rabbit polyclonal anti trpm4 antibody  (Alomone Labs)


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    Structured Review

    Alomone Labs rabbit polyclonal anti trpm4 antibody
    Treatment of cells with cholesterol increases, but with lovastatin decreases, <t>TRPM4</t> channel activity by regulating its sensitivity to Ca 2+ in mpkCCD c14 Cells. (A) Representative single channel recording from inside-out patches exposed the patch membrane to the bath containing different concentrations of free Ca 2+ . The cells were treated with 30 μg/ml cholesterol for 48 hrs. From top to bottom: 1 μM, 10 μM, 200 μM, 1 mM, and 5 mM free bath Ca 2+ . “C” indicates channel at the closed state; “O” indicates single-level openings. (B) Representative single channel recording from inside-out patches exposed the patch membrane to the bath containing different concentrations of free Ca 2+ . The cells were treated with 5 μM lovastatin for 48 hrs. From top to bottom: 10 μM, 200 μM, 1 mM, 5 mM and 10 mM free bath Ca 2+ . “C” indicates channel at the closed state; “O” indicates single-level openings. (C) The effect of membrane cholesterol on Ca 2+ -dependence of channel opening. Channel P O was plotted as a function of free Ca 2+ concentration in the bath. P o values are shown for patches either with exogenous cholesterol treatment (black line) or lovastatin treatment (red line). n = 4–7 cells for different data points.
    Rabbit Polyclonal Anti Trpm4 Antibody, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/rabbit polyclonal anti trpm4 antibody/product/Alomone Labs
    Average 94 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    rabbit polyclonal anti trpm4 antibody - by Bioz Stars, 2023-02
    94/100 stars

    Images

    1) Product Images from "Cholesterol Stimulates the Transient Receptor Potential Melastatin 4 Channel in mpkCCD c14 Cells"

    Article Title: Cholesterol Stimulates the Transient Receptor Potential Melastatin 4 Channel in mpkCCD c14 Cells

    Journal: Frontiers in Pharmacology

    doi: 10.3389/fphar.2021.627875

    Treatment of cells with cholesterol increases, but with lovastatin decreases, TRPM4 channel activity by regulating its sensitivity to Ca 2+ in mpkCCD c14 Cells. (A) Representative single channel recording from inside-out patches exposed the patch membrane to the bath containing different concentrations of free Ca 2+ . The cells were treated with 30 μg/ml cholesterol for 48 hrs. From top to bottom: 1 μM, 10 μM, 200 μM, 1 mM, and 5 mM free bath Ca 2+ . “C” indicates channel at the closed state; “O” indicates single-level openings. (B) Representative single channel recording from inside-out patches exposed the patch membrane to the bath containing different concentrations of free Ca 2+ . The cells were treated with 5 μM lovastatin for 48 hrs. From top to bottom: 10 μM, 200 μM, 1 mM, 5 mM and 10 mM free bath Ca 2+ . “C” indicates channel at the closed state; “O” indicates single-level openings. (C) The effect of membrane cholesterol on Ca 2+ -dependence of channel opening. Channel P O was plotted as a function of free Ca 2+ concentration in the bath. P o values are shown for patches either with exogenous cholesterol treatment (black line) or lovastatin treatment (red line). n = 4–7 cells for different data points.
    Figure Legend Snippet: Treatment of cells with cholesterol increases, but with lovastatin decreases, TRPM4 channel activity by regulating its sensitivity to Ca 2+ in mpkCCD c14 Cells. (A) Representative single channel recording from inside-out patches exposed the patch membrane to the bath containing different concentrations of free Ca 2+ . The cells were treated with 30 μg/ml cholesterol for 48 hrs. From top to bottom: 1 μM, 10 μM, 200 μM, 1 mM, and 5 mM free bath Ca 2+ . “C” indicates channel at the closed state; “O” indicates single-level openings. (B) Representative single channel recording from inside-out patches exposed the patch membrane to the bath containing different concentrations of free Ca 2+ . The cells were treated with 5 μM lovastatin for 48 hrs. From top to bottom: 10 μM, 200 μM, 1 mM, 5 mM and 10 mM free bath Ca 2+ . “C” indicates channel at the closed state; “O” indicates single-level openings. (C) The effect of membrane cholesterol on Ca 2+ -dependence of channel opening. Channel P O was plotted as a function of free Ca 2+ concentration in the bath. P o values are shown for patches either with exogenous cholesterol treatment (black line) or lovastatin treatment (red line). n = 4–7 cells for different data points.

    Techniques Used: Activity Assay, Concentration Assay

    Application of MβCD to the cytoplasmic bath decreases TRPM4 channel activity which is reversed by exogenous cholesterol. (A) A representative single channel recording from an inside-out patch shows TRPM4 activity before and after replacement of control cytoplasmic bath solution first with a solution containing 0.5 mM MβCD and then with a solution containing 30 μg/ml cholesterol. “C” indicates channel at the closed state. “O” indicates single-level openings. “I, II, and III” are zoom-ins of the single-channel recording. (B) Summary plots of TRPM4 channel P O under each indicted condition. n = 5 paired experiments, ** P < 0.01, significantly different with control; ## P < 0.01, significantly different with cells treated with 0.5 mM MβCD.
    Figure Legend Snippet: Application of MβCD to the cytoplasmic bath decreases TRPM4 channel activity which is reversed by exogenous cholesterol. (A) A representative single channel recording from an inside-out patch shows TRPM4 activity before and after replacement of control cytoplasmic bath solution first with a solution containing 0.5 mM MβCD and then with a solution containing 30 μg/ml cholesterol. “C” indicates channel at the closed state. “O” indicates single-level openings. “I, II, and III” are zoom-ins of the single-channel recording. (B) Summary plots of TRPM4 channel P O under each indicted condition. n = 5 paired experiments, ** P < 0.01, significantly different with control; ## P < 0.01, significantly different with cells treated with 0.5 mM MβCD.

    Techniques Used: Activity Assay

    Cholesterol stimulates TRPM4 via a PI(4, 5)P 2 -dependent mechanism. (A) A representative single channel recording shows that treatment of mpkCCD c14 cells with 20 nM wortmannin had no effect on cholesterol-induced TRPM4 channel activity. (B) Summary plots of TRPM4 channel P O under each indicated conditions. n = 6 paired experiments, ** P < 0.01, significantly different with cells treated with 20 nm wortmannin. (C) A representative single channel recording shows that depletion of PI(4, 5)P 2 with 20 μM wortmannin abrogated cholesterol-induced TRPM4 channel activity. (D) Summary plots of TRPM4 channel P O under indicated conditions. n = 5 paired experiments. (E) A representative single channel recording shows that application of 20 μM diC8-PI(4,5)P 2 induced TRPM4 channel activity after cholesterol failed to stimulate TRPM4 in the presence of 2 μM PGE2. (F) Summary plots of TRPM4 channel P O under indicated conditions. n = 4 paired experiments. ** P < 0.01, significantly different with cells treated with 2 μM PGE2 or 30 μg/ml cholesterol.
    Figure Legend Snippet: Cholesterol stimulates TRPM4 via a PI(4, 5)P 2 -dependent mechanism. (A) A representative single channel recording shows that treatment of mpkCCD c14 cells with 20 nM wortmannin had no effect on cholesterol-induced TRPM4 channel activity. (B) Summary plots of TRPM4 channel P O under each indicated conditions. n = 6 paired experiments, ** P < 0.01, significantly different with cells treated with 20 nm wortmannin. (C) A representative single channel recording shows that depletion of PI(4, 5)P 2 with 20 μM wortmannin abrogated cholesterol-induced TRPM4 channel activity. (D) Summary plots of TRPM4 channel P O under indicated conditions. n = 5 paired experiments. (E) A representative single channel recording shows that application of 20 μM diC8-PI(4,5)P 2 induced TRPM4 channel activity after cholesterol failed to stimulate TRPM4 in the presence of 2 μM PGE2. (F) Summary plots of TRPM4 channel P O under indicated conditions. n = 4 paired experiments. ** P < 0.01, significantly different with cells treated with 2 μM PGE2 or 30 μg/ml cholesterol.

    Techniques Used: Activity Assay

    PI(4, 5)P 2 stimulates TRPM4 via a physical interaction. (A) A representative single channel recording shows that diC8-PI(4,5)P 2 significantly increased TRPM4 activity. (B) Summary plots of TRPM4 channel P O under each indicated conditions. n = 5 paired experiments. ** P < 0.01, significantly different with control. (C) TRPM4 was detected in dots where most anionic phospholipids including PI(4, 5)P 2 are located on PIP Strips membrane. Data represent three individual experiments showing consistent results.
    Figure Legend Snippet: PI(4, 5)P 2 stimulates TRPM4 via a physical interaction. (A) A representative single channel recording shows that diC8-PI(4,5)P 2 significantly increased TRPM4 activity. (B) Summary plots of TRPM4 channel P O under each indicated conditions. n = 5 paired experiments. ** P < 0.01, significantly different with control. (C) TRPM4 was detected in dots where most anionic phospholipids including PI(4, 5)P 2 are located on PIP Strips membrane. Data represent three individual experiments showing consistent results.

    Techniques Used: Activity Assay

    TRPM4 channels are mainly located in lipid rafts. (A) Representative confocal microscopy images indicate that majority of TRPM4 (green) was co-localized with cholera toxin B (red) in the apical membrane; white rectangular box indicate zoomed-in areas shown in the Zoom-in panels. Data represent five individual experiments showing consistent results (B) Majority of TRPM4 was detected in low-density regions in sucrose gradient experiments; Caveolin-1 was used as a control protein that is known to be located in lipid rafts. Data represent three individual experiments showing consistent results.
    Figure Legend Snippet: TRPM4 channels are mainly located in lipid rafts. (A) Representative confocal microscopy images indicate that majority of TRPM4 (green) was co-localized with cholera toxin B (red) in the apical membrane; white rectangular box indicate zoomed-in areas shown in the Zoom-in panels. Data represent five individual experiments showing consistent results (B) Majority of TRPM4 was detected in low-density regions in sucrose gradient experiments; Caveolin-1 was used as a control protein that is known to be located in lipid rafts. Data represent three individual experiments showing consistent results.

