Journal: bioRxiv

Article Title: Differential expression and localization of thermosensitive Transient Receptor Potential Vanilloid (TRPV) channels in the mature sperm of white pekin duck ( Anas platyrhynchos )

doi: 10.1101/2020.02.10.941732

Figure Lengend Snippet: Western blot analysis of duck sperm extracts probed with different TRPV-specific antibodies are shown. A. TRPV1 specific band is detected by a specific antibody (directed against the C-terminus of TRPV1, Alomone Labs) in absence but not in presence of its blocking peptide; B. Western blot analysis with antibody that detects TRPV2 (raised against the C-terminus, Alomone Labs). C. Two different antibodies detecting TRPV3 [raised against the C-terminus, (Ab1: Alomone Labs) and N-terminus (Ab3: Sigma Aldrich)] detect similar expression pattern of TRPV3. D. Two different antibodies raised against the TRPV4 [raised against C-terminus, Ab1: Alomone Labs) and N-terminus (Ab3: Sigma Aldrich)] detect TRPV4 at the expected size. E. Two different antibodies raised against the C-terminus of TRPV5 (Ab1: Alomone Labs and Ab2: Sigma-Aldrich) detects TRPV5 at expected size. F. A specific antibody raised against the C-terminus of TRPV6 (Ab-1: Alomone Labs) detects TRPV6 in absence but not in presence of its blocking peptide.

Article Snippet: Primary antibodies against TRPV channels were Anti-TRPV1 antibody (Ab1: Alomone Labs, Cat. No. ACC-030; Ab2: Sigma Aldrich, Cat. No. WH0007442M1); Anti-TRPV2 antibody (Ab1: Alomone Labs, Cat. No. ACC-032; Ab2: Calbiochem, Cat. No. PC421); Anti-TRPV3 antibody (Ab1: Alomone Labs, Cat. No. ACC-033; Ab2: Sigma Aldrich, Cat. No. SAB-1300540; Ab3: Sigma Aldrich, Cat. No. SAB1300539); Anti-TRPV4 antibody (Ab1: Alomone Labs, Cat. No. ACC-034; Ab2: Sigma Aldrich, Cat. No. T9075; Ab3: Sigma Aldrich, Cat. No. HPA 007150); Anti-TRPV5 antibody (Ab1: Alomone Labs, Cat. No. ACC-035; Ab2: Sigma Aldrich, Cat. No. T3076); Anti-TRPV6 antibody (Ab1: Alomone Labs, Cat. No. ACC-036).

Techniques: Western Blot, Blocking Assay, Expressing

Journal: bioRxiv

Article Title: Differential expression and localization of thermosensitive Transient Receptor Potential Vanilloid (TRPV) channels in the mature sperm of white pekin duck ( Anas platyrhynchos )

doi: 10.1101/2020.02.10.941732

Figure Lengend Snippet: Flow cytometric evaluation of duck sperm stained for physiologically relevant thermosensitive TRPV channels are shown. A. Representative dot-plots showing percentage of cells expressing TRPV1, TRPV3, TRPV4 channels detected by Ab-1 (antibodies from Alomone labs) antibody specific for each TRPV channel. B. Histograms showing percentage of cells expressing TRPV channels and corresponding Mean Fluorescence Intensity (MFI) of TRPV channels detected by Ab2 antibody of each channel (from Sigma Aldrich), expressed as fold change in comparison to MFI of unstained cells. n = 3, unpaired T-test. ** = P <0.01, *** = P <0.001.

Article Snippet: Primary antibodies against TRPV channels were Anti-TRPV1 antibody (Ab1: Alomone Labs, Cat. No. ACC-030; Ab2: Sigma Aldrich, Cat. No. WH0007442M1); Anti-TRPV2 antibody (Ab1: Alomone Labs, Cat. No. ACC-032; Ab2: Calbiochem, Cat. No. PC421); Anti-TRPV3 antibody (Ab1: Alomone Labs, Cat. No. ACC-033; Ab2: Sigma Aldrich, Cat. No. SAB-1300540; Ab3: Sigma Aldrich, Cat. No. SAB1300539); Anti-TRPV4 antibody (Ab1: Alomone Labs, Cat. No. ACC-034; Ab2: Sigma Aldrich, Cat. No. T9075; Ab3: Sigma Aldrich, Cat. No. HPA 007150); Anti-TRPV5 antibody (Ab1: Alomone Labs, Cat. No. ACC-035; Ab2: Sigma Aldrich, Cat. No. T3076); Anti-TRPV6 antibody (Ab1: Alomone Labs, Cat. No. ACC-036).

