Identification of a saturable uptake system for deoxyribonucleosides at the blood-brain and blood-cerebrospinal fluid barriers

Substances can enter the brain either directly across the blood-brain barrier or indirectly across the choroid plexuses and arachnoid membrane (blood-CSF barrier) into the CSF and then by diffusion into the brain. Earlier studies have demonstrated a saturable thymidine uptake across the blood-CSF barrier, but not across the blood-brain barrier. In this study transport of [³H]thymidine across both barriers was measured in vivo by means of a bilateral vascular brain perfusion technique in the anaesthetised guinea-pig. This method allows simultaneous and quantitative measurement of slowly penetrating solutes into both brain and CSF, under controlled conditions of arterial inflow. The results of the present study carried out over perfusion periods of up to 30 min indicated a progressive uptake of [³H]thymidine into brain and CSF, which was found to be significantly greater than the transport ofd-[¹⁴C]mannitol (a plasma space marker). Furthermore, the addition of 1 mM unlabelled thymidine in the perfusate caused saturation of [³H]thymidine uptake into both brain and CSF. In conclusion, these findings suggest that thymidine can cross both the blood-brain and blood-CSF barriers in the guinea-pig by carrier-mediated transport systems.     

The inulin space of the lamprey spinal cord

The distribution of [¹⁴C]inulin was measured in isolated spinal cords of larval and feeding stage adult forms of sea lamprey (Petromyzon marinus) and expressed in per cent of total cord wet weight. In larval cord the apparent inulin space reached a plateau value of 32–33% within 2.5 min. This correlates well with electrophysiological experiments in which 10⁻⁷ M tetrodotoxin added to the perfusion fluid blocked the responses of giant interneurons to both intracellular and rostral cord stimulation in 1 to 2 min. Thus the plateau level of inulin space probably represents the extracellular space.[¹⁴C]Mannitol did not reach a steady distribution space even after 30 min of incubation. Therefore, mannitol is not an accurate extracellular space indicator in the isolated lamprey spinal cord.The inulin space increased with increasing temperature of incubation. Average inulin spaces for larval spinal cords incubated at 5, 10 and 22 °C were approximately 26%, 33% and 42% respectively. The inulin space of isolated adult lamprey spinal cords was about 18–19%. Since in larvae the inulin space did not vary consistently with the sizes (and therefore presumably the ages) of the animals, it is likely that the reduction in inulin space during maturation does not occur gradually during the larval phase, but probably occurs during transformation. The difference between the inulin spaces of isolated larval and adult spinal cords is reflected qualitatively in the electron microscopic appearance of the extracellular space.We conclude that the inulin space in the lamprey spinal cord behaves similarly to the picture of the mammalian brain extracellular space which has emerged in recent years. Because of the rapidity of inulin diffusion in the lamprey cord and the unambiguous time-dependent behavior of the inulin space of the isolated lamprey cord, the latter would seem to be a useful model for the extracellular space of the vertebrate central nervous system.     

Passage of MHPG from plasma to CSF in a non-human primate

In experimental protocols with humans and non-human primates, cerebrospinal fluid (CSF) concentrations of 3-methoxy-4-hydroxyphenylethylene glycol (MHPG), the predominant end-product of norepinephrine metabolism in the mammalian central nervous system (CNS), have been widely used as an index of the rate of CNS norepinephrine metabolism. However, an earlier investigation showed that there was slow but free exchange between plasma and CSF MHPG. To define more precisely the time-course of equilibration of plasma and CSF MHPG, we intravenously administered 100 micrograms/kg of [2H3]-MHPG to drug-naive squirrel monkeys. Measurements were made of the concentrations of [2H3]- and [1H]-MHPG in plasma and cervical CSF samples collected at time points from 10 min to 4 hr thereafter. The results indicated that neither plasma nor CSF concentrations of [1H]-MHPG changed during the course of the experiment, and that [2H3]-MHPG appeared in the CSF within 10 min of intravenous administration. The maximal plasma and CSF concentrations of [2H3]-MHPG were 7.6- and 2.3-fold higher than the respective concentrations of [1H]-MHPG. The plasma and CSF pools of [2H3]-MHPG reached concentration equilibrium within 30 min, and thereafter the temporal decline in concentration of [2H3]-MHPG was the same in plasma and CSF. These results demonstrate that MHPG rapidly crosses from plasma to CSF, and support the suggestion that this factor be included in any attempts to estimate norepinephrine turnover in the CNS from measurements of steady-state MHPG concentrations in CSF or plasma.     

