The effects of GABA in the CNS are mediated by three different GABA receptors: GABA(A), GABA(B) and GABA(C) receptors. GABA(A) and GABA(B) receptors, but not yet GABA(C) receptors, have been demonstrated in the enteric nervous system, where GABA has been proposed to be a transmitter. The purpose of this study was to determine whether GABA(C) receptors are present and thus may play a role in mediating the effects of GABA in the myenteric plexus of the rat gastrointestinal tract. We examined the expression of the three known GABA(C) receptor subunits, rho1, rho2 and rho3, in the rat duodenum, ileum and colon using the reverse transcriptase-polymerase chain reaction. We determined the localization of GABA(C) receptors in the myenteric plexus of these regions using two different antisera directed against GABA(C) receptor subunits. The polymerase chain reaction revealed that all three subunits were expressed in the gastrointestinal tract. When the layers of the intestine were separated and the layer containing myenteric neurons was assayed, the rho3 subunit was found in the ileum and colon, whereas rho1 was expressed in the duodenum and weakly in the colon and rho2 was expressed in the ileum. Immunocytochemistry revealed numerous labeled neurons in the myenteric plexus of each region. Colocalization showed that a large proportion of calbindin plus calretinin immunoreactive neurons (intrinsic primary afferent neurons) were immunoreactive for the GABA(C) receptor, and that 56% of nitric oxide synthase immunoreactive neurons (inhibitory motor neurons) exhibited the receptor. These results indicate that GABA(C) receptors of differing subunit compositions are expressed by neurons in the rat gastrointestinal tract. The effects of GABA on intrinsic sensory and on inhibitory motor neurons are likely to be mediated in part through GABA(C) receptors.