bluuebunny/crossref_metadata_2025 huggingface dataset:
A Method for 14C and 3H Double-Label Autoradiography
Authors: Juhler M. , Diemer N. H.
Published: April 2011

The present study describes and validates a 3H/14C double-label autoradiographic method in which separation of the labels was obtained by sequential film exposures to film types sensitive to 14C only and to both 3H and 14C, respectively. The error in assuming a pure 14C image on the first film was 2–3%. A linear subtraction equation was developed for calculation of the 3H activity expressed in 14C equivalents as the difference in tissue activity between the second and the first film exposure. The actual 3H activity in the tissue could be obtained by multiplying the result by 25 (14C to 3H conversion factor). The subtraction procedure was validated for absolute 14C and 3H activities of 100–1,300 nCi/g and 600–11,000 nCi/g, respectively, and for relative 3H/14C activities between 5 and 10. Self-absorption of 3H in white matter was corrected for by multiplication by 1.61 (self-absorption coefficient). This factor was close to unity for 14C.
Light and Electron Microscopic Evaluation of Hydrogen Ion-Induced Brain Necrosis
Authors: Petito C. K. , Kraig R. P. , Pulsinelli W. A.
Published: April 2011

Excessive accumulation of hydrogen ions in the brain may play a pivotal role in initiating the necrosis seen in infarction and following hyperglycemic augmentation of ischemic brain damage. To examine possible mechanisms involved in hydrogen ion-induced necrosis, sequential structural changes in rat brain were examined following intracortical injection of sodium lactate solution (pH 4.5), as compared with injections at pH 7.3. Following pH 7.3 injection, neuronal swelling developed between 1 and 6 h, but only a needle track wound surrounded by a thin rim of necrotic neurons and vacuolated neuropil was present 24 h after injection. In contrast, pH 4.5 injection produced neuronal necrosis as soon as 1 h after injection, followed by necrosis of astrocytes and intravascular thrombi at 3 and 6 h. Alterations common to both groups included vascular permeability to horseradish peroxidase, dilation of extracellular spaces, astrocyte swelling, capillary compression, and vascular stasis. These data suggest that neurons, astrocytes, and endothelia can be directly damaged by increased acid in the interstitial space. Lethal injury initially appeared to affect neurons, while subsequent astrocyte necrosis and vascular occlusion may damage tissue by secondary ischemia.
Time-Dependent Changes of Lumped and Rate Constants in the Deoxyglucose Method in Experimental Cerebral Ischemia
Authors: Nakai Hirofumi , Yamamoto Y. Lucas , Diksic Mirko , Matsuda Hiroshi , Takara Eiichi , Meyer Ernst , Redies Christoph
Published: April 2011

Time-dependent changes in the lumped and rate constants in a bilateral middle cerebral artery (MCA) occlusion in cats were evaluated. These variables were measured in 11 cats after a sham operation, in five after a 1-h occlusion, in two after a 2-h occlusion, in five after a 4-h occlusion, and in four after a 16-h occlusion. The time course of the cerebral tissue radioactivity [Ci* ( t)] was monitored by external coincidence counting during a programmed infusion of [18F]2-fluorodeoxyglucose (FDG). Arterial plasma concentration [Cp* ( t)] of tracer was kept constant during the first 45 min. Comparison of k2* and k3* in the sham-operated group, estimated by external coincidence counting, and by the ratio of extraction fractions of glucose and [18F]2-FDG, demonstrated no significant difference between these rate constants in these two groups of animals. The rate and lumped constants were also estimated from Ci* ( t) and Cp* ( t), as well as from the ratio of extraction fractions of glucose and [18F]2-FDG, respectively, in the MCA occlusion group. Significant decrease in k3* was observed after 1 h of occlusion (20% lower than in the sham operation, p < 0.05); in ki* decrease occurred within 4 h of occlusion (21% lower than in the sham operation, p < 0.05). However, decrease in k2* was observed only after 16 h of occlusion (26% lower than in the sham operation, p < 0.05). Namely, decrease of rate constants occurred first in k3* then in k1* and k2*. In contrast to the rate constants where decrease was observed, a significant increase in the lumped constant occurred between 1 and 4 hours after occlusion (55% higher than in the 1-h occlusion, p < 0.05). However, the values of the lumped constant were not significantly different between the 4-h occlusion group and the 16-h occlusion group.
Scaled Subprofile Model: A Statistical Approach to the Analysis of Functional Patterns in Positron Emission Tomographic Data
Authors: Moeller J. R. , Strother S. C. , Sidtis J. J. , Rottenberg D. A.
Published: April 2011

The data obtained from measurements of regional rCMRglu using [18F]fluorodeoxyglucose (FDG)/positron emission tomographic (PET) data contain more structure than can be identified with group mean rCMRglu profiles or regional correlation coefficients. This additional structure is revealed by a novel mathematical-statistical model of regional metabolic interactions that explicitly represents rCMRglu profiles as a combination of region-independent global effects, a group mean pattern and a mosaic of interacting networks. In its application to FDG/PET data, this model removes global subject effects [global scaling factors (GSFs)] and a group mean pattern (profile) so as to maximize statistical power for the detection and simultaneous discovery of all networks of two or more regions that form a significant and consistent linearly covarying pattern. The model approach presented here was applied to the combined rCMRglu data from 12 demented AIDS patients and 18 normal controls: Two significant metabolic covariance pattern descriptors that together accounted for 71 to 96% of the rCMRglu/GSF variation across subjects for 22/28 regions in the AIDS group were extracted. Each descriptor was found to be highly correlated with performance on several neuropsychological tests, providing independent validation of the analysis technique as a means of discovering and describing behaviorally related components of group rCMRglu profiles.
Hematocrit in Cerebral Capillaries and Veins under Control and Ischemic Conditions
Authors: McHedlishvili George , Varazashvili Manana
Published: April 2011

