Vesicular monoamine transporter-1 (VMAT-1) mRNA and immunoreactive proteins in mouse brain

Karen M. Ashe, Wan Ling Chiu, Ahmed M. Khalifa, Antoine N. Nicolas, Bonnie L. Brown, Randall R De Martino, Clayton P. Alexander, Christopher T. Waggener, Krista Fischer-Stenger, Jennifer K. Stewart

Research output: Contribution to journalArticle

7 Citations (Scopus)

Abstract

OBJECTIVE: Vesicular monoamine transporter 1 (VMAT-1) mRNA and protein were examined (1) to determine whether adult mouse brain expresses full-length VMAT-1 mRNA that can be translated to functional transporter protein and (2) to compare immunoreactive VMAT-1 proteins in brain and adrenal. METHODS: VMAT-1 mRNA was detected in mouse brain with RT-PCR. The cDNA was sequenced, cloned into an expression vector, transfected into COS-1 cells, and cell protein was assayed for VMAT-1 activity. Immunoreactive proteins were examined on western blots probed with four different antibodies to VMAT-1. RESULTS: Sequencing confirmed identity of the entire coding sequences of VMAT-1 cDNA from mouse medulla oblongata/pons and adrenal to a Gen-Bank reference sequence. Transfection of the brain cDNA into COS-1 cells resulted in transporter activity that was blocked by the VMAT inhibitor reserpine and a proton ionophore, but not by tetrabenazine, which has a high affinity for VMAT-2. Antibodies to either the C- or N- terminus of VMAT-1 detected two proteins (73 and 55 kD) in transfected COS-1 cells. The C-terminal antibodies detected both proteins in extracts of mouse medulla/pons, cortex, hypothalamus, and cerebellum but only the 73 kD protein and higher molecular weight immunoreactive proteins in mouse adrenal and rat PC12 cells, which are positive controls for rodent VMAT-1. CONCLUSIONS: These findings demonstrate that a functional VMAT-1 mRNA coding sequence is expressed in mouse brain and suggest processing of VMAT-1 protein differs in mouse adrenal and brain.

Original languageEnglish (US)
Pages (from-to)253-258
Number of pages6
JournalNeuroendocrinology Letters
Volume32
Issue number3
StatePublished - 2011
Externally publishedYes

Fingerprint

Vesicular Monoamine Transport Proteins
Messenger RNA
Brain
Proteins
COS Cells
Pons
Complementary DNA
Antibodies
Proton Ionophores
Rodent Control
Tetrabenazine
Medulla Oblongata
PC12 Cells
Reserpine

Keywords

  • Brain
  • Schizophrenia
  • Solute carrier family 18 member 1
  • Vesicular monoamine transporter
  • VMAT-1

ASJC Scopus subject areas

  • Endocrinology
  • Endocrine and Autonomic Systems
  • Endocrinology, Diabetes and Metabolism
  • Clinical Neurology
  • Psychiatry and Mental health
  • Neurology

Cite this

Ashe, K. M., Chiu, W. L., Khalifa, A. M., Nicolas, A. N., Brown, B. L., De Martino, R. R., ... Stewart, J. K. (2011). Vesicular monoamine transporter-1 (VMAT-1) mRNA and immunoreactive proteins in mouse brain. Neuroendocrinology Letters, 32(3), 253-258.

Vesicular monoamine transporter-1 (VMAT-1) mRNA and immunoreactive proteins in mouse brain. / Ashe, Karen M.; Chiu, Wan Ling; Khalifa, Ahmed M.; Nicolas, Antoine N.; Brown, Bonnie L.; De Martino, Randall R; Alexander, Clayton P.; Waggener, Christopher T.; Fischer-Stenger, Krista; Stewart, Jennifer K.

In: Neuroendocrinology Letters, Vol. 32, No. 3, 2011, p. 253-258.