    Techniques Used: Confocal Microscopy

    Treatment with cholesterol or lovastatin does not alter expression levels of TRPM4 in mpkCCDc14 Cells. (A) Representative confocal microscopy images of mpkCCDc14 cells stained with TRPM4 antibody under each indicated conditions. (B) Summary plots of fluorescence intensity of TRPM4. Data are from 24 cells in four sets of separate experiments. (C) Representative Western blots from cell-surface biotinylated and the total proteins of TRPM4 protein. (D) Summary plots of relative expression of TRPM4. Cells were either under control conditions or treated with 30 μg/ml cholesterol alone, 30 μg/ml cholesterol plus 5 μM lovastatin, or 5 μM lovastatin alone for 48 hrs, respectively. n = 5. (E) Representative single channel recording from inside-out patches exposed the patch membrane to the bath containing 5 mM CaCl 2 , followed by a bath solution with 10 mM EGTA and no calcium. Cells were either under control conditions or treated with 30 μg/ml cholesterol, or 5 μM lovastatin for 48 hrs, respectively. (F) Summary plots of the number of active channels in the patches under each indicated condition. n = 5 for control cells, n = 5 for cells treated with cholesterol, n = 4 for cells treated with lovastatin.
    Figure Legend Snippet: Treatment with cholesterol or lovastatin does not alter expression levels of TRPM4 in mpkCCDc14 Cells. (A) Representative confocal microscopy images of mpkCCDc14 cells stained with TRPM4 antibody under each indicated conditions. (B) Summary plots of fluorescence intensity of TRPM4. Data are from 24 cells in four sets of separate experiments. (C) Representative Western blots from cell-surface biotinylated and the total proteins of TRPM4 protein. (D) Summary plots of relative expression of TRPM4. Cells were either under control conditions or treated with 30 μg/ml cholesterol alone, 30 μg/ml cholesterol plus 5 μM lovastatin, or 5 μM lovastatin alone for 48 hrs, respectively. n = 5. (E) Representative single channel recording from inside-out patches exposed the patch membrane to the bath containing 5 mM CaCl 2 , followed by a bath solution with 10 mM EGTA and no calcium. Cells were either under control conditions or treated with 30 μg/ml cholesterol, or 5 μM lovastatin for 48 hrs, respectively. (F) Summary plots of the number of active channels in the patches under each indicated condition. n = 5 for control cells, n = 5 for cells treated with cholesterol, n = 4 for cells treated with lovastatin.

    Techniques Used: Expressing, Confocal Microscopy, Staining, Fluorescence, Western Blot

    rabbit polyclonal anti trpm4 antibody  (Alomone Labs)


    Bioz Verified Symbol Alomone Labs is a verified supplier
    Bioz Manufacturer Symbol Alomone Labs manufactures this product  
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  • 94

    Structured Review

    Alomone Labs rabbit polyclonal anti trpm4 antibody
    Treatment of cells with cholesterol increases, but with lovastatin decreases, <t>TRPM4</t> channel activity by regulating its sensitivity to Ca 2+ in mpkCCD c14 Cells. (A) Representative single channel recording from inside-out patches exposed the patch membrane to the bath containing different concentrations of free Ca 2+ . The cells were treated with 30 μg/ml cholesterol for 48 hrs. From top to bottom: 1 μM, 10 μM, 200 μM, 1 mM, and 5 mM free bath Ca 2+ . “C” indicates channel at the closed state; “O” indicates single-level openings. (B) Representative single channel recording from inside-out patches exposed the patch membrane to the bath containing different concentrations of free Ca 2+ . The cells were treated with 5 μM lovastatin for 48 hrs. From top to bottom: 10 μM, 200 μM, 1 mM, 5 mM and 10 mM free bath Ca 2+ . “C” indicates channel at the closed state; “O” indicates single-level openings. (C) The effect of membrane cholesterol on Ca 2+ -dependence of channel opening. Channel P O was plotted as a function of free Ca 2+ concentration in the bath. P o values are shown for patches either with exogenous cholesterol treatment (black line) or lovastatin treatment (red line). n = 4–7 cells for different data points.
    Rabbit Polyclonal Anti Trpm4 Antibody, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/rabbit polyclonal anti trpm4 antibody/product/Alomone Labs
    Average 94 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    rabbit polyclonal anti trpm4 antibody - by Bioz Stars, 2023-02
    94/100 stars

    Images

    1) Product Images from "Cholesterol Stimulates the Transient Receptor Potential Melastatin 4 Channel in mpkCCD c14 Cells"

    Article Title: Cholesterol Stimulates the Transient Receptor Potential Melastatin 4 Channel in mpkCCD c14 Cells

    Journal: Frontiers in Pharmacology

    doi: 10.3389/fphar.2021.627875

    Treatment of cells with cholesterol increases, but with lovastatin decreases, TRPM4 channel activity by regulating its sensitivity to Ca 2+ in mpkCCD c14 Cells. (A) Representative single channel recording from inside-out patches exposed the patch membrane to the bath containing different concentrations of free Ca 2+ . The cells were treated with 30 μg/ml cholesterol for 48 hrs. From top to bottom: 1 μM, 10 μM, 200 μM, 1 mM, and 5 mM free bath Ca 2+ . “C” indicates channel at the closed state; “O” indicates single-level openings. (B) Representative single channel recording from inside-out patches exposed the patch membrane to the bath containing different concentrations of free Ca 2+ . The cells were treated with 5 μM lovastatin for 48 hrs. From top to bottom: 10 μM, 200 μM, 1 mM, 5 mM and 10 mM free bath Ca 2+ . “C” indicates channel at the closed state; “O” indicates single-level openings. (C) The effect of membrane cholesterol on Ca 2+ -dependence of channel opening. Channel P O was plotted as a function of free Ca 2+ concentration in the bath. P o values are shown for patches either with exogenous cholesterol treatment (black line) or lovastatin treatment (red line). n = 4–7 cells for different data points.
    Figure Legend Snippet: Treatment of cells with cholesterol increases, but with lovastatin decreases, TRPM4 channel activity by regulating its sensitivity to Ca 2+ in mpkCCD c14 Cells. (A) Representative single channel recording from inside-out patches exposed the patch membrane to the bath containing different concentrations of free Ca 2+ . The cells were treated with 30 μg/ml cholesterol for 48 hrs. From top to bottom: 1 μM, 10 μM, 200 μM, 1 mM, and 5 mM free bath Ca 2+ . “C” indicates channel at the closed state; “O” indicates single-level openings. (B) Representative single channel recording from inside-out patches exposed the patch membrane to the bath containing different concentrations of free Ca 2+ . The cells were treated with 5 μM lovastatin for 48 hrs. From top to bottom: 10 μM, 200 μM, 1 mM, 5 mM and 10 mM free bath Ca 2+ . “C” indicates channel at the closed state; “O” indicates single-level openings. (C) The effect of membrane cholesterol on Ca 2+ -dependence of channel opening. Channel P O was plotted as a function of free Ca 2+ concentration in the bath. P o values are shown for patches either with exogenous cholesterol treatment (black line) or lovastatin treatment (red line). n = 4–7 cells for different data points.

    Techniques Used: Activity Assay, Concentration Assay

    Application of MβCD to the cytoplasmic bath decreases TRPM4 channel activity which is reversed by exogenous cholesterol. (A) A representative single channel recording from an inside-out patch shows TRPM4 activity before and after replacement of control cytoplasmic bath solution first with a solution containing 0.5 mM MβCD and then with a solution containing 30 μg/ml cholesterol. “C” indicates channel at the closed state. “O” indicates single-level openings. “I, II, and III” are zoom-ins of the single-channel recording. (B) Summary plots of TRPM4 channel P O under each indicted condition. n = 5 paired experiments, ** P < 0.01, significantly different with control; ## P < 0.01, significantly different with cells treated with 0.5 mM MβCD.
    Figure Legend Snippet: Application of MβCD to the cytoplasmic bath decreases TRPM4 channel activity which is reversed by exogenous cholesterol. (A) A representative single channel recording from an inside-out patch shows TRPM4 activity before and after replacement of control cytoplasmic bath solution first with a solution containing 0.5 mM MβCD and then with a solution containing 30 μg/ml cholesterol. “C” indicates channel at the closed state. “O” indicates single-level openings. “I, II, and III” are zoom-ins of the single-channel recording. (B) Summary plots of TRPM4 channel P O under each indicted condition. n = 5 paired experiments, ** P < 0.01, significantly different with control; ## P < 0.01, significantly different with cells treated with 0.5 mM MβCD.

    Techniques Used: Activity Assay

    Cholesterol stimulates TRPM4 via a PI(4, 5)P 2 -dependent mechanism. (A) A representative single channel recording shows that treatment of mpkCCD c14 cells with 20 nM wortmannin had no effect on cholesterol-induced TRPM4 channel activity. (B) Summary plots of TRPM4 channel P O under each indicated conditions. n = 6 paired experiments, ** P < 0.01, significantly different with cells treated with 20 nm wortmannin. (C) A representative single channel recording shows that depletion of PI(4, 5)P 2 with 20 μM wortmannin abrogated cholesterol-induced TRPM4 channel activity. (D) Summary plots of TRPM4 channel P O under indicated conditions. n = 5 paired experiments. (E) A representative single channel recording shows that application of 20 μM diC8-PI(4,5)P 2 induced TRPM4 channel activity after cholesterol failed to stimulate TRPM4 in the presence of 2 μM PGE2. (F) Summary plots of TRPM4 channel P O under indicated conditions. n = 4 paired experiments. ** P < 0.01, significantly different with cells treated with 2 μM PGE2 or 30 μg/ml cholesterol.
    Figure Legend Snippet: Cholesterol stimulates TRPM4 via a PI(4, 5)P 2 -dependent mechanism. (A) A representative single channel recording shows that treatment of mpkCCD c14 cells with 20 nM wortmannin had no effect on cholesterol-induced TRPM4 channel activity. (B) Summary plots of TRPM4 channel P O under each indicated conditions. n = 6 paired experiments, ** P < 0.01, significantly different with cells treated with 20 nm wortmannin. (C) A representative single channel recording shows that depletion of PI(4, 5)P 2 with 20 μM wortmannin abrogated cholesterol-induced TRPM4 channel activity. (D) Summary plots of TRPM4 channel P O under indicated conditions. n = 5 paired experiments. (E) A representative single channel recording shows that application of 20 μM diC8-PI(4,5)P 2 induced TRPM4 channel activity after cholesterol failed to stimulate TRPM4 in the presence of 2 μM PGE2. (F) Summary plots of TRPM4 channel P O under indicated conditions. n = 4 paired experiments. ** P < 0.01, significantly different with cells treated with 2 μM PGE2 or 30 μg/ml cholesterol.

