Membrane-bound catechol-O-methyl transferase in cortical neurons and glial cells is intracellularly oriented
Catechol-O-methyl transferase (COMT) is involved in the inactivation of dopamine in brain regions in which the dopamine transporter (DAT1) is sparsely expressed. The membrane-bound isoform of COMT (MB-COMT) is the predominantly expressed form in the mammalian central nervous system (CNS). It has bee...
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doaj-0e22a2d14a0e44bcbed48377bbe267c82020-11-25T01:03:25ZengFrontiers Media S.A.Frontiers in Psychiatry1664-06402010-10-01110.3389/fpsyt.2010.001422180Membrane-bound catechol-O-methyl transferase in cortical neurons and glial cells is intracellularly orientedBjörn H Schott0Björn H Schott1Björn H Schott2Renato Frischknecht3Grazyna eDebska-Vielhaber4Grazyna eDebska-Vielhaber5Nora John6Gusalija Behnisch7Emrah eDüzel8Emrah eDüzel9Emrah eDüzel10Eckart D Gundelfinger11Constanze I Seidenbecher12Leibniz Institute for NeurobiologyUniversity of MagdeburgCharité University Hospital BerlinLeibniz Institute for NeurobiologyLeibniz Institute for NeurobiologyUniversity of MagdeburgLeibniz Institute for NeurobiologyLeibniz Institute for NeurobiologyUniversity of MagdeburgUniversity of MagdeburgUniversity College LondonLeibniz Institute for NeurobiologyLeibniz Institute for NeurobiologyCatechol-O-methyl transferase (COMT) is involved in the inactivation of dopamine in brain regions in which the dopamine transporter (DAT1) is sparsely expressed. The membrane-bound isoform of COMT (MB-COMT) is the predominantly expressed form in the mammalian central nervous system (CNS). It has been a matter of debate whether in neural cells of the CNS the enzymatic domain of MB-COMT is oriented towards the cytoplasmic or the extracellular compartment. Here we used live immunocytochemistry on cultured neocortical neurons and glial cells to investigate the expression and membrane orientation of native COMT and of transfected MB-COMT fused to green fluorescent protein (GFP). After live staining, COMT immunoreactivity was reliably detected in both neurons and glial cells after permeabilization, but not on unpermeabilized cells. Similarly, autofluorescence of COMT-GFP fusion protein and antibody fluorescence showed overlap only in permeabilized neurons. Our data provide converging evidence for an intracellular membrane orientation of MB-COMT in neurons and glial cells, suggesting the presence of a DAT1-independent postsynaptic uptake mechanism for dopamine, prior to its degradation via COMT.http://journal.frontiersin.org/Journal/10.3389/fpsyt.2010.00142/fullDopaminemembraneneuronal cell culturecatechol-O-methyl transferaseimmnunocytochemistry |
collection |
DOAJ |
language |
English |
format |
Article |
sources |
DOAJ |
author |
Björn H Schott Björn H Schott Björn H Schott Renato Frischknecht Grazyna eDebska-Vielhaber Grazyna eDebska-Vielhaber Nora John Gusalija Behnisch Emrah eDüzel Emrah eDüzel Emrah eDüzel Eckart D Gundelfinger Constanze I Seidenbecher |
spellingShingle |
Björn H Schott Björn H Schott Björn H Schott Renato Frischknecht Grazyna eDebska-Vielhaber Grazyna eDebska-Vielhaber Nora John Gusalija Behnisch Emrah eDüzel Emrah eDüzel Emrah eDüzel Eckart D Gundelfinger Constanze I Seidenbecher Membrane-bound catechol-O-methyl transferase in cortical neurons and glial cells is intracellularly oriented Frontiers in Psychiatry Dopamine membrane neuronal cell culture catechol-O-methyl transferase immnunocytochemistry |
author_facet |
Björn H Schott Björn H Schott Björn H Schott Renato Frischknecht Grazyna eDebska-Vielhaber Grazyna eDebska-Vielhaber Nora John Gusalija Behnisch Emrah eDüzel Emrah eDüzel Emrah eDüzel Eckart D Gundelfinger Constanze I Seidenbecher |
author_sort |
Björn H Schott |
title |
Membrane-bound catechol-O-methyl transferase in cortical neurons and glial cells is intracellularly oriented |
title_short |
Membrane-bound catechol-O-methyl transferase in cortical neurons and glial cells is intracellularly oriented |
title_full |
Membrane-bound catechol-O-methyl transferase in cortical neurons and glial cells is intracellularly oriented |
title_fullStr |
Membrane-bound catechol-O-methyl transferase in cortical neurons and glial cells is intracellularly oriented |
title_full_unstemmed |
Membrane-bound catechol-O-methyl transferase in cortical neurons and glial cells is intracellularly oriented |
title_sort |
membrane-bound catechol-o-methyl transferase in cortical neurons and glial cells is intracellularly oriented |
publisher |
Frontiers Media S.A. |
series |
Frontiers in Psychiatry |
issn |
1664-0640 |
publishDate |
2010-10-01 |
description |
Catechol-O-methyl transferase (COMT) is involved in the inactivation of dopamine in brain regions in which the dopamine transporter (DAT1) is sparsely expressed. The membrane-bound isoform of COMT (MB-COMT) is the predominantly expressed form in the mammalian central nervous system (CNS). It has been a matter of debate whether in neural cells of the CNS the enzymatic domain of MB-COMT is oriented towards the cytoplasmic or the extracellular compartment. Here we used live immunocytochemistry on cultured neocortical neurons and glial cells to investigate the expression and membrane orientation of native COMT and of transfected MB-COMT fused to green fluorescent protein (GFP). After live staining, COMT immunoreactivity was reliably detected in both neurons and glial cells after permeabilization, but not on unpermeabilized cells. Similarly, autofluorescence of COMT-GFP fusion protein and antibody fluorescence showed overlap only in permeabilized neurons. Our data provide converging evidence for an intracellular membrane orientation of MB-COMT in neurons and glial cells, suggesting the presence of a DAT1-independent postsynaptic uptake mechanism for dopamine, prior to its degradation via COMT. |
topic |
Dopamine membrane neuronal cell culture catechol-O-methyl transferase immnunocytochemistry |
url |
http://journal.frontiersin.org/Journal/10.3389/fpsyt.2010.00142/full |
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