Techniques: Staining, Expressing, Fluorescence, Comparison

Journal: bioRxiv

Article Title: Differential expression and localization of thermosensitive Transient Receptor Potential Vanilloid (TRPV) channels in the mature sperm of white pekin duck ( Anas platyrhynchos )

doi: 10.1101/2020.02.10.941732

Figure Lengend Snippet: A-B. Confocal microscopic images depicting the localization of TRPV3 (green) as detected by two different antibodies and Nucleus (blue) by DAPI. Mitochondria (red) is labelled by Mitotracker Red dye in A and C to highlight the channel expression in the mitochondrial region. C . SR-SIM images of TRPV3 localization (using Ab1 antibody) at the head (left) and tail (right) of duck sperm is shown. D. Zoomed up image of neck region of sperm depicting the presence of TRPV3 (green) in the neck region. The head (blue) and arrows mark the start- and end-point of mitochondrial region.

Article Snippet: Primary antibodies against TRPV channels were Anti-TRPV1 antibody (Ab1: Alomone Labs, Cat. No. ACC-030; Ab2: Sigma Aldrich, Cat. No. WH0007442M1); Anti-TRPV2 antibody (Ab1: Alomone Labs, Cat. No. ACC-032; Ab2: Calbiochem, Cat. No. PC421); Anti-TRPV3 antibody (Ab1: Alomone Labs, Cat. No. ACC-033; Ab2: Sigma Aldrich, Cat. No. SAB-1300540; Ab3: Sigma Aldrich, Cat. No. SAB1300539); Anti-TRPV4 antibody (Ab1: Alomone Labs, Cat. No. ACC-034; Ab2: Sigma Aldrich, Cat. No. T9075; Ab3: Sigma Aldrich, Cat. No. HPA 007150); Anti-TRPV5 antibody (Ab1: Alomone Labs, Cat. No. ACC-035; Ab2: Sigma Aldrich, Cat. No. T3076); Anti-TRPV6 antibody (Ab1: Alomone Labs, Cat. No. ACC-036).

Techniques: Expressing

Journal: Molecular Pain

Article Title: TRPV3 and TRPV4 ion channels are not major contributors to mouse heat sensation

doi: 10.1186/1744-8069-7-37

Figure Lengend Snippet: Temperature preference and heat-evoked acute nociceptive behavior of TRPV3 knockout mice on the C57BL6 background . (A) Thermal preference behavior of freely moving wild-type (black solid line, n = 12) and TRPV3 knockout (KO, red dashed line, n = 12) mice on a floor temperature gradient of 0.8°C to 48.8°C over 120 minutes. Percent of time spent (mean ± SEM) within the indicated 2°C bins was monitored during the indicated time periods. (B) Thermal preference behavior of freely moving wild-type (filled black squares, n = 10-12) and TRPV3 knockout (KO, open red circles, n = 10-12) mice in a two-temperature selection task. Percent of time spent (mean ± SEM) at the indicated floor temperature was monitored for 5 min intervals over 60 min. Floor temperature choices were 33°C vs 37°C (left), 35°C vs. 24°C (middle), and 34°C vs. 28°C (right). (C) Heat-evoked acute nociceptive behavior of wild-type (filled bars) and TRPV3 KO (open bars) mice. Latency to response was measured in the tail immersion (left, n = 13) and hot plate (middle, n = 10) assays at the indicated temperatures. Latency to response in the radiant paw heating assay (n = 13-14) was measured during stimulation at a fixed lamp intensity. Data represent mean ± SEM.

