Review



trpm8 channel plasmid  (Addgene inc)


Bioz Verified Symbol Addgene inc is a verified supplier  
  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
    • 90
      Buy from Supplier

    Structured Review

    Addgene inc trpm8 channel plasmid
    FIGURE 1 Rapamycin activates recombinant <t>TRPM8</t> channels and potentiates cold-evoked responses. (a) Average ± SEM Fura2 ratio time course in HEK293 cells stably expressing rat TRPM8 (CR#1 cells) during application of rapamycin (RAP) at different concentrations. A single dose was applied in individual experiments (n = 38 to 185 cells, eight experiments). (b) Dose– response curve of rapamycin effects. Data shown are means ± SEM obtained from individual peak amplitudes from experiments shown in a. (c) Average ± SEM Fura2 ratio time course in HEK293 cells stably expressing rat TRPM8 during a protocol in which rapamycin responses were monitored in the presence (2.4-mM Ca2+) and in the absence (0 Ca2+) of extracellular calcium (n = 103 cells). (d) Bar histogram summarizing the amplitude ± SEM of rapamycin responses in the presence and absence of extracellular Ca2+ (n = 257 cells, five experiments). Data shown are means ± SEM. *P<0.05, significantly different as indicated; one-way ANOVA for repeated measures followed by Bonferroni post- hoc test. (e) Average ± SEM Fura2 ratio of cold-evoked responses in CR#1 HEK293 cells in control and 30-μM rapamycin (n = 98). (f) Bar histogram summarizing the effect of vehicle or 1-, 10-, 30-μM rapamycin on cold-evoked responses. Individual cold responses have been normalized to the amplitude during the first cold ramp (71 to 92 cells per condition). Ratios >20 have been excluded from the analysis. Data shown are individual values with means ± SEM. *P<0.05, significantly different from vehicle; one-way ANOVA followed by Bonferroni post-hoc test.
    Trpm8 Channel Plasmid, supplied by Addgene inc, used in various techniques. Bioz Stars score: 90/100, based on 4 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/trpm8 channel plasmid/product/Addgene inc
    Average 90 stars, based on 4 article reviews
    trpm8 channel plasmid - by Bioz Stars, 2026-04
    90/100 stars

    Images

    1) Product Images from "The ion channel TRPM8 is a direct target of the immunosuppressant rapamycin in primary sensory neurons."

    Article Title: The ion channel TRPM8 is a direct target of the immunosuppressant rapamycin in primary sensory neurons.

    Journal: British journal of pharmacology

    doi: 10.1111/bph.16402

    FIGURE 1 Rapamycin activates recombinant TRPM8 channels and potentiates cold-evoked responses. (a) Average ± SEM Fura2 ratio time course in HEK293 cells stably expressing rat TRPM8 (CR#1 cells) during application of rapamycin (RAP) at different concentrations. A single dose was applied in individual experiments (n = 38 to 185 cells, eight experiments). (b) Dose– response curve of rapamycin effects. Data shown are means ± SEM obtained from individual peak amplitudes from experiments shown in a. (c) Average ± SEM Fura2 ratio time course in HEK293 cells stably expressing rat TRPM8 during a protocol in which rapamycin responses were monitored in the presence (2.4-mM Ca2+) and in the absence (0 Ca2+) of extracellular calcium (n = 103 cells). (d) Bar histogram summarizing the amplitude ± SEM of rapamycin responses in the presence and absence of extracellular Ca2+ (n = 257 cells, five experiments). Data shown are means ± SEM. *P<0.05, significantly different as indicated; one-way ANOVA for repeated measures followed by Bonferroni post- hoc test. (e) Average ± SEM Fura2 ratio of cold-evoked responses in CR#1 HEK293 cells in control and 30-μM rapamycin (n = 98). (f) Bar histogram summarizing the effect of vehicle or 1-, 10-, 30-μM rapamycin on cold-evoked responses. Individual cold responses have been normalized to the amplitude during the first cold ramp (71 to 92 cells per condition). Ratios >20 have been excluded from the analysis. Data shown are individual values with means ± SEM. *P<0.05, significantly different from vehicle; one-way ANOVA followed by Bonferroni post-hoc test.
    Figure Legend Snippet: FIGURE 1 Rapamycin activates recombinant TRPM8 channels and potentiates cold-evoked responses. (a) Average ± SEM Fura2 ratio time course in HEK293 cells stably expressing rat TRPM8 (CR#1 cells) during application of rapamycin (RAP) at different concentrations. A single dose was applied in individual experiments (n = 38 to 185 cells, eight experiments). (b) Dose– response curve of rapamycin effects. Data shown are means ± SEM obtained from individual peak amplitudes from experiments shown in a. (c) Average ± SEM Fura2 ratio time course in HEK293 cells stably expressing rat TRPM8 during a protocol in which rapamycin responses were monitored in the presence (2.4-mM Ca2+) and in the absence (0 Ca2+) of extracellular calcium (n = 103 cells). (d) Bar histogram summarizing the amplitude ± SEM of rapamycin responses in the presence and absence of extracellular Ca2+ (n = 257 cells, five experiments). Data shown are means ± SEM. *P<0.05, significantly different as indicated; one-way ANOVA for repeated measures followed by Bonferroni post- hoc test. (e) Average ± SEM Fura2 ratio of cold-evoked responses in CR#1 HEK293 cells in control and 30-μM rapamycin (n = 98). (f) Bar histogram summarizing the effect of vehicle or 1-, 10-, 30-μM rapamycin on cold-evoked responses. Individual cold responses have been normalized to the amplitude during the first cold ramp (71 to 92 cells per condition). Ratios >20 have been excluded from the analysis. Data shown are individual values with means ± SEM. *P<0.05, significantly different from vehicle; one-way ANOVA followed by Bonferroni post-hoc test.