Diffusion-weighted imaging in noncompressive myelopathies: a 33-patient prospective study

Diffusion-weighted imaging (DWI) is frequently used to differentiate cerebral lesions. The aim of our study was to evaluate the diagnostic value of DWI and the measurement of the apparent diffusion coefficient (ADC) in noncompressive myelopathy explorations. Thirty-three patients presenting a spinal cord syndrome due to a noncompressive myelopathy underwent spinal cord MRI between September 2005 and November 2008. For each patient, the ADC was calculated in the pathological spinal cord. ADC values were also measured in the healthy spinal cord of ten control subjects. Statistical analysis was based on the Student’s t test. Twenty-one patients presented an inflammatory myelopathy: Nine patients presented multiple sclerosis, three patients presented a parainfectious myelopathy, two patients acute disseminated encephalomyelitis, one patient neuromyelitis optica, one patient systemic lupus erythematosus, and five patients a myelopathy of unknown aetiology. Six patients presented a spinal cord infarction. ADC values were significantly lower in spinal cord infarct (mean ADC = 0.81 ± 0.08 × 10⁻³ mm²/s) than in inflammatory spinal cord lesions (mean ADC = 1.37 ± 0.23 × 10⁻³ mm²/s) and in healthy control spinal cord (mean ADC = 0.93 ± 0.07 × 10⁻³ mm²/s). These results are important to differentiate ischaemic from inflammatory myelopathies, especially at the acute phase when clinical presentation and extensive work-up are not able to show an aetiologic diagnosis. Although these results are similar to those described in cerebral explorations, ADC measurements remain technically limited for the moment.     

Acute treatment with pulsed electromagnetic fields and its effect on fast axonal transport in normal and regenerating nerve

The mechanism whereby low-frequency electromagnetic fields accelerate axonal regrowth and regeneration of peripheral nerve after crush lesion is not known. One candidate is an alteration in axonal transport. In this study we exposed unoperated rats for 15 min/day, and rats that had undergone a crush lesion of the sciatic nerve, for 1 hr/day for 2 days, to 2-Hz pulsed electromagnetic fields. To label fast transported proteins, [³H]-proline was microinjected into the spinal cord, and the sciatic nerves were removed 2, 3.5, and 5 hr later. The rates of fast axonal transport were obtained for animals in all groups by counting sequential 2-mm segments of nerves. The following transport rates were found: in unoperated normal sciatic nerve not exposed to PEMF, 373 ± 14 mm/day; in unoperated normal nerve exposed to PEMF, 383 ± 14 mm/day; in sham crush nerves not exposed to PEMF, 379 ± 19 mm/day; in sham crush nerve exposed to PEMF, 385 ± 17 mm/day; in crushed nerves not exposed to PEMF, 393 ± 16 mm/ day; and in crushed nerves exposed to PEMF, 392 ± 15 mm/day. The results of these experiments indicate that (1) a crush injury to the sciatic nerve does not alter the rate of fast axonal transport, and (2) low-frequency pulsed electromagnetic fields do not alter fast axonal transport rates in operated (crush) or unoperated sciatic nerves. © 1995 Wiley-Liss, Inc.     

Intracellular transport and neuronal activation of phospholipid and glycoprotein synthesis during axonal regeneration of cranio-spinal nerves

