Levels of amino acids in 52 discrete areas of postmortem brain of adult and aged humans

Summary In a series of studies we have analyzed the regional distribution of the free amino acid pool in 52 discrete areas of postmortem brain of adult and aged humans. Here we show the distribution of eleven amino acids: alanine, methionine, valine, leucine, isoleucine, glutamine, asparagine, lysine, arginine, ornithine, and histidine. As found previously for other amino acids, the distribution of these amino acids was seen to be heterogeneous, the level of the area of highest level being 3.4 to 10.7 times that of the area of the lowest level. On average we found a five- or six-fold difference in concentration between the highest and lowest level areas in the brain samples from adult and old respectively. The distribution patterns were found to be different for each amino acid; they were not similar even in the same class (amides, branched chain, basic amino acids), and they were different from those recently found in rat brain. Only a few changes, mostly increases, were found in the aged brain, such as increases in alanine and valine levels in cortical areas. In studies of changes in cerebral amino acid levels, the great regional heterogeneity of distribution has to be taken into account since changes in whole brain values may not reflect regional changes. The functional significance and the control of this regional heterogeneity are under investigation.     

Changes with aging in the levels of amino acids in rat CNS structural elements II. Taurine and small neutral amino acids

Taurine (Tau) and the small neutral amino acids glycine (Gly), serine (Ser), threonine (Thr), and alanine (Ala) were measured in 53 brain areas of 3- and 29-month-old male Fisher 344 rats. The ratio of highest to lowest level was 34 for Tau, 9.1 for Thr, 7.6 for Gly and Ser, and 6.5 for Ala. The heterogeneity was found in numerous areas; for example, Tau levels were more than 90 nmol/mg protein in 6 areas, and less than 20 nmol/mg protein in 10 areas. Similar heterogeneity was found with the other amino acids. The relative distribution of the small neutral amino acids showed several similarities; Tau distribution was different. With age, four amino acids decreased in 10–18 areas, and increased in only 1–3, while Thr increased in more areas than it decreased. The five amino acids of this paper, and the four of the previous paper, are among the amino acids at highest level in the brain; the sequence in their levels shows considerable regional heterogeneity.     

Long-lasting effects of audiogenic seizures on neurotransmitter amino acids in Rb mice

The existence of long-lasting (15–18 h) alterations of neurotrasmitter amino acid levels following a single or repeated acoustic stimulations in audiogenic seizure-prone Rb1 and Rb2 mice and suizure-resistant Rb3 mice were investigated. The levels of glutamate, aspartate, glycine, taurine, and of some of their precursors: glutamine and serine were determined. Fourteen brain areas were examined. Alterations were found only in 6 brain areas (pons, olfactory bulbs, superior colliculus, inferior colliculus, olfactory tubercles and raphe). Most frequent occuring changes were observed in pons and olfactory tubercles. These changes concerned mainly the excitatory amino acids, glutamate, and aspartate. Alterations of taurine, glycine and serine were also recorded.Abbreviations GABA 4-aminobutyrate - Tau taurine - Gly glycine - Asp aspertate - Glu glutamate - Gln glutamine - Ser serine - OB olfactory bulbs - OT olfactory tubercles - Sr striatum - Se septum - Hy hypothalamus - Th thalamus - Hi hippocampus - A amygdala - SC superior colliculus - IC inferior colliculus - FC frontal cortex - C cerebellum - P pons medulla - Ra raphe - AA neurotransmitter amino acids - I inhibitory - E excitatory - SSL steady-state level Plesant memories of Lawrence Austin's sojourn in my group at Strasbourg gather upon me when I dedicate this article on this occasion for the contribution that Lawrence Austin has made for the cause of neurochemical researchers.     