Red blood cell concentration and hematocrit were assessed in the blood flowing in the capillaries of rabbit cerebral cortex (in unstained histologic sections prepared after in situ fixation of the tissue) and in the blood samples taken from the respective veins using direct, quantitative techniques. The values appeared to be significantly higher in veins than in capillaries under control conditions. With the development of cerebral ischemia, when the blood flow in the cortex was reduced to almost half of the control level, red cell concentration and hematocrit in the blood were found to decrease markedly both in the capillaries and veins of the brain, while in the femoral veins of the same animals the values remained essentially unchanged.
Ischemia-Induced Neuronal Cell Death, Calcium Accumulation, and Glial Response in the Hippocampus of the Mongolian Gerbil and Protection by Propentofylline (HWA 285)
Authors: DeLeo J. , Toth L. , Schubert P. , Rudolphi K. , Kreutzberg G. W.
Published: April 2011

The localization and timing of cellular calcium loading and glial cell reaction in relation to selective death of hippocampal neurons was studied in Mongolian gerbils following transient forebrain ichemia. Two days after a 5-min period of ischemia, heavy calcium staining was histochemically demonstrated in circumscribed groups of nerve cells, located in the transition zone between the CA1 and CA3 areas. This preceded complete neuronal cell death that was quantitatively assessed by measuring the intensity of Nissl staining. After a 12-min period of ischemia, extensive calcium loading was observed in conjunction with severe neuronal damage throughout the CA1 region as well in the dorsal nuclei of the thalamus. The extent of calcium staining decreased with time and was not seen at stages later than 7 days. Already at 2 days after a 5-min period of ischemia, a strong increase of glial fibrillary acidic protein immunoreactivity was seen. This indicates a marked and early hypertrophy of astrocytes that was not accompanied by an obvious proliferation. Neither the astrocytic response nor the neuronal calcium accumulation were observed in gerbils pretreated with propentofylline, HWA 285 (10 mg/kg, i.p.) 15 min before bilateral carotid artery occlusion. Also, the decrease of Nissl staining in the CA1 area after 5 and 12 min of ischemia was considerably less pronounced and did not significantly differ from sham-operated controls.
Cerebral Ischemia and Reperfusion: Prevention of Brain Mitochondrial Injury by Lidoflazine
Authors: Rosenthal Robert E. , Hamud Fozia , Fiskum Gary , Varghese P. Jacob , Sharpe Steven
Published: April 2011

Mitochondrial degradation is implicated in the irreversible cell damage that can occur during cerebral ischemia and reperfusion. In this study, the effects of 10 min of ventricular fibrillation and 100 min of spontaneous circulation on brain mitochondrial function was studied in dogs in the absence and presence of pretreatment with the Ca2+ antagonist lidoflazine. Twenty-three beagles were separated into four experimental groups: (i) nonischemic controls (ii) those undergoing 10-min ventricular fibrillation, (iii) those undergoing 10-min ventricular fibrillation pretreated with 1 mg/kg lidoflazine i.v., and (iv) those undergoing 10-min ventricular fibrillation followed by spontaneous circulation for 100 min. Brain mitochondria were isolated and tested for their ability to respire and accumulate calcium in a physiological test medium. There was a 35% decrease in the rate of phosphorylating respiration (ATP production) following 10 min of complete cerebral ischemia. Those animals pretreated with lidoflazine showed significantly less decline in phosphorylating respiration (16%) when compared with nontreated dogs. Resting and uncoupled respiration also declined following 10 min of fibrillatory arrest. One hundred minutes of spontaneous circulation following 10 min of ventricular fibrillation and 3 min of open-chest cardiac massage provided complete recovery of normal mitochondrial respiration. Energy-dependent Ca2+ accumulation by isolated brain mitochondria was unimpaired by 10 min of complete cerebral ischemia. However, by 100 min after resuscitation, there was a small, but significant rise in the capacity for mitochondrial Ca2+ sequestration when compared to either control or fibrillated groups. These findings indicate that: (a) 10 min of complete cerebral ischemia causes a substantial decline in the rate at which cortical brain mitochondria can synthesize ATP; (b) pretreatment with lidoflazine significantly protects the ability of brain mitochondria to synthesize ATP following 10-min ventricular fibrillation, (c) mitochondrial damage is completely reversible by 100 min following restoration of circulation, (d) mitochondrial Ca2+ uptake is relatively insensitive to the adverse effects of ischemia.
Experimental Model for Repetitive Ischemic Attacks in the Gerbil: The Cumulative Effect of Repeated Ischemic Insults
Authors: Tomida Shuichi , Nowak Thaddeus S. , Vass Karl , Lohr Julia M. , Klatzo Igor
Published: April 2011

