Research Project
3494361
Opioid peptides and drugs cause multiple effects in the CNS and throughout the entire body. Several distinct opioid receptors have been described biochemically and pharmacologically, but the genes encoding them remain unknown. We propose to identify and clone the opioid receptor genes and transfect each into mammalian cells. Because we have identified and thoroughly characterized a human neuroblastoma cell line (SH-SY5Y) that expresses abundant mu opioid receptors, studies will focus on the mu opioid receptor during phase I. The strategies for gene isolation include 1. the use of oligonucleotide probes derived from peptide fragments of the mu receptor protein; 2. direct mammalian expression cloning of cDNA libraries from SH-SY5Y cells, or alternatively, expression cloning in frog oocytes micro-injected with receptor mRNA 3. screening of cDNA libraries for closely related genes with probes derived from any known opioid receptor gene. The extended search will focus on the main opioid receptor classes, mu, delta and kappa, and their putative subtypes. Knowledge of the number of distinct opioid receptor genes and their molecular structure, and the availability of stably transfected cell lines, will permit the development of unequivocal assays of opioid effects at each of the receptor subtypes, and thereby, provide a powerful tool for the development of highly specific novel drugs. These general objectives will be addressed in phase II of this project.