Article Snippet: TRPV3 knockout 129S6 mice were derived by backcrossing TRPV3 +/- C57BL6 mice first with HATRPV3 C57BL6 transgenic mice [ ], then over 6 generations with wild-type 129S6 mice (Taconic Labs).

Techniques: Knock-Out, Selection

Journal: Molecular Pain

Article Title: TRPV3 and TRPV4 ion channels are not major contributors to mouse heat sensation

doi: 10.1186/1744-8069-7-37

Figure Lengend Snippet: Temperature preference and heat-evoked nociceptive behavior of TRPV3 knockout mice on the 129S6 background . (A) Thermal preference behavior of wild-type (black solid line, n = 12) and TRPV3 knockout (KO, red dashed line, n = 10) mice on a floor temperature gradient of 0.8 to 48.8°C. Percent of time (mean ± SEM) spent within the indicated 2°C bins was monitored during the indicated time periods. Right panel shows percent of time (mean ± SEM) spent by wild-type (filled bar) and knockout (open bar) mice between 19°C and 29°C during the final 30 min. (B) Preference index of individual mice for temperatures > 29°C in the thermal gradient. Wild-type C57BL6 mice were those presented in Figure 1A (grp 1, n = 11) or a separate group (grp 2, n = 7) assayed at the same time as the 129S6 mice from Figure 2A (129S6 wild-type n = 12, 129S6 TRPV3 KO n = 10). (C) Thermal preference of wild-type (filled black squares, n = 10-12) and TRPV3 KO (open red circles, n = 10-12) mice in two-temperature selection tasks. Percent of time spent (mean ± SEM) at the indicated temperature was monitored at 5 min intervals. (D) Heat-evoked acute nociceptive behavior of wild-type (filled bars, n = 12) and TRPV3 KO (open bars, n = 10) mice. Latency to response was measured in the tail immersion (left), hot plate (middle), and radiant paw heating (right) assays. Data represent mean ± SEM. (* p < 0.05, wild-type vs. knockout, unpaired t-test).

Article Snippet: TRPV3 knockout 129S6 mice were derived by backcrossing TRPV3 +/- C57BL6 mice first with HATRPV3 C57BL6 transgenic mice [ ], then over 6 generations with wild-type 129S6 mice (Taconic Labs).

Techniques: Knock-Out, Selection

Journal: Molecular Pain

Article Title: TRPV3 and TRPV4 ion channels are not major contributors to mouse heat sensation

doi: 10.1186/1744-8069-7-37

Figure Lengend Snippet: Temperature preference and heat-evoked nociceptive behavior of TRPV3/TRPV4 double knockout mice on the C57BL6 background . (A) Thermal preference behavior of freely moving wild-type (black solid line, n = 12) and TRPV3/TRPV4 double knockout (V3V4 KO, red dashed line, n = 10) mice on a floor temperature gradient of 0.8°C to 48.8°C over 120 minutes. Percent of time spent (mean ± SEM) within the indicated 2°C bins was monitored during the indicated time periods. (B) Heat-evoked acute nociceptive behavior of wild-type (filled bars) and TRPV3/TRPV4 double knockout (open bars) mice. Latency to response was measured in the tail immersion (left, n = 12-13) and hot plate (middle, n = 12) assays at the indicated temperatures. Latency to response in the radiant paw heating assay (n = 19-20) was measured during stimulation at a fixed lamp intensity. Data represent mean ± SEM. (*p < 0.05 wild-type vs. knockout, unpaired t-test).

Article Snippet: TRPV3 knockout 129S6 mice were derived by backcrossing TRPV3 +/- C57BL6 mice first with HATRPV3 C57BL6 transgenic mice [ ], then over 6 generations with wild-type 129S6 mice (Taconic Labs).

