bluuebunny/crossref_metadata_2025 huggingface dataset:
Muscarinic—but Not Nicotinic—Acetylcholine Receptors Mediate a Nitric Oxide-Dependent Dilation in Brain Cortical Arterioles: A Possible Role for the M5 Receptor Subtype
Authors: Elhusseiny Ahmed , Hamel Edith
Published: November 2003

Increases in cortical cerebral blood flow are induced by stimulation of basal forebrain cholinergic neurons. This response is mediated in part by nitric oxide (NO) and reportedly involves both nicotinic and muscarinic receptors, some of which are possibly located in the vessel wall. In the present study, the vasomotor response(s) elicited by acetylcholine (ACh) on isolated and pressurized bovine and/or human intracortical penetrating arterioles were investigated, and pharmacological characterization of the receptor involved in this response was carried out. Acetylcholine (10−11 to 10−4 mol/L) dose dependently dilated bovine and human intracortical arterioles at spontaneous tone (respective pD2 values of 6.4 ± 0.3 and 7.2 ± 0.3 and EAmax of 65.0 ± 26.8 and 43.2 ± 30.1% of the maximal dilation obtained with papaverine) and bovine arterioles after preconstriction with serotonin (pD2 = 6.3 ± 0.1, EAmax = 80.0 ± 17.9% of induced tone). In contrast, nicotine (10−8 to 10−4 mol/L) failed to induce any vasomotor response in bovine vessels whether at spontaneous or at pharmacologically induced tone. Application of the nitric oxide synthase (NOS) inhibitor Nω-nitro-L-arginine (L-NNA; 10−5 mol/L) elicited a gradual constriction (∼20%) of the arterioles, indicating the presence of constitutive NO release in these vessels. Nω-Nitro-L-argigine (10−5 to 10−4 mol/L) also significantly blocked the dilation induced by ACh. The muscarinic ACh receptor (mAChR) antagonists pirenzepine, 4-DAMP, and AF-DX 384 dose dependently inhibited the dilatation induced by ACh (10−5 mol/L) with the following rank order of potency: 4-DAMP (pIC50 = 9.2 ± 0.3) ≫ pirenzepine (pIC50 = 6.7 ± 0.4) > AF-DX 384 (pIC50 = 5.9 ± 0.2). These results suggest that ACh can induce a potent, dose-dependent, and NO-mediated dilation of bovine and/or human intracortical arterioles via interaction with an mAChR that best corresponds to the M5 subtype.
Interplay Between the Gamma Isoform of PKC and Calcineurin in Regulation of Vulnerability to Focal Cerebral Ischemia
Authors: Aronowski Jaroslaw , Grotta James C. , Strong Roger , Waxham M. Neal
Published: November 2003

Protein phosphorylation and dephosphorylation mediated by protein kinases and protein phosphatases, respectively, represent essential steps in a variety of vital neuronal processes that could affect susceptibility to ischemic stroke. In this study, the role of the neuron-specific γ isoform of protein kinase C (γPKC) in reversible focal ischemia was examined using mutant mice in which the gene for γPKC was knockedout (γPKC-KO). A period of 150 minutes of unilateral middle cerebral artery and common carotid artery (MCA/CCA) occlusion followed by 21.5 hours of reperfusion resulted in significantly larger ( P < 0.005) infarct volumes (n = 10; 31.1 ± 4.2 mm3) in γPKC-KO than in wild-type (WT) animals (n = 12; 22.6 ± 7.4 mm3). To control for possible differences related to genetic background, the authors analyzed Balb/cJ, C57BL/6J, and 129SVJ WT in the MCA/CCA model of focal ischemia. No significant differences in stroke volume were detected between these WT strains. Impaired substrate phosphorylation as a consequence of γPKC-KO might be corrected by inhibition of protein dephosphorylation. To test this possibility, γPKC-KO mice were treated with the protein phosphatase 2B (calcineurin) inhibitor, FK-506, before ischemia. FK-506 reduced ( P < 0.008) the infarct volume in γPKC-KO mice (n = 7; 24.6 ± 4.6 mm3), but at this dose in this model, had no effect on the infarct volume in WT mice (n = 7; 20.5 ± 10.7 mm3). These results indicate that γPKC plays some neuroprotective role in reversible focal ischemia.
Hypoxic but Not Ischemic Neurotoxicity of Free Radicals Revealed by Dynamic Changes in Glucose Metabolism of Fresh Rat Brain Slices on Positron Autoradiography
Authors: Omata Naoto , Murata Tetsuhito , Fujibayashi Yasuhisa , Waki Atsuo , Sadato Norihiro , Yoshimoto Mitsuyoshi , Wada Yuji , Yonekura Yoshiharu
Published: November 2003

Dynamic changes in the regional cerebral glucose metabolic rate induced by hypoxia/reoxygenation or ischemia/reperfusion were investigated with a positron autoradiography technique. Fresh rat brain slices were incubated with [18F]2-fluoro-2-deoxy-D-glucose ([18F]FDG) in oxygenated Krebs-Ringer solution at 36°C, and serial two-dimensional time-resolved images of [18F]FDG uptake in the slices were obtained. In the case of loading hypoxia (oxygen deprivation)/pseudoischemia (oxygen and glucose deprivation) for various periods of time, the net influx constant ( K) of [18F]FDG at preloading and after reoxygenation/pseudoreperfusion (post-loading) was quantitatively evaluated by applying the Patlak graphical method to the image data. Regardless of the brain region, with hypoxia lasting ≥20 minutes, the postloading K value was decreased compared with the unloaded control, whereas with pseudoischemia of ≤40 minutes, approximately the same level as the unloaded control was maintained. Next, the neuroprotective effect against hypoxia/pseudoischemia loading induced by the addition of a free radical scavenger or an N-methyl-D-aspartate (NMDA) antagonist was assessed by determining whether a decrease in the postloading K value was prevented. Whereas with 20-minute hypoxia, both agents exhibited a neuroprotective effect, in the case of 50-minute pseudoischemia, only the NMDA antagonist did so, with the free radical scavenger being ineffective. These results demonstrate that hypoxia causes irreversible neuronal damage within a shorter period than ischemia, with both free radicals and glutamate suggested to be involved in tandem in the neurotoxicity induced by hypoxia, whereas glutamate alone is involved in ischemic neurotoxicity.
Evidence for Apoptosis After Intracerebral Hemorrhage in Rat Striatum
Authors: Matsushita Kohji , Meng Wei , Wang Xiaoying , Asahi Minoru , Asahi Kazuko , Moskowitz Michael A. , Lo Eng H.
Published: November 2003

