Header

UZH-Logo

Maintenance Infos

Regulation of Amino Acid Homeostasis in the Brain Interstitial Fluid


Dolgodilina, Elena. Regulation of Amino Acid Homeostasis in the Brain Interstitial Fluid. 2015, University of Zurich, Faculty of Science.

Abstract

Homeostatic control of the surrounding microenvironment is a requirement for proper neuronal function. Their high metabolism requires an efficient supply of nutrients (sugars, amino acids, nucleosides, etc.) and removal of potentially toxic metabolites. A central role in balancing these two processes belongs to the barriers of the central nervous system (CNS). The Blood Brain barrier (BBB) is a dynamic interface between the peripheral circulation and the CNS that tightly regulates transport of compounds into and out of the brain. According to the currently accepted hypothesis the BBB plays a significant role in the production of the brain interstitial fluid (ISF) that directly surrounds neurons and other brain cells. However, since the ISF is only ~ 20% of the total brain volume and is distributed throughout the brain, it is difficult to sample unlike the cerebrospinal fluid (CSF). Therefore, rather less is known about its content. Amino acids (AA) are vitally necessary compounds with multiple functions. Additionally, in the CNS AAs are neurotransmitters or neurotransmitter precursors. For example, glutamine (Gln) serves as a precursor for both the major excitatory (glutamate) and inhibitory (γ-aminobutyric acid (GABA)) neurotransmitters and as a result levels are altered during some pathologies. The conditionally essential AA Gln is the most abundant AA in plasma and CSF. Moreover, in CSF Gln concentration is ~ 80% of that in plasma, while all other standard AAs are ~ 10-fold less concentrated [1]. Nevertheless, data about concentrations of Gln and other AAs in brain ISF are infrequent and inconsistent. The following Gln transporters: LAT1 (SLC7A5), SNAT2 (SLC38A2), SNAT3 (SLC38A3), and SNAT5 (SLC38A5), are significantly expressed in the BBB [2]. Moreover, LAT1 and SNAT3 have been shown to be localized on both luminal and abluminal membranes of BBB endothelial cells [3].

For my dissertation I focused on the brain ISF AAs (in particular Gln) and possible mechanisms of regulation by BBB expressed amino acid transporters (AAT)s. Using in vivo intracranial microdialysis in awake freely moving mice we determined the levels of 14 standard and 2 non-standard AA. For all AAs a ~10 fold concentration gradient between ISF and CSF was observed. Furthermore the acute administration of valine by intraperitoneal (IP) injection caused a significant increase of its concentration not only in plasma, but also in the brain ISF. In contrast, 15N2 Gln (hGln) IP administration resulted in an elevation only in plasma, but not ISF. Nonetheless this labeled form of Gln represented ~ 4% of total Gln in microdialysate samples. Thus, we demonstrated efficient transendothelial transport of both AA from the blood. Competitive inhibition of system L and system A AATs in the brain by inhibitors (2-aminobicyclo-(2,2,1)-heptane-2-carboxylic acid (BCH) and α-(methylamino)-isobutyric acid (MeAIB), respectively) introduced via the microdialysis probe led to Gln increase in the brain ISF. The Gln elevation produced due to BCH infusion was twice higher than that caused by MeAIB. To study whether Gln was released from brain cells or its raise in ISF was due to increased BBB transport, IP injection of 15N2 Gln was combined with BCH administration. The data showed a significant increase of hGln in the brain ISF in presence of BCH in comparison to its IP administration alone indicating that brain perfusion with BCH increased Gln influx via the BBB, presumably by transstimulating Gln uptake via obligatory exchanger LAT1.

Since a high level of Snat5 mRNA was found in BBB its localization in brain endothelial cells (BEC) membranes was studied. Unfortunately, an antibody-dependent approach did not allow distinction of SNAT5 in the BBB endothelium.

Therefore, first of all, a steep AA concentration gradient between the brain ISF and CSF was demonstrated. Second, the data are consistent with LAT1 playing a major role in the regulation of Gln in brain ISF. Since LAT1 is an obligatory exchanger, we suggest it acts together with BBB expressed SNAT3 as key regulators of Gln in brain ISF. We consider that studies using inducible endothelial specific SNAT3 knockout animals will clarify the contributions of BBB expressed SNAT3 in Gln ISF homeostasis.

