bluuebunny/crossref_metadata_2025 huggingface dataset:
Preconditioning In Vivo Ischemia Inhibits Anoxic Long-Term Potentiation and Functionally Protects CA1 Neurons in the Gerbil
Authors: Kawai Kensuke , Nakagomi Tadayoshi , Kirino Takaaki , Tamura Akira , Kawai Nobufumi
Published: November 2003

Preconditioning with sublethal ischemia induces tolerance to subsequent lethal ischemia in neurons. We investigated electrophysiologic aspects of the ischemic tolerance phenomenon in the gerbil hippocampus. Gerbils were subjected to 2 minutes of forebrain ischemia (preconditioning ischemia). Some of them were subjected to a subsequent 5 minutes of forebrain ischemia 2 to 3 days after the preconditioning ischemia (double ischemia). Hippocampal slices were prepared from these gerbils subjected to the preconditioning or double ischemia, and field excitatory postsynaptic potentials were recorded from CA1 pyramidal neurons. Capacity for long-term potentiation triggered by tetanic stimulation (tetanic LTP) was transiently inhibited 1 to 2 days after the double ischemia but then recovered. Latency of anoxic depolarization was not significantly different between slices from preconditioned gerbils and those from sham-operated gerbils when these slices were subjected to in vitro anoxia. Postanoxic potentiation of N-methyl-D-aspartate (NMDA) receptor-mediated transmission (anoxic LTP) was inhibited in slices from gerbils 2 to 3 days after the preconditioning ischemia, whereas it was observed in slices from sham-operated gerbils and gerbils 9 days after the preconditioning ischemia. These results suggest that protection by induced tolerance is (1) not only morphologic but also functional, and (2) expressed in inhibiting postischemic overactivation of NMDA receptor-mediated synaptic responses.
Evidence that Functional Glutamate Receptors are not Expressed on Rat or Human Cerebromicrovascular Endothelial Cells
Authors: Morley Paul , Small Daniel L. , Murray Christine L. , Mealing Geoffrey A. , Poulter Michael O. , Durkin Jon P. , Stanimirovic Danica B.
Published: November 2003

Excitatory amino acids can modify the tone of cerebral vessels and permeability of the blood-brain barrier (BBB) by acting directly on endothelial cells of cerebral vessels or indirectly by activating receptors expressed on other brain cells. In this study we examined whether rat or human cerebromicrovascular endothelial cells (CEC) express ionotropic and metabotropic glutamate receptors. Glutamate and the glutamate receptor agonists N-methyl-d-aspartate (NMDA), α-amino-3-hydroxy-5-methyl-isoxazole-4-propionic acid (AMPA), and kainate failed to increase [Ca2+]i in either rat or human microvascular and capillary CEC but elicited robust responses in primary rat cortical neurons, as measured by fura-2 fluorescence. The absence of NMDA and AMPA receptors in rat and human CEC was further confirmed by the lack of immunocytochemical staining of cells by antibodies specific for the AMPA receptor subunits GluR1, GluR2/3, and GluR4 and the NMDA receptor subunits NR1, NR2A, and NR2B. We failed to detect mRNA expression of the AMPA receptor subunits GluR1 to GluR4 or the NMDA receptor subunits NR11XX, NR10XX, and NR2A to NR2C in both freshly isolated rat and human microvessels and cultured CEC using reverse transcriptase polymerase chain reaction (RT-PCR). Cultured rat CEC expressed mRNA for KA1 or KA2 and GluR5 subunits. Primary rat cortical neurons were found to express GluR1 to GluR3 and NR1, NR2A, and NR2B by both immunocytochemistry and RT-PCR and KA1, KA2, GluR5, GluR6, and GluR7 by RT-PCR. Moreover, the metabotropic glutamate receptor agonist 1-amino-cyclopentyl-1 S, 3 R-dicorboxylate (1 S,3 R-trans-ACPD), while eliciting both inositol trisphosphate and [Ca2+]i increases and inhibiting forskolin-stimulated cyclic AMP in cortical neurons, was unable to induce either of these responses in rat or human CEC. These results strongly suggest that both rat and human CEC do not express functional glutamate receptors. Therefore, excitatory amino acid-induced changes in the cerebral microvascular tone and BBB permeability must be affected indirectly, most likely by mediators released from the adjacent glutamate-responsive cells.
Absolute Cerebral Blood Flow and Blood Volume Measured by Magnetic Resonance Imaging Bolus Tracking: Comparison with Positron Emission Tomography Values
Authors: Østergaard Leif , Smith Donald F. , Vestergaard-Poulsen Peter , Hansen SørenB. , Gee Antony D. , Gjedde Albert , Gyldensted Carsten
Published: November 2003

The authors determined cerebral blood flow (CBF) with magnetic resonance imaging (MRI) of contrast agent bolus passage and compared the results with those obtained by O-15 labeled water (H215O) and positron emission tomography (PET). Six pigs were examined by MRI and PET under normo- and hypercapnic conditions. After dose normalization and introduction of an empirical constant ΦGd, absolute regional CBF was calculated from MRI. The spatial resolution and the signal-to-noise ratio of CBF measurements by MRI were better than by the H215O-PET protocol. Magnetic resonance imaging cerebral blood volume (CBV) estimates obtained using this normalization constant correlated well with values obtained by O-15 labeled carbonmonooxide (C15O) PET. However, PET CBV values were approximately 2.5 times larger than absolute MRI CBV values, supporting the hypothesized sensitivity of MRI to small vessels.
Noninvasive Measurement of Regional Cerebral Blood Flow by Near-Infrared Spectroscopy and Indocyanine Green
Authors: Kuebler Wolfgang M. , Sckell Axel , Habler Oliver , Kleen Martin , Kuhnle Gerhard E. H. , Welte Martin , Messmer Konrad , Goetz Alwin E.
Published: November 2003

