D-Aspartate Uptake and Release in the Guinea Pig Spinal Cord After Partial Ablation of the Cerebral Cortex

This study attempts to determine if L-glutamate and L-aspartate may be transmitters of the guinea pig corticospinal tract. Unilateral ablations were made of the frontal and parietal neocortex which destroyed most of the motor and somatosensory areas in the right cerebral hemisphere. In lesioned animals, transverse sections of the cervical enlargement of the spinal cord (segments C6--T1) were stained to reveal degenerating fibers. Degeneration of axons first appeared 4 days after surgery, reached a maximum on the seventh day, and began to wane by the ninth day. The most prominent loss of axons appeared deep in the dorsal funiculus and in laminae IV-IX of the gray matter contralateral to the cortical lesion. Ipsilaterally, there was very sparse degeneration of fibers in the dorsal and ventral funiculi and in the spinal gray matter. The uptake and release of D-[3H]aspartate, a putative nonmetabolizable marker for L-glutamate and L-aspartate, were measured in dissected quadrants of the cervical enlargement taken from intact and lesioned animals. The uptake and the electrically evoked, Ca2+-dependent release of D-[3H]aspartate were depressed by 29-35% at 4 and 7 days after surgery, but only in tissue that was contralateral to the cortical ablation. The lesion had no effect on the uptake and release of exogenous gamma-[14C]aminobutyric acid, which were measured as indices of the postlesion integrity of neurons in the spinal gray matter. These findings suggest that the synaptic endings of corticospinal fibers probably mediate the uptake and release of D-[3H]aspartate and, therefore, may use L-glutamate and/or L-aspartate as a transmitter.     

Decreased Uptake and Release of D-Aspartate in the Guinea Pig Spinal Cord After Partial Cordotomy

This study attempts to determine if L-glutamate and/or L-aspartate may be transmitters of neural tracts descending from the brain to the spinal cord. The uptake and electrically evoked release of D-[3H]aspartate, a putative marker for L-glutamate and L-aspartate, were measured in the cervical enlargement of the guinea pig spinal cord. These activities were compared using unlesioned animals and others with a lesion on the right side of the spinal cord. Partial cordotomy (segment C5) produced a heavy loss of descending fibers, a small loss of primary sensory fibers, and a depression of the uptake and the Ca2+ -dependent, electrically evoked release of D-aspartate ipsilateral and caudal to the lesion. Contralaterally, there was a moderate loss of corticospinal fibers, some loss of other descending axons, and a depression of D-aspartate release. Dorsal rhizotomy (segments C4-T1) produced a heavy loss of primary sensory fibers ipsilateral to the lesion. Ipsilaterally, but not contralaterally, the uptake and release of D-aspartate were depressed. Degeneration after partial cordotomy in combination with dorsal rhizotomy was assumed to be the sum of that produced by each lesion separately. This combined lesion depressed D-aspartate uptake ipsilaterally and depressed D-aspartate release on both sides of the cervical enlargement. None of the lesions altered the uptake and the evoked release of [3H]GABA. These findings support the hypothesis that the synaptic endings of one or more neural tracts descending from the brain to the spinal cord mediate the uptake and release of D-aspartate and, therefore, may use L-glutamate or L-aspartate as a transmitter.     

Evidence for Glutamatergic Projections from the Cochlear Nucleus to the Superior Olive and the Ventral Nucleus of the Lateral Lemniscus

