bluuebunny/crossref_metadata_2025 huggingface dataset:
Changes in Pial Arteriolar Diameter and CSF Adenosine Concentrations during Hypoxia
Authors: Meno Joseph R. , Ngai Al C. , Winn H. Richard
Published: April 2011

We measured the changes in pial arteriolar diameter and CSF concentrations of adenosine, inosine, and hypoxanthine during hypoxia in the absence and presence of topically applied dipyridamole (10−6 M) and erythro-9-(2-hydroxy-3-nonyl)adenine (EHNA; 10−5 M). Closed cranial windows were implanted in halothane-anesthetized adult male Sprague–Dawley rats for the observation of the pial circulation and collection of CSF. The mean resting arteriolar diameter in mock CSF was 31.2 ± 5.9 μm. Topically applied dipyridamole and EHNA, in combination, caused a slight but significant ( p < 0.05) increase in resting arteriolar diameter (33.8 ± 4.3 μm). With mock CSF, moderate hypoxia caused a 22.1 ± 9.7% increase in pial vessel diameter. Topically applied dipyridamole and EHNA significantly ( p < 0.01) potentiated pial arteriolar vasodilation in response to hypoxia. Moreover, the potentiating effects of dipyridamole and EHNA during hypoxia were completely abolished by theophylline (0.20 μmol/g, i.p.; p < 0.05), an adenosine receptor antagonist. Resting concentrations of adenosine, inosine, and hypoxanthine in the subwindow CSF were 0.18 ± 0.09, 0.35 ± 0.21, and 0.62 ± 0.12 μ M, respectively. In the absence of dipyridamole and EHNA, these levels were not affected by sustained moderate hypoxia (Pao2 = 36 ± 6 mm Hg). However, in the presence of dipyridamole and EHNA, the concentration of adenosine in the CSF during hypoxia was significantly ( p < 0.05) increased. Our data indicate that dipyridamole and EHNA potentiate hypoxic vasodilation of pial arterioles while simultaneously increasing extracellular adenosine levels, thus supporting the hypothesis that adenosine is involved in the regulation of cerebral blood flow.
NMDA and Glycine Receptor mRNA Expression following Transient Global Ischemia in the Gerbil Brain
Authors: Gass Peter , Muelhardt Cornel , Sommer Clemens , Becker Cord-Michael , Kiessling Marika
Published: April 2011

Excitotoxic activation of postsynaptic N-methyl-d-aspartate (NMDA) receptors is thought to be a key event for the molecular pathogenesis of postischemic delayed neuronal death in CA1 hippocampus. To assess a possible interference of ischemia with NMDA receptor expression, transcription of the NMDA receptor 1 (NMDA-R1) gene was examined by in situ hybridization in the gerbil brain after 5 min of global ischemia and various recirculation intervals. In normal gerbil brain, NMDA-R1 was strongly expressed and equally abundant in CA1 and CA3 neurons. After ischemia, expression remained unchanged for 24 h, followed by a selective decline in mRNA levels in CA1 neurons, resulting in the complete disappearance of hybridization signals after 4 days. NMDA-R1 expression in forebrain neurons less vulnerable or resistant to ischemia including CA3 and dentate granule cells remained unchanged. Similar in situ data were obtained for the β subunit of the inhibitory glycine receptor (Gly-R). This subunit is also abundantly expressed in the pyramidal cell layer of the hippocampus, but not known to be involved in the mechanisms of postischemic excitotoxicity.
Intraischemic but Not Postischemic Brain Hypothermia Protects Chronically following Global Forebrain Ischemia in Rats
Authors: Dietrich W. Dalton , Busto Raul , Alonso Ofelia , Globus Mordecai Y.-T. , Ginsberg Myron D.
Published: April 2011

We investigated whether postischemic brain hypothermia (30°C) would permanently protect the hippocampus following global forebrain ischemia. Global ischemia was produced in anesthetized rats by bilateral carotid artery occlusion plus hypotension (50 mm Hg). In the postischemic hypothermic group, brain temperature was maintained at 37°C during the 10-min ischemic insult but reduced to 30°C starting 3 min into the recirculation period and maintained at 30°C for 3 h. In normothermic animals, intra- and postischemic brain temperature was maintained at 37°C. After recovery for 3 days, 7 days, or 2 months, the extent of CA1 hippocampal histologic injury was quantitated. At 3 days after ischemia, postischemic hypothermia significantly protected the hippocampal CA1 sector compared with normothermic animals. For example, within the medial, middle, and lateral CA1 subsectors, the numbers of normal neurons were increased 20-, 13-, and 9-fold by postischemic hypothermia (p < 0.01). At 7 days after the ischemic insult, however, the degree of postischemic hypothermic protection was significantly reduced. In this case, the numbers of normal neurons were increased an average of only threefold compared with normothermia. Ultrastructural analysis of 7-day postischemic hypothermic rats demonstrated CA1 pyramidal neurons showing variable degrees of injury surrounded by reactive astrocytes and microglial cells. At 2 months after the ischemic insult, no trend for protection was demonstrated. In contrast to postischemic hypothermia, significant protection was seen at 2 months following intraischemic hypothermia. These data indicate that intraischemic, but not postischemic, brain hypothermia provides chronic protection to the hippocampus after transient brain ischemia. The inability of postischemic hypothermia to protect chronically after 3 days could indicate that (a) postischemic hypothermia merely delays ischemic cell death and/or (b) the postischemic brain undergoes a secondary insult. In postischemic treatment protocols, chronic survival studies are required to determine accurately the ultimate histopathological outcome following global cerebral ischemia.
Endotoxin Decreases the Contractile Responses of the Porcine Basilar Artery to Vasoactive Substances
Authors: Ueno Masami , Lee Tony J.-F.
Published: April 2011

