Pial Artery Diameter Research Articles

Localization of TRPA1 channels and characterization of TRPA1 mediated responses in dural and pial arteries in vivo after intracarotid infusion of Na2S.

The Transient Receptor Potential Ankyrin 1 (TRPA1) channel might play a role in migraine. However, different mechanisms for this have been suggested. The purpose of our study was to investigate the localization and significance of TRPA1 channels in rat pial and dural arteries. Immunofluorescence microscopy was used to localize TRPA1 channels in dural arteries, pial arteries, dura mater and trigeminal ganglion. The genuine closed cranial window model was used to examine the effect of Na2S, a donor of the TRPA1 channel opener H2S, on the diameter of pial and dural arteries. Further, we performed blocking experiments with TRPA1 antagonist HC-030031, calcitonin gene-related peptide (CGRP) receptor antagonist olcegepant and KCa3.1 channel blocker TRAM-34. TRPA1 channels were localized to the endothelium of both dural and pial arteries and in nerve fibers in dura mater. Further, we found TRPA1 expression in the membrane of trigeminal ganglia neuronal cells, some of them also staining for CGRP. Na2S caused dilation of both dural and pial arteries. In dural arteries, this was inhibited by HC-030031 and olcegepant. In pial arteries, the dilation was inhibited by TRAM-34, suggesting involvement of the KCa3.1 channel. Na2S causes a TRPA1- and CGRP-dependent dilation of dural arteries and a KCa3.1 channel-dependent dilation of pial arteries in rats.

Increased Oxyhemoglobin Binding Affinity Improves Cerebrovascular Responses To Hypoxia In Sprague-Dawley Rats

PURPOSE: This study evaluated the effect of increased oxyhemoglobin (O-H) affinity on cerebrovascular response to normobaric hypoxia. METHODS: Cerebrovascular response was assessed in 18 mechanically-ventilated Sprague-Dawley rats during acute exposure to normobaric hyperoxia (FiO2 1.0), normoxic air (FiO2 0.21), and hypoxic air (FiO2 0.10). Prior to exposure, the rats were administered GBT-1118 (GBT, 100mg/kg) or normal saline via oral gavage. Pial microcirculation was assessed and quantified through a cranial window created in the parietal bone of anesthetized rats. Cardiopulmonary parameters were measured throughout the exposures. Oxyhemoglobin binding affinity (p50) was assessed following the final gas exposure using a Hemox Analyzer. RESULTS: The mean p50 of GBT- and saline-treated animals was 31.46 and 37.63, respectively, demonstrating increased O-H affinity with GBT (p=0.0017). During steady state normoxia (FiO2 0.21), mean pial artery diameter decreased from baseline (FiO2 1.0) by 0.26% in saline animals compared to a 3.99% increase in GBT animals (p=0.1807). Additionally, GBT-treated animals demonstrated an 11.7% increase in blood O2 saturation (p<0.0001), an 11.4% increase in peak CO2 (p=0.0692), and an 8% increase in heart rate (HR) (p=0.015) compared to saline controls. During steady state hypoxia (FiO2 0.10), mean pial artery diameter decreased from baseline by 23.83% in saline animals compared to a 3.78% increase in GBT animals (p<0.0001). Additionally, GBT-treated animals demonstrated an 11.4% increase in blood O2 saturation (p=0.012), a 14.1% increase in peak CO2 (p=0.074), and a 9.3% decrease in HR (p=0.018) compared to saline controls. CONCLUSIONS: Collectively, the data show that impairments in cerebrovascular and cardiopulmonary function resulting from exposure to severe hypoxia can be mitigated through increased O-H binding and subsequent increases in blood oxygenation.

Contribution of Hydrogen Sulfide to Dilation of Rat Cerebral Arteries after Ischemia/Reperfusion Injury.

The study examined the effect of H2S on the tone of cerebral arteries in rats after global cerebral ischemia/reperfusion injury and cooperation between NO and H2S in the control over cerebral circulation during the postischemic period. In control sham-operated and in experimental rats with ischemia/reperfusion injury, the diameter of pial arteries was repeatedly measured in vivo under a light microscope after removal of parietal bone and dura mater in 1 h and in 7 days after the surgery. The study established that H2S is an important signaling molecule in pial arteries, where it is responsible for vasodilation. Interaction of H2S and NO augmented dilation of pial arteries; in these arteries, H2S up-regulated the effect of NO/cGMP/sGC/PKG signaling pathways. Partially, the dilating effect of H2S is realized via activation of ATP-sensitive K+ channels in plasmalemma of smooth muscle cells. In the brain, ischemia/reperfusion injury degrades the ability of pial arteries to dilate via inhibition of NO-mediated signaling pathway.

Effect of TRPA1 activator allyl isothiocyanate (AITC) on rat dural and pial arteries.