An experimental model for repeated ischemic attacks, which allows easy induction of cerebral ischemia of any desired duration and frequency, has been developed in the gerbil. With this procedure, a pronounced cumulative effect on development of edema and tissue injury was observed using 3 separate, 5-min bilateral occlusions of the common carotid arteries spaced at various time intervals. This effect was most evident when the occlusions were carried out at 1-h intervals, i.e., during the period of marked postischemic hypoperfusion. Such animals, killed after 24 h of recirculation, showed significantly more severe edema and brain tissue injury in the areas exposed to ischemia than was observed in animals killed 24 h after single 5- or 15-min occlusions. The changes of regional CBF, assayed with a [3H]nicotine method, indicated a relatively rapid onset of hypoperfusion of similar degree after each release of arterial occlusion. The hypoperfusion recovered significantly within 6 h of recirculation following either single or multiple occlusions, and no residual hypoperfusion was observed in animals which, when killed at 24 h, showed severe edema and brain tissue injury. This model should prove useful in elucidating the pathophysiological mechanisms operative in repetitive cerebral ischemia.
Electrolyte Shifts between Brain and Plasma in Hypoglycemic Coma
Authors: Siesjö Bo K. , Deshpande Jayant K.
Published: April 2011

Hypoglycemia of sufficient severity to cause cessation of EEG activity (coma) is accompanied by energy failure and by loss of ion homeostasis, the latter encompassing a marked rise in extracellular fluid (ECF) K+ concentration and a fall in ECF Ca2+ concentration. Presumably, ECF Na+ concentration decreases as well. In the present study, the extent that the altered ECF-plasma gradients give rise to net ion fluxes between plasma and tissue is explored. Accordingly, whole tissue contents of Ca2+, Mg2+, K+, and Na+ were measured. The experiments were carried out in anaesthetized and artificially ventilated rats given insulin i.p.; cerebral cortical tissue was sampled at the stage of slow-wave EEG activity, after 10, 30, and 60 min of coma (defined as isoelectric EEG), as well as after 1.5, 6, and 24 h of recovery. In the precomatose animals (with a slow-wave EEG pattern), no changes in electrolyte contents were observed. During coma, tissue Na+ content increased progressively and the K+ content fell (each by 20 μmol g−1 during 60 min). During recovery, these alterations were reversed within the first 6 h. The Mg2+ content remained unchanged. In spite of the appreciable plasma to ECF Ca2+ gradient, no significant calcium accumulation was observed. It is concluded that hypoglycemia leads to irreversible neuronal necrosis in the absence of gross accumulation of calcium in the tissue.
Positron Emission Tomography in Minor Ischemic Stroke Using Oxygen-15 Steady-State Technique
Authors: Pozzilli C. , Itoh M. , Matsuzawa T. , Fukuda H. , Abe Y. , Sato T. , Takeda S. , Ido T.
Published: April 2011

A study with positron emission tomography (PET) was performed on 10 patients with ischemic stroke and mild disability. The patients underwent cerebral angiography, x-ray computed tomography (CT) scan and regional cerebral measurements of CBF, CMRO2, oxygen extraction ratio (OER), and cerebral blood volume (CBV). Only minor arterial involvement was detected by angiography. In all patients, PET images of functional defects were more extensive than the corresponding CT hypodensity, and there were statistically significant reductions in CBF, CMRO2, and CBF/CBV ratio as compared with control subjects. Half of the regions analyzed in the affected hemisphere demonstrated a disruption of the normal coupling between CBF and CMRO2 as reflected by OER values significantly higher or lower than those of the corresponding region of the contralateral hemisphere. The pathophysiological pattern of high OER combined with a reduction in CBF proportionally greater than the reduction in CMRO2 was particularly indicative of regional chronic hemodynamic compromise in these patients.
Regional Differences in Arachidonic Acid Release in Rat Hippocampal CA1 and CA3 Regions during Cerebral Ischemia
Authors: Westerberg Eva , Deshpande Jayant K. , Wieloch Tadeusz
Published: April 2011

Changes in the levels of arachidonic acid during ischemia in selectively vulnerable areas of the hippocampus were studied in the rat brain. Since neurons in the CA, region are more vulnerable to ischemia than neurons in the adjacent CA3 region, the release of arachidonic acid in these two regions was measured during decapitation ischemia of 4- to 12-min duration. The concentration of free arachidonic acid increased with the duration of ischemia in both regions. However, the level was significantly higher in CA1, than in CA3 after 8 and 12 min of ischemia. This difference in arachidonic acid accumulation may reflect differences between the regions in agonist-dependent phospholipid breakdown as well as calcium-dependent phospholipase activity. The importance for the development of neuronal necrosis is discussed.
The Influence of Tissue Heterogeneity on Results of Fitting Nonlinear Model Equations to Regional Tracer Uptake Curves: With an Application to Compartmental Models Used in Positron Emission Tomography
Authors: Herholz K. , Patlak C. S.
Published: April 2011

An analytical method based on Taylor expansions was developed to analyze errors caused by tissue heterogeneity in dynamic positron emission tomography (PET) measurements. Some general rules concerning the effect of parameter variances and covariances were derived. The method was further applied to various compartmental models currently used for measurement of blood flow, capillary permeability, glucose metabolism, and tracer binding. Blood flow and capillary permeability are shown to be generally underestimated in heterogenous tissue, the underestimation being more severe for slowly decaying, constant or increasing input functions rather than for bolus input, and increasing with measurement time. Typical errors caused by the heterogeneity due to insufficient separation between gray and white matter by a PET scanner with full width at half-maximum (FWHM)= 5 to 10 mm resolution range between–0.9 and–6% in dynamic CBF measurements with intravenous (i. v.) bolus injection of 15O-water or inhalation of 18F-fluoromethane and total measurement times of6 or 10 min, respectively. Binding or metabolic rates determined with tracers that are essentially trapped in tissue (e.g., FDG for measurement of cerebral glucose metabolism) are only slightly overestimated (0.5–3.0%) at typical measurement times and are essentially independent of the shape of the input function. The error increases considerably if tracer accumulation is very slow, however, or if short measurement times [<5/(k2 + k3)] are used. Some rate constants are also subject to larger errors.
A Double-Radionuclide Autoradiographic Method Using N-Isopropyl-Iodoamphetamine for Sequential Measurements of Local Cerebral Blood Flow
Authors: Obrenovitch Tihomir Paul , Clayton Charles Barry , Strong Anthony John
Published: April 2011