The effects of endotoxin (lipopolysaccharide; LPS) on the reactivity of isolated porcine basilar artery were examined using in vitro tissue bath techniques. The active muscle tone of the basilar arterial rings with or without endothelial cells induced by U46619 (1 μ M) reached a plateau in 15 min, which was stable for the first hour and gradually decreased during the next 5 h. This time-dependent decrease in tone was significantly potentiated in the presence of LPS (20 μg/ml). The potentiation by LPS was blocked by Nw-nitro-l-arginine (l-NNA; 60 μ M), methylene blue (10 μ M), and dexamethasone (1 μ M) but not by hemoglobin (1 μ M). The effect of l-NNA was readily reversed by l-arginine but not by d-arginine. Furthermore, the contractile responses of porcine basilar arterial rings with or without intact endothelium to U46619 and KCl were decreased following incubation with LPS (20 μg/ml) for 4 h. Similar hyporeactivity was observed in cold storage–denervated cerebral arteries incubated with LPS for 4 h. This decrease in contractile responses in LPS-treated rings was reversed by 60 μ M l-NNA and 1 μ M dexamethasone. These results indicate that LPS treatment renders the porcine basilar arteries hyporesponsive to vasoconstrictors. Since effects of LPS were not modified by the presence of endothelial cells and perivascular neurons, the alteration in cerebral arterial reactivity may be due in part to an enhanced formation of nitric oxide from l-arginine in the vascular smooth muscle cells.
Indomethacin Abolishes Cerebral Blood Flow Increase in Response to Acetazolamide-Induced Extracellular Acidosis: A Mechanism for its Effect on Hypercapnia?
Authors: Wang Qiong , Paulson Olaf B. , Lassen Niels A.
Published: April 2011

Indomethacin is known to attenuate quite markedly the increase in CBF during hypercapnia. Hypercapnia is, in all likelihood, mediated by the acid shift at the level of the smooth muscle cells of the cerebral arterioles. We therefore investigated the effect of indomethacin on the CBF increase caused by acetazolamide (Az), a drug that induces brain extracellular acidosis, which triggers its effect on CBF. We compared the results to the inhibitory effect of indomethacin on the CBF increase during hypercapnia. Indomethacin but not diclofenac, another potent cyclooxygenase inhibitor, was found to block almost completely the CBF increase caused by Az-induced extracellular acidosis or by CO2, but it did not influence the CBF increase produced by sodium nitroprusside or papaverine. The results suggest that indomethacin exerts its action on CO2 reactivity by a nonprostaglandin-mediated mechanism that directly interferes with the regulation of cerebrovascular tone mediated by extracellular pH.
Adenosine, a “Retaliatory” Metabolite, Promotes Anoxia Tolerance in Turtle Brain
Authors: Pérez-Pinzón Miguel A. , Lutz Peter L. , Sick Thomas J. , Rosenthal Myron
Published: April 2011

Contrary to what is found in most vertebrates, the brains of certain turtle species maintain ATP levels and ion homeostasis and survive prolonged anoxia. The hypothesis tested here is that the release of adenosine and its binding to A1 receptors are essential for this anoxic tolerance. Studies were conducted in the isolated turtle cerebellum, which did release adenosine to the extracellular space during anoxia. When adenosine receptor antagonists [theophylline, 8-cyclopentyltheophylline (CPT), or 8-cyclopentyl-1,3-dipropylxanthine (DPCPX)] were added to the superfusate under control conditions, they had no effect on extracellular potassium ion activity ([K+]o). During anoxia, however, these antagonists provoked maximal efflux of K+ (anoxic depolarization). Anoxic depolarization occurred earlier during anoxia with theophylline (a nonspecific adenosine receptor antagonist) than with CPT or DPCPX, which specifically block A1 receptors. Therefore, adenosine release and effects mediated by A1 receptors are essential to anoxia tolerance in turtle brain.
Changes of Mitochondrial DNA and Heat Shock Protein Gene Expressions in Gerbil Hippocampus after Transient Forebrain Ischemia
Authors: Abe K. , Kawagoe J. , Aoki M. , Kogure K.
Published: April 2011

Hippocampal CA1 neurons are the most vulnerable to transient cerebral ischemia. However, the mechanism has not been fully understood. The level of mRNA for cytochrome C oxidase (COX) subunit I (COX-I), which is encoded by mitochondrial (mt) DNA, progressively decreased in the hippocampal CA1 neurons of gerbils from 3 h of reperfusion after 3.5 min of transient forebrain ischemia and completely disappeared at 7 days. The activity of COX protein also showed an early decrease in CA1 cells and was followed by reduction of the level of COX-I DNA after 2 days. However, succinic dehydrogenase, an mt enzyme encoded by nuclear DNA, maintained normal activity until 1 day in the CA1 cells and significantly decreased at 7 days. The mRNA for mt heat shock protein (HSP) 60 began to increase at 3 h in the CA1 cells and was sustained until 1 day. The mRNAs for 72-kDa heat shock protein and 73-kDa heat shock cognate protein, which are located mainly in the cytoplasm, were induced together in the CA1 cells with a peak at 1–2 days. These results suggest that a disturbance of mt DNA expression occurred in the CA1 neurons at the early stage of reperfusion and was aggravated over the course of time. The disturbance could cause progressive failure of energy production of the cells that eventually results in neuronal cell death.
Prevention of Periinfarct Direct Current Shifts with Glutamate Antagonist NBQX following Occlusion of the Middle Cerebral Artery in the Rat
Authors: Mies Günter , Kohno Kanehisa , Hossmann Konstantin-Alexander
Published: April 2011