    Techniques Used: Recombinant, Stable Transfection, Expressing, Control

    FIGURE 2 Rapamycin activates mouse and human TRPM8 orthologues and the menthol-insensitive mutant. (a) Average ± SEM Fura2 ratio time course in HEK293 cells expressing mouse TRPM8. Three different behaviours were observed; cells which responded to a first cold ramp in control conditions (blue trace), cells which did not respond to this cold ramp in control conditions but responded to cold in the presence of rapamycin (RAP) or menthol (Ment; green trace), and cells which did not show any response to the applied stimuli (black trace). (b) Bar histogram summarizing the individual and mean responses of the cold-sensitive HEK293 cells (n = 162, five experiments). Data shown are individual values with means ± SEM. *P<0.05, significantly different as indicated; one-way ANOVA followed by Bonferroni post-test. (c) Average ± SEM Fura2 ratio time course in HEK293 cells expressing human TRPM8. The different traces represented the three behaviours described in a. (d) Bar histogram summarizing the responses of the cold-sensitive HEK293 cells (n = 178, five experiments) transfected with human TRPM8. Data shown are individual values with means ± SEM. *P<0.05, significantly different as indicated; one-way ANOVA followed by Bonferroni post-test. (e) Average ± SEM Fura2 ratio time course in HEK293 cells expressing mouse TRPM8-Y745H. The different traces represented the three behaviours described in a. (f) Bar histogram summarizing the responses of the cold-sensitive HEK293 cells (n = 182, five experiments) transfected with mouse TRPM8-Y745H. Data shown are individual values with means ± SEM. *P<0.05, significantly different as indicated, n.s., not significant; one-way ANOVA followed by Bonferroni post-test.
    Figure Legend Snippet: FIGURE 2 Rapamycin activates mouse and human TRPM8 orthologues and the menthol-insensitive mutant. (a) Average ± SEM Fura2 ratio time course in HEK293 cells expressing mouse TRPM8. Three different behaviours were observed; cells which responded to a first cold ramp in control conditions (blue trace), cells which did not respond to this cold ramp in control conditions but responded to cold in the presence of rapamycin (RAP) or menthol (Ment; green trace), and cells which did not show any response to the applied stimuli (black trace). (b) Bar histogram summarizing the individual and mean responses of the cold-sensitive HEK293 cells (n = 162, five experiments). Data shown are individual values with means ± SEM. *P<0.05, significantly different as indicated; one-way ANOVA followed by Bonferroni post-test. (c) Average ± SEM Fura2 ratio time course in HEK293 cells expressing human TRPM8. The different traces represented the three behaviours described in a. (d) Bar histogram summarizing the responses of the cold-sensitive HEK293 cells (n = 178, five experiments) transfected with human TRPM8. Data shown are individual values with means ± SEM. *P<0.05, significantly different as indicated; one-way ANOVA followed by Bonferroni post-test. (e) Average ± SEM Fura2 ratio time course in HEK293 cells expressing mouse TRPM8-Y745H. The different traces represented the three behaviours described in a. (f) Bar histogram summarizing the responses of the cold-sensitive HEK293 cells (n = 182, five experiments) transfected with mouse TRPM8-Y745H. Data shown are individual values with means ± SEM. *P<0.05, significantly different as indicated, n.s., not significant; one-way ANOVA followed by Bonferroni post-test.

    Techniques Used: Mutagenesis, Expressing, Control, Transfection

    FIGURE 3 Rapamycin (RAP) activates TRPM8 currents. (a) Representative time course of whole-cell current at 100 and +100 mV in a HEK293 cell transiently transfected with mTRPM8 during the sequential application of different agonists. The bottom trace shows the simultaneous recording of the bath temperature. (b) Current–voltage (I-V) relationship obtained by a 400-ms voltage ramp from 100 to +150 mV during the experiment shown in (a). The colour of individual traces matches the colour at each particular time point in (a). (c) Bar histogram of current density values at 100 and +100 mV evoked by the different stimuli shown in (a), with the same colour code (n = 9 cells). Data shown are individual values with means ± SEM. *P<0.05, significantly different as indicated; one-way ANOVA, followed by Bonferroni's post-hoc test. (d) V1/2 values calculated from fitting individual I-V curves to a linearized Boltzmann equation. Data shown are individual values with means ± SEM. *P<0.05, significantly different from control, &P<0.05, significantly different as indicated; one-way ANOVA, followed by a Bonferroni's post-hoc test. (e) Representative whole-cell current at 100 and +100 mV in a HEK293 cell transiently transfected with mTRPM8 during a protocol in which the effect of AMTB on rapamycin response was explored. Bottom trace corresponds to the simultaneous recording of bath temperature. (f) Current–voltage relationship (I-V) obtained with a 400 ms voltage ramp from 100 to +150 mV of the responses plotted in e. the colour of the I-V curves matches the coloured time points in e. (g) Bar histogram of individual and mean ± SEM current density values at +100 mV evoked by the different stimuli shown in ( e). Data shown are individual values with means ± SEM; n = 5 cells. *P<0.05, significantly different from control, &P<0.05, significantly different as indicated; one-way ANOVA, followed by Bonferroni's post-hoc test.
    Figure Legend Snippet: FIGURE 3 Rapamycin (RAP) activates TRPM8 currents. (a) Representative time course of whole-cell current at 100 and +100 mV in a HEK293 cell transiently transfected with mTRPM8 during the sequential application of different agonists. The bottom trace shows the simultaneous recording of the bath temperature. (b) Current–voltage (I-V) relationship obtained by a 400-ms voltage ramp from 100 to +150 mV during the experiment shown in (a). The colour of individual traces matches the colour at each particular time point in (a). (c) Bar histogram of current density values at 100 and +100 mV evoked by the different stimuli shown in (a), with the same colour code (n = 9 cells). Data shown are individual values with means ± SEM. *P<0.05, significantly different as indicated; one-way ANOVA, followed by Bonferroni's post-hoc test. (d) V1/2 values calculated from fitting individual I-V curves to a linearized Boltzmann equation. Data shown are individual values with means ± SEM. *P<0.05, significantly different from control, &P<0.05, significantly different as indicated; one-way ANOVA, followed by a Bonferroni's post-hoc test. (e) Representative whole-cell current at 100 and +100 mV in a HEK293 cell transiently transfected with mTRPM8 during a protocol in which the effect of AMTB on rapamycin response was explored. Bottom trace corresponds to the simultaneous recording of bath temperature. (f) Current–voltage relationship (I-V) obtained with a 400 ms voltage ramp from 100 to +150 mV of the responses plotted in e. the colour of the I-V curves matches the coloured time points in e. (g) Bar histogram of individual and mean ± SEM current density values at +100 mV evoked by the different stimuli shown in ( e). Data shown are individual values with means ± SEM; n = 5 cells. *P<0.05, significantly different from control, &P<0.05, significantly different as indicated; one-way ANOVA, followed by Bonferroni's post-hoc test.

    Techniques Used: Transfection, Control

    FIGURE 4 Rapamycin activates single TRPM8 channels. (a) Single channel recordings at +100 mV in the cell-attached configuration in HEK293 cells transfected with mTRPM8. The same patch in control (i.e., high K+), rapamycin (RAP; 30 μM) and WS-12 (10 μM). The coloured ticks mark the expanded traces shown below. (b) All-point histograms of current amplitudes for the different recording conditions. The histograms have been fitted with the sum of two Gaussians (red line). (c) Single channel amplitudes of individual patches at different membrane potentials. The dotted lines represent linear fits to the data. (d) Open probability of channel current at different potentials. The dotted lines are fits to the Boltzmann function.
    Figure Legend Snippet: FIGURE 4 Rapamycin activates single TRPM8 channels. (a) Single channel recordings at +100 mV in the cell-attached configuration in HEK293 cells transfected with mTRPM8. The same patch in control (i.e., high K+), rapamycin (RAP; 30 μM) and WS-12 (10 μM). The coloured ticks mark the expanded traces shown below. (b) All-point histograms of current amplitudes for the different recording conditions. The histograms have been fitted with the sum of two Gaussians (red line). (c) Single channel amplitudes of individual patches at different membrane potentials. The dotted lines represent linear fits to the data. (d) Open probability of channel current at different potentials. The dotted lines are fits to the Boltzmann function.