Techniques: Double Knockout, Knock-Out

Journal: Molecular Pain

Article Title: TRPV3 and TRPV4 ion channels are not major contributors to mouse heat sensation

doi: 10.1186/1744-8069-7-37

Figure Lengend Snippet: Effect of TRPV1 antagonism on acute and inflammatory heat-evoked nociceptive behavior of TRPV3/TRPV4 double knockout mice on the C57BL6 background . (A) Effect of TRPV1 antagonist (JNJ-17203212, 40 mg/kg, i.p., open symbols, n = 7 per genotype) or vehicle (filled symbols, n = 7 per genotype), administered at the indicated time, on tail withdrawal latencies from a 50°C water bath in naïve wild-type (squares) and TRPV3/TRPV4 double knockout (circles) mice. (B) Ipsilateral paw withdrawal latencies to radiant heat in naïve wild-type (filled bars) and TRPV3/TRPV4 double knockout (open bars) mice prior to and 48 hr after intraplantar injection of complete Freund's adjuvant (CFA, 50% in saline) in the left hindpaw (10 μl) (wild-type, n = 30; TRPV3/TRPV4 double knockout, n = 32). (C) Effect of TRPV1 antagonist (JNJ-17203212, 40 mg/kg, i.p., open symbols) or vehicle (filled symbols), on post-CFA paw withdrawal latencies in the wild-type (squares, n = 15 per treatment) and TRPV3/TRPV4 double knockout (circles, n = 16 per treatment) mice shown in panel (B). Data shown represent mean ± SEM.

Article Snippet: TRPV3 knockout 129S6 mice were derived by backcrossing TRPV3 +/- C57BL6 mice first with HATRPV3 C57BL6 transgenic mice [ ], then over 6 generations with wild-type 129S6 mice (Taconic Labs).

Techniques: Double Knockout, Injection, Adjuvant, Saline

Journal: Molecular Pharmacology

Article Title: Activation of TRPV3 channels in bladder cancer cells stimulates ATP release

doi: 10.1016/j.molpha.2025.100096

Figure Lengend Snippet: AV3-1 activated human and mouse TRPV3. (A, B) Concentration-response curves depicting TRPV3 activation by AV3-1 in HEK cells expressing either mouse TRPV3 (A, black squares) or human TRPV3 (B, black squares). Parental HEK cells lacking TRPV3 expression (A, black triangles) served as a negative control and showed no response to AV3-1. Fluorescence signals (F) were normalized to the fluorescence before compound addition (F 0 ). Data are presented as mean ± SD from 5 independent experiments, with duplicates each. Chemical structure of AV3-1 is shown as in inset in (A). (C–E) Microfluorometric single-cell analysis of [Ca 2+ ] i in fura-2-loaded cells. Shown are representative traces of [Ca 2+ ]ᵢ over time in parental HEK cells in response to application of 50 μ M AV3-1 (C) and in HEK cells stably expressing mouse TRPV3 (HEK mTRPV3 ), in response to application of 5 μ M AV3-1, followed by addition of the inhibitor 26E01 (50 μ M) (left panel) or without inhibitor (right panel) (D, E). Individual cell traces are shown in gray; the average trace is overlaid in black. Application periods are indicated by colored bars. (E) Quantification of peak [Ca 2+ ]ᵢ responses from HEK mTRPV3 cells under the indicated treatment conditions. A baseline was recorded in all experiments, followed by AV3-1 application; in 50% of experiments, 26E01 was subsequently added to the cells. Because of this design, sample sizes differ across conditions; the exact n for each box is indicated in the figure. Quantification for parental HEK cells is shown as an inset in (C) (n = 5 independent experiments). Statistical significance was determined using one-way ANOVA with Tukey post hoc analysis; ∗ P < .05. ns, not significant.

Article Snippet: Human TRPV3 ( NM_145068 , purchased from OriGene) and mouse TRPV3 ( NM_001371006 ), rat TRPV1, mouse TRPV2, mouse TRPV4, human TRPM2, human TRPM8, and human TRPA1 were stably transfected in HEK293 cells as described before.