In the present work, the hypothesis that the increased rapid intracellular transport of newly-synthesized material along the axons of a regenerating system is sustained by an alteration of the transport of proteolipid complexes through subcellular compartments of a neuronal cell body was tested by a biochemical methodology. The motoneurons of spinal cord ventral horn, 4 wk after unilateral lesion (crush) of cervico-thoracic nerves of the rabbit at the level of brachial plexus, were chosen as the model system of regeneration. A time-staggered procedure of in vivo and in vitro double labeling with metabolic precursors, such as [³H]-choline, [¹⁴C]-choline, [³H]-fucose, and [¹⁴C]-fucose, was used. Subcellular fractions (RER, SER, Golgi apparatus, and plasma membranes) of ventral horn tissue, taken from spinal cord hemisections (regenerating and contralateral side), were further isolated. Twenty-eight days after axotomy, we did not observe any change of intracellular transport kinetics (¹⁴C/³H ratio) of newly-synthesized choline-phospholipids and glycoproteins in regenerating motoneurons compared to controls. However, associated with regenerating phenomenon in Golgi apparatus, we observed an increase of labeled choline-phospholipid and glycoprotein material that could contribute to the increased fast axonal transport and delivery of membrane proteolipid complexes to plasma membrane and axonal compartments. The increase of glycoprotein labeling was more pronounced in the SER portion (vesicles and elements of smooth membranes). This result is in favor of the hypothesis that membrane-bound proteins are transported from the Golgi to the axon through the perikaryal SER.     

Measurement of tritiated norepinephrine metabolism in intact rat brain

A procedure for the study of NE metabolism in the intact rat brain is described. The method involves ventriculocisternal perfusion of the adult male rat with artificial CSF containing [³H]NE. Radioactivity in the perfusate associated with NE and its metabolites 3,4-dihydroxymandelic acid (DOMA), 3,4-dihydroxyphenylethyleneglycol (DHPG), 3-methoxy-4-hydroxymandelic acid (VMA), 3-methoxy-4-hydroxyphenylethyleneglycol (MHPG), and normetanephrine (NMN) is separated using high-performance liquid chromatography (HPLC). After 80 min the radioactivity in the perfusate reaches an apparent steady-state. Analysis of the steady-state samples shows higher activity in the fractions corresponding to DHPG and MHPG than in those corresponding to DOMA and VMA, confirming glycol formation as the major pathway of NE metabolism in rat brain. Pretreatment with an MAO inhibitor (tranylcypromine) results in a marked decrease in the deaminated metabolites DHPG and MHPG and a concurrent increase in NMN. The results indicate this to be a sensitive procedure for the in vivo determination of changes in NE metabolism.     

Melatonin from Cerebrospinal Fluid but Not from Blood Reaches Sheep Cerebral Tissues Under Physiological Conditions

The pineal gland secretes melatonin (MLT) that circulates in the blood and cerebrospinal fluid (CSF). We provide data to support the hypothesis that, in sheep and possibly in humans, only the CSF MLT, and not the blood MLT, can provide most of MLT to the cerebral tissue in high concentrations, particularly in the periventricular area. The MLT content of sheep brain, our chosen animal model, was found in significant concentration gradients oriented from the ventricle (close to the CSF) to the cerebral tissue, with concentrations varying by a factor of 1–125. The highest concentrations were observed close to the ventricle wall, whereas the lowest concentrations were furthest from the ventricles (407.0 ± 71.5 pg/ml compared to 84.7 ± 5.2 pg/ml around the third ventricle). This concentration gradient was measured in brain tissue collected at mid‐day and at the end of the night. Nocturnal concentrations were higher than daytime concentrations, reflecting the diurnal variation in the pineal gland. The concentration gradient was not detected when MLT was delivered to the brain via the bloodstream. The diffusion of MLT to cerebral tissues via CSF was supported by in vivo scintigraphy and autoradiography. 2‐[¹²³I]‐MLT infused into the CSF quickly and efficiently diffused into the brain tissues, whereas [¹²³I]‐iodine (control) was mostly washed away by the CSF flow and [¹²³I]‐bovine serum albumin remained mostly in the CSF. Taken together, these data support a critical role of CSF in providing the brain with MLT.     