The overall hypothesis that cell death after intracerebral hemorrhage is mediated in part by apoptotic mechanisms was tested. Intracerebral hemorrhage was induced in rats using stereotactic infusions of 0.5 U of collagenase (1-μL volume) into the striatum. After 24 hours, large numbers of TUNEL-positive stained cells with morphologies suggestive of apoptosis were present in the center and periphery of the hemorrhage. Double staining with Nissl and immunocytochemical labeling with antibodies against neuronal nuclei and glial fibrillary acidic protein suggested that these TUNEL-positive cells were mostly neurons and astrocytes. Electrophoresis of hemorrhagic brain extracts showed evidence of DNA laddering into ∼200-bp fragments. Western blots showed cleavage of the cytosolic caspase substrate gelsolin. The density of TUNEL-positive cells at 24 and 48 hours after hemorrhage was significantly reduced by treatment with the broad-spectrum caspase inhibitor zVADfmk. It was unlikely that apoptotic changes were due to neurotoxicity of injected collagenase because TUNEL-positive cells and DNA laddering were also obtained in an alternative model of hemorrhage where autologous blood was infused into the striatum. Furthermore, equivalent doses of collagenase did not induce cell death in primary neuronal cultures. These results provide initial evidence that apoptotic mechanisms may mediate some of the injury in brain after intracerebral hemorrhage.
Intraventricular Infusion of Apolipoprotein E Ameliorates Acute Neuronal Damage after Global Cerebral Ischemia in Mice
Authors: Horsburgh Karen , McCulloch James , Nilsen Margaret , McCracken Eileen , Large Charles , Roses Allen D. , Nicoll James A. R.
Published: November 2003

The ability of intraventricular infusion of apolipoprotein E (apoE) to reduce neuronal damage after global cerebral ischemia was investigated in apoE-deficient and wild-type mice. ApoE (5 μg/mL lipid-conjugated derived from human plasma; 1 μL/h, continuous infusion) significantly reduced neuronal damage in the caudate nucleus and CA2 pyramidal cell layer by approximately 50% in apoE-deficient mice after global ischemia compared to vehicle infusion. In wild-type mice infused with apoE, there was a trend for ischemic neuronal damage to be reduced. ApoE-infused mice had a marked reduction in 4-hydroxynonenal immunoreactivity, as a marker of lipid peroxidation. The results show that the presence of apoE at or after the time of injury can be neuroprotective, possibly via an anti-oxidant mechanism.
Administration of Selective Endothelin Receptor Type a Antagonist Ro 61-1790 Does Not Improve Outcome in Focal Cerebral Ischemia in Cat
Authors: Bhardwaj Anish , Wu Ying , Hurn Patricia D. , Kirsch Jeffrey R. , Traystman Richard J.
Published: November 2003

The authors examined the effect of selective endothelin (ET) receptor type A (ETA) antagonism on histological and functional recovery in cat at 24 hours after reversible middle cerebral artery occlusion (MCAO). A novel and specific ETA antagonist, Ro 61-1790 [5-methylpyridine-2-sulfonic acid-6-(2-hydroxyethoxy)-5-(2-methoxyphenoxy)-2-(2-1 H-tetrazol-5-yl-pyridin-4-yl)-pyrimidin-4-ylamide sodium salt (1:2)] (Roche, Basel, Switzerland), was used at doses that produced steady-state plasma concentrations and abolished ET-induced pial arteriolar vasoconstriction. In a cranial window preparation, 8 nmol/L ET constricted pial arterioles by 33 ± 18% (mean ± SD), but this response was ablated by intravenous Ro 61-1790 treatment (10-mg/kg bolus, 4-mg/kg/h infusion). In additional animal cohorts, halothane-anesthetized cats were treated with 90 minutes of MCAO and 24 hours of reperfusion. Animals received Ro 61-1790 infusion beginning at the onset of reperfusion and continuing for 6 or 24 hours (n = 41). Control cats were treated with 0.9% saline by intravenous infusion throughout reperfusion. There was no difference in injury volume or neurologic evaluation score in saline-treated cats (n = 11; caudate 24 ± 28%, cortical injury 7.5 ± 5% of ipsilateral structure; score 52 ± 8) versus the results in cats treated with Ro 61-1790 for either 24 hours (n = 6; caudate 22 ± 23%, cortex 6 ± 5%, injury volume of ipsilateral structure; score 55 ± 3) or 6 hours (n = 11; caudate 33 ± 30%, cortex 12 ± 14%, injury volume of ipsilateral structure; score 50 ± 10). Mortality was greatest in the 24-hour drug treatment group. These data suggest that blockade of ETA receptor activity is not beneficial to tissue or functional outcomes from experimental stroke in cat.
Transcription Factor Nuclear Factor-Kappa B is Activated in Neurons after Focal Cerebral Ischemia
Authors: Stephenson Diane , Yin Tinggui , Smalstig E. Barry , Hsu Mei Ann , Panetta Jill , Little Sheila , Clemens James
Published: November 2003