Abstract: This study attempts to determine if projections ascending from the guinea pig cochlear nucleus (CN) could be glutamatergic and/or aspartatergic. Multiple radio frequency lesions were made to ablate the right CN. The ablation was verified histologically. To identify the principal targets of CN efferents, silver impregnation methods were used to localize the preterminal degeneration of fibers in transverse sections of the brainstem 5 and 7 days after CN ablation. CN efferents projected heavily to the lateral superior olive (LSO) ipsilaterally, the medial superior olive (MSO) bilaterally, and contralaterally to the medial (MNTB) and ventral (VNTB) nuclei of the trapezoid body, the ventral (VNLL) and intermediate nuclei of the lateral lemniscus and the central nucleus of the inferior colliculus (ICc). There were smaller projections to the lateral nucleus of the trapezoid body ipsilaterally, the dorsal and dorsomedial periolivary nuclei bilaterally, and the dorsal nucleus of the lateral lemniscus contralaterally. There were sparse projections to the VNLL and ICc ipsilaterally and the CN contralaterally, and a very sparse projection to the contralateral LSO. To determine if CN efferents were glutamatergic and/or aspartatergic, the fresh brainstem was sectioned transversely and samples of the LSO, MSO, MNTB, VNLL, and ICc were taken to measure the electrically evoked release and the uptake of d -[³H]Asp and [¹⁴C]Gly or [¹⁴C]GABA 3–5 days after the CN ablation. The release studies suggest that only certain of the histologically identified projections ascending from the CN may be glutamatergic and/or aspartatergic. CN ablation depressed d -[³H]Asp release in the MSO bilaterally and in the contralateral MNTB and VNLL, suggesting that the CN efferents to these nuclei may use glutamate or aspartate as a transmitter. It was unclear whether a marginal depression of d -[³H]Asp release in the ipsilateral LSO reflected the presence of glutamatergic CN projections to this nucleus. d -[³H]Asp release in the ICc was unaffected, suggesting that CN efferents to this nucleus may not be glutamatergic. There were no deficits in d -[³H]Asp uptake. [¹⁴C]Gly release from the LSO and MSO was unchanged. [¹⁴C]Gly uptake was unchanged in the MSO and depressed only in the contralateral LSO, possibly reflecting subnormal uptake activity in endings contributed by contralateral MNTB cells that had lost their CN efferents. [¹⁴C]GABA uptake in the MNTB, VNLL, and ICc was unchanged. [¹⁴C]GABA release was unchanged in the VNLL and ICc. [¹⁴C]GABA release was depressed only in the contralateral MNTB, possibly reflecting the loss of a small complement of GABAergic CN efferents and the reaction of GABAergic projections from the contralateral VNTB to their loss of CN efferents.     

Decreased Uptake and Release of D-Aspartate in the Guinea Pig Spinal Cord After Dorsal Root Section

This study attempts to determine if L-glutamate and/or L-aspartate may be transmitters of dorsal sensory neurons. The uptake and the electrically evoked release of D-[3H]aspartate, a putative marker for L-glutamate and L-aspartate, were measured in the cervical enlargement (segments C4-T1) of the guinea pig spinal cord before and after cutting dorsal roots C5-T1 on the right side. The uptake and the release of gamma-aminobutyric acid (GABA) also were measured as indices of the integrity of GABAergic neurons in the spinal cord. The cervical enlargement was excised and divided into left and right halves, then into dorsal and ventral quadrants. Quadrants from unlesioned animals took up D-aspartate and GABA, achieving concentrations in the tissues which were 14-25 times that in the medium. Subsequently, electrical stimulation evoked a Ca2+-dependent release of D-aspartate and of GABA. The uptake and release of D-aspartate and GABA were similar in tissues taken from intact and sham-operated animals. However, dorsal rhizotomy, without damage to dorsal radicular or spinal blood vessels, depressed the uptake (by 22-29%) and the release (by 50%) of D-aspartate only in quadrants ipsilateral to the lesion. The uptake and the release of GABA were unchanged. In transverse sections of the cervical enlargement, stained to reveal degenerating fibers, by far the heaviest loss of axons occurred in the cuneate fasciculus and in the gray matter ipsilateral to the cut dorsal roots. These findings suggest that the synaptic endings of dorsal sensory neurons probably mediate the uptake and the release of D-aspartate and, therefore, may use L-glutamate or L-aspartate as a transmitter. When spinal blood vessels were damaged during dorsal rhizotomy, the deficits in D-aspartate uptake and release were larger than those in the absence of vascular damage and were accompanied by deficits in GABA uptake and release. These findings imply that vascular damage results in the loss of intraspinal neurons, some of which probably mediate the uptake and release of D-aspartate and, therefore, may use L-glutamate and/or L-aspartate as a transmitter.     