    Techniques Used: Transfection, Control, Membrane

    FIGURE 6 TRPM8 is the principal determinant of rapamycin responses in mouse DRG neurons. (a) Representative traces of Fura2 ratio fluorescence in a Trpm8EGFPf/+ DRG culture. Consecutive applications of cold, rapamycin (RAP, 30 μM), menthol (100 μM), AITC (100 μM), capsaicin (100 nM) and high K+ (30 mM) were used to phenotype each neuron. The green trace corresponds to a EGFPf(+) neuron, which responds to cold, rapamycin, menthol and capsaicin. The other two traces (blue and red) correspond to EGFPf() neurons. Bottom trace shows the simultaneous recording of bath temperature. (b) Venn diagram summarizing the responses to rapamycin in EGFPf(+) and EGFPf() neurons in Trpm8EGFPf/+ DRG cultures. (c) Bar histograms of the amplitude of the responses to different agonists in EGFPf(+) rapamycin-sensitive neurons (n = 20, five experiments). Data shown are individual values with means ± SEM. *P<0.05, significantly different as indicated; one-way ANOVA for repeated measures followed by a Bonferroni's post-hoc test. (d) Correlation between the amplitude of the cold-evoked response and the rapamycin response (n = 20, six experiments). The horizontal dotted line marks the threshold level for rapamycin sensitivity. Green circles represent the three EGFPf(+), cold-sensitive, rapamycin-insensitive neurons recorded. (e) Representative traces of Fura2 ratios in cultured neurons from Trpm8 KO mouse. The protocol was the same as in (a). Orange trace corresponds to a EGFPf(+) neuron that was rapamycin- sensitive. Green trace represents a EGFPf(+) neuron which was not sensitive to any of the agonists tested and the red and blue traces are examples of two EGFPf() neurons. Bottom trace corresponds to the simultaneous recording of the bath temperature during the experiment. (f) Venn diagram summarizing the responses to rapamycin in EGFPf(+) and EGFPf() neurons in Trpm8EGFPf/EGFPf DRG cultures (eight experiments).
    Figure Legend Snippet: FIGURE 6 TRPM8 is the principal determinant of rapamycin responses in mouse DRG neurons. (a) Representative traces of Fura2 ratio fluorescence in a Trpm8EGFPf/+ DRG culture. Consecutive applications of cold, rapamycin (RAP, 30 μM), menthol (100 μM), AITC (100 μM), capsaicin (100 nM) and high K+ (30 mM) were used to phenotype each neuron. The green trace corresponds to a EGFPf(+) neuron, which responds to cold, rapamycin, menthol and capsaicin. The other two traces (blue and red) correspond to EGFPf() neurons. Bottom trace shows the simultaneous recording of bath temperature. (b) Venn diagram summarizing the responses to rapamycin in EGFPf(+) and EGFPf() neurons in Trpm8EGFPf/+ DRG cultures. (c) Bar histograms of the amplitude of the responses to different agonists in EGFPf(+) rapamycin-sensitive neurons (n = 20, five experiments). Data shown are individual values with means ± SEM. *P<0.05, significantly different as indicated; one-way ANOVA for repeated measures followed by a Bonferroni's post-hoc test. (d) Correlation between the amplitude of the cold-evoked response and the rapamycin response (n = 20, six experiments). The horizontal dotted line marks the threshold level for rapamycin sensitivity. Green circles represent the three EGFPf(+), cold-sensitive, rapamycin-insensitive neurons recorded. (e) Representative traces of Fura2 ratios in cultured neurons from Trpm8 KO mouse. The protocol was the same as in (a). Orange trace corresponds to a EGFPf(+) neuron that was rapamycin- sensitive. Green trace represents a EGFPf(+) neuron which was not sensitive to any of the agonists tested and the red and blue traces are examples of two EGFPf() neurons. Bottom trace corresponds to the simultaneous recording of the bath temperature during the experiment. (f) Venn diagram summarizing the responses to rapamycin in EGFPf(+) and EGFPf() neurons in Trpm8EGFPf/EGFPf DRG cultures (eight experiments).

    Techniques Used: Fluorescence, Cell Culture

    FIGURE 7 Rapamycin (RAP) increases the excitability of cold-sensitive DRG neurons. (a) Representative whole-cell recording in the voltage- clamp configuration (Vhold = 60 mV) of a TRPM8-expressing, cold-sensitive DRG neuron identified by EYFP expression. Bottom trace corresponds to the simultaneous recording of bath temperature. Rapamycin activates a small inward current and potentiates the response to cold. (b) Temperature dependence of the cold-evoked current in the same neuron in control conditions and in the presence of 30 μM rapamycin. (c) Bar histogram of peak inward current density values evoked by cold, rapamycin and cold in the presence of rapamycin (n = 12). Data shown are individual values with means ± SEM. #P<0.05, significantly different as indicated; one-way ANOVA for repeated measures followed by Bonferroni's post-hoc test. (d) Representative recording of a cold-sensitive neuron in the whole-cell current-clamp configuration. Cold and rapamycin elicited the firing of action potentials. The combined application of cold and rapamycin led to faster firing, followed by a strong depolarization and the blockade of spikes. (e) Bar histogram of responses, measured as average firing frequency, during the different stimuli applied (n = 11). Firing frequency for cold was the average from the first to the last spike during the cooling ramp. Firing frequency in control conditions was calculated during the 60 s previous to rapamycin application. Rapamycin-evoked firing was calculated from the first spike during rapamycin application to the start of the cold ramp. *P<0.05, significantly different from control, #P<0.05, significantly different as indicated; one- way ANOVA for repeated measures followed by Bonferroni's post-hoc test.
    Figure Legend Snippet: FIGURE 7 Rapamycin (RAP) increases the excitability of cold-sensitive DRG neurons. (a) Representative whole-cell recording in the voltage- clamp configuration (Vhold = 60 mV) of a TRPM8-expressing, cold-sensitive DRG neuron identified by EYFP expression. Bottom trace corresponds to the simultaneous recording of bath temperature. Rapamycin activates a small inward current and potentiates the response to cold. (b) Temperature dependence of the cold-evoked current in the same neuron in control conditions and in the presence of 30 μM rapamycin. (c) Bar histogram of peak inward current density values evoked by cold, rapamycin and cold in the presence of rapamycin (n = 12). Data shown are individual values with means ± SEM. #P<0.05, significantly different as indicated; one-way ANOVA for repeated measures followed by Bonferroni's post-hoc test. (d) Representative recording of a cold-sensitive neuron in the whole-cell current-clamp configuration. Cold and rapamycin elicited the firing of action potentials. The combined application of cold and rapamycin led to faster firing, followed by a strong depolarization and the blockade of spikes. (e) Bar histogram of responses, measured as average firing frequency, during the different stimuli applied (n = 11). Firing frequency for cold was the average from the first to the last spike during the cooling ramp. Firing frequency in control conditions was calculated during the 60 s previous to rapamycin application. Rapamycin-evoked firing was calculated from the first spike during rapamycin application to the start of the cold ramp. *P<0.05, significantly different from control, #P<0.05, significantly different as indicated; one- way ANOVA for repeated measures followed by Bonferroni's post-hoc test.