Techniques: Concentration Assay, Activation Assay, Expressing, Negative Control, Fluorescence, Single-cell Analysis, Stable Transfection

Journal: Molecular Pharmacology

Article Title: Activation of TRPV3 channels in bladder cancer cells stimulates ATP release

doi: 10.1016/j.molpha.2025.100096

Figure Lengend Snippet: Electrophysiological analysis of AV3-1–activated TRPV3 currents in HEK m TRPV3 cells. (A) Representative time course of a whole-cell patch-clamp recording obtained from a HEK m TRPV3 cell, showing current responses to repeated applications of the TRPV3 agonist AV3-1 (5 μ M, blue bars, 1 minute each), followed by a washout period. The TRPV3 inhibitor 26E01 (50 μ M, red bar) was applied during the third AV3-1 stimulation. Current amplitudes were recorded using voltage ramps from −100 mV to +100 mV (500-millisecond duration) such as in B and expressed as current density (pA/pF) measured at +100 mV and −100 mV. The labels (1, 6–8) indicate time points for statistical analysis as shown in (C). (B) Representative current-voltage (I–V) curves from the same cell shown in (A), taken at baseline (1, black), at peak response to AV3-1 prior to 26E01 addition (6, blue), and during coapplication with 26E01 (7, red). (C) Quantification of current densities (pA/pF) at −100 mV (inward current) and +100 mV (outward current) at the time points (1, 6–8) as indicated in (A). Box plot shows data of n = 5 independent recordings. Statistical significance was determined using Kruskal-Wallis ANOVA followed by Conover test; ∗ P < .05. Asterisks indicate significance for both outward and inward current components.

Article Snippet: Human TRPV3 ( NM_145068 , purchased from OriGene) and mouse TRPV3 ( NM_001371006 ), rat TRPV1, mouse TRPV2, mouse TRPV4, human TRPM2, human TRPM8, and human TRPA1 were stably transfected in HEK293 cells as described before.

Techniques: Patch Clamp

Journal: Molecular Pharmacology

Article Title: Activation of TRPV3 channels in bladder cancer cells stimulates ATP release

doi: 10.1016/j.molpha.2025.100096

Figure Lengend Snippet: TRPV3 is functionally expressed in bladder cancer cells. (A–D) Activation of TRPV3 in KU-19-19 (A, B) and CAL-29 (C, D) bladder cancer cells by AV3-1, assessed by fura-2-based Ca 2+ assays. The experimental layout was as in , D and . Shown are representative traces of [Ca 2+ ]ᵢ over time in response to application of 50 μ M AV3-1, followed by addition of the inhibitor 26E01 (50 μ M) (left panel) or without inhibitor (right panel). Individual cell traces are shown in gray; the average trace is overlaid in black. Application periods are indicated by colored bars. (B, D) Quantification of peak [Ca 2+ ]ᵢ responses from KU-19-19 (B) and CAL-29 (D) cells under the indicated treatment conditions. The exact n for each box is indicated in the figure. Statistical significance was determined using one-way ANOVA with Tukey post hoc analysis; ∗ P < .05. (E–G) Electrophysiological whole-cell recordings performed on KU-19-19 cells. (E) Representative time course of a whole-cell patch-clamp recording obtained from a KU-19-19 cell, showing current responses to repeated applications of the TRPV3 agonist AV3-1 (50 μ M, blue bars, 1 minute each). The TRPV3 inhibitor 26E01 (50 μ M, red bar) was applied during the third AV3-1 stimulation. Current amplitudes were recorded using voltage ramps from −100 mV to +100 mV (500-millisecond duration) such as in F and expressed as current density (pA/pF) measured at +100 mV and −100 mV. The labels (1, 6–8) indicate time points for statistical analysis as shown in (G). (F) Representative current-voltage (I–V) curves from the same cell shown in (E), taken at baseline (1, black), at peak response to AV3-1 prior to 26E01 addition (6, blue), and during c-application with 26E01 (7, red). (G) Quantification of current densities (pA/pF) at −100 mV (inward current) and +100 mV (outward current) at the time points (1, 6–8) as indicated in (E). Box plot shows data of n = 5 independent recordings. Statistical significance was determined using Kruskal-Wallis ANOVA followed by Conover test; ∗ P < .05. Asterisks indicate significance for both outward and inward current components.