Clinicians lack a practical method for measuring CBF rapidly, repeatedly, and noninvasively at the bedside. A new noninvasive technique for estimation of cerebral hemodynamics by use of near-infrared spectroscopy (NIRS) and an intravenously infused tracer dye is proposed. Kinetics of the infrared tracer indocyanine green were monitored on the intact skull in pigs. According to an algorithm derived from fluorescein flowmetry, a relative blood flow index (BFI) was calculated. Data obtained were compared with cerebral and galeal blood flow values assessed by radioactive microspheres under baseline conditions and during hemorrhagic shock and resuscitation. Blood flow index correlated significantly ( rs = 0.814, P < 0.001) with cortical blood flow but not with galeal blood flow ( rs = 0.258). However, limits of agreement between BFI and CBF are rather wide (± 38.2 ± 6.4 mL 100 g−1 min−1) and require further studies. Data presented demonstrate that detection of tracer kinetics in the cerebrovasculature by NIRS may serve as valuable tool for the noninvasive estimation of regional CBF. Indocyanine green dilution curves monitored noninvasively on the intact skull by NIRS reflect dye passage through the cerebral, not extracerebral, circulation.
Long-Term Neuroprotection by Benzodiazepine: Full Versus Partial Agonists after Transient Cerebral Ischemia in the Gerbil
Authors: Schwartz-Bloom Rochelle D. , McDonough Kelly J. , Chase Peter J. , Chadwick Laura E. , Inglefield Jon R. , Levin Edward D.
Published: November 2003

The ability of diazepam, a benzodiazepine full agonist, and imidazenil, a benzodiazepine partial agonist, to protect hippocampal area CA1 neurons from death for at least 35 days after cerebral ischemia was investigated. Diazepam (10 mg/kg) administered to gerbils 30 and 90 minutes after forebrain ischemia produced significant protection of hippocampal area CA1 pyramidal neurons 7 days later. In gerbils surviving for 35 days, diazepam produced the same degree of neuroprotection (70% ± 30%) in the hippocampus compared with 7 days after ischemia. The therapeutic window for diazepam was short; there was no significant neuroprotection when the administration of diazepam was delayed to 4 hours after ischemia. The neuroprotective dose of diazepam also produced hypothermia (~32°C) for several hours after injection. To assess the role of hypothermia in neuroprotection by diazepam, hypothermia depth and duration was simulated using a cold-water spray in separate gerbils. Seven days after ischemia, neuroprotection by hypothermia was similar to that produced by diazepam. However, 35 days after ischemia, there was no significant protection by hypothermia, suggesting that hypothermia does not play a significant role in long-term diazepam neuroprotection. Imidazenil (3 mg/kg), which produced only minimal hypothermia, protected area CA1 of hippocampus to the same degree as that by diazepam 7 days after ischemia. At 35 days after ischemia, significant protection remained, but it was considerably reduced compared with 7 days. Like diazepam, the therapeutic window for imidazenil was short. Imidazenil neuroprotection was lost when the drug was administered as early as 2 hours after ischemia. The ability of ischemia to produce deficits in working memory and of benzodiazepines to prevent the deficits also was investigated. Gerbils trained on an eight-arm radial maze before ischemia demonstrated a significant increase in the number of working errors 1 month after ischemia. The ischemia-induced deficits in working memory were completely prevented by diazepam but not by imidazenil. There was a significant, but weak, negative correlation between the degree of CA1 pyramidal cell survival and the number of working errors in both the diazepam and imidazenil groups. Thus, if given early enough during reperfusion, both benzodiazepine full and partial agonists are neuroprotective for at least 35 days, but the lack of sedating side effects of imidazenil must be weighed against its reduced efficacy.
Quinolinic Acid is Increased in CSF and Associated with Mortality after Traumatic Brain Injury in Humans
Authors: Sinz Elizabeth H. , Kochanek Patrick M. , Heyes Melvyn P. , Wisniewski Stephen R. , Bell Michael J. , Clark Robert S. B. , DeKosky Steven T. , Blight Andrew R. , Marion Donald W.
Published: November 2003

We tested the hypothesis that quinolinic acid, a tryptophan-derived N-methyl-d-aspartate agonist produced by macrophages and microglia, would be increased in CSF after severe traumatic brain injury (TBI) in humans, and that this increase would be associated with outcome. We also sought to determine whether therapeutic hypothermia reduced CSF quinolinic acid after injury. Samples of CSF ( n = 230) were collected from ventricular catheters in 39 patients (16 to 73 years old) during the first week after TBI, (Glasgow Coma Scale [GCS] < 8). As part of an ongoing study, patients were randomized within 6 hours after injury to either hypothermia (32°C) or normothermia (37°C) treatments for 24 hours. Oth-erwise, patients received standard neurointensive care. Quinolinic acid was measured by mass spectrometry. Univariate and multivariate analyses were used to compare CSF quinolinic acid concentrations with age, gender, GCS, time after injury, mortality, and treatment (hypothermia versus normothermia). Quinolinic acid concentration in CSF increased maximally to 463 ± 128 nmol/L (mean ± SEM) at 72 to 83 hours after TBI. Normal values for quinolinic acid concentration in CSF are less than 50 nmol/L. Quinolinic acid concentration was increased 5-to 50-fold in many patients. There was a powerful association between time after TBI and increased quinolinic acid ( P < 0.00001), and quinolinic acid was higher in patients who died than in survivors ( P = 0.003). Age, gender, GCS, and treatment (32°C versus 37°C) did not correlate with CSF quinolinic acid. These data reveal a large increase in quinolinic acid concentration in CSF after TBI in humans and raise the possibility that this macrophage-derived excitotoxin may contribute to secondary damage.
Cerebral Circulation in Sleep: Vasodilatory Response to Cerebral Hypotension
Authors: Grant Daniel A. , Franzini Carlo , Wild Jennene , Walker Adrian M.
Published: November 2003

