title: Pharmacological profiling of the TRPV3 channel in recombinant and native assays.
abstract: OBJECTIVE
  Transient receptor potential vanilloid subtype 3 (TRPV3) is implicated in nociception and certain skin conditions. As such, it is an attractive target for pharmaceutical research. Understanding of endogenous TRPV3 function and pharmacology remains elusive as selective compounds and native preparations utilizing higher throughput methodologies are lacking. In this study, we developed medium-throughput recombinant and native cellular assays to assess the detailed pharmacological profile of human, rat and mouse TRPV3 channels.
  
  
  METHODS
  Medium-throughput cellular assays were developed using a Ca(2+) -sensitive dye and a fluorescent imaging plate reader. Human and rat TRPV3 pharmacology was examined in recombinant cell lines, while the mouse 308 keratinocyte cell line was used to assess endogenous TRPV3 activity.
  
  
  RESULTS
  A recombinant rat TRPV3 cellular assay was successfully developed after solving a discrepancy in the published rat TRPV3 protein sequence. A medium-throughput, native, mouse TRPV3 keratinocyte assay was also developed and confirmed using genetic approaches. Whereas the recombinant human and rat TRPV3 assays exhibited similar agonist and antagonist profiles, the native mouse assay showed important differences, namely, TRPV3 activity was detected only in the presence of potentiator or during agonist synergy. Furthermore, the native assay was more sensitive to block by some antagonists.
  
  
  CONCLUSIONS
  Our findings demonstrate similarities but also notable differences in TRPV3 pharmacology between recombinant and native systems. These findings offer insights into TRPV3 function and these assays should aid further research towards developing TRPV3 therapies.
authors.affiliation: Neuroscience Research Division, Lilly Research Centre, Eli Lilly & Co. Ltd., Windlesham, UK.
authors.forename: Olivera
authors.initials: O
authors.lastname: Grubisha
authors.forename: Adrian J
authors.initials: AJ
authors.lastname: Mogg
authors.forename: Jessica L
authors.initials: JL
authors.lastname: Sorge
authors.forename: Laura-Jayne
authors.initials: LJ
authors.lastname: Ball
authors.forename: Helen
authors.initials: H
authors.lastname: Sanger
authors.forename: Cara L A
authors.initials: CL
authors.lastname: Ruble
authors.forename: Elizabeth A
authors.initials: EA
authors.lastname: Folly
authors.forename: Daniel
authors.initials: D
authors.lastname: Ursu
authors.forename: Lisa M
authors.initials: LM
authors.lastname: Broad
mesh_terms: D000818:Animals; D020013:Calcium Signaling / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D057809:HEK293 Cells; D006801:Humans; D008024:Ligands; D049990:Membrane Transport Modulators / Q000494:pharmacology*; D051379:Mice; D051381:Rats; D011994:Recombinant Proteins / Q000187:drug effects / Q000378:metabolism; D050916:TRPV Cation Channels / Q000187:drug effects* / Q000235:genetics / Q000378:metabolism; D014162:Transfection
chemical_list: D008024:Ligands; D049990:Membrane Transport Modulators; D011994:Recombinant Proteins; D050916:TRPV Cation Channels; C459445:TRPV3 protein, human; C505767:TRPV3 protein, rat; C486760:Trpv3 protein, mouse
keywords: FLIPR; TRP channel; TRPV3; mouse 308 keratinocytes