Transient receptor potential ankyrin 1 (TRPA1) channels may have a role in migraine as some substances known to cause headache activate the channel. In the craniovascular system such activation causes a calcitonin gene-related peptide (CGRP)-dependent increase in meningeal blood flow. TRPA1 channels in the endothelium of cerebral arteries cause vasodilation when activated. The headache preventive substance feverfew inhibits activation of TRPA1 channels. In this study we aim to compare and characterize the effect of the TRPA1 agonist allyl isothiocyanate (AITC) on the diameter of rat dural and pial arteries in vivo. The genuine closed-cranial window technique in rats was used to examine changes in dural and pial artery diameter and mean arterial blood pressure (MABP) after intracarotid infusion of AITC. Blockade experiments were performed by intravenous infusion of olcegepant, HC-030031, sumatriptan or capsazepine immediately after infusion of AITC, in four different groups of rats. AITC caused a significant dilation of dural arteries, which was inhibited by HC-030031, olcegepant and sumatriptan, but not by capsazepine. In pial arteries AITC caused a significant dilation, which was not inhibited by any of the pre-treatments, suggesting a poor penetration of the blood-brain barrier or autoregulation due to dimethyl sulfoxide (DMSO) mediated decrease in MABP during HC-030031 infusion. AITC did not cause a significant change in MABP. AITC causes dilation of dural arteries via a mechanism dependent on CGRP and TRPA1 that is sensitive to sumatriptan. AITC causes a small but significant dilation of pial arteries.

Vascular Gap Junctions Contribute to Forepaw Stimulation-Induced Vasodilation Differentially in the Pial and Penetrating Arteries in Isoflurane-Anesthetized Rats.

Somatosensory stimulation causes dilation of the pial and penetrating arteries and an increase in cerebral blood flow (CBF) in the representative region of the somatosensory cortex. As an underlying mechanism for such stimulation-induced increases in CBF, cerebral artery dilation has been thought to propagate in the vascular endothelium from the parenchyma to the brain surface. Vascular gap junctions may propagate vasodilation. However, the contribution of vascular gap junctions to cerebrovascular regulation induced by somatosensory stimulation is largely unknown. The aim of the present study was to investigate the contribution of vascular gap junctions to the regulation of the pial and penetrating arteries during neuronal activity attributed to somatosensory stimulation. Experiments were performed on male Wistar rats (age: 7–10 weeks) with artificial ventilation under isoflurane anesthesia. For somatosensory stimulation, the left forepaw was electrically stimulated (1.5 mA, 0.5 ms and 10 Hz, for 5 s). The artery in the forelimb area of the right somatosensory cortex was imaged through a cranial window using a two-photon microscope and the diameter was measured. Carbenoxolone (CBX) was intravenously (i.v.) administered, at a dose of 100 mg/kg, to block vascular gap junctions. The forepaw electrical stimulation increased the diameter of the pial and penetrating arteries by 7.0% and 5.0% of the pre-stimulus diameter, respectively, without changing the arterial pressure. After CBX administration, the change in pial artery diameter during forepaw stimulation was attenuated to 3.2%. However, changes in the penetrating artery were not significantly affected. CBF was measured using a laser speckle flowmeter, together with somatosensory-evoked potential (SEP) recorded in the somatosensory cortex. The extent of CBF increase (by 24.1% of the pre-stimulus level) and amplitude of SEP were not affected by CBX administration. The present results suggest that vascular gap junctions, possibly on the endothelium, contribute to pial artery dilation during neuronal activity induced by somatosensory stimulation.