The effect of the glutamate (AMPA subtype) receptor antagonist NBQX on periinfarct direct current (DC) shifts and cortical ATP depletion volume was examined in rats subjected to 3 h of occlusion of the middle cerebral artery (MCA). MCA occlusion produced an immediate DC shift in the periphery of the ischemic territory. Vehicle-treated (untreated) animals developed one to five additional DC shifts (median, 2) during the 3-h occlusion time. NBQX treatment (2 × 30 mg/kg i.v. immediately after MCA occlusion and 1 h later) significantly reduced the number of DC deflections (median, 0; range, 0–2; p < 0.05) without changing blood flow in the border zone of the infarct (untreated, 50.6 ± 10.6%; NBQX-treated: 51.9 ± 7.7% of control; mean ± SD). NBQX treatment significantly decreased the cortical volume of ATP depletion (untreated, 75.3 ± 11.4 mm3; NBQX-treated, 47.9 ± 10.1 mm3; p < 0.05). Moreover, a significant linear relationship between the number of periinfarct DC shifts and the volume of cortical ATP depletion was obtained ( y = 38.3 + 9.4x; r = 0.866; p < 0.001). The reduction of brain infarct volume by NBQX treatment is explained by the suppression of DC shifts and the decrease of metabolic workload in hemodynamically compromised cortex.
Calcium Accumulation following Middle Cerebral Artery Occlusion in Stroke-Prone Spontaneously Hypertensive Rats
Authors: Shirotani Toshiki , Shima Katsuji , Iwata Miwako , Kita Hideyuki , Chigasaki Hiroo
Published: April 2011

Delayed neuronal damage in the ischemic region of the rat brain following middle cerebral artery (MCA) occlusion in stroke-prone spontaneously hypertensive rats was studied. The distribution of neuronal damage was determined by 45Ca autoradiography. Accumulation of 45Ca was observed in the corpus callosum and ipsilateral cerebral cortex immediately following MCA occlusion. After 3 days of occlusion, 45Ca had accumulated in the ipsilateral pyramidal tract, the ventral posterior nucleus of the thalamus, and the lateral portion of the striatum. Significant accumulation of 45Ca was observed in the same areas after 7 and 14 days of occlusion. Next the effect of MK-801 on accumulation of 45Ca after MCA occlusion was examined using the same technique. MK-801 (0.5–10 mg/kg i.v.) or saline was administered 15 min before MCA occlusion, and volumes of accumulation of 45Ca were calculated 1 week after ischemic insults. MK-801 significantly reduced 45Ca uptake in the cortex, striatum, and thalamus. Furthermore, there was a strong statistical correlation between the volume of accumulation of 45Ca in the cortex and that in the thalamus ( r = 0.8974; p < 0.001; n = 25). We speculate that delayed neuronal damage in the corpus callosum, ipsilateral pyramidal tract, and thalamus may be caused by secondary neuronal degeneration. However, neuronal damage in the striatum, a segment not supplied by the MCA, may be related to excessive release of glutamate.
Cytokine-Mediated Activation of Cultured CNS Microvessels: A System for Examining Antigenic Modulation of CNS Endothelial Cells, and Evidence for Long-Term Expression of the Adhesion Protein E-Selectin
Authors: Dore-Duffy Paula , Washington Ruth A. , Balabanov Roumen
Published: April 2011

Much of what is known of endothelial responses to cytokines has been derived from in vitro studies using cultured human umbilical vein endothelial cells (EC). Less is known of CNS EC responses and whether intact endothelium responds similarly to cultured cells. We have used techniques by which rat CNS microvessels can be isolated, then cultured in vitro, to study the response of intact endothelium to activation with cytokines. These microvessels are composed of viable EC and perivascular cells, predominantly pericytes. Expression of EC activation antigens in multicellular systems such as cultured microvessels can be assessed quantitatively using immunofluorescence laser cytometry. Interferon gamma increased immunologically reactive major histocompatibility complex class II antigens (<300 to 2,398 ± 225 average fluorescence intensity), while tumor necrosis factor alpha induced an increase in vascular cell adhesion molecule-1 (2,167 ± 171) and E-selectin (1,628 ± 315). CNS EC appeared to respond similarly to cultured EC with the exception that E-selectin expression was not transiently expressed but was maintained by microvessel EC for 24 and 48 h. Cultured CNS microvessels provide a good system for studying EC activation.
Bradykinin Selectively Opens Blood-Tumor Barrier in Experimental Brain Tumors
Authors: Inamura Takanori , Black Keith L.
Published: April 2011

Bradykinin, infused in low doses (10 μg/kg/min) through the carotid artery ipsilateral to RG2 glioma in rats, significantly increased the permeability in tumor capillaries to six different tracers of varying molecular weights compared with intracarotid infusion of saline alone. Permeability in normal brain capillaries was not significantly increased by intracarotid bradykinin infusion. Tracers used to examined permeability included radiolabeled α-aminoisobutyric acid (AIB; MW 103), sucrose (MW 342.3), inulin (MW 5000), and dextran (MW 70,000), horseradish peroxidase (HRP) and Evans blue (EB). Permeability was expressed as the unidirectional transfer constant Ki (μl/g/min). The permeabilities ( Ki) of tumors in the bradykinin group versus the control saline group for AIB, sucrose, inulin, and dextran were 25.91 ± 6.78 vs. 13.95 ± 4.29 (p < 0.01), 17.90 ± 2.65 vs. 10.75 ± 4.55 (p < 0.01), 23.92 ± 6.99 vs. 6.20 ± 4.37 (p < 0.01), and 17.84 ± 1.00 vs. 1.47 ± 1.24 (p < 0.001), respectively (mean ± SD). Permeability of RG2 gliomas to high molecular weight dextran (70,000) was 12-fold higher in the bradykinin group than in the saline infusion group. Intracarotid infusion of bradykinin did not significantly increase the blood volume in tumor or brain tissue despite its known vasodilative effect. The permeability of normal brain capillaries was unaffected by intracarotid bradykinin infusion. The increased permeability was reversed 20 min after stopping the intracarotid infusion. Electron microscopic and gross qualitative analysis was performed using HRP and EB. Intracarotid bradykinin infusion increased HRP and EB within tumor tissue but not normal tissue. We believe that intracarotid infusion of bradykinin will be a useful technique for selective delivery of antitumor compounds to brain tumors.
Simultaneous Assessment of Vasoreactivity Using Transcranial Doppler Ultrasound and Cerebral Blood Flow in Healthy Subjects
Authors: Dahl Arve , Russell David , Nyberg-Hansen Rolf , Rootwelt Kjell , Mowinckel Petter
Published: April 2011

