Anders Hamberger

Ischemia-Induced Shift of Inhibitory and Excitatory Amino Acids from Intra- to Extracellular Compartments

Brain ischemia was induced for 10 or 30 min by clamping the common carotid arteries in rabbits whose vertebral arteries had previously been electrocauterized. EEG and tissue content of high energy phosphates were used to verify the ischemic state and to evaluate the degree of postischemic recovery. Extracellular levels and total contents of amino acids were followed in the hippocampus during ischemia and 4 h of recirculation. At the end of a 30-min ischemic period, GABA had increased 250 times, glutamate 160 times, and aspartate and taurine 30 times in the extracellular phase. The levels returned to normal within 30 min of reflow. A delayed increase of extracellular phosphoethanolamine and ethanolamine peaked after 1–2 h of reflow. Ten minutes of ischemia elicited considerably smaller but similar effects. With respect to total amino acids in the hippocampus, glutamate and aspartate decreased to 30–50% of control while GABA appeared unaffected after 4 h of reflow. Alanine, valine, phenylalanine, leucine, and isoleucine increased severalfold. The importance of toxic extracellular levels of excitatory amino acids, as well as of high extracellular levels of inhibitory amino acids, are considered in relation to the pathophysiology of neuronal cell loss during cerebral ischemia.

Mass transfer in brain dialysis devices—a new method for the estimation of extracellular amino acids concentration

A simple mathematical description of mass transfer in brain dialysis devices is presented. The validity of this model is supported by results from both in vitro and in vivo dialysis experiments. Based on this description, an alternative approach for estimating the extracellular concentration of amino acids is outlined.

Effects of vagus nerve stimulation on amino acids and other metabolites in the CSF of patients with partial seizures

Electrical stimulation of the vagus nerve (VNS) is a new method for the treatment of patients with medically intractable epilepsy. Sixteen patients, ten of whom participated in a larger multicenter double-blind trial on the efficacy of VNS in epilepsy, and six who participated in pilot studies, consented to participate in the present study. Ten patients received HIGH stimulation and six patients LOW stimulation for the 3-month trial. Cerebrospinal fluid (CSF) samples (16 ml) were collected both before and after 3 months of VNS. Amino acid and neurotransmitter metabolites were analyzed. Four patients responded to VS with more than a 25% seizure reduction after 3 months. Mean and median concentrations of phosphoethanolamine (PEA) increased in responders and decreased in nonresponders. Free GABA increased in both groups but more so in the nonresponders. After 9 months of VS (6-9 months on HIGH stimulation) 4 of 15 patients had more than 40% seizure reduction. There were significant correlations between seizure reduction and increases in asparagine, phenylalanine, PEA, alanine and tryptophan concentrations. Comparison between patients with HIGH or LOW stimulation showed a significant increase in ethanolamine (EA) in the HIGH group and a decrease in glutamine in the LOW group. All patients regardless of response or stimulation intensity showed significantly increased total and free GABA levels. A decrease in CSF aspartate was marginally significant. Other trends were decreases in glutamate and increases in 5-hydroxyindoleacetic acid. Chronic VNS appears to have an effect on various amino acids pools in the brain.(ABSTRACT TRUNCATED AT 250 WORDS)

Glial cell function: active control of extracellular K + concentration

Abstract To investigate the ability of the glial cell to control the extracellular K+ ion concentration in the CNS 3 kinds of experiment were carried out. (1) The ability of isolated cell fractions enriched in glia or neurons to reaccumulate K+ was measured. (2) The activation of the Na+ K+-ATPase by K+ was measured on these isolated cell suspensions. (3) Plasma membrane fractions were prepared from neurons and glia and the Na+ K+-ATPase activity was studied as a function of K+ ion concentration. The glial cell Na+ K+-ATPase was always more active, approximately 2.5- to 4-fold, and markedly more sensitive to variations in the K+ ion concentration than the neuronal ATPase. These results support the idea that control of the extracellular K+ ion concentration in the CNS is at least in part dependent on the metabolically coupled sodium-potassium pump of the glial cell.

Effects of In Vivo Administration of Kainic Acid on the Extracellular Amino Acid Pool in the Rabbit Hippocampus

Abstract: The effect of local administration of kainic acid in the rabbit hippocampus was studied; the hippocampus was perfused continuously in the freely moving animal with an implanted 0.3‐mm dialysis fiber. The pattern of endogenous amino acids in the perfusate, reflecting extracellular amino acids, was monitored with liquid chromatography separation and fluorimetric detection of amino acid derivatives. Kainic acid was included in the perfusion medium for up to 70 min at 0.1–1.0 mM and, with time, induced epileptiform activity. Endogenous glutamic acid, taurine, and phosphoethanolamine levels were increased selectively at the lower perfusion concentrations of kainic acid. Long perfusion periods with higher concentrations increased the levels of virtually all amino acids. Perfusion of the hippocampus with depolarizing concentrations of potassium gave an amino acid response partly similar to that seen with kainic acid treatment. However, one notable difference between the two responses was that the extracellular concentration of glutamine, although not influenced by kainic acid, was significantly decreased after high potassium concentrations. These results confirm previous notions that kainic acid has a primarily excitatory effect, one manifestation of this effect being the release of glutamic acid.