Nuclear factor-kappa B (NF-kB) is a multisubunit transcription factor that when activated induces the expression of genes encoding acute-phase proteins, cell adhesion molecules, cell surface receptors, and cytokines. NF-kB is composed of a variety of protein subunits of which p50-and p65-kDa (RelA) are the most widely studied. Under resting conditions, these subunits reside in the cytoplasm as an inactive complex bound by inhibitor proteins, IkBα and IkBβ. On activation, IkB is phosphorylated by IkB kinase and ubiquitinated and degraded by the proteasome; simultaneously, the active heterodimer translocates to the nucleus where it can initiate gene transcription. In the periphery, NF-kB is involved in inflammation through stimulation of the production of inflammatory mediators. The role of NF-kB in the brain is unclear. In vitro, NF-kB activation can be either protective or deleterious. The role of NF-kB in ischemic neuronal cell death in vivo was investigated. Adult male rats were subjected to 2 hours of focal ischemia induced by middle cerebral artery occlusion (MCAO). At 2, 6, and 12 hours after reperfusion, the expression and transactivation of NF-kB in ischemic versus nonischemic cortex and striatum were determined by immunocytochemistry and by electrophoretic mobility gel-shift analysis. At all time points studied, p50 and p65 immunoreactivity was found exclusively in the nuclei of cortical and striatal neurons in the ischemic hemisphere. The contralateral nonischemic hemisphere showed no evidence of nuclear NF-kB immunoreactivity. Double immunofluorescence confirmed expression of p50 in nuclei of neurons. Increased NF-kB DNA-binding activity in nuclear extracts prepared from the ischemic hemisphere was further substantiated by electrophoretic mobility gel-shift analysis. Because the activation of NF-kB by many stimuli can be blocked by antioxidants in vitro, the effect of the antioxidant, LY341122, previously shown to be neuroprotective, on NF-kB activation in the MCAO model was evaluated. No significant activation of NF-kB was found by electrophoretic mobility gel-shift analysis in animals treated with LY341122. These results demonstrate that transient focal cerebral ischemia results in activation of NF-kB in neurons and supports previous observations that neuroprotective antioxidants may inhibit neuronal death by preventing the activation of NF-kB.
Hypercarbia and Mild Hypothermia, Only When Not Combined, Improve Postischemic Bioenergetic Recovery in Neonatal Rat Brain Slices
Authors: Tasker Robert C. , Sahota Sati K. , Williams Stephen R.
Published: November 2003

In the immature brain, postischemic metabolism may be influenced beneficially by the effect of inducing hypercarbia or hypothermia. With use of 31P nuclear magnetic resonance spectroscopy, intracellular pH (pHi) and cellular energy metabolites in ex vivo neonatal rat cerebral cortex were measured before, during, and after substrate and oxygen deprivation in in vitro ischemia. Early postischemic hypothermia (fall in temperature −3.2 ± 1.0°C) delayed the normalization of pHi after ischemia by inducing an acid shift in pHi ( P < 0.01). Postischemic hypercarbia (Krebs—Henseleit bicarbonate buffer equilibrated with 10% carbon dioxide in oxygen) and hypothermia induced separate, but potentially additive, reversible decreases in pHi, each of approximately −0.16 pH unit ( P < 0.05). When these postischemic perturbations were applied in isolation, there was significant improvement of ∼20% in the recovery of β-ATP ( P < 0.05). In combination, however, hypercarbia and hypothermia worsened recovery in ATP by ∼20% ( P < 0.05). In control tissue, which had not been exposed to ischemia, ATP content was also significantly reduced by co-administration of the two treatments ( P < 0.05), an effect that persisted even after discontinuing the perturbing conditions. Therefore, in this vascular-independent neonatal preparation, early postischemic modulation of metabolism by hypercarbia or hypothermia appears to confer improved bioenergetic recovery, but only if they are not administered together.
Estrogen as a Neuroprotectant in Stroke
Authors: Hurn Patricia D. , Macrae I. Mhairi
Published: November 2003

Recent evidence suggests that reproductive steroids are important players in shaping stroke outcome and cerebrovascular pathophysiologic features. Although women are at lower risk for stroke than men, this native protection is lost in the postmenopausal years. Therefore, aging women sustain a large burden for stroke, contrary to a popular misconception that cancer is the main killer of women. Further, the value of hormone replacement therapy in stroke prevention or in improving outcome remains controversial. Estrogen has been the best studied of the sex steroids in both laboratory and clinical settings and is considered increasingly to be an endogenous neuroprotective agent. A growing number of studies demonstrate that exogenous estradiol reduces tissue damage resulting from experimental ischemic stroke in both sexes. This new concept suggests that dissecting interactions between estrogen and cerebral ischemia will yield novel insights into generalized cellular mechanisms of injury. Less is known about estrogen's undesirable effects in brain, for example, the potential for increasing seizure susceptibility and migraine. This review summarizes gender-specific aspects of clinical and experimental stroke and results of estrogen treatment on outcome in animal models of cerebral ischemia, and briefly discusses potential vascular and parenchymal mechanisms by which estrogen salvages brain.
Endothelial Nitric Oxide Synthase-Dependent Cerebral Blood Flow Augmentation by L-Arginine After Chronic Statin Treatment
Authors: Yamada Masaru , Huang Zhihong , Dalkara Turgay , Endres Matthias , Laufs Ulrich , Waeber Christian , Huang Paul L. , Liao James K. , Moskowitz Michael A.
Published: November 2003