Blood flow velocities in both middle cerebral arteries and regional cerebral blood flow in their perfusion territories were measured simultaneously in 36 healthy subjects. In 20 subjects, the measurements were first performed under basal conditions and then repeated 15–20 min after an intravenous injection of 1 g of acetazolamide. Reproducibility of simultaneous blood flow and velocity measurements was tested by examining 16 subjects on two occasions under basal conditions with an interval of 20 min. Relatively good reproducibility was found for repeated measurements of velocity and blood flow, being best when side-to-side asymmetry was assessed. The increase in blood flow velocities after acetazolamide was symmetrical (right side, 34.2%; and left side, 35.5%), and the velocity increase was significantly correlated with basal values. The increase in cerebral blood flow was also symmetrical (right side, 29.8%; left side, 30.1%) but not correlated with basal flow values. No significant relationship was found between velocity increase and increase in regional cerebral blood flow. This finding is probably not only due to methodological inaccuracies but may suggest that acetazolamide has an effect on the diameter of the middle cerebral artery or on the magnitude of this artery's perfusion territory. This study supports the use of acetazolamide for assessing cerebral vasoreactivity following the definition of lower limits for velocity and flow increase and for asymmetry of the response.
3-Acetylpyridine Produces Age-Dependent Excitotoxic Lesions in Rat Striatum
Authors: Schulz Jörg B. , Henshaw D. Ross , Jenkins Bruce G. , Ferrante Robert J. , Kowall Neil W. , Rosen Bruce R. , Beal M. Flint
Published: April 2011

The effects of 3-acetylpyridine (3-AP) were studied in rat striatum. Striatal injections of 3-AP produced dose-dependent lesions. The lesion size was significantly increased in 4- and 12-month-old rats compared to 1-month-old rats. Coinjection of the competitive N-methyl-d-aspartate (NMDA) antagonist 2-amino-5-phosphonovaleric acid (APV) or systemic administration of the noncompetitive NMDA antagonist MK-801, the competitive NMDA antagonist LY274614, or the glutamate release inhibitor lamotrigine partially but significantly attenuated striatal lesion volume. Consistent with an NMDA receptor-mediated excitotoxic effect, histologic studies showed that 3-AP lesions result in relative sparing of NADPH-diaphorase neurons. Using freeze clamp, 3-AP resulted in a marked depletion of ATP. Two-dimensional water-suppressed proton chemical shift magnetic resonance imaging showed a striatal depletion of the neuronal marker N-acetylaspartate but no focal increase in lactate during the first 3 h after intrastriatal 3-AP injections. Pretreatment with fructose-1,6-biphosphate attenuated the lesion volume significantly, which may be due to its ability to serve as a substrate for glycolytic metabolism, with resulting ATP production. The results of the present studies support the hypothesis that 3-AP produces an impairment of energy metabolism due to its substitution for niacinamide in the formation of NAD(P). Furthermore, 3-AP toxicity may involve a secondary excitotoxic mechanism mediated by NMDA receptors.
Swelling, Acidosis, and Irreversible Damage of Glial Cells from Exposure to Arachidonic Acid in vitro
Authors: Staub F. , Winkler A. , Peters J. , Kempski O. , Kachel V. , Baethmann A.
Published: April 2011

Swelling and damage of C6 glioma cells and of primary cultured astrocytes were analyzed in vitro during incubation with arachidonic acid (AA; 20:4). The cells were suspended in a physiological medium supplemented with AA at concentrations of 0.001–1.0 m M. Cell swelling was quantified by flow cytometry with hydrodynamic focusing. Flow cytometry was also utilized for assessment of cell viability by exclusion of the fluorescent dye propidium iodide and for measurement of the intracellular pH (pHi) by 2′,7′-bis-(2-carboxyethyl)−5(and −6)carboxyfluorescein. Administration of AA caused an immediate dose-dependent swelling of C6 glioma cells, even at a concentration of 0.01 m M. At this level cell volume increased within 20 min to 105.0% of control, at 0.1 m M to 111.0%, while at 1.0 m M to 123.7%. Following a phase of rapid cell volume increase, swelling leveled off during the subsequent observation period of 70 min. Viability of the C6 glioma cells was 90% under control conditions. It remained unchanged after raising AA concentrations to 0.1 m M. At 0.5 m M, however, cell viability fell to 72.8%, and at 1.0 m M to 32.7%. pHi of the glioma cells was 7.3 under control conditions. In parallel with the early swelling phase, AA led to a dose-dependent decrease of the intracellular pH and an elevated lactate production of the cells. During incubation with 0.1 m M AA, pHi decreased to 7.06 after 5 min, but recovered to normal subsequently. In addition, swelling-inducing properties of linoleic (18:2) or stearic (18:0) acid were analyzed for evaluation of the specificity of glial swelling induced by AA. Whereas stearic acid (0.1 m M) failed to induce a swelling response, linoleic acid (0.1 m M) was found to be effective. The volume increase of the glial cells, however, was only half of that found during exposure to AA at the same concentration. Further, glial swelling from AA or linoleic acid was completely inhibited by the aminosteroid U-74389F, an antagonist of lipid peroxidation. Finally, omission of Na+ ions in the suspension medium with replacement by choline led also to inhibition of the cell volume increase by AA. Experiments using astrocytes from primary culture confirmed the swelling-inducing properties of AA at a quantitative level, whereas vulnerability of the cells to AA was increased. The present results demonstrate an important role of AA in cytotoxic swelling and irreversible damage of glial cells at concentrations that occur in vivo in cerebral ischemia or trauma. The damaging potential of AA might be enhanced by a concurrently evolving intracellular acidosis, stimulating the formation of oxygen-derived free radicals and lipid peroxidation.
Digital Imaging of Umbelliferone Clearance: A Method for Repeated Measurements of Cerebral Cortical Blood Flow with High Temporal and Spatial Resolution
Authors: Kirkpatrick P. J. , Bucknall R. , Naylor P. , Rudman S. , Strong A. J.
Published: April 2011