A double-radionuclide autoradiographic method has been assessed for sequential determinations of local CBF (LCBF). It is based on two successive intravascular injections of N-isopropyl- p-iodoamphetamine (IMP) labelled with different radionuclides, whose concentrations can later be differentiated in the same tissue section using double-radionuclide autoradiography. Previous studies suggested that the distribution of IMP, up to 30 min after its administration, still represents LCBFs. Our data indicate that, provided the tracer is injected directly into the left ventricle, (a) there is little back diffusion from normal brain to blood under physiological conditions for at least 35 min following the tracer injection and (b) an injection of unlabelled IMP, in a dose larger than that used for blood flow determination, does not displace any labelled IMP previously taken up by the brain, nor does it displace any labelled IMP previously accumulated in the lung that would lead to secondary brain uptake. On the basis of these results, we conclude that sequential autoradiographic determinations of LCBF using IMP labelled with different radionuclides is possible. This is a promising experimental method for the simultaneous investigation of changes in LCBF in several CNS structures.
Autoradiographic Study of Regional Protein Synthesis in Focal Cerebral Ischemia with TCA Wash and Image Subtraction Techniques
Authors: Yoshimine T. , Hayakawa T. , Kato A. , Yamada K. , Matsumoto K. , Ushio Y. , Mogami H.
Published: April 2011

The standard biochemical method of trichloracetic acid (TCA) wash and the image processing technique were combined to differentiate and visualize the distributions of polypeptide-incorporated and unincorporated tracers in an autoradiographic study of regional protein synthesis, The validity of applying TCA wash procedures to cryostat sections was considered by histologic and chemical evaluations, For the autoradiographic study of in vivo protein synthesis, a tracer dose of L-[14C]valine was administered 30 min after occlusion of the posterior communicating artery in gerbils. Images of total (polypeptide-incorporated and unincorporated) radioactivity and of polypeptide-incorporated radioactivity were obtained from an identical cryostat section before and after TCA wash. The polypeptide-unincorporated radioactivity image was produced with an image processing system by subtracting pixel by pixel the polypeptide-incorporated radioactivity from the total radioactivity. The present study clearly demonstrated that in spite of the sufficient delivery of tracer amino acids, the polypeptide synthesis was completely lost in the ischemic focus. Free tracer was markedly accumulated in the brain adjacent to the ischemic focus. This kind of autoradiographic technique seems to be indispensable in studying the topographical complexity of the altered protein metabolism in the pathologic brain.
Cerebral Blood Volume Measured with Inhaled C15O and Positron Emission Tomography
Authors: Martin W. R. Wayne , Powers William J. , Raichle Marcus E.
Published: April 2011

Local cerebral blood volume (CBV) has been measured previously with inhaled 11CO and positron emission tomography (PET). The model used assumes that equilibrium in tracer concentration has occurred between arterial and systemic venous blood before the PET measurement is made. To verify that this model may be used with the much shorter half-lived C15O, we have simultaneously measured arterial and venous blood radioactivity following C15O inhalation. Equilibrium occurred 95 ± 39 s after inhalation (n = 7). If the PET measurement is commenced prior to arteriovenous equilibrium, significant errors occur in calculated CBV. These data indicate that C15O may be used as a tracer for CBV measurement provided that emission data collection commences at ∼120 s after inhalation. Strict quality control measures must be maintained to minimize the contamination of administered C15O with 15O-labeled CO2.
Cerebrocortical Microcirculation in Different Stages of Hypoxic Hypoxia
Authors: Kozniewska Ewa , Weller Lutz , Höper Jens , Harrison David K. , Kessler Manfred
Published: April 2011

This study investigated the relation between local cerebrocortical oxygen tension (PO2) and cerebrocortical microflow (CMF) during normoxia (Fio2 = 0.3) and hypoxic hypoxemia (Fio2 = 0.16 and 0.1). Measurements were performed on mechanically ventilated rats and rabbits anesthetized with 0.8% Ethrane and maintained within normocapnic limits. Polarographic techniques based on the use of multiwire surface electrodes were applied for measurements of local PO2 and CMF. In both species the mean tissue PO2 values were similar under normoxia (26.0 and 31.5 mm Hg for rats and rabbits, respectively). CMF histograms showed pronounced heterogeneity. The highest CMF values exceeded the lowest ones by a factor of 36 in the rat and by a factor of 26 in the rabbit. Mean CMF values were 6.67 ± 0.72 and 4.09 ± 0.14 relative units (for definition see text) in rats and rabbits, respectively. During hypoxemia, if the mean tissue Po2 was >5 mm Hg, mean CMF did not change but a change in the pattern of microflow distribution was observed with increases in some CMF values (up to 670% of control) and decreases in others (down to 12% of control). When mean tissue Po2 values of <5 mm Hg were observed during hypoxemia, mean CMF increased in both species by ∼50% on average. The increase in CMF could be seen in each individual CMF recording. We conclude that in the brain cortex local regulatory mechanisms are responsible for the change in the pattern of distribution of microcirculation during moderate tissue hypoxia. The same or similar mechanisms produce an increase in microflow if the local tissue Po2 falls below a critical level. Our experiments show that this critical level lies within the range of 0–5 mm Hg.
Thick Brain Slices Model the Ischemic Penumbra
Authors: Newman George C. , Hospod Frank E. , Wu Priscilla
Published: April 2011

