MOLECULAR STUDIES OF A NEW OPIOID RECEPTOR

Three major classes of opioid receptors, mu(mu), delta (delta) and kappa
(kappa), have been defined based on differences in their pharmacology,
physiology and tissue distribution. When stimulated in vivo the opioid
receptors activate a cascade of intracellular reactions involving
adenylyl cyclase, calcium channels and potassium channels, which result
in many of the classical effects of opiate intoxication including
euphoria, analgesia and physical dependence. The molecular
characterization of the opioid receptors has been slow due to several
factors, perhaps the most important of which being that they are
intrinsic membrane proteins which are difficult to solubilize in active
form and they are expressed in relatively low amounts. Recently these
difficulties were overcome by the expression cloning of a mouse delta
opioid receptor subtype from the neuroblastoma X glioma cell line NG108-
15. The primary known as the G protein-coupled receptors. In light of
these recent reports we re-examined the sequence of an orphan receptor
clone which we had obtained by degenerate PCR. This cDNA clone, referred
to as R21, encodes a novel G protein-coupled receptor which shares
significant sequence identity with the recently published mouse delta
opioid receptor. Based on the conservation of key amino acid residues
and the overall homology between R21 and the mouse delta opioid receptor
we predict that R21 is a member of the opioid receptor family. To test
this hypothesis we propose to pharmacologically characterize the receptor
encoded by R21 and investigate the affect its stimulation has on adenylyl
cyclase and a voltage-dependent outwardly rectifying potassium
conductance. Once pharmacologically defined the tissue regulated the rat
R21 gene will be characterized. The human homologue of the R21 gene will
also be characterized to identify markers that can be used i genetic
linkage and association studies of HR21 and human disease. Eventually
the mouse gene, MR21, will be target and knocked out. These transgenic
mice will be a valuable model system in which to evaluate the receptor's
role in processes ranging from synaptic transmission to behavior.