We have developed a procedure for digital imaging of the exposed cerebral cortex during elution of a fluorescent dye. This avoids disturbing the cortex and has provided a method for the repeated estimation of regional CBF (rCBF) with a high topographical resolution. Under varying conditions of MABP and arterial blood gases, grey-level images of the exposed cortex irradiated with ultraviolet light (340 or 370 nm) were digitised (8 bits) at 15-s intervals after the injection of 1–2 ml of saturated umbelliferone solution into the lingual or external carotid artery of anaesthetised cats and rabbits. Specifically designed software allowed (a) regions of interest (ROIs) in the exposed cortex to be defined that were automatically applied to the sequence of images in a selected clearance and (b) solution of the initial slope equation for rCBF from the decay in grey-level fluorescence by exponential regression. Separate software that solved the equation at the level of a single pixel allowed a pseudocolour map of cortical rCBF to be generated. The factors affecting the resolution of this technique have been identified and quantified. Thus consistent and reproducible results were obtained provided that the fluorescence enhancement exceeded 20 grey levels and the r2 coefficient for regression was 90% or above. Mean rCBF values of 99.5 [95% confidence interval (CI), 89.4–110] ml 100 g−1 min−1 were obtained for rabbits ( N = 12; mean MABP = 75.2; mean Paco2 = 32.9; Pao2 = 111.8; pH 7.38) and 65.1 (95% CI, 55.1–75.1) ml 100 g−1 min−1 for cats ( N = 8; mean MABP = 92.8; Paco2 = 31.5; Pao2 = 114.6, pH 7.40). The method registers the cerebrovascular responses of autoregulation and CO2 reactivity and can detect transient changes in rCBF induced by direct cortical stimulation. Early experience suggests potential use of the method for the monitoring of changes in regional rCBF during ischaemia and for sequential measurement of changes in cortical perfusion in any study where a high topographical resolution is needed.
The Human Brain GLUT1 Glucose Transporter: Ultrastructural Localization to the Blood—Brain Barrier Endothelia
Authors: Cornford Eain M. , Hyman Shigeyo , Swartz Barbara E.
Published: April 2011

Immunogold electron microscopy was used to examine human brain resections to localize the GLUT1 glucose transporter. The tissue examined was obtained from a patient undergoing surgery for treatment of seizures, and the capillary profiles examined had characteristics identical to those described previously for active, epileptogenic sites (confirmed by EEG analyses). A rabbit polyclonal antiserum to the full-length human erythrocyte glucose transporter (GLUT1) was labeled with 10-nm gold particle-secondary antibody conjugates and localized immunoreactive GLUT1 molecules in human brain capillary endothelia, with <0.25% of the particles beyond the capillary profile. Erythrocyte membranes were also highly immunoreactive, whereas macrophage membranes were GLUT1-negative. The number of immunoreactive sites per capillary profile was observed to be 10-fold greater in humans than in previous studies of rat and rabbit brain capillaries. In addition, half of the total number of immunoreactive gold particles were localized to the luminal capillary membrane. We suggest that the blood–brain barrier GLUT1 glucose transporter is up-regulated in seizures, and this elevated transporter activity is characterized by increased GLUT1 transporters, particularly on the luminal capillary membranes. In addition, acute modulation of glucose transporter activity is presumed to involve translocation of GLUT1 from cytoplasmic to luminal membrane sites, demonstrable with quantitative immunogold electron microscopy.
SPECT Quantification of [123I]Iomazenil Binding to Benzodiazepine Receptors in Nonhuman Primates: I. Kinetic Modeling of Single Bolus Experiments
Authors: Laruelle Marc , Baldwin Ronald M. , Rattner Zachary , Al-Tikriti Mohammed S. , Zea-Ponce Yolanda , Zoghbi Sami S. , Charney Dennis S. , Price Julie C. , Frost J. James , Hoffer Paul B. , Innis Robert B.
Published: April 2011

