Amino Acid Transport in Isolated Neurons and Glia

Hamberger, Anders; Nyström, Britta; Sellström, Åke; Woiler, C. T.

doi:10.1007/978-1-4684-3264-0_17

Cited by 29 publications

(6 citation statements)

References 51 publications

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“…There is no doubt that the neuropil samples constituted less pure glial samples than present-day astrocytic preparations, but nevertheless many of Hydén’s observations are presently being confirmed. A different technique to separate neurons and glia, gradient centrifugation ( 121 ) was used in his laboratory by Hamberger and coworkers to show glial uptake of glutamate and GABA for the first time and many features of their subsequent metabolic fate ( 122 , 123 ). Such studies are suitable for determination of K m values, but the cells are too damaged to evaluate V max .…”

Section: Cell Separation Techniques Bac Transgenic Mice and Methodomentioning

confidence: 99%

Methodological Limitations in Determining Astrocytic Gene Expression

et al. 2013

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Traditionally, astrocytic mRNA and protein expression are studied by in situ hybridization (ISH) and immunohistochemically. This led to the concept that astrocytes lack aralar, a component of the malate-aspartate-shuttle. At least similar aralar mRNA and protein expression in astrocytes and neurons isolated by fluorescence-assisted cell sorting (FACS) reversed this opinion. Demonstration of expression of other astrocytic genes may also be erroneous. Literature data based on morphological methods were therefore compared with mRNA expression in cells obtained by recently developed methods for determination of cell-specific gene expression. All Na,K-ATPase-α subunits were demonstrated by immunohistochemistry (IHC), but there are problems with the cotransporter NKCC1. Glutamate and GABA transporter gene expression was well determined immunohistochemically. The same applies to expression of many genes of glucose metabolism, whereas a single study based on findings in bacterial artificial chromosome (BAC) transgenic animals showed very low astrocytic expression of hexokinase. Gene expression of the equilibrative nucleoside transporters ENT1 and ENT2 was recognized by ISH, but ENT3 was not. The same applies to the concentrative transporters CNT2 and CNT3. All were clearly expressed in FACS-isolated cells, followed by biochemical analysis. ENT3 was enriched in astrocytes. Expression of many nucleoside transporter genes were shown by microarray analysis, whereas other important genes were not. Results in cultured astrocytes resembled those obtained by FACS. These findings call for reappraisal of cellular nucleoside transporter expression. FACS cell yield is small. Further development of cell separation methods to render methods more easily available and less animal and cost consuming and parallel studies of astrocytic mRNA and protein expression by ISH/IHC and other methods are necessary, but new methods also need to be thoroughly checked.

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Section: Cell Separation Techniques Bac Transgenic Mice and Methodomentioning

confidence: 99%

Methodological Limitations in Determining Astrocytic Gene Expression

et al. 2013

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“…Uptake sites for amino-acid transmitters have been identified on glial cells, and it is expected that these cells play an important role in the uptake processes of these transmitters (Hamberger et al, 1976;Iversen and Kelly, 1975). Glutamate is metabolized in astrocytes to glutamine and subsequently channeled back to the neurons.…”

Section: Nonneuronal Poolsmentioning

confidence: 99%

Brain microdialysis of GABA and glutamate: What does it signify?

Timmerman

Westerink

1997

Synapse

375

194

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Microdialysis has become a frequently used method to study extracellular levels of GABA and glutamate in the central nervous system. However, the fact that the major part of GABA and glutamate as measured by microdialysis does not fulfill the classical criteria for exocytotic release questions the vesicular origin of the amino acids in dialysates. Glial metabolism or reversal of the (re)uptake sites has been suggested to be responsible for the pool of nonexocytotically released amino-acid transmitters that seem to predominate over the neuronal exocytotic pool. The origin of extracellular GABA and glutamate levels and, as a consequence, the implications of changes in these levels upon manipulations are therefore obscure. This review critically analyzes what microdialysis data signify, i.e., whether amino-acid neurotransmitters sampled by microdialysis represent synaptic release, carrier-mediated release, or glial metabolism. The basal levels of GABA and glutamate are virtually tetrodotoxin- and calcium-independent. Given the fact that evidence for nonexocytotic release mediated by reversal of the uptake sites as a release mechanism relevant for normal neurotransmission is so far limited to conditions of "excessive stimulation," basal levels most likely reflect a nonneuronal pool of amino acids. Extracellular GABA and glutamate concentrations can be enhanced by a wide variety of pharmacological and physiological manipulations. However, it is presently impossible to ascertain that the stimulated GABA and glutamate in dialysates are of neuronal origin. On the other hand, under certain stimulatory conditions, increases in amino-acid transmitters can be obtained in the presence of tetrodotoxin, again suggesting that aspecific factors not directly related to neurotransmission underlie these changes in extracellular levels. It is concluded that synaptic transmission of GABA and glutamate is strictly compartmentalized and as a result, these amino acids can hardly leak out of the synaptic cleft and reach the extracellular space where the dialysis probe samples.