Ascending and descending projections of the lateral vestibular nucleus in the rat

The tracer neurobiotin was injected into the lateral vestibular nucleus in rat and the efferent fiber connections of the nucleus were studied. The labeled fibers reached the diencephalon rostrally and the sacral segments of the spinal cord caudally. In the diencephalon, the ventral posteromedial and the gustatory nuclei received the most numerous labeled fibers. In the mesencephalon, the inferior colliculus, the interstitial nucleus of Cajal, the nucleus of Darkschewitch, the periaqueductal gray matter and the red nucleus received large numbers of labeled fibers. In the rhombencephalon, commissural and internuclear connections originated from the lateral vestibular nucleus to all other vestibular nuclei. The medioventral (motor) part of the reticular formation was richly supplied, whereas fewer fibers were seen in the lateral (vegetative) part. In the spinal cord, the descending fibers were densely packed in the anterior funiculus and in the ventral part of the lateral funiculus. Collaterals invaded the entire gray matter from lamina IX up to lamina III; the fibers and terminals were most numerous in laminae VII and VIII. Collateral projections were rich in the cervical and lumbosacral segments, whereas they were relatively poor in the thoracic segments of the spinal cord. It was concluded that the fiber projection in the rostral direction was primarily aimed at sensory-motor centers; in the rhombencephalon and spinal cord, fibers projected onto structures subserving various motor functions.     

Quisqualic Acid Modulates Kainate Responses in Cultured Cerebellar Granule Cells

The activation of kainic acid and quisqualic acid receptors in cultured cerebellar granule cells stimulated the release of preaccumulated D-[3H]aspartate. The effect of kainate could be distinguished from that of quisqualate by its sensitivity to the antagonists kynurenic acid and 2,3-cis-piperidine dicarboxylic acid. At a concentration of kainic acid (50 microM) close to its half-maximal releasing effect, simultaneous addition of quisqualic acid (10-50 microM) resulted in a significant dose-dependent inhibition of the kainate-induced component of D-[3H]aspartate release, which was monitored by the progressive decrease in sensitivity of the evoked release to kynurenic acid. In contrast, when kainic acid was used at a subeffective concentration (10 microM), addition of low doses of quisqualate (2-5 microM) resulted in a synergistic effect on D-[3H]aspartate release. Under these conditions, the effect of the two agonists was sensitive to kynurenic acid. Kainic acid (50-100 microM) also caused a dose-dependent, kynurenic acid-sensitive accumulation of cyclic GMP (cGMP) in granule cell cultures. Quisqualic acid was, by itself, ineffective and prevented, in a dose-dependent manner, the kainate-induced cGMP formation (IC50 = 5 microM). Finally, the guanylate cyclase activator sodium nitroprusside greatly enhanced cGMP formation but had no effect on D-[3H]aspartate release. Together, these results demonstrate the existence of complex interactions between quisqualic and kainic acids and indicate that the effects of the two glutamate agonists on D-[3H]aspartate release and on cGMP accumulation are independent.     

Glutamate Receptor Subtypes in Cultured Cerebellar Neurons: Modulation of Glutamate and γ-Aminobutyric Acid Release

Using cerebellar, neuron-enriched primary cultures, we have studied the glutamate receptor subtypes coupled to neurotransmitter amino acid release. Acute exposure of the cultures to micromolar concentrations of kainate and quisqualate stimulated D-[3H]aspartate release, whereas N-methyl-D-aspartate, as well as dihydrokainic acid, were ineffective. The effect of kainic acid was concentration dependent in the concentration range of 20-100 microM. Quisqualic acid was effective at lower concentrations, with maximal releasing activity at about 50 microM. Kainate and dihydrokainate (20-100 microM) inhibited the initial rate of D-[3H]aspartate uptake into cultured granule cells, whereas quisqualate and N-methyl-DL-aspartate were ineffective. D-[3H]Aspartate uptake into confluent cerebellar astrocyte cultures was not affected by kainic acid. The stimulatory effect of kainic acid on D-[3H]aspartate release was Na+ independent, and partly Ca2+ dependent; the effect of quisqualate was Na+ and Ca2+ independent. Kynurenic acid (50-200 microM) and, to a lesser extent, 2,3-cis-piperidine dicarboxylic acid (100-200 microM) antagonized the stimulatory effect of kainate but not that of quisqualate. Kainic and quisqualic acid (20-100 microM) also stimulated gamma-[3H]-aminobutyric acid release from cerebellar cultures, and kynurenic acid antagonized the effect of kainate but not that of quisqualate. In conclusion, kainic acid and quisqualic acid appear to activate two different excitatory amino acid receptor subtypes, both coupled to neurotransmitter amino acid release. Moreover, kainate inhibits D-[3H]aspartate neuronal uptake by interfering with the acidic amino acid high-affinity transport system.     

