Gastrin is an important stimulant of acid secretion by gastric parietal cells and is structurally related to the peptide hormone cholecystokinin (CCK).

Dopamine is an important neurotransmitter in the central nervous system of both Drosophila and mammals. Despite the evolutionary distance, functional parallels exist between the fly and mammalian dopaminergic systems, with both playing roles in modulating locomotor activity, sexual function, and the response to drugs of abuse. In mammals, dopamine exerts its effects through either dopamine 1-like (D1-like) or D2-like G protein-coupled receptors. Although pharmacologic data suggest the presence of both receptor subtypes in insects, only cDNAs encoding D1-like proteins have been isolated previously. Here we report the cloning and characterization of a newly discovered Drosophila dopamine receptor. Sequence analysis reveals that this putative protein shares highest homology with known mammalian dopamine 2-like receptors. Eight isoforms of the Drosophila D2-like receptor (DD2R) transcript have been identified, each the result of alternative splicing. The encoded heptahelical receptors range in size from 461 to 606 aa, with variability in the length and sequence of the third intracellular loop. Pharmacologic assessment of three DD2R isoforms, DD2R-606, DD2R-506, and DD2R-461, revealed that among the endogenous biogenic amines, dopamine is most potent at each receptor. As established for mammalian D2-like receptors, stimulation of the Drosophila homologs with dopamine triggers pertussis toxin-sensitive Gi͞o-mediated signaling. The D2-like receptor agonist, bromocriptine, has nanomolar potency at DD2R-606, -506, and -461, whereas multiple D2-like receptor antagonists (as established with mammalian receptors) have markedly reduced if any affinity when assessed at the fly receptor isoforms. The isolation of cDNAs encoding Drosophila D2-like receptors extends the range of apparent parallels between the dopaminergic system in flies and mammals. Pharmacologic and genetic manipulation of the DD2Rs will provide the opportunity to better define the physiologic role of these proteins in vivo and further explore the utility of invertebrates as a model system for understanding dopaminergic function in higher organisms. Dopamine is an essential catecholamine in the central nervous system of both mammals (1) and Drosophila (2). In mammals, dopamine modulates neurologic, cardiovascular, endocrine, and renal functions. In addition, this neurotransmitter regulates motor activity, sexual behavior, and the response to drugs of abuse (1, 3-6). In Drosophila, known dopamine-mediated functions overlap with those of mammals. Experimental evidence suggests that in flies dopamine modulates locomotor activity, sexual function and the response to cocaine, nicotine, and alcohol (7-11).Dopaminergic signaling is mediated through seven transmembrane domain (TM) G protein-coupled receptors that fall within the class A or rhodopsin family based on amino acid homology and conservation of amino acid signature motifs. The dopamine receptor family is divided into two major subfamilies: the D1-like receptors (D1 and D5) and D2-like receptors (D2, D3, and ...

The brain cholecystokinin-B/gastrin receptor (CCK-B/gastrin) has been implicated in mediating anxiety, panic attacks, satiety, and the perception of pain. The canine and human CCK-B/gastrin receptors share 90% amino-acid identity and have similar agonist affinities. These receptors can be selectively blocked by the non-peptide benzodiazepine-based antagonists L365260 (ref. 8) and L364718 (ref. 9); however, the binding of these antagonists to the human and canine receptors differs by up to 20-fold, resulting in a reversal of affinity rank order. Here we report the identification of a single amino acid in the sixth transmembrane domain of the CCK-B/gastrin receptor that corresponds to valine 319 in the human homologue and which is critical in determining the binding affinity for these non-peptide antagonists. We show that it is the variability in the aliphatic side chain of the amino acid in position 319 that confers antagonist specificity. Substitution of valine 319 with a leucine residue decreases the affinity for L365260 20-fold while concomitantly increasing the affinity for L364718. An isoleucine in the same position of the human receptor selectively increases affinity for L364718. Interspecies differences in the aliphatic amino acid occupying this single position selectively affect antagonist affinities without altering the agonist binding profile. We therefore conclude that the residues underlying non-peptide antagonist affinity must differ from those that confer agonist specificity. To our knowledge, these findings are the first example in which a critical antagonist binding determinant for a seven-transmembrane-domain peptide hormone receptor has been identified.