    Techniques Used: Expressing, Control

    FIGURE 9 Mathematical modelling of the effects of rapamycin (RAP) on cold thermoreceptor activity. (a) Membrane potential (blue trace), temperature (red trace), and action potential firing frequency (blue dots), as a function of time. The green bar indicates the period of application of agonist (i.e. a change of -100 mV in V1/2 in this example). The red dots in the temperature trace indicate where the temperature threshold has been reached. The black dashed lines in the lower panel indicate the value of the average firing frequency, and the time range where it has been calculated (i.e. the duration of the temperature ramps). (b) Firing temperature threshold as a function of the shift in V1/2. Each trace corresponds to a cell with different parameters, which have been selected in order to encompass a wide range of thresholds under control conditions. The larger dot corresponds to the cell and conditions simulated in panel (a). (c) Same as in panel B for the average firing frequency during cold stimulation. (d) TRPM8 current density (blue trace) as a function of time, and changes in temperature (red trace) in voltage-clamp simulations. The green horizontal bar represents the application of agonist, i.e. a shift of -100 mV in V1/2 as in (a). (e) TRPM8 current density as a function of temperature with (green trace) or without agonist (blue trace). (f) Peak cold-evoked current as a function of the shift in V1/2 for the same cells as shown above with the corresponding colours. Values for V1/2 = 0 correspond to the cold-evoked current in the absence of agonist. The magenta line corresponds to a neuron with very small TRPM8 currents that did not fire action potentials and thus is not shown in the current-clamp panels above. Color code is the same for panels b, c, e and f. The set numbers in panel f correspond to the row numbers within the table found in Extended Data Figure 11-1 of Rivera et al. (2021), detailing the simulation parameter values.
    Figure Legend Snippet: FIGURE 9 Mathematical modelling of the effects of rapamycin (RAP) on cold thermoreceptor activity. (a) Membrane potential (blue trace), temperature (red trace), and action potential firing frequency (blue dots), as a function of time. The green bar indicates the period of application of agonist (i.e. a change of -100 mV in V1/2 in this example). The red dots in the temperature trace indicate where the temperature threshold has been reached. The black dashed lines in the lower panel indicate the value of the average firing frequency, and the time range where it has been calculated (i.e. the duration of the temperature ramps). (b) Firing temperature threshold as a function of the shift in V1/2. Each trace corresponds to a cell with different parameters, which have been selected in order to encompass a wide range of thresholds under control conditions. The larger dot corresponds to the cell and conditions simulated in panel (a). (c) Same as in panel B for the average firing frequency during cold stimulation. (d) TRPM8 current density (blue trace) as a function of time, and changes in temperature (red trace) in voltage-clamp simulations. The green horizontal bar represents the application of agonist, i.e. a shift of -100 mV in V1/2 as in (a). (e) TRPM8 current density as a function of temperature with (green trace) or without agonist (blue trace). (f) Peak cold-evoked current as a function of the shift in V1/2 for the same cells as shown above with the corresponding colours. Values for V1/2 = 0 correspond to the cold-evoked current in the absence of agonist. The magenta line corresponds to a neuron with very small TRPM8 currents that did not fire action potentials and thus is not shown in the current-clamp panels above. Color code is the same for panels b, c, e and f. The set numbers in panel f correspond to the row numbers within the table found in Extended Data Figure 11-1 of Rivera et al. (2021), detailing the simulation parameter values.

    Techniques Used: Activity Assay, Membrane, Control

    FIGURE 10 Rapamycin stimulates tearing by a TRPM8-dependent mechanism. Effect of topical solutions of rapamycin (RAP; 1%) or vehicle on tearing in mice. Tearing is represented as the length of staining in the threads (in mm) placed in the eye. Each dot corresponds to the tearing measured in an eye of an individual mouse (15 WT and 15 Trpm8 KO). Data shown are individual values of evoked tearing, with means ± SEM. *P < 0.05, significantly different as indicated; n.s., not significant; Mann– Whitney test).
    Figure Legend Snippet: FIGURE 10 Rapamycin stimulates tearing by a TRPM8-dependent mechanism. Effect of topical solutions of rapamycin (RAP; 1%) or vehicle on tearing in mice. Tearing is represented as the length of staining in the threads (in mm) placed in the eye. Each dot corresponds to the tearing measured in an eye of an individual mouse (15 WT and 15 Trpm8 KO). Data shown are individual values of evoked tearing, with means ± SEM. *P < 0.05, significantly different as indicated; n.s., not significant; Mann– Whitney test).

    Techniques Used: Staining, MANN-WHITNEY



    Similar Products

    trpm8 channel plasmid  (Addgene inc)
    90
    Addgene inc trpm8 channel plasmid
    FIGURE 1 Rapamycin activates recombinant <t>TRPM8</t> channels and potentiates cold-evoked responses. (a) Average ± SEM Fura2 ratio time course in HEK293 cells stably expressing rat TRPM8 (CR#1 cells) during application of rapamycin (RAP) at different concentrations. A single dose was applied in individual experiments (n = 38 to 185 cells, eight experiments). (b) Dose– response curve of rapamycin effects. Data shown are means ± SEM obtained from individual peak amplitudes from experiments shown in a. (c) Average ± SEM Fura2 ratio time course in HEK293 cells stably expressing rat TRPM8 during a protocol in which rapamycin responses were monitored in the presence (2.4-mM Ca2+) and in the absence (0 Ca2+) of extracellular calcium (n = 103 cells). (d) Bar histogram summarizing the amplitude ± SEM of rapamycin responses in the presence and absence of extracellular Ca2+ (n = 257 cells, five experiments). Data shown are means ± SEM. *P<0.05, significantly different as indicated; one-way ANOVA for repeated measures followed by Bonferroni post- hoc test. (e) Average ± SEM Fura2 ratio of cold-evoked responses in CR#1 HEK293 cells in control and 30-μM rapamycin (n = 98). (f) Bar histogram summarizing the effect of vehicle or 1-, 10-, 30-μM rapamycin on cold-evoked responses. Individual cold responses have been normalized to the amplitude during the first cold ramp (71 to 92 cells per condition). Ratios >20 have been excluded from the analysis. Data shown are individual values with means ± SEM. *P<0.05, significantly different from vehicle; one-way ANOVA followed by Bonferroni post-hoc test.
    Trpm8 Channel Plasmid, supplied by Addgene inc, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/trpm8 channel plasmid/product/Addgene inc
    Average 90 stars, based on 1 article reviews
    trpm8 channel plasmid - by Bioz Stars, 2026-04
    90/100 stars
    		  Buy from Supplier