Little is known of the factors that regulate CBF in sleep. We therefore studied 10 lambs to assess the vasodilatory processes that underlie cerebral autoregulation during sleep. Lambs, instrumented to measure CBF (flow probe on the superior sagittal sinus), sleep state, and cerebral perfusion pressure (CPP), were rapidly made hypotensive by inflating a cuff around the brachiocephalic artery to reduce CPP to 30 mm Hg in each state. During control periods, cerebral vascular resistance (CVR in mm Hg/mL/min) was lower in active sleep (2.8±0.3, mean±SD, P ≤ 0.001) than in wakefulness (3.9±0.6) and quiet sleep (4.3±0.6). The CVR decreased promptly in each state as CPP was lowered. The time (seconds) required for maximal cerebral vasodilation to occur was longer in active sleep (35±11) than in quiet sleep (20±6, P ≤ 0.001) and wakefulness (27±11, P ≤ 0.05). The CVR decreased less in active sleep (0.6±0.3, P ≤ 0.001) than in quiet sleep (1.5±0.3), although the changes in CPP induced with brachiocephalic occlusion were equal in each state. In conclusion, our studies provide the first evidence that the vasoactive mechanisms that underlie autoregulation of the cerebral circulation function during sleep. Moreover, our data reveal that the speed and the magnitude of the vasodilatory reserves available for autoregulation are significantly less in active sleep than in quiet sleep.
Reoxygenation Increases the Release of Reactive Oxygen Intermediates in Murine Microglia
Authors: Spranger Matthias , Kiprianova Irina , Krempien Sebastian , Schwab Stefan
Published: November 2003

Respiratory burst activity of murine microglial cells was investigated in vitro under normoxic and hypoxic conditions with a chemoluminometric assay. Hypoxia for 24 hours reduced the release of extracellular reactive oxygen intermediates (ROIs), whereas reoxygenation increased the chemoluminescence more than sevenfold. Blockade of potassium channels inhibited the increase of oxidative burst after reoxygenation, indicating that potassium ions, which were increased in the supernatant of hypoxic microglial cells, were involved in this activation process. Also, blockade of voltage-gated calcium channels with nifedipine attenuated the increased release of ROIs. With fura-2 analysis, it was shown that the activation of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase by potassium ions was mediated by calcium influx via voltage-gated calcium channels. Thus, influx of calcium ions through voltage-gated channels activates the NADPH oxidase in microglial cells during reoxygenation. By the increased production of ROIs, microglial cells may add to the reperfusion injury after ischemia in vivo.
Brain Networks Affected by Synchronized Sleep Visualized by Positron Emission Tomography
Authors: Andersson Jesper L. R. , Onoe Hirotaka , Hetta Jerker , Lidström Karin , Valind Sven , Lilja Anders , Sundin Anders , Fasth Karl-Johan , Westerberg Göran , Broman Jan-Erik , Watanabe Yasuyoshi , Långström Bengt
Published: November 2003

Nineteen lightly sleep-deprived healthy volunteers were examined with H215O and positron emission tomography (PET). Scanning was performed during wakefulness and after the subjects had fallen asleep. Sleep stage was graded retrospectively from electroencephalogram (EEG) recordings, and scans were divided into two groups: wakefulness or synchronized sleep. Global flow was quantified, revealing no difference between sleep and wakefulness. A pixel-by-pixel—blocked one-way analysis of variance (ANOVA) was performed after correcting for differences in anatomy and global flow. The sum of squares of the z-score distribution showed a highly significant ( P < 0.00001) omnibus difference between sleep and wakefulness. The z-score images indicated decreased flow in the thalamus and the frontal and parietal association cortices and increased flow in the cerebellum during sleep. A principal component (PC) analysis was performed on data after correction for global flow and block effects, and a multivariate analysis of variance (MANOVA) on all PC scores revealed significant ( P = 0.00004) differences between sleep and wakefulness. Principal component's 2 and 5 correlated to sleep and revealed distinct networks consisting of PC 2, cerebellum and frontal and parietal association cortices, and PC 5, thalamus.
Noninvasive Quantification of the Cerebral Metabolic Rate for Glucose Using Positron Emission Tomography, 18F-Fluoro-2-Deoxyglucose, the Patlak Method, and an Image-Derived Input Function
Authors: Chen Kewei , Bandy Daniel , Reiman Eric , Huang Sung-Cheng , Lawson Michael , Feng Dagan , Yun Lang-sheng , Palant Anita
Published: November 2003

The authors developed and tested a method for the noninvasive quantification of the cerebral metabolic rate for glucose (CMRglc) using positron emission tomography (PET), 18F-fluoro-2-deoxyglucose, the Patlak method, and an image-derived input function. Dynamic PET data acquired 12 to 48 seconds after rapid tracer injection were summed to identify carotid artery regions of interest (ROIs). The input function then was generated from the carotid artery ROIs. To correct spillover, the early summed image was superimposed over the last PET frame, a tissue ROI was drawn around the carotid arteries, and a tissue time activity curve (TAC) was generated. Three venous samples were drawn from the tracer injection site at a later time and used for the spillover and partial volume correction by non-negative least squares method. Twenty-six patient data sets were studied. It was found that the image-derived input function was comparable in shape and magnitude to the one obtained by arterial blood sampling. Moreover, no significant difference was found between CMRglc estimated by the Patlak method using either the arterial blood sampling data or the image-derived input function.
Temporal Patterns of Evoked Cerebral Blood Flow during Reading
Authors: Tiecks Frank P. , Haberl Roman L. , Newell David W.
Published: November 2003