An Excitant Amino Acid Projection from the Medial Prefrontal Cortex to the Anterior Part of Nucleus Accumbens in the Rat

High-affinity uptake of neurotransmitter substrates in synaptosome-containing homogenates and tissue concentrations of amino acids were examined in subcortical areas 5-6 days after bilateral N-methyl-D-aspartate lesions confined to rat medial prefrontal cortex. D-[3H]Aspartate (32% of control) and [3H] gamma-aminobutyric acid ( [3H]GABA) (60% of control) uptakes were significantly reduced in medial prefrontal cortex, whereas [3H]choline (110% of control) uptake was unchanged, suggesting the production of axon-sparing lesions. The uptake of D-[3H]aspartate (76% of control), but not of [3H]GABA or [3H]choline, was significantly reduced in nucleus accumbens, with no concomitant reduction in amino acid concentrations. When examined in serial coronal sections, reduced D-[3H]aspartate uptake was confined to the most anterior 500 micron of nucleus accumbens (67% of contralateral sample). No significant reductions of uptake or amino acid concentrations were observed in caudate putamen or ventral tegmental area. These results suggest a role for glutamate or aspartate as neurotransmitters in projections from medial prefrontal cortex to anterior nucleus accumbens. Medial prefrontal cortex may represent the major excitatory cortical input to the nucleus accumbens.     

Projections of the optic tectum and the mesencephalic nucleus of the trigeminal nerve in the tegu lizard (Tupinambis nigropunctatus)

Summary Fibers undergoing Wallerian degeneration following tectal lesions were demonstrated with the Nauta and Fink-Heimer methods and traced to their termination. Four of the five distinct fiber paths originating in the optic tectum appear related to vision, while one is related to the mesencephalic nucleus of the trigeminus. The latter component of the tectal efferents distributes fibers to 1) the main sensory nucleus of the trigeminus, 2) the motor nucleus of the trigeminus, 3) the nucleus of tractus solitarius, and 4) the intermediate gray of the cervical spinal cord.The principal ascending bundle projects to the nucleus rotundus, three components of the ventral geniculate nucleus and the nucleus ventromedialis anterior ipsilaterally, before it crosses in the supraoptic commissure and terminates in the contralateral nucleus rotundus, ventral geniculate nucleus and a hitherto unnamed region dorsal to the nucleus of the posterior accessory optic tract.Fibers leaving the tectum dorso-medially terminate in the posterodorsal nucleus ipsilaterally and the stratum griseum periventriculare of the contralateral tectum. The descending fiber paths terminate in medial reticular cell groups and the rostral spinal cord contralaterally and in the torus and the lateral reticular regions ipsilaterally. The ipsilateral fascicle also issues fibers to the magnocellular nucleus isthmi.     

Kainic Acid Differentially Affects the Synaptosomal Release of Endogenous and Exogenous Amino Acidic Neurotransmitters

