RESUMEN
Transient receptor potential melastatin-3 (TRPM3) is a broadly expressed Ca(2+)-permeable nonselective cation channel. Previous work has demonstrated robust activation of TRPM3 by the neuroactive steroid pregnenolone sulfate (PS), but its in vivo gating mechanisms and functions remained poorly understood. Here, we provide evidence that TRPM3 functions as a chemo- and thermosensor in the somatosensory system. TRPM3 is molecularly and functionally expressed in a large subset of small-diameter sensory neurons from dorsal root and trigeminal ganglia, and mediates the aversive and nocifensive behavioral responses to PS. Moreover, we demonstrate that TRPM3 is steeply activated by heating and underlies heat sensitivity in a subset of sensory neurons. TRPM3-deficient mice exhibited clear deficits in their avoidance responses to noxious heat and in the development of inflammatory heat hyperalgesia. These experiments reveal an unanticipated role for TRPM3 as a thermosensitive nociceptor channel implicated in the detection of noxious heat.
Asunto(s)
Calor/efectos adversos , Hiperalgesia/metabolismo , Umbral del Dolor/fisiología , Células Receptoras Sensoriales/metabolismo , Canales Catiónicos TRPM/metabolismo , Acrilamidas/uso terapéutico , Animales , Conducta Animal/efectos de los fármacos , Glucemia/efectos de los fármacos , Glucemia/genética , Compuestos Bicíclicos Heterocíclicos con Puentes/uso terapéutico , Calcio/metabolismo , Bloqueadores de los Canales de Calcio/farmacología , Capsaicina/farmacología , Línea Celular Transformada , Modelos Animales de Enfermedad , Relación Dosis-Respuesta a Droga , Adyuvante de Freund/efectos adversos , Ganglios Espinales/citología , Regulación de la Expresión Génica/genética , Regulación de la Expresión Génica/efectos de la radiación , Humanos , Hiperalgesia/inducido químicamente , Hiperalgesia/tratamiento farmacológico , Hiperalgesia/genética , Inflamación/inducido químicamente , Inflamación/genética , Inflamación/metabolismo , Potenciales de la Membrana/efectos de los fármacos , Potenciales de la Membrana/genética , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Planta de la Mostaza , Nifedipino/farmacología , Dimensión del Dolor/efectos de los fármacos , Dimensión del Dolor/métodos , Umbral del Dolor/efectos de los fármacos , Técnicas de Placa-Clamp , Aceites de Plantas/farmacología , Pregnenolona/efectos adversos , Células Receptoras Sensoriales/efectos de los fármacos , Canal Catiónico TRPA1 , Canales Catiónicos TRPM/antagonistas & inhibidores , Canales Catiónicos TRPM/deficiencia , Canales Catiónicos TRPM/genética , Telemetría/métodos , Factores de Tiempo , Transfección/métodos , Canales de Potencial de Receptor Transitorio/deficiencia , Canales de Potencial de Receptor Transitorio/genética , Ganglio del Trigémino/citologíaRESUMEN
The transient receptor potential channel of the ankyrin-binding repeat subfamily, TRPA1, is a Ca(2+)-permeable non-selective cation channel that depolarizes the plasma membrane and causes Ca(2+) influx. A typical feature of TRPA1 is its rapid desensitization following activation by agonists such as mustard oil (MO), cinnamaldehyde, and a high intracellular Ca(2+) concentration. In whole-cell recordings on Chinese hamster ovary (CHO) cells expressing TRPA1, desensitization was delayed when phosphatidylinositol 4,5-biphosphate (PIP(2)) was supplemented via the patch pipette, whereas the PIP(2) scavenger neomycin accelerated desensitization. Preincubation with the PI-4 kinase inhibitor wortmannin reduced both constitutive TRPA1 channels activity and the response to MO. Run down was also accelerated by high intracellular Mg(2+) concentrations, whereas chelating intracellular Mg(2+) with 10 mM ethylenedinitrilotetraacetic acid (EDTA) increased the basal channel activity. In inside-out patches, we observed a rapid run down of TRPA1 activity, which could be prevented by application of diC8-PIP(2) or 2 mM Mg-ATP but not Na(2)-ATP to the cytosolic side of the excised patches. In isolated trigeminal ganglion neurons, preincubation with wortmannin resulted in inhibition of endogenous TRPA1 activation by MO. Taken together, our data indicate that PIP(2) modulates TRPA1, albeit to a lesser extent than other known PIP(2)-dependent TRP channels, and that tools modifying the plasma membrane PIP(2) content often have direct effects on this channel.