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Opioids in skin function during stress response, regeneration, ageing and, particularly in regulating sensation. In chronic pruritus, topical treatment with Naltrexone changes μ-opioid receptor (μ-OR) localization to relieve itch. The molecular mechanisms behind the effects of Naltrexone on μ-OR function in reduction of itching behavior has not been studied. There is an immediate need to understand the endogenous complexity of μ-OR dynamics in normal and pathological skin conditions. Here we evaluate real-time behavior of μ-OR-Endomorphine complexes in the presence of agonist and antagonists. The μ-OR ligand Endomorphine-1 (EM) was conjugated to the fluorescent dye Tetramethylrhodamine (TAMRA) to investigate the effects of agonist and antagonists in N/TERT-1 keratinocytes. The cellular localization of the EM-TAMRA was followed through time resolved confocal microscopy and population analysis was performed by flow cytometry. The in vitro analyses demonstrate fast internalization and trafficking of the endogenous EM-TAMRA-μ-OR interactions in a qualitative manner. Competition with Endomorphine-1, Naltrexone and CTOP show both canonical and non-canonical effects in basal and differentiated keratinocytes. Acute and chronic treatment with Naltrexone and Endomorphine-1 increases EM-TAMRA binding to skin cells. Although Naltrexone is clinically effective in relieving itch, the mechanisms behind re-distribution of μ-ORs during clinical treatments are not known. Our study has given insight into cellular mechanisms of μ-OR ligand-receptor interactions after opioid agonist and antagonist treatments in vitro. These findings potentially offer opportunities in using novel treatment strategies for skin and peripheral sensory disorders. [ABSTRACT FROM AUTHOR]

Investigation of the genetic functions in complex biological systems is a challenging step in recent year. Hence, several valuable and interesting research projects have been developed with novel ideas to find out the unknown functions of genes or proteins. To validate the applicability of their novel ideas, various approaches are built up. To date, the most promising and commonly used approach for discovering the target proteins from biological system using small molecule is well known a forward chemical genetics which is considered to be more convenient than the classical genetics. Although, the forward chemical genetics consists of the three basic components, the target identification is the most challenging step to chemical biology researchers. Hence, the diverse target identification methods have been developed and adopted to disclose the small molecule bound protein. Herein, in this review, we briefly described the first two parts chemical toolbox and screening, and then the target identifications in forward chemical genetics are thoroughly described along with the illustrative real example case study. In the tabular form, the different biological active small molecules which are the successful examples of target identifications are accounted in this research review. [ABSTRACT FROM AUTHOR]