    Image Search Results


    FIGURE 1 Rapamycin activates recombinant TRPM8 channels and potentiates cold-evoked responses. (a) Average ± SEM Fura2 ratio time course in HEK293 cells stably expressing rat TRPM8 (CR#1 cells) during application of rapamycin (RAP) at different concentrations. A single dose was applied in individual experiments (n = 38 to 185 cells, eight experiments). (b) Dose– response curve of rapamycin effects. Data shown are means ± SEM obtained from individual peak amplitudes from experiments shown in a. (c) Average ± SEM Fura2 ratio time course in HEK293 cells stably expressing rat TRPM8 during a protocol in which rapamycin responses were monitored in the presence (2.4-mM Ca2+) and in the absence (0 Ca2+) of extracellular calcium (n = 103 cells). (d) Bar histogram summarizing the amplitude ± SEM of rapamycin responses in the presence and absence of extracellular Ca2+ (n = 257 cells, five experiments). Data shown are means ± SEM. *P<0.05, significantly different as indicated; one-way ANOVA for repeated measures followed by Bonferroni post- hoc test. (e) Average ± SEM Fura2 ratio of cold-evoked responses in CR#1 HEK293 cells in control and 30-μM rapamycin (n = 98). (f) Bar histogram summarizing the effect of vehicle or 1-, 10-, 30-μM rapamycin on cold-evoked responses. Individual cold responses have been normalized to the amplitude during the first cold ramp (71 to 92 cells per condition). Ratios >20 have been excluded from the analysis. Data shown are individual values with means ± SEM. *P<0.05, significantly different from vehicle; one-way ANOVA followed by Bonferroni post-hoc test.

    Journal: British journal of pharmacology

    Article Title: The ion channel TRPM8 is a direct target of the immunosuppressant rapamycin in primary sensory neurons.

    doi: 10.1111/bph.16402

    Figure Lengend Snippet: FIGURE 1 Rapamycin activates recombinant TRPM8 channels and potentiates cold-evoked responses. (a) Average ± SEM Fura2 ratio time course in HEK293 cells stably expressing rat TRPM8 (CR#1 cells) during application of rapamycin (RAP) at different concentrations. A single dose was applied in individual experiments (n = 38 to 185 cells, eight experiments). (b) Dose– response curve of rapamycin effects. Data shown are means ± SEM obtained from individual peak amplitudes from experiments shown in a. (c) Average ± SEM Fura2 ratio time course in HEK293 cells stably expressing rat TRPM8 during a protocol in which rapamycin responses were monitored in the presence (2.4-mM Ca2+) and in the absence (0 Ca2+) of extracellular calcium (n = 103 cells). (d) Bar histogram summarizing the amplitude ± SEM of rapamycin responses in the presence and absence of extracellular Ca2+ (n = 257 cells, five experiments). Data shown are means ± SEM. *P<0.05, significantly different as indicated; one-way ANOVA for repeated measures followed by Bonferroni post- hoc test. (e) Average ± SEM Fura2 ratio of cold-evoked responses in CR#1 HEK293 cells in control and 30-μM rapamycin (n = 98). (f) Bar histogram summarizing the effect of vehicle or 1-, 10-, 30-μM rapamycin on cold-evoked responses. Individual cold responses have been normalized to the amplitude during the first cold ramp (71 to 92 cells per condition). Ratios >20 have been excluded from the analysis. Data shown are individual values with means ± SEM. *P<0.05, significantly different from vehicle; one-way ANOVA followed by Bonferroni post-hoc test.

    Article Snippet: When necessary, we co-transfected the cells with 1 μg of TRPM8 channel plasmid (from different species) and 0.5 μg of EGFP or mCherry plasmid (Addgene_84329) (van Unen et al., 2016).

    Techniques: Recombinant, Stable Transfection, Expressing, Control

    FIGURE 2 Rapamycin activates mouse and human TRPM8 orthologues and the menthol-insensitive mutant. (a) Average ± SEM Fura2 ratio time course in HEK293 cells expressing mouse TRPM8. Three different behaviours were observed; cells which responded to a first cold ramp in control conditions (blue trace), cells which did not respond to this cold ramp in control conditions but responded to cold in the presence of rapamycin (RAP) or menthol (Ment; green trace), and cells which did not show any response to the applied stimuli (black trace). (b) Bar histogram summarizing the individual and mean responses of the cold-sensitive HEK293 cells (n = 162, five experiments). Data shown are individual values with means ± SEM. *P<0.05, significantly different as indicated; one-way ANOVA followed by Bonferroni post-test. (c) Average ± SEM Fura2 ratio time course in HEK293 cells expressing human TRPM8. The different traces represented the three behaviours described in a. (d) Bar histogram summarizing the responses of the cold-sensitive HEK293 cells (n = 178, five experiments) transfected with human TRPM8. Data shown are individual values with means ± SEM. *P<0.05, significantly different as indicated; one-way ANOVA followed by Bonferroni post-test. (e) Average ± SEM Fura2 ratio time course in HEK293 cells expressing mouse TRPM8-Y745H. The different traces represented the three behaviours described in a. (f) Bar histogram summarizing the responses of the cold-sensitive HEK293 cells (n = 182, five experiments) transfected with mouse TRPM8-Y745H. Data shown are individual values with means ± SEM. *P<0.05, significantly different as indicated, n.s., not significant; one-way ANOVA followed by Bonferroni post-test.