    Techniques Used: Activity Assay

    PI(4, 5)P 2 stimulates TRPM4 via a physical interaction. (A) A representative single channel recording shows that diC8-PI(4,5)P 2 significantly increased TRPM4 activity. (B) Summary plots of TRPM4 channel P O under each indicated conditions. n = 5 paired experiments. ** P < 0.01, significantly different with control. (C) TRPM4 was detected in dots where most anionic phospholipids including PI(4, 5)P 2 are located on PIP Strips membrane. Data represent three individual experiments showing consistent results.
    Figure Legend Snippet: PI(4, 5)P 2 stimulates TRPM4 via a physical interaction. (A) A representative single channel recording shows that diC8-PI(4,5)P 2 significantly increased TRPM4 activity. (B) Summary plots of TRPM4 channel P O under each indicated conditions. n = 5 paired experiments. ** P < 0.01, significantly different with control. (C) TRPM4 was detected in dots where most anionic phospholipids including PI(4, 5)P 2 are located on PIP Strips membrane. Data represent three individual experiments showing consistent results.

    Techniques Used: Activity Assay

    TRPM4 channels are mainly located in lipid rafts. (A) Representative confocal microscopy images indicate that majority of TRPM4 (green) was co-localized with cholera toxin B (red) in the apical membrane; white rectangular box indicate zoomed-in areas shown in the Zoom-in panels. Data represent five individual experiments showing consistent results (B) Majority of TRPM4 was detected in low-density regions in sucrose gradient experiments; Caveolin-1 was used as a control protein that is known to be located in lipid rafts. Data represent three individual experiments showing consistent results.
    Figure Legend Snippet: TRPM4 channels are mainly located in lipid rafts. (A) Representative confocal microscopy images indicate that majority of TRPM4 (green) was co-localized with cholera toxin B (red) in the apical membrane; white rectangular box indicate zoomed-in areas shown in the Zoom-in panels. Data represent five individual experiments showing consistent results (B) Majority of TRPM4 was detected in low-density regions in sucrose gradient experiments; Caveolin-1 was used as a control protein that is known to be located in lipid rafts. Data represent three individual experiments showing consistent results.

    Techniques Used: Confocal Microscopy

    Treatment with cholesterol or lovastatin does not alter expression levels of TRPM4 in mpkCCDc14 Cells. (A) Representative confocal microscopy images of mpkCCDc14 cells stained with TRPM4 antibody under each indicated conditions. (B) Summary plots of fluorescence intensity of TRPM4. Data are from 24 cells in four sets of separate experiments. (C) Representative Western blots from cell-surface biotinylated and the total proteins of TRPM4 protein. (D) Summary plots of relative expression of TRPM4. Cells were either under control conditions or treated with 30 μg/ml cholesterol alone, 30 μg/ml cholesterol plus 5 μM lovastatin, or 5 μM lovastatin alone for 48 hrs, respectively. n = 5. (E) Representative single channel recording from inside-out patches exposed the patch membrane to the bath containing 5 mM CaCl 2 , followed by a bath solution with 10 mM EGTA and no calcium. Cells were either under control conditions or treated with 30 μg/ml cholesterol, or 5 μM lovastatin for 48 hrs, respectively. (F) Summary plots of the number of active channels in the patches under each indicated condition. n = 5 for control cells, n = 5 for cells treated with cholesterol, n = 4 for cells treated with lovastatin.
    Figure Legend Snippet: Treatment with cholesterol or lovastatin does not alter expression levels of TRPM4 in mpkCCDc14 Cells. (A) Representative confocal microscopy images of mpkCCDc14 cells stained with TRPM4 antibody under each indicated conditions. (B) Summary plots of fluorescence intensity of TRPM4. Data are from 24 cells in four sets of separate experiments. (C) Representative Western blots from cell-surface biotinylated and the total proteins of TRPM4 protein. (D) Summary plots of relative expression of TRPM4. Cells were either under control conditions or treated with 30 μg/ml cholesterol alone, 30 μg/ml cholesterol plus 5 μM lovastatin, or 5 μM lovastatin alone for 48 hrs, respectively. n = 5. (E) Representative single channel recording from inside-out patches exposed the patch membrane to the bath containing 5 mM CaCl 2 , followed by a bath solution with 10 mM EGTA and no calcium. Cells were either under control conditions or treated with 30 μg/ml cholesterol, or 5 μM lovastatin for 48 hrs, respectively. (F) Summary plots of the number of active channels in the patches under each indicated condition. n = 5 for control cells, n = 5 for cells treated with cholesterol, n = 4 for cells treated with lovastatin.

    Techniques Used: Expressing, Confocal Microscopy, Staining, Fluorescence, Western Blot

    rabbit polyclonal antibodies  (Alomone Labs)


    Bioz Verified Symbol Alomone Labs is a verified supplier
    Bioz Manufacturer Symbol Alomone Labs manufactures this product  
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  • 94

    Structured Review

    Alomone Labs rabbit polyclonal antibodies
    Rabbit Polyclonal Antibodies, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/rabbit polyclonal antibodies/product/Alomone Labs
    Average 94 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    rabbit polyclonal antibodies - by Bioz Stars, 2023-02
    94/100 stars

    Images

    rabbit polyclonal anti trpm4 antibody  (Alomone Labs)


    Bioz Verified Symbol Alomone Labs is a verified supplier
    Bioz Manufacturer Symbol Alomone Labs manufactures this product  
  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 94

    Structured Review

    Alomone Labs rabbit polyclonal anti trpm4 antibody
    Treatment of cells with cholesterol increases, but with lovastatin decreases, <t>TRPM4</t> channel activity by regulating its sensitivity to Ca 2+ in mpkCCD c14 Cells. (A) Representative single channel recording from inside-out patches exposed the patch membrane to the bath containing different concentrations of free Ca 2+ . The cells were treated with 30 μg/ml cholesterol for 48 hrs. From top to bottom: 1 μM, 10 μM, 200 μM, 1 mM, and 5 mM free bath Ca 2+ . “C” indicates channel at the closed state; “O” indicates single-level openings. (B) Representative single channel recording from inside-out patches exposed the patch membrane to the bath containing different concentrations of free Ca 2+ . The cells were treated with 5 μM lovastatin for 48 hrs. From top to bottom: 10 μM, 200 μM, 1 mM, 5 mM and 10 mM free bath Ca 2+ . “C” indicates channel at the closed state; “O” indicates single-level openings. (C) The effect of membrane cholesterol on Ca 2+ -dependence of channel opening. Channel P O was plotted as a function of free Ca 2+ concentration in the bath. P o values are shown for patches either with exogenous cholesterol treatment (black line) or lovastatin treatment (red line). n = 4–7 cells for different data points.
    Rabbit Polyclonal Anti Trpm4 Antibody, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/rabbit polyclonal anti trpm4 antibody/product/Alomone Labs
    Average 94 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    rabbit polyclonal anti trpm4 antibody - by Bioz Stars, 2023-02
    94/100 stars

    Images

    1) Product Images from "Cholesterol Stimulates the Transient Receptor Potential Melastatin 4 Channel in mpkCCD c14 Cells"

    Article Title: Cholesterol Stimulates the Transient Receptor Potential Melastatin 4 Channel in mpkCCD c14 Cells

    Journal: Frontiers in Pharmacology

    doi: 10.3389/fphar.2021.627875

    Treatment of cells with cholesterol increases, but with lovastatin decreases, TRPM4 channel activity by regulating its sensitivity to Ca 2+ in mpkCCD c14 Cells. (A) Representative single channel recording from inside-out patches exposed the patch membrane to the bath containing different concentrations of free Ca 2+ . The cells were treated with 30 μg/ml cholesterol for 48 hrs. From top to bottom: 1 μM, 10 μM, 200 μM, 1 mM, and 5 mM free bath Ca 2+ . “C” indicates channel at the closed state; “O” indicates single-level openings. (B) Representative single channel recording from inside-out patches exposed the patch membrane to the bath containing different concentrations of free Ca 2+ . The cells were treated with 5 μM lovastatin for 48 hrs. From top to bottom: 10 μM, 200 μM, 1 mM, 5 mM and 10 mM free bath Ca 2+ . “C” indicates channel at the closed state; “O” indicates single-level openings. (C) The effect of membrane cholesterol on Ca 2+ -dependence of channel opening. Channel P O was plotted as a function of free Ca 2+ concentration in the bath. P o values are shown for patches either with exogenous cholesterol treatment (black line) or lovastatin treatment (red line). n = 4–7 cells for different data points.
    Figure Legend Snippet: Treatment of cells with cholesterol increases, but with lovastatin decreases, TRPM4 channel activity by regulating its sensitivity to Ca 2+ in mpkCCD c14 Cells. (A) Representative single channel recording from inside-out patches exposed the patch membrane to the bath containing different concentrations of free Ca 2+ . The cells were treated with 30 μg/ml cholesterol for 48 hrs. From top to bottom: 1 μM, 10 μM, 200 μM, 1 mM, and 5 mM free bath Ca 2+ . “C” indicates channel at the closed state; “O” indicates single-level openings. (B) Representative single channel recording from inside-out patches exposed the patch membrane to the bath containing different concentrations of free Ca 2+ . The cells were treated with 5 μM lovastatin for 48 hrs. From top to bottom: 10 μM, 200 μM, 1 mM, 5 mM and 10 mM free bath Ca 2+ . “C” indicates channel at the closed state; “O” indicates single-level openings. (C) The effect of membrane cholesterol on Ca 2+ -dependence of channel opening. Channel P O was plotted as a function of free Ca 2+ concentration in the bath. P o values are shown for patches either with exogenous cholesterol treatment (black line) or lovastatin treatment (red line). n = 4–7 cells for different data points.