Presynaptic actions of kainic acid have been tested on uptake and release mechanisms in synaptosome-enriched preparations from rat hippocampus and goldfish brain. Kainic acid increased in a Ca2+-dependent way the basal release of endogenous glutamate and aspartate from both synaptosomal preparations, with the maximum effect (40-80%) being reached at the highest concentration tested (1 mM). In addition, kainic acid potentiated, in an additive or synergic way, the release of excitatory amino acids stimulated by high K+ concentrations. Kainic acid at 1 mM showed a completely opposite effect on the release of exogenously accumulated D-[3H]aspartate. The drug, in fact, caused a marked inhibition of both the basal and the high K+-stimulated release. Kainic acid at 0.1 mM had no clear-cut effect, whereas at 0.01 mM it caused a small stimulation of the basal release. The present results suggest that kainic acid differentially affects two neurotransmitter pools that are not readily miscible in the synaptic terminals. The release from an endogenous, possibly vesiculate, pool of excitatory amino acids is stimulated, whereas the release from an exogenously accumulated, possibly cytoplasmic and carrier-mediated, pool is inhibited or slightly stimulated, depending on the external concentration of kainic acid. Kainic acid, in addition, strongly inhibits the high-affinity uptake of L-glutamate and D-aspartate in synaptic terminals. All these effects appear specific for excitatory amino acids, making it likely that they are mediated through specific recognition sites present on the membranes of glutamatergic and aspartatergic terminals. The relevance of the present findings to the mechanism of excitotoxicity of kainic acid is discussed.     

The hypothalamo-cerebellar projection in the rat: origin and transmitter.

Hypothalamic neurons projecting to cerebellum were identified by retrograde tracing with wheat germ agglutinin-horseradish peroxidase (WGA-HRP) in the rat. Selective D-[3H]aspartate labelling was used to investigate whether any of these connections may use excitatory amino acids as transmitters. The WGA-HRP experiments revealed that the hypothalamo-cerebellar fibers have their main origins in the lateral, dorsal and posterior hypothalamic areas, and the tubero-mammillary nucleus, while smaller numbers of cells were observed in tuber cinereum, the anterior hypothalamic area, and the periventricular and paraventricular nuclei. After injections of D-[3H]aspartate into the cerebellar cortex, intense labelling of the olivocerebellar climbing fiber system was observed, but hypothalamic cells were not retrogradely labelled with this selective tracer. The absence of D-[3H]aspartate labelling indicates that hypothalamo-cerebellar neurons lack specific uptake mechanisms for excitatory amino acids, but it does not entirely preclude the possibility that some of these hypothalamic neurons may use such transmitters. Many cerebellar projecting cells were located in the tubero-mammillary nucleus, which is known to contain histaminergic and GABAergic neurons, and it was concluded that part of the hypothalamo-cerebellar pathways may use histamine and/or GABA as transmitters. The transmitter remains unknown for other parts of the hypothalamo-cerebellar pathways.     

Evidence for a Glutamatergic Pathway from the Guinea Pig Auditory Cortex to the Inferior Colliculus

Abstract: We attempt to provide evidence that the projection from the guinea pig auditory cortex (AC) to the inferior colliculus (IC) may contain glutamatergic or GABAergic fibers. Seven days after unilateral AC aspiration, histological studies indicated almost complete AC destruction and preterminal degeneration of fibers and terminal fields in the dorsal cortex (DCIC), external cortex (ECIC), and central nucleus (CNIC) of the IC ipsilateral to the ablated AC. Contralaterally, degeneration appeared in the DCIC. AC ablation depressed the electrically evoked Ca²⁺-dependent release of d -[³H]aspartate ( d -[³H]Asp) in the ipsilateral DCIC, ECIC, and CNIC, and d -[³H]Asp uptake in the CNIC. Together with other evidence that the corticocollicular pathway is excitatory, these findings suggest that this projection may contain glufamatergic and/or aspartatergic (Glu/Asp-ergic) fibers. Glutamic acid decarboxylase immunoreactivity was not apparent in presumed pyramidal cells of layer V of the AC retrogradely labeled with biotinylated dextran injected into the ipsilateral IC. Thus, corticocollicular neurons probably do not synthesize GABA and may not be GABAergic. However, AC ablation depressed [¹⁴C]GABA release from the ipsilateral DCIC and ECIC, and [¹⁴C]GABA uptake in the DCIC. These findings are consistent with the atrophy or down-regulation of some subcortical neurons that mediate GABAergic transmission in the IC.     