SOD3-Mimetic As a Complement for Genetic Therapy in Sickle Cell Disease

Background and Objectives: Genetic therapy relieves transfusion-dependence in patients 12 years and older with ß-Thalassemia (1) There is an urgent need to develop a complementary approach to genetic therapy for children younger than 12 years of age. The objective of this report is to describe the potential development of an extracellular superoxide dismutase-mimetic (SOD3-mimetic) to relieve sickle cell disease (SCD) children susceptible to stroke from being transfusion-dependent in order to avoid stroke. SanFlow [aka polynitroxylated pegylated hemoglobin (PNPH)], a SOD3-mimetic has been evaluated and approved by the FDA for further development as drug for the treatment of traumatic brain injury (TBI) complicated by hemorrhagic shock (HS), stroke, and SCD. PK&PD studies demonstrated that SanFlow works as a macromolecular SOD3-mimetic by protecting the endogenous vascular nitric oxide (NO) leading to enhanced blood flow. SanFlow has been shown to protect against superoxide induced hypoxia and inflammation in transgenic SCD and rat model of ischemic stroke as well as a mouse model of TBI+HS. In a rat middle cerebral artery occlusion (MCAO) model of stroke the spreading of the hypoxic core of the ischemic brain as measured by Pial arterial diameter is maintained over 2 hours post onset of stroke with associated reduction in inflammatory markers and brain infraction (3). In a mouse TBI+SH model, SanFlow was shown to be superior to fresh shed whole blood. The safety and efficacy of SanFlow was tested over a 50 fold dose range (i.e. from 0.1 to 5 times the shed blood volume). SanFlow was shown to work at extremely low volume in conjunction with volume replacement crystalloid to substitute whole blood resuscitation (3).Experimental Results: Using Berkeley model of sickle mice (Hba0/0 Hbb0/0Tg (HuminiLCRα1GγAγδβS) we have measures the PK&PD of SanFlow as a complementary or a substitute for genetic therapy. We have non-invasively measured the blood flow and vasoconstriction using transgenic SS mice (N=7) against WT littermates (N=4) as control. A single bolus dose of SanFlow (20ml/kg at hemoglobin of 4g/dl) significantly corrected the aortic stiffness and pulmonary flow of the SS mice to that of WT littermates (P<0.05). This is also correlated with the decrease of superoxide level in the lung as measure by Luminol activity assays (which fluoresces in the presence of superoxide) (P<0.05). Plasma cGMP (downstream effector of NO and natriuretic peptide activity) and NOx (breakdown product of NO and a measure of NOS activity) concentrations were measured in plasma of mice after infusion of SanFlow. SanFlow infusion was shown to significantly increased plasma cGMP concentrations in plasma. SanFlow infusion also resulted in a significant increase in NOx in plasma after infusion (P<0.05). SanFlow resulted in an increase in plasma cGMP and NOx in SS mice. Thus, the pathophysiological defects or difference of SS mice and WT littermates are corrected by SanFlow infusion.Conclusions:The present results support development of SanFlow, delivered through continuous infusion, for anemic SCD children to prevent the development of blood transfusion dependency in order to avoid stroke and painful vaso-occlusive crisis (VOC). These results also demonstrated that SanFlow can be used safely and effectively in the elimination of serious painful vaso-occlusive crisis and protect silent and major strokes. Clinical trials of SanFlow in SCD children, prior to their transfusion dependence, as well as in transfusion-dependent teenagers and adults with SCD patients are warranted. By extension, ß-Thalassemia patients can also be treated like SCD patients using SanFlow to relieve them from dependence on life-long blood exchange transfusion. However, these patients would be treated with SOD3-mimetic while waiting for genetic therapy.

Abstract TP251: Release of IL-6 After Stroke Contributes to Impaired Cerebral Autoregulation and Hippocampal Neuronal Necrosis Through NMDA Receptor Activation and Upregulation of ET-1 and JNK

Introduction: The sole FDA approved drug treatment for acute stroke is tissue type plasminogen activator (tPA). However, its brief therapeutic window and post-treatment complications markedly constrain its use. Upregulation of JNK MAPK and ET-1 by exogenous tPA after thrombotic stroke contributes to impaired cerebrovascular autoregulation, which exacerbates ischemic injury. Wild type (wt) tPA can bind to either the lipoprotein-related receptor (LRP), which mediates vasodilation, or NMDA-receptors (NMDA-Rs), exacerbating vasoconstriction and ischemia. tPA-A 296-299 , a variant that is fibrinolytic and binds to LRP but not to NMDA-Rs, does not induce upregulation of ET-1 and JNK and thereby does not exacerbate loss of autoregulation and the resultant histopathology after stroke. Elevations in IL-6, a marker of the inflammation that accompanies ischemic stroke, is reported to be an adverse prognostic factor. Hypothesis: We hypothesized that IL-6 released after stroke by wt tPA through activation of NMDA-Rs and upregulation of ET-1 and JNK contributes to impairment of cerebral autoregulation and increased histopathology. Methods: Pial artery diameter, CSF ET-1, JNK, and IL-6 measured and histopathology assessed in anesthetized pigs. Stroke was induced by photothrombosis. Results: IL-6 was increased post-stroke, which was increased further by wt tPA. Co-administration of the IL-6 antagonist LMT-28 with wt tPA prevented impairment of cerebral autoregulation and necrosis of hippocampal CA1, CA3 cells. Isoproterenol dilation was unchanged by stroke. tPA-A 296-299 , wt tPA co-administered with the JNK inhibitor SP 600125 and the ET-1 antagonist BQ 123 blocked stroke induced elevation of IL-6. Co-administration of LMT-28 with wt tPA blocked the augmentation of JNK and ET-1 post-stroke. Conclusions: IL-6 released after stroke, which is enhanced by wt tPA through activation of NMDA-Rs and upregulation of ET-1 and JNK, impairs cerebral autoregulation and increased histopathology. Strategies that promote fibrinolysis while limiting activation of NMDA-Rs and release of IL-6, such as tPA-A 296-299 , may improve the benefit/risk ratio compared to wt tPA in treatment of stroke.

