Norepinephrine transporter immunoblotting and radioligand binding in cocaine abusers

Deborah C Mash, Qinjie Ouyang, Yujing Qin, John Pablo

Research output: Contribution to journalArticle

30 Citations (Scopus)

Abstract

The norepinephrine transporter (NET) is a membrane protein responsible for transporting extracellular norepinephrine. The cocaine and tricyclic antidepressant-sensitive NET belongs to a family of sodium and chloride coupled transporters that include the monoamines dopamine and serotonin and the amino acids GABA and glycine. The regional distribution of the NET has been defined by synaptosomal uptake of norepinephrine and by autoradiographic approaches in rodent and primate brain. However, the NET has not been well characterized in the human brain due to the overall low abundance of protein expressed in axon terminals. Recently, immunolocalization studies have been used to identify the regional distribution of the cytoplasmic NET epitope in rodent brain. We report here on the characteristics of drug interactions with the native NET protein in human postmortem brain. Antisera raised against a 17-amino acid peptide from the N-terminus of the hNET recognized an 80 kDa species in human cerebral cortex. Chronic exposure to cocaine upregulated NET protein expression and [ 3H]nisoxetine binding sites in the insular cortex from brains of cocaine addicts. These results demonstrate that immunologic and radioligand binding approaches afford specific labeling of the native transport protein in postmortem human brain.

Original languageEnglish
Pages (from-to)79-85
Number of pages7
JournalJournal of Neuroscience Methods
Volume143
Issue number1
DOIs
StatePublished - Apr 15 2005

Fingerprint

Norepinephrine Plasma Membrane Transport Proteins
Cocaine
Immunoblotting
Brain
nisoxetine
Cerebral Cortex
Sodium Chloride Symporters
Rodentia
Norepinephrine
Amino Acids
Tricyclic Antidepressive Agents
Presynaptic Terminals
Drug Interactions
Glycine
gamma-Aminobutyric Acid
Primates
Immune Sera
Epitopes
Dopamine
Serotonin

Keywords

  • Brain
  • Cocaine
  • Insula
  • NET
  • Postmortem

ASJC Scopus subject areas

  • Neuroscience(all)

Cite this

Norepinephrine transporter immunoblotting and radioligand binding in cocaine abusers. / Mash, Deborah C; Ouyang, Qinjie; Qin, Yujing; Pablo, John.

In: Journal of Neuroscience Methods, Vol. 143, No. 1, 15.04.2005, p. 79-85.

Research output: Contribution to journalArticle

Mash, Deborah C ; Ouyang, Qinjie ; Qin, Yujing ; Pablo, John. / Norepinephrine transporter immunoblotting and radioligand binding in cocaine abusers. In: Journal of Neuroscience Methods. 2005 ; Vol. 143, No. 1. pp. 79-85.
@article{d345e58165ad44cb9b6dcf91ded96b5d,
title = "Norepinephrine transporter immunoblotting and radioligand binding in cocaine abusers",
abstract = "The norepinephrine transporter (NET) is a membrane protein responsible for transporting extracellular norepinephrine. The cocaine and tricyclic antidepressant-sensitive NET belongs to a family of sodium and chloride coupled transporters that include the monoamines dopamine and serotonin and the amino acids GABA and glycine. The regional distribution of the NET has been defined by synaptosomal uptake of norepinephrine and by autoradiographic approaches in rodent and primate brain. However, the NET has not been well characterized in the human brain due to the overall low abundance of protein expressed in axon terminals. Recently, immunolocalization studies have been used to identify the regional distribution of the cytoplasmic NET epitope in rodent brain. We report here on the characteristics of drug interactions with the native NET protein in human postmortem brain. Antisera raised against a 17-amino acid peptide from the N-terminus of the hNET recognized an 80 kDa species in human cerebral cortex. Chronic exposure to cocaine upregulated NET protein expression and [ 3H]nisoxetine binding sites in the insular cortex from brains of cocaine addicts. These results demonstrate that immunologic and radioligand binding approaches afford specific labeling of the native transport protein in postmortem human brain.",
keywords = "Brain, Cocaine, Insula, NET, Postmortem",
author = "Mash, {Deborah C} and Qinjie Ouyang and Yujing Qin and John Pablo",
year = "2005",
month = "4",
day = "15",
doi = "10.1016/j.jneumeth.2004.09.013",
language = "English",
volume = "143",
pages = "79--85",
journal = "Journal of Neuroscience Methods",
issn = "0165-0270",
publisher = "Elsevier",
number = "1",

}

TY - JOUR

T1 - Norepinephrine transporter immunoblotting and radioligand binding in cocaine abusers

AU - Mash, Deborah C

AU - Ouyang, Qinjie

AU - Qin, Yujing

AU - Pablo, John

PY - 2005/4/15

Y1 - 2005/4/15

N2 - The norepinephrine transporter (NET) is a membrane protein responsible for transporting extracellular norepinephrine. The cocaine and tricyclic antidepressant-sensitive NET belongs to a family of sodium and chloride coupled transporters that include the monoamines dopamine and serotonin and the amino acids GABA and glycine. The regional distribution of the NET has been defined by synaptosomal uptake of norepinephrine and by autoradiographic approaches in rodent and primate brain. However, the NET has not been well characterized in the human brain due to the overall low abundance of protein expressed in axon terminals. Recently, immunolocalization studies have been used to identify the regional distribution of the cytoplasmic NET epitope in rodent brain. We report here on the characteristics of drug interactions with the native NET protein in human postmortem brain. Antisera raised against a 17-amino acid peptide from the N-terminus of the hNET recognized an 80 kDa species in human cerebral cortex. Chronic exposure to cocaine upregulated NET protein expression and [ 3H]nisoxetine binding sites in the insular cortex from brains of cocaine addicts. These results demonstrate that immunologic and radioligand binding approaches afford specific labeling of the native transport protein in postmortem human brain.

AB - The norepinephrine transporter (NET) is a membrane protein responsible for transporting extracellular norepinephrine. The cocaine and tricyclic antidepressant-sensitive NET belongs to a family of sodium and chloride coupled transporters that include the monoamines dopamine and serotonin and the amino acids GABA and glycine. The regional distribution of the NET has been defined by synaptosomal uptake of norepinephrine and by autoradiographic approaches in rodent and primate brain. However, the NET has not been well characterized in the human brain due to the overall low abundance of protein expressed in axon terminals. Recently, immunolocalization studies have been used to identify the regional distribution of the cytoplasmic NET epitope in rodent brain. We report here on the characteristics of drug interactions with the native NET protein in human postmortem brain. Antisera raised against a 17-amino acid peptide from the N-terminus of the hNET recognized an 80 kDa species in human cerebral cortex. Chronic exposure to cocaine upregulated NET protein expression and [ 3H]nisoxetine binding sites in the insular cortex from brains of cocaine addicts. These results demonstrate that immunologic and radioligand binding approaches afford specific labeling of the native transport protein in postmortem human brain.

KW - Brain

KW - Cocaine

KW - Insula

KW - NET

KW - Postmortem

UR - http://www.scopus.com/inward/record.url?scp=14844313683&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=14844313683&partnerID=8YFLogxK

U2 - 10.1016/j.jneumeth.2004.09.013

DO - 10.1016/j.jneumeth.2004.09.013

M3 - Article

VL - 143

SP - 79

EP - 85

JO - Journal of Neuroscience Methods

JF - Journal of Neuroscience Methods

SN - 0165-0270

IS - 1

ER -