The aim of this work was to study the feasibility and reproducibility of in vivo measurement of benzodiazepine receptors with single photon emission computerized tomography (SPECT) in the baboon brain. Arterial and brain regional activities were measured for 420 min in three baboons after single bolus injection of the benzodiazepine antagonist [123I]iomazenil. Data were fit to a three-compartment model to derive the regional binding potential (BP), which corresponds to the product of the receptor density, (Bmax) and affinity (1/KD). Regional BP values (from 114 in striatum to 241 in occipital) were in good agreement with values predicted from in vitro studies. Constraining the regional volume of distribution of the nondisplaceable compartment to the value measured during tracer constant infusion experiments in baboons (Laruelle et al., 1993) improved the identifiability of the rate constants. Each experiment was repeated to investigate the reproducibility of the measurement. The regional average reproducibility was 10 ± 5%, expressed as coefficient of variation (CV). Results of equilibrium analysis at peak uptake were in good agreement with results of kinetic analysis. Empirical counts ratio methods were found to be poorly sensitive to benzodiazepine receptor density. These studies suggest the feasibility of quantitative measurement of benzodiazepine receptors by kinetic analysis of SPECT data and the inadequacy of empirical methods of analysis, such as counts ratios, to evaluate differences in receptor density.
Transient Spinal Ischemia in the Rat: Characterization of Behavioral and Histopathological Consequences as a Function of the Duration of Aortic Occlusion
Authors: Marsala Martin , Yaksh Tony L.
Published: April 2011

To characterize the behavioral and histopathological changes that occur in spinal cord after transient ischemia, reversible occlusion of the descending aorta was achieved in the halothane (1–1.5%)-anesthetized rat by the insertion and subsequent inflation of a 2F Fogarty catheter for 10, 15, 20, or 30 min, Neurological recovery was tested during 8 h of reperfusion. After reflow, animals undergoing 30 min of ischemia displayed an initial flaccidity at 1 h, spasticity at 4 h, and flaccidity at the end of 8 h. Following 20 min of ischemia the initial flaccidity was followed by hindlimb spasticity that persisted for 8 h. Shorter intervals of ischemia had minimal effects on motor function. After reflow, animals developed a prominent allodynea, the incidence of which was dependent on the duration of ischemia. A clear correlation of histopathological changes with the degree of neurological deficit was noted. In spastic animals, small and medium-sized interneurons localized in laminae III to VII were affected. Animals with flaccidity at 8 h additionally displayed a significant incidence of argyrophilic A motoneurons in the ventral horns. Corresponding to the frequent appearance of allodynea, these animals also showed a significant number of damaged neurons in lamina II.
A Comparison of the Early Development of Ischaemic Damage following Permanent Middle Cerebral Artery Occlusion in Rats as Assessed Using Magnetic Resonance Imaging and Histology
Authors: Gill R. , Sibson N. R. , Hatfield R. H. , Burdett N. G. , Carpenter T. A. , Hall L. D. , Pickard J. D.
Published: April 2011

Recent developments in diffusion-weighted imaging (DWI) have enabled the pathological changes that occur during cerebral ischaemia to be studied. The present studies utilised DWI to investigate the development of early ischaemic changes following permanent middle cerebral artery (MCA) occlusion in the rat, which represents a model of stroke. An increased DWI signal was seen in the region of the occluded MCA and this was detectable as early as 1 h postocclusion. DWI images were obtained at nine stereotactic levels throughout the brain, providing a quantifiable measure of the volume of increased signal intensity in each animal. At 1 h post-MCA occlusion the hyperintense areas were seen in the frontoparietal cortex and lateral caudate nucleus; these areas represent the core of the infarct and no protection is seen with any compounds in these areas. There was a progressive increase in the area of hyperintensity up to 4 h post-MCA occlusion, and at this time point the hyperintensity was seen in the dorsolateral cortex and caudate nucleus. At 4 h post-MCA occlusion there was a significant correlation between the volume of hemispheric and cortical ischaemic damage measured using DWI and histology. Thus, it appears that the increased DWI signal seen during the early time points after MCA occlusion was demarcating tissue that was destined for infarction. The area beyond the hyperintense region at 1 h represents the so-called “penumbral” region, because with increasing time (post-MCA occlusion) this area became incorporated into the infarct. There was also a slight increase in infarct size between 4 and 24 h, when assessed using DWI or histology, although two groups of animals were being compared, as opposed to the time-course study, in which just one group of animals was used. At 24 h post-MCA occlusion there was a good correlation between DWI, histology, and conventional T2 weighted imaging. There was no further increase in size of the infarct between 24 h and 7 days as assessed using histology and T2-weighted imaging. DWI could not be used to quantify infarct volume at 7 days because there was no uniform signal in the damaged area. At 7 days the area of infarction actually appeared to be darker in the diffusion-weighted images. The hyperintensity seen in diffusion-weighted images appears to decrease some time between 24 h and 7 days. These data provide evidence that DWI can be used to detect early ischaemic changes and it appears that the hyperintense areas seen at 1 h delineates tissue that is destined to become the core of the infarct. Thus, DWI could be used to identify the so-called penumbral areas, which are salvageable if treated within the therapeutic window of 2–3 h and may therefore have an important role in detecting such changes in the clinical setting.
Coupled Glucose Transport and Metabolism in Cultured Neuronal Cells: Determination of the Rate-Limiting Step
Authors: Whitesell Richard R. , Ward Michael , McCall Anthony L. , Granner Daryl K. , May James M.
Published: April 2011

In brain and nerves the phosphorylation of glucose, rather than its transport, is generally considered the major rate-limiting step in metabolism. Since little is known regarding the kinetic coupling between these processes in neuronal tissues, we investigated the transport and phosphorylation of [2-3H]glucose in two neuronal cell models: a stable neuroblastoma cell line (NCB20), and a primary culture of isolated rat dorsal root ganglia cells. When transport and phosphorylation were measured in series, phosphorylation was the limiting step, because intracellular glucose concentrations were the same as those outside of cells, and because the apparent Km for glucose utilization was lower than expected for the transport step. However, the apparent Km was still severalfold higher than the Km of hexokinase I. When [2-3H]glucose efflux and phosphorylation were measured from the same intracellular glucose pool in a parallel assay, rates of glucose efflux were three- to-fivefold greater than rates of phosphorylation. With the parallel assay, we observed that activation of glucose utilization by the sodium channel blocker veratridine caused a selective increase in glucose phosphorylation and was without effect on glucose transport. In contrast to results with glucose, both cell types accumulated 2-deoxy-d-[14C]glucose to concentrations severalfold greater than extracellular concentrations. We conclude from these studies that glucose utilization in neuronal cells is phosphorylation-limited, and that the coupling between transport and phosphorylation depends on the type of hexose used.
Global Cerebral Ischemia in the Rat: Online Monitoring of Oxygen Free Radical Production Using Chemiluminescence in vivo
Authors: Dirnagl Ulrich , Lindauer Ute , Them Andreas , Schreiber Stefan , Pfister Hans-Walter , Koedel Uwe , Reszka Regine , Freyer Dorette , Villringer Arno
Published: April 2011

