Evidence for reactive synaptogenesis in the ventrolateral thalamus and red nucleus of the rat: changes in high affinity glutamate uptake and numbers of corticofugal fiber terminals

Summary High affinity glutamate uptake into corticofugal fiber terminals was measured in the ventrolateral thalamus and red nucleus at varying time intervals after lesions were made by kainic acid in the contralateral interpositus nucleus of the cerebellum in rats. Under similar conditions the density of cortical fiber terminals was estimated using the Fink-Heimer impregnation technique. 1. Glutamate uptake steadily increased in the ventrolateral thalamus up to 60 days after lesions in the contralateral cerebellum. 2. Similar changes were noted in the red nucleus. 3. The changes were dependent on the integrity of corticofugal fibers to the thalamus and red nucleus. 4. No changes in uptake of gammaaminobutyric acid were noted. 5. Saturation curves for glutamate uptake suggested a change in the maximal number of transport sites. 6. Fink-Heimer degeneration studies showed an increase in cortical terminals in the ipsilateral ventrolateral thalamus and in both rostral and caudal regions of the red nucleus following lesions in the contralateral interpositus nucleus. The data are consistent with an increase in the number of cortical fiber terminals in reaction to loss of cerebellar input to the ventrolateral thalamus and red nucleus. This study correlates anatomical and biochemical evidence for collateral sprouting in a model based on electrophysiologic data in the red nucleus and extends the model to include the thalamus.     

Distribution of glutamine-like immunoreactivity in the cerebellum of rat and baboon (Papio anubis) with reference to the issue of metabolic compartmentation

Summary The cellular and subcellular localization of glutamine, a major glutamate precursor, was studied by means of an antiserum raised against glutaraldehydefixed glutamine. Ultrathin sections from the cerebellar cortex of rat and baboon (Papio anubis) were incubated sequentially in the primary antiserum and in a secondary antibody coupled to colloidal gold particles. The labelling intensity was quantified by computer-aided calculation of gold particle densities. High levels of immunoreactivity occurred in glial cells (Bergmann fibres, astrocytes, and oligodendrocytes), intermediate levels in cell bodies and processes of granule cells, and low levels in terminals of presumed GABAergic or glutamatergic fibres (terminals of basket and Golgi cells, and of parallel, mossy, and climbing fibres). The labelling intensity of Purkinje cells showed some variation, but never exceeded that in glial cells. Within the nerve fibre terminals, the glutamine-like immunoreactivity showed some preference for mitochondria, but was otherwise evenly distributed. The predominant glial localization of glutamine was also obvious in light microscopic preparations processed according to the postembedding peroxidase-antiperoxidase procedure. Gold particle densities over different types of profile in glutamine immunolabelled sections were compared with particle densities over the corresponding types of profiles in neighbouring sections labelled with an antiserum to glutaraldehyde-fixed glutamate. The glutamate/glutamine ratio, expressed arbitrarily by the ratio between the respective gold particle densities, varied by a factor of about 6, with the highest ratio in the putative glutamatergic mossy and parallel fibre terminals, and the lowest ratio in glial elements. The remaining tissue components displayed intermediate ratios. The present study provides direct morphological evidence for the existence in the brain of distinct compartments with differing glutamate/glutamine ratios.This paper is dedicated to Professor Fred Walberg on the occasion of his 70th birthdayOn leave of absence from Department of Anatomy, Capital Institute of Medicine, You An Men Street, Beijing, China     

Functional AMPA/kainate receptors in human embryonic and foetal central nervous system

Here, functional AMPA/kainate receptors in human embryonic (5.5–7.5 gestational weeks) and foetal (8–10 gestational weeks) central nervous system tissue, shown by the cobalt labeling method, are reported. Specific agonist-induced cobalt incorporation was detected in brainstem and spinal cord cells, even in the youngest embryo studied. T-AMPA or kainate, but also vegetal toxins such as L-BOAA or acromelate, induced accumulation of cobalt. In contrast, no labeling was observed after exposure to KCl or NMDA. Cobalt labeled cells were particularly prominent in motor regions of brainstem and spinal cord. Co-application of the diuretic agent cyclothiazide, a desensitization blocker at AMPA receptors, dramatically increased the number of stained cells, which was particularly obvious in sensory regions, suggesting different receptor properties in motor versus sensory regions. This is the first study providing evidence for functional AMPA/kainate receptors, permeable to divalent cations, in brainstem and spinal cord at an early stage of human central nervous system development. Since many developmental processes are influenced by the modulation of cytosolic calcium, exposure at critical stages of embryogenesis to food or drug substances modifying the activity of AMPA/kainate receptors may alter brain development.     

