic AMP-Dependent Protein Kinases; Cyclic GMP; Cyclic GMP-Dependent Protein Kinases; Dogs; Female; Guanylate Cyclase; Ischemic Attack, Transient; Kinetics; Male; Muscle, Smooth, Vascular; Signal Transduction; Subarachnoid Hemorrhage; Vasodilation

Nitric oxide (NO) increases 3',5'-cyclic guanosine monophosphate (cGMP) in vascular smooth muscle and increases cerebral blood flow (CBF). In early stages of cerebral ischemia, NO plays a beneficial role in sustaining CBF. Subarachnoid hemorrhage (SAH), one of the main causes of ischemia, may impair vascular reactivity to NO. To test the hypothesis, 48 h after SAH was induced in rats, we examined the CBF response to the NO donor, SIN-1 (3-morpholinosydnonimine). We measured CBF by laser-Doppler flowmetry in association with: (1) intracarotid injection (for 30 min) of SIN-1 (1.5 mg/kg), 8-bromo-cGMP (7.5 mg/kg), papaverin (1.5 mg/kg) or vehicle; (2) cortical superfusion (for 90 min) of SIN-1 (10(-5) M) or vehicle through the cranial window. Hypotension produced by these vasodilators was controlled with phenylephrine. Vehicle alone did not change CBF throughout the measurement. Intracarotid infusion of SIN-1 (n = 6/group) increased CBF up to 128.6 +/- 3.9% and 111.9 +/- 2.9% in the control group and the SAH group, respectively. SAH significantly attenuated the response (P < 0.05, ANOVA). SAH did not affect the CBF increases elicited by intracarotid administration of cGMP or papaverin, or cortical superfusion of SIN-1. We conclude that during chronic vasospasm SAH disturbs the pathway between NO release and cGMP production in large cerebral arteries. The impairment accounts for the fragility of the brain in the face of ischemia following SAH.

Topics: Animals; Brain; Cerebral Arteries; Cerebral Cortex; Cerebrovascular Circulation; Cyclic GMP; Injections, Intra-Articular; Male; Molsidomine; Muscle, Smooth, Vascular; Nitric Oxide; Papaverine; Phenylephrine; Rats; Rats, Sprague-Dawley; Regional Blood Flow; Subarachnoid Hemorrhage; Time Factors; Vasodilator Agents

We hypothesize that the interaction between protein kinase C (PKC) and nitric oxide (NO) plays a role in the modulation of cerebral vascular tone, and the disturbance of this interaction following subarachnoid hemorrhage (SAH) results in vasospasm. To prove this hypothesis with direct evidence, PKC activities of smooth muscle cells of canine basilar arteries in the control and in the SAH groups were measured by an enzyme immunoassay method. N omega-nitro-L arginine (L-NA), an inhibitor of NO production, enhanced PKC activity. This enhancement was inhibited neither by 8-bromo-guanosine 3',5'-cyclic monophosphate (8-bromo-cGMP) nor SIN-1, a NO releasing agent. PKC activity in the SAH was significantly higher than in the control; however, no further enhancement was produced with L-NA. In the SAH, PKC activity was not inhibited either by 8-bromo-cGMP or SIN-1. We conclude that NO maintains an appropriate vascular tone through inactivation of PKC, and that this effect is disturbed following SAH, resulting in PKC-dependent vascular contraction, such as vasospasm. On the other hand, once PKC has been activated, NO precursors do not inhibit PKC. These facts indicate NO inactivates PKC through the inhibition of phosphatidylinositol breakdown.