Changes in regional levels of putative neurotransmitter amino acids in brain under unilateral forebrain ischemia

The levels of the neurotransmitter amino acids glutamate, aspartate, and GABA were determined in different brain regions during ischemia and post-ischemic recirculation periods using the unilateral carotid artery occlusion model of stroke in gerbils. The levels of glutamate, aspartate and GABA in ischemic hemisphere were increased significantly by 10 min of ischemia and later declined with time. Reperfusion for 30 min following 10 min. of ischemia further enhanced the levels of glutamate and aspartate. Increase in GABA levels were found during early periods of reperfusion. Regional variations in the changes of amino acids' levels were noticed following ischemia. Hippocampus showed the highest increase in glutamate levels followed by striatum and cerebral cortex. Aspartate levels in striatum and hippocampus increased during 10 min ischemia (46% and 30%) and recirculation (70% and 79%), whereas in cerebral cortex the levels were doubled only during recirculation. Ischemia induced elevations of GABA levels were observed in cerebral cortex (68%) and in hippocampus (30%), and the levels were normalized during recirculation. No changes in GABA levels were found in striatum. It is suggested that the large increase in the levels of excitatory neurotransmitter amino acids in brain regions specially in hippocampus during ischemia and recirculation may be one of the causal factors for ischemic brain damage.     

Changes in brain temperature and free amino acids in normal and protein deprived suckling rats exposed to room temperature

Previous reports on early-induced protein-calorie malnutrition (PCM) in rats have indicated alterations in the concentration of free amino acids and of protein synthesis in the brain. Recently it was shown that early-induced protein deprivation (PD) retards the development of thermoregulation. This resulted in a failure to maintain a normal rectal temperature after short exposure to room temperature (+22°C) still at the age of 20–25 days corresponding to changes seen in normal rats at an age of 10–15 days. In the present study, 20-day old PD and normal rats where examined with regard to the effect of exposure to room temperature on brain temperature and on brain free amino acids. The results show a similar reduction in brain and rectal temperature of the PD rats occuring within 30 minutes after exposure to room temperature. The reduction was in the range of 5°C. PD rats kept in room temperature for 5 hours and then allowed to recover at 32.5°C showed a slow increase in brain and rectal temperature but normal temperatures were not reached even after 1 hour. The concentration of free amino acids in the brain was examined in rats kept for 1 hour at room temperature or at 32.5°C. In the PD rats kept at 32.5°C, free aspartate and glutamate were reduced whereas taurine, GABA and glycine were increased as compared to their corresponding control rats. As a result of the reduced brain temperature in PD rats exposed to room temperature there was a reduction in free asparagine. The lability of the pool of asparagine may be related to the low levels of aspartate and glutamate in PD rats. On the basis of the present findings it is recommended that temperature-sensitive parameters are examined in PCM rats at a normal body temperature.Special Issue dedicated to Prof. Holger Hydén     

Changes with aging in the levels of amino acids in rat CNS structural elements I. Glutamate and related amino acids

Glutamate and related amino acids were determined in 53 discrete brain areas of 3-and 29-month-old male Fischer 344 rats microdissected with the punch technique. The levels of amino acids showed high regional variation-the ratio of the highest to lowest level was 9 for aspartate, 5 for glutamate, 6 for glutamine, and 21 for GABA. Several areas were found to have all four amino acids at very high or at very low level, but also some areas had some amino acids at high, others at low level. With age, in more than half of the areas, significant changes could be observed, decrease occurred 5 times more frequently than increase. Changes occurred more often in levels of aspartate and GABA than in those of glutamate or glutamine. The regional levels of glutamate and its related amino acids show severalfold variations, with the levels tending to decrease in the aged brain.     

Effect of Altered Blood Plasma Osmolalities on Regional Brain Amino Acid Concentrations and Focal Seizure Susceptibility in the Rat

Blood plasma hypo- or hyperosmolality alters significantly the concentration of some amino acids in brain tissues of the medial septum and hippocampus of adult Sprague-Dawley rats. With some notable exceptions, brain amino acid concentrations decreased under hypoosmotic conditions and increased under hyperosmotic conditions. Osmotic changes and brain amino acid changes appear to be related to each other in an almost linear fashion. A comparison of rats and toads indicates that the patterns of changes in brain amino acid concentrations in response to a hypoosmotic plasma osmolality were almost identical for both species. Changes achievable under hyperosmotic conditions were considerably greater in toads. When rats with kindled epileptogenic foci were made hypoosmotic by water-loading, seizure thresholds decreased dramatically. Our data suggest a possible relationship between the hypoosmotically induced biochemical changes in brain tissues (especially some amino acid neurotransmitters and neurotransmitter precursors) and the hypoosmotically induced increase in seizure susceptibility.     

