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Gary A. Quamme - One of the best experts on this subject based on the ideXlab platform.

  • Immunosuppressants inhibit hormone-stimulated Mg2+ uptake in mouse Distal Convoluted Tubule cells.
    Biochemical and biophysical research communications, 2006
    Co-Authors: Shang-jin Kim, Hyung Sub Kang, Chang-woo Jeong, Sang-yeol Park, In-shik Kim, Nam-soo Kim, Sung-zoo Kim, Yong-geun Kwak, Jin-shang Kim, Gary A. Quamme
    Abstract:

    Abstract Immunosuppressants such as cyclosporinA and FK506 (tacrolimus) are widely prescribed to treat numerous disorders and to treat organ transplant recipients. However, cyclosporine A and FK506 are both known to produce hypomagnesaemia. The mechanism of this effect is still unclear. The present study determined the effects of immunosuppressant treatment on the parathyroid hormone (PTH) mediated Mg2+ uptake and the mitogen-activated protein kinase (MAPK) activation in mouse Distal Convoluted Tubule (MDCT) cells. The intracellular Ca2+ and Mg2+ concentrations in a single MDCT cell were measured by using the fluorescentdye Fura-2 AM and Mag-fura-2 AM, respectively. Cyclosporine A and FK506 illicited a transient increase of intracellular Ca2+ from a basal level of 99 ± 16 nM to 685 ± 105 and 608 ± 96 nM, respectively. In order to determine the Mg2+ transport, the MDCT cells were Mg2+-depleted by culturing them in Mg2+-free media for 16 h, and the Mg2+ uptake was measured by microfluorescence after placing the depleted cells in 1.5 mM MgCl2. The mean rate of Mg2+ uptake, d([Mg2+]i)/dt, was 140 ± 16 nM/s in the control MDCT cells. PTH increased the Mg2+ uptake more than 2 times in this cell. Cyclosporine A (10 μM) and FK506 (0.1 μM) did not affect the basal Mg2+uptake (140 ± 16 and 142 ± 14 nM/s, respectively), but they inhibited the PTH-stimulated Mg2+ entry, decreasing it from 248 ± 12 to 147 ± 7 and 148 ± 14 nM/s, respectively. These effects were inhibited by L685818, which is a potent competitive antagonist of FK506. PTH stimulated the extracellular signal-regulated kinase1/2 (ERK1/2) protein synthesis. This PTH-stimulated ERK1/2 activation was inhibited by cyclosporine A and FK506. In the present study, the role of ERK1/2 activation on the PTH-dependent magnesium uptake was examined in MDCT cells, and we showed that immunosuppressants inhibit the hormone-stimulated Mg2+ uptake into the MDCT cells by inhibiting the MAPK pathway.

  • Adenosine modulates Mg2+ uptake in Distal Convoluted Tubule cells via A1 and A2purinoceptors
    American journal of physiology. Renal physiology, 2001
    Co-Authors: Hyung Sub Kang, Long-jun Dai, Gordon Ritchie, Dirk Kerstan, Gary A. Quamme
    Abstract:

    tk;1Adenosine plays a role in the control of water and electrolyte reabsorption in the Distal Tubule. As the Distal Convoluted Tubule is important in the regulation of renal Mg2+ balance, we determ...

  • β-Adrenergic agonists stimulate Mg2+ uptake in mouse Distal Convoluted Tubule cells
    American journal of physiology. Renal physiology, 2000
    Co-Authors: Hyung Sub Kang, Long-jun Dai, Gordon Ritchie, Dirk Kerstan, Gary A. Quamme
    Abstract:

    β-Adrenergic agonists influence electrolyte reabsorption in the proximal Tubule, loop of Henle, and Distal Tubule. Although isoproterenol enhances magnesium absorption in the thick ascending limb, it is unclear what effect, if any, β-adrenergic agonists have on tubular magnesium handling. The effects of isoproterenol were studied in immortalized mouse Distal Convoluted Tubule (MDCT) cells by measuring cellular cAMP formation with radioimmunoassays and Mg2+ uptake with fluorescence techniques. Intracellular free Mg2+ concentration ([Mg2+]i) was measured in single MDCT cells by using microfluorescence with mag-fura-2. To assess Mg2+uptake, MDCT cells were first Mg2+ depleted to 0.22 ± 0.01 mM by culturing in Mg2+-free media for 16 h and then placed in 1.5 mM MgCl2, and the changes in [Mg2+]i were determined. [Mg2+]i returned to basal levels, 0.53 ± 0.02 mM, with a mean refill rate, d([Mg2+]i)/dt, of 168 ± 11 nM/s. Isoproterenol stimulated Mg2+ entry in a concentration-dependent manner, with a maximal respon...