Hypothalamic brain slices, varying in thickness from 400μ to 1,000μ, were assessed by studying 2-deoxyglucose (2DG) metabolism, lactate accumulation, inulin spaces, and morphology at the light and ultrastructural levels. Evidence of increased glycolytic flux due to anaerobic metabolism is found at thicknesses greater than 600μ in association with a progressive increase in the inulin-exclusion space. The metabolic profiles, as a function of depth into the slices, reveal that 700-μ slices function in a manner similar to 540-μ slices at the surfaces, but with a core of increased 2DG phosphorylation at the slice center. In contrast, the 1000-μ slices show significant reductions of 2DG and increases in 2DG6P relative to the 540-μ slices at the slice surface as well as in the slice interior, suggesting impaired transport of 2DG into cells and spread of ischemic injury from the slice interior to the slice surface. Despite these metabolic changes, only minor morphologic changes of ischemic injury were found at the center of thicker slices, and in vitro glucose utilization of 1000-μ slices remained constant for up to 15 h. These three slice thicknesses should provide a useful model for studying the neurochemistry and neuropharmacology of the ischemic penumbra.
Effects of Cerebral Ischemia and Reperfusion on Prostanoid Accumulation in Unanesthetized and Pentobarbital-Treated Gerbils
Authors: Dorman Robert V.
Published: April 2011

Cerebral ischemia was induced in unanesthetized gerbils using bilateral carotid artery ligations. The effects of 20 min of global ischemia on the concentrations of prostaglandin F2α (PGF2α), PGE2, 6-keto-prostaglandin F1α (6-keto-PGF1α), and thromboxane B2 were determined after 0–24 h of reperfusion. Ischemia had little effect on eicosanoid production, but significant increases were observed by 5 min of reperfusion, with maximal levels reached by 15 min of reperfusion. PGF2α was the most concentrated prostaglandin in postischemic brain, whereas PGE2 was most concentrated in control cerebra. Pretreatment with anesthetic doses of pentobarbital supported increased accumulation of PGF2α in postischemic cerebra, increased accumulation of 6-keto-PGF1α during the ischemic episode, and decreased accumulation of PGE2 at 120 min of reperfusion. It appears that the protective effects of barbiturate anesthesia are not expressed by the reduced accumulation of the above eicosanoids.
Quantitation of Photochemically Induced Focal Cerebral Ischemia in the Rat
Authors: Grome John J. , Gojowczyk Gerlinde , Hofmann Wolfgang , Graham David I.
Published: April 2011

This study was carried out with a recently developed model of focal cerebral ischemia in the rat based on the photochemical induction of thrombotic stroke using the dye Rose Bengal. We examined the change in the volume of the lesion and brain water content, in separate groups of rats, at different times (1, 4, 24, 72, and 168 h) after the induction of the ischemic lesion. The volume of ischemic damage increased rapidly between 1 and 24 h after the ischemic insult and decreased between 24 and 168 h. The lesion at 168 h was significantly larger than that following 1 h of ischemia and similar to that obtained at 4 h, suggesting that the maximum extent of tissue damage (without the involvement of significant edema) was reached within the first 4 h in this model. The enlargement of the lesion after 4 h correlated closely with changes in brain water content.
The Retention Mechanism of Technetium-99m-HM-PAO: Intracellular Reaction with Glutathione
Authors: Neirinckx Rudi D. , Burke James F. , Harrison Roger C. , Forster Alan M. , Andersen Allan R. , Lassen Niels A.
Published: April 2011

Preparations of d,l- and meso-hexamethylpropyleneamine oxime (HM-PAO) labeled with technetium-99m were added to rat brain homogenates diluted with phosphate buffer (l: 10). The conversion of d,l-HM-PAO to hydrophilic forms took place with an initial rate constant of 0.12 min−1. Incubation of the brain homogenate with 2% diethyl maleate for 5 h decreased the homogenate's measured glutathione (GSH) concentration from 160 to 16 μ M and decreased the conversion rate to 0.012 min−1. Buffered aqueous solutions of glutathione rapidly converted the HM-PAO tracers to hydrophilic forms having the same chromatographic characteristics as found in the brain homogenates. The rate constant for the conversion reaction of d,l-HM-PAO in GSH aqueous solution was 208 and 317 L/mol/min in two different assay systems and for meso-HM-PAO the values were 14.7 and 23.2 L/mol/min, respectively. Rat brain has a GSH concentration of about 2.3 m M and the conversion of the d,l-HM-PAO due to GSH alone should proceed with a rate constant of 0.48 to 0.73 min−1 and be correspondingly 14-fold slower for meso-HM-PAO. In human brain, the in vivo data of Lassen et al. show a conversion rate constant of 0.80 min−1. This correspondence of values supports the notion that GSH may be important for the in vivo conversion of 99mTc-labeled HM-PAO to hydrophilic forms and may be the mechanism of trapping in brain and other cells. A kinetic model for the trapping of d,l- and meso-HM-PAO in tissue is developed that is based on data of GSH concentration in various organs. This model predicts that the d,l form rapidly reaches a steady state in tissue and the tissue distribution reflects a pattern dominated by blood flow. For the meso form, the model predicts that steady state is reached more slowly and the tissue distribution reflects a pattern dominated by glutathione concentration.
Determination of Flow and Rate Constants in a Kinetic Model of [99mTc]-Hexamethyl-Propylene Amine Oxime in the Human Brain
Authors: Matsuda Hiroshi , Oba Hiroshi , Seki Hiroyasu , Higashi Sotaro , Sumiya Hisashi , Tsuji Shiro , Terada Hitoshi , Imai Keiko , Shiba Kazuhiro , Mori Hirofumi , Hisada Kinichi
Published: April 2011