This study describes the dynamics of flow activation by reading and investigates the potential use of repeated flow velocity measurements for the lateralization of speech. Using simultaneous transcranial Doppler recordings from both middle cerebral arteries and averaging techniques in 25 healthy volunteers, we describe the changes in blood flow velocity caused by repetitive reading tasks of variable duration in comparison with a resting state. Reading aloud evoked a characteristic temporal flow pattern in both hemispheres, consisting of three relative maxima in flow velocity during and after activation. Flow velocities lower than baseline were common during longer lasting activation. The amplitudes of two of the observed peaks decreased depending on the duration of the task. Reading silently produced a markedly different temporal pattern of activation than reading aloud. There were individually reproducible significant side to side differences. Right-handed persons ( n = 15) almost without exception showed a significantly higher increase in flow velocity on the left hemisphere (e.g., reading silently 8.7% versus 5.3%; P < 0.0001). Three out of ten left-handed individuals, however, exhibited no significant side to side difference or exhibited lateralization to the right during one or more of the tasks. These findings suggest that reading induces task-specific temporal patterns of regional neuronal activity, which show habituation with longer duration of activation. Additionally, the observed side to side differences could be useful to predict language dominance.
Characterization of Neuroprotection from Excitotoxicity by Moderate and Profound Hypothermia in Cultured Cortical Neurons Unmasks a Temperature-Insensitive Component of Glutamate Neurotoxicity
Authors: Tymianski Michael , Sattler Rita , Zabramski Joseph M. , Spetzler Robert F.
Published: November 2003

Although profound hypothermia has been used for decades to protect the human brain from hypoxic or ischemic insults, little is known about the underlying mechanism. We therefore report the first characterization of the effects of moderate (30°C) and profound hypothermia (12° to 20°C) on excitotoxicity in cultured cortical neurons exposed to excitatory amino acids (EAA; glutamate, N-methyl-D-aspartate [NMDA], AMPA, or kainate) at different temperatures (12° to 37°C). Cooling neurons to 30°C and 20°C was neuroprotective, but cooling to 12°C was toxic. The extent of protection depended on the temperature, the EAA receptor agonist employed, and the duration of the EAA challenge. Neurons challenged briefly (5 minutes) with all EAA were protected, as were neurons challenged for 60 minutes with NMDA, AMPA, or kainate. The protective effects of hypothermia (20° and 30°C) persisted after rewarming to 37°C, but rewarming from 12°C was deleterious. Surprisingly, however, prolonged (60 minutes) exposures to glutamate unmasked a temperature-insensitive component of glutamate neurotoxicity that was not seen with the other, synthetic EAA; this component was still mediated via NMDA receptors, not by ionotropic or metabotropic non-NMDA receptors. The temperature-insensitivity of glutamate toxicity was not explained by effects of hypothermia on EAA-evoked [Ca2+]i increases measured using high- and low-affinity Ca2+ indicators, nor by effects on mitochondrial production of reactive oxygen species. This first characterization of excitotoxicity at profoundly hypothermic temperatures reveals a previously unnoticed feature of glutamate neurotoxicity unseen with the other EAA, and also suggests that hypothermia protects the brain at the level of neurons by blocking, rather than slowing, excitotoxicity.
Changes of Cerebral Blood Flow during Short-Term Exposure to Normobaric Hypoxia
Authors: Buck Alfred , Schirlo Christian , Jasinsky Valeska , Weber Bruno , Burger Cyrill , von Schulthess Gustav K. , Koller Erwin A. , Pavlicek Vojtech
Published: November 2003

Decreased arterial partial oxygen pressure (PaO2) below a certain level presents a strong stimulus for increasing cerebral blood flow. Although several field studies examined the time course of global cerebral blood flow (gCBF) changes during hypoxia at high altitude, little was known about the regional differences in the flow pattern. Positron emission tomography (PET) with [15O]H2O was used on eight healthy volunteers to assess regional cerebral blood flow (rCBF) during short-term exposure to hypoxia corresponding to simulated altitudes of 3,000 and 4,500 m. Scans at the simulated altitudes were preceded and followed by baseline scans at the altitude of Zurich (450 m, baseline-1 and baseline-2). Each altitude stage lasted 20 minutes. From baseline to 4,500 m, gCBF increased from 34.4 ± 5.9 to 41.6 ± 9.0 mL · minute−1 · 100 g−1 (mean ± SD), whereas no significant change was noted at 3,000 m. During baseline-2 the flow values returned to those of baseline-1. Statistical parametric mapping identified the hypothalamus as the only region with excessively increased blood flow at 4,500 m (+32.8% ± 21.9% relative to baseline-1). The corresponding value for the thalamus, the structure with the second largest increase, was 19.2% ± 16.3%. Compared with the rest of the brain, an excessive increase of blood flow during acute exposure to hypoxia is found in the hypothalamus. The functional implications are at present unclear. Further studies of this finding should elucidate its meaning and especially focus on a potential association with the symptoms of acute mountain sickness.
Validation of the Three-Dimensional Acquisition Mode in Positron Emission Tomography for the Quantitation of [18F]Fluoro-DOPA Uptake in the Human Striata
Authors: Trébossen Régine , Bendriem Bernard , Ribeiro Maria-Joao , Fontaine Anne , Frouin Vincent , Remy Philippe
Published: November 2003