    Techniques Used: Activity Assay, Concentration Assay

    Application of MβCD to the cytoplasmic bath decreases TRPM4 channel activity which is reversed by exogenous cholesterol. (A) A representative single channel recording from an inside-out patch shows TRPM4 activity before and after replacement of control cytoplasmic bath solution first with a solution containing 0.5 mM MβCD and then with a solution containing 30 μg/ml cholesterol. “C” indicates channel at the closed state. “O” indicates single-level openings. “I, II, and III” are zoom-ins of the single-channel recording. (B) Summary plots of TRPM4 channel P O under each indicted condition. n = 5 paired experiments, ** P < 0.01, significantly different with control; ## P < 0.01, significantly different with cells treated with 0.5 mM MβCD.
    Figure Legend Snippet: Application of MβCD to the cytoplasmic bath decreases TRPM4 channel activity which is reversed by exogenous cholesterol. (A) A representative single channel recording from an inside-out patch shows TRPM4 activity before and after replacement of control cytoplasmic bath solution first with a solution containing 0.5 mM MβCD and then with a solution containing 30 μg/ml cholesterol. “C” indicates channel at the closed state. “O” indicates single-level openings. “I, II, and III” are zoom-ins of the single-channel recording. (B) Summary plots of TRPM4 channel P O under each indicted condition. n = 5 paired experiments, ** P < 0.01, significantly different with control; ## P < 0.01, significantly different with cells treated with 0.5 mM MβCD.

    Techniques Used: Activity Assay

    Cholesterol stimulates TRPM4 via a PI(4, 5)P 2 -dependent mechanism. (A) A representative single channel recording shows that treatment of mpkCCD c14 cells with 20 nM wortmannin had no effect on cholesterol-induced TRPM4 channel activity. (B) Summary plots of TRPM4 channel P O under each indicated conditions. n = 6 paired experiments, ** P < 0.01, significantly different with cells treated with 20 nm wortmannin. (C) A representative single channel recording shows that depletion of PI(4, 5)P 2 with 20 μM wortmannin abrogated cholesterol-induced TRPM4 channel activity. (D) Summary plots of TRPM4 channel P O under indicated conditions. n = 5 paired experiments. (E) A representative single channel recording shows that application of 20 μM diC8-PI(4,5)P 2 induced TRPM4 channel activity after cholesterol failed to stimulate TRPM4 in the presence of 2 μM PGE2. (F) Summary plots of TRPM4 channel P O under indicated conditions. n = 4 paired experiments. ** P < 0.01, significantly different with cells treated with 2 μM PGE2 or 30 μg/ml cholesterol.
    Figure Legend Snippet: Cholesterol stimulates TRPM4 via a PI(4, 5)P 2 -dependent mechanism. (A) A representative single channel recording shows that treatment of mpkCCD c14 cells with 20 nM wortmannin had no effect on cholesterol-induced TRPM4 channel activity. (B) Summary plots of TRPM4 channel P O under each indicated conditions. n = 6 paired experiments, ** P < 0.01, significantly different with cells treated with 20 nm wortmannin. (C) A representative single channel recording shows that depletion of PI(4, 5)P 2 with 20 μM wortmannin abrogated cholesterol-induced TRPM4 channel activity. (D) Summary plots of TRPM4 channel P O under indicated conditions. n = 5 paired experiments. (E) A representative single channel recording shows that application of 20 μM diC8-PI(4,5)P 2 induced TRPM4 channel activity after cholesterol failed to stimulate TRPM4 in the presence of 2 μM PGE2. (F) Summary plots of TRPM4 channel P O under indicated conditions. n = 4 paired experiments. ** P < 0.01, significantly different with cells treated with 2 μM PGE2 or 30 μg/ml cholesterol.

    Techniques Used: Activity Assay

    PI(4, 5)P 2 stimulates TRPM4 via a physical interaction. (A) A representative single channel recording shows that diC8-PI(4,5)P 2 significantly increased TRPM4 activity. (B) Summary plots of TRPM4 channel P O under each indicated conditions. n = 5 paired experiments. ** P < 0.01, significantly different with control. (C) TRPM4 was detected in dots where most anionic phospholipids including PI(4, 5)P 2 are located on PIP Strips membrane. Data represent three individual experiments showing consistent results.
    Figure Legend Snippet: PI(4, 5)P 2 stimulates TRPM4 via a physical interaction. (A) A representative single channel recording shows that diC8-PI(4,5)P 2 significantly increased TRPM4 activity. (B) Summary plots of TRPM4 channel P O under each indicated conditions. n = 5 paired experiments. ** P < 0.01, significantly different with control. (C) TRPM4 was detected in dots where most anionic phospholipids including PI(4, 5)P 2 are located on PIP Strips membrane. Data represent three individual experiments showing consistent results.

    Techniques Used: Activity Assay

    TRPM4 channels are mainly located in lipid rafts. (A) Representative confocal microscopy images indicate that majority of TRPM4 (green) was co-localized with cholera toxin B (red) in the apical membrane; white rectangular box indicate zoomed-in areas shown in the Zoom-in panels. Data represent five individual experiments showing consistent results (B) Majority of TRPM4 was detected in low-density regions in sucrose gradient experiments; Caveolin-1 was used as a control protein that is known to be located in lipid rafts. Data represent three individual experiments showing consistent results.
    Figure Legend Snippet: TRPM4 channels are mainly located in lipid rafts. (A) Representative confocal microscopy images indicate that majority of TRPM4 (green) was co-localized with cholera toxin B (red) in the apical membrane; white rectangular box indicate zoomed-in areas shown in the Zoom-in panels. Data represent five individual experiments showing consistent results (B) Majority of TRPM4 was detected in low-density regions in sucrose gradient experiments; Caveolin-1 was used as a control protein that is known to be located in lipid rafts. Data represent three individual experiments showing consistent results.

    Techniques Used: Confocal Microscopy

    Treatment with cholesterol or lovastatin does not alter expression levels of TRPM4 in mpkCCDc14 Cells. (A) Representative confocal microscopy images of mpkCCDc14 cells stained with TRPM4 antibody under each indicated conditions. (B) Summary plots of fluorescence intensity of TRPM4. Data are from 24 cells in four sets of separate experiments. (C) Representative Western blots from cell-surface biotinylated and the total proteins of TRPM4 protein. (D) Summary plots of relative expression of TRPM4. Cells were either under control conditions or treated with 30 μg/ml cholesterol alone, 30 μg/ml cholesterol plus 5 μM lovastatin, or 5 μM lovastatin alone for 48 hrs, respectively. n = 5. (E) Representative single channel recording from inside-out patches exposed the patch membrane to the bath containing 5 mM CaCl 2 , followed by a bath solution with 10 mM EGTA and no calcium. Cells were either under control conditions or treated with 30 μg/ml cholesterol, or 5 μM lovastatin for 48 hrs, respectively. (F) Summary plots of the number of active channels in the patches under each indicated condition. n = 5 for control cells, n = 5 for cells treated with cholesterol, n = 4 for cells treated with lovastatin.
    Figure Legend Snippet: Treatment with cholesterol or lovastatin does not alter expression levels of TRPM4 in mpkCCDc14 Cells. (A) Representative confocal microscopy images of mpkCCDc14 cells stained with TRPM4 antibody under each indicated conditions. (B) Summary plots of fluorescence intensity of TRPM4. Data are from 24 cells in four sets of separate experiments. (C) Representative Western blots from cell-surface biotinylated and the total proteins of TRPM4 protein. (D) Summary plots of relative expression of TRPM4. Cells were either under control conditions or treated with 30 μg/ml cholesterol alone, 30 μg/ml cholesterol plus 5 μM lovastatin, or 5 μM lovastatin alone for 48 hrs, respectively. n = 5. (E) Representative single channel recording from inside-out patches exposed the patch membrane to the bath containing 5 mM CaCl 2 , followed by a bath solution with 10 mM EGTA and no calcium. Cells were either under control conditions or treated with 30 μg/ml cholesterol, or 5 μM lovastatin for 48 hrs, respectively. (F) Summary plots of the number of active channels in the patches under each indicated condition. n = 5 for control cells, n = 5 for cells treated with cholesterol, n = 4 for cells treated with lovastatin.