Research output: Contribution to journalArticle

Ashe, KM, Chiu, WL, Khalifa, AM, Nicolas, AN, Brown, BL, De Martino, RR, Alexander, CP, Waggener, CT, Fischer-Stenger, K & Stewart, JK 2011, 'Vesicular monoamine transporter-1 (VMAT-1) mRNA and immunoreactive proteins in mouse brain', Neuroendocrinology Letters, vol. 32, no. 3, pp. 253-258.
Ashe, Karen M. ; Chiu, Wan Ling ; Khalifa, Ahmed M. ; Nicolas, Antoine N. ; Brown, Bonnie L. ; De Martino, Randall R ; Alexander, Clayton P. ; Waggener, Christopher T. ; Fischer-Stenger, Krista ; Stewart, Jennifer K. / Vesicular monoamine transporter-1 (VMAT-1) mRNA and immunoreactive proteins in mouse brain. In: Neuroendocrinology Letters. 2011 ; Vol. 32, No. 3. pp. 253-258.
@article{b21a6b00c96b4a499334de077093dde1,
title = "Vesicular monoamine transporter-1 (VMAT-1) mRNA and immunoreactive proteins in mouse brain",
abstract = "OBJECTIVE: Vesicular monoamine transporter 1 (VMAT-1) mRNA and protein were examined (1) to determine whether adult mouse brain expresses full-length VMAT-1 mRNA that can be translated to functional transporter protein and (2) to compare immunoreactive VMAT-1 proteins in brain and adrenal. METHODS: VMAT-1 mRNA was detected in mouse brain with RT-PCR. The cDNA was sequenced, cloned into an expression vector, transfected into COS-1 cells, and cell protein was assayed for VMAT-1 activity. Immunoreactive proteins were examined on western blots probed with four different antibodies to VMAT-1. RESULTS: Sequencing confirmed identity of the entire coding sequences of VMAT-1 cDNA from mouse medulla oblongata/pons and adrenal to a Gen-Bank reference sequence. Transfection of the brain cDNA into COS-1 cells resulted in transporter activity that was blocked by the VMAT inhibitor reserpine and a proton ionophore, but not by tetrabenazine, which has a high affinity for VMAT-2. Antibodies to either the C- or N- terminus of VMAT-1 detected two proteins (73 and 55 kD) in transfected COS-1 cells. The C-terminal antibodies detected both proteins in extracts of mouse medulla/pons, cortex, hypothalamus, and cerebellum but only the 73 kD protein and higher molecular weight immunoreactive proteins in mouse adrenal and rat PC12 cells, which are positive controls for rodent VMAT-1. CONCLUSIONS: These findings demonstrate that a functional VMAT-1 mRNA coding sequence is expressed in mouse brain and suggest processing of VMAT-1 protein differs in mouse adrenal and brain.",
keywords = "Brain, Schizophrenia, Solute carrier family 18 member 1, Vesicular monoamine transporter, VMAT-1",
author = "Ashe, {Karen M.} and Chiu, {Wan Ling} and Khalifa, {Ahmed M.} and Nicolas, {Antoine N.} and Brown, {Bonnie L.} and {De Martino}, {Randall R} and Alexander, {Clayton P.} and Waggener, {Christopher T.} and Krista Fischer-Stenger and Stewart, {Jennifer K.}",
year = "2011",
language = "English (US)",
volume = "32",
pages = "253--258",
journal = "Neuroendocrinology Letters",
issn = "0172-780X",
publisher = "Maghira and Maas Publications",
number = "3",

}

TY - JOUR

T1 - Vesicular monoamine transporter-1 (VMAT-1) mRNA and immunoreactive proteins in mouse brain

AU - Ashe, Karen M.

AU - Chiu, Wan Ling

AU - Khalifa, Ahmed M.

AU - Nicolas, Antoine N.

AU - Brown, Bonnie L.

AU - De Martino, Randall R

AU - Alexander, Clayton P.

AU - Waggener, Christopher T.