Supersensitivity of the GABA receptor in the frog spinal cord, as induced by kainic acid

In the isolated frog spinal cord perfused with kainic acid (KA, 5 X 10(-4) M) containing Ringer's solution, within 2 hr there were increases in the amplitude of the dorsal root depolarization, as induced by the GABA-agonists. KA perfusion produced increases in the specific binding of [3H]muscimol to crude synaptic membranes and incubation with KA for 3 hr did not increase [3H]muscimol binding. [3H]GABA was released from KA-treated spinal cord slices in the presence of high K+. KA-induced supersensitivity of the dorsal root to GABA may relate to direct actions on primary afferent terminals and not to denervation of GABAergic neurons.     

γ-Aminobutyric Acid and Glycine Modulate Each Other''s Release Through Heterocarriers Sited on the Releasing Axon Terminals of Rat CNS

The ability of gamma-aminobutyric acid (GABA) and glycine (Gly) to modulate each other's release was studied in synaptosomes from rat spinal cord, cerebellum, cerebral cortex, or hippocampus, prelabeled with [3H]GABA or [3H]Gly and exposed in superfusion to Gly or to GABA, respectively. GABA increased the spontaneous outflow of [3H]Gly (EC50, 20.8 microM) from spinal cord synaptosomes. Neither muscimol nor (-)-baclofen, up to 300 microM, mimicked the effect of GABA, which was not antagonized by either bicuculline or picrotoxin. However, the effect of GABA was counteracted by the GABA uptake inhibitors nipecotic acid and N-(4,4-diphenyl-3-butenyl)nipecotic acid. Moreover, the GABA-induced [3H]Gly release was Na+ dependent and disappeared when the medium contained 23 mM Na+. The effect of GABA was Ca2+ independent and tetrodotoxin insensitive. Conversely, Gly enhanced the outflow of [3H]GABA from rat spinal cord synaptosomes (EC50, 100.9 microM). This effect was insensitive to both strychnine and 7-chlorokynurenic acid, antagonists at Gly receptors, but it was strongly Na+ dependent. Also, the Gly-evoked [3H]GABA release was Ca2+ independent and tetrodotoxin insensitive. GABA increased the outflow of [3H]Gly (EC50, 11.1 microM) from cerebellar synaptosomes; the effect was not mimicked by either muscimol or (-)-baclofen nor was it prevented by bicuculline or picrotoxin. The GABA effect was, however, blocked by GABA uptake inhibitors and was Na+ dependent. Gly increased [3H]GABA release from cerebellar synaptosomes (EC50, 110.7 microM) in a strychnine- and 7-chlorokynurenic acid-insensitive manner. This effect was Na+ dependent. The effects of GABA on [3H]Gly release seen in spinal cord and cerebellum could be reproduced also with cerebrocortical synaptosomes.(ABSTRACT TRUNCATED AT 250 WORDS)     

The effect of GABA on lumbar terminations of rubrospinal neurons in the cat spinal cord

Although GABA and piperidine-4-sulphonic acid depolarize I a afferent terminations in the cat spinal cord by activation of bicuculline-sensitive GABA receptors, no evidence was obtained for a bicuculline-sensitive alteration by either gabamimetic of the electrical threshold of rubrospinal terminations in the spinal intermediate nucleus. The terminal axonal arborizations in the spinal cord of neurons in the red nucleus thus do not have GABA receptors similar to those on the cell bodies. The results are discussed in relation to the depolarizing action of GABA on some central neurons, and on neurons with peripheral cell bodies, and to probable differences in the intracellular chloride content of neurons having peripheral or central cell bodies, and thus of different embryological origin. A presynaptic depolarizing inhibitory process mediated by GABA appears to be confined to the terminals of primary afferent fibres in the mammalian central nervous system.     

