This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. The underlying mechanism(s) of opiate tolerance remains to be elucidated. Neuropeptide FF (NPFF, Phe-Leu-Phe-Gln,Pro-Gln,Arg,Phe-NH2), is a mammalian octapeptide that has been shown to attenuate various opiate-induced effects, like antinociception and the development of morphine tolerance and dependence. It is hypothesized that the administration of morphine releases anti-opiates as part of a homeostatic mechanism. As greater quantities of morphine are administered, increasing quantities of anti-opiates are released into the cerebral spinal fluid. As a direct consequence mu opioid receptor (MOR) complex is down-regulated, producing progressively higher degrees of agonist subsensitivity (tolerance). The present project will examine the mechanism(s) by which NPFF attenuates morphine-induced tolerance via uncoupling of the G protein and/or altering a shared signaling transduction pathway between NPFF and MOR. Rats will be made tolerant by chronic (13 days) intraventricular infusion of morphine sulfate, via the Alzet 2001 osmotic mini-pumps. Because previous findings have shown that chronic i.c.v. infusion of morphine and NPFF alone were able to down-regulate MOR density, the present proposal represents an excellent model to further examine the mechanism(s) by which NPFF attenuates morphine tolerance in spinal cord and discrete brain regions (Cerebral Cortex, Striatum, and Thalamus) that are associated with opiate tolerance. To test this hypothesis, studies will be conducted to test the chronic effects of NPFF on morphine tolerance as measured at several indices related to the MOR signaling transduction pathway (GTP-gamma-S activity, cAMP levels, protein kinases A and C activities). Additional experiments will determine if NPFF modulation at the MOR receptor density is associated with changes at the transcriptional level and/or expression of MOR and NPFF receptor protein. These findings are expected to delineate the mechanism(s) by which NPFF attenuates morphine tolerance by providing a clearer understanding as to where the interaction is occurring within the signaling transduction pathway of the MOR system.