The values for flow and rate constants for a kinetic model of [99mTc]–hexamethylpropyleneamine oxime (HM-PAO) distribution in the human brain were determined. The single-pass extraction ratio of HM-PAO was also determined in the rat brain by the indicator diffusion method; a value of 0.90 ± 0.02 (mean ± SEM, n = 5) was obtained. Time course data of brain activity and arterial blood activity of the tracer were fitted to a four compartment model: Values of blood flow and the firstorder rate constants for backdiffusion of the diffusible tracer from brain to blood (k2), conversion of the lipophilic tracer to the hydrophilic one in brain ( k3), and conversion of the diffusible tracer to the nondiffusible one in blood ( k5) were determined. Conversion of hydrophilic tracer back to a lipophilic form in both blood and brain was assumed to be negligible during the course of the experiment. The values obtained for blood flow, k2, and k3 were, respectively, 0.40 ± 0.03 ml/g/min, 0.38 ± 0.04 min−1, and 0.92 ± 0.05 min−1 in the gray matter ( n = 4), and 0.23 ± 0.01 ml/g/min, 0.17 ± 0.01 min−1, and 1.01 ± 0.05 min−1 in the white matter ( n = 2) in patients with cerebrovascular disorder. The k5 value was 1.14 ± 0.06 min−1 ( n = 4). These experimentally determined values agree well with the theoretical ones previously reported by Lassen et al. The results suggest the relative constancy of the k3 and k5 values and the more prominent initial backdiffusion of the lipophilic HM-PAO from brain to blood in high flow regions compared to low flow regions.
The Effect of the GABA-A Agonist THIP on Regional Cortical Blood Flow in Humans. A New Test of Hemispheric Dominance
Authors: Roland Per E. , Friberg Lars
Published: April 2011

We studied the effect of a γ-aminobutyric acid (GABA)–A receptor–induced postsynaptic inhibition on regional CBF (rCBF) in awake humans. For this purpose we used a new specific GABA–A agonist, 4,5,6,7-tetrahydroisoxazolo(5,4)-pyridin-3-ol (THIP). As part of a new diagnostic procedure for the determination of which hemisphere subserved language, THIP was infused into the internal carotid artery 20 s before measurement of the rCBF. Administered by this route the THIP is distributed to the neocortex and neostriatum. THIP induced a dosage-dependent decrease of the rCBF. The rCBF decrease was not due to any direct effect on the cerebral vessels. The efficiency of the THIP-induced postsynaptic inhibition was tested by having the subjects voluntarily activate the inhibited cortex. During submaximal inhibition the subjects were able voluntarily to counteract decreases of rCBF in superior frontal cortex and motor cortex. Larger doses of THIP abolished this response and depressed the rCBF to baseline levels (20 ml/100 g/min). This was associated with 10-min total depression of function of the anterior two-thirds of the injected hemisphere. An analysis of the changes of rCBF in activated and nonactivated cortex—with and without THIP-induced inhibition—showed that it would be very unlikely that average increases in synaptic inhibition would increase rCBF in neocortical areas. Intracarotid injection of the water-soluble, nonirritative THIP is a very useful alternative to sodium amytal injection for determination of hemispheric dominance.
The Effect of p-Chlorophenylalanine on Cerebral Metabolism and Biogenic Amine Content of Traumatized Brain
Authors: Pappius Hanna M. , Dadoun Ralph , McHugh Michael
Published: April 2011

It was shown previously that focal cortical freezing lesions in rats cause widespread decrease in local cerebral glucose utilization (LCGU) in cortical areas of the lesioned hemisphere. This was interpreted as reflecting a depression of cortical activity. It was then demonstrated that cortical serotonin (5-HT) metabolism was increased throughout the lesioned hemisphere of a focally injured brain. To find out if the changes in the serotonergic system are of functional importance and mediate the observed changes in LCGU, the effects of the inhibition of 5-HT synthesis with p-chlorophenylalanine (PCPA) on cerebral metabolism and biogenic amine content in injured brain were studied. PCPA in doses up to 300 mg/kg had little, if any, effect on LCGU in intact brain and in doses up to 100 mg/kg did not modify the depressed LCGU in injured brain. In doses of 200 and 300 mg/kg, PCPA selectively increased cortical glucose utilization in the lesioned hemisphere where it was depressed following injury. PCPA decreased 5-HT levels in cortical and raphe areas of both intact and injured brain in a dose-dependent manner. However, at doses of PCPA ineffective on LCGU (50 and 100 mg/kg), traumatization still resulted in increased 5-HT metabolism. Doses of PCPA that ameliorated the depression of LCGU in injured brain completely prevented increases in both 5-HT and its metabolite 5-hydroxyindoleacetic acid seen following traumatization in untreated animals. These results provide evidence that decreased LCGU in lesioned brain is due to an activation of the serotonergic system by traumatization. The data are in agreement with the postulated inhibitory role of serotonin in the cortex and its involvement in functional alterations associated with injury. They suggest that blockage of this neurotransmitter system may have a potential in the development of novel therapeutic approaches to brain injury.
Local Cerebral Glucose Utilization in Controlled Graded Levels of Hyperglycemia in the Conscious Rat
Authors: Orzi F. , Lucignani G. , Dow-Edwards D. , Namba H. , Nehlig A. , Patlak C. S. , Pettigrew K. , Schuier F. , Sokoloff L.
Published: April 2011