Amino Acid Levels in the Guinea Pig Spinal Gray Matter After Axotomy of Primary Sensory and Descending Tracts

This study attempts to determine if the axonal endings of dorsal root sensory fibers and of descending axons to the spinal gray matter in the guinea pig store glutamate and/or aspartate. Bilateral dorsal rhizotomy (spinal segments C5-T1) and partial cordotomy (segment C5, right side) were used to interrupt primary sensory and descending tracts, respectively. At 1 and 2 days after surgery, amino acid levels were determined in regions microdissected from areas of the gray matter of spinal segment C7 that receive heavy projections from the primary sensory and the descending tracts. These regions were identified by visualizing the degeneration of axons and their terminal fields in silver-impregnated light microscopic preparations of the spinal cord. After dorsal rhizotomy, the heaviest degeneration in the spinal gray appeared centrally in laminae II-IV and medially in laminae IV-VI. The levels of aspartate, glutamate, and gamma-aminobutyrate were reduced by 34, 21, and 26% in laminae II-IV and 28, 33, and 23% in medial laminae IV-VI. The levels of glycine, alanine, and threonine-serine-glutamine (unseparated) were increased. After partial cordotomy, the heaviest degeneration in the spinal gray appeared laterally in laminae IV-VI, dorsolaterally in lamina VII, and in lamina IX. The levels of aspartate and glutamate were reduced by 22 and 28% in lateral laminae IV-VI and by 26 and 28% in dorsolateral laminae VII and IX. Glycine levels were reduced by 9% in dorsolateral laminae VII and IX. The levels of gamma-aminobutyrate, alanine, and threonine-serine-glutamine were either unchanged or raised. These findings suggest that the axonal endings of the primary sensory and of one or more of the descending tracts probably contain relatively high levels of glutamate and aspartate, and that they may use these amino acids as transmitters. The partial deafferentation of spinal interneurons and the destruction of some propriospinal fibers probably caused the losses of gamma-aminobutyrate and glycine, and contributed modestly to those of aspartate.     

Endogenous Content and Release of [3H]-GABA and [3H]-Glutamate in the Spinal Cord of Chronically Undernourished Rat

The aim of this study was to determine the effect of chronic undernutrition on the content and release of γ-amino butyric acid (GABA) and glutamate (GLU) transmitters in the rat spinal cord. The release of [³H]-GABA and [³H]-GLU was determined by radioactive liquid scintillation techniques, and the concentrations of GABA and GLU in spinal cord preparations from control and undernourished young rats (50–60 days old) were measured by reverse-phase HPLC. The GABA and GLU contents in the lumbar spinal dorsal horn (L6 segment) were significantly lower in undernourished rats relative to control rats (22.2 ± 3.7 and 10.7 ± 1.9 %, respectively; P < 0.05). Spinal cord blocks from undernourished animals also showed lower rates of [³H]-GABA and [³H]-GLU release than controls (27.6 ± 3.5 and 12.8 ± 2.5 %, respectively; P < 0.01). We propose that the decreases in GLU content and release are consistent with a reduced activation of either afferent fibers, spinal glutaminergic neurons, or both. Furthermore, we propose that the decreased content and release of GABA in undernourished animals are related to a depression in pre- and post-synaptic inhibition. In addition, we hypothesize that the reductions in GABA content and release serve as compensatory mechanisms to counterbalance decreases in sensory transmission and GLU content in the spinal cord of the chronically undernourished rat.     

Release of γ-[3H]Aminobutyric Acid from Rat Olfactory Bulb and Substantia Nigra: Differential Modulation by Glutamic Acid

We have studied the glutamate modulation of gamma-[3H]aminobutyric acid ([3H]GABA) release from GABAergic dendrites of the external plexiform layer of the olfactory bulb and from GABAergic axons of the substantia nigra. In the olfactory bulb, [3H]GABA release was induced by high K+ and kainate, and not by aspartate and glutamate alone. However, when the tissue was conditioned by a previous K+ depolarization, glutamate and aspartate caused [3H]GABA release. The effect of glutamate was significantly enhanced when the GABA uptake mechanism was blocked by nipecotic acid. N-Methyl-D-aspartate and quisqualate did not cause [3H]GABA release under the same conditions. The acidic amino acid receptor antagonist 2-amino-4-phosphonobutyric acid and the N-methyl-D-aspartate receptor antagonist 2-amino-5-phosphonovaleric acid significantly inhibited the K+-glutamate- and the kainate-induced [3H]GABA release. Mg2+ (5 mM), which blocks the N-methyl-D-aspartate receptors, significantly inhibited the K+-glutamate-induced but not the kainic acid-induced [3H]GABA release. The K+-glutamate-stimulated release, but not the K+-stimulated [3H]GABA release, was strongly inhibited by Na+-free solutions or by 300 nM tetrodotoxin. Apparently the glutamate-induced release of [3H]GABA occurs through an interneuron because it is dependent on the presence of nerve conduction. In the substantia nigra no [3H]GABA release was elicited by any of the glutamate agonists tested. The present results clearly differentiate between the effects of glutamate on the release of [3H]GABA from the substantia nigra and from the olfactory bulb.(ABSTRACT TRUNCATED AT 250 WORDS)     