    Journal: British journal of pharmacology

    Article Title: The ion channel TRPM8 is a direct target of the immunosuppressant rapamycin in primary sensory neurons.

    doi: 10.1111/bph.16402

    Figure Lengend Snippet: FIGURE 2 Rapamycin activates mouse and human TRPM8 orthologues and the menthol-insensitive mutant. (a) Average ± SEM Fura2 ratio time course in HEK293 cells expressing mouse TRPM8. Three different behaviours were observed; cells which responded to a first cold ramp in control conditions (blue trace), cells which did not respond to this cold ramp in control conditions but responded to cold in the presence of rapamycin (RAP) or menthol (Ment; green trace), and cells which did not show any response to the applied stimuli (black trace). (b) Bar histogram summarizing the individual and mean responses of the cold-sensitive HEK293 cells (n = 162, five experiments). Data shown are individual values with means ± SEM. *P<0.05, significantly different as indicated; one-way ANOVA followed by Bonferroni post-test. (c) Average ± SEM Fura2 ratio time course in HEK293 cells expressing human TRPM8. The different traces represented the three behaviours described in a. (d) Bar histogram summarizing the responses of the cold-sensitive HEK293 cells (n = 178, five experiments) transfected with human TRPM8. Data shown are individual values with means ± SEM. *P<0.05, significantly different as indicated; one-way ANOVA followed by Bonferroni post-test. (e) Average ± SEM Fura2 ratio time course in HEK293 cells expressing mouse TRPM8-Y745H. The different traces represented the three behaviours described in a. (f) Bar histogram summarizing the responses of the cold-sensitive HEK293 cells (n = 182, five experiments) transfected with mouse TRPM8-Y745H. Data shown are individual values with means ± SEM. *P<0.05, significantly different as indicated, n.s., not significant; one-way ANOVA followed by Bonferroni post-test.

    Article Snippet: When necessary, we co-transfected the cells with 1 μg of TRPM8 channel plasmid (from different species) and 0.5 μg of EGFP or mCherry plasmid (Addgene_84329) (van Unen et al., 2016).

    Techniques: Mutagenesis, Expressing, Control, Transfection

    FIGURE 3 Rapamycin (RAP) activates TRPM8 currents. (a) Representative time course of whole-cell current at 100 and +100 mV in a HEK293 cell transiently transfected with mTRPM8 during the sequential application of different agonists. The bottom trace shows the simultaneous recording of the bath temperature. (b) Current–voltage (I-V) relationship obtained by a 400-ms voltage ramp from 100 to +150 mV during the experiment shown in (a). The colour of individual traces matches the colour at each particular time point in (a). (c) Bar histogram of current density values at 100 and +100 mV evoked by the different stimuli shown in (a), with the same colour code (n = 9 cells). Data shown are individual values with means ± SEM. *P<0.05, significantly different as indicated; one-way ANOVA, followed by Bonferroni's post-hoc test. (d) V1/2 values calculated from fitting individual I-V curves to a linearized Boltzmann equation. Data shown are individual values with means ± SEM. *P<0.05, significantly different from control, &P<0.05, significantly different as indicated; one-way ANOVA, followed by a Bonferroni's post-hoc test. (e) Representative whole-cell current at 100 and +100 mV in a HEK293 cell transiently transfected with mTRPM8 during a protocol in which the effect of AMTB on rapamycin response was explored. Bottom trace corresponds to the simultaneous recording of bath temperature. (f) Current–voltage relationship (I-V) obtained with a 400 ms voltage ramp from 100 to +150 mV of the responses plotted in e. the colour of the I-V curves matches the coloured time points in e. (g) Bar histogram of individual and mean ± SEM current density values at +100 mV evoked by the different stimuli shown in ( e). Data shown are individual values with means ± SEM; n = 5 cells. *P<0.05, significantly different from control, &P<0.05, significantly different as indicated; one-way ANOVA, followed by Bonferroni's post-hoc test.

    Journal: British journal of pharmacology

    Article Title: The ion channel TRPM8 is a direct target of the immunosuppressant rapamycin in primary sensory neurons.

    doi: 10.1111/bph.16402

    Figure Lengend Snippet: FIGURE 3 Rapamycin (RAP) activates TRPM8 currents. (a) Representative time course of whole-cell current at 100 and +100 mV in a HEK293 cell transiently transfected with mTRPM8 during the sequential application of different agonists. The bottom trace shows the simultaneous recording of the bath temperature. (b) Current–voltage (I-V) relationship obtained by a 400-ms voltage ramp from 100 to +150 mV during the experiment shown in (a). The colour of individual traces matches the colour at each particular time point in (a). (c) Bar histogram of current density values at 100 and +100 mV evoked by the different stimuli shown in (a), with the same colour code (n = 9 cells). Data shown are individual values with means ± SEM. *P<0.05, significantly different as indicated; one-way ANOVA, followed by Bonferroni's post-hoc test. (d) V1/2 values calculated from fitting individual I-V curves to a linearized Boltzmann equation. Data shown are individual values with means ± SEM. *P<0.05, significantly different from control, &P<0.05, significantly different as indicated; one-way ANOVA, followed by a Bonferroni's post-hoc test. (e) Representative whole-cell current at 100 and +100 mV in a HEK293 cell transiently transfected with mTRPM8 during a protocol in which the effect of AMTB on rapamycin response was explored. Bottom trace corresponds to the simultaneous recording of bath temperature. (f) Current–voltage relationship (I-V) obtained with a 400 ms voltage ramp from 100 to +150 mV of the responses plotted in e. the colour of the I-V curves matches the coloured time points in e. (g) Bar histogram of individual and mean ± SEM current density values at +100 mV evoked by the different stimuli shown in ( e). Data shown are individual values with means ± SEM; n = 5 cells. *P<0.05, significantly different from control, &P<0.05, significantly different as indicated; one-way ANOVA, followed by Bonferroni's post-hoc test.

    Article Snippet: When necessary, we co-transfected the cells with 1 μg of TRPM8 channel plasmid (from different species) and 0.5 μg of EGFP or mCherry plasmid (Addgene_84329) (van Unen et al., 2016).

    Techniques: Transfection, Control

    FIGURE 4 Rapamycin activates single TRPM8 channels. (a) Single channel recordings at +100 mV in the cell-attached configuration in HEK293 cells transfected with mTRPM8. The same patch in control (i.e., high K+), rapamycin (RAP; 30 μM) and WS-12 (10 μM). The coloured ticks mark the expanded traces shown below. (b) All-point histograms of current amplitudes for the different recording conditions. The histograms have been fitted with the sum of two Gaussians (red line). (c) Single channel amplitudes of individual patches at different membrane potentials. The dotted lines represent linear fits to the data. (d) Open probability of channel current at different potentials. The dotted lines are fits to the Boltzmann function.

    Journal: British journal of pharmacology

    Article Title: The ion channel TRPM8 is a direct target of the immunosuppressant rapamycin in primary sensory neurons.

    doi: 10.1111/bph.16402

    Figure Lengend Snippet: FIGURE 4 Rapamycin activates single TRPM8 channels. (a) Single channel recordings at +100 mV in the cell-attached configuration in HEK293 cells transfected with mTRPM8. The same patch in control (i.e., high K+), rapamycin (RAP; 30 μM) and WS-12 (10 μM). The coloured ticks mark the expanded traces shown below. (b) All-point histograms of current amplitudes for the different recording conditions. The histograms have been fitted with the sum of two Gaussians (red line). (c) Single channel amplitudes of individual patches at different membrane potentials. The dotted lines represent linear fits to the data. (d) Open probability of channel current at different potentials. The dotted lines are fits to the Boltzmann function.