Three-dimensional (3D) positron emission tomography (PET) is attractive for [18F]fluoro-DOPA studies, since the sensitivity improvement is maximal for radioactive sources located in central planes, which is usually the case for the human striata. However, the image quantitation in that mode must be assessed because of the nearly threefold increase in scattered coincidences. We report the results of [18F]fluoro-DOPA studies performed on six normal volunteers. Each one was scanned in the 3D and two-dimensional(2D) modes on the same tomograph. The quantitation in the 3D and 2D modes was compared for a Patlak graphical analysis with the occipital counts as the input function (Ki) and a striatooccipital ratio analysis. We find that, in 3D PET, a scatter correction is required to preserve the same quantitation as in 2D PET. When the 3D data sets are corrected for scatter, the quantitation of the [18F]fluoro-DOPA uptake, using the Patlak analysis, is similar in the 2D and 3D acquisition modes. Conversely, analysis of the striatooccipital ratio leads to higher values in 3D PET because of a better in-plane resolution. Finally, using the 3D mode, the dose injected to the subjects can be reduced by a factor greater than 1.5 without any loss in accuracy compared to the 2D mode.
Nitric Oxide Scavenging by Hemoglobin or Nitric Oxide Synthase Inhibition by N-Nitro-L-Arginine Induces Cortical Spreading Ischemia When K+ Is Increased in the Subarachnoid Space
Authors: Dreier Jens P. , Körner Katrin , Ebert Nathalie , Görner Astrid , Rubin Inger , Back Tobias , Lindauer Ute , Wolf Tilo , Villringer Arno , Einhäupl Karl Max , Lauritzen Martin , Dirnagl Ulrich
Published: November 2003

We investigated the combined effect of increased brain topical K+ concentration and reduction of the nitric oxide(NO.) level caused by nitric oxide scavenging or nitric oxide synthase (NOS) inhibition on regional cerebral blood flow and subarachnoid direct current (DC) potential. Using thiopental-anesthetized male Wistar rats with a closed cranial window preparation, brain topical superfusion of a combination of the NO. scavenger hemoglobin(Hb; 2 mmol/L) and increased K+ concentration in the artificial cerebrospinal fluid ([K+]ACSF) at 35 mmol/L led to sudden spontaneous transient ischemic events with a decrease of CBF to 14 ± 7% (n = 4) compared with the baseline (100%). The ischemic events lasted for 53 ± 17 minutes and were associated with a negative subarachnoid DC shift of −7.3 ± 0.6 mV of 49 ± 12 minutes' duration. The combination of the NOS inhibitor N-nitro-L-arginine(L-NA, 1 mmol/L) with [K+]ACSF at 35 mmol/L caused similar spontaneous transient ischemic events in 13 rats. When cortical spreading depression was induced by KCl at a 5-mm distance, a typical cortical spreading hyperemia (CSH) and negative DC shift were measured at the closed cranial window during brain topical superfusion with either physiologic artificial CSF (n = 5), or artificial CSF containing increased [K+]ACSF at 20 mmol/L (n = 4), [K+]ACSF at 3 mmol/L combined with L-NA (n = 10), [K+]ACSF at 10 mmol/L combined with L-NA (five of six animals) or [K+]ACSF at 3 mmol/L combined with Hb (three of four animals). Cortical spreading depression induced long-lasting transient ischemia instead of CSH, when brain was superfused with either[K+]ACSF at 20 mmol/L combined with Hb (CBF decrease to 20± 20% duration 25 ± 21 minutes, n = 4), or [K+]ACSF at 20 mmol/L combined with L-NA (n = 19). Transient ischemia induced by NOS inhibition and [K+]ACSF at 20 mmol/L propagated at a speed of 3.4 ± 0.6 mm/min, indicating cortical spreading ischemia (CSI). Although CSH did not change oxygen free radical production, as measured on-line by in vivo lucigenin-enhanced chemiluminescence, CSI resulted in the typical radical production pattern of ischemia and reperfusion suggestive of brain damage (n = 4). Nimodipine (2 μg/kg body weight/min intravenously) transformed CSI back to CSH (n = 4). Vehicle had no effect on CSI(n = 4). Our data suggest that the combination of decreased NO. levels and increased subarachnoid K+ levels induces spreading depression with acute ischemic CBF response. Thus, a disturbed coupling of metabolism and CBF can cause ischemia. We speculate that CSI may be related to delayed ischemic deficits after subarachnoid hemorrhage, a clinical condition in which the release of Hb and K+ from erythrocytes creates a microenvironment similar to the one investigated here.
Postischemic Enhancements of N-Methyl-D-Aspartic Acid (NMDA) and Non-NMDA Receptor-Mediated Responses in Hippocampal CA1 Pyramidal Neurons
Authors: Mitani Akira , Namba Shigeru , Ikemune Keizou , Yanase Hisato , Arai Tatsuru , Kataoka Kiyoshi
Published: November 2003