Nitric oxide, a product of nitric oxide synthase activity, relaxes vascular smooth muscle and elevates brain blood flow. We evaluated the importance of eNOS to cerebral blood flow augmentation after L-arginine infusion and increases in flow after eNOS upregulation in SV-129 mice. Blood flow was measured by laser-Doppler flowmetry before and after L-arginine infusion (450 mg/kg during a 15-minute period) or measured by 14C-iodoamphetamine indicator fractionation or 14C-iodoantipyrine tissue equilibration techniques. rCBF increased by 26% (laser Doppler flowmetry) after L-arginine infusion but did not change in mutant mice deficient in eNOS expression. After eNOS upregulation by chronic simvastatin treatment (2 mg/kg subcutaneously, daily for 14 days), L-arginine amplified and sustained the hyperemia (38%) and increased absolute brain blood flow from 86 ± 7 to 119 ± 10 mL/100 g per minute. Furthermore, pretreatment with simvastatin enhanced blood flow within ischemic brain tissue after middle cerebral artery occlusion. Together, these findings suggest that eNOS activity is critical for blood flow augmentation during acute L-arginine infusion, and chronic eNOS upregulation combined with L-arginine administration provides a novel strategy to elevate cerebral blood flow in the normal and ischemic brain.
NMDA Receptor Blockade Fails to Alter Axonal Injury in Focal Cerebral Ischemia
Authors: Yam Philippa S. , Dunn Laurence T. , Graham David I. , Dewar Deborah , McCulloch James
Published: November 2003

The ability of the NMDA receptor antagonist, MK-801, to protect myelinated axons after focal cerebral ischemia has been examined. Amyloid precursor protein (APP) immunocytochemistry was used to assess the anatomic extent of axonal injury, and conventional histopathology was used to assess the volume of ischemic damage to neuronal perikarya. The middle cerebral artery was permanently occluded in 16 cats. The cats were treated with either vehicle or MK-801 as a 0.5-mg/kg bolus at 15 minutes before middle cerebral artery occlusion, followed by an infusion of 0.14 mg/kg per hour. After 6 hours, the animals were killed and the brains processed for histology and immunocytochemistry. The volume of neuronal necrosis was determined from 16 preselected coronal levels of the brain. The circumscribed zones of APP accumulation in axons were mapped onto images at the same 16 coronal levels, and quantitative analysis was performed using a transparent counting grid, randomly placed over each image. The histologic appearance and anatomic location of axons with increased APP immunoreactivity was similar in animals treated with vehicle and MK-801. MK-801 failed to reduce the hemispheric APP score significantly. In vehicle-treated animals, there was a significant association between the volume of neuronal necrosis and the amount of APP immunoreactivity. MK-801 significantly reduced the slope of the association between the volume of neuronal necrosis and the amount of APP immunoreactivity compared with that observed in vehicle-treated animals. As a result, the ratio of hemispheric APP score and volume of neuronal necrosis was significantly increased with MK-801 treatment. The inability of NMDA receptor antagonists to protect axons may limit their functional efficacy in improving functional outcome after stroke.
Excitotoxic Hippocampal Injury is Attenuated in Thioredoxin Transgenic Mice
Authors: Takagi Yasushi , Hattori Itaro , Nozaki Kazuhiko , Mitsui Akira , Ishikawa Masatsune , Hashimoto Nobuo , Yodoi Junji
Published: November 2003

Thioredoxin is a small, multifunctional protein with a redox-active disulfide/dithiol in the active site. Thioredoxin plays several important biologic roles both in intracellular and extracellular compartments with its redox-regulating and reactive oxygen intermediates scavenging activities. We assayed the seizure response and the excitotoxic hippocampal injury in thioredoxin transgenic and wild-type C57BL/6 mice. Seizure score after kainic acid treatment was significantly lower in thioredoxin transgenic mice. Seven days after kainic acid administration, the damage in the hippocampal CA1 and CA3 regions was significantly attenuated in thioredoxin transgenic mice. Thioredoxin and redox regulation play an important role in excitotoxic brain damage.
Dexmedetomidine-Induced Stimulation of Glutamine Oxidation in Astrocytes: A Possible Mechanism for its Neuroprotective Activity
Authors: Huang Rong , Chen Ye , Yu A. C. H. , Hertz Leif
Published: November 2003

Dexmedetomidine is a highly specific α2-adrenergic agonist, which is used clinically as an anesthetic adjuvant and in animal experiments has a neuroprotective effect during ischemia. The current study showed that dexmedetomidine enhances glutamine disposal by oxidative metabolism in astrocytes. This effect occurs at pharmacologically relevant concentrations. It is exerted on α2-adrenergic receptors and not on imidazoline-preferring sites, and it is large enough to reduce the availability of glutamine as a precursor of neurotoxic glutamate.
Kinetic Analysis of the 5-HT2A Ligand [11C]MDL 100,907
Authors: Watabe Hiroshi , Channing Michael A. , Der Margaret G. , Adams H. Richard , Jagoda Elaine , Herscovitch Peter , Eckelman William C. , Carson Richard E.
Published: November 2003

The goal of this study was to develop a suitable kinetic analysis method for quantification of 5-HT2A receptor parameters with [11C]MDL 100,907. Twelve control studies and four preblocking studies (400 nmol/kg unlabeled MDL 100,907) were performed in isoflurane-anesthetized rhesus monkeys. The plasma input function was determined from arterial blood samples with metabolite measurements by extraction in ethyl acetate. The preblocking studies showed that a two-tissue compartment model was necessary to fit the time activity curves of all brain regions including the cerebellum—in other words, the need for two compartments is not proof of specific binding. Therefore, a three-tissue compartment model was used to analyze the control studies, with three parameters fixed based on the preblocking data. Reliable fits of control data could be obtained only if no more than three parameters were allowed to vary. For routine use of [11C]MDL 100,907, several simplified methods were evaluated. A two-tissue (2T‘) compartment with one fixed parameter was the most reliable compartmental approach; a one-compartment model failed to fit the data adequately. The Logan graphical approach was also tested and produced comparable results to the 2T’ model. However, a simulation study showed that Logan analysis produced a larger bias at higher noise levels. Thus, the 2T' model is the best choice for analysis of [11C]MDL 100,907 studies.
Cerebral Hemodynamics in Human Acute Ischemic Stroke: A Study with Diffusion- and Perfusion-Weighted Magnetic Resonance Imaging and SPECT
Authors: Liu Yawu , Karonen Jari O. , Vanninen Ritva L. , Østergaard Leif , Roivainen Reina , Nuutinen Juho , Perkio Jussi , Könönen Mervi , Hämäläinen Anne , Vanninen Esko J. , Soimakallio Seppo , Kuikka Jyrki T. , Aronen Hannu J.
Published: November 2003