Impaired Ca-signaling in astrocytes from the Ts16 mouse model of Down syndrome

The trisomy 16 mouse model of Down syndrome has been used to compare calcium (Ca)-homeostasis and Ca-signaling in astrocytes from trisomic mice and from diploid littermates. Ratio calcium-imaging of Fura-2/AM loaded primary astroglial cultures prepared from the hippocampus shows that resting Ca levels are on average significantly higher in trisomic than in the control astrocytes (280 vs. 120 nM). Serotonin (3 μM) and glutamate (30–300 μM) evoked transient Ca-increases from 400 to 600 nM in euploid but from only 20 to 150 nM in trisomic astrocytes. Imaging of ATP-driven Ca-accumulation in cellular organelles revealed a significantly stronger uptake of Ca in trisomic astrocytes that might buffer cytosolic Ca-increases. Our results demonstrate major disturbances in Ca-signaling in trisomic astrocytes that are likely to be of pathophysiological relevance.     

The involvement of glutamatergic transmission in the mechanism of movement disorders induced by reversive rotation of white mice

The ability of the selective non-competitive NMDA receptor blocker MK-801 and a series of new glutamate antagonists—the adamantane derivatives IEM-1754 and IEM-1857 and phencyclidine (IEM-1925)—to prevent movement disorders induced by reversive rotation in mice was studied. I.p. MK-801 at a dose of 0.15 ml and IEM-1754 at a dose of 5.0 mg/kg prevented the development of akinesia in response to reversive rotation, as effectively as scopolamine, a known agent which provides effective prophylaxis for movement diseases. IEM-1857, the quaternary analog of IEM-1754, was not effective. IEM-1925 significantly increased the responses of mice to reversive rotation, possibly because of its high activity in relation to other subtypes of glutamate receptors. These data provide evidence for the involvement of glutamatergic transmission in the mechanism of movement disorders of vestibular origin. Translated from Rossiiskii Fiziologicheskii Zhurnal imeni I. M. Sechenova, Vol. 85, No. 4, pp. 497–501, May, 1999.     

Dependence of the anterior olfactory area self-stimulation upon the lateral hypothalamic area

The functional relation between the anterior olfactory area (AO) and the lateral hypothalamic area (LH) was examined in a self-stimulation situation. Bar-pressing responses for AO sitmulation were suppressed by unilateral injection of procaine, and enhanced by glutamate, into LH. Neither procaine nor glutamate injected into AO had any influence upon LH self-stimulation. It is unlikely that the procaine effect was due to motor disturbance because similar injection of procaine into LH did not disturb the performance of a one-way avoidance task. It appears that the rewarding effect of AO stimulation is dependent upon the excitation of the more caudal structures including LH.     

Reversal or reduction of glutamate and GABA transport in CNS pathology and therapy

A dysfunction of amino acid neurotransmitter transporters occurs in a number of central nervous system disorders, including stroke, epilepsy, cerebral palsy and amyotrophic lateral sclerosis. This dysfunction can comprise a reversal of transport direction, leading to the release of neurotransmitter into the extracellular space, or an alteration in transporter expression level. This review analyses the role of glutamate and GABA transporters in the pathogenesis and therapy of a number of acute and chronic neurological disorders.     