Stimulation of [3H]GABA and β-[3H]Alanine Release from Rat Brain Slices by cis-4-Aminocrotonic Acid

Abstract: cis -4-Aminocrotonic acid (CACA; 100 µ M ), an analogue of GABA in a folded conformation, stimulated the passive release of [³H]GABA from slices of rat cerebellum, cerebral cortex, retina, and spinal cord and of β-[³H]alanine from slices of cerebellum and spinal cord without influencing potassium-evoked release. In contrast, CACA (100 µ M ) did not stimulate the passive release of [³H]taurine from slices of cerebellum and spinal cord or of d -[³H]aspartate from slices of cerebellum and did not influence potassium-evoked release of [³H]taurine from the cerebellum and spinal cord and d -[³H]aspartate from the cerebellum. These results suggest that the effects of CACA on GABA and β-alanine release are due to CACA acting as a substrate for a β-alanine-sensitive GABA transport system, consistent with CACA inhibiting the uptake of β-[³H]alanine into slices of rat cerebellum and cerebral cortex. The observed K _i for CACA against β-[³H]alanine uptake in the cerebellum was 750 ± 60 µ M . CACA appears to be 10-fold weaker as a substrate for the transporter system than as an agonist for the GABA_c receptor. The effects of CACA on GABA and β-alanine release provide indirect evidence for a GABA transporter in cerebellum, cerebral cortex, retina, and spinal cord that transports GABA, β-alanine, CACA, and nipecotic acid that has a similar pharmacological profile to that of the GABA transporter, GAT-3, cloned from rat CNS. The structural similarities of GABA, β-alanine, CACA, and nipecotic acid are demonstrated by computer-aided molecular modeling, providing information on the possible conformations of these substances being transported by a common carrier protein.     

Potassium-Stimulated Release of [3HJTaurine from Cultured GABAergic and Glutamatergic Neurons

The effect of depolarizing concentrations of potassium (56 mM) on the release of [3H]taurine was examined in two types of cultured neurons from mouse brain: cerebral cortex neurons, which are largely GABAergic, and cerebellar neurons, which after treatment with kainate consist almost entirely of glutamatergic granule cells. The release of [3H]taurine was compared to that of gamma-[3H]aminobutyric acid [( 3H]GABA) in cortical neurons and to that of D-[3H]aspartate in granule cells. Cortical neurons responded to potassium stimulation (1 min or continuously) by an immediate increase in [3H]GABA efflux of more than six times over the basal efflux, followed by a sharp decline despite the persistence of the stimulatory agent. The potassium-induced release of [3H]GABA was largely calcium-dependent. The release of [3H]taurine was considerably less in magnitude, only doubling after the stimulus, with a time course delayed in both onset and decline. The release of [3H]taurine was partially calcium-dependent and was also decreased in low-chloride solutions. In cerebellar granule cells, exposure to potassium resulted in a large (sixfold) and prompt release of D-[3H]aspartate, largely calcium-dependent. A totally different pattern was observed for the release of [3H]taurine. A stimulatory effect occurred only when cells were exposed continuously to potassium. Taurine efflux was very delayed, with a broad stimulus plateau reached after 15-20 min of stimulation. Taurine release was unaffected by omission of calcium, but it was abolished in a low-chloride medium. These results suggest that taurine is released from cells handling other neuroactive amino acids as neurotransmitters.(ABSTRACT TRUNCATED AT 250 WORDS)