Local cerebral glucose utilization assayed by the [14C]deoxyglucose ([14C]DG) method and calculated by means of its operational equation with values for the rate constants and lumped constant determined in rats under physiological conditions remains relatively stable with variations in arterial plasma glucose concentration within the normoglycemic range. Large changes in arterial plasma glucose level may, however, significantly alter the values of these constants and lead to artifactual results. Values for the lumped constant have been measured and reported for a wide range of arterial plasma glucose concentrations ranging from hypoglycemia to hyperglycemia in the rat (Schuier et al., 1981; Suda et al., 1981; Pettigrew et al., 1983). In the present study we have redetermined the rate constants in rats with arterial plasma glucose levels clamped at ∼350, 450, and 550 mg/dl (i.e., 19, 25, and 31 m M) by a glucose clamp technique. The rate constants for the transport of DG from plasma to brain, K*1, and its phosphorylation in tissue, k*3, were found to decline with increasing plasma glucose levels, while the rate constant for its transport back from brain to plasma, k*2, remained relatively unchanged from its value in normoglycemia. These rate constants were used together with the previously determined values for the lumped constants to calculate local rates of cerebral glucose utilization in three groups of rats in which arterial plasma glucose levels were clamped at ∼350, 450, and 550 mg/dl (i.e., 19, 25, and 31 m M). Average glucose utilization in the brain as a whole was unchanged in hyperglycemia from the values calculated in normoglycemic rats with the standard normal set of constants. Changes in the rate of glucose utilization were found, however, in the hypothalamus, globus pallidus, and amygdala during hyperglycemia.
Measurement of End-Capillary Po2 with Positron Emission Tomography
Authors: Alpert N. M. , Buxton R. B. , Correia J. A. , Katz R. M. , Ackerman R. H.
Published: April 2011

The analysis of positron emission tomography measurements of oxygen metabolism has been extended to provide a quantitative estimate of end-capillary Po2. The principle of this extension rests on the idea that the oxygen extraction fraction can be used to calculate the end-capillary oxygen saturation of the blood. The relation between oxygen saturation and Po2 is obtained through the oxygen dissociation curve. Our studies show that in addition to the local oxygen extraction fraction, arterial Po2 and pH values are needed in the calculation, whereas fairly large variations in factors such as Pco2, hematocrit, hemoglobin, and plasma protein levels have little or no effect. Rough estimates of end-capillary Po2 can be made using standard o2 dissociation nomograms. Blood gas and acid-base properties of blood have been known for decades, making it possible to account accurately for individual differences that may be encountered when studying patients. Measurements in nine normal subjects yielded a mean end-capillary Po2 value of 31.2 mm Hg. The ability to make a quantitative visualization of altered patterns of end-capillary Po2 provides an additional dimension to the investigation of stroke disease and tumor metabolism.
Anatomical-Functional Correlation Using an Adjustable MRI-Based Region of Interest Atlas with Positron Emission Tomography
Authors: Evans A. C. , Beil C. , Marrett S. , Thompson C. J. , Hakim A.
Published: April 2011

A procedure is described for combining anatomical information from magnetic resonance imaging (MRI) or computerized tomography (CT) and functional information from positron emission tomography (PET) in a rapid fashion. MRI data are combined with a procedure for the definition, storage, and recall of anatomically based regions of interest. An atlas of standard regions of interest, defined for a set of 18 parallel planes spaced at 6-mm intervals, provides an initial region of interest template for each patient slice. Global adjustments to scale, orientation, and position are applied to obtain an initial match. Individual regions of interest may then be moved, deleted, or redrawn as needed. The ability to store region of interest templates ensures reproducibility of analysis over long periods and introduces a standardization of analysis technique. In 25 brain structures, the mean coefficient of variation in cerebral glucose utilization rate (CMRGlc) measurements among five neuroanatomically trained observers was reduced from 8.1% for manual region of interest definition to 4.0% using the template approach with MRI. Template analysis for space-occupying lesions such as tumors or infarcts is illustrated with PET data from a stroke study, emphasizing the facility for rapid, reproducible analysis of multifunctional studies. MRI-PET matching for a structurally intact caudate nucleus having reduced CMRGlc in Huntington's disease emphasizes the accuracy of anatomical localization required to quantify small structures.
A Double-Injection Technique for in vivo Measurement of Dopamine D2-Receptor Density in Monkeys with 3-(2'-[18F] Fluoroethyl)Spiperone and Dynamic Positron Emission Tomography
Authors: Huang Sung-Cheng , Bahn Mark M. , Barrio Jorge R. , Hoffman John M. , Satyamurthy Nagichettiar , Hawkins Randall A. , Mazziotta John C. , Phelps Michael E.
Published: April 2011