AU - Fischer-Stenger, Krista

AU - Stewart, Jennifer K.

PY - 2011

Y1 - 2011

N2 - OBJECTIVE: Vesicular monoamine transporter 1 (VMAT-1) mRNA and protein were examined (1) to determine whether adult mouse brain expresses full-length VMAT-1 mRNA that can be translated to functional transporter protein and (2) to compare immunoreactive VMAT-1 proteins in brain and adrenal. METHODS: VMAT-1 mRNA was detected in mouse brain with RT-PCR. The cDNA was sequenced, cloned into an expression vector, transfected into COS-1 cells, and cell protein was assayed for VMAT-1 activity. Immunoreactive proteins were examined on western blots probed with four different antibodies to VMAT-1. RESULTS: Sequencing confirmed identity of the entire coding sequences of VMAT-1 cDNA from mouse medulla oblongata/pons and adrenal to a Gen-Bank reference sequence. Transfection of the brain cDNA into COS-1 cells resulted in transporter activity that was blocked by the VMAT inhibitor reserpine and a proton ionophore, but not by tetrabenazine, which has a high affinity for VMAT-2. Antibodies to either the C- or N- terminus of VMAT-1 detected two proteins (73 and 55 kD) in transfected COS-1 cells. The C-terminal antibodies detected both proteins in extracts of mouse medulla/pons, cortex, hypothalamus, and cerebellum but only the 73 kD protein and higher molecular weight immunoreactive proteins in mouse adrenal and rat PC12 cells, which are positive controls for rodent VMAT-1. CONCLUSIONS: These findings demonstrate that a functional VMAT-1 mRNA coding sequence is expressed in mouse brain and suggest processing of VMAT-1 protein differs in mouse adrenal and brain.

AB - OBJECTIVE: Vesicular monoamine transporter 1 (VMAT-1) mRNA and protein were examined (1) to determine whether adult mouse brain expresses full-length VMAT-1 mRNA that can be translated to functional transporter protein and (2) to compare immunoreactive VMAT-1 proteins in brain and adrenal. METHODS: VMAT-1 mRNA was detected in mouse brain with RT-PCR. The cDNA was sequenced, cloned into an expression vector, transfected into COS-1 cells, and cell protein was assayed for VMAT-1 activity. Immunoreactive proteins were examined on western blots probed with four different antibodies to VMAT-1. RESULTS: Sequencing confirmed identity of the entire coding sequences of VMAT-1 cDNA from mouse medulla oblongata/pons and adrenal to a Gen-Bank reference sequence. Transfection of the brain cDNA into COS-1 cells resulted in transporter activity that was blocked by the VMAT inhibitor reserpine and a proton ionophore, but not by tetrabenazine, which has a high affinity for VMAT-2. Antibodies to either the C- or N- terminus of VMAT-1 detected two proteins (73 and 55 kD) in transfected COS-1 cells. The C-terminal antibodies detected both proteins in extracts of mouse medulla/pons, cortex, hypothalamus, and cerebellum but only the 73 kD protein and higher molecular weight immunoreactive proteins in mouse adrenal and rat PC12 cells, which are positive controls for rodent VMAT-1. CONCLUSIONS: These findings demonstrate that a functional VMAT-1 mRNA coding sequence is expressed in mouse brain and suggest processing of VMAT-1 protein differs in mouse adrenal and brain.

KW - Brain

KW - Schizophrenia

KW - Solute carrier family 18 member 1

KW - Vesicular monoamine transporter

KW - VMAT-1

UR - http://www.scopus.com/inward/record.url?scp=80051955980&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=80051955980&partnerID=8YFLogxK

M3 - Article

C2 - 21712771

AN - SCOPUS:80051955980

VL - 32

SP - 253

EP - 258

JO - Neuroendocrinology Letters

JF - Neuroendocrinology Letters

SN - 0172-780X

IS - 3

ER -