Microarray profiling provides the capability to analyze complex changes in gene expression that accompany specific genetic and physiologic alterations. Our combined laboratories have produced multiple mutations in the murine opioid system and have developed technology to characterize gene expression in these models using microarrays. We propose two aims in which opioid system KO mice will be used in gene array studies. We will first extend preliminary data that have identified genes that are up- and down-regulated in several opioid receptor mutants. We will identify and characterize genes affected by opioid system disruption in the brain and spinal cord of individual and combinatorial KO mice. Once changes in expression have been confirmed, bioinformatic approaches will be used to determine whether specific functional classes of genes are altered and in situ hybridization will be used identify their cellular sites of expression. Together, these approaches will identify specific genes whose expression is altered by opioid system mutation, reveal the extent of compensatory change that accompanies these mutations, either alone or in combination, and provide initial indications of the functional significance of any changes. Second, we will explore changes in gene expression that accompany morphine administration during the development of analgesic tolerance and dependence to this drug using the above models. We propose to screen acute and chronically morphine-treated wild-type, MOR-1, KOR-1, DOR-1, ORL-l and ENK knock-out mice using microarrays to identify morphine-regulated genes and to determine whether changes in some or all of these morphine-regulated genes are absent in any of the above mutant strains, which all demonstrate deficits in the development of morphine tolerance, dependence, or both. We will extend preliminary data indicating that multiple gene expression changes can be identified following chronic morphine exposure and that at least some of these changes do not arise in MOR-1 mutant mice; we expect that more restricted sets of genes may be altered in other strains, such as DOR-1, ENK, or ORL-l KOs, in which development of analgesic tolerance following chronic morphine treatment is either delayed or abolished. Taken together, these studies should provide detailed, novel information about the relationship between opioid system gene expression and that of other neurotransmitter systems, as well as illuminate the molecular basis for tolerance, dependence and sensitization, which are critical questions in drug abuse research.
