MOLECULAR-CLONING AND EXPRESSION OF A HIGH-AFFINITY L-PROLINE TRANSPORTER EXPRESSED IN PUTATIVE GLUTAMATERGIC PATHWAYS OF RAT-BRAIN

We have used the polymerase chain reaction (PCR) with degenerate oligonucleotides derived from two conserved regions of the norepinephrine and gamma-aminobutyric acid transporters to identify novel Na+-dependent transporters in rat brain. One PCR product hybridized to a 4.0 kb RNA concentrated in subpopulations of putative glutamatergic neurons including mitral cells of the olfactory bulb, pyramidal cells of layer V of the cerebral cortex, pyramidal cells of the piriform cortex, and pyramidal cells of field CA3 of the hippocampus. Transient expression of the cognate cDNA conferred Na+-dependent L-proline uptake in HeLa cells that was saturable (K(m) = 9.7-mu-M) and exhibited a pharmacological profile similar to that for high affinity L-proline transport in rat brain slices. The cloned transporter cDNA predicts a 637 aa protein with 12 putative transmembrane domains and exhibits 44%-45% amino acid sequence identity with other members of the emerging family of neurotransmitter transporters. These findings support a synaptic role for L-proline in specific excitatory pathways in the CNS.