title: "In-bone" utricle cultures--a simplified, atraumatic technique for in situ cultures of the adult mouse (Mus musculus) utricle.
abstract: OBJECTIVE
  The "in-bone" method of culturing utricles described here is a reliable and atraumatic technique for culturing mature mouse hair cells and studying hair cell death and protection.
  
  
  BACKGROUND
  The current in vitro technique for studying hair cells of the mature mouse utricle involves removal from the temporal bone and free floating culture in media. This technique can be problematic because of variability in the preservation of the sensory epithelium and a steep learning curve that results in injury of the sensory epithelium in less experienced hands. We present a new atraumatic technique of culturing the utricle in situ within the temporal bone.
  
  
  METHODS
  Leaving the temporal bone largely intact, a window is opened in the bony vestibule overlying the mouse utricle. The entire temporal bone is then placed into culture media. Utricles were cultured in situ for several days with minimal damage to the epithelium. The utricles are then fixed in situ, removed from the temporal bone, and processed. A standardized aminoglycoside-induced hair cell damage protocol was developed.
  
  
  RESULTS
  Mature mouse utricles maintained hair cell numbers for 3 days in culture. Exposure to neomycin resulted in significant dose-dependent hair cell toxicity (p < 0.0001, 1-way analysis of variance). Exposure to the protective drug tacrine resulted in significant protection against neomycin (p < 0.05, 3-way analysis of variance).
  
  
  CONCLUSIONS
  The "in-bone" technique is a reliable and atraumatic method for culturing mature mouse utricles and studying hair cell death and protection. It is easily mastered and can make in vitro study of hair cells accessible to more research groups.
authors: Virginia Merrill Bloedel Hearing Research Center, Department of Otolaryngology/Head and Neck Surgery, University of Washington, Seattle, WA 98195, USA. henryou@u.washington.edu
authors: Henry C
authors: HC
authors: Ou
authors: Vincent
authors: V
authors: Lin
authors: Edwin W
authors: EW
authors: Rubel
mesh_terms: D000818:Animals; D000900:Anti-Bacterial Agents / Q000037:antagonists & inhibitors / Q000494:pharmacology / Q000633:toxicity; D002452:Cell Count; D016923:Cell Death / Q000187:drug effects; D003470:Culture Media; D004305:Dose-Response Relationship, Drug; D006198:Hair Cells, Auditory / Q000473:pathology; D007759:Labyrinth Diseases / Q000139:chemically induced / Q000473:pathology / Q000517:prevention & control; D008297:Male; D051379:Mice; D008808:Mice, Inbred CBA; D018613:Microscopy, Confocal; D008855:Microscopy, Electron, Scanning; D009355:Neomycin / Q000037:antagonists & inhibitors / Q000633:toxicity; D018697:Nootropic Agents / Q000627:therapeutic use; D015242:Ofloxacin / Q000494:pharmacology; D009924:Organ Culture Techniques / Q000379:methods*; D012444:Saccule and Utricle / Q000502:physiology*; D013619:Tacrine / Q000627:therapeutic use; D013701:Temporal Bone / Q000502:physiology
chemical_list: D000900:Anti-Bacterial Agents; D003470:Culture Media; D018697:Nootropic Agents; D013619:Tacrine; D015242:Ofloxacin; D009355:Neomycin