Topics: Animals; Basilar Artery; Cyclic GMP; Dogs; Enzyme Activation; Enzyme Inhibitors; Female; Immunoenzyme Techniques; Ischemic Attack, Transient; Male; Molsidomine; Muscle, Smooth, Vascular; Nitric Oxide; Nitric Oxide Synthase; Nitroarginine; Protein Kinase C; Reference Values; Subarachnoid Hemorrhage

This study was undertaken to investigate how protein kinase C (PKC) and nitric oxide (NO) interact to regulate the vascular tone, and how their interaction contributes to the development of vasospasm after subarachnoid hemorrhage (SAH). For these purposes, vasospasm was conducted with a canine model. We investigated the following subjects with arteries from intact animals and those from the SAH model, and compared the results between the two; tension at rest of isometric tension study, the effect of PKC inhibitors and of an inhibitor of NO synthesis on the tension at rest, and levels of guanosine 3',5'-cyclic monophosphate (cGMP) as an indicator of NO production. The tension at rest was enhanced in the artery from the SAH model compared to that from intact animals, and it was PKC-dependent. Arteries from intact animals but not those from the SAH model developed tonic tensions by NO inhibitors, and these tonic tensions were suppressed by PKC inhibitors, and also by cGMP. An enzyme immunoassay revealed a decreased cGMP level in the SAH model. The evidence indicates that NO exerts a negative feedback control on PKC activation. Subarachnoid hemorrhage interferes with this feedback control, resulting in PKC-dependent enhanced vascular tone and vasospasm.

Topics: 1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; Alkaloids; Animals; Basilar Artery; Cyclic GMP; Dogs; Enzyme Inhibitors; Female; In Vitro Techniques; Ischemic Attack, Transient; Isometric Contraction; Isoquinolines; Male; Models, Cardiovascular; Muscle Tonus; Muscle, Smooth, Vascular; Nitric Oxide; Piperazines; Protein Kinase C; Staurosporine; Subarachnoid Hemorrhage

Nitroglycerin, as well as nitric oxide, causes hyperpolarization and cGMP elevation in vascular smooth muscle cells. It is unknown whether nitroglycerin ameliorates vasospasm by an increase in cGMP levels after subarachnoid hemorrhage (SAH). The purpose of the present study was to measure the levels of both cGMP and cAMP in the cerebral arteries and parietal cerebral cortices in a primate model and to determine the effect of nitroglycerin on vasospasm after SAH.. Chronic vasospasm was induced by clot placement around the right middle cerebral artery (MCA). Seven days after the surgery, angiography was repeated and either nitroglycerin (3 micrograms/kg per hour) or saline was administered intravenously. Angiography and regional cerebral blood flow (rCBF) measurements in the bilateral parietal cortices were performed before and after each treatment. Both cGMP and cAMP levels were measured in the cerebral arteries and bilateral parietal cortices.. A significant vasospasm occurred in the cerebral arteries on both sides, more prominently on the right side. Concomitantly, rCBF on the right side was significantly decreased (P < .05). In the right MCA, cGMP levels were significantly lower than in the normal MCA (P < .05). After the administration of nitroglycerin for 3 hours, the cerebral vessels were significantly dilated on both sides (P < .05), and rCBF was significantly increased on the right side (P < .05) but not on the left side. Although depressed cGMP levels in the right MCA were not recovered by nitroglycerin, a significant increase in cGMP levels was observed in the basilar artery (P < .05). In both parietal cortices, cGMP levels were significantly decreased after SAH (P < .05) and unchanged after nitroglycerin treatment. There were no significant changes in cAMP levels in SAH and after nitroglycerin treatment.. The vasodilator effect of nitroglycerin in spastic MCA may not be mediated by an increase in cGMP levels, suggesting an involvement of hyperpolarization of the smooth muscle cells. Given the increase in rCBF, nitroglycerin may be therapeutic for the treatment of vasospasm.

Topics: Animals; Blood Pressure; Cerebral Angiography; Cerebral Arteries; Cerebral Cortex; Cerebrovascular Circulation; Cyclic GMP; Heart Rate; Ischemic Attack, Transient; Macaca; Macaca fascicularis; Nervous System; Nitroglycerin; Nucleotides, Cyclic; Regional Blood Flow; Subarachnoid Hemorrhage; Vasodilator Agents