Using online in vivo chemiluminescence (CL), we studied for the first time continuously the production of reactive oxygen species (ROS) after global cerebral ischemia and the relationship of ROS production to CBF. In anesthetized rats equipped with a closed cranial window, the CL enhancer, lucigenin (1 m M), was superfused onto the brain topically. CL was measured through the cranial window with a cooled photomultiplier, and CBF was measured simultaneously with laser–Doppler flowmetry. Reperfusion after 10 min (n = 8) of global cerebral ischemia led to a CL peak to 188 ± 77% (baseline = 100%) within 10 ± 4 min. After 2 h of reperfusion, CL had returned to 102 ± 28%. Reperfusion after 20 min (n = 8) of ischemia increased CL to 225 ± 48% within 12 ± 3 min. After 2 h, CL was still increased (150 ± 44%, p < 0.05 compared with 10 min of ischemia). CL after 10 min of ischemia was neither affected by brain topical free CuZn-superoxide dismutase (SOD) (100 U/ml, n = 3) nor by i.v. administration of free CuZn-SOD (104 U/kg, followed by 104 U/kg/h, n = 3). The CBF hyperperfusion peak on reperfusion preceded the CL peak in all experiments by several minutes. In additional in vitro experiments we investigated the source of CL: Intracellular loading of lucigenin was demonstrated in cultured CNS cells, and a very similar pattern of CL as in the in vivo preparation after ischemia developed in rat brain slices after 15 min of hypoxia, which was unaffected by free CuZn-SOD (100 U/ml) but strongly attenuated by liposome-entrapped CuZn-SOD. We conclude that lucigenin-enhanced CL is a promising tool to study ROS production continuously from the in vivo brain of experimental animals and brain slices, and that the CL signal most likely derives from the intracellular production of superoxide. The production of ROS is preceded by reperfusion, is burst-like, and is dependent on the duration of the ischemic interval.
Reexpression of Developmentally Regulated MAP2c mRNA after Ischemia: Colocalization with hsp72 mRNA in Vulnerable Neurons
Authors: Saito Nobuhito , Kawai Kensuke , Nowak Thaddeus S.
Published: April 2011

Levels of mRNAs encoding the microtubule-associated proteins MAP2b and MAP2c as well as the 70-kDa stress protein [72-kDa heat shock protein (hsp72)] were evaluated in postischemic rat brain by in situ hybridization with oligonucleotide probes corresponding to the known rat sequences. Rats were subjected to 10-min cardiac arrest, produced by compression of major thoracic vessels, followed by resuscitation. The normally expressed MAP2b mRNA showed transient twofold elevations in all hippocampal neuron populations at 6-h recirculation, followed by a return to control levels by 24 h. MAP2b hybridization was progressively lost thereafter from the vulnerable CA1 and outer cortical layers, preceding both the fall in immunoreactive MAP2b and the eventual cell loss in these regions. The depletion of MAP2b mRNA coincided with an increase in the alternatively spliced MAP2c in vulnerable regions during 12–48 h of recirculation, precisely overlapping the late component of hsp72 expression that persisted in these cell populations. Previous studies have suggested that the initial induction of hsp72 provides an index of potential postischemic injury in neuron populations that may or may not be injured, while lasting hsp72 mRNA expression is associated with cell damage. In contrast, the present results demonstrate that MAP2c expression under these conditions occurs uniquely in neuron populations subject to injury. Available evidence suggests that MAP2c expression represents a plastic response in subpopulations of neurons that will survive in these regions, although it remains to be explicitly determined whether it may also be transiently expressed in dying cells. In any case, these observations demonstrate that reexpression of developmentally regulated MAP2c mRNA is a relatively late postischemic response in vulnerable cell populations, indicating that pathways regulating MAP2 splicing may be closely associated with mechanisms of neuron injury and/or recovery.
D2 Receptor Blockade by Flunarizine and Cinnarizine Explains Extrapyramidal Side Effects. A SPECT Study
Authors: Brücke T. , Wöber Ch. , Podreka I. , Wöber-Bingöl C. , Asenbaum S. , Aull S. , Wenger S. , Ilieva D. , van der Meer C. Harasko , Wessely P. , Deecke L.
Published: April 2011

Twenty-six patients under treatment with the calcium channel blockers flunarizine (Fz) or cinnarizine (Cz) were examined with single-photon emission computed tomography using [123I]iodobenzamide as a ligand. The striatal dopamine D2 receptor-binding potential was determined and found to be reduced by 14 to 63% (39.5 ± 15.0%; p < 0.0001) in patients compared with age-matched control values. This reduction was larger in 12 patients with extrapyramidal symptoms and was only slowly reversible after discontinuation of treatment. Patients treated for >6 months had significantly larger reductions than patients treated for a shorter period. Parkinsonian symptoms were only seen in patients older than 50 years. Our findings prove a neuroleptic-like action of Fz and Cz, which seems to be the major reason for their extrapyramidal side effects. Older age and long-term treatment are predisposing factors for these effects.
Poor Hemodynamic and Metabolic Recovery after Global Incomplete Cerebral Ischemia Associated with Short-Term Diabetes in Dogs
Authors: Palmon Sally C. , Sieber Frederick E. , Brown P. Randall , Koehler Raymond C. , Eleff Scott M. , Traystman Richard J.
Published: April 2011