    Techniques Used: Expressing, Confocal Microscopy, Staining, Fluorescence, Western Blot

    rabbit polyclonal anti trpm4  (Alomone Labs)


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    Alomone Labs rabbit polyclonal anti trpm4
    Rabbit Polyclonal Anti Trpm4, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/rabbit polyclonal anti trpm4/product/Alomone Labs
    Average 94 stars, based on 1 article reviews
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    rabbit polyclonal antibody  (Alomone Labs)


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    Alomone Labs rabbit polyclonal antibody
    Rabbit Polyclonal Antibody, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    rabbit polyclonal antibody to anti trpm4  (Alomone Labs)


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    Alomone Labs rabbit polyclonal antibody to anti trpm4
    Rabbit Polyclonal Antibody To Anti Trpm4, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/rabbit polyclonal antibody to anti trpm4/product/Alomone Labs
    Average 94 stars, based on 1 article reviews
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    rabbit polyclonal anti trpm4 antibody  (Alomone Labs)


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    Alomone Labs rabbit polyclonal anti trpm4 antibody
    Transient receptor potential channel, the melastatin subfamily 4 <t>(TRPM4)</t> blockade and knockdown reduce channel activity. A: representative single-channel current recorded from an inside-out patch before and after replacing the bath solution with a similar solution but containing 100 μM 9-phenanthrol (a specific TRPM4 blocker). B: summary plots of mean PO. C: representative single-channel currents recorded from either a control cell (top trace) or a TRPM4 knockdown cell (bottom trace). D: summary plots of percentage of patches containing channels in control and TRPM4 knockdown cells. E: Western blot of TRPM4 from cells under control conditions or transfected with either control RNA or TRPM4 short hairpin (sh) RNA (left). Data represent 3 separate experiments, showing that TRPM4 shRNA significantly reduced TRPM4 expression (right).
    Rabbit Polyclonal Anti Trpm4 Antibody, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/rabbit polyclonal anti trpm4 antibody/product/Alomone Labs
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    rabbit polyclonal anti trpm4 antibody - by Bioz Stars, 2023-02
    86/100 stars

    Images

    1) Product Images from "Hydrogen peroxide suppresses TRPM4 trafficking to the apical membrane in mouse cortical collecting duct principal cells"

    Article Title: Hydrogen peroxide suppresses TRPM4 trafficking to the apical membrane in mouse cortical collecting duct principal cells

    Journal: American Journal of Physiology - Renal Physiology

    doi: 10.1152/ajprenal.00439.2016

    Transient receptor potential channel, the melastatin subfamily 4 (TRPM4) blockade and knockdown reduce channel activity. A: representative single-channel current recorded from an inside-out patch before and after replacing the bath solution with a similar solution but containing 100 μM 9-phenanthrol (a specific TRPM4 blocker). B: summary plots of mean PO. C: representative single-channel currents recorded from either a control cell (top trace) or a TRPM4 knockdown cell (bottom trace). D: summary plots of percentage of patches containing channels in control and TRPM4 knockdown cells. E: Western blot of TRPM4 from cells under control conditions or transfected with either control RNA or TRPM4 short hairpin (sh) RNA (left). Data represent 3 separate experiments, showing that TRPM4 shRNA significantly reduced TRPM4 expression (right).
    Figure Legend Snippet: Transient receptor potential channel, the melastatin subfamily 4 (TRPM4) blockade and knockdown reduce channel activity. A: representative single-channel current recorded from an inside-out patch before and after replacing the bath solution with a similar solution but containing 100 μM 9-phenanthrol (a specific TRPM4 blocker). B: summary plots of mean PO. C: representative single-channel currents recorded from either a control cell (top trace) or a TRPM4 knockdown cell (bottom trace). D: summary plots of percentage of patches containing channels in control and TRPM4 knockdown cells. E: Western blot of TRPM4 from cells under control conditions or transfected with either control RNA or TRPM4 short hairpin (sh) RNA (left). Data represent 3 separate experiments, showing that TRPM4 shRNA significantly reduced TRPM4 expression (right).

    Techniques Used: Activity Assay, Western Blot, Transfection, shRNA, Expressing

    H2O2 does not affect TRPM4 channel activity in mpkCCDc14 cells. A: representative single-channel currents recorded from 3 inside-out patches. Addition of 100 or 500 μM H2O2 to the bath did not alter channel activity, no matter whether the channel was strongly activated by 10−3 M Ca2+ (top and middle traces) or slightly activated by 10−6 M Ca2+ (bottom trace). B: summary plots of PO calculated from recordings before and after application of H2O2 under each condition.
    Figure Legend Snippet: H2O2 does not affect TRPM4 channel activity in mpkCCDc14 cells. A: representative single-channel currents recorded from 3 inside-out patches. Addition of 100 or 500 μM H2O2 to the bath did not alter channel activity, no matter whether the channel was strongly activated by 10−3 M Ca2+ (top and middle traces) or slightly activated by 10−6 M Ca2+ (bottom trace). B: summary plots of PO calculated from recordings before and after application of H2O2 under each condition.

    Techniques Used: Activity Assay

    H2O2 reduces apical TRPM4 density in mpkCCDc14 cells. A: representative confocal microscopy images taken from either control mpkCCDc14 cells (top) or the cells treated with 100 μM H2O2 for 24 h (bottom). TRPM4 levels in mpkCCDc14 cell monolayer were examined through immunostaining the cells with its antibody (shown in red). Optical sections were set at or near the apical membrane according to where the tight junction protein ZO-1 was localized (shown in green). Scale bar = 20 μm. B: representative Western blots of either total or biotinylated TRPM4 protein in control cells or the cells treated with 100 μM H2O2 for 24 h; the data represent 5 experiments, showing that biotinylated (apical) TRPM4 was significantly reduced by H2O2. C: representative TRPM4 single-channel currents recorded from control cells or the cells treated with 100 μM H2O2 for 24 h. D: summary plots of patches containing TRPM4 channels under control condition or after treatment of the cells with H2O2.
    Figure Legend Snippet: H2O2 reduces apical TRPM4 density in mpkCCDc14 cells. A: representative confocal microscopy images taken from either control mpkCCDc14 cells (top) or the cells treated with 100 μM H2O2 for 24 h (bottom). TRPM4 levels in mpkCCDc14 cell monolayer were examined through immunostaining the cells with its antibody (shown in red). Optical sections were set at or near the apical membrane according to where the tight junction protein ZO-1 was localized (shown in green). Scale bar = 20 μm. B: representative Western blots of either total or biotinylated TRPM4 protein in control cells or the cells treated with 100 μM H2O2 for 24 h; the data represent 5 experiments, showing that biotinylated (apical) TRPM4 was significantly reduced by H2O2. C: representative TRPM4 single-channel currents recorded from control cells or the cells treated with 100 μM H2O2 for 24 h. D: summary plots of patches containing TRPM4 channels under control condition or after treatment of the cells with H2O2.

    Techniques Used: Confocal Microscopy, Immunostaining, Western Blot

    rabbit polyclonal anti trpm4 antibody  (Alomone Labs)


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    Alomone Labs rabbit polyclonal anti trpm4 antibody
    Transient receptor potential channel, the melastatin subfamily 4 <t>(TRPM4)</t> blockade and knockdown reduce channel activity. A: representative single-channel current recorded from an inside-out patch before and after replacing the bath solution with a similar solution but containing 100 μM 9-phenanthrol (a specific TRPM4 blocker). B: summary plots of mean PO. C: representative single-channel currents recorded from either a control cell (top trace) or a TRPM4 knockdown cell (bottom trace). D: summary plots of percentage of patches containing channels in control and TRPM4 knockdown cells. E: Western blot of TRPM4 from cells under control conditions or transfected with either control RNA or TRPM4 short hairpin (sh) RNA (left). Data represent 3 separate experiments, showing that TRPM4 shRNA significantly reduced TRPM4 expression (right).
    Rabbit Polyclonal Anti Trpm4 Antibody, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/rabbit polyclonal anti trpm4 antibody/product/Alomone Labs
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    rabbit polyclonal anti trpm4 antibody - by Bioz Stars, 2023-02
    86/100 stars

    Images

    1) Product Images from "Hydrogen peroxide suppresses TRPM4 trafficking to the apical membrane in mouse cortical collecting duct principal cells"

    Article Title: Hydrogen peroxide suppresses TRPM4 trafficking to the apical membrane in mouse cortical collecting duct principal cells

    Journal: American Journal of Physiology - Renal Physiology

    doi: 10.1152/ajprenal.00439.2016

    Transient receptor potential channel, the melastatin subfamily 4 (TRPM4) blockade and knockdown reduce channel activity. A: representative single-channel current recorded from an inside-out patch before and after replacing the bath solution with a similar solution but containing 100 μM 9-phenanthrol (a specific TRPM4 blocker). B: summary plots of mean PO. C: representative single-channel currents recorded from either a control cell (top trace) or a TRPM4 knockdown cell (bottom trace). D: summary plots of percentage of patches containing channels in control and TRPM4 knockdown cells. E: Western blot of TRPM4 from cells under control conditions or transfected with either control RNA or TRPM4 short hairpin (sh) RNA (left). Data represent 3 separate experiments, showing that TRPM4 shRNA significantly reduced TRPM4 expression (right).
    Figure Legend Snippet: Transient receptor potential channel, the melastatin subfamily 4 (TRPM4) blockade and knockdown reduce channel activity. A: representative single-channel current recorded from an inside-out patch before and after replacing the bath solution with a similar solution but containing 100 μM 9-phenanthrol (a specific TRPM4 blocker). B: summary plots of mean PO. C: representative single-channel currents recorded from either a control cell (top trace) or a TRPM4 knockdown cell (bottom trace). D: summary plots of percentage of patches containing channels in control and TRPM4 knockdown cells. E: Western blot of TRPM4 from cells under control conditions or transfected with either control RNA or TRPM4 short hairpin (sh) RNA (left). Data represent 3 separate experiments, showing that TRPM4 shRNA significantly reduced TRPM4 expression (right).

    Techniques Used: Activity Assay, Western Blot, Transfection, shRNA, Expressing

    H2O2 does not affect TRPM4 channel activity in mpkCCDc14 cells. A: representative single-channel currents recorded from 3 inside-out patches. Addition of 100 or 500 μM H2O2 to the bath did not alter channel activity, no matter whether the channel was strongly activated by 10−3 M Ca2+ (top and middle traces) or slightly activated by 10−6 M Ca2+ (bottom trace). B: summary plots of PO calculated from recordings before and after application of H2O2 under each condition.
    Figure Legend Snippet: H2O2 does not affect TRPM4 channel activity in mpkCCDc14 cells. A: representative single-channel currents recorded from 3 inside-out patches. Addition of 100 or 500 μM H2O2 to the bath did not alter channel activity, no matter whether the channel was strongly activated by 10−3 M Ca2+ (top and middle traces) or slightly activated by 10−6 M Ca2+ (bottom trace). B: summary plots of PO calculated from recordings before and after application of H2O2 under each condition.