The present study was undertaken to determine whether oxyhemoglobin (OxyHb) is responsible for the functional alterations in the cerebral arteries observed during chronic vasospasm after subarachnoid hemorrhage. Vascular strips of canine basilar arteries were kept in organ culture for 3 days with or without repetitive exposure to OxyHb (OxyHb-treated and control strips). Contractions elicited by high levels of potassium (80 mM) and uridine 5'-triphosphate (3 x 10(-4) M) were reduced in the OxyHb-treated group in a concentration-dependent manner. The relaxations evoked by nitric oxide and 8-bromo-cyclic guanosine monophosphate (8-bromo-cGMP) were not affected. Relaxations elicited by the calcium channel blocker, diltiazem, were attenuated in the OxyHb-treated rings. When the extracellular calcium concentration ([Ca2+]e) was changed from a concentration in the external solution of 10(-8) M to 10(-3) M, myogenic tension developed. Myogenic tension, expressed as a percentage of the maximum contraction in each segment, was augmented in the OxyHb-treated group at [Ca2+]e of 10(-5) M and 10(-4) M. There were no significant differences in passive compliance of the arterial wall between the two groups. These results demonstrated that prolonged exposure to OxyHb in vitro results in a decrease in contractile capacity and an increase in sensitivity to [Ca2+]e, in agreement with previous findings in spastic arteries. By contrast, impairment of the 8-bromo-cGMP-mediated relaxation pathway and increased stiffness of the arterial wall, which have been reported to occur in spastic arteries, were not induced by prolonged exposure to OxyHb in vitro.

Topics: Analysis of Variance; Animals; Basilar Artery; Calcium; Calcium Channel Blockers; Compliance; Cyclic GMP; Diltiazem; Dogs; Female; Ischemic Attack, Transient; Male; Muscle Contraction; Nitric Oxide; Organ Culture Techniques; Oxyhemoglobins; Potassium; Subarachnoid Hemorrhage; Uridine Triphosphate; Vasoconstriction; Vasodilation

We hypothesized that nitric oxide exerts a negative feedback control on protein kinase C (PKC) activation, and the disturbance of the feedback control after subarachnoid hemorrhage results in vasospasm due to PKC activation. This study was undertaken to verify this hypothesis.. Different dogs were prepared for three separate experiments: measurement of the angiographic diameter of the basilar artery and determination of cGMP and PKC activity in vascular smooth muscle cells. In each experiment, two models were used: the single-hemorrhage model for mild vasospasm and the two-hemorrhage model for severe vasospasm. In both models, chronological changes of these three parameters were examined from day 1 until day 7.. In the single-hemorrhage model, mild vasospasm and a slight decrease of the cGMP level were noted on day 4, then both returned to the baseline levels on day 7. PKC activity was slightly enhanced throughout the study period. In the two-hemorrhage model, severe vasospasm and a significant decrease of the cGMP level were observed on day 5 and persisted until day 7. PKC activity was remarkably enhanced from day 5 until day 7. The differences between the two models with regard to the three parameters were statistically significant.. The decrease of cGMP level and the enhancement of PKC activity were obviously associated with the development of severe vasospasm. We conclude that subarachnoid hemorrhage disturbed the feedback control exerted by nitric oxide on PKC activation, leading to PKC-dependent vasospasm.

Topics: Animals; Basilar Artery; Cerebellum; Cerebral Angiography; Cyclic GMP; Disease Models, Animal; Dogs; Enzyme Activation; Feedback; Female; Ischemic Attack, Transient; Male; Muscle, Smooth, Vascular; Nitric Oxide; Protein Kinase C; Subarachnoid Hemorrhage; Time Factors