    Article Snippet: When necessary, we co-transfected the cells with 1 μg of TRPM8 channel plasmid (from different species) and 0.5 μg of EGFP or mCherry plasmid (Addgene_84329) (van Unen et al., 2016).

    Techniques: Transfection, Control, Membrane

    FIGURE 6 TRPM8 is the principal determinant of rapamycin responses in mouse DRG neurons. (a) Representative traces of Fura2 ratio fluorescence in a Trpm8EGFPf/+ DRG culture. Consecutive applications of cold, rapamycin (RAP, 30 μM), menthol (100 μM), AITC (100 μM), capsaicin (100 nM) and high K+ (30 mM) were used to phenotype each neuron. The green trace corresponds to a EGFPf(+) neuron, which responds to cold, rapamycin, menthol and capsaicin. The other two traces (blue and red) correspond to EGFPf() neurons. Bottom trace shows the simultaneous recording of bath temperature. (b) Venn diagram summarizing the responses to rapamycin in EGFPf(+) and EGFPf() neurons in Trpm8EGFPf/+ DRG cultures. (c) Bar histograms of the amplitude of the responses to different agonists in EGFPf(+) rapamycin-sensitive neurons (n = 20, five experiments). Data shown are individual values with means ± SEM. *P<0.05, significantly different as indicated; one-way ANOVA for repeated measures followed by a Bonferroni's post-hoc test. (d) Correlation between the amplitude of the cold-evoked response and the rapamycin response (n = 20, six experiments). The horizontal dotted line marks the threshold level for rapamycin sensitivity. Green circles represent the three EGFPf(+), cold-sensitive, rapamycin-insensitive neurons recorded. (e) Representative traces of Fura2 ratios in cultured neurons from Trpm8 KO mouse. The protocol was the same as in (a). Orange trace corresponds to a EGFPf(+) neuron that was rapamycin- sensitive. Green trace represents a EGFPf(+) neuron which was not sensitive to any of the agonists tested and the red and blue traces are examples of two EGFPf() neurons. Bottom trace corresponds to the simultaneous recording of the bath temperature during the experiment. (f) Venn diagram summarizing the responses to rapamycin in EGFPf(+) and EGFPf() neurons in Trpm8EGFPf/EGFPf DRG cultures (eight experiments).

    Journal: British journal of pharmacology

    Article Title: The ion channel TRPM8 is a direct target of the immunosuppressant rapamycin in primary sensory neurons.

    doi: 10.1111/bph.16402

    Figure Lengend Snippet: FIGURE 6 TRPM8 is the principal determinant of rapamycin responses in mouse DRG neurons. (a) Representative traces of Fura2 ratio fluorescence in a Trpm8EGFPf/+ DRG culture. Consecutive applications of cold, rapamycin (RAP, 30 μM), menthol (100 μM), AITC (100 μM), capsaicin (100 nM) and high K+ (30 mM) were used to phenotype each neuron. The green trace corresponds to a EGFPf(+) neuron, which responds to cold, rapamycin, menthol and capsaicin. The other two traces (blue and red) correspond to EGFPf() neurons. Bottom trace shows the simultaneous recording of bath temperature. (b) Venn diagram summarizing the responses to rapamycin in EGFPf(+) and EGFPf() neurons in Trpm8EGFPf/+ DRG cultures. (c) Bar histograms of the amplitude of the responses to different agonists in EGFPf(+) rapamycin-sensitive neurons (n = 20, five experiments). Data shown are individual values with means ± SEM. *P<0.05, significantly different as indicated; one-way ANOVA for repeated measures followed by a Bonferroni's post-hoc test. (d) Correlation between the amplitude of the cold-evoked response and the rapamycin response (n = 20, six experiments). The horizontal dotted line marks the threshold level for rapamycin sensitivity. Green circles represent the three EGFPf(+), cold-sensitive, rapamycin-insensitive neurons recorded. (e) Representative traces of Fura2 ratios in cultured neurons from Trpm8 KO mouse. The protocol was the same as in (a). Orange trace corresponds to a EGFPf(+) neuron that was rapamycin- sensitive. Green trace represents a EGFPf(+) neuron which was not sensitive to any of the agonists tested and the red and blue traces are examples of two EGFPf() neurons. Bottom trace corresponds to the simultaneous recording of the bath temperature during the experiment. (f) Venn diagram summarizing the responses to rapamycin in EGFPf(+) and EGFPf() neurons in Trpm8EGFPf/EGFPf DRG cultures (eight experiments).

    Article Snippet: When necessary, we co-transfected the cells with 1 μg of TRPM8 channel plasmid (from different species) and 0.5 μg of EGFP or mCherry plasmid (Addgene_84329) (van Unen et al., 2016).

    Techniques: Fluorescence, Cell Culture

    FIGURE 7 Rapamycin (RAP) increases the excitability of cold-sensitive DRG neurons. (a) Representative whole-cell recording in the voltage- clamp configuration (Vhold = 60 mV) of a TRPM8-expressing, cold-sensitive DRG neuron identified by EYFP expression. Bottom trace corresponds to the simultaneous recording of bath temperature. Rapamycin activates a small inward current and potentiates the response to cold. (b) Temperature dependence of the cold-evoked current in the same neuron in control conditions and in the presence of 30 μM rapamycin. (c) Bar histogram of peak inward current density values evoked by cold, rapamycin and cold in the presence of rapamycin (n = 12). Data shown are individual values with means ± SEM. #P<0.05, significantly different as indicated; one-way ANOVA for repeated measures followed by Bonferroni's post-hoc test. (d) Representative recording of a cold-sensitive neuron in the whole-cell current-clamp configuration. Cold and rapamycin elicited the firing of action potentials. The combined application of cold and rapamycin led to faster firing, followed by a strong depolarization and the blockade of spikes. (e) Bar histogram of responses, measured as average firing frequency, during the different stimuli applied (n = 11). Firing frequency for cold was the average from the first to the last spike during the cooling ramp. Firing frequency in control conditions was calculated during the 60 s previous to rapamycin application. Rapamycin-evoked firing was calculated from the first spike during rapamycin application to the start of the cold ramp. *P<0.05, significantly different from control, #P<0.05, significantly different as indicated; one- way ANOVA for repeated measures followed by Bonferroni's post-hoc test.