Gastrointestinal motor activity associated with postoperative ileus and emesis

The gastrointestinal motor activity associated with post-operative ileus and emesis has not been fully elucidated. This study has evaluated gastric and small-bowel motility in six patients before and after cholecystectomy and in six healthy volunteers, by solid-state manometry. Nausea and vomiting were recorded post-operatively. After surgery, fasting motor abnormalities including (a) total gastric quiescence and (b) small-bowel ‘phasic-bursts’ of contractions were observed in all patients. Phasic bursts (PB) resembled phase III of the migrating motor complex (MMC) on initial visual inspection, but further analysis revealed that they were of shorter duration (3.4 ±.2 min [PB] vs 6.4 ± 0.8 min [MMC], [mean ± SEM] P < 0.01), lower contraction frequency (6.4 ± 0.1 contractions min ^?1 [PBj vs 10 ± 0.3 contractions min^?1 [MMC] [mean ± SEM] P < 0.01) and shorter periodicity (36.4 ± 3 min [PB] vs 70.0 ± 6 min [MMC] [mean ± SEM] P < 0.01). Four patients experienced nausea during phasic burst activity. Vomiting was only observed in association with retrograde phasic-bursts, which migrated through the duodenum to the stomach. This study has shown consistent gastrointestinal motor abnormalities in the immediate post-operative state.     

Disposition and metabolism of MHPG in humans: application to studies in depression

MHPG is formed from norepinephrine metabolized throughout the body; its levels in plasma reflect total norepinephrine metabolism. Over half of the MHPG, is converted to VMA. Less than 20% of MHPG is derived from brain norepinephrine and urinary excretion of this metabolite cannot be used as a measure of brain norepinephrine metabolism. Because unconjugated MHPG is readily diffusable, there is a free exchange of this metabolite among plasma, cerebrospinal fluid, and nerve tissues (including brain and spinal cord). The CSF MHPG levels, after appropriately correction for the plasma contribution of the metabolite, reflect its rate of formation in the central nervous system.     

Uptake,release and metabolism of alaume in neurons and astrocytes in primary cultures

The uptake, release, and metabolism of alanine were studied in primary cultures of cerebral cortical neurons or astrocytes and cerebellar granule neurons. All three cell types exhibited a saturable, sodiumdependent uptake of alanine with K_M values (ULM) of 256 ± 30, 463 ± 39, and 292 ± 39, respectively, and V values (nmol/min/mg) of 15.9 ± 0.7,7.9 ± 0.01, and 17.4 ± 0.8, respectively. The corresponding val' ues < nmol/min/mg for the specific activity of alanine aminotransferase were 4.7 ± 0.4, 17.1 ± 2.5, and 4.5 ± 0.9 (all values represent the mean ± SEM). Release of alanine from the cells was rectilinear with time over a 10 hr period in case of astrocytes (40 nmol/hr/ mg) and cerebellar granule neurons (21 nmol/hr/mg). In cortical neurons the release rate declined from an initial value of 19 nmol/hr/mg during the first 3 hr to a value of less than 3 nmol/hr/mg during the subsequent 7 hr of incubation. Metabolism of [¹⁴C]alanine to ¹⁴CO₂ was found to have a lag period of 15 min and subsequently the rate of CO₂ production was constant over a 45 min period with a value of 0.5 nmol/min/mg in granule neurons and about 0.3 nmol/ min/mg in the other two cell types. Altogether the results show that alanine is preferentially produced in and released from astrocytes and accumulated into both GABAergic cortical neurons and gintamatergic cerebellar granule neurons. © 1993 Wiley-Liss, Inc.     

A precise and sensitive method for measurement of spinal cord blood flow

Spinal cord blood flow can be accurately and rapidly measured in rabbits by intravenous injection of [¹⁴C]iodoantipyrine, collection of sequential arterial samples, and abrupt termination of blood flow to the spinal cord by severing the aorta with a previously implanted ligature. The radioactive tracer concentrations in arterial and spinal cord samples can be rapidly measured in a scintillation counter. The spinal cord blood flow can then be calculated from these blood and tissue tracer concentrations. This method is suitable for use in anesthetized or unanesthetized small animals.     

Effect of acute administration of morphine on newly synthesized 5-hydroxytryptamine in spinal cord of the rat

The effect of morphine (10 mg/kg, s.c.) on the rate of [³H]5-HT synthesis in brain and spinal cord following intravenous injection of [³H]tryptophan was studied in the rat.

(1) In the spinal cord morphine induced an increase in [³H]TRP uptake which was significant 30 and 60 min after the injection, and a clear enhancement of the formation of [³H]5-HT which was significant 15 min after the injection and which peaked at 30 min.

