Cloning and molecular characterization of the orphan carrier protein Slc10a4: expression in cholinergic neurons of the rat central nervous system.

We report on the cloning and molecular characterization of the rat carrier Slc10a4 and its cellular localization in the CNS by immunohistochemistry. Slc10a4 is the rat counterpart of the human orphan carrier SLC10A4, which was recently reported to be highly expressed in brain and placenta. Both carriers belong to the solute carrier family SLC10, formerly named the "sodium/bile acid cotransporter family." So SLC10A4/Slc10a4 has a phylogenetic relationship to the Na+/taurocholate cotransporting polypeptide Ntcp (Slc10a1) and the apical sodium-dependent bile acid transporter Asbt (Slc10a2). The rat Slc10a4 protein consists of 437 amino acids and exhibits a seven transmembrane domain topology with N(exo)/C(cyt)trans-orientation of the N- and C-terminal ends. Expression of the Slc10a4 protein was detected in motor regions of the spinal cord and rhombencephalon, as well as in mesopontine cholinergic neurons, the medial habenula, cholinergic areas of the forebrain, and the gut myenteric plexus. Co-localization studies with the cholinergic marker proteins choline acetyltransferase (ChAT), vesicular acetylcholine transporter (VAChT), and high-affinity choline transporter (CHT1) demonstrated expression of Slc10a4 in cholinergic neurons. Despite its close phylogenetic relationship to Ntcp, Slc10a4 showed no transport activity for the Ntcp substrates taurocholate, estrone-3-sulfate, dehydroepiandrosterone sulfate, and pregnenolone sulfate when expressed in HEK293 cells or Xenopus laevis oocytes. Slc10a4 also did not transport choline, which is a substrate of CHT1. Although the functional properties of Slc10a4 could not be elucidated in this study, Slc10a4 is regarded as a new marker protein for cholinergic neurons in the rat CNS.