Glutamate receptor-mediated responses were investigated by using a whole-cell recording and an intracellular calcium ion ([Ca2+]i) imaging in gerbil postischemic hippocampal slices prepared at 1, 3, 6, 9, 12, and 24 hours after 5-minute ischemia. Bath application of N-methyl-D-aspartic acid (NMDA), α-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA), and kainate showed that NMDA-, AMPA- and kainate-induced currents were enhanced in postischemic CA1 pyramidal neurons at 1 to 12 hours after 5-minute ischemia. NMDA and non-NMDA receptor-mediated excitatory postsynaptic currents (EPSC) were examined in postischemic CA1 pyramidal neurons at 3 hours after 5-minute ischemia to confirm whether synaptic responses are enhanced in the postischemic CA1 pyramidal neurons. The amplitudes of NMDA- and non-NMDA-receptor-mediated EPSC were enhanced in the postischemic CA1 pyramidal neurons. NMDA-, AMPA-, and kainate-induced [Ca2+]i elevations were also examined to determine whether the enhancement of currents is accompanied by the enhancement of [Ca2+]i elevation. The enhancements of NMDA-, AMPA-, and kainate-induced [Ca2+]i elevations were shown in the postischemic CA1. These results indicate that NMDA and non-NMDA receptor-mediated responses are persistently enhanced in the CA1 pyramidal neurons 1 to 12 hours after transient ischemia, and suggest that the enhancement of glutamate receptor-mediated responses may act as one of crucial factors in the pathologic mechanism responsible for leading postischemic CA1 pyramidal neurons to irreversible neuronal injury.
Delayed Treatment with Aminoguanidine Decreases Focal Cerebral Ischemic Damage and Enhances Neurologic Recovery in Rats
Authors: Nagayama Masao , Zhang Fangyi , Iadecola Costantino
Published: November 2003

Delayed treatment with aminoguanidine (AG), a relatively selective inhibitor of inducible nitric oxide synthase, ameliorates brain damage produced by occlusion of the rat's middle cerebral artery (MCA). We investigated whether the protection exerted by AG is dose-dependent and whether it is associated with improved neurologic outcome. We also studied the effect of the timing of administration of AG relative to the induction of cerebral ischemia. Halothane-anesthetized spontaneously hypertensive rats underwent permanent MCA occlusion distal to the lenticulostriate branches. Neurologic deficits were assessed daily by the postural reflex test and beam balance test. Infarct volume was determined in thionin- stained sections 96 hours after ischemia and values corrected for swelling. Treatment with AG (intraperitoneally, twice daily), starting 24 hours after MCA occlusion, decreased neocortical infarct volume in comparison to vehicle-treated rats. After correction for swelling, the decrease was 8 ± 12% at 50 mg/kg (n = 8; P > .05; analysis of variance), 25 ± 13% at 100 mg/kg (n = 7; P < .05), 30 ± 16% at 200 mg/kg (n = 7; P < .05) and 32 ± 9% at 400 mg/kg (n = 5; P < .05). Twenty-four hours after induction of ischemia neurologic deficits scores did not differ between treated and untreated rats ( P > .05). However, from 48 to 96 hours after ischemia, neurologic deficits improved significantly in rats treated with AG (100 to 400 mg/kg) compared to rats in which vehicle was administered ( P < .05). The decrease in neocortical infarct volume was greatest when AG (100 mg/kg; twice daily) was administered 12 (26 ± 17%; n = 9) or 24 hours (25 ± 13; n = 7) after MCA occlusion. The findings show that AG decreases ischemic brain damage dose-dependently and improves neurologic recovery. Delayed treatment with AG may be a therapeutic strategy to selectively target the evolution of ischemic damage that occurs in the post-ischemic period.
Spatial Stability of Extracellular Potassium Ion and Blood Flow Distribution in Rat Cerebral Cortex after Permanent Middle Cerebral Artery Occlusion
Authors: Sick Thomas J. , Feng Zi-Cai , Rosenthal Myron
Published: November 2003

Extracellular potassium ion activity ([K+]o) increases precipitously during brain ischemia when blood flow falls below threshold values less than approximately 15 mL/100 g/min. This flow threshold for increase of [K+]o occurs also in focal ischemia producing gradient from ischemic core to adjacent normally perfused brain. In this study we investigated the spatial and temporal stability of extracellular potassium ion and blood flow gradients after permanent middle cerebral artery occlusion (MCAO) in rats. [K+]o and regional CBF were measured, respectively, with K+-sensitive and polarographic hydrogen-sensitive microelectrodes at different cortical locations in the middle cerebral artery distribution region. Spatial assessment of [K+]o and regional CBF was conducted at 30, 90, and 180 minutes after MCAO. [K+]o in the more lateral cortex (core) increased from near 3 mmol/L before MCAO to greater than 50 mmol/L and was associated with flow values less than 25% of pre-ischemic levels. Measurements medial to the core (penumbra) indicated progressively decreasing levels of [K+]o and improvement of CBF. There was a tendency for [K+]o in penumbral zones to decrease toward normal levels with time, but there was little dissipation of [K+]o in core regions. In contrast, the spatial CBF profile remained remarkably constant for the entire recording period. Thus, unlike infarction which has been reported to expand with time after focal ischemia, the spatial [K+]o disturbance tends to contract primarily due to decreasing [K+]o with time in the penumbra. Thus, steady state levels of [K+]o after focal ischemia may not be a valuable predictor of cell viability.
Validation of a Less-Invasive Method for Measurement of Serotonin Synthesis Rate with α-[11C]Methyl-Tryptophan
Authors: Nishizawa Sadahiko , Leyton Marco , Okazawa Hidehiko , Benkelfat Chawki , Mzengeza Shadreck , Diksic Mirko
Published: November 2003