The effects of Y-27632 on pial microvessels during global brain ischemia and reperfusion in rabbits

BackgroundGlobal brain ischemia-reperfusion during propofol anesthesia provokes persistent cerebral pial constriction. Constriction is likely mediated by Rho-kinase. Cerebral vasoconstriction possibly exacerbates ischemic brain injury. Because Y-27632 is a potent Rho-kinase inhibitor, it should be necessary to evaluate its effects on cerebral pial vessels during ischemia—reperfusion period. We therefore tested the hypotheses that Y-27632 dilates cerebral pial arterioles after the ischemia-reperfusion injury, and evaluated the time-course of cerebral pial arteriolar status after the ischemia-reperfusion.MethodsJapanese white rabbits were anesthetized with propofol, and a closed cranial window inserted over the left hemisphere. Global brain ischemia was produced by clamping the brachiocephalic, left common carotid, and left subclavian arteries for 15 min. Rabbits were assigned to cranial window perfusion with: (1) artificial cerebrospinal fluid (Control group, n = 7); (2) topical infusion of Y-27632 10−6 mol · L−1 for 30 min before the initiation of global brain ischemia (Pre group, n = 7); (3) topical infusion of Y-27632 10−6 mol · L−1 starting 30 min before ischemia and continuing throughout the study period (Continuous group, n = 7); and, (4) topical infusion of Y-27632 10−6 mol · L−1 starting 10 min after the ischemia and continuing until the end of the study (Post group, n = 7). Cerebral pial arterial and venule diameters were recorded 30 min before ischemia, just before arterial clamping, 10 min after clamping, and 5, 10, 20, 40, 60, 80, 100, and 120 min after unclamping.ResultsMean arterial blood pressure and blood glucose concentration increased significantly after global brain ischemia except in the Continuous group. In the Pre and Continuous groups, topical application of Y-27632 produced dilation of large (mean 18–19%) and small (mean; 25–29%) pial arteries, without apparent effect on venules. Compared with the Control and Pre groups, arterioles were significantly dilated during the reperfusion period in the Continuous and Post groups (mean at 120 min: 5–8% in large arterioles and 11–12% in small arterioles).ConclusionsY-27632 dilated cerebral pial arterioles during reperfusion. Y-27632 may enhance recovery from ischemia by preventing arteriolar vasoconstriction during reperfusion.

Ketamine modulation of the haemodynamic response to spreading depolarization in the gyrencephalic swine brain

Spreading depolarization (SD) generates significant alterations in cerebral haemodynamics, which can have detrimental consequences on brain function and integrity. Ketamine has shown an important capacity to modulate SD; however, its impact on SD haemodynamic response is incompletely understood. We investigated the effect of two therapeutic ketamine dosages, a low-dose of 2 mg/kg/h and a high-dose of 4 mg/kg/h, on the haemodynamic response to SD in the gyrencephalic swine brain. Cerebral blood volume, pial arterial diameter and cerebral blood flow were assessed through intrinsic optical signal imaging and laser-Doppler flowmetry. Our findings indicate that frequent SDs caused a persistent increase in the baseline pial arterial diameter, which can lead to a diminished capacity to further dilate. Ketamine infused at a low-dose reduced the hyperemic/vasodilative response to SD; however, it did not alter the subsequent oligemic/vasoconstrictive response. This low-dose did not prevent the baseline diameter increase and the diminished dilative capacity. Only infusion of ketamine at a high-dose suppressed SD and the coupled haemodynamic response. Therefore, the haemodynamic response to SD can be modulated by continuous infusion of ketamine. However, its use in pathological models needs to be explored to corroborate its possible clinical benefit.

Preferential Protection of Cerebral Autoregulation and Reduction of Hippocampal Necrosis With Norepinephrine After Traumatic Brain Injury in Female Piglets.

Traumatic brain injury contributes to morbidity in children and boys is disproportionately represented. Cerebral autoregulation is impaired after traumatic brain injury, contributing to poor outcome. Cerebral perfusion pressure is often normalized by the use of vasopressors to increase mean arterial pressure. In prior studies, we observed that phenylephrine prevented impairment of autoregulation in female but exacerbated in male piglets after fluid percussion injury. In contrast, dopamine prevented impairment of autoregulation in both sexes after fluid percussion injury, suggesting that pressor choice impacts outcome. The extracellular signal-regulated kinase isoform of mitogen-activated protein kinase produces hemodynamic impairment after fluid percussion injury, but the role of the cytokine interleukin-6 is unknown. We investigated whether norepinephrine sex-dependently protects autoregulation and limits histopathology after fluid percussion injury and the role of extracellular signal-regulated kinase and interleukin-6 in that outcome. Prospective, randomized animal study. University laboratory. Newborn (1-5 d old) pigs. Cerebral perfusion pressure, cerebral blood flow, and pial artery diameter were determined before and after fluid percussion injury in piglets equipped with a closed cranial window and post-treated with norepinephrine. Cerebrospinal fluid extracellular-signal-regulated kinase mitogen-activated protein kinase was determined by enzyme-linked immunosorbent assay. Norepinephrine does not protect autoregulation or prevent reduction in cerebral blood flow in male but fully protects autoregulation in female piglets after fluid percussion injury. Papaverine-induced dilation was unchanged by fluid percussion injury and norepinephrine. Norepinephrine increased extracellular signal-regulated kinase mitogen-activated protein kinase up-regulation in male but blocked such up-regulation in female piglets after fluid percussion injury. Norepinephrine aggravated interleukin-6 upregulation in males in an extracellular signal-regulated kinase mitogen-activated protein kinase-dependent mechanism but blocked interleukin-6 up-regulation in females after fluid percussion injury. Norepinephrine augments loss of neurons in CA1 and CA3 hippocampus of male piglets after fluid percussion injury in an extracellular signal-regulated kinase mitogen-activated protein kinase-dependent and interleukin-6-dependent manner but prevents loss of neurons in females after fluid percussion injury. Norepinephrine protects autoregulation and limits hippocampal neuronal cell necrosis via modulation of extracellular signal-regulated kinase mitogen-activated protein kinase and interleukin-6 after fluid percussion injury in a sex-dependent manner.