Serum amino acid profiles and dopamine in schizophrenic patients and healthy subjects: Window to the brain?

Summary Altered dopamine turnover has been postulated as underlying cause for schizophrenia. This is partially inferred from pharmacological studies and from changes in serum dopamine and dopamine metabolite levels. It is not clear whether the serum amino acid precursors' availability and neurotransmitter-mediated hormonal release could be indicative of the neurotransmitter turnover. We speculate in this context that the profile of serum amino acids and neurotransmitters reflects differences of neurotransmitter activity in the central nervous system and may be considered in a broad sense window to the brain.We analyzed basal serum amino acids (including monoamine precursors), and monoamines in schizophrenic patients after a drug holiday of 3 or more days, and in healthy subjects.Asparagine, phenylalanine, and cystine were higher and tyrosine, tryptophan, and the ratio of tryptophan to competing amino acids lower in schizophrenic patients than in healthy subjects (P < 0.05). Dopamine was increased in schizophrenic patients compared to healthy subjects.We speculate that these results sustain the notion for dopamine overactivity in schizophrenia, which might be caused by altered amino acid precursor availability.Presented at the 2nd International Congress on Amino Acids and Analogues, Vienna, Austria August 5–9, 1991     

Amino acid neurotransmitter alterations in three sublines of Rb mice differing by their susceptibility to audiogenic seizures

The levels of inhibitory amino acids (Tau, Gly), or excitatory amino acids (Glu, Asp) and Gln, precursor of GABA, have been determined, under resting conditions, in 17 brain areas of 3 sublines of inbred Rb mice displaying different responses to an acoustic stimulus. Rb1 mice were clonictonic seizure-prone, Rb2 mice were clonic seizure-prone and Rb3 mice were seizure resistant. Profile of distribution in the brain of each one of these amino acids differed. Maximum to minimum level ratio was higher for Tau (3.8) than for Glu or Asp or Gln (2). The level of Gly was similar in 13 out of the 17 areas examined. Multiple inter-subline differences were recorded for each amino acid. These differences have been analyzed considering the seizure susceptibility or severity of the three Rb sublines. Common lower levels (approximately –20%: Rb1/Rb3, Rb2/Rb3) of Gln in Temporal Cortex may be implicated in seizure susceptibility. Seirure severity (Rb1/Rb2) seems to correlate, in some areas, with additional lower amounts of GABA already reported and, to a lower extent, of Asp (–19% in striatum, inferior colliculus and cerebellum), of Tau and Gly; a tendency for a rise in Gln content was observed in certain others (10–20% in olfactory bulb, thalamus, hypothalamus, substantia nigra, and frontal, temporal and occipital cortex). The data and correlations recorded provide guidelines for further investigations for synaptosomal and metabolic alterations in the three sublines of the same strain of Rb mice.Abbreviations used GABA 4-aminobutyrate - Tau taurine - Gly glycine - Asp aspartate - Glu glutamate - Gln glutamine - GEPR genetically epilepsy-prone rat - OB olfactory bulbs - OT olfactory tubercles - Sr striatum - Se septum - Hy hypothalamus - Hi hippocampus - Th thalamus - A amygdala - SC superior colliculus - IC interior colliculus - SN substantia nigra - FCx frontal cortex - TCx temporal cortex - OCx occipital cortex - C cerebellum - P pons - Ra raphe     

A Regional Study of 4-Aminobutyrate Metabolism and Amino Acid Levels in Rat Brain Following Chronic Oral Administration of Ethanolamine O-Sulphate

Abstract: Ethanolamine O-sulphate (EOS) dissolved in the drinking water (5mg-ml⁻¹) was administered ad libitum to rats for 26 days. At the end of this period, glutamate decarboxylase (GAD) and GABA-transaminase (GABA-T) activities, 4-aminobutyrate (GABA) concentration, and the levels of six other amino acids were measured in various brain regions. Significant inhibition of GABA-T accompanied by significant increases in GABA content were observed throughout the brain, although the magnitudes of these effects varied according to region. GAD activity was significantly reduced in most brain regions, although this effect was apparently not related to cofactor availability or the direct actions of EOS or increased GABA concentration. Glutamine levels were significantly reduced to approximately 72% of control values in all brain regions. Aspartate levels were significantly reduced to approximately 84% of control values in all regions except the striatum and cerebellum. Minor changes in other amino acid levels were also detected. These neurochemical changes which accompanied the primary effect of EOS on GABA-T are discussed in terms of indirect secondary metabolic changes rather than nonspecific enzyme inhibition by EOS.     

