Functional characterization of Na+-coupled citrate transporter NaC2/NaCT expressed in primary cultures of neurons from mouse cerebral cortex

Neurons are known to express a high-affinity Na+-coupled dicarboxylate transporter(s) for uptake of tricarboxylic acid cycle intermediates, such as alpha-ketoglutarate and malate, which are precursors for neuro transmitters including glutamate and gamma-aminobutyric acid. There is, however, little information available on the molecular identity of the transporters responsible for this uptake process in neurons. In the present study, we investigated the characteristics of Na+-dependent citrate transport in primary cultures of neurons from mouse cerebral cortex and established the molecular identity of this transport system as the Na+-coupled citrate transporter (NaC2/NaCT). Reverse transcriptase (RT)-PCR and immunocytochemical analyses revealed that only NaC2/NaCT was expressed in mouse cerebrocortical neurons but not in astrocytes. Uptake of citrate in neurons was Na+-dependent, Li+-sensitive, and saturable with the K-t value of 12.3 mu M. This K-t value was comparable with that in the case of Na+-dependent succinate transport (K-t = 9.2 mu M). Na+-activation kinetics revealed that the Na+-to-citrate stoichiometry was 3.4:1 and concentration of Na+ necessary for half-maximal activation (K-0.5(Na)) was 45.7 mM. Na+-dependent uptake of [C-14]citrate (18 mu M) was significantly inhibited by unlabeled citrate as well as dicarboxylates such as succinate, malate, fumarate, and a-ketoglutarate. This is the first report demonstrating the molecular identity of the Na+-coupled di/tricarboxylate transport system expressed in neurons as NaC2/NaCT, which can transport the tricarboxylate citrate as well as dicarboxylates such as succinate, alpha-ketoglutarate, and malate. (c) 2006 Elsevier B.V. All rights reserved.