Comparison of glutamate metabolism in low K (LK), high K and high glutathione (HK/HG) and high K and low glutathione (HK/LG) dog red blood cells

The reasons for the high accumulation of glutamate (Glu), aspartate (Asp) and glutamine (Gin) in high K and high glutathione (HK/HG) dog red blood cells (DRBCs) have been explained as due to enhanced Glu/Asp influxes. However, in our study, Glu/Asp influxes in high K and low glutathione (HK/LG) DRBCs were low, whereas their cellular Asp and Gin contents were high. In low K (LK) DRBCs, there were also other variant cells with high Asp accumulation, but extremely low Glu/Asp influxes. So, the high amino acid accumulation in DRBCs of these new variants might not be due to Glu/Asp influxes. To examine the high accumulation of these amino acids in these variant DRBCs, first, LK and HK/LG DRBCs were classified into two subgroups with their Na-dependent Glu/Asp influxes; one had clear Na-dependent Glu/Asp transport (GAT⁺), and the other failed to have any transport (GAT⁻). The influxes of both Glu and Asp in HK/HG DRBCs were the highest, and the order was HK/HG>LK/GAT⁺>HK/LG/GAT⁺>>LK/GAT⁻=HK/LG/GAT⁻. LK/GAT⁺ cells represented normal DRBCs. Glu/Asp influxes were only trace in both LK/GAT⁻ and HK/LG/GAT⁻ cells, but Glu and Asp concentration was high in HK/LG/GAT⁻ cells whereas Asp concentration was high in LK/GAT⁻ cells. In HK/HG cells, the conversion of Glu into Gin in whole cells was several fold higher than in the other cell groups due to the differing amount of the substrate of glutamine synthetase, Glu, but glutamine synthetase activity itself was not different among these cell groups. Furthermore, glutamine synthetase and glutaminase activities were not different among the cell groups. Therefore, these enzymes were not involved in the high amino acid accumulation.     

Serotonergic inhibition of the mossy fibre—granule cell glutamate transmission in rat cerebellar slices

Summary The glutamatergic mossy fibre granule cell pathway has been investigated in rat cerebellar slices. Exposure to 35 mM KCI, a concentration of K⁺ known to elicit Ca²⁺-dependent releases of excitatory amino acids from cerebellar slices, raised cGMP levels. The cGMP response was decreased in a concentration-dependent manner by D-(–)-2-amino-5-phosphonopentanoic acid (D-AP5) and by 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) indicating the involvement of ionotropic glutamate receptors of both the N-methyl-D-aspartate (NMDA) and the non-NMDA type. The K⁺-evoked production of cGMP was potently inhibited (EC₅₀ = 1.21 nM) by 1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane (DOI), a selective 5-HT₂ receptor agonist. The effect of DOI (0.01 M) was antagonized by 0.03 M of the 5-HT₂ receptor antagonists ketanserin and methiothepin. At concentrations higher than 0.1 M, both antagonists increased on their own the cGMP response elicited by high-K⁺. This effect was insensitive to tetrodotoxin.It had been previously shown that rat mossy fibre endings release glutamate upon depolarization and that such release can be inhibited by activation of 5-HT₂ receptors sited on the mossy fibre endings. Altogether the available data suggest the following conclusions: (a) the glutamate/aspartate endogenously released in cerebellar slices during K⁺ depolarization increase cGMP synthesis through the activation of both NMDA and non-NMDA receptors; (b) a portion of the cGMP response can be prevented by 5-HT₂ receptor activation and may reflect the activity of the mossy fibre-granule cell pathway. Thus serotonin is likely to exert a potent inhibitory control of the excitatory mossy fibre input to the cerebellum by acting at receptors of the 5-HT₂ type. Correspondence to M. Raiteri at the above address     

Glutamate: its role in learning,memory, and the aging brain

l-Glutamate is the most abundant of a group of endogenous amino acids in the mammalian central nervous system which presumably function as excitatory neurotransmitters and under abnormal conditions may behave as neurotoxins. As neurotransmitters, these compounds are thought to play an important role in functions of learning and memory. As neurotoxins, they are believed to be involved in the pathogenesis of a variety of neurodegenerative disorders in which cognition is impaired. Moreover, brain structures which are considered anatomical substrata for learning and memory may be particularly vulnerable to the neurotoxic actions of these excitatory amino acids, especially in the elderly who are also the segment of the population most susceptible to impairments of mnemonic function. This paper is a review of data concerning the role of excitatory amino acids in the processes of learning and memory and in the pathogenesis and treatment of disorders thereof.