Partitioning of 14C-Photosynthate of Leaves in Roots,Rhizome, and in Essential Oil and Curcumin in Turmeric (Curcuma longa L.)

Incorporation of photosynthetically fixed ¹⁴C was studied at different time intervals of 12, 24, and 36 h in various plant parts—leaf 1 to 4 from apex, roots, and rhizome—into primary metabolites—sugars, amino acids, and organic acids, and secondary metabolites—essential oil and curcumin—in turmeric. The youngest leaves were most active in fixing ¹⁴C at 24 h. Fixation capacity into primary metabolites decreased with leaf position and time. The primary metabolite levels in leaves were maximal in sugars and organic acids and lowest in amino acids. Roots as well as rhizome received maximum photoassimilate from leaves at 24 h; this declined with time. The maximum metabolite concentrations in the roots and rhizome were high in sugars and organic acids and least in amino acids. ¹⁴C incorporation into oil in leaf and into curcumin in rhizome was maximal at 24 h and declined with time. These studies highlight importance of time-dependent translocation of ¹⁴C-primary metabolites from leaves to roots and rhizome and their subsequent biosynthesis into secondary metabolite, curcumin, in rhizome. This might be one of factors regulating the secondary metabolite accumulation and rhizome development.     

Co-variation of free amino acids in human epileptogenic cortex

The concentration of free amino acids was measured in 41 surgically removed samples of human epileptogenic brain and in 7 specimens of non-epileptic brain tissue, removed during surgery for meningiomas, etc. The material was subdivided according to the neuropathological diagnosis: mild cortical dysplasia (MCD), gliosis astrocytoma infiltration and a histologically heterogeneous group. The non-tumoral epileptogenic samples had five times higher than normal concentration of ethanolamine and 50% elevated concentration of glycine. The concentration of other neurotransmitter amino acids did not differ markedly between epileptogenic and non-epileptic samples. The concentration of neurotransmitter amino acids showed a strong correlation with the enzyme neuron specific enolase (NSE) and were low in most samples with astrocytoma infiltration. On the other hand, tyrosine and leucine had higher concentrations in samples with lower NSE concentration. Factor analysis of the amino acids revealed four groups of covarying compounds in the brain samples, first, a neurotransmitter group, including aspartate, glutamate, GABA and phosphoethanolamine. Another group contained ethanolamine, glutamine, glycine and taurine. Factor analysis on corresponding extracellular amino acids showed two groups, the first being a neurotransmitter group, containing serine, taurine phosphoethanolamine and ethanolamine in addition to aspartate and glutamate. The other group consisted of asparagine, glycine, alanine, tyrosine, valine, phenylalanine, isoleucine and leucine.Special issue dedicated to Dr. Claude Baxter.     

Effect of Impact Trauma on Neurotransmitter and Nonneurotransmitter Amino Acids in Rat Spinal Cord

N-Methyl-D-aspartate (NMDA) administration exacerbates neurological dysfunction after traumatic spinal cord injury in rats, whereas NMDA antagonists improve outcome in this model. These observations suggest that release of excitatory amino acids contributes to secondary tissue damage after traumatic spinal cord injury. To further examine this hypothesis, concentrations of free amino acids were measured in spinal cord samples from anesthetized rats subjected to various degrees of impact trauma to the T9 spinal segment. Levels of excitatory and inhibitory neurotransmitter amino acids [gamma-aminobutyric acid (GABA), glutamate, aspartate, glycine, taurine] and levels of nonneurotransmitter amino acids (asparagine, glutamine, alanine, threonine, serine) were determined at 5 min, 4 h, and 24 h posttrauma. Uninjured surgical (laminectomy) control animals showed modest but significant declines in aspartate and glutamate levels, but not in other amino acids, at all time points. In injured animals, the excitatory amino acids glutamate and aspartate were significantly decreased by 5 min posttrauma, and remained depressed at 4 h and 24 h as compared with corresponding laminectomy controls. In contrast, the inhibitory amino acids, glycine, GABA, and taurine, were decreased at 5 min postinjury, had partially recovered at 4 h, and were almost fully recovered at 24 h. The nonneurotransmitter amino acids were unchanged at 5 min posttrauma and significantly increased at 4 h, with partial recovery at 24 h. At 4 h postinjury, severe trauma caused significantly greater decreases in aspartate and glutamate than did either mild or moderate injury. These findings are consistent with the postulated role of excitatory amino acids in CNS trauma.     