Abstract

Homeostatic control of the surrounding microenvironment is a requirement for proper neuronal function. Their high metabolism requires an efficient supply of nutrients (sugars, amino acids, nucleosides, etc.) and removal of potentially toxic metabolites. A central role in balancing these two processes belongs to the barriers of the central nervous system (CNS). The Blood Brain barrier (BBB) is a dynamic interface between the peripheral circulation and the CNS that tightly regulates transport of compounds into and out of the brain. According to the currently accepted hypothesis the BBB plays a significant role in the production of the brain interstitial fluid (ISF) that directly surrounds neurons and other brain cells. However, since the ISF is only ~ 20% of the total brain volume and is distributed throughout the brain, it is difficult to sample unlike the cerebrospinal fluid (CSF). Therefore, rather less is known about its content. Amino acids (AA) are vitally necessary compounds with multiple functions. Additionally, in the CNS AAs are neurotransmitters or neurotransmitter precursors. For example, glutamine (Gln) serves as a precursor for both the major excitatory (glutamate) and inhibitory (γ-aminobutyric acid (GABA)) neurotransmitters and as a result levels are altered during some pathologies. The conditionally essential AA Gln is the most abundant AA in plasma and CSF. Moreover, in CSF Gln concentration is ~ 80% of that in plasma, while all other standard AAs are ~ 10-fold less concentrated [1]. Nevertheless, data about concentrations of Gln and other AAs in brain ISF are infrequent and inconsistent. The following Gln transporters: LAT1 (SLC7A5), SNAT2 (SLC38A2), SNAT3 (SLC38A3), and SNAT5 (SLC38A5), are significantly expressed in the BBB [2]. Moreover, LAT1 and SNAT3 have been shown to be localized on both luminal and abluminal membranes of BBB endothelial cells [3].

For my dissertation I focused on the brain ISF AAs (in particular Gln) and possible mechanisms of regulation by BBB expressed amino acid transporters (AAT)s. Using in vivo intracranial microdialysis in awake freely moving mice we determined the levels of 14 standard and 2 non-standard AA. For all AAs a ~10 fold concentration gradient between ISF and CSF was observed. Furthermore the acute administration of valine by intraperitoneal (IP) injection caused a significant increase of its concentration not only in plasma, but also in the brain ISF. In contrast, 15N2 Gln (hGln) IP administration resulted in an elevation only in plasma, but not ISF. Nonetheless this labeled form of Gln represented ~ 4% of total Gln in microdialysate samples. Thus, we demonstrated efficient transendothelial transport of both AA from the blood. Competitive inhibition of system L and system A AATs in the brain by inhibitors (2-aminobicyclo-(2,2,1)-heptane-2-carboxylic acid (BCH) and α-(methylamino)-isobutyric acid (MeAIB), respectively) introduced via the microdialysis probe led to Gln increase in the brain ISF. The Gln elevation produced due to BCH infusion was twice higher than that caused by MeAIB. To study whether Gln was released from brain cells or its raise in ISF was due to increased BBB transport, IP injection of 15N2 Gln was combined with BCH administration. The data showed a significant increase of hGln in the brain ISF in presence of BCH in comparison to its IP administration alone indicating that brain perfusion with BCH increased Gln influx via the BBB, presumably by transstimulating Gln uptake via obligatory exchanger LAT1.

Since a high level of Snat5 mRNA was found in BBB its localization in brain endothelial cells (BEC) membranes was studied. Unfortunately, an antibody-dependent approach did not allow distinction of SNAT5 in the BBB endothelium.

Therefore, first of all, a steep AA concentration gradient between the brain ISF and CSF was demonstrated. Second, the data are consistent with LAT1 playing a major role in the regulation of Gln in brain ISF. Since LAT1 is an obligatory exchanger, we suggest it acts together with BBB expressed SNAT3 as key regulators of Gln in brain ISF. We consider that studies using inducible endothelial specific SNAT3 knockout animals will clarify the contributions of BBB expressed SNAT3 in Gln ISF homeostasis.

Statistics

Additional indexing

Other titles:Dissertation zu Erlangung der naturwissenschaftlichen Doktorwürde (Dr.sc.nat.) vorgelegt der Mathematisch-naturwissenschaftlichen Fakultät der Universität Zürich
Item Type:Dissertation
Referees:Verrey François, Wagner Carsten A, Fritschy Jean-Marc, Engelhardt Britta
Communities & Collections:04 Faculty of Medicine > Institute of Physiology
07 Faculty of Science > Institute of Physiology
Dewey Decimal Classification:570 Life sciences; biology
610 Medicine & health
Language:English
Date:2015
Deposited On:22 Jan 2016 13:56
Last Modified:05 Apr 2016 20:00
Number of Pages:143

Download

Full text not available from this repository.

Article Networks

TrendTerms

TrendTerms displays relevant terms of the abstract of this publication and related documents on a map. The terms and their relations were extracted from ZORA using word statistics. Their timelines are taken from ZORA as well. The bubble size of a term is proportional to the number of documents where the term occurs. Red, orange, yellow and green colors are used for terms that occur in the current document; red indicates high interlinkedness of a term with other terms, orange, yellow and green decreasing interlinkedness. Blue is used for terms that have a relation with the terms in this document, but occur in other documents.
You can navigate and zoom the map. Mouse-hovering a term displays its timeline, clicking it yields the associated documents.

Author Collaborations