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“…The alcohol-induced increase in K + efflux increases glial depolarization [45] and glucose demand by neurons and astrocytes as additional glucose is required by astrocytes to take up the excess K + and store it [46]. An increase in K + efflux can stimulate GABA release [47], in order to protect against brain impairment caused by an increase in neuronal depolarization [48]. Alcohol's potentiation of GABA is specifically linked to the GABA A receptor gated chloride channel [49] and this may be due to Clbeing needed by astrocytes in order to maintain electrical neutrality [46].…”

Section: Discussionmentioning

confidence: 99%

“…An increase in K + efflux could eventually develop into an extracellular acidosis [50], which in turn could promote a significant alteration in the activity of the HPA axis as a result of chronic osmotic stimulation [51]. A decrease in HPA axis activity under these conditions would be beneficial to the survival of the organism because cortisol decreases GABA activity at higher concentrations [52,53], which could potentially increase the risk of brain injury due to an increase in neuronal depolarization [48]. Additionally, cortisol (similar to alcohol) can promote the loss of body potassium due to the release of K + from intracellular stores [41] and reduce glucose utilization and transport in neurons and astrocytes [54][55][56], which could increase the risk of hypoglycemia and hypoxia [57].…”

Section: Discussionmentioning

confidence: 99%

Salivary or Serum Cortisol: Possible Implications for Alcohol Research

Kokavec¹

2012

Steroids - From Physiology to Clinical Medicine

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Alcohol consumption can induce the development of nutritional disorders as alcohol ingestion often replaces food intake [ ]. The long-term intake of alcohol decreases the amount of food consumed when food is freely available [ ], and the degree of malnutrition may be related to the irregularity of feeding habits and intensity of alcohol intake [ ]. The repercussions of alcohol abuse over time can involve damage to most of the major organs and systems in the body [ ]. However, despite the overwhelming evidence linking alcohol to ill health the role if any alcohol plays in the development of disease remains uncertain.The hypothalamic-pituitary-adrenal HPA axis is responsible for the synthesis and release of steroid hormones, the most abundant being dehydroepiandrosterone DHEA , DHEA sulfate DHEAS , cortisol, and aldosterone [e.g. ]. The release of either corticotropin-releasing factor or arguinine vasopressin by the hypothalamus stimulates the anterior pituitary to release adrenocorticotropin ACTH , which promotes the synthesis and release of steroid hormones that have glucocorticoid i.e. cortisol , mineralocorticoid i.e. aldosterone , and androgenic i.e. DHEA, DHEAS functions [ ].Steroid hormones have a diverse and highly important role in the body and any dysregulation in steroid activity can lead to the development of disease. The adrenocortical system is markedly altered by food availability and an elevation in cortisol is commonly observed under fasting conditions [ -]. Cortisol plays a major role in the regulation of carbohydrate, protein, and lipid metabolism [ , ] and during prolonged fasting by stimulating gluconeogenesis acts to protect the body from cellular damage until food once again becomes available [ , , -].© 2012 Kokavec; licensee InTech. This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.It is well accepted that alcohol consumption can significantly reduce DHEAS [ , ] and aldosterone [ , ]. However, the literature is highly contradictory with respect to the effect of oral alcohol intake on cortisol. Investigations have shown that blood alcohol concentrations exceeding g/L can elevate plasma cortisol [ ]. Moreover, it has also been reported that low to moderate alcohol intake has little [ ], if any effect on [ -] or may even significantly reduce [ ], cortisol concentration. Furthermore, early work showed that while alcohol consumption may promote a significant decrease in cortisol initially this is later followed by a significant elevation in plasma cortisol concentration [ ].In the past it has been proposed that the discrepancy in cortisol, noted in the alcohol literature, could be due to differences in stress levels associated with the testing procedure [ ].Cortisol is rapidly released in response to stress and the stress associated with blood sampling alone can falsely incr...

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Amino Acid Transport in Isolated Neurons and Glia

Cited by 29 publications

References 51 publications

Methodological Limitations in Determining Astrocytic Gene Expression

Methodological Limitations in Determining Astrocytic Gene Expression

Brain microdialysis of GABA and glutamate: What does it signify?

Salivary or Serum Cortisol: Possible Implications for Alcohol Research

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