This is a request for an ADAMHA Research Scientist Development Award (K02). the long term goal is to develop neuropeptide analogs with oral activity, enzymatic stability and long duration of action in inhibiting morphine tolerance-dependence and abstinence processes. In the present proposal, based on the studies from this laboratory, the following hypotheses will be verified: (a) neuropeptides like Pro-Leu-Gly-NH2 and analogs inhibit morphine tolerance-dependence by acting on the central nervous system (CNS), (b) dopamine (DA) and multiple opiate receptors of specific regions of the CNS are involved in morphine tolerance-dependence and abstinence processes and (c) peptides like Pro-Leu-Gly-NH2 (MIF) and cyclo(Leu-Gly) (CLG) inhibit morphine tolerance-dependence and abstinence processes by affecting DA and opiate receptors. Studies will be carried out in mice and rats to establish if it is a general phenomena or is species dependent. The animals will be made tolerant to and dependent on morphine by subcutaneous implantation of morphine pellets. The degree of tolerance will be assessed by measuring the responses (e.g. analgesia and hypothermia) to varying doses of morphine in morphine and placebo pellet implanted rodents. the degree of physical dependence will be assessed by determining the intensity of symptoms like hypothermia, stereotyped jumping and weight loss during the withdrawal of morphine. The effect of peptides on the development of tolerance to and dependence on morphine and on the symptoms of morphine abstinence will be determined. the binding of DA receptor ligands 3H-SCH 23390 and 3H-domperidone (D1 and D2 receptors, respectively) and of opiate receptor ligands, 3H-DAMGO (mu), 3H-DPDPE (sigma) and 3H-U-69,593 (k) to CNS regions (spinal cord, amygdala, hippocampus, hypothalamus, corpus striatum, pons and medulla, midbrain and cerebral cortex) will be determined. Preliminary studies show that in tolerance-dependence and abstinence processes DA and opiate receptors are affected differentially in CNS regions, hence studies will be carried out with specific brain regions. In order to establish whether CNS changes in DA and opiate receptors are mediated via opiate mechanism the effect of naltrexone on such changes will be determined. Once the specificity of DA and opiate receptor changes is established, then the involvement of second messenger systems (adenylate cyclase and phosphoinositol) will be determined. the effect of different degrees of tolerance induced by implanting different number of pellets during different time intervals on the above biochemical parameters will be monitored. Effect of peptides given intracerebroventricularly on morphine tolerance-dependence will be determined to establish central or peripheral mechanism of action. To test the hypothesis that peptides inhibit morphine tolerance by modifying DA and opiate receptors, their effect on morphine induced changes in specific regions of the CNS will be determined. These studies may lead not only to better understanding of the mechanisms in opiate addiction processes but also to the development of safer drugs in the management of opioid addiction and distressing withdrawal syndrome.
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