A Na(+)/Cl(-)-dependent transporter for catecholamines, identified as a norepinephrine transporter, is expressed in the brain of the teleost fish medaka (Oryzias latipes).
We report the isolation, functional characterization, and localization of a Na(+)/Cl(-)-dependent catecholamine transporter (meNET) present in the brain of the teleost fish medaka. This carrier is very similar to the human neuronal norepinephrine transporter (NET) and the human neuronal dopamine transporter (DAT), showing 70 and 64% amino acid identity, respectively. When expressed in COS-7 cells, this transporter mediates the high-affinity uptake of dopamine (K(M) = 290 nM) and norepinephrine (K(M) = 640 nM). Its pharmacological profile reveals more similarities with NET, including a high affinity for the tricyclic antidepressants desipramine (IC(50) = 0.92 nM) and nortriptyline (IC(50) = 16 nM). In situ hybridization on the medaka brain shows that meNET mRNA is present only in a subset of tyrosine hydroxylase-positive neurons found in the noradrenergic areas of the hindbrain, such as the locus ceruleus and area postrema. None of the dopaminergic areas anterior to the isthmus contains any labeled neurons. Neither reverse transcriptase-polymerase chain reaction with degenerate primers specific for gamma-aminobutyric acid transporter/NET nor autoradiographic experiments with [(125)I]3b-(4-iodophenyl)-tropane-2b-carboxylic acid methyl ester revealed an additional catecholamine transporter in the medaka brain. Uptake experiments with medaka brain synaptosomes show an endogenous transport with a pharmacological profile identical to that of the recombinant meNET. Thus, meNET is probably the predominant--if not the only--catecholamine transporter in the medaka fish brain. In view of the highly conserved primary structures and pharmacological properties of meNET, it is tempting to speculate that a specific dopamine transport developed later in vertebrate evolution and probably accompanied the tremendous enlargement of the meso-telencephalic dopaminergic pathways in amniotes.
Roubert
C
C
Institut National de la Santé et de la Recherche Médicale U-513, Faculté de Médecine de Créteil, Créteil, France.
Sagné
C
C
Kapsimali
M
M
Vernier
P
P
Bourrat
F
F
Giros
B
B
D000595:Amino Acid Sequence; D000818:Animals; D001692:Biological Transport; D001921:Brain / Q000378:metabolism*; D019556:COS Cells; D002352:Carrier Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism; D050482:Catecholamine Plasma Membrane Transport Proteins; D002395:Catecholamines / Q000378:metabolism*; D003001:Cloning, Molecular; D026901:Membrane Transport Proteins; D008969:Molecular Sequence Data; D050484:Norepinephrine Plasma Membrane Transport Proteins; D009990:Oryzias; D010802:Phylogeny; D051381:Rats; D017386:Sequence Homology, Amino Acid; D012965:Sodium Chloride / Q000378:metabolism*; D027981:Symporters; D014162:Transfection
D002352:Carrier Proteins; D050482:Catecholamine Plasma Membrane Transport Proteins; D002395:Catecholamines; D026901:Membrane Transport Proteins; D050484:Norepinephrine Plasma Membrane Transport Proteins; C493212:Slc22a1 protein, rat; C493226:Slc6a2 protein, rat; D027981:Symporters; D012965:Sodium Chloride