In addition to their physiological function, metabotropic receptors for neurotransmitter ␥-aminobutyric acid (GABA), the GABAB receptors, may play a role in the behavioral actions of addictive compounds. Recently, GABAB receptors were cloned in fruit flies (Drosophila melanogaster), indicating that the advantages of this experimental model could be applied to GABAB receptor research. RNA interference (RNAi) is an endogenous process triggered by double-stranded RNA and is being used as a tool for functional gene silencing and functional genomics. Here we show how cell-nonautonomous RNAi can be induced in adult fruit flies to silence a subtype of GABAB receptors, GABABR1, and how RNAi combined with pharmacobehavioral techniques (including intraabdominal injections of active compounds and a computer-assisted quantification of behavior) can be used to functionally characterize these receptors. We observed that injection of double-stranded RNA complementary to GABABR1 into adult Drosophila selectively destroys GABABR1 mRNA and attenuates the behavioral actions of the GABAB agonist, 3-aminopropyl-(methyl)phosphinic acid. Moreover, both GABABR1 RNAi and the GABAB antagonist CGP 54626 reduced the behavior-impairing effects of ethanol, suggesting a putative role for the Drosophila GABAB receptors in alcohol's mechanism of action. The Drosophila model we have developed can be used for further in vivo functional characterization of GABAB receptor subunits and their involvement in the molecular and systemic actions of addictive substances.fruit fly ͉ GABAB receptors ͉ ethanol ͉ CGP 54626 ͉ 3-APMPA F ruit flies are used for research primarily because of the homology of Drosophila genes to those of mammals and because Drosophila is amenable to genetic manipulation including gene silencing through a process known as RNA interference (RNAi) (1-7). Typically, RNAi is triggered by double-stranded RNA (dsRNA), which is first processed by an RNase, Dicer (8), into 21-to 23-nt fragments. These fragments form a silencing complex that binds specifically to the dsRNA-complementary endogenous mRNA and leads to the destruction of the mRNA (1, 9). Injecting adult Drosophila intraabdominally with dsRNA results in the cell-nonautonomous silencing of the complementary endogenous mRNA throughout the body, including the CNS (10). Because this method does not interfere in the normal development of the animal but can replicate typical phenotypes produced by gene mutations (11), here we have used adult RNAi to silence ␥-aminobutyric acid (GABA) B receptors.The slow inhibitory GABA synaptic neurotransmission is mediated by the metabotropic G protein-coupled and cAMPlinked GABA B receptors (12, 13) that have been cloned in humans (14) and also recently in Drosophila (15). In addition to their physiological function, the GABA B receptors may play a role in the behavioral actions of addictive compounds such as ethanol (16)(17)(18)(19). Although Drosophila has been used to study the mechanisms of action of alcohol (20)(21)(22)(23), as yet, this model ...