    Journal: British journal of pharmacology

    Article Title: The ion channel TRPM8 is a direct target of the immunosuppressant rapamycin in primary sensory neurons.

    doi: 10.1111/bph.16402

    Figure Lengend Snippet: FIGURE 7 Rapamycin (RAP) increases the excitability of cold-sensitive DRG neurons. (a) Representative whole-cell recording in the voltage- clamp configuration (Vhold = 60 mV) of a TRPM8-expressing, cold-sensitive DRG neuron identified by EYFP expression. Bottom trace corresponds to the simultaneous recording of bath temperature. Rapamycin activates a small inward current and potentiates the response to cold. (b) Temperature dependence of the cold-evoked current in the same neuron in control conditions and in the presence of 30 μM rapamycin. (c) Bar histogram of peak inward current density values evoked by cold, rapamycin and cold in the presence of rapamycin (n = 12). Data shown are individual values with means ± SEM. #P<0.05, significantly different as indicated; one-way ANOVA for repeated measures followed by Bonferroni's post-hoc test. (d) Representative recording of a cold-sensitive neuron in the whole-cell current-clamp configuration. Cold and rapamycin elicited the firing of action potentials. The combined application of cold and rapamycin led to faster firing, followed by a strong depolarization and the blockade of spikes. (e) Bar histogram of responses, measured as average firing frequency, during the different stimuli applied (n = 11). Firing frequency for cold was the average from the first to the last spike during the cooling ramp. Firing frequency in control conditions was calculated during the 60 s previous to rapamycin application. Rapamycin-evoked firing was calculated from the first spike during rapamycin application to the start of the cold ramp. *P<0.05, significantly different from control, #P<0.05, significantly different as indicated; one- way ANOVA for repeated measures followed by Bonferroni's post-hoc test.

    Article Snippet: When necessary, we co-transfected the cells with 1 μg of TRPM8 channel plasmid (from different species) and 0.5 μg of EGFP or mCherry plasmid (Addgene_84329) (van Unen et al., 2016).

    Techniques: Expressing, Control

    FIGURE 9 Mathematical modelling of the effects of rapamycin (RAP) on cold thermoreceptor activity. (a) Membrane potential (blue trace), temperature (red trace), and action potential firing frequency (blue dots), as a function of time. The green bar indicates the period of application of agonist (i.e. a change of -100 mV in V1/2 in this example). The red dots in the temperature trace indicate where the temperature threshold has been reached. The black dashed lines in the lower panel indicate the value of the average firing frequency, and the time range where it has been calculated (i.e. the duration of the temperature ramps). (b) Firing temperature threshold as a function of the shift in V1/2. Each trace corresponds to a cell with different parameters, which have been selected in order to encompass a wide range of thresholds under control conditions. The larger dot corresponds to the cell and conditions simulated in panel (a). (c) Same as in panel B for the average firing frequency during cold stimulation. (d) TRPM8 current density (blue trace) as a function of time, and changes in temperature (red trace) in voltage-clamp simulations. The green horizontal bar represents the application of agonist, i.e. a shift of -100 mV in V1/2 as in (a). (e) TRPM8 current density as a function of temperature with (green trace) or without agonist (blue trace). (f) Peak cold-evoked current as a function of the shift in V1/2 for the same cells as shown above with the corresponding colours. Values for V1/2 = 0 correspond to the cold-evoked current in the absence of agonist. The magenta line corresponds to a neuron with very small TRPM8 currents that did not fire action potentials and thus is not shown in the current-clamp panels above. Color code is the same for panels b, c, e and f. The set numbers in panel f correspond to the row numbers within the table found in Extended Data Figure 11-1 of Rivera et al. (2021), detailing the simulation parameter values.

    Journal: British journal of pharmacology

    Article Title: The ion channel TRPM8 is a direct target of the immunosuppressant rapamycin in primary sensory neurons.

    doi: 10.1111/bph.16402

    Figure Lengend Snippet: FIGURE 9 Mathematical modelling of the effects of rapamycin (RAP) on cold thermoreceptor activity. (a) Membrane potential (blue trace), temperature (red trace), and action potential firing frequency (blue dots), as a function of time. The green bar indicates the period of application of agonist (i.e. a change of -100 mV in V1/2 in this example). The red dots in the temperature trace indicate where the temperature threshold has been reached. The black dashed lines in the lower panel indicate the value of the average firing frequency, and the time range where it has been calculated (i.e. the duration of the temperature ramps). (b) Firing temperature threshold as a function of the shift in V1/2. Each trace corresponds to a cell with different parameters, which have been selected in order to encompass a wide range of thresholds under control conditions. The larger dot corresponds to the cell and conditions simulated in panel (a). (c) Same as in panel B for the average firing frequency during cold stimulation. (d) TRPM8 current density (blue trace) as a function of time, and changes in temperature (red trace) in voltage-clamp simulations. The green horizontal bar represents the application of agonist, i.e. a shift of -100 mV in V1/2 as in (a). (e) TRPM8 current density as a function of temperature with (green trace) or without agonist (blue trace). (f) Peak cold-evoked current as a function of the shift in V1/2 for the same cells as shown above with the corresponding colours. Values for V1/2 = 0 correspond to the cold-evoked current in the absence of agonist. The magenta line corresponds to a neuron with very small TRPM8 currents that did not fire action potentials and thus is not shown in the current-clamp panels above. Color code is the same for panels b, c, e and f. The set numbers in panel f correspond to the row numbers within the table found in Extended Data Figure 11-1 of Rivera et al. (2021), detailing the simulation parameter values.

    Article Snippet: When necessary, we co-transfected the cells with 1 μg of TRPM8 channel plasmid (from different species) and 0.5 μg of EGFP or mCherry plasmid (Addgene_84329) (van Unen et al., 2016).

    Techniques: Activity Assay, Membrane, Control

    FIGURE 10 Rapamycin stimulates tearing by a TRPM8-dependent mechanism. Effect of topical solutions of rapamycin (RAP; 1%) or vehicle on tearing in mice. Tearing is represented as the length of staining in the threads (in mm) placed in the eye. Each dot corresponds to the tearing measured in an eye of an individual mouse (15 WT and 15 Trpm8 KO). Data shown are individual values of evoked tearing, with means ± SEM. *P < 0.05, significantly different as indicated; n.s., not significant; Mann– Whitney test).

    Journal: British journal of pharmacology

    Article Title: The ion channel TRPM8 is a direct target of the immunosuppressant rapamycin in primary sensory neurons.

    doi: 10.1111/bph.16402

    Figure Lengend Snippet: FIGURE 10 Rapamycin stimulates tearing by a TRPM8-dependent mechanism. Effect of topical solutions of rapamycin (RAP; 1%) or vehicle on tearing in mice. Tearing is represented as the length of staining in the threads (in mm) placed in the eye. Each dot corresponds to the tearing measured in an eye of an individual mouse (15 WT and 15 Trpm8 KO). Data shown are individual values of evoked tearing, with means ± SEM. *P < 0.05, significantly different as indicated; n.s., not significant; Mann– Whitney test).

    Article Snippet: When necessary, we co-transfected the cells with 1 μg of TRPM8 channel plasmid (from different species) and 0.5 μg of EGFP or mCherry plasmid (Addgene_84329) (van Unen et al., 2016).

    Techniques: Staining, MANN-WHITNEY