We tested in normal human subjects a less invasive method to obtain plasma input function required in the calculation of the brain serotonin synthesis rate measured with positron emission tomography (PET) and α-[11C]methyl-tryptophan (α-MTrp). The synthesis rates derived with the arterial input function were compared to those derived from venous plasma and venous sinus time-radioactivity curves obtained from dynamic PET images. Dynamic PET images were obtained for the lengths up to 90 minutes after an injection of α-MTrp (400 to 800 MBq). Input functions were generated from both artery and vein in three subjects, and from artery only in two subjects. Net unidirectional uptake constants of α-MTrp (K*; mL/g/min) were calculated in several brain regions graphically using data between 20 and 60 minutes after injection with different input functions. In the five subjects with arterial sampling, we tested two methods for correcting the input functions from the venous samples: (1) normalization to the mean exposure time at 20 minutes from arterial curve; and (2) the use of the venous sinus curve for the first 20 minutes. Venous curves coincided with the arterial ones after about 20 minutes. When the venous curves were used, there was an underestimation of the area under the curves up to 20 minutes, resulting in a 5% to 30% overestimation of K* values. Combined use of the sinus curve up to 20 minutes and venous curve from 20 to 60 minutes as an input function resulted in the K* (mL/g/min) values larger by 7.1 ± 3.8% than the K* values estimated with the arterial input function. Normalization of the venous curve to the exposure time at 20 minutes obtained from the arterial plasma curve resulted in a bias in the K* of about −0.34 ± 3.32%. The bias from the K* values was propagated to the serotonin synthesis rates. The use of a combination of the venous blood samples and venous sinus as the input function resulted in an acceptable bias in the serotonin synthesis rates from the tissue time-radioactivity curves generated by PET.
Cerebral Blood Flow and Cerebrovascular Reserve Capacity: Estimation by Dynamic Magnetic Resonance Imaging
Authors: Schreiber Wolfgang G. , Gückel Friedemann , Stritzke Peter , Schmiedek Peter , Schwartz Andreas , Brix Gunnar
Published: November 2003

We have developed a new method for estimation of regional CBF (rCBF) and cerebrovascular reserve capacity on a pixel-by-pixel basis by means of dynamic magnetic resonance imaging (MRI). Thirteen healthy volunteers, 8 patients with occlusion and/or high grade stenosis of the internal carotid artery (ICA), and 2 patients with acute stroke underwent dynamic susceptibility-weighted contrast enhanced MRI. Using principles of indicator dilution theory and deconvolution analysis, maps of rCBF, regional cerebral blood volume, and of the mean transit time (MTT) were calculated. In patients with ICA occlusion/stenosis, cerebrovascular reserve capacity was assessed by the rCBF increase after acetazolamide stimulation. Mean gray and white matter rCBF values in normals were 67.1 and 23.7 mL · 100 g−1 · min−1, respectively. Before acetazolamide stimulation, six of eight patients with ICA occlusions showed decreased rCBF values; and in seven patients increased MTT values were observed in tissue ipsilateral to the occlusion. After acetazolamide stimulation, decreased cerebrovascular reserve capacity was observed in five of eight patients with ICA occlusion. In acute stroke, rCBF in the central core of ischemia was less than 8 mL · 100 g−1 · min−1. In peri-infarct tissue, rCBF and MTT were higher than in unaffected tissue but rCBF was normal. Dynamic MRI provides important clinical information on the hemodynamic state of brain tissue in patients with occlusive cerebrovascular disease or acute stroke.
Matrix Metalloproteinases in Cerebrovascular Disease
Authors: Mun-Bryce Sheila , Rosenberg Gary A.
Published: November 2003

Cerebral ischemia and intracerebral hemorrhage cause extensive damage to neurons, disrupt the extracellular matrix, and increase capillary permeability. Multiple substrates participate in the cellular damage, including free radicals and proteases. Matrix metalloproteinases and serine proteases are two classes of proteases that are normally present in brain in latent forms, but once activated, contribute to the injury process. These enzymes have a unique role in the remodeling of the extracellular matrix and in the modulation of the capillary permeability. Intracerebral injection of the matrix metalloproteinase, type IV collagenase, attacks the basal lamina around the capillary and opens the blood—brain barrier, Extracellular matrix-degrading proteases are induced by immediate early genes and cytokines, and regulated by growth factors. Activity of the matrix metalloproteinases is tightly controlled by activation mechanisms and tissue inhibitors of metalloproteinases. During ischemia and hemorrhage, multiple matrix metalloproteinases and serine proteases are produced along with their inhibitors. These proteolytic enzymes are involved in the delayed injury that accompanies the neuroinflammatory response. Synthetic inhibitors to metalloproteinases reduce proteolytic tissue damage, and may limit secondary neuroinflammation.
Estradiol Protects against Ischemic Injury
Authors: Dubal Dena B. , Kashon Michael L. , Pettigrew L. Creed , Ren Jing M. , Finklestein Seth P. , Rau Shane W. , Wise Phyllis M.
Published: November 2003

Clinical studies demonstrate that estrogen replacement therapy in postmenopausal women may enhance cognitive function and reduce neurodegeneration associated with Alzheimer's disease and stroke, This study assesses whether physiologic levels of estradiol prevent brain injury in an in vivo model of permanent focal ischemia. Sprague-Dawley rats were ovariectomized; they then were implanted, immediately or at the onset of ischemia, with capsules that produced physiologically low or physiologically high 17β-estradiol levels in serum (10 or 60 pg/mL, respectively), One week after ovariectomy, ischemia was induced. Estradiol pretreatment significantly reduced overall infarct volume compared with oil-pretreated controls (mean ± SD: oil = 241 ± 88; low = 139 ± 91; high = 132 ±88 mm3); this protective effect was regionally specific to the cortex, since no protection was observed in the striatum. Baseline and ischemic regional CBF did not differ between oil and estradiol pretreated rats, as measured by laser Doppler flowmetry. Acute estradiol treatment did not protect against ischemic injury. Our finding that estradiol pretreatment reduces injury demonstrates that physiologic levels of estradiol can protect against neurodegeneration.
BCL-2 Overexpression Attenuates Cortical Cell Loss after Traumatic Brain Injury in Transgenic Mice
Authors: Raghupathi Ramesh , Fernandez Seamus C. , Murai Hisayuki , Trusko Stephen P. , Scott Richard W. , Nishioka Walter K. , McIntosh Tracy K.
Published: November 2003