    Techniques Used: Activity Assay

    H2O2 reduces apical TRPM4 density in mpkCCDc14 cells. A: representative confocal microscopy images taken from either control mpkCCDc14 cells (top) or the cells treated with 100 μM H2O2 for 24 h (bottom). TRPM4 levels in mpkCCDc14 cell monolayer were examined through immunostaining the cells with its antibody (shown in red). Optical sections were set at or near the apical membrane according to where the tight junction protein ZO-1 was localized (shown in green). Scale bar = 20 μm. B: representative Western blots of either total or biotinylated TRPM4 protein in control cells or the cells treated with 100 μM H2O2 for 24 h; the data represent 5 experiments, showing that biotinylated (apical) TRPM4 was significantly reduced by H2O2. C: representative TRPM4 single-channel currents recorded from control cells or the cells treated with 100 μM H2O2 for 24 h. D: summary plots of patches containing TRPM4 channels under control condition or after treatment of the cells with H2O2.
    Figure Legend Snippet: H2O2 reduces apical TRPM4 density in mpkCCDc14 cells. A: representative confocal microscopy images taken from either control mpkCCDc14 cells (top) or the cells treated with 100 μM H2O2 for 24 h (bottom). TRPM4 levels in mpkCCDc14 cell monolayer were examined through immunostaining the cells with its antibody (shown in red). Optical sections were set at or near the apical membrane according to where the tight junction protein ZO-1 was localized (shown in green). Scale bar = 20 μm. B: representative Western blots of either total or biotinylated TRPM4 protein in control cells or the cells treated with 100 μM H2O2 for 24 h; the data represent 5 experiments, showing that biotinylated (apical) TRPM4 was significantly reduced by H2O2. C: representative TRPM4 single-channel currents recorded from control cells or the cells treated with 100 μM H2O2 for 24 h. D: summary plots of patches containing TRPM4 channels under control condition or after treatment of the cells with H2O2.

    Techniques Used: Confocal Microscopy, Immunostaining, Western Blot

    rabbit polyclonal anti trpm4 antibody  (Alomone Labs)


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    Alomone Labs rabbit polyclonal anti trpm4 antibody
    Transient receptor potential channel, the melastatin subfamily 4 <t>(TRPM4)</t> blockade and knockdown reduce channel activity. A: representative single-channel current recorded from an inside-out patch before and after replacing the bath solution with a similar solution but containing 100 μM 9-phenanthrol (a specific TRPM4 blocker). B: summary plots of mean PO. C: representative single-channel currents recorded from either a control cell (top trace) or a TRPM4 knockdown cell (bottom trace). D: summary plots of percentage of patches containing channels in control and TRPM4 knockdown cells. E: Western blot of TRPM4 from cells under control conditions or transfected with either control RNA or TRPM4 short hairpin (sh) RNA (left). Data represent 3 separate experiments, showing that TRPM4 shRNA significantly reduced TRPM4 expression (right).
    Rabbit Polyclonal Anti Trpm4 Antibody, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/rabbit polyclonal anti trpm4 antibody/product/Alomone Labs
    Average 94 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    rabbit polyclonal anti trpm4 antibody - by Bioz Stars, 2023-02
    94/100 stars

    Images

    1) Product Images from "Hydrogen peroxide suppresses TRPM4 trafficking to the apical membrane in mouse cortical collecting duct principal cells"

    Article Title: Hydrogen peroxide suppresses TRPM4 trafficking to the apical membrane in mouse cortical collecting duct principal cells

    Journal: American Journal of Physiology - Renal Physiology

    doi: 10.1152/ajprenal.00439.2016

    Transient receptor potential channel, the melastatin subfamily 4 (TRPM4) blockade and knockdown reduce channel activity. A: representative single-channel current recorded from an inside-out patch before and after replacing the bath solution with a similar solution but containing 100 μM 9-phenanthrol (a specific TRPM4 blocker). B: summary plots of mean PO. C: representative single-channel currents recorded from either a control cell (top trace) or a TRPM4 knockdown cell (bottom trace). D: summary plots of percentage of patches containing channels in control and TRPM4 knockdown cells. E: Western blot of TRPM4 from cells under control conditions or transfected with either control RNA or TRPM4 short hairpin (sh) RNA (left). Data represent 3 separate experiments, showing that TRPM4 shRNA significantly reduced TRPM4 expression (right).
    Figure Legend Snippet: Transient receptor potential channel, the melastatin subfamily 4 (TRPM4) blockade and knockdown reduce channel activity. A: representative single-channel current recorded from an inside-out patch before and after replacing the bath solution with a similar solution but containing 100 μM 9-phenanthrol (a specific TRPM4 blocker). B: summary plots of mean PO. C: representative single-channel currents recorded from either a control cell (top trace) or a TRPM4 knockdown cell (bottom trace). D: summary plots of percentage of patches containing channels in control and TRPM4 knockdown cells. E: Western blot of TRPM4 from cells under control conditions or transfected with either control RNA or TRPM4 short hairpin (sh) RNA (left). Data represent 3 separate experiments, showing that TRPM4 shRNA significantly reduced TRPM4 expression (right).

    Techniques Used: Activity Assay, Western Blot, Transfection, shRNA, Expressing

    H2O2 does not affect TRPM4 channel activity in mpkCCDc14 cells. A: representative single-channel currents recorded from 3 inside-out patches. Addition of 100 or 500 μM H2O2 to the bath did not alter channel activity, no matter whether the channel was strongly activated by 10−3 M Ca2+ (top and middle traces) or slightly activated by 10−6 M Ca2+ (bottom trace). B: summary plots of PO calculated from recordings before and after application of H2O2 under each condition.
    Figure Legend Snippet: H2O2 does not affect TRPM4 channel activity in mpkCCDc14 cells. A: representative single-channel currents recorded from 3 inside-out patches. Addition of 100 or 500 μM H2O2 to the bath did not alter channel activity, no matter whether the channel was strongly activated by 10−3 M Ca2+ (top and middle traces) or slightly activated by 10−6 M Ca2+ (bottom trace). B: summary plots of PO calculated from recordings before and after application of H2O2 under each condition.

    Techniques Used: Activity Assay

    H2O2 reduces apical TRPM4 density in mpkCCDc14 cells. A: representative confocal microscopy images taken from either control mpkCCDc14 cells (top) or the cells treated with 100 μM H2O2 for 24 h (bottom). TRPM4 levels in mpkCCDc14 cell monolayer were examined through immunostaining the cells with its antibody (shown in red). Optical sections were set at or near the apical membrane according to where the tight junction protein ZO-1 was localized (shown in green). Scale bar = 20 μm. B: representative Western blots of either total or biotinylated TRPM4 protein in control cells or the cells treated with 100 μM H2O2 for 24 h; the data represent 5 experiments, showing that biotinylated (apical) TRPM4 was significantly reduced by H2O2. C: representative TRPM4 single-channel currents recorded from control cells or the cells treated with 100 μM H2O2 for 24 h. D: summary plots of patches containing TRPM4 channels under control condition or after treatment of the cells with H2O2.
    Figure Legend Snippet: H2O2 reduces apical TRPM4 density in mpkCCDc14 cells. A: representative confocal microscopy images taken from either control mpkCCDc14 cells (top) or the cells treated with 100 μM H2O2 for 24 h (bottom). TRPM4 levels in mpkCCDc14 cell monolayer were examined through immunostaining the cells with its antibody (shown in red). Optical sections were set at or near the apical membrane according to where the tight junction protein ZO-1 was localized (shown in green). Scale bar = 20 μm. B: representative Western blots of either total or biotinylated TRPM4 protein in control cells or the cells treated with 100 μM H2O2 for 24 h; the data represent 5 experiments, showing that biotinylated (apical) TRPM4 was significantly reduced by H2O2. C: representative TRPM4 single-channel currents recorded from control cells or the cells treated with 100 μM H2O2 for 24 h. D: summary plots of patches containing TRPM4 channels under control condition or after treatment of the cells with H2O2.

    Techniques Used: Confocal Microscopy, Immunostaining, Western Blot

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  • 94
    Alomone Labs rabbit polyclonal anti trpm4 antibody
    Treatment of cells with cholesterol increases, but with lovastatin decreases, <t>TRPM4</t> channel activity by regulating its sensitivity to Ca 2+ in mpkCCD c14 Cells. (A) Representative single channel recording from inside-out patches exposed the patch membrane to the bath containing different concentrations of free Ca 2+ . The cells were treated with 30 μg/ml cholesterol for 48 hrs. From top to bottom: 1 μM, 10 μM, 200 μM, 1 mM, and 5 mM free bath Ca 2+ . “C” indicates channel at the closed state; “O” indicates single-level openings. (B) Representative single channel recording from inside-out patches exposed the patch membrane to the bath containing different concentrations of free Ca 2+ . The cells were treated with 5 μM lovastatin for 48 hrs. From top to bottom: 10 μM, 200 μM, 1 mM, 5 mM and 10 mM free bath Ca 2+ . “C” indicates channel at the closed state; “O” indicates single-level openings. (C) The effect of membrane cholesterol on Ca 2+ -dependence of channel opening. Channel P O was plotted as a function of free Ca 2+ concentration in the bath. P o values are shown for patches either with exogenous cholesterol treatment (black line) or lovastatin treatment (red line). n = 4–7 cells for different data points.
    Rabbit Polyclonal Anti Trpm4 Antibody, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/rabbit polyclonal anti trpm4 antibody/product/Alomone Labs
    Average 94 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    rabbit polyclonal anti trpm4 antibody - by Bioz Stars, 2023-02
    94/100 stars
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    94
    Alomone Labs rabbit polyclonal antibodies
    Treatment of cells with cholesterol increases, but with lovastatin decreases, <t>TRPM4</t> channel activity by regulating its sensitivity to Ca 2+ in mpkCCD c14 Cells. (A) Representative single channel recording from inside-out patches exposed the patch membrane to the bath containing different concentrations of free Ca 2+ . The cells were treated with 30 μg/ml cholesterol for 48 hrs. From top to bottom: 1 μM, 10 μM, 200 μM, 1 mM, and 5 mM free bath Ca 2+ . “C” indicates channel at the closed state; “O” indicates single-level openings. (B) Representative single channel recording from inside-out patches exposed the patch membrane to the bath containing different concentrations of free Ca 2+ . The cells were treated with 5 μM lovastatin for 48 hrs. From top to bottom: 10 μM, 200 μM, 1 mM, 5 mM and 10 mM free bath Ca 2+ . “C” indicates channel at the closed state; “O” indicates single-level openings. (C) The effect of membrane cholesterol on Ca 2+ -dependence of channel opening. Channel P O was plotted as a function of free Ca 2+ concentration in the bath. P o values are shown for patches either with exogenous cholesterol treatment (black line) or lovastatin treatment (red line). n = 4–7 cells for different data points.
    Rabbit Polyclonal Antibodies, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/rabbit polyclonal antibodies/product/Alomone Labs
    Average 94 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    rabbit polyclonal antibodies - by Bioz Stars, 2023-02
    94/100 stars
      Buy from Supplier