Dynamics of pial vessels reactivity after brief cerebral ischemia

Cerebral blood vessel reactivity is one of the main determinants of final outcome of brain ischemia. Most of studies on the vascular mechanisms of ischemic brain injury, however, focus on the acute changes within ischemic period or several hours after it. Dilatatory capacity of cerebral arterioles (perfusion reservoir) is considered as an important factor of brain perfusion elevation in critical situations.The aim of the present study was to examine the pial vessel reactivity in response to hypercapnia in rats, subjected to transient global cerebral ischemia, at 7, 14 and 21 days after ischemia. Materials and methods. Transient global cerebral ischemia was induced in anesthetized Wistar rats by bilateral common carotid artery occlusion for 12 min with simultaneous controlled hypotension to 45±3 mm Hg, followed by blood reinfusion and recovery from anesthesia. Three different groups of rats were re-anesthetized at 7, 14 or 21 days after ischemia and subjected to microvascular reactivity studies using in vivo video microscopy. Hypercapnia was caused by i.v. injection of acetazolamide. The changes in diameter of pial arteries and veins in response to hypercapnia were measured. Results and discussion. Global cerebral ischemia led to marked decrease in pial vessels (both arteries and veins) reactivity in response to hypercapnia, caused by i.v. injection of acetazolamide. In intact rats, i.v. injection of acetazolamide led to pial arteries dilation and pial veins constriction; in animals subjected to ischemia-reperfuion. the numbers of dilated large arteries and constricted small veins were much less, as well as the extent of arterial dilation. Reactivity changes were observed in all time points studied. Conclusions. Thus, transient global cerebral ischemia cause marked and long lasting (3 weeks) decrease in pial vessel reactivity in response to hypercapnia.

Dopamine prevents impairment of autoregulation after traumatic brain injury in the newborn pig through inhibition of Up-regulation of endothelin-1 and extracellular signal-regulated kinase mitogen-activated protein kinase.

Traumatic brain injury contributes to morbidity in children and boys are disproportionately represented. Autoregulation is impaired more in male compared with female piglets after traumatic brain injury through sex-dependent up-regulation of the spasmogen endothelin-1 and extracellular signal-regulated kinase (ERK) mitogen-activated protein kinase (MAPK), a family of three kinases: ERK, p38, and JNK). Elevation of mean arterial pressure leading to increased cerebral perfusion pressure via phenylephrine improves impairment of autoregulation after traumatic brain injury in female but not male piglets through modulation of endothelin-1 and ERK MAPK up-regulation, blocked in females, but aggravated in males. We hypothesized that pressor choice to elevate cerebral perfusion pressure is important in improving cerebral hemodynamics after traumatic brain injury and that dopamine will prevent impairment of autoregulation in both male and female piglets through blockade of endothelin-1 and ERK MAPK. Prospective, randomized animal study. University laboratory. Newborn (1-5 days old) pigs. Cerebral perfusion pressure and pial artery diameter were determined before and after lateral fluid percussion brain injury was produced in piglets equipped with a closed cranial window. Dopamine (15 µg/kg/min IV) was administered 30 mins post fluid percussion injury. Cerebrospinal fluid ERK MAPK was determined by enzyme-linked immunosorbent assay. Dopamine increased cerebral perfusion pressure equivalently in both sexes and prevented sex-dependent reductions in pial artery diameter after fluid percussion injury. Loss of pial artery dilation during hypotension was greater in male than in female piglets after fluid percussion injury, but dopamine prevented such impairment equivalently in both sexes post injury. endothelin-1 and ERK MAPK release was greater in male compared to female piglets after fluid percussion injury, but dopamine also blocked their up-regulation equivalently in male and female piglets after fluid percussion injury. These data indicate that dopamine is protective of autoregulation after fluid percussion injury in both sexes. These observations advocate for the consideration of development of sex based therapies for treatment of hemodynamic sequalae of pediatric traumatic brain injury.

