Fig. 3: TRPV1 enhances the responsiveness of TRPM8-expressing cells to cold.

a, b TRPV1 antagonist AMG9810 (AMG, 300 nM, 3 min pre-treatment before cold stimulation) attenuated calcium responses to cold in transfected KNRK cells that co-express TRPM8 and TRPV1. As a control, the vehicle treatment did not change their response to cold or capsaicin (Cap, 1 µM). Cold stimulation was generated by bath temperature drop from 32 °C to 15 °C. Each dot in b represents a KNRK cell that co-expresses TRPM8 and TRPV1. c Representative calcium transients of sensory neurons to cold stimuli after 10 min pre-treatment of vehicle (0.0006% DMSO in calcium imaging buffer) or AMG9810 (300 nM). Imaged neurons were located within the cornea-projection area in the ophthalmic division of the whole trigeminal ganglion explants (n = 7) from Pirt^GCaMP3/+ transgenic mice, in which a calcium indicator GCaMP3 was expressed in primary sensory neurons. The expression of TRPV1 in tested neurons was determined by their responses to capsaicin (10 µM). d Quantification of calcium responses of trigeminal neurons to cold. Each dot represents one cold-sensing neuron. e–g Cold treatments elicited greater membrane potential changes (ΔMP) and more action potentials (APs) in a subset of TRPM8^EGFPf/+ sensory neurons that are sensitive to capsaicin (1 µM, identified by calcium imaging) than those insensitive to capsaicin. h–j Repeated cold treatments did not desensitize TRPM8^EGFPf/+; TRPV1⁺ sensory neurons in the vehicle control group, as shown by stable membrane potential changes and neuronal firing. k–m Application of TRPV1 antagonist AMG9810 (300 nM, 10 min pre-treatment before cold stimulation) effectively suppressed depolarization and neuronal firing in capsaicin-sensitive TRPM8^EGFPf/+ neurons upon subsequent cold treatment. Data are expressed as mean ± s.e.m. Statistical analysis by two tailed Student’s t-test. **P < 0.01; ***P < 0.001. Source data are provided as a Source Data file.