Nineteen patients with acute ischemic stroke (<24 hours) underwent diffusion-weighted and perfusion-weighted (PWI) magnetic resonance imaging at the acute stage and 1 week later. Eleven patients also underwent technetium-99m ethyl cysteinate dimer single-photon emission computed tomography (SPECT) at the acute stage. Relative (ischemic vs. contralateral control) cerebral blood flow (relCBF), relative cerebral blood volume, and relative mean transit time were measured in the ischemic core, in the area of infarct growth, and in the eventually viable ischemic tissue on PWI maps. The relCBF was also measured from SPECT. There was a curvilinear relationship between the relCBF measured from PWI and SPECT ( r = 0.854; P < 0.001). The tissue proceeding to infarction during the follow-up had significantly lower initial CBF and cerebral blood volume values on PWI maps ( P < 0.001) than the eventually viable ischemic tissue had. The best value for discriminating the area of infarct growth from the eventually viable ischemic tissue was 48% for PWI relCBF and 87% for PWI relative cerebral blood volume. Combined diffusion and perfusion-weighted imaging enables one to detect hemodynamically different subregions inside the initial perfusion abnormality. Tissue survival may be different in these subregions and may be predicted.
Multiple Molecular Penumbras after Focal Cerebral Ischemia
Authors: Sharp Frank R. , Lu Aigang , Tang Yang , Millhorn David E.
Published: November 2003

Though the ischemic penumbra has been classically described on the basis of blood flow and physiologic parameters, a variety of ischemic penumbras can be described in molecular terms. Apoptosis-related genes induced after focal ischemia may contribute to cell death in the core and the selective cell death adjacent to an infarct. The HSP70 heat shock protein is induced in glia at the edges of an infarct and in neurons often at some distance from the infarct. HSP70 proteins are induced in cells in response to denatured proteins that occur as a result of temporary energy failure. Hypoxia-inducible factor (HIF) is also induced after focal ischemia in regions that can extend beyond the HSP70 induction. The region of HIF induction is proposed to represent the areas of decreased cerebral blood flow and decreased oxygen delivery. Immediate early genes are induced in cortex, hippocampus, thalamus, and other brain regions. These distant changes in gene expression occur because of ischemia-induced spreading depression or depolarization and could contribute to plastic changes in brain after stroke.
Quantitation of Regional Cerebral Blood Flow Corrected for Partial Volume Effect Using O-15 Water and PET: II. Normal Values and Gray Matter Blood Flow Response to Visual Activation
Authors: Law Ian , Iida Hidehiro , Holm Søren , Nour Sam , Rostrup Egill , Svarer Claus , Paulson Olaf B.
Published: November 2003

One of the most limiting factors for the accurate quantification of physiologic parameters with positron emission tomography (PET) is the partial volume effect (PVE). To assess the magnitude of this contribution to the measurement of regional cerebral blood flow (rCBF), the authors have formulated four kinetic models each including a parameter defining the perfusable tissue fraction (PTF). The four kinetic models used were 2 one-tissue compartment models with (Model A) and without (Model B) a vascular term and 2 two-tissue compartment models with fixed (Model C) or variable (Model D) white matter flow. Furthermore, rCBF based on the autoradiographic method was measured. The goals of the study were to determine the following in normal humans: (1) the optimal model, (2) the optimal length of fit, (3) the model parameters and their reproducibility, and (4) the effects of data acquisition (2D or 3D). Furthermore, the authors wanted to measure the activation response in the occipital gray matter compartment, and in doing so test the stability of the PTF, during perturbations of rCBF induced by visual stimulation. Eight dynamic PET scans were acquired per subject (n = 8), each for a duration of 6 minutes after IV bolus injection of H215O. Four of these scans were performed using 2D and four using 3D acquisition. Visual stimulation was presented in four scans, and four scans were during rest. Model C was found optimal based on Akaike's Information Criteria (AIC) and had the smallest coefficient of variance after a 6-minute length of fit. Using this model the average PVE corrected rCBF during rest in gray matter was 1.07 mL·min−1·g−1 (0.11 SD), with an average coefficient of variance of 6%. Acquisition mode did not affect the estimated parameters, with the exception of a significant increase in the white matter rCBF using the autoradiographic method (2D: 0.17 mL·min−1·g−1 (0.02 SD); 3D: 0.21 mL·min−1·g−1 (0.02 SD)). At a 6-minute fit the average gray matter CBF using Models C and D were increased by 100% to 150% compared with Models A and B and the autoradiographic method. There were no significant changes in the perfusable tissue fraction by the activation induced rCBF increases. The largest activation response was found using Model C (median = 39.1%). The current study clearly demonstrates the importance of PVE correction in the quantitation of rCBF in normal humans. The potential use of this method is to cost-effectively deliver PVE corrected measures of rCBF and tissue volumes without reference to imaging modalities other than PET.
Serine–Threonine Protein Kinase Akt does Not Mediate Ischemic Tolerance after Global Ischemia in the Gerbil
Authors: Namura Shobu , Nagata Izumi , Kikuchi Haruhiko , Andreucci Michele , Alessandrini Alessandro
Published: November 2003