Red blood cell-coupled tissue plasminogen activator prevents impairment of cerebral vasodilatory responses through inhibition of c-Jun-N-terminal kinase and potentiation of p38 mitogen-activated protein kinase after cerebral photothrombosis in the newborn pig

Pediatric ischemic stroke is a poorly understood, yet clinically important, problem. The sole approved treatment for acute stroke is tissue-type plasminogen activator. However, tissue plasminogen activator vasoactivity aggravates hypoxia/ischemia-induced impairment of cerebrovasodilation in response to hypercapnia and hypotension in newborn pigs. Mitogen-activated protein kinase (a family of 3 kinases, extracellular signal-related kinase, p38, and c-Jun-N-terminal kinase) is upregulated after hypoxia/ischemia. Coupling of tissue plasminogen activator to red blood cells prevented hypoxia/ischemia-induced impairment of dilation and suppressed extracellular signal-related kinase mitogen-activated protein kinase activation. This study investigated the differential roles of mitogen-activated protein kinase isoforms in the effects of red blood cells-tissue plasminogen activator on cerebrovasodilation in a translationally relevant injury model, photothrombosis. Prospective, randomized animal study. : University laboratory. Newborn (1- to 5-day-old) pigs. Cerebral blood flow and pial artery diameter were determined before and after photothrombotic injury (laser 532 nm and erythrosine B) was produced in piglets equipped with a closed cranial window. Cerebral blood flow extracellular signal-related kinase, p38, and c-Jun-N-terminal kinase mitogen-activated protein kinase were determined by enzyme-linked immunosorbent assay. Tissue plasminogen activator and red blood cells-tissue plasminogen activator alleviated reduction of cerebral blood flow after photothrombotic injury. Cerebrovasodilation was blunted by photothrombotic injury, reversed to vasoconstriction by tissue plasminogen activator, but dilation was maintained by red blood cells-tissue plasminogen activator. Cerebral blood flow c-Jun-N-terminal kinase and p38 mitogen-activated protein kinase but not extracellular signal-related kinase mitogen-activated protein kinase was elevated by photothrombotic injury, an effect potentiated by tissue plasminogen activator. Red blood cells-tissue plasminogen activator blocked c-Jun-N-terminal kinase but potentiated p38 mitogen-activated protein kinase upregulation after photothrombotic injury. A c-Jun-N-terminal kinase mitogen-activated protein kinase antagonist prevented, a p38 mitogen-activated protein kinase antagonist potentiated, whereas an extracellular signal-related kinase mitogen-activated protein kinase antagonist had no effect on dilator impairment after photothrombotic injury. These data indicate that in addition to restoring perfusion, red blood cells-tissue plasminogen activator prevents impairment of cerebrovasodilation after photothrombotic injury through blockade of c-Jun-N-terminal kinase and potentiation of p38 mitogen-activated protein kinase. These data suggest tissue plasminogen activator coupling to red blood cells offers a novel approach to increase the benefit/risk ratio of thrombolytic therapy to treat central nervous system ischemic disorders.

TPA contributes to impaired NMDA cerebrovasodilation after traumatic brain injury through activation of JNK MAPK

<title/>Objective: N-methyl-D-aspartate (NMDA)-induced pial artery dilation (PAD) is reversed to vasoconstriction after fluid percussion brain injury (FPI). Tissue type plasminogen activator (tPA) is up-regulated and the tPA antagonist, EEIIMD, prevents impaired NMDA PAD after FPI. Mitogen-activated protein kinase (MAPK), a family of at least three kinases, ERK, p38, and JNK, is also up-regulated after traumatic brain injury (TBI). We hypothesize that tPA impairs NMDA-induced cerebrovasodilation after FPI in a MAPK isoform-dependent mechanism.Methods: Lateral FPI was induced in newborn pigs. The closed cranial window technique was used to measure pial artery diameter and to collect cerebrospinal fluid (CSF). ERK, p38, and JNK MAPK concentrations in CSF were quantified by ELISA.Results: CSF JNK MAPK was increased by FPI, increased further by tPA, but blocked by JNK antagonists SP600125 and D-JNKI1. FPI modestly increased p38 and ERK isoforms of MAPK. NMDA-induced PAD was reversed to vasoconstriction after FPI, whereas dilator responses to papaverine were unchanged. tPA, in post-FPI CSF concentration, potentiated NMDA-induced vasoconstriction while papaverine dilation was unchanged. SP 600125 and D-JNKI1, blocked NMDA-induced vasoconstriction and fully restored PAD. The ERK antagonist U 0126 partially restored NMDA-induced PAD, while the p38 inhibitor SB203580 aggravated NMDA-induced vasoconstriction observed in the presence of tPA after FPI.Discussion: These data indicate that tPA contributes to impairment of NMDA-mediated cerebrovasodilation after FPI through JNK, while p38 may be protective. These data suggest that inhibition of the endogenous plasminogen activator system and JNK may improve cerebral hemodynamic outcome post-TBI.