Changes in Free Amino Acid Levels in Developing Human Foetal Brain Regions

The levels of free amino acids were determined in human foetal brain regions during prenatal development. Variation in the distribution of amino acids and their rate of change in five segments of the CNS at different stages of ontogeny was observed. Striking developmental changes were found in the levels of aspartic acid in medulla-pons and spinal cord, glycine in the spinal cord, gamma-aminobutyric acid in the cerebral cortex, glutamic acid in the cerebral cortex, midbrain, and spinal cord, and taurine in the medulla-pons and spinal cord. At a late gestational period, glutamic acid was found most abundantly over all the brain regions, whereas the level of taurine was highest at an early gestational stage but not in spinal cord.     

Perinatal changes of transport systems for amino acids in slices of mouse brain

Perinatal changes in the uptake of amino acids were measured in slices of fetal (15- and 19-day) and newborn (4-, 24-, and 48-hr-old) mouse brain. Uptake increased with age; smaller changes occurred with basic and neutral amino acid transport systems, and the largest changes occurred in fetal brain with amino acids of putative neurotransmitter function (taurine, glycine, GABA, and the acidic amino acids). The pattern of increase in uptake was similar at high and at low external amino acid concentrations. Developmental changes in tissue content of Na⁺, K⁺, or ATP were small during this period, and so are unlikely to be responsible for the observed changes in uptake. It appears that by the 15th day of fetal life, the transport systems for essential amino acids are fairly well developed in the brain, and the transport systems for neurotransmitter amino acids are not so well developed, but undergo a rapid increase in the 15–19-day period. From birth to adulthood, the concentrative capacity of slices of mouse brain for nonessential (putative neurotransmitter) amino acids is much greater than for essential amino acids.This research was supported in part by NIH Grant No. RR05707.     

Effects of Dietary Amino Acids on Brain Amino Acids and Transmitter Amines in Rats with a Portacaval Shunt

The etiologic relationship between disturbances in metabolism of amino acids and amines and hepatic coma was investigated by examining the effects of diets containing various mixtures of amino acids on brain amine metabolism in rats with a portacaval shunt, using a method for simultaneous analysis of amino acids and amines. Rats with a portacaval shunt were fed on four different amino acid compositions with increased amounts of various amino acids suspected to be etiologically related to hepatic coma, such as methionine, phenylalanine, tyrosine, and tryptophan. The animals were killed 4 weeks after operation. During the experimental period, these animals did not become comatose, but exhibited various behavioral abnormalities. Marked increase in the plasma and brain levels of the augmented amino acids, especially methionine and tyrosine, were observed in rats with a portacaval shunt. Brain noradrenaline, dopamine, and serotonin levels were significantly decreased when the brain tyrosine level was increased. These results indicate that in rats with a portacaval shunt the dietary levels of amino acids greatly influence the brain levels of both amino acids and transmitter amines.     

Amino Acid Levels and γ-Aminobutyric AcidA Receptors in Rat Neostriatum, Cortex, and Thalamus After Neonatal 6-Hydroxydopamine Lesion