In the forebrain, the increase in [³H]TRP accumulation was significant as early as 15 min after morphine. At this time, the rate of 5-HT synthesis was not modified, but it was significantly increased 30 and 60 min after the injection.

(2) Increases in [³H]5-HT synthesis rate were suppressed by naloxone (1 mg/kg, i.m.). However this narcotic antagonist did not significantly reduce the increased accumulation of [³H]TRP in brain and spinal cord due to morphine treatment.

These results demonstrate that morphine induces a fast and marked increase in 5-HT synthesis, and suggest that this increase is only partly related to an increase in the availability of tryptophan to the central nervous system.These results are in good agreement with recent investigations showing the involvement of the raphe-spinal system in morphine analgesia.     

Long-Term Changes in Thecal Sac Compression and Decreased Cerebrospinal Fluid Space Following Paddle Lead Spinal Cord Stimulation at T9: A Long-Term Follow-Up via Three-Dimensional Myelographic Computed Tomography

ObjectivesTo investigate the long-term changes in thecal sac compression following T9 paddle lead spinal cord stimulation (SCS) using three-dimensional myelographic computed tomography (CT).Materials and MethodsSeventeen patients with five-column paddle lead SCS at T9 underwent three-dimensional myelographic CT scans preoperatively, immediately after surgery, and after an average of 11 months. The cross-sectional areas of thecal sac and spinal cord and the widths of anterior and posterior cerebrospinal fluid (CSF) spaces were repeatedly measured and compared. The contact angle of the lead with long-term pain relief was assessed.ResultsThe cross-sectional areas of thecal sac and spinal cord decreased significantly after lead placement (30.47 ± 9.21% and 4.71 ± 9.84%, respectively). Even after 11 months, a significant reduction was found with the preoperative values (17.97 ± 12.32% and 2.88 ± 7.09%). The widths of anterior and posterior CSF spaces decreased significantly after surgery (43.53 ± 13.17% and 57.13 ± 13.17%, respectively) and the severe decrease persisted long-term (29.13 ± 21.54% and 50.99 ± 16.07%). The average pain relief was 42.27 ± 17.50% with no correlation between the rate of reduction in cross-sectional areas of thecal sac and the widths of CSF spaces.ConclusionsSignificant early reduction and late partial restoration occurred in the thecal sac and spinal cord and the width of the anterior and posterior CSF spaces in the T9 5-column paddle lead SCS. Thecal sac compromise was expected to some extent after paddle lead implantation, but the degree is significant, and the cross-sectional area of the spinal cord as well as the thecal sac is affected. Fortunately, these anatomical changes did not cause any clinical problems except for intercostal root irritation. The shape and flat contours of the five-column paddle leads clearly affected the results.     

Neuropeptide receptors in developing and adult rat spinal cord: An in vitro quantitative autoradiography study of calcitonin gene-related peptide,neurokinins, μ-opioid,galanin, somatostatin,neurotensin and vasoactive intestinal polypeptide receptors

