title: TRPA1 and TRPV4 activation in human odontoblasts stimulates ATP release.
abstract: The mechanism of pain in dentine hypersensitivity is poorly understood but proposed to result from the activation of dental sensory neurons in response to dentinal fluid movements. Odontoblasts have been suggested to contribute to thermal and mechanosensation in the tooth via expression of transient receptor potential (TRP) channels. However, a mechanism by which odontoblasts could modulate neuronal activity has not been demonstrated. In this study, we investigated functional TRP channel expression in human odontoblast-like cells and measured ATP release in response to TRP channel activation. Human immortalized dental pulp cells were driven toward an odontoblast phenotype by culture in conditioned media. Functional expression of TRP channels was determined with reverse transcription polymerase chain reaction and ratiometric calcium imaging with Fura-2. ATP release was measured using a luciferin-luciferase assay. Expression of mRNA for TRPA1, TRPV1, and TRPV4 but not TRPM8 was detected in odontoblasts by reverse transcription polymerase chain reaction. Expression of TRPV4 protein was detected by Western blotting and immunocytochemistry. The TRPA1 agonists allyl isothiocyanate and cinnamaldehyde and the TRPV4 agonist GSK1016790A caused a concentration-dependent increase in intracellular Ca(2+) concentration that was inhibited by the selective antagonists HC030031, AP18, and HC067047, respectively. In contrast, exposure to the TRPV1 agonist capsaicin or the TRPM8 agonist icilin had no effect on intracellular Ca(2+) concentration. Treatment with allyl isothiocyanate, cinnamaldehyde, or GSK1016790A caused an increase in ATP concentration in culture medium that was abolished by preincubation with TRP channel antagonists. These data demonstrate that activation of TRPA1 and TRPV4 channels in human odontoblast-like cells can stimulate ATP release. We were unable to confirm the presence of thermosensitive TRPV1 and TRPM8 that has previously been reported in odontoblasts.
authors: Biomaterials, Tissue Engineering, and Imaging, King's College London, London, UK Department of Oral Surgery, Dental Institute, King's College London, London, UK.
authors: O
authors: O
authors: Egbuniwe
authors: Wolfson Centre for Age-Related Diseases, King's College London, London, UK.
authors: S
authors: S
authors: Grover
authors: Wolfson Centre for Age-Related Diseases, King's College London, London, UK.
authors: A K
authors: AK
authors: Duggal
authors: Wolfson Centre for Age-Related Diseases, King's College London, London, UK.
authors: A
authors: A
authors: Mavroudis
authors: Department of Oral Surgery, Dental Institute, King's College London, London, UK.
authors: M
authors: M
authors: Yazdi
authors: Department of Oral Surgery, Dental Institute, King's College London, London, UK.
authors: T
authors: T
authors: Renton
authors: Biomaterials, Tissue Engineering, and Imaging, King's College London, London, UK.
authors: L
authors: L
authors: Di Silvio
authors: Wolfson Centre for Age-Related Diseases, King's College London, London, UK andrew.2.grant@kcl.ac.uk.
authors: A D
authors: AD
authors: Grant
mesh_terms: D000083:Acetanilides / Q000494:pharmacology; D000171:Acrolein / Q000031:analogs & derivatives / Q000494:pharmacology; D000255:Adenosine Triphosphate / Q000378:metabolism*; D015220:Calcium Channels / Q000502:physiology*; D020013:Calcium Signaling / Q000187:drug effects; D002211:Capsaicin / Q000494:pharmacology; D018929:Cell Culture Techniques; D002460:Cell Line; D017077:Culture Media, Conditioned; D003782:Dental Pulp / Q000166:cytology; D006801:Humans; D017879:Isothiocyanates / Q000494:pharmacology; D007930:Leucine / Q000031:analogs & derivatives / Q000494:pharmacology; D009419:Nerve Tissue Proteins / Q000819:agonists / Q000037:antagonists & inhibitors / Q000502:physiology*; D009619:Nociceptors / Q000502:physiology*; D009804:Odontoblasts / Q000378:metabolism / Q000502:physiology*; D011687:Purines / Q000494:pharmacology; D011744:Pyrimidinones / Q000494:pharmacology; D018689:Sensory System Agents / Q000494:pharmacology; D013449:Sulfonamides / Q000494:pharmacology; D000074025:TRPA1 Cation Channel; D050053:TRPM Cation Channels / Q000819:agonists; D050916:TRPV Cation Channels / Q000819:agonists / Q000502:physiology*; D050051:Transient Receptor Potential Channels / Q000819:agonists / Q000037:antagonists & inhibitors / Q000502:physiology*
chemical_list: C552888:2-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydro-7H-purin-7-yl)-N-(4-isopropylphenyl)acetamide; D000083:Acetanilides; D015220:Calcium Channels; D017077:Culture Media, Conditioned; D017879:Isothiocyanates; C530602:N-(1-((4-(2-(((2,4-dichlorophenyl)sulfonyl)amino)-3-hydroxypropanoyl)-1-piperazinyl)carbonyl)-3-methylbutyl)-1-benzothiophene-2-carboxamide; D009419:Nerve Tissue Proteins; D011687:Purines; D011744:Pyrimidinones; D018689:Sensory System Agents; D013449:Sulfonamides; D000074025:TRPA1 Cation Channel; C473350:TRPA1 protein, human; D050053:TRPM Cation Channels; C425966:TRPM8 protein, human; D050916:TRPV Cation Channels; C494482:TRPV1 protein, human; C416986:TRPV4 protein, human; D050051:Transient Receptor Potential Channels; D000171:Acrolein; D000255:Adenosine Triphosphate; C004471:allyl isothiocyanate; C490483:icilin; D007930:Leucine; D002211:Capsaicin; C012843:cinnamaldehyde
keywords: dentin; ion channels; orofacial pain; sensation; signal transduction; transient receptor potential channel