  • aminoglycosides inhibit hormone stimulated mg2 uptake in mouse Distal Convoluted Tubule cells
    Canadian Journal of Physiology and Pharmacology, 2000
    Co-Authors: Hyung Sub Kang, Long-jun Dai, Gordon Ritchie, Dirk Kerstan, Gary A. Quamme
    Abstract:

    The clinical use of aminoglycosides often leads to renal magnesium wasting and hypomagnesemia. Of the nephron segments, both the thick ascending limb of Henle's loop and the Distal Tubule play significant roles in renal magnesium conservation but the Distal Convoluted Tubule exerts the final control of urinary excretion. An immortalized mouse Distal Convoluted Tubule (MDCT) cell line has been extensively used to study the cellular mechanisms of magnesium transport in this nephron segment. Peptide hormones, such as parathyroid hormone (PTH), glucagon, calcitonin, and arginine vasopressin (AVP) stimulate Mg2+ uptake in MDCT cells that is modulated by extracellular polyvalent cations, Ca2+ and Mg2+. The present studies determined the effect of aminoglycosides on parathyroid hormone (PTH)-mediated cAMP formation and Mg2+ uptake in MDCT cells. Gentamicin, a prototypic aminoglycoside, elicited transient increases in intracellular Ca2+ from basal levels of 102 +/- 13 nM to 713 +/- 125 nM, suggesting a receptor-mediated response. In order to determine Mg2+ transport, MDCT cells were Mg(2+)-depleted by culturing in Mg(2+)-free media for 16 h and Mg2+ uptake was measured by microfluorescence after placing the depleted cells in 1.0 mM MgCl2. The mean rate of Mg2+ uptake, d([Mg2+]i)/dt, was 138 +/- 24 nM/s in control MDCT cells. Gentamicin (50 microM) did not affect basal Mg2+ uptake (105 +/- 29 nM/s), but inhibited PTH stimulated Mg2+ entry, decreasing it from 257 +/- 36 nM/s to 108 +/- 42 nM/s. This was associated with diminished PTH-stimulated cAMP formation, from 80 +/- 2.5 to 23 +/- 1 pmol/mg protein x 5 min. Other aminoglycosides such as tobramycin, streptomycin, and neomycin also inhibited PTH-stimulated Mg2+ entry and cAMP formation. As these antibiotics are positively charged, the data suggest that aminoglycosides act through an extracellular polyvalent cation-sensing receptor present in Distal Convoluted Tubule cells. We infer from these studies that aminoglycosides inhibit hormone-stimulated Mg2+ absorption in the Distal Convoluted Tubule that may contribute to the renal magnesium wasting frequently observed with the clinical use of these antibiotics.

  • Aminoglycosides inhibit hormone-stimulated Mg 2+ uptake in mouse Distal Convoluted Tubule cells
    Canadian journal of physiology and pharmacology, 2000
    Co-Authors: Hyung Sub Kang, Long-jun Dai, Gordon Ritchie, Dirk Kerstan, Gary A. Quamme
    Abstract:

    The clinical use of aminoglycosides often leads to renal magnesium wasting and hypomagnesemia. Of the nephron segments, both the thick ascending limb of Henle's loop and the Distal Tubule play significant roles in renal magnesium conservation but the Distal Convoluted Tubule exerts the final control of urinary excretion. An immortalized mouse Distal Convoluted Tubule (MDCT) cell line has been extensively used to study the cellular mechanisms of magnesium transport in this nephron segment. Peptide hormones, such as parathyroid hormone (PTH), glucagon, calcitonin, and arginine vasopressin (AVP) stimulate Mg 2+ uptake in MDCT cells that is modulated by extracellular polyvalent cations, Ca 2+ and Mg 2+ . The present studies determined the effect of aminoglycosides on parathyroid hormone (PTH)-mediated cAMP formation and Mg 2+ uptake in MDCT cells. Gentamicin, a prototypic aminoglycoside, illicited transient increases in intracellular Ca 2+ from basal levels of 102 ± 13 nM to 713 ± 125 nM, suggesting a receptor-mediated response. In order to determine Mg 2+ transport, MDCT cells were Mg 2+ -depleted by culturing in Mg 2+ -free media for 16 h and Mg 2+ uptake was measured by microfluorescence after placing the depleted cells in 1.0 mM MgCl2. The mean rate of Mg 2+ uptake, d((Mg 2+ )i)/dt, was 138 ± 24 nM/s in control MDCT cells. Gentamicin (50 μM) did not affect basal Mg 2+ uptake (105 ± 29 nM/s), but inhibited PTH stimulated Mg 2+ entry, decreasing it from 257 ± 36 nM/s to 108 ± 42 nM/s. This was associated with diminished PTH-stimulated cAMP formation, from 80 ±2 .5 to 23 ±1p mol/mgprotein·5 min. Other aminoglycosides such as tobramycin, streptomycin, and neomycin also inhibited PTH-stimulated Mg 2+ entry and cAMP formation. As these antibiotics are positively charged, the data suggest that aminoglycosides act through an extracellular polyvalent cation-sensing receptor present in Distal Convoluted Tubule cells. We infer from these studies that aminoglycosides inhibit hormone-stimulated Mg 2+ absorption in the Distal Convoluted Tubule that may contribute to the renal magnesium wasting frequently observed with the clinical use of

Long-jun Dai - One of the best experts on this subject based on the ideXlab platform.

  • Adenosine modulates Mg2+ uptake in Distal Convoluted Tubule cells via A1 and A2purinoceptors
    American journal of physiology. Renal physiology, 2001
    Co-Authors: Hyung Sub Kang, Long-jun Dai, Gordon Ritchie, Dirk Kerstan, Gary A. Quamme
    Abstract:

    tk;1Adenosine plays a role in the control of water and electrolyte reabsorption in the Distal Tubule. As the Distal Convoluted Tubule is important in the regulation of renal Mg2+ balance, we determ...

  • β-Adrenergic agonists stimulate Mg2+ uptake in mouse Distal Convoluted Tubule cells
    American journal of physiology. Renal physiology, 2000
    Co-Authors: Hyung Sub Kang, Long-jun Dai, Gordon Ritchie, Dirk Kerstan, Gary A. Quamme
    Abstract:

    β-Adrenergic agonists influence electrolyte reabsorption in the proximal Tubule, loop of Henle, and Distal Tubule. Although isoproterenol enhances magnesium absorption in the thick ascending limb, it is unclear what effect, if any, β-adrenergic agonists have on tubular magnesium handling. The effects of isoproterenol were studied in immortalized mouse Distal Convoluted Tubule (MDCT) cells by measuring cellular cAMP formation with radioimmunoassays and Mg2+ uptake with fluorescence techniques. Intracellular free Mg2+ concentration ([Mg2+]i) was measured in single MDCT cells by using microfluorescence with mag-fura-2. To assess Mg2+uptake, MDCT cells were first Mg2+ depleted to 0.22 ± 0.01 mM by culturing in Mg2+-free media for 16 h and then placed in 1.5 mM MgCl2, and the changes in [Mg2+]i were determined. [Mg2+]i returned to basal levels, 0.53 ± 0.02 mM, with a mean refill rate, d([Mg2+]i)/dt, of 168 ± 11 nM/s. Isoproterenol stimulated Mg2+ entry in a concentration-dependent manner, with a maximal respon...

  • aminoglycosides inhibit hormone stimulated mg2 uptake in mouse Distal Convoluted Tubule cells
    Canadian Journal of Physiology and Pharmacology, 2000
    Co-Authors: Hyung Sub Kang, Long-jun Dai, Gordon Ritchie, Dirk Kerstan, Gary A. Quamme
    Abstract:

    The clinical use of aminoglycosides often leads to renal magnesium wasting and hypomagnesemia. Of the nephron segments, both the thick ascending limb of Henle's loop and the Distal Tubule play significant roles in renal magnesium conservation but the Distal Convoluted Tubule exerts the final control of urinary excretion. An immortalized mouse Distal Convoluted Tubule (MDCT) cell line has been extensively used to study the cellular mechanisms of magnesium transport in this nephron segment. Peptide hormones, such as parathyroid hormone (PTH), glucagon, calcitonin, and arginine vasopressin (AVP) stimulate Mg2+ uptake in MDCT cells that is modulated by extracellular polyvalent cations, Ca2+ and Mg2+. The present studies determined the effect of aminoglycosides on parathyroid hormone (PTH)-mediated cAMP formation and Mg2+ uptake in MDCT cells. Gentamicin, a prototypic aminoglycoside, elicited transient increases in intracellular Ca2+ from basal levels of 102 +/- 13 nM to 713 +/- 125 nM, suggesting a receptor-mediated response. In order to determine Mg2+ transport, MDCT cells were Mg(2+)-depleted by culturing in Mg(2+)-free media for 16 h and Mg2+ uptake was measured by microfluorescence after placing the depleted cells in 1.0 mM MgCl2. The mean rate of Mg2+ uptake, d([Mg2+]i)/dt, was 138 +/- 24 nM/s in control MDCT cells. Gentamicin (50 microM) did not affect basal Mg2+ uptake (105 +/- 29 nM/s), but inhibited PTH stimulated Mg2+ entry, decreasing it from 257 +/- 36 nM/s to 108 +/- 42 nM/s. This was associated with diminished PTH-stimulated cAMP formation, from 80 +/- 2.5 to 23 +/- 1 pmol/mg protein x 5 min. Other aminoglycosides such as tobramycin, streptomycin, and neomycin also inhibited PTH-stimulated Mg2+ entry and cAMP formation. As these antibiotics are positively charged, the data suggest that aminoglycosides act through an extracellular polyvalent cation-sensing receptor present in Distal Convoluted Tubule cells. We infer from these studies that aminoglycosides inhibit hormone-stimulated Mg2+ absorption in the Distal Convoluted Tubule that may contribute to the renal magnesium wasting frequently observed with the clinical use of these antibiotics.

  • Aminoglycosides inhibit hormone-stimulated Mg 2+ uptake in mouse Distal Convoluted Tubule cells
    Canadian journal of physiology and pharmacology, 2000
    Co-Authors: Hyung Sub Kang, Long-jun Dai, Gordon Ritchie, Dirk Kerstan, Gary A. Quamme
    Abstract:

    The clinical use of aminoglycosides often leads to renal magnesium wasting and hypomagnesemia. Of the nephron segments, both the thick ascending limb of Henle's loop and the Distal Tubule play significant roles in renal magnesium conservation but the Distal Convoluted Tubule exerts the final control of urinary excretion. An immortalized mouse Distal Convoluted Tubule (MDCT) cell line has been extensively used to study the cellular mechanisms of magnesium transport in this nephron segment. Peptide hormones, such as parathyroid hormone (PTH), glucagon, calcitonin, and arginine vasopressin (AVP) stimulate Mg 2+ uptake in MDCT cells that is modulated by extracellular polyvalent cations, Ca 2+ and Mg 2+ . The present studies determined the effect of aminoglycosides on parathyroid hormone (PTH)-mediated cAMP formation and Mg 2+ uptake in MDCT cells. Gentamicin, a prototypic aminoglycoside, illicited transient increases in intracellular Ca 2+ from basal levels of 102 ± 13 nM to 713 ± 125 nM, suggesting a receptor-mediated response. In order to determine Mg 2+ transport, MDCT cells were Mg 2+ -depleted by culturing in Mg 2+ -free media for 16 h and Mg 2+ uptake was measured by microfluorescence after placing the depleted cells in 1.0 mM MgCl2. The mean rate of Mg 2+ uptake, d((Mg 2+ )i)/dt, was 138 ± 24 nM/s in control MDCT cells. Gentamicin (50 μM) did not affect basal Mg 2+ uptake (105 ± 29 nM/s), but inhibited PTH stimulated Mg 2+ entry, decreasing it from 257 ± 36 nM/s to 108 ± 42 nM/s. This was associated with diminished PTH-stimulated cAMP formation, from 80 ±2 .5 to 23 ±1p mol/mgprotein·5 min. Other aminoglycosides such as tobramycin, streptomycin, and neomycin also inhibited PTH-stimulated Mg 2+ entry and cAMP formation. As these antibiotics are positively charged, the data suggest that aminoglycosides act through an extracellular polyvalent cation-sensing receptor present in Distal Convoluted Tubule cells. We infer from these studies that aminoglycosides inhibit hormone-stimulated Mg 2+ absorption in the Distal Convoluted Tubule that may contribute to the renal magnesium wasting frequently observed with the clinical use of