The proto-oncogene, BCL-2, has been suggested to participate in cell survival during development of, and after injury to, the CNS. Transgenic (TG) mice overexpressing human Bcl-2 (n = 21) and their wild-type (WT) littermates (n = 18) were subjected to lateral controlled cortical impact brain injury. Lateral controlled cortical impact brain injury resulted in the formation of a contusion in the injured cortex at 2 days, which developed into a well-defined cavity by 7 days in both WT and TG mice. At 7 days after injury, brain-injured TG mice had a significantly reduced cortical lesion (volume = 1.99 mm3) compared with that of the injured WT mice (volume = 5.1 mm3, P <0.01). In contrast, overexpression of BCL-2 did not affect the extent of hippocampal cell death after lateral controlled cortical impact brain injury. Analysis of motor function revealed that both brain-injured WT and TG mice exhibited significant right-sided deficits at 2 and 7 days after injury ( P < 0.05 compared with the uninjured controls). Although composite neuroscores (sum of scores from forelimb and hind limb flexion, lateral pulsion, and inclined plane tests) were not different between WT and TG brain-injured mice, TG mice had a slightly but significantly reduced deficit in the inclined plane test ( P < 0.05 compared to the WT mice). These data suggest that the cell death regulatory gene, BCL-2, may play a protective role in the pathophysiology of traumatic brain injury.
Regulation of a Blood-Brain Barrier—Specific Enzyme Expressed by Cerebral Pericytes (Pericytic Aminopeptidase N/pAPN) under Cell Culture Conditions
Authors: Ramsauer Markus , Kunz Jörg , Krause Dorothee , Dermietzel Rolf
Published: November 2003

In this study we show that the aminopeptidase N of cerebral pericytes (pAPN) associated with the blood—brain barrier (BBB) is downregulated in pericytic cell cultures. This observation is in accordance with previous data describing comparable in vitro effects for BBB-specific enzymes of endothelial or pericytic origin, such as γ-glutamyl transpeptidase or alkaline phosphatase. By polymerase chain reaction and in situ hybridization we were able to determine that the downregulation of pAPN occurs at the posttranscriptional level. The mRNA of pAPN was found to be constitutively expressed even when the protein is no longer detectable. Culturing the pericytes in an endothelial cell—conditioned medium allowed pAPN to be reexpressed. However, the reexpression effect depended largely on the culturing conditions of the pericytes. Although purified pericytes deprived of endothelial cells did not reveal a reexpression effect, pericytes that were kept in contact with endothelial cells were able to acquire a pAPN-positive phenotype, indicating that endothelial cells constitute an essential requirement for the in vitro reexpression of pAPN. Astrocytes, however, were insufficient in exerting any reexpression effect.
Ischemic and Excitotoxic Brain Injury is Enhanced in Mice Lacking the p55 Tumor Necrosis Factor Receptor
Authors: Gary Devin S. , Bruce-Keller Annadora J. , Kindy Mark S. , Mattson Mark P.
Published: November 2003

Ischemic and excitotoxic insults to the brain induce rapid production of tumor necrosis factor-α (TNF), but the role of TNF in neuronal responses to brain injury are unclear. Two different TNF receptors (p55 and p75) are expressed in neurons and glia, To understand the role of TNF in brain injury, we generated mice that lack p55, p75, or both receptors, We report that neuronal damage after focal cerebral ischemia—reperfusion is significantly increased in mice lacking p55 receptors (85 ± 7 mm3 infarct volume; mean ± SD) compared with wild-type mice (70 ± 8 mm3) and mice lacking p75 receptors (72 ± 6 mm3). Moreover, mice lacking p55 receptors exhibited increased degeneration of CA3 hippocampal neurons after administration of the excitotoxin kainic acid compared with wild-type mice and mice lacking p75 receptors. When taken together with recent data showing that TNF can prevent apoptosis of cultured neurons exposed to oxidative and metabolic insults, our findings suggest that TNF plays a neuroprotective role after acute brain insults.
Astrocyte Regulation of Endothelial Tissue Plasminogen Activator in a Blood-Brain Barrier Model
Authors: Tran Nam D. , Schreiber Steven S. , Fisher Mark
Published: November 2003

Expression of tissue plasminogen activator (tPA) substantially determines endothelial-dependent fibrinolysis. We used a blood-brain barrier (BBB) model to analyze regulation of brain capillary endothelial tPA and its inhibitor, plasminogen activator inhibitor-1 (PAI-1). This model consists of coculture of murine astrocytes with bovine brain capillary endothelial cells grown as capillary-like structures (CS); after 1 week, astrocytes become extensively associated with CS, and the BBB-associated enzyme γ-glutamyl transpeptidase is present. We measured tPA and PAI-1 mRNA and tPA activity in this model. Reverse transcription-polymerase chain reaction (RT-PCR) studies showed similar tPA and PAI-1 mRNA levels after 1 day mono-culture (endothelial cells only) versus astrocyte-endothelial coculture preparations. After 7 days (i.e., when elements of the BBB are present), astrocyte-endothelial cocultures (compared with endothelial mono-cultures) showed a 50.7% ± 27.1% (mean ± SD) reduction in tPA mRNA ( P < 0.03) and a 183.3% ± 86.9% increase in PAI-1 mRNA expression ( P < 0.02). Moreover, 7-day cocultures demonstrated reduced tPA activity compared with mono-cultures (14.6 ± 2.9 IU/mL versus 30.2 ± 7.7 IU/mL, P < 0.01); 1-day cocultures and mono-cultures had similar tPA activity. These findings demonstrate that astrocytes regulate brain capillary endothelial expression of tPA when elements of the BBB phenotype are present in this model. These data suggest an important role for astrocytes in the regulation of brain capillary endothelial fibrinolysis.