Cloning and functional expression of GABA sub(B) receptors from Drosophila

The neurotransmitter GABA ( gamma -aminobutyric acid) functions as the major inhibitory neurotransmitter in the central nervous system of vertebrates and invertebrates. In vertebrates GABA signals both through ionotropic receptors (GABA sub(A), GABA sub(C)), which induce fast synaptic inhibitory responses, and through metabotropic receptors (GABA sub(B)), which play a fundamental role in the reduction of presynaptic transmitter release and postsynaptic inhibitory potentials. Whilst GABA sub(A) and GABA sub(C) receptors have been cloned from vertebrates as well as invertebrates, GABA sub(B) receptors have only been identified in vertebrate species to date, although indirect evidence suggests their existence in arthropods, too. Here we report the cloning of three putative invertebrate GABA sub(B) receptor subtypes (D-GABA sub(B)R1, R2 and R3) isolated from Drosophila melanogaster. Whilst D-GABA sub(B)R1 and R2 show high sequence identity to mammalian GABA sub(B)R1 and R2, respectively, the receptor D-GABA sub(B)R3 seems to be an insect-specific subtype with no known mammalian counterpart so far. All three D-GABA sub(B)R subtypes are expressed in the embryonic central nervous system. In situ hybridization of Drosophila melanogaster embryos shows that two of the D-GABA sub(B)Rs (D-GABA sub(B)R1 and R2) are expressed in similar regions, suggesting a coexpression of the two receptors, whilst the third D-GABA sub(B)R (D-GABA sub(B)R3) displays a unique expression pattern. In agreement with these results we have only been able to functionally characterize D-GABA sub(B)R1 and R2 when the two subtypes are coexpressed either in Xenopus laevis oocytes or mammalian cell lines, whilst D-GABA sub(B)R3 was inactive in any combination. The pharmacology of the coexpressed D-GABA sub(B)R1/2 receptor was different from the mammalian GABA sub(B)Rs: e.g. baclofen, an agonist of mammalian GABA sub(B)Rs, showed no effect.