Article Snippet: Human TRPV3 ( NM_145068 , purchased from OriGene) and mouse TRPV3 ( NM_001371006 ), rat TRPV1, mouse TRPV2, mouse TRPV4, human TRPM2, human TRPM8, and human TRPA1 were stably transfected in HEK293 cells as described before.

Techniques: Activation Assay, Patch Clamp

Journal: Molecular Pharmacology

Article Title: Activation of TRPV3 channels in bladder cancer cells stimulates ATP release

doi: 10.1016/j.molpha.2025.100096

Figure Lengend Snippet: TRPV3 activation by AV3-1 was enhanced by cholesterol supplementation and causes ATP release in KU-19-19 bladder cancer cells. (A–C) Electrophysiological whole-cell recordings performed on KU-19-19 cells without (A) and with (B) cholesterol supplementation. (A, B) Representative time courses of whole-cell patch-clamp recordings obtained from a KU-19-19 cell, showing current responses to repeated applications of the TRPV3 agonist AV3-1 (50 μ M, blue bars, 1 minute each). Current amplitudes were recorded using voltage ramps from −100 mV to +100 mV (500-millisecond duration) and expressed as current density (pA/pF) measured at +100 mV and −100 mV. (C) Quantification of current densities (pA/pF) at −100 mV (inward current) and +100 mV (outward current) at the time points (1, 2) as indicated in (A, B). Box plot show data of n = 10 independent recordings. Statistical significance was determined using Kruskal-Wallis ANOVA followed by Conover test; ∗ P < .05. Asterisks indicate significance for both outward and inward current components. (D) Concentration-response curves depicting TRPV3 activation in KU-19-19 cells without (open squares) or with (gray circles) cholesterol supplementation. Experiments were conducted as described in , A and , and depict means ± SD of 5 independent experiments. Inset: Box-and-whisker plots of AV3-1 potencies under control conditions (open box) and following cholesterol supplementation (gray box), derived from individual concentration-response fits and expressed as pEC 50 (-log 10 (EC 50 [M])). Each diamond represents an individual experiment (n = 5 per condition). Group differences were tested on pEC 50 values using a two-tailed unpaired Student t test; ∗ P < .05. (E) Concentration response analysis of AV3-1 in MTT assays with KU-19-19 cells depicting cell viability and proliferation after 24 hours of stimulation with AV3-1 at concentrations as indicated. Data represent means ± SD of n = 5 independent experiments. (F) Box plot quantification of extracellular ATP levels in KU-19-19 cells after stimulation with AV3-1 (50 μM, blue), measured by a luciferin-luciferase–based luminescence assay. Luminescence values are normalized to control values observed in wells containing only media (L/L control). Pretreatment with the TRPV3 antagonist 26E01 (50 μ M, red) or the nonselective TRP channel blocker ruthenium red (RR; 10 μM, red diagonal hatched fill) reduced AV3-1–evoked ATP release. Statistical analysis of n = 6 independent measurements was performed using one-way ANOVA with Tukey post hoc test; ∗ P < .05. ns, not significant.

Article Snippet: Human TRPV3 ( NM_145068 , purchased from OriGene) and mouse TRPV3 ( NM_001371006 ), rat TRPV1, mouse TRPV2, mouse TRPV4, human TRPM2, human TRPM8, and human TRPA1 were stably transfected in HEK293 cells as described before.

Techniques: Activation Assay, Patch Clamp, Concentration Assay, Whisker Assay, Control, Derivative Assay, Two Tailed Test, Luciferase, Luminescence Assay

Journal: eLife

Article Title: TRPV3 channel activity helps cortical neurons stay active during fever

doi: 10.7554/eLife.102412

Figure Lengend Snippet: ( A ) TRPV3 immunostaining (green) in mouse primary somatosensory (S1) cortex, striatum, thalamus, and hippocampus, with DAPI counterstain (blue). Top row left: Postnatal day (P)7; bottom row left: P14; bottom row right: P21. Top row right: Sections incubated with TRPV3 antibody plus TRPV3 blocking peptide in S1 cortex, striatum, thalamus, and hippocampus (green) with DAPI counterstain (blue). Remaining TRPV3 staining is shown in green with DAPI in blue. ( B ) TRPV3 and TRPV4 immunostaining in S1 cortex, striatum, thalamus, and hippocampus at P14. Top row left: TRPV3 (green) and TRPV4 (brick red). Top row right: TRPV3 (green) and TRPV4 (brick red) with DAPI counterstain (blue). Bottom row left: TRPV4 immunostaining in S1BF (inset from top row left) at 10x. Bottom row right: TRPV3 immunostaining in S1BF (inset from top row left) at 10x.