The protein kinase Akt/PKB has been implicated in antiapoptosis and neuronal survival. The authors now show that Akt is phosphorylated in the hippocampus during the early reperfusion period after 3.5 minutes bilateral carotid artery occlusion (BCAO) in the gerbil. Repeated sublethal ischemia induces ischemic tolerance, which is known as ischemic preconditioning. Ischemic preconditioning does not affect the amount of Akt protein, but rather decreases the phosphorylation of Akt at Ser-473 after 10 minutes reperfusion after 3.5 minutes BCAO. These results suggest that although Akt may play a role in neuronal survival after ischemia, it may not play a role in ischemic tolerance by preconditioning.
Quantitative Comparison of Functional Contrast from BOLD-Weighted Spin-Echo and Gradient-Echo Echoplanar Imaging at 1.5 Tesla and H215O PET in the Whole Brain
Authors: Lowe Mark J. , Lurito Joseph T. , Mathews Vincent P. , Phillips Micheal D. , Hutchins Gary D.
Published: November 2003

Spin-echo and gradient-echo echoplanar functional magnetic resonance imaging (fMRI) studies at 1.5 Tesla (T) were used to obtain blood oxygenation level-dependent (BOLD) contrast images of the whole brain in seven strongly right-handed women during execution of a complex motor task. Five subjects underwent subsequent H215O positron emission tomography (PET) studies while performing the same task. Group-averaged results for changes in the MRI relaxation rates R2* and R2 at 1.5T in response to neuronal activation in nine cortical, subcortical, and cerebellar motor regions are reported. Results for each method are grouped according to tissue type—cerebral cortex (precentral gyrus and supplementary motor area), subcortical regions (thalamus and putamen), and cerebellar cortex (superior lobule). The observed changes in R2* from activation-induced oxygenation changes were more variable across brain regions with different tissue characteristics than observed changes in R2. The ratio of ΔR2* to ΔR2 was 3.3 ± 0.9 for cerebral cortex and 2.0 ± 0.6 for subcortical tissue. ΔR2*, ΔR2, and relative blood flow changes were ΔR2* = −0.201 ± 0.040 s−1, ΔR2 = −0.064 ± 0.011 s−1, and Δf/f = 16.7 ± 0.8% in the cerebral cortex; ΔR2* = −0.100 ± 0.026 s−1, ΔR2 = −0.049 ± 0.009 s−1, and Δf/f = 9.4 ± 0.7% in the subcortical regions; and ΔR2* = −0.215 ± 0.093 s−1, ΔR2 = −0.069 ± 0.012 s−1, and Δf/f = 16.2 ± 1.2% in the cerebellar cortex.
Functional Imaging Correlates of Recovery after Stroke in Humans
Authors: Herholz Karl , Heiss Wolf-Dieter
Published: November 2003

The mechanisms that are responsible for the remarkable potential for functional recovery from stroke in humans remain unclear, and functional tomographic neuroimaging techniques increasingly are being used to investigate this issue. Such studies confirmed that recovery of function is related to the volume of penumbra tissue that escapes infarction. For language, reactivation of the primary functional areas in the dominant hemisphere is associated with the best prognosis. Evidence for functional plasticity in the immediate vicinity of infarcts, as demonstrated under experimental conditions with invasive methods, is still limited after stroke in humans, probably because of the limitations of spatial resolution of most currently available methods. Often, functional changes in the large-scale networks that support motor (for example, supplementary and premotor cortex) and language functions (for example, prefrontal cortex) have been found, more extensively after lesions acquired during childhood than at adult age. A frequent finding is an increase in the cerebral blood flow response in corresponding regions of the healthy hemisphere during unilateral motor activation or language activation. It is, however, not yet clear whether that is related to functional recovery, and there are several observations indicating that it is often inefficient. Further systematic follow-up studies and therapeutic intervention trials are needed to clarify these issues.
Serial Changes in Cerebral Blood Flow and Flow—Metabolism Uncoupling in Primates with Acute Thromboembolic Stroke
Authors: Kuge Yuji , Yokota Chiaki , Tagaya Masafumi , Hasegawa Yasuhiro , Nishimura Akira , Kito Go , Tamaki Nagara , Hashimoto Naoto , Yamaguchi Takenori , Minematsu Kazuo
Published: November 2003

The authors recently developed a primate thromboembolic stroke model. To characterize the primate model, the authors determined serial changes in cerebral blood flow (CBF) and the relation between CBF and cerebral metabolic rate of glucose (CMRglc) using high-resolution positron emission tomography. Thromboembolic stroke was produced in male cynomolgus monkeys (n = 4). Acute obstruction of the left middle cerebral artery was achieved by injecting an autologous blood clot into the left internal carotid artery. Cerebral blood flow was measured with [15O]H2O before and 1, 2, 4, 6, and 24 hours after embolization. CMRglc was measured with 2-[18F]fluoro-2-deoxy-D-glucose ([18F]FDG) 24 hours after embolization. Lesion size and location 24 hours after embolization was determined by the 2,3,5-triphenyltetrazolium chloride (TTC) staining method. The results are summarized as follows: (1) 1 hour after embolization, CBF in the temporal cortex and the basal ganglia decreased to < 40% of the contralateral values. In these regions, regarded as an ischemic core, CBF decreased further with time and CMRglc at 24 hours also decreased. Infarcted lesions as indicated by being unstained with TTC were consistently observed in these regions. (2) In the parietal cortex and several regions surrounding the ischemic core, CBF was >40% of the contralateral values 1 hour after embolization and recovered gradually with time (ischemic penumbra). In these regions, CMRglc at 24 hours increased compared with that in the contralateral regions, indicating an uncoupling of CBF and CMRglc. No obvious TTC-unstained lesions were detected in these regions. The authors demonstrated a gradual recovery of reduced CBF, an elevated CMRglc and a CBF-CMRglc uncoupling in the penumbra regions of the primate model. Positron emission tomography investigations using this model will provide better understanding of the pathophysiology of thromboembolic stroke in humans.
bcl-2 Antisense Treatment Prevents Induction of Tolerance to Focal Ischemia in the Rat Brain
Authors: Shimizu Shigetoshi , Nagayama Tetsuya , Jin Kun Lin , Zhu Li , Loeffert J. Eric , Watkins Simon C. , Graham Steven H. , Simon Roger P.
Published: November 2003