A number of neuroactive peptides including calcitonin gene-related peptide (CGRP), substance P, neurokinin B, opioids, somatostatin (SRIF), galanin, neurotensin and vasoactive intestinal polypeptide (VIP) have been localized in adult rat spinal cord and are considered to participate either directly and/or indirectly in the processing of sensory, motor and autonomic functions. Most of these peptides appear early during development, leading to the suggestion that peptides, in addition to their neurotransmitter/neuromodulator roles, may possibly be involved in the normal growth and maturation of the spinal cord. To provide an anatomical substrate for a better understanding of the possible roles of peptides in the ontogenic development of the cord, we investigated the topographical profile as well as variation in densities of [¹²⁵I]hCGRPα, [¹²⁵I]substance P/neurokinin-1 (NK-1), [¹²⁵I]eledoisin/neurokinin-3 (NK-3), [¹²⁵I]FK 33–824 ([D-Ala², Me-Phe⁴, Met(O)ol⁵]enkephalin)/μ-opioid, [¹²⁵I]galanin, [¹²⁵I]T₀D₈-SRIF₁₄ (an analog of sornatostatin), [¹²⁵I]neurotensin and [¹²⁵I]VIP binding sites in postnatal and adult rat spinal cord using in vitro quantitative receptor autoradiography. Receptor binding sites recognized by each radioligand are found to be distributed widely during early stages of postnatal development and then to undergo selective modification to attain their adult profile of distribution during the third week of postnatal development. The apparent density of various receptor sites, however, are differently regulated depending on the lamina and the stage of development studied. For example, the density of μ-opioid binding sites, following a peak at postnatal day 4 (N), declines gradually in almost all regions of the spinal cord with the increasing age of the animal [¹²⁵I]substance P/NK-1 binding sites, on the other hand, show very little variation until P14 and then subsequently decrease as the development proceeds. In the adult rat, most of these peptide receptor binding sites are localized in relatively high amounts in the superficial laminae of the dorsal horn. To varying extents, moderate to low density of various peptide receptor binding sites are also found to be present in the ventral horn, intermediolateral cell column and around the central canal. Taken together, these results suggest that each receptor-ligand system is regulated differently during development and may each uniquely be involved in cellular growth, differentiation and in maturation of the normal neural circuits of the spinal cord, Furthermore, the selective localization of various receptor binding sites in adult rat spinal cord over a wide variety of functionally distinct regions reinforces the neurotransmitter/ modulator roles of these peptides in sensory, motor and autonomic functions associated with the spinal cord. © 1995 Wiley-Liss, Inc.     

Comparison of radio-iodinated serum albumin and blue dextran as indicators to measure rate of formation of cerebrospinal fluid.

The characteristics of two nondiffusible indicators, ¹²⁵I-labeled albumin (RISA-125) and blue dextran, were compared by using them simultaneously to measure rate of formation of cerebrospinal fluid (CSF) in in vitro experiments and in a series of ventriculocisternal perfusion experiments in rhesus monkeys. Mean CSF formation calculated from the dilution of RISA-125 was 37.6 μl/min (±0.74 sem); the CSF formation calculated from the corresponding dilution of blue dextran was 37.1 μl/min (±0.70 sem). The difference is not statistically significant. As measured by counts per minute, the concentration of RISA-125 in aliquots of a dilute RISA-125 solution varied considerably. In the in vitro experiments, simulated CSF formation calculated from these counts varied around the mean by more than ± 10% at the 95% confidence limits. Counting variability is probably related to the low energy levels of the ¹²⁵I isotope. Cerebrospinal fluid formation calculated from the dilution of blue dextran had one-tenth the coefficient of variation of that calculated from the dilution of RISA-125. We have concluded that blue dextran is preferable to RISA-125 to measure the rate of formation of cerebrospinal fluid in the experimental setting because it is easier to detect accurately small changes of CSF formation with blue dextran.     

Axonal transport of polyamines: a reappraisal.

Axonal transport of putrescine and the polyamines in normal and transected motor axons of adult rat sciatic nerves was studied. [³H]Putrescine injected into the ventral horn region of the spinal cord was not transported distally along the course of the nerve as a function of time. l-[³H]ornithine, the immediate precursor of putrescine, when injected into the ventral horn of the spinal cord, was followed by rapid axonal transport of acid-insoluble material, but no transport of acid-soluble material, such as putrescine or the polyamines, was detected despite the demonstration that radioactive spermidine was synthesized at the site of the l-[³H]ornithine injection in the spinal cord. Furthermore, acid-soluble radioactive material failed to accumulate proximal to the site of a ligature of the sciatic nerve in acute and chronic experiments. Ligation of the sciatic nerve also failed to cause an accumulation of endogenous putrescine proximal to the ligature. The data showed no evidence for fast axonal transport of putrescine and the polyamines in normal and transected motor axons of the sciatic nerve in adult rats.     

