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Amino acid transport and receptor binding properties in neuropsychiatric disorders using the fibroblast cell model
Örebro University, School of Health and Medical Sciences.
2011 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Altered transport of the catecholamines and serotonin precursor amino acids tyrosine and tryptophan, might be one explanation for the dysfunctional neurotransmission implicated in the pathophysiology of bipolar disorder and Attention Deficit/Hyperactivity Disorder (ADHD). In previous studies, an altered amino acid transport has been found in schizophrenia and autism, when using the fibroblast cell model. The aim of this thesis was to investigate if the transport of precursor amino acids also may be altered in bipolar disorder and ADHD, and to relate the pre-synaptic activity (transport) with post-synaptic activity (receptors). A functional characterization of tryptophan transport in fibroblasts was also motivated, since the transport of tryptophan in fibroblast cells has not been fully explored.

Fibroblast cell lines from patients with bipolar type-1 disorder, from children with ADHD and from controls were included in the studies. The maximal transport capacity (Vmax) and affinity constant (Km) of tyrosine, tryptophan and alanine transport in bipolar patients and ADHD children were determined. Tryptophan transport characterization included; 1) measuring the uptake of tryptophan at high and low concentrations in the presence or absence of transporter selective inhibitors; 2) determination of Vmax and Km of tryptophan transport at high and low concentrations; 3) sodium dependency studies of tryptophan uptake. All transport studies were done using the cluster tray method. Furthermore, the maximal binding capacity (Bmax) and the equilibrium dissociation constant (KD) of muscarinic acetylcholine receptors (mAChRs) were determined in the ADHD children by a radioligand binding assay, using the mAChRs antagonist QNB.

In patients with bipolar disorder a decreased Vmax in the transport of tyrosine was observed (p=0.027), while the children with ADHD had a decreased Vmax of tryptophan transport (p=0.039) and an increased Vmax of alanine transport (p=0.031). Children with a hereditary ADHD also had a significantly decreased Bmax (p=0.01). The uptake of tryptophan at both high and low concentrations was partly sodium dependent and the inhibitors had different inhibitory effects on the tryptophan uptake. The uptake of tryptophan at high concentration had low affinity and high Vmax, whilst at low concentration the transport was with high affinity and low Vmax.

Altered amino acid transport was observed in fibroblasts of both bipolar disorder patients and ADHD children, which might indicate that the availability of precursor amino acid in the brain is altered. This could lead to disturbances, directly or indirectly, in the catecholaminergic and serotonergic systems. Children with hereditary ADHD might also have reduced levels of mAChRs in the CNS that could indirectly affect the dopaminergic activity. The uptake of tryptophan was through multiple transporters and was different at different substrate concentrations in terms of sodium dependency, affects of inhibitors and kinetic parameters.

Place, publisher, year, edition, pages
Örebro: Örebro universitet , 2011. , p. 68
Series
Örebro Studies in Medicine, ISSN 1652-4063 ; 57
Keywords [en]
Fibroblasts, Bipolar disorder, ADHD, Tyrosine, Tryptophan, Alanine, Transport, mAChRs
National Category
Medical and Health Sciences
Research subject
Biomedicine
Identifiers
URN: urn:nbn:se:oru:diva-17261ISBN: 978-91-7668-818-2 (print)OAI: oai:DiVA.org:oru-17261DiVA, id: diva2:441545
Public defence
2011-10-28, Örebro universitet, Prismahuset, hörsal P2, Fakultetsgatan 1, Örebro, 13:15
Opponent
Supervisors
Available from: 2011-09-16 Created: 2011-09-16 Last updated: 2017-10-17Bibliographically approved
List of papers
1. Aberrant amino acid transport in fibroblasts from patients with bipolar disorder
Open this publication in new window or tab >>Aberrant amino acid transport in fibroblasts from patients with bipolar disorder
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2009 (English)In: Neuroscience Letters, ISSN 0304-3940, E-ISSN 1872-7972, Vol. 457, no 1, p. 49-52Article in journal (Refereed) Published
Abstract [en]

Aberrant tyrosine transport is a repeated finding in fibroblasts from schizophrenic patients. The transport aberration could lead to disturbances in the dopaminergic and noradrenergic neurotransmitter systems. Tyrosine and tryptophan are the precursors of the neurotransmitters dopamine and serotonin. Disturbed dopaminergic, noradrenergic and serotoninergic systems are implicated as causes of bipolar disorder. Hence, the aim of this study was to explore whether patients with bipolar disorder have an aberrant transport of tyrosine and/or tryptophan. Fibroblast cell lines from patients with bipolar type-1 disorder (n = 10) and healthy controls (n = 10) were included in this study. All patients fulfilled the DSM-IV diagnostic criteria. The transport of amino acids across the cell membranes was measured by the cluster tray method. The kinetic parameters, maximal transport velocity (Vmax) and affinity constant (Km) were determined. A significantly lower Vmax for tyrosine (p = 0.027) was found in patients with bipolar type-1 disorder in comparison to healthy controls. No significant differences in Km for tyrosine and in the kinetic parameters of tryptophan between patients with bipolar type-1 disorder and healthy controls were observed. The decreased tyrosine transport (low Vmax) found in this study may indicate less access of dopamine in the brain, resulting in disturbed dopaminergic and/or noradrenergic neurotransmission, that secondarily could lead to disturbances in other central neurotransmitter systems, such as the serotoninergic system. However, as sample size was small in this study and an age difference between patients and controls existed, the present findings should be considered as pilot data. Further studies with larger sample number are needed to elucidate the transport aberration and the significance of these findings.