Capillary Remodeling and Collateral Growth Without Angiogenesis After Unilateral Common Carotid Artery Occlusion in Mice

To clarify the mechanisms of blood flow restoration after major artery occlusion, we presented first dynamic changes in cortical vessel morphology observed through a cranial window in mice after unilateral common carotid artery (CCA) occlusion. The density and diameter of capillaries, as well as diameters of pial arteries, were measured by confocal laser-scanning microscopy and fluorescent microscopy, respectively. Possible angiogenesis was evaluated by detecting any outgrowth of endothelial cells from pre-existing vessels or intussusception in Tie2-GFP mice. Immediately after unilateral CCA occlusion, cerebral blood flow (CBF) index, the reciprocal of mean transit time, reduced significantly and returned to the previous level after 14 days. Repeated observation of the cortical vessels did not reveal any angiogenesis, whereas the cortical capillary diameter increased by 74% after 14 days. The anterior cerebral artery (ACA) and collateral vessels connecting ACA and middle cerebral artery also dilated significantly. The capillary dilatation to the size of arteriole in the settings of collateral growth and CBF restoration suggested capillary remodeling. Our results indicate that capillary remodeling, pial artery dilatation and collateral growth without angiogenesis are sufficient mechanisms to restore normal cerebral blood flow after unilateral CCA occlusion.

Endothelial SKCa and IKCa Channels Regulate Brain Parenchymal Arteriolar Diameter and Cortical Cerebral Blood Flow

Calcium-sensitive potassium (K(Ca)) channels have been shown to modulate the diameter of cerebral pial arteries; however, little is known regarding their roles in controlling cerebral parenchymal arterioles (PAs). We explored the function and cellular distribution of small-conductance (SK(Ca)) and intermediate-conductance (IK(Ca)) K(Ca) channels and large-conductance K(Ca) (BK(Ca)) channels in endothelial cells (ECs) and smooth muscle cells (SMCs) of PAs. Both SK(Ca) and IK(Ca) channels conducted the outward current in isolated PA ECs (current densities, ~20 pA/pF and ~28 pA/pF at +40 mV, respectively), but these currents were not detected in PA SMCs. In contrast, BK(Ca) currents were prominent in PA SMCs (~154 pA/pF), but were undetectable in PA ECs. Pressurized PAs constricted to inhibition of SK(Ca) (~16%) and IK(Ca) (~16%) channels, but were only modestly affected by inhibition of BK(Ca) channels (~5%). Blockade of SK(Ca) and IK(Ca) channels decreased resting cortical cerebral blood flow (CBF) by ~15%. NS309 (6,7-dichloro-1H-indole-2,3-dione3-oxime), a SK(Ca)/IK(Ca) channel opener, hyperpolarized PA SMCs by ~27 mV, maximally dilated pressurized PAs, and increased CBF by ~40%. In conclusion, these data show that SK(Ca) and IK(Ca) channels in ECs profoundly modulate PA tone and CBF, whereas BK(Ca) channels in SMCs only modestly influence PA diameter.

Impaired cerebral blood flow autoregulation during posttraumatic arterial hypotension after fluid percussion brain injury is prevented by phenylephrine in female but exacerbated in male piglets by extracellular signal-related kinase mitogen-activated protein kinase upregulation.

Traumatic brain injury contributes to morbidity and mortality in children and boys are disproportionately represented. Hypotension is common and worsens outcome after traumatic brain injury. Extracellular signal-related kinase mitogen-activated protein kinase is upregulated and reduces cerebral blood flow after fluid percussion brain injury in piglets. We hypothesized that increased cerebral perfusion pressure through phenylephrine sex dependently reduces impairment of cerebral autoregulation during hypotension after fluid percussion brain injury through modulation of extracellular signal-related kinase mitogen-activated protein kinase. Prospective, randomized animal study. University laboratory. Newborn (1- to 5-day-old) pigs. Cerebral blood flow, pial artery diameter, intracranial pressure, and autoregulatory index were determined before and after fluid percussion brain injury in untreated, preinjury, and postinjury phenylephrine (1 microg/kg/min intravenously) treated male and female pigs during normotension and hemorrhagic hypotension. Cerebrospinal fluid extracellular signal-related kinase mitogen-activated protein kinase was determined by enzyme-linked immunosorbent assay. Reductions in pial artery diameter, cerebral blood flow, cerebral perfusion pressure, and elevated intracranial pressure after fluid percussion brain injury were greater in males, which were blunted by phenylephrine pre- or postfluid percussion brain injury. During hypotension and fluid percussion brain injury, pial artery dilation was impaired more in males. Phenylephrine decreased impairment of hypotensive pial artery dilation after fluid percussion brain injury in females, but paradoxically caused vasoconstriction after fluid percussion brain injury in males. Papaverine-induced pial artery vasodilation was unchanged by fluid percussion brain injury and phenylephrine. Cerebral blood flow, cerebral perfusion pressure, and autoregulatory index decreased markedly during hypotension and fluid percussion brain injury in males but less in females. Phenylephrine prevented reductions in cerebral blood flow, cerebral perfusion pressure, and autoregulatory index during hypotension in females but increased reductions in males. Cerebrospinal fluid extracellular signal-related kinase mitogen-activated protein kinase was increased more in males than females after fluid percussion brain injury. Phenylephrine blunted extracellular signal-related kinase mitogen-activated protein kinase upregulation in females but increased extracellular signal-related kinase mitogen-activated protein kinase upregulation in males after fluid percussion brain injury. These data indicate that elevation of cerebral perfusion pressure with phenylephrine sex dependently prevents impairment of cerebral autoregulation during hypotension after fluid percussion brain injury through modulation of extracellular signal-related kinase mitogen-activated protein kinase. These data suggest the potential role for sex-dependent mechanisms in cerebral autoregulation after pediatric traumatic brain injury.