Calcitonin gene-related peptide (CGRP) is a potent vasodilator and a primary signaling molecule in neurovascular communication. In the present study, the authors examined cerebrovascular responses to CGRP and its related second messenger systems during cerebral vasospasm induced by subarachnoid hemorrhage (SAH). Tension measurements were performed in vitro on ring strips of basilar arteries obtained from rabbits subjected to artificial SAH and from control (non-SAH) animals. In vessels from SAH animals, which were preconstricted with serotonin, the vasorelaxant response to CGRP was attenuated. Because it has been suggested that vasodilation elicited by CGRP is mediated by cyclic 3',5'-adenosine monophosphate (cAMP) and/or cyclic 3',5'-guanosine monophosphate (cGMP), the vascular effects of directly activating these second messenger systems were also examined. The relaxant effect of forskolin, which activates adenylate cyclase directly, was slightly enhanced after SAH. In contrast, the relaxant effect of nitroglycerin (GTN), which activates soluble guanylate cyclase directly, was unchanged after SAH. The attenuation of CGRP-induced vasorelaxation could be the result of a modification in its ability to stimulate the production of second messengers. Experiments testing the capacity of CGRP to elevate cAMP levels showed no significant differences between vessels from non-SAH and SAH animals. Similarly, the resting levels of cAMP and the forskolin-induced elevations of cAMP did not differ between non-SAH and SAH animals. In contrast, cGMP levels were lower in resting and CGRP-treated vessels from SAH animals than in those from non-SAH animals. No significant differences in the levels of cGMP were observed between non-SAH and SAH vessels treated with GTN. This study indicates that CGRP-induced vasodilation is attenuated during vasospasm in a rabbit model of SAH. The findings also demonstrate that vasodilatory responses mediated by cAMP and cGMP are intact, although the levels of cGMP in SAH vessels are reduced. Together, these observations suggest that an attenuation in the capacity of vessels to dilate in response to CGRP occurs during cerebral vasospasm, and this change in CGRP vasoactivity is a result of modifications prior to, or independent of, the elevation of cyclic nucleotide second messengers.

Topics: Analysis of Variance; Animals; Basilar Artery; Calcitonin Gene-Related Peptide; Colforsin; Cyclic AMP; Cyclic GMP; Dose-Response Relationship, Drug; In Vitro Techniques; Ischemic Attack, Transient; Isometric Contraction; Male; Nitroglycerin; Potassium; Rabbits; Serotonin; Subarachnoid Hemorrhage; Vasodilation

The vasoactive effects of substance P (SP), as well as the content of cyclic guanine monophosphate (cGMP), were determined in the rabbit basilar artery after subarachnoid haemorrhage (SAH). Out of 47 rabbits, 24 were subjected to a SAH, induced by injecting 5 ml of autologous arterial blood into the cisterna magna; 23 were used as controls. In 20 animals (10 SAH and 10 controls), isometric tension recording of isolated rings of the basilar artery--dissected 2 days after SAH--was employed to assess the dose-dependent vasodilatation to SP (10(-10) to 10(-6) M) after precontraction with serotonin (10(-8) to 10(-5) M). In 15 animals (8 SAH and 7 controls), the basal cGMP content was measured in the basilar artery 2 days after SAH. In the other 12 animals (6 SAH and 6 controls), the increase in cGMP content was measured in the basilar artery after a 10-minute incubation with SP (10(-6) M). SP caused significantly less dilatation in animals subjected to SAH than in controls, especially for concentrations between 10(-9) and 10(-6) M (p < 0.001). The cGMP content in the arteries 2 days after SAH was significantly lower than in control arteries (31.5 +/- 7.3 against 57.3 +/- 4.3 pmoles/g tissue). In the preparations incubated with SP, the increase of cGMP was 440 +/- 115% in the control arteries, and only 97 +/- 30% in the arteries after SAH. It is concluded that the vasodilator activity of SP is significantly impaired after SAH.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Animals; Basilar Artery; Cyclic GMP; Dose-Response Relationship, Drug; Ischemic Attack, Transient; Male; Nitric Oxide; Rabbits; Subarachnoid Hemorrhage; Substance P; Vasoconstriction; Vasodilation

