title: Phosphoproteomics and bioinformatics analyses of spinal cord proteins in rats with morphine tolerance.
abstract: BACKGROUND
  Morphine is the most effective pain-relieving drug, but it can cause unwanted side effects. Direct neuraxial administration of morphine to spinal cord not only can provide effective, reliable pain relief but also can prevent the development of supraspinal side effects. However, repeated neuraxial administration of morphine may still lead to morphine tolerance.
  
  
  METHODS
  To better understand the mechanism that causes morphine tolerance, we induced tolerance in rats at the spinal cord level by giving them twice-daily injections of morphine (20 µg/10 µL) for 4 days. We confirmed tolerance by measuring paw withdrawal latencies and maximal possible analgesic effect of morphine on day 5. We then carried out phosphoproteomic analysis to investigate the global phosphorylation of spinal proteins associated with morphine tolerance. Finally, pull-down assays were used to identify phosphorylated types and sites of 14-3-3 proteins, and bioinformatics was applied to predict biological networks impacted by the morphine-regulated proteins.
  
  
  RESULTS
  Our proteomics data showed that repeated morphine treatment altered phosphorylation of 10 proteins in the spinal cord. Pull-down assays identified 2 serine/threonine phosphorylated sites in 14-3-3 proteins. Bioinformatics further revealed that morphine impacted on cytoskeletal reorganization, neuroplasticity, protein folding and modulation, signal transduction and biomolecular metabolism.
  
  
  CONCLUSIONS
  Repeated morphine administration may affect multiple biological networks by altering protein phosphorylation. These data may provide insight into the mechanism that underlies the development of morphine tolerance.
authors.affiliation: Department of Anesthesiology, Tungs' Taichung MetroHarbor Hospital, Taichung, Taiwan ; Department of Anesthesiology, Tri-Service General Hospital, National Defense Medical Center, Taipei, Taiwan ; Department of Pharmacology, National Defense Medical Center, Taipei, Taiwan.
authors.forename: Wen-Jinn
authors.initials: WJ
authors.lastname: Liaw
authors.affiliation: Department of Anesthesiology, Tri-Service General Hospital, National Defense Medical Center, Taipei, Taiwan ; Department of Anesthesiology, Taipei Veterans General Hospital and National Yang- Ming University, Taipei, Taiwan.
authors.forename: Cheng-Ming
authors.initials: CM
authors.lastname: Tsao
authors.affiliation: Department of Anesthesiology, Tri-Service General Hospital, National Defense Medical Center, Taipei, Taiwan.
authors.forename: Go-Shine
authors.initials: GS
authors.lastname: Huang
authors.affiliation: Department of Pharmacology, National Defense Medical Center, Taipei, Taiwan ; Department of Pharmacology, College of Medicine, Taipei Medical University, Taipei, Taiwan.
authors.forename: Chin-Chen
authors.initials: CC
authors.lastname: Wu
authors.affiliation: Department of Anesthesiology, Taipei Veterans General Hospital and National Yang- Ming University, Taipei, Taiwan.
authors.forename: Shung-Tai
authors.initials: ST
authors.lastname: Ho
authors.affiliation: Departments of Anesthesiology, Medical Research, Chi Mei Medical Center, Tainan, Taiwan.
authors.forename: Jhi-Joung
authors.initials: JJ
authors.lastname: Wang
authors.affiliation: Department of Anesthesiology, Rutgers, The State University of New Jersey, New Jersey Medical School, Newark, New Jersey, United States of America.
authors.forename: Yuan-Xiang
authors.initials: YX
authors.lastname: Tao
authors.affiliation: Graduate Institute of Medical Sciences, National Defense Medical Center, Taipei, Taiwan.
authors.forename: Hao-Ai
authors.initials: HA
authors.lastname: Shui
mesh_terms: D000818:Animals; D019295:Computational Biology / Q000379:methods; D004361:Drug Tolerance / Q000502:physiology*; D008297:Male; D009020:Morphine / Q000494:pharmacology*; D009294:Narcotics / Q000494:pharmacology*; D009473:Neuronal Plasticity / Q000502:physiology; D010748:Phosphopeptides / Q000378:metabolism; D010750:Phosphoproteins / Q000378:metabolism*; D025941:Protein Interaction Mapping / Q000379:methods; D060066:Protein Interaction Maps; D020543:Proteome; D040901:Proteomics / Q000379:methods; D051381:Rats; D013116:Spinal Cord / Q000187:drug effects* / Q000378:metabolism*
chemical_list: D009294:Narcotics; D010748:Phosphopeptides; D010750:Phosphoproteins; D020543:Proteome; D009020:Morphine