We are attempting to characterize the vesicular gamma-aminobutyric acid (GABA) and glutamate transporters. Vesicular transporters sequester neurotransmitter into synaptic vesicles. In previous genetic and pharmacological studies of Caenorhabditis elegans, we identified
unc-47 as a putative vesicular GABA transporter. In collaboration with the Jorgensen laboratory, both mouse and rat cDNAs with substantial sequence similarity to
unc-47 have been isolated. Overlapping expression of the rat homologue (
runc-47) and glutamic acid decarboxylase (GAD) in GABAergic cell populations in rat brain has been demonstrated by in situ hybridization, supporting the role of
runc-47 in GABA transport. By expressing full length cDNAs in heterologous cell systems (PC12 and CHO cells) we have histochemically demonstrated a vesicular pattern of staining of
unc-47 and
runc-47. Finally, using a membrane preparation from transfected PC12 cells we have demonstrated and characterized GABA transport activity. These results indicate that
unc-47 and
runc-47 encode vesicular transporters of GABA. Additional C. elegans genes with sequence similarity to
unc-47 and multiple transmembrane domains were identified in a database search. Rat cDNAs for the putative vertebrate homologues of these
unc-47-related genes have also been isolated. Expression of one of these genes,
rvt2, has been demonstrated in glutamatergic cell populations by in situ hybridization. We are now attempting to demonstrate glutamate transport in transfected PC12 cells. The results of this work should define the molecular basis for packaging of the major inhibitory and excitatory neurotransmitters.