Endothelium-dependent relaxations are inhibited during chronic vasospasm after subarachnoid hemorrhage in the canine basilar artery, although the luminal release of endothelium-derived relaxing factor (EDRF) is maintained. The present study investigated the mechanisms underlying the impaired vascular reactivity and in particular whether the loss of responsiveness of the smooth muscle to EDRF is due to an impaired production of cGMP. Bradykinin and nitric oxide evoked concentration-dependent relaxations in isolated canine basilar arteries with and without endothelium, respectively, which were reduced in the subarachnoid hemorrhage group. Relaxations evoked by M&B22,948 (an inhibitor of cGMP phosphodiesterases) were smaller, but those evoked by the lipophilic cGMP analogue 8-bromo-cGMP were potentiated slightly in the subarachnoid hemorrhage group. The resting levels of cGMP in rings with endothelium (reflecting the effect of spontaneous release of EDRF) and those evoked by bradykinin in rings with endothelium and by nitric oxide in rings without endothelium were diminished in the subarachnoid hemorrhage group. These data indicate that the altered endothelium-mediated relaxations of the smooth muscle after subarachnoid hemorrhage is due, at least in part, to an impaired activation of soluble guanylate cyclase leading to a reduced production of cGMP in the smooth muscle.

Topics: 3',5'-Cyclic-GMP Phosphodiesterases; Animals; Basilar Artery; Bradykinin; Cyclic GMP; Dogs; Endothelium, Vascular; Female; Male; Nitric Oxide; Purinones; Radioimmunoassay; Subarachnoid Hemorrhage; Vasodilation

The authors have investigated the hypothesis that loss of endothelium-derived relaxing factor (EDRF) activity contributes to cerebral vasospasm after subarachnoid hemorrhage. Adventitial exposure to hemoglobin was studied angiographically by injecting purified hemoglobin solution or autologous whole blood into the cisterna magna of anesthetized pigs. Both interventions induced intra- but not extracerebral vasoconstriction, which persisted for 2 and 7 days, respectively. Cyclic guanosine monophosphate (cGMP) levels were measured in isolated buffer-perfused pig intrathecal arteries to quantify inhibition of basal EDRF activity by hemoglobin. Adventitial exposure was less effective than intimal exposure, 10 microM hemoglobin applied adventitially for 30 minutes having an effect equivalent to that of 1 microM applied intraluminally for 5 minutes. The depression of cGMP levels by hemoglobin was reversible and equivalent to the effect of endothelial denudation or incubation with NG-nitro-L-arginine methyl ester, so that the effects of hemoglobin can be attributed to a specific action on EDRF rather than interaction with a nitric oxide-like substance produced by vascular smooth muscle or adventitial nerves. Cyclic GMP levels in isolated arteries were unchanged after in vivo exposure to hemoglobin for either 2 or 7 days or to whole blood for 2 days, and were reduced by intraluminal perfusion with 1 microM hemoglobin. In contrast, after 7 days of in vivo exposure to whole blood, cGMP levels were already depressed, and not further reduced by intraluminal perfusion with 1 microM hemoglobin. The findings support the view that adventitially applied hemoglobin can inhibit basal EDRF activity and that in vivo adventitial exposure to whole blood leads to a reduction in basal cGMP levels in association with vasoconstriction of intrathecal arteries. Both mechanisms could contribute to the clinical syndrome of cerebral vasospasm after subarachnoid hemorrhage.

Topics: Animals; Cerebral Arteries; Chronic Disease; Cyclic GMP; Hemoglobins; Ischemic Attack, Transient; Nitric Oxide; Subarachnoid Hemorrhage; Subarachnoid Space; Swine; Time Factors; Vasoconstriction

c-AMP, c-GMP, HVA and 5 HIAA cerebrospinal fluid levels were investigated in 18 patients with subarachnoid haemorrhage (SAH). The main findings in the acute stage after SAH were represented by a marked increase of c-AMP and 5 HIAA values, whereas HVA levels were only slightly higher. In the chronic phase c-GMP levels turned out to be significantly increased, and were clearly related to intracranial hypertension. 5 HIAA and particularly HVA levels were decreased, probably due to the functional and anatomical lesion of the periventricular adrenergic structures, following the raised intracranial pressure.

Topics: Acute Disease; Adult; Chronic Disease; Cyclic AMP; Cyclic GMP; Female; Homovanillic Acid; Humans; Hydroxyindoleacetic Acid; Intracranial Pressure; Male; Middle Aged; Phenylacetates; Subarachnoid Hemorrhage