Monoamine metabolites in lumbar CSF: the question of their origin in relation to clinical studies

The sources of homovanillic acid (HVA), 3-methoxy-4-hydroxyphenylethylene glycol (MHPG) and 5-hydroxyindoleacetic acid (5HIAA) in lumbar CSF of man are discussed. Although HVA in lumbar CSF is derived from the brain, and MHPG seems to be mainly from the spinal cord, the origin of 5HIAA is uncertain. The evidence for and against a brain contribution to 5HIAA of the lumbar CSF is evaluated. It is concluded that the brain can contribute 5HIAA to lumbar CSF under some circumstances. Measurement of 5HIAA concentrations in lumbar CSF can be of clinical value in detecting changes of 5-hydroxytryptamine (5HT) metabolism in the CNS if the changes are general throughout the CNS.     

Time course of IL-6 expression in experimental CNS ischemia

Abstract

lnterleukin-6 (IL-6) appears to be an important modulator of the inflammatory response associated with CNS ischemia. Clinically, IL-6 values obtained in the first week post-stroke have been shown to correlate with infarct size and outcome. In this study we used a focal reversible stroke model to investigate the time course and relationship to outcome of IL-6 production in plasma, brain and CSF. Reversible middle cerebral artery occlusion or sham surgery was produced in 50 adult Swiss Webster mice by advancing an 8-0 filament into the internal carotid artery for 2 h (sham 1 min). At 3, 6, 12, 24, and 72 h (8 each ischemia; 2 each sham) groups of animals were evaluated on a 28 point clinical scale, blood and CSF obtainedand the brains were evaluated for infarct volume and IL-6 mRNA levels. Serum levels of IL-6 (ELISA mean ± SD; undetectable in controls) overall sham group, 102 ±87; 3 h, 908 ±494*pg ml^–1; 6 h, 1079±468*pgml^–1; 12h, 980±221 * pg ml^–1; 24 h, 320± 314*pgml^–1; 72 h, 20±30*pg ml^–1 (*p < 0.05 to sham). CSF levels (ELISA) overall sham group, 10±18; 3 h, 379±210*pgml^–1; 6 h, 157±61*pg ml^–1; 12 h, 136± 88*pgml^–1; 24 h, 127± 99 pg ml^–1; 72 h, 72±9* pg ml^–1 (*p < 0.05 to sham). Brain IL-6 mRNA levels overall sham group, 20; 3 h, 480; 6 h, 599; 12 h, 7960; 24 h, 20267; 72 h, 0. There was an overall R² of 0.20 between plasma and CSF IL-6. There was an overall R² of 0.13 and 0.20 between infarct size and serum and CSF IL-6 level respectively, and an overall R2 of 0.10 and 0.17 between neurologic function and serum and CSF IL-6 level respectively. These findings confirm that IL-6 values increase following CNS ischemia with peak serum and CSF levels occurring before brain values. CSF IL-6 levels had a stronger correlation with neurologic function and infarct size than serum. [Neurol Res 1999; 21: 287-292]     

Comparison of binding affinities and capacities of gamma-aminobutyric acid and glycine in synaptosome-enriched fractions of rat cerebral cortex and spinal cord.

The “binding” affinities and capacities of [³H]-γ-aminobutyric acid (³H-GABA) and [¹⁴C]-glycine to synaptosome-enriched fractions of rat cerebral cortex and spinal cord have been compared over a 10^?8 to 10^?3m concentration range in isosmotic sucrose solutions containing 32 mm NaCl. At 0 C, the binding of both GABA and glycine to particles of the cerebral cortex appeared to take place by “triple-affinity” processes. The respective binding affinities (reciprocals of K_B values) of GABA for particles of the cortex were generally greater than those for glycine, whereas maximal binding capacities (B_max values) for GABA were about twice as great as those for glycine. In fractions of spinal cord, at 0 C, the binding of both GABA and glycine occurred by “double-affinity” processes, and though the binding affinities of GABA were about twice those of glycine, B_max values for GABA were smaller than those for glycine. Striking changes occurred when particles were allowed to stand in contact with [³H]-GABA plus [¹⁴C]-glycine at 23 C for 45 min. Although double-affinity mechanisms persisted at the higher temperature for the binding of both amino acids to particles of spinal cord (GABA still being bound with greater affinities than glycine), the binding of both amino acids to particles of cerebral cortex was best resolved as “single-affinity” processes. The differences between B_max values for GABA and glycine in particles of cerebral cortex and spinal cord appeared to reflect their different endogenous concentrations.