Place, publisher, year, edition, pages
Elsevier, 2009
National Category
Medical and Health Sciences
Research subject
Biomedicine
Identifiers
urn:nbn:se:oru:diva-6392 (URN)10.1016/j.neulet.2009.03.095 (DOI)000266360300012 ()19429160 (PubMedID)2-s2.0-67349103112 (Scopus ID)
Available from: 2009-04-27 Created: 2009-04-27 Last updated: 2017-12-13Bibliographically approved
2. Tryptophan transport in human fibroblast cells: a functional characterization
Open this publication in new window or tab >>Tryptophan transport in human fibroblast cells: a functional characterization
2011 (English)In: International Journal of Tryptophan Research, ISSN 1178-6469, no 4, p. 19-27Article in journal (Refereed) Published
Abstract [en]

There are indications that serotonergic neurotransmission is disturbed in several psychiatric disorders. One explanation may be disturbed transport of tryptophan (precursor for serotonin synthesis) across cell membranes. Human fibroblast cells offer an advantageous model to study the transport of amino acids across cell membranes, since they are easy to propagate and the environmental factors can be controlled. The aim of this study was to functionally characterize tryptophan transport and to identify the main transporters of tryptophan in fibroblast cell lines from healthy controls.

Tryptophan kinetic parameters (Vmax and Km) at low and high concentrations were measured in fibroblasts using the cluster tray method. Uptake of 3H (5)-L-tryptophan at different concentrations in the presence and absence of excess concentrations of inhibitors or combinations of inhibitors of amino acid transporters were also measured. Tryptophan transport at high concentration (0.5 mM) had low affinity and high Vmax and the LAT1 isoform of system-L was responsible for approximately 40% of the total uptake of tryptophan. In comparison, tryptophan transport at low concentration (50 nM) had higher affinity, lower Vmax and approximately 80% of tryptophan uptake was transported by system-L with LAT1 as the major isoform. The uptake of tryptophan at the low concentration was mainly sodium (Na+) dependent, while uptake at high substrate concentration was mainly Na+ independent. A series of different transporter inhibitors had varying inhibitory effects on tryptophan uptake.

This study indicates that tryptophan is transported by multiple transporters that are active at different substrate concentrations in human fibroblast cells. The tryptophan transport trough system-L was mainly facilitated by the LAT1 isoform, at both low and high substrate concentrations of tryptophan.

Place, publisher, year, edition, pages
Libertas Academica Ltd., 2011
Keywords
Fibroblasts, tryptophan, serotonin, LAT1, amino acid transporters
National Category
Medical and Health Sciences
Research subject
Biomedicine
Identifiers
urn:nbn:se:oru:diva-17365 (URN)10.4137/IJTR.S6913 (DOI)
Available from: 2011-09-27 Created: 2011-09-27 Last updated: 2018-05-03Bibliographically approved
3. Altered tryptophan and alanine transport in fibroblasts from boys with Attention Deficit/Hyperactivity Disorder (ADHD): an in vitro study
Open this publication in new window or tab >>Altered tryptophan and alanine transport in fibroblasts from boys with Attention Deficit/Hyperactivity Disorder (ADHD): an in vitro study
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(English)Manuscript (preprint) (Other academic)
National Category
Medical and Health Sciences
Research subject
Biomedicine
Identifiers
urn:nbn:se:oru:diva-17366 (URN)
Available from: 2011-09-27 Created: 2011-09-27 Last updated: 2017-10-17Bibliographically approved
4. Decreased density of muscarinic acetylcholine receptors in fibroblasts from children with Attention Deficit/Hyperactivity Disorder (ADHD)
Open this publication in new window or tab >>Decreased density of muscarinic acetylcholine receptors in fibroblasts from children with Attention Deficit/Hyperactivity Disorder (ADHD)
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(English)Manuscript (preprint) (Other academic)
National Category
Medical and Health Sciences
Research subject
Biomedicine
Identifiers
urn:nbn:se:oru:diva-17367 (URN)
Available from: 2011-09-27 Created: 2011-09-27 Last updated: 2017-10-17Bibliographically approved

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