In the rat, 60 minutes of transient ischemia to the middle cerebral artery results in infarction of the caudate putamen. Ischemic preconditioning with 20 minutes of transient focal ischemia produced tolerance (attenuated infarction volume) to 60 minutes of subsequent focal ischemia administered three days, five days, or seven days later. Western blots from tolerant caudate putamen demonstrated increased bcl-2 expression, maximum at 3 days and persisting through 7 days. Immunocytochemical examination found that bcl-2 was expressed in cells with both neuronal and nonneuronal morphology in striatum after preconditioning ischemia. bcl-2 antisense oligodeoxynucleotides (ODNs), bcl-2 sense ODNs, or artificial cerebrospinal fluid (CSF, vehicle) was infused into the lateral ventricle for the 72 hours between the 20-minute ischemic preconditioning and the 60-minute period of ischemia. Antisense ODN treatment reduced expression of bcl-2 in the striatum and blocked the induction of tolerance by preconditioning ischemia. Sense and CSF treatments had no effect on either bcl-2 expression or tolerance. In this model of induced tolerance to focal ischemia, bcl-2 appears to be a major determinant.
Amyloid β Peptide–Induced Cerebral Endothelial Cell Death Involves Mitochondrial Dysfunction and Caspase Activation
Authors: Xu Jan , Chen Shawei , Ku Grace , Ahmed S. Hinan , Xu Jinming , Chen Hong , Hsu Chung Y.
Published: November 2003

Amyloid β peptide (Aβ), a 39 to 43 amino acid fragment of the β-amyloid precursor protein (βAPP), forms insoluble fibrillar accumulation in neurofibrillary tangles and vascular plaques. Aβ has been implicated in neuronal and vascular degeneration in brain regions susceptible to plaque formation because of its cytotoxic effect on neurons and endothelial cells (ECs). The authors used a murine cerebral endothelial cell (CEC) line and primary cultures of bovine CECs to explore the cytotoxic mechanism of Aβ. Aβ 1–40 and Aβ 25–35 peptides caused cell death in a dose-dependent and time-dependent manner. Exposure to either Aβ 25–35 or Aβ 1–40 at 10 μmol/L for 48 hours caused at least 40% cell death. Cerebral endothelial cell death was characterized by nuclear condensation, mitochondrial dysfunction, and nuclear and mitochondrial DNA damage. Aβ 25–35 activated both caspase-8 and caspase-3 in murine CECs. zVAD-fmk, a broad-spectrum caspase inhibitor, prevented Aβ 25–35-induced increase in caspase-3 activity and CEC death. N-acetyl-cysteine, an antioxidant, also prevented Aβ-induced cell death. Together, these findings indicate that Aβ-mediated CEC death is an apoptotic process that is characterized by increased oxidative stress, caspase activation, mitochondrial dysfunction, and nuclear and mitochondrial DNA damage.
Induction of the HSP110/105 Family in the Rat Hippocampus in Cerebral Ischemia and Ischemic Tolerance
Authors: Yagita Yoshiki , Kitagawa Kazuo , Ohtsuki Toshiho , Tanaka Shigeru , Hori Masatsugu , Matsumoto Masayasu
Published: November 2003

Recently, the authors isolated a novel gene of the HSP110 family, ischemia responsive protein 94 kDa (irp94), and demonstrated the expression of this gene after transient forebrain ischemia. In the current study, the authors investigated the expression profiles of all HSP110 family members including hsp110/105 and osp94/apg-1, after transient forebrain ischemia using rat four-vessel occlusion model. Among three members of the HSP110 family, induction of hsp110/105 was the most prominent after ischemia. hsp110/105 mRNA expression was clearly enhanced from 4 to 24 hours after a 6-minute or longer ischemic period. First, hsp110/105 mRNA expression was induced in the dentate gyrus, and later in the pyramidal layer. HSP110/105 protein expression also was enhanced by a 6-minute or longer period of ischemia. Profiles of HSP110/105 expression after ischemia were similar to those of inducible HSP70. After transient forebrain ischemia for 10 minutes, HSP110/105 protein was induced in the dentate gyrus and the CA3 pyramidal layer, but not in the CA1 pyramidal neurons. However, 6 minutes of ischemia induced the HSP110/105 protein, as well as the HSP70 protein, in the CA1 region. CA1 pyramidal neurons expressing HSP110/105 acquired tolerance against subsequent severe ischemia. In conclusion, HSP110/105 showed the most prominent induction after ischemia among the three HSP110 gene family members. Colocalization of HSP110/105 and HSP70 in the CA1 neurons that acquired tolerance suggested that induced HSP110/105 might contribute to ischemic tolerance together with HSP70.
Delayed Hemorrhagic Hypotension Exacerbates the Hemodynamic and Histopathologic Consequences of Traumatic Brain Injury in Rats
Authors: Matsushita Yoshitaro , Bramlett Helen M. , Kuluz John W. , Alonso Ofelia , Dietrich W. Dalton
Published: November 2003