    94
    Alomone Labs rabbit polyclonal anti trpm4
    Treatment of cells with cholesterol increases, but with lovastatin decreases, <t>TRPM4</t> channel activity by regulating its sensitivity to Ca 2+ in mpkCCD c14 Cells. (A) Representative single channel recording from inside-out patches exposed the patch membrane to the bath containing different concentrations of free Ca 2+ . The cells were treated with 30 μg/ml cholesterol for 48 hrs. From top to bottom: 1 μM, 10 μM, 200 μM, 1 mM, and 5 mM free bath Ca 2+ . “C” indicates channel at the closed state; “O” indicates single-level openings. (B) Representative single channel recording from inside-out patches exposed the patch membrane to the bath containing different concentrations of free Ca 2+ . The cells were treated with 5 μM lovastatin for 48 hrs. From top to bottom: 10 μM, 200 μM, 1 mM, 5 mM and 10 mM free bath Ca 2+ . “C” indicates channel at the closed state; “O” indicates single-level openings. (C) The effect of membrane cholesterol on Ca 2+ -dependence of channel opening. Channel P O was plotted as a function of free Ca 2+ concentration in the bath. P o values are shown for patches either with exogenous cholesterol treatment (black line) or lovastatin treatment (red line). n = 4–7 cells for different data points.
    Rabbit Polyclonal Anti Trpm4, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/rabbit polyclonal anti trpm4/product/Alomone Labs
    Average 94 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    rabbit polyclonal anti trpm4 - by Bioz Stars, 2023-02
    94/100 stars
      Buy from Supplier

    94
    Alomone Labs rabbit polyclonal antibody
    Treatment of cells with cholesterol increases, but with lovastatin decreases, <t>TRPM4</t> channel activity by regulating its sensitivity to Ca 2+ in mpkCCD c14 Cells. (A) Representative single channel recording from inside-out patches exposed the patch membrane to the bath containing different concentrations of free Ca 2+ . The cells were treated with 30 μg/ml cholesterol for 48 hrs. From top to bottom: 1 μM, 10 μM, 200 μM, 1 mM, and 5 mM free bath Ca 2+ . “C” indicates channel at the closed state; “O” indicates single-level openings. (B) Representative single channel recording from inside-out patches exposed the patch membrane to the bath containing different concentrations of free Ca 2+ . The cells were treated with 5 μM lovastatin for 48 hrs. From top to bottom: 10 μM, 200 μM, 1 mM, 5 mM and 10 mM free bath Ca 2+ . “C” indicates channel at the closed state; “O” indicates single-level openings. (C) The effect of membrane cholesterol on Ca 2+ -dependence of channel opening. Channel P O was plotted as a function of free Ca 2+ concentration in the bath. P o values are shown for patches either with exogenous cholesterol treatment (black line) or lovastatin treatment (red line). n = 4–7 cells for different data points.
    Rabbit Polyclonal Antibody, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/rabbit polyclonal antibody/product/Alomone Labs
    Average 94 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    rabbit polyclonal antibody - by Bioz Stars, 2023-02
    94/100 stars
      Buy from Supplier

    94
    Alomone Labs rabbit polyclonal antibody to anti trpm4
    Treatment of cells with cholesterol increases, but with lovastatin decreases, <t>TRPM4</t> channel activity by regulating its sensitivity to Ca 2+ in mpkCCD c14 Cells. (A) Representative single channel recording from inside-out patches exposed the patch membrane to the bath containing different concentrations of free Ca 2+ . The cells were treated with 30 μg/ml cholesterol for 48 hrs. From top to bottom: 1 μM, 10 μM, 200 μM, 1 mM, and 5 mM free bath Ca 2+ . “C” indicates channel at the closed state; “O” indicates single-level openings. (B) Representative single channel recording from inside-out patches exposed the patch membrane to the bath containing different concentrations of free Ca 2+ . The cells were treated with 5 μM lovastatin for 48 hrs. From top to bottom: 10 μM, 200 μM, 1 mM, 5 mM and 10 mM free bath Ca 2+ . “C” indicates channel at the closed state; “O” indicates single-level openings. (C) The effect of membrane cholesterol on Ca 2+ -dependence of channel opening. Channel P O was plotted as a function of free Ca 2+ concentration in the bath. P o values are shown for patches either with exogenous cholesterol treatment (black line) or lovastatin treatment (red line). n = 4–7 cells for different data points.
    Rabbit Polyclonal Antibody To Anti Trpm4, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/rabbit polyclonal antibody to anti trpm4/product/Alomone Labs
    Average 94 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    rabbit polyclonal antibody to anti trpm4 - by Bioz Stars, 2023-02
    94/100 stars
      Buy from Supplier

    Image Search Results


    Treatment of cells with cholesterol increases, but with lovastatin decreases, TRPM4 channel activity by regulating its sensitivity to Ca 2+ in mpkCCD c14 Cells. (A) Representative single channel recording from inside-out patches exposed the patch membrane to the bath containing different concentrations of free Ca 2+ . The cells were treated with 30 μg/ml cholesterol for 48 hrs. From top to bottom: 1 μM, 10 μM, 200 μM, 1 mM, and 5 mM free bath Ca 2+ . “C” indicates channel at the closed state; “O” indicates single-level openings. (B) Representative single channel recording from inside-out patches exposed the patch membrane to the bath containing different concentrations of free Ca 2+ . The cells were treated with 5 μM lovastatin for 48 hrs. From top to bottom: 10 μM, 200 μM, 1 mM, 5 mM and 10 mM free bath Ca 2+ . “C” indicates channel at the closed state; “O” indicates single-level openings. (C) The effect of membrane cholesterol on Ca 2+ -dependence of channel opening. Channel P O was plotted as a function of free Ca 2+ concentration in the bath. P o values are shown for patches either with exogenous cholesterol treatment (black line) or lovastatin treatment (red line). n = 4–7 cells for different data points.

    Journal: Frontiers in Pharmacology

    Article Title: Cholesterol Stimulates the Transient Receptor Potential Melastatin 4 Channel in mpkCCD c14 Cells

    doi: 10.3389/fphar.2021.627875

    Figure Lengend Snippet: Treatment of cells with cholesterol increases, but with lovastatin decreases, TRPM4 channel activity by regulating its sensitivity to Ca 2+ in mpkCCD c14 Cells. (A) Representative single channel recording from inside-out patches exposed the patch membrane to the bath containing different concentrations of free Ca 2+ . The cells were treated with 30 μg/ml cholesterol for 48 hrs. From top to bottom: 1 μM, 10 μM, 200 μM, 1 mM, and 5 mM free bath Ca 2+ . “C” indicates channel at the closed state; “O” indicates single-level openings. (B) Representative single channel recording from inside-out patches exposed the patch membrane to the bath containing different concentrations of free Ca 2+ . The cells were treated with 5 μM lovastatin for 48 hrs. From top to bottom: 10 μM, 200 μM, 1 mM, 5 mM and 10 mM free bath Ca 2+ . “C” indicates channel at the closed state; “O” indicates single-level openings. (C) The effect of membrane cholesterol on Ca 2+ -dependence of channel opening. Channel P O was plotted as a function of free Ca 2+ concentration in the bath. P o values are shown for patches either with exogenous cholesterol treatment (black line) or lovastatin treatment (red line). n = 4–7 cells for different data points.

    Article Snippet: Briefly, after fixation with 4% paraformaldehyde at room temperature for 10 min, the cells were permeabilized with 0.2% Triton X-100 in PBS for 10 min and blocked with 5% BSA/PBS-T for 30 min. Rabbit polyclonal anti-TRPM4 antibody (alomone ACC-044; 1:100 dilution) was added in 1% BSA/TBS-T for overnight at 4°C.

    Techniques: Activity Assay, Concentration Assay

    Application of MβCD to the cytoplasmic bath decreases TRPM4 channel activity which is reversed by exogenous cholesterol. (A) A representative single channel recording from an inside-out patch shows TRPM4 activity before and after replacement of control cytoplasmic bath solution first with a solution containing 0.5 mM MβCD and then with a solution containing 30 μg/ml cholesterol. “C” indicates channel at the closed state. “O” indicates single-level openings. “I, II, and III” are zoom-ins of the single-channel recording. (B) Summary plots of TRPM4 channel P O under each indicted condition. n = 5 paired experiments, ** P < 0.01, significantly different with control; ## P < 0.01, significantly different with cells treated with 0.5 mM MβCD.

    Journal: Frontiers in Pharmacology

    Article Title: Cholesterol Stimulates the Transient Receptor Potential Melastatin 4 Channel in mpkCCD c14 Cells

    doi: 10.3389/fphar.2021.627875

    Figure Lengend Snippet: Application of MβCD to the cytoplasmic bath decreases TRPM4 channel activity which is reversed by exogenous cholesterol. (A) A representative single channel recording from an inside-out patch shows TRPM4 activity before and after replacement of control cytoplasmic bath solution first with a solution containing 0.5 mM MβCD and then with a solution containing 30 μg/ml cholesterol. “C” indicates channel at the closed state. “O” indicates single-level openings. “I, II, and III” are zoom-ins of the single-channel recording. (B) Summary plots of TRPM4 channel P O under each indicted condition. n = 5 paired experiments, ** P < 0.01, significantly different with control; ## P < 0.01, significantly different with cells treated with 0.5 mM MβCD.