We determined the effect of 4–5 weeks of diabetes on ATP recovery following global incomplete cerebral ischemia. 31P magnetic resonance spectra of ATP, intracellular pH (pHi), and CBF (radiolabeled microspheres) were measured in three groups of anesthetized dogs (n = 8/group): chronic hyperglycemic diabetes (pancreatectomy followed by blood glucose of >10 m M for 4–5 weeks); acute hyperglycemia (blood glucose of >10 m M) during ischemia and reperfusion in nondiabetic dogs; and normoglycemic controls. Twenty minutes of incomplete ischemia was produced by ventricular fluid infusion to keep cerebral perfusion pressure (CPP) at 10 mm Hg during spontaneous variations in MABP. Intracranial pressure was increased initially to similar levels, resulting in a similar Cushing response among the groups. However, during the final 8 min of ischemia, MABP decreased to a greater extent in diabetic (86 ± 42 mm Hg) than in hyperglycemic (162 ± 30 mm Hg) and normoglycemic (135 ± 54 mm Hg) groups and remained lower throughout 3 h of reperfusion. CPP was kept constant during ischemia, but was lower throughout reperfusion in diabetic dogs. During ischemia CBF was reduced similarly among groups: 5 ± 3 ml · min−1 · 100 g−1 in hyperglycemic and normoglycemic and 4 ± 3 ml · min−1 · 100 g−1 in diabetic dogs. During reperfusion early hyperemia was attenuated and delayed hypoperfusion was augmented (7 ± 17 ml · min−1 · 100 g−1 by 180 min) as a result of low perfusion pressure in diabetics. However, medullary blood flow was similar among groups. Endischemic ATP was similar among groups (25 ± 34, 30 ± 25, and 4 ± 11% of baseline in normoglycemic, hyperglycemic and diabetic dogs, respectively). Following 180 min of reperfusion ATP was 87 ± 15, 45 ± 38, and 7 ± 19% of control in normoglycemic, hyperglycemic and diabetic dogs, respectively. End-ischemic pHi was lower in hyperglycemic (5.94 ± 0.16) and diabetic (5.96 ± 0.08) than normoglycemic (6.32 ± 0.04) groups. By 45 min of reperfusion, pHi recovered in normoglycemic (7.02 ± 0.09) but not in diabetic (6.13 ± 0.38) or hyperglycemic dogs (6.44 ± 0.28). This study shows that MABP, CBF, ATP, and pHi following global incomplete ischemia did not recover in diabetic dogs. In diabetics poor metabolic recovery was related to the MABP decreases during ischemia and reperfusion. These results contrast with previous studies from our laboratory showing good metabolic recovery with longer-term diabetes (3–4 months). Our findings indicate that in this diabetic model, the cerebral ischemic response is dependent on duration of diabetes.
One-pot synthesis of benzofused heteroaryl azoles via tandem C-heteroatom coupling/C–H activation of azoles
Authors: Qin Xurong , Cong Xuefeng , Zhao Dongbing , You Jingsong , Lan Jingbo
Published: April 2011

The Cu(i) or Pd(ii)-catalyzed cross-couplings of gem-dihaloolefins with azoles via tandem C-heteroatom coupling/C–H activation for the preparation of benzofused heteroaryl azoles have been developed.
Which, if any, hydrates will crystallise? Predicting hydrate formation of two dihydroxybenzoic acids
Authors: Braun Doris E. , Karamertzanis Panagiotis G. , Price Sarah L.
Published: April 2011

A study of two dihydroxybenzoic acid isomers shows that computational methods can predict which hydrates are formed and provide information that helps in characterising hydrate forms.
A versatile precursor for non-aqueous neptunyl(v) chemistry
Authors: Copping Roy , Mougel Victor , Petit Sebastien , Auwer Christophe Den , Moisy Philippe , Mazzanti Marinella
Published: April 2011

The polymeric complex [(NpO2Py5)(KI2Py2)]n is prepared from dry “NpO2Cl” by anion exchange with potassium iodide in pyridine affording the first convenient starting material for the development of NpO2+ coordination chemistry in anhydrous organic media.
The importance of methylation in the binding of (ferrocenylmethyl)tempammonium guests by cucurbit[n]uril (n = 7, 8) hosts
Authors: Yi Song , Captain Burjor , Kaifer Angel E.
Published: April 2011

The extent of methylation of a guest's ammonium nitrogen controls the predominant binding site with the cucurbit[8]uril host.
Basic analysis on a thermoacoustic engine with gas and liquid
Authors: Tang Ke , Lei Tian , de Waele A. T. A. M , Jin Tao
Published: April 2011

This paper analyzes the basics of a thermoacoustic engine with gas and liquid as working fluids. The governing equations for the engine are deduced from the dynamics of each individual component. From the governing equations, analytical expressions are obtained for oscillation frequency and onset temperature. The relations for the dependence of the displacement amplitude of liquid column, the velocity amplitude at the end of resonant tube, the pressure amplitude gradient and the enthalpy flow in stack, on the pressure amplitude in resonant tube are formulated. The calculation with the deduced formulae shows that an oscillation frequency below 10 Hz can be achieved in the thermoacoustic engine with gas and liquid. Meanwhile, a lower oscillation frequency, as well as a lower onset temperature, requires a larger liquid mass and a lower mean pressure. Experiments, focusing on the oscillation frequency and the onset temperature, have been performed to validate the computation.