Alterations in cerebral autoregulation and cerebrovascular reactivity after traumatic brain injury (TBI) may increase the susceptibility of the brain to secondary insults, including arterial hypotension. The purpose of this study was to evaluate the consequences of mild hemorrhagic hypotension on hemodynamic and histopathologic outcome after TBI. Intubated, anesthetized male rats were subjected to moderate (1.94 to 2.18 atm) parasagittal fluid–percussion (FP) brain injury. After TBI, animals were exposed to either normotension (group 1: TBI alone, n = 6) or hypotension (group 2: TBI + hypotension, n = 6). Moderate hypotension (60 mm Hg/30 min) was induced 5 minutes after TBI or sham procedures by hemorrhage. Sham-operated controls (group 3, n = 7) underwent an induced hypotensive period, whereas normotensive controls (group 4, n = 4) did not. For measuring regional cerebral blood flow (rCBF), radiolabeled microspheres were injected before, 20 minutes after, and 60 minutes after TBI (n = 23). For quantitative histopathologic evaluation, separate groups of animals were perfusion-fixed 3 days after TBI (n = 22). At 20 minutes after TBI, rCBF was bilaterally reduced by 57% ± 6% and 48% ± 11% in cortical and subcortical brain regions, respectively, under normotensive conditions. Compared with normotensive TBI rats, hemodynamic depression was significantly greater with induced hypotension in the histopathologically vulnerable (P1) posterior parietal cortex ( P < 0.01). Secondary hypotension also increased contusion area at specific bregma levels compared with normotensive TBI rats ( P < 0.05), as well as overall contusion volume (0.96 ± 0.46 mm3 vs. 2.02 ± 0.51 mm3, mean ± SD, P < 0.05). These findings demonstrate that mild hemorrhagic hypotension after FP injury worsens local histopathologic outcome, possibly through vascular mechanisms.
Protein Aggregation after Focal Brain Ischemia and Reperfusion
Authors: Hu Bing-Ren , Janelidze Shorena , Ginsberg Myron D. , Busto Raul , Perez-Pinzon Miguel , Sick Thomas J. , Siesjö Bo K. , Liu C. L.
Published: November 2003

Two hours of transient focal brain ischemia causes acute neuronal death in the striatal core region and a somewhat more delayed type of neuronal death in neocortex. The objective of the current study was to investigate protein aggregation and neuronal death after focal brain ischemia in rats. Brain ischemia was induced by 2 hours of middle cerebral artery occlusion. Protein aggregation was analyzed by electron microscopy, laser-scanning confocal microscopy, and Western blotting. Two hours of focal brain ischemia induced protein aggregation in ischemic neocortical neurons at 1 hour of reperfusion, and protein aggregation persisted until neuronal death at 24 hours of reperfusion. Protein aggregates were found in the neuronal soma, dendrites, and axons, and they were associated with intracellular membranous structures during the postischemic phase. High-resolution confocal microscopy showed that clumped protein aggregates surrounding nuclei and along dendrites were formed after brain ischemia. On Western blots, ubiquitinated proteins (ubi-proteins) were dramatically increased in neocortical tissues in the postischemic phase. The ubi-proteins were Triton-insoluble, indicating that they might be irreversibly aggregated. The formation of ubi-protein aggregates after ischemia correlated well with the observed decrease in free ubiquitin and neuronal death. The authors concluded that proteins are severely damaged and aggregated in neurons after focal ischemia. The authors propose that protein damage or aggregation may contribute to ischemic neuronal death.
Quantification of Neuroreceptors in Living Human Brain. V. Endogenous Neurotransmitter Inhibition of Haloperidol Binding in Psychosis
Authors: Gjedde Albert , Wong Dean F.
Published: November 2003

The half-inhibition concentration (IC50) of a drug indicates its ability to inhibit the binding of other ligands of a receptor. The authors used positron emission tomography to test the hypothesis that haloperidol's IC50 toward the binding of tracer N-[11C]methylspiperone ([11C]NMSP) in brain must be increased in patients in whom more dopamine is bound to receptors than in healthy volunteers. The IC50 of haloperidol was significantly elevated from 1.5 nmol/L in healthy volunteers and patients with bipolar disease without psychosis to 4.5 nmol/L in patients with schizophrenia or bipolar disease with psychosis. The higher IC50 values in psychosis are consistent with an 8-fold increased binding of dopamine and a 16-fold elevated concentration of synaptic dopamine in psychosis. At the 80% haloperidol blockade of the receptors, the calculated amount of neurotransmitter bound in the patients with psychosis declined to twice the value estimated in the nonpsychotic subjects, that is, 5 pmol cm−3.
Effects of Moderate Hypothermia on IL-1β–Induced Leukocyte Rolling and Adhesion in Pial Microcirculation of Mice and on Proinflammatory Gene Expression in Human Cerebral Endothelial Cells
Authors: Sutcliffe Ian T. , Smith Holly A. , Stanimirovic Danica , Hutchison James S.
Published: November 2003

Although the neuroprotective effects of hypothermia have been known for a long time, the molecular correlates of this neuroprotection are poorly understood. In this study, the authors investigated how hypothermia affects inflammatory responses in the brain elicited by systemic injection of IL-1β. Leukocyte rolling and adhesion were quantified in pial venules (20 to 50 μm) of C57/Bl6 mice 4 hours after intraperitoneal injection of IL-1β (5 μg/kg) using an open cranial window and intravital microscopy. Animals were subjected to moderate hypothermia (32°C) or normothermia (37°C) for 1 or 4 hours after IL-1β injection. Significant increases in leukocyte rolling and adhesion were observed in IL-1β–injected animals as compared with sham controls. Whereas 1-hour hypothermia did not affect IL-1β–induced leukocyte rolling and adhesion, 4-hour hypothermia caused a reduction in both rolling and adhesion. Molecular mechanisms of hypothermic effects were investigated in cultured human cerebral endothelial cells exposed to IL-1β (50 U/mL) for 4 hours at 37°C or 32°C followed by 18 hours at 37°C. Human cerebral endothelial cells exposed to IL-1β at 32°C showed attenuated NF-κB activation determined by the Luciferase yellow reporter gene assay and reduced expression of IL-8 and IL-1β measured by reverse transcriptase–polymerase chain reaction and enzyme-linked immunosorbent assay. Intracellular adhesion molecule-1 was induced to similar levels (threefold over control) at both temperatures. The expression of CD18 on neutrophils in vitro was not affected by either IL-1β or hypothermia. These findings suggest that mechanisms by which hypothermia reduces leukocyte rolling and adhesion include suppression of inflammatory gene transcription in brain endothelial cells.