    Article Snippet: Briefly, after fixation with 4% paraformaldehyde at room temperature for 10 min, the cells were permeabilized with 0.2% Triton X-100 in PBS for 10 min and blocked with 5% BSA/PBS-T for 30 min. Rabbit polyclonal anti-TRPM4 antibody (alomone ACC-044; 1:100 dilution) was added in 1% BSA/TBS-T for overnight at 4°C.

    Techniques: Activity Assay

    Cholesterol stimulates TRPM4 via a PI(4, 5)P 2 -dependent mechanism. (A) A representative single channel recording shows that treatment of mpkCCD c14 cells with 20 nM wortmannin had no effect on cholesterol-induced TRPM4 channel activity. (B) Summary plots of TRPM4 channel P O under each indicated conditions. n = 6 paired experiments, ** P < 0.01, significantly different with cells treated with 20 nm wortmannin. (C) A representative single channel recording shows that depletion of PI(4, 5)P 2 with 20 μM wortmannin abrogated cholesterol-induced TRPM4 channel activity. (D) Summary plots of TRPM4 channel P O under indicated conditions. n = 5 paired experiments. (E) A representative single channel recording shows that application of 20 μM diC8-PI(4,5)P 2 induced TRPM4 channel activity after cholesterol failed to stimulate TRPM4 in the presence of 2 μM PGE2. (F) Summary plots of TRPM4 channel P O under indicated conditions. n = 4 paired experiments. ** P < 0.01, significantly different with cells treated with 2 μM PGE2 or 30 μg/ml cholesterol.

    Journal: Frontiers in Pharmacology

    Article Title: Cholesterol Stimulates the Transient Receptor Potential Melastatin 4 Channel in mpkCCD c14 Cells

    doi: 10.3389/fphar.2021.627875

    Figure Lengend Snippet: Cholesterol stimulates TRPM4 via a PI(4, 5)P 2 -dependent mechanism. (A) A representative single channel recording shows that treatment of mpkCCD c14 cells with 20 nM wortmannin had no effect on cholesterol-induced TRPM4 channel activity. (B) Summary plots of TRPM4 channel P O under each indicated conditions. n = 6 paired experiments, ** P < 0.01, significantly different with cells treated with 20 nm wortmannin. (C) A representative single channel recording shows that depletion of PI(4, 5)P 2 with 20 μM wortmannin abrogated cholesterol-induced TRPM4 channel activity. (D) Summary plots of TRPM4 channel P O under indicated conditions. n = 5 paired experiments. (E) A representative single channel recording shows that application of 20 μM diC8-PI(4,5)P 2 induced TRPM4 channel activity after cholesterol failed to stimulate TRPM4 in the presence of 2 μM PGE2. (F) Summary plots of TRPM4 channel P O under indicated conditions. n = 4 paired experiments. ** P < 0.01, significantly different with cells treated with 2 μM PGE2 or 30 μg/ml cholesterol.

    Article Snippet: Briefly, after fixation with 4% paraformaldehyde at room temperature for 10 min, the cells were permeabilized with 0.2% Triton X-100 in PBS for 10 min and blocked with 5% BSA/PBS-T for 30 min. Rabbit polyclonal anti-TRPM4 antibody (alomone ACC-044; 1:100 dilution) was added in 1% BSA/TBS-T for overnight at 4°C.

    Techniques: Activity Assay

    PI(4, 5)P 2 stimulates TRPM4 via a physical interaction. (A) A representative single channel recording shows that diC8-PI(4,5)P 2 significantly increased TRPM4 activity. (B) Summary plots of TRPM4 channel P O under each indicated conditions. n = 5 paired experiments. ** P < 0.01, significantly different with control. (C) TRPM4 was detected in dots where most anionic phospholipids including PI(4, 5)P 2 are located on PIP Strips membrane. Data represent three individual experiments showing consistent results.

    Journal: Frontiers in Pharmacology

    Article Title: Cholesterol Stimulates the Transient Receptor Potential Melastatin 4 Channel in mpkCCD c14 Cells

    doi: 10.3389/fphar.2021.627875

    Figure Lengend Snippet: PI(4, 5)P 2 stimulates TRPM4 via a physical interaction. (A) A representative single channel recording shows that diC8-PI(4,5)P 2 significantly increased TRPM4 activity. (B) Summary plots of TRPM4 channel P O under each indicated conditions. n = 5 paired experiments. ** P < 0.01, significantly different with control. (C) TRPM4 was detected in dots where most anionic phospholipids including PI(4, 5)P 2 are located on PIP Strips membrane. Data represent three individual experiments showing consistent results.

    Article Snippet: Briefly, after fixation with 4% paraformaldehyde at room temperature for 10 min, the cells were permeabilized with 0.2% Triton X-100 in PBS for 10 min and blocked with 5% BSA/PBS-T for 30 min. Rabbit polyclonal anti-TRPM4 antibody (alomone ACC-044; 1:100 dilution) was added in 1% BSA/TBS-T for overnight at 4°C.

    Techniques: Activity Assay

    TRPM4 channels are mainly located in lipid rafts. (A) Representative confocal microscopy images indicate that majority of TRPM4 (green) was co-localized with cholera toxin B (red) in the apical membrane; white rectangular box indicate zoomed-in areas shown in the Zoom-in panels. Data represent five individual experiments showing consistent results (B) Majority of TRPM4 was detected in low-density regions in sucrose gradient experiments; Caveolin-1 was used as a control protein that is known to be located in lipid rafts. Data represent three individual experiments showing consistent results.

    Journal: Frontiers in Pharmacology

    Article Title: Cholesterol Stimulates the Transient Receptor Potential Melastatin 4 Channel in mpkCCD c14 Cells

    doi: 10.3389/fphar.2021.627875

    Figure Lengend Snippet: TRPM4 channels are mainly located in lipid rafts. (A) Representative confocal microscopy images indicate that majority of TRPM4 (green) was co-localized with cholera toxin B (red) in the apical membrane; white rectangular box indicate zoomed-in areas shown in the Zoom-in panels. Data represent five individual experiments showing consistent results (B) Majority of TRPM4 was detected in low-density regions in sucrose gradient experiments; Caveolin-1 was used as a control protein that is known to be located in lipid rafts. Data represent three individual experiments showing consistent results.

    Article Snippet: Briefly, after fixation with 4% paraformaldehyde at room temperature for 10 min, the cells were permeabilized with 0.2% Triton X-100 in PBS for 10 min and blocked with 5% BSA/PBS-T for 30 min. Rabbit polyclonal anti-TRPM4 antibody (alomone ACC-044; 1:100 dilution) was added in 1% BSA/TBS-T for overnight at 4°C.

    Techniques: Confocal Microscopy

    Treatment with cholesterol or lovastatin does not alter expression levels of TRPM4 in mpkCCDc14 Cells. (A) Representative confocal microscopy images of mpkCCDc14 cells stained with TRPM4 antibody under each indicated conditions. (B) Summary plots of fluorescence intensity of TRPM4. Data are from 24 cells in four sets of separate experiments. (C) Representative Western blots from cell-surface biotinylated and the total proteins of TRPM4 protein. (D) Summary plots of relative expression of TRPM4. Cells were either under control conditions or treated with 30 μg/ml cholesterol alone, 30 μg/ml cholesterol plus 5 μM lovastatin, or 5 μM lovastatin alone for 48 hrs, respectively. n = 5. (E) Representative single channel recording from inside-out patches exposed the patch membrane to the bath containing 5 mM CaCl 2 , followed by a bath solution with 10 mM EGTA and no calcium. Cells were either under control conditions or treated with 30 μg/ml cholesterol, or 5 μM lovastatin for 48 hrs, respectively. (F) Summary plots of the number of active channels in the patches under each indicated condition. n = 5 for control cells, n = 5 for cells treated with cholesterol, n = 4 for cells treated with lovastatin.

    Journal: Frontiers in Pharmacology

    Article Title: Cholesterol Stimulates the Transient Receptor Potential Melastatin 4 Channel in mpkCCD c14 Cells

    doi: 10.3389/fphar.2021.627875

    Figure Lengend Snippet: Treatment with cholesterol or lovastatin does not alter expression levels of TRPM4 in mpkCCDc14 Cells. (A) Representative confocal microscopy images of mpkCCDc14 cells stained with TRPM4 antibody under each indicated conditions. (B) Summary plots of fluorescence intensity of TRPM4. Data are from 24 cells in four sets of separate experiments. (C) Representative Western blots from cell-surface biotinylated and the total proteins of TRPM4 protein. (D) Summary plots of relative expression of TRPM4. Cells were either under control conditions or treated with 30 μg/ml cholesterol alone, 30 μg/ml cholesterol plus 5 μM lovastatin, or 5 μM lovastatin alone for 48 hrs, respectively. n = 5. (E) Representative single channel recording from inside-out patches exposed the patch membrane to the bath containing 5 mM CaCl 2 , followed by a bath solution with 10 mM EGTA and no calcium. Cells were either under control conditions or treated with 30 μg/ml cholesterol, or 5 μM lovastatin for 48 hrs, respectively. (F) Summary plots of the number of active channels in the patches under each indicated condition. n = 5 for control cells, n = 5 for cells treated with cholesterol, n = 4 for cells treated with lovastatin.

    Article Snippet: Briefly, after fixation with 4% paraformaldehyde at room temperature for 10 min, the cells were permeabilized with 0.2% Triton X-100 in PBS for 10 min and blocked with 5% BSA/PBS-T for 30 min. Rabbit polyclonal anti-TRPM4 antibody (alomone ACC-044; 1:100 dilution) was added in 1% BSA/TBS-T for overnight at 4°C.

    Techniques: Expressing, Confocal Microscopy, Staining, Fluorescence, Western Blot