A sensitive ELISA for glial fibrillary acidic protein: application in CSF of children

In the present study we describe a sensitive ELISA for determination of glial fibrillary acidic protein (GFAP). To validate the method combined determinations of GFAP and S-100 protein were performed in cerebrospinal fluid (CSF) of normal children and children with autism. The GFAP ELISA is of sandwich type and uses the biotin-avidin system. Sensitivity was 16 pg/ml. Between-day precision was 0.079 (coeff. of variance). S-100 protein concentrations were measured using a commercially available ELISA kit. Normal CSF from children and young adults were analysed. The CSF levels of GFAP in normal children were low (16-163 pg/ml). Both GFAP and S-100 protein concentrations correlated with age (P < 0.01 and P < 0.05, respectively), but the GFAP increment was more pronounced, probably reflecting the age-dependent expansion of the fibrillary astrocytes in the central nervous system (CNS). GFAP levels in children with infantile autism were higher than those in normal children of the same age range. S-100 protein concentrations were similar in both groups. High levels of GFAP in combination with normal S-100 protein concentrations in CSF indicates reactive astrogliosis in the CNS. In conclusion, the sensitive ELISA described makes it possible to measure low levels of GFAP present in the CSF of children. Combined assays of GFAP and S-100 protein can be used to discriminate between acute and chronic brain disorders in children.

Potassium-stimulated γ-aminobutyric acid release from neurons and glia

Summary Potassium-stimulated [ 3 H]GABA release has been studied with fractions of glial cells, neuronal perikarya and synaptosomes using a superfusion technique. A monotonic increase in the [ 3 H]GABA release was observed when external K + concentration was raised from5 to 15m M . However, there was no further increase in the [ 3 H]GABA release when K + was raised above15m M . The KCl-stimulated release was saturable at the level of 60–100% stimulation. Potassium-stimulated [ 3 H]GABA release from glial cells was unchanged in the absence of calcium while release from synaptosomes and neuronal perikarya demonstrated Ca 2+ -dependence. The potassium-stimulated [ 3 H]GABA release was enhanced when superfusion was performed in the presence of10 −5 M ouabain.

Excitatory amino acids in the cerebrospinal fluid of asphyxiated infants: relationship to hypoxic‐ischemic encephalopathy

Asphyxiated (n = 27) and control infants (n = 25) were subjected to spinal taps. Amino acids were measured with liquid chromatography and the degree of hypoxic‐ischemic cncephalopathy was determined in each case. In asphyxiated infants, the concentrations of aspartate and glutamate were 286% and 387% (p0.01 and p 0.05) of the control values. respectively. The Cerebrospinal fluid aspartate levels were significantly (p 0.05) higher in the group with severe (3.4 μmol/l) compared with the group with mild hypoxic‐ischemic encephalopathy (1.0 μmol/l). Glutamate was also higher in the group with severe (12.3 μmol/l) than in the groups with mild (2.7 μmol/l) or moderate (3.2 μmol/l) hypoxic‐ischemic enccphalopathy (p 0.05). High concentrations of excitatory amino acids were present in the CSF of asphyxiated infants which may exert excitotoxic effects.

A role for taurine in the maintenance of homeostasis in the central nervous system during hyperexcitation

Employing the brain dialysis technique, we demonstrate that the aspartate congener N-methyl-D,L-aspartic acid (NMA) stimulates the release of endogenous taurine in vivo in a Ca2+-dependent manner. Furthermore, exogenous taurine (1-10 mM) inhibits the NMA-induced Ca2+ influx into intracellular compartments. This suggests that the extracellular taurine concentration may control Ca2+ movement and thereby provide a homeostatic mechanism in situations of excessive excitation.

Neuropathology and Pressure in the Pig Brain Resulting from Low-Impulse Noise Exposure

Military personnel are exposed to occupational levels of blast overpressure during training. This study characterizes the pressure-time histories of air, underwater, and localized blast, and correlates blast parameters with neuropathology. Blast overpressure was produced by a howitzer, a bazooka, an automatic rifle, underwater explosives, or a shock tube. Anesthetized pigs were exposed in positions that simulated real training scenarios. Underwater exposures were performed using explosives at distances recommended by safety requirements. In other experiments, rats were exposed via a shock tube. The pressure changes were recorded with a hydrophone sensor in the brain of the pig and in rats with an optical fiber sensor. Histological examination of porcine brains revealed small parenchymal and subarachnoid hemorrhages, predominately in the occipital lobe, cerebellum, and medulla oblongata. Relative to the peak pressure in air, that in porcine brain (Pmax brain/air) was 0.7 for the bazooka and 0.5 and 0.7, respectively, for the 9- and 30-kPa howitzer. The attenuation was stronger in water: the detonation pulse had a brain/water ratio of 0.1, and the secondary pulses had ratios of 0.3-0.4. The results indicate that low-frequency spectra penetrate easily from air or water into the brain, but high-frequency spectra appear to be filtered by body structures. In addition, blast waves were recorded in the brain and abdomen of pigs after local exposure via shock tube to either the abdomen or the top of the skull. When the abdomen was exposed, the maximal peak value in the brain was only 3% of that in the abdomen. Moreover, part of this pressure could have been derived from the air outside the head. The results gave little support to significant transmission of pressure within the body.