Abstract: The amino acid γ-aminobutyric acid (GABA) is the main inhibitory neurotransmitter in brain, and GABAergic neurons have been proposed to play a major role in basal ganglia physiology. In the neostriatum (caudate putamen), medium-sized aspiny interneurons, as well as neostriatal output neurons that project to several brain regions, use GABA as their neurotransmitter. Dopamine fibers arising from the substantia nigra represent a major input to the neostriatum where, besides their classic neurotransmitter role, they are seemingly involved in the regulation of amino acid neurotransmitter release. To further characterize the nature of some of the amino acid/dopamine interactions, selective dopaminergic deafferentations were produced in neonatal rats (3 days postnatal) by intraventricular administration of the neurotoxin 6-hydroxydopamine (6-OHDA); the noradrenergic neurons were protected by prior administration of desmethylimipramine. After a 3-month survival, levels of catecholamines, indoleamines, and amino acids were determined in cingulate cortex, thalamus, and neostriatum. In addition, GABA_A receptors were measured in membrane preparations from these three regions, using the specific agonist [³H]muscimol. In the 6-hydroxydopamine-lesioned rats, levels of dopamine and its metabolites homovanillic acid, 3,4-dihydroxyphenylacetic acid, and 3-methoxytyramine were decreased, as expected, in cortex and neostriatum, but remained unmodified in thalamus. In all three regions, serotonin content was increased; its metabolite, 5-hydroxyindole-3-acetic acid, was also elevated, but only in cortex and neostriatum. The levels of GABA were increased in neostriatum and thalamus, but remained unmodified in cortex. Glycine was increased in all three regions examined. There were also increases of phosphatylethanolamine and serine in thalamus, and of aspartic acid and alanine in neostriatum. The density of GABA_A binding sites was increased in neostriatum, but remained unchanged in cortex and thalamus. The changes in amino acid levels and [³H]muscimol binding sites induced by a neonatal 6-hydroxydopamine treatment differ from those found after similar lesions in adult animals, possibly because of the plastic and synaptic rearrangements that can still occur during early postnatal development. The present results also demonstrate that adaptations occur in response to a dopaminergic deafferentation at an early age and that these exhibit a regional specificity.     

The establishment of essential and metabolically derived amino acid profiles in developing chick embryo organs

The accumulation of certain essential and metabolically derived amino acids in the free amino acid pools of three excitable tissues has been studied in the chick embryo. Valine together with leucine are at the onset present in the yolk at higher concentrations than any of the other essential amino acids. By 15 days all the amino acids studied have accumulated in the embryonic pools at a higher rate than valine, although certain amino acids, such as phenylalanine or methionine, always remain at lower relative concentrations than valine. This reflects their low supply in the yolk, rather than a more rapid rate of disappearance (utilization). During early embryogenesis (E₂–E₄), tissues preferentially concentrate glutamic acid, besides taurine and phosphoethanolamine (6). The next distinct stage of development (E₄–E₇) is marked in the brain by a gradual rise in glutamic acid, glutamine and aspartic acid; the same three amino, acids do not demonstrate a further increase in the pool of the heart, while in the whole eye the amino acid profile begins to resemble the blood. Leucine in all three tissues declines rapidly, to reach isoleucine levels by day 7 of development; tyrosine increases slowly in apparent reciprocity to an equally gradual phenylalanine decrease. Into the second week of embryo growth (E₇–E₁₅), GABA appears in the mesencephalon (E₇) and the eye (E₉–E₁₀). In the mesencephalon, the free amino acid pool composition exhibits a rather sudden increase of most metabolically derived amino acids. Glutamic acid and glutamine in the brain increase in parallel; the rate of GABA and aspartic acid accumulation is slower, and for GABA stabilizes on day 14, as does glutamine. In the eye, by day 15, GABA levels are more closely aligned with the aspartic acid content. Finally, throughout embryogenesis serine fluctuations in blood and tissues are in parallel with those of threonine, and different from glycine or alanine which also change in tandem.     

Regional distribution of biogenic amines, amino acids and cholinergic markers in the CNS of the C57BL/6 strain

Summary. A reliable extrapolation of neurochemical alterations from a mouse model to human metabolic brain disease requires knowledge of neurotransmitter levels and related compounds in control mouse brain. C57BL/6 is a widely used background strain for knockout and transgenic mouse models. A prerequisite for reliable extrapolation from mouse brain to the human condition is the existence of analogous distribution patterns of neurotransmitters and related compounds in control mouse and human brain. We analysed regional distribution patterns of biogenic amines, neurotransmitter and non-neurotransmitter amino acids, and cholinergic markers. Distribution patterns were compared with known neurotransmitter pathways in human brain. The present study provides a reference work for future analyses of neurotransmitters and related compounds in mouse models bred in a C57BL/6 background strain.