Article Snippet: Slides were blocked in 10% normal goat serum for 1 hr at room temperature, then incubated overnight at 4°C with anti-TRPV3-Biotin antibody (#ACC-033-B), TRPV3 blocking peptide (#BLP-CC033), and/or anti-TRPV4 antibody (#ACC-034) (Alomone Labs, Israel).

Techniques: Immunostaining, Incubation, Blocking Assay, Staining

Journal: eLife

Article Title: TRPV3 channel activity helps cortical neurons stay active during fever

doi: 10.7554/eLife.102412

Figure Lengend Snippet: ( A ) Setup for recording whole-cell TRPV3 currents at 30°C (black), 36°C (gray), and 39°C (red) in cortical excitatory pyramidal neurons (PNs) with bath application of camphor (5 mM), a TPRV3 agonist. ( B ) Current density-voltage (I–V) relationship of TRPV3 currents at 30°C (black), 36°C (gray) and 39°C (red) in wild-type (WT) mice: 11 cells from four mice. ( C ) Scatter dot plots of the current density-voltage measurements. ( D ) Current density-voltage (I–V) relationship of TRPV3 currents at 30°C (black) in the presence of camphor (5 mM), a TPRV3 agonist, or camphor (5 mM) + TRPV3 blocker (Forsythoside B, 50 µM) (blue). ( E ) Same as ( D ) but for 36°C. ( F ) Same as ( D ) but for 39°C. ( G ) Current density-voltage (I–V) plot showing the net TRPV3 current (opener – (opener+blocker) condition). In B - F , statistical significance was assessed using a two-way repeated-measures ANOVA with Tukey’s or Sidak post-hoc test ( α =0.05).

Article Snippet: Slides were blocked in 10% normal goat serum for 1 hr at room temperature, then incubated overnight at 4°C with anti-TRPV3-Biotin antibody (#ACC-033-B), TRPV3 blocking peptide (#BLP-CC033), and/or anti-TRPV4 antibody (#ACC-034) (Alomone Labs, Israel).

Techniques:

Journal: eLife

Article Title: TRPV3 channel activity helps cortical neurons stay active during fever

doi: 10.7554/eLife.102412

Figure Lengend Snippet: ( A ) Setup for recording L4-evoked post-synaptic potential and spiking in an excitatory cortical pyramidal neuron (PN) with an intracellular blocker of TRPV3 channels (Forsythoside B, 50 µM) (left) or TRPV4 channels (RN1734, 10 µM) (right) at just-subthreshold V m at 30°C, 36°C, and 39°C in mouse S1 cortex. ( B ) Percentages of cell types obtained from experiment in A . ( C ) Evoked spikes in L2/3 cortical PNs during temperature elevations to 30°C, 36°C, and 39°C under three conditions: no blockers, TRPV3 blocker (Forsythoside B, 50 µM), or TRPV4 blocker (RN1734, 10 µM). ( D ) Correlation between post-synaptic potential (PSP) peak and spike threshold (ST). r =Pearson correlation coefficient with Deming linear regression. ( E ) Same as ( C ) for the L4-evoked late PSP peak. ( F ) Same as ( C ) for input resistance (R in ). Each data point in C – F represents an individual cell. Data were collected from 26 cells in 7 animals for the TRPV3 blocker, 24 cells in 6 animals for the TRPV4 blocker, and 37 cells in 14 animals for the no-block condition. Mean ± SEM is shown in C , E , and F . Statistical significance was assessed using one- or two-way repeated-measures ANOVA with Tukey’s or Sidak post-hoc test ( α =0.05). In D , correlations were evaluated using Pearson’s r with Deming linear regression.