SNP improves cerebral hemodynamics during normotension but fails to prevent sex dependent impaired cerebral autoregulation during hypotension after brain injury

Traumatic brain injury (TBI) is a leading cause of morbidity in children and boys are disproportionately represented. Hypotension is common and worsens outcome after TBI. Previous studies show that adrenomedullin, a cerebrovasodilator, prevented sex dependent impairment of autoregulation during hypotension after piglet fluid percussion brain injury (FPI). We hypothesized that this concept was generalizable and that administration of another vasodilator, sodium nitroprusside (SNP), may equally improve CBF and cerebral autoregulation in a sex dependent manner after FPI. SNP produced equivalent percent cerebrovasodilation in male and female piglets. Reductions in pial artery diameter, cortical CBF, and cerebral perfusion pressure (CPP) concomitant with elevated intracranial pressure (ICP) after FPI were greater in male compared to female piglets during normotension which was blunted by SNP. During hypotension, pial artery dilation (PAD) was impaired more in the male than the female after FPI. However, SNP did not improve hypotensive PAD after FPI in females and paradoxically caused vasoconstriction in males. SNP did not prevent reductions in CBF, CPP or autoregulatory index during combined hypotension and FPI in either sex. SNP aggravated ERK MAPK upregulation after FPI. These data indicate that despite prevention of reductions in CBF after FPI, SNP does not prevent impairment of autoregulation during hypotension after FPI. These data suggest that therapies directed at a purely hemodynamic increase in CPP will fail to improve outcome during combined TBI and hypotension.

Effect of two novel CGRP-binding compounds in a closed cranial window rat model

We investigated the in vivo effects of two novel calcitonin gene-related peptide (CGRP) binding molecules in the genuine closed cranial window model in the rat. The RNA-Spiegelmer (NOX-C89) and the monoclonal CGRP antibody are CGRP scavengers and might be used as an alternative to CGRP-receptor antagonists in the treatment of migraine. Rats were anaesthetized and a closed cranial window established. Changes in dural and pial artery diameter and mean arterial blood pressure were measured simultaneously. Infusion of the RNA-Spiegelmer or the CGRP antibody alone had no effect on the arteries or the mean arterial blood pressure. We then used a bolus of 0.3 μg/kg CGRP (n = 6) or electrical stimulation (25 V, 5 Hz, 1 ms pulse width and of 10 s of duration) (n = 6) to induce dilatation of dural and pial arteries (mediated via CGRP-receptors). Pre-treatment with the RNA-Spiegelmer inhibited CGRP-induced vasodilatation of the dural artery (from 38 ± 17% to 7 ± 3%) and the pial artery (from 14 ± 1% to 3 ± 2%) (P < 0.05). The RNA-Spiegelmer, however, did not significantly inhibit dilatation induced by electrical stimulation (P > 0.05). The CGRP antibody caused a significant reduction of the dural artery diameter caused by intravenous CGRP-infusion (from 23 ± 5% to 12 ± 3%) (P < 0.05), but did not inhibit dilatation caused by electrical stimulation (P > 0.05). In conclusion, the CGRP scavengers effectively inhibited the effect of circulating CGRP but do not modify the effect of electrical stimulation and the consequent liberation of CGRP from perivascular sensory nerve fibres.

Mode Changes of Cerebrovascular Pressure Transmission Induced by Cerebral Vasodilation

Changes in the modes of cerebrovascular pressure transmission during cerebral vasodilation induced by hypercapnic challenge were examined as a means for developing the basis for a bedside method to evaluate regulation of cerebral blood flow. Recordings of arterial blood pressure (ABP) and intracranial pressure (ICP) obtained from a piglet preparation equipped with a cranial window were used to determine serial changes of the highest modal frequency (HMF) and dampening factor (DF) of a numerical system identification model of cerebrovascular pressure transmission. Resistance and compliance elements of a Windkessel model of ICP dynamics selected to provide the mathematical structure for the system identification modeling approach were also manipulated to obtain a match with HMF, DF, and the experimental and simulated recordings of ICP. During hypercapnic challenge, significant increases of ICP, pial arterial diameter (PAD) and partial pressure of arterial blood carbon dioxide increases, and a decrease of arterial pH were observed. Vasodilation changed the modes of the system identification model of cerebrovascular pressure transmission from a dominantly over-damped process to an under-damped one with a significant increase in HMF and decrease in DF. Simulations of the Windkessel circuit model required a decrease in the relative resistance and an increase in relative compliance of the arterial-arteriolar vascular bed consistent with the observed increases in PAD induced by vasodilation. Evaluation of changes in the modes of cerebrovascular pressure transmission may provide means of assessing the state of cerebrovascular vasodilation and autoregulation of cerebral blood flow in the clinical setting.