Dopamine D2-receptor density in striatum of monkey was measured with 3-(2'-[18F]fluoroethyl)spiperone (FESP) and dynamic positron emission tomography (PET), using a double-injection technique. A first bolus of high specific activity (SA) FESP (5 mCi; ≃ 1 Ci/μmol) was injected i.v.; 90 min later, a second bolus of lower SA FESP (5 mCi; ≃ 0.04 Ci/μmol) was injected. A dynamic PET study was performed to measure the kinetics of FESP in striatum over 180 min, and the metabolite-corrected concentration of FESP in plasma as a function of time was obtained from arterial blood samples. A nonlinear compartmental model that took into account the saturability of the receptor binding was used to describe the kinetics of FESP in striatum. Model parameters were estimated by regression with a constraint based on information about the equilibrium dissociation constant of the ligand–receptor binding. Dopamine D2-receptor density in striatum was estimated to be 25.9 ± 12.7 pmol/g in seven Macaca nemestrina monkeys. The method does not require the use of cerebellum as a reference tissue region and an estimate of dopamine D2-receptor density can be obtained from a single study.
Cytosolic Free Calcium, NAD/NADH Redox State and Hemodynamic Changes in the Cat Cortex during Severe Hypoglycemia
Authors: Uematsu Daisuke , Greenberg Joel H. , Reivich Martin , Karp Andrea
Published: April 2011

Summary Using indo-1, a fluorescent Ca2+ indicator, in vivo fluorometric measurements were made of changes in cytosolic free Ca2+, NAD/NADH redox state, and hemodynamics directly from the cat cortex during and after severe insulin-induced hypoglycemia. Cytosolic free Ca2+ started to increase when the EEG became isoelectric, remained at a significantly high level (p < 0.05) during the period of isoelectric EEG (IEEG), and recovered to the control level 6 min following an intravenous infusion of glucose. The NAD/NADH redox state oxidized significantly during IEEG and then recovered rapidly to the control level after the glucose infusion. Local cortical blood volume (LCBV) increased gradually during the progression of hypoglycemia, reaching the maximal level (146 ± 7%) at the end of IEEG, and then started to recover. The mean transit time (MTT) through the cortical microcirculation was shortened during the IEEG (control: 3.84 ± 0.41 s versus IEEG: 2.73 ± 0.17 s, p < 0.05), whereas it was prolonged during the 30-min recovery period (5.68 ± 0.58 s, p < 0.05). Local cortical blood flow calculated from the LCBV and MTT showed a twofold increase 5 min into IEEG (201 ± 27% of control, p < 0.05), recovered 15 min into the recovery period, and then decreased to 77% of control (p < 0.05) by 30 min. The data support the hypothesis that hypoglycemic brain damage might be mediated by an elevation of cytosolic free calcium.
Postischemic Administration of Idazoxan, an α-2 Adrenergic Receptor Antagonist, Decreases Neuronal Damage in the Rat Brain
Authors: Gustafson Ingvar , Miyauchi Yoshitoyo , Wieloch Tadeusz W.
Published: April 2011

The effect of an α-2 receptor antagonist, idazoxan, on ischemic neuronal damage in the hippocampus and neocortex was studied in rats following 10 min of forebrain ischemia. Idazoxan was given 0.1 mg/kg i. v. immediately after recirculation, followed by 48 h of continuous infusion at a rate of 10 μg/kg/min. A histopathological examination of the CA1 region of the dorsal hippocampus and neocortex from each hemisphere was made on paraffin-embedded sections following 7 days of survival. In ischemic animals receiving an infusion of saline, 71% of the neurons in the hippocampal CA1 region were degenerated. In contrast, in the idazoxan-treated animals only 31% of the neurons were irreversibly damaged (p < 0.01). We conclude that postischemic administration of the α-2 antagonist idazoxan protects neurons against damage following cerebral ischemia. Rapid postischemic administration of α-2 adrenergic receptor antagonists could be an effective treatment after stroke and cardiac arrest.
CNS Stimulation and PGE2 Release. III. Pentamethylenetetrazole-Induced Seizures
Authors: Navarro E. , Romero S. D. , Yaksh T. L.
Published: April 2011

Measurement of prostaglandin E2 (PGE2) in the ventriculocisternal perfusate of the halothane anesthetized, artificially ventilated cat has revealed low but measurable levels of the prostanoid (64 ± 5 pg/min). Administration of pentamethylenetetrazole (PTZ) resulted in a rapid appearance of paroxysmal bursting, the magnitude and duration of which was dose dependent. During the 30-min interval after seizure initiation, PGE2 secretion rates into the ventriculocisternal perfusate rose by five- to sevenfold. Though the initial rate of PGE2 secretion correlated closely with the initial magnitude of bursting, there were significant differences, viz. the time courses. Thus, after a low dose of PTZ (200 mg/kg) the increase and return to normal of PGE2 secretion was time locked with the onset and offset of seizures. In contrast, after high doses of PTZ (250 mg/kg i. v.), seizure activity returned to near baseline by 90 min, while the levels of PGE2 secretion remained elevated for periods in excess of 150 min. Pretreatment with clonazepam (CLP: 3 mg/kg i. v. infusion) blocked seizures otherwise induced by PTZ (250 mg/kg) and the increase in PGE2 secretion. CLP administration 60 min after the initiation of seizures, blocked further seizure activity but did not alter the elevated secretion of PGE2. We thus believe these data jointly support the hypothesis that under intense paroxysmal bursting there is a change in neuronal state such that large stores of free fatty acids are available either because they have accumulated during the seizure because of a continued Ca2+ influx or the presence of large and continuing concentrations of Ca2+ accumulating in the cytosol secondary to energy failure.