Article Snippet: Slides were blocked in 10% normal goat serum for 1 hr at room temperature, then incubated overnight at 4°C with anti-TRPV3-Biotin antibody (#ACC-033-B), TRPV3 blocking peptide (#BLP-CC033), and/or anti-TRPV4 antibody (#ACC-034) (Alomone Labs, Israel).

Techniques: Blocking Assay

Journal: eLife

Article Title: TRPV3 channel activity helps cortical neurons stay active during fever

doi: 10.7554/eLife.102412

Figure Lengend Snippet: ( A ) Setup for recording L4-evoked post-synaptic potentials and spiking in excitatory cortical pyramidal neurons (PNs) at just-subthreshold Vm at 30°C, 36°C, and 39°C in mouse S1 cortex of wild-type ( Trpv3 +/+ ) and Trpv3 knockout ( Trpv3 -/- ) mice. ( B ) Depolarization required to reach spike threshold (ST) in Trpv3 +/+ and Trpv3 -/- mice at 30°C, 36°C, and 39°C. ( C ) Same as ( B ) but for input resistance (R in ). ( D ) Same as ( B ) but for number of spikes. ( E ) Same as ( B ) but for post-synaptic potential (PSP). ( F ) Setup for recording mouse body temperature (T b ) at room temperature and during fever-range and higher, using an implanted transponder for non-invasive measurement, with exposure to infrared light. ( G ) Time to loss of postural control (LPC), defined as collapse and failure to maintain upright posture, in wild-type ( Trpv3 +/+ ), heterozygous ( Trpv3 +/- ), and Trpv3 knockout ( Trpv3 -/- ) mice. ( H ) Same as in ( G ) but showing T b at seizure onset. ( I ) Same as in ( G ) but for the time from LPC to seizure onset. Each data point in B – E represents an individual cell (three animals per genotype). Statistical significance was assessed using two-way repeated-measures ANOVA with Tukey’s or Sidak post-hoc test ( α =0.05). Each data point in G – I represents an individual animal. Statistical significance was assessed using one-way repeated-measures ANOVA with Tukey’s or Sidak post-hoc test ( α =0.05).

Article Snippet: Slides were blocked in 10% normal goat serum for 1 hr at room temperature, then incubated overnight at 4°C with anti-TRPV3-Biotin antibody (#ACC-033-B), TRPV3 blocking peptide (#BLP-CC033), and/or anti-TRPV4 antibody (#ACC-034) (Alomone Labs, Israel).

Techniques: Knock-Out, Control

Journal: eLife

Article Title: TRPV3 channel activity helps cortical neurons stay active during fever

doi: 10.7554/eLife.102412

Figure Lengend Snippet: ( A ) In cortical L2/3 pyramidal neurons (PNs) with synaptically evoked spiking, gradual increases in brain slice temperature from 30 °C to 36°C to 39°C result in four possible outcomes: neurons remain inactive, continue spiking (STAY), stop spiking, or initiate spiking. To spike, PNs must reach the spike threshold (ST), defined as the minimal V m that elicits an action potential, which requires sufficient depolarization via the post-synaptic potential (PSP). STAY neurons consistently reach ST and continue spiking. Neurons that stop spiking fall below the level of depolarization required to reach ST, while neurons that initiate spiking achieve sufficient depolarization to newly reach ST. ( B ) STAY neurons may contain unique ion channels, such as TRPV3. TRPV3 channels are highly permeable to Ca² + ions, and Ca² + influx contributes to PN depolarization. The presence of TRPV3 facilitates greater ion entry, enabling depolarization sufficient to reach ST and sustain spiking, whereas its absence reduces depolarization to levels insufficient for spiking.

Article Snippet: Slides were blocked in 10% normal goat serum for 1 hr at room temperature, then incubated overnight at 4°C with anti-TRPV3-Biotin antibody (#ACC-033-B), TRPV3 blocking peptide (#BLP-CC033), and/or anti-TRPV4 antibody (#ACC-034) (Alomone Labs, Israel).

Techniques: Slice Preparation