  • b-Adrenergic agonists stimulate Mg 21 uptake in mouse Distal Convoluted Tubule cells
    2000
    Co-Authors: Hyung Sub Kang, Long-jun Dai, Gordon Ritchie, Dirk Kerstan, Gary A. Quamme
    Abstract:

    Kang, Hyung Sub, Dirk Kerstan, Long-Jun Dai, Gordon Ritchie, and Gary A. Quamme. b-Adrenergic agonists stimulate Mg uptake in mouse Distal Convoluted Tubule cells. Am J Physiol Renal Physiol 279: F1116–F1123, 2000.—b-Adrenergic agonists influence electrolyte reabsorption in the proximal Tubule, loop of Henle, and Distal Tubule. Although isoproterenol enhances magnesium absorption in the thick ascending limb, it is unclear what effect, if any, b-adrenergic agonists have on tubular magnesium handling. The effects of isoproterenol were studied in immortalized mouse Distal Convoluted Tubule (MDCT) cells by measuring cellular cAMP formation with radioimmunoassays and Mg uptake with fluorescence techniques. Intracellular free Mg concentration ([Mg]i) was measured in single MDCT cells by using microfluorescence with mag-fura-2. To assess Mg uptake, MDCT cells were first Mg depleted to 0.22 6 0.01 mM by culturing in Mg-free media for 16 h and then placed in 1.5 mM MgCl2, and the changes in [Mg ]i were determined. [Mg]i returned to basal levels, 0.53 6 0.02 mM, with a mean refill rate, d([Mg]i)/dt, of 168 6 11 nM/s. Isoproterenol stimulated Mg entry in a concentration-dependent manner, with a maximal response of 252 6 11 nM/s, at a concentration of 10 M, that represented a 50 6 7% increase in uptake rate above control values. This was associated with a sixfold increase in intracellular cAMP generation. Isoproterenol-stimulated Mg uptake was completely inhibited with RpcAMPS, a protein kinase A inhibitor, and U-73122, a phospholipase C inhibitor, and partially blocked by RO 31– 822, a protein kinase C inhibitor. Accordingly, isoproterenolmediated Mg entry rates involve multiple intracellular signaling pathways. Aldosterone potentiated isoproterenolstimulated Mg uptake (326 6 31 nM/s), whereas elevation of extracellular Ca inhibited isoproterenol-mediated cAMP accumulation and Mg uptake, 117 6 37 nM/s. These studies demonstrate that isoproterenol stimulates Mg uptake in a cell line of mouse Distal Convoluted Tubules that is modulated by hormonal and extracellular influences.

David H. Ellison - One of the best experts on this subject based on the ideXlab platform.

  • Distal Convoluted Tubule sexual dimorphism revealed by advanced 3D imaging
    American journal of physiology. Renal physiology, 2020
    Co-Authors: Ebrahim Tahaei, David H. Ellison, Richard Coleman, Turgay Saritas, Paul A. Welling
    Abstract:

    The thiazide-sensitive Na+-Cl− cotransporter (NCC) is more abundant in kidneys of female subjects than of male subjects. Because morphological remodeling of the Distal Convoluted Tubule (DCT) is de...

  • Distal Convoluted Tubule Cl- concentration is modulated via K+ channels and transporters
    American journal of physiology. Renal physiology, 2020
    Co-Authors: Nathan J. Klett, Wen Hui Wang, Avika Sharma, Charles N. Allen, Chao Ling Yang, David H. Ellison
    Abstract:

    Cl--sensitive with-no-lysine kinase (WNK) plays a key role in regulating the thiazide-sensitive Na+-Cl- cotransporter (NCC) in the Distal Convoluted Tubule (DCT). Cl- enters DCT cells through NCC and leaves the cell across the basolateral membrane via the Cl- channel ClC-K2 or K+-Cl- cotransporter (KCC). While KCC is electroneutral, Cl- exit via ClC-K2 is electrogenic. Therefore, an alteration in DCT basolateral K+ channel activity is expected to influence Cl- movement across the basolateral membrane. Although a role for intracellular Cl- in the regulation of WNK and NCC has been established, intracellular Cl- concentrations ([Cl-]i) have not been directly measured in the mammalian DCT. Therefore, to measure [Cl-]i in DCT cells, we generated a transgenic mouse model expressing an optogenetic kidney-specific Cl-Sensor and measured Cl- fluorescent imaging in the isolated DCT. Basal measurements indicated that the mean [Cl-]i was ~7 mM. Stimulation of Cl- exit with low-Cl- hypotonic solutions decreased [Cl-]i, whereas inhibition of KCC by DIOA or inhibition of ClC-K2 by NPPB increased [Cl-]i, suggesting roles for both KCC and ClC-K2 in the modulation of [Cl-]i . Blockade of basolateral K+ channels (Kir4.1/5.1) with barium significantly increased [Cl-]i. Finally, a decrease in extracellular K+ concentration transiently decreased [Cl-]i, whereas raising extracellular K+ transiently increased [Cl-]i, further suggesting a role for Kir4.1/5.1 in the regulation of [Cl-]i. We conclude that the alteration in ClC-K2, KCC, and Kir4.1/5.1 activity influences [Cl-]i in the DCT.

  • A novel Distal Convoluted Tubule-specific Cre-recombinase driven by the NaCl cotransporter gene.
    American journal of physiology. Renal physiology, 2020
    Co-Authors: Ryan J. Cornelius, David H. Ellison, Avika Sharma, Jin Jin Guo, Jill A. Mcmahon, Andrew P. Mcmahon, James A. Mccormick
    Abstract:

    Cre-lox technology has revolutionized research in renal physiology by allowing site-specific genetic recombination in individual nephron segments. The Distal Convoluted Tubule (DCT), consisting of distinct early (DCT1) and late (DCT2) segments, plays a central role in Na+ and K+ homeostasis. The only established Cre line targeting the DCT is Pvalb-Cre, which is limited by noninducibility, activity along DCT1 only, and activity in neurons. Here, we report the characterization of the first Cre line specific to the entire DCT. CRISPR/Cas9 targeting was used to introduce a tamoxifen-inducible IRES-Cre-ERT2 cassette downstream of the coding region of the Slc12a3 gene encoding the NaCl cotransporter (NCC). The resulting Slc12a3-Cre-ERT2 mice were crossed with R26R-YFP reporter mice, which revealed minimal leakiness with 6.3% of NCC-positive cells expressing yellow fluorescent protein (YFP) in the absence of tamoxifen. After tamoxifen injection, YFP expression was observed in 91.2% of NCC-positive cells and only in NCC-positive cells, revealing high recombination efficiency and DCT specificity. Crossing to R26R-TdTomato mice revealed higher leakiness (64.5%), suggesting differential sensitivity of the floxed site. Western blot analysis revealed no differences in abundances of total NCC or the active phosphorylated form of NCC in Slc12a3-Cre-ERT2 mice of either sex compared with controls. Plasma K+ and Mg2+ concentrations and thiazide-sensitive Na+ and K+ excretion did not differ in Slc12a3-Cre-ERT2 mice compared with controls when sex matched. These data suggest genetic modification had no obvious effect on NCC function. Slc12a3-Cre-ERT2 mice are the first line generated demonstrating inducible Cre recombinase activity along the entire DCT and will be a useful tool to study DCT function.

  • Effect of Angiotensin II on ENaC in the Distal Convoluted Tubule and in the Cortical Collecting Duct of Mineralocorticoid Receptor Deficient Mice
    Journal of the American Heart Association, 2020
    Co-Authors: Zhong Xiuzi Gao, David H. Ellison, Andrew S. Terker, Dan-dan Zhang, Xin Peng Duan, Dao Hong Lin, Wen Hui Wang
    Abstract:

    Background Angiotensin II stimulates epithelial Na+ channel (ENaC) by aldosterone‐independent mechanism. We now test the effect of angiotensin II on ENaC in the Distal Convoluted Tubule (DCT) and c...

  • Distal Convoluted Tubule.
    Comprehensive Physiology, 2014
    Co-Authors: James A. Mccormick, David H. Ellison
    Abstract:

    The Distal Convoluted Tubule (DCT) is a short nephron segment, interposed between the macula densa and collecting duct. Even though it is short, it plays a key role in regulating extracellular fluid volume and electrolyte homeostasis. DCT cells are rich in mitochondria, and possess the highest density of Na+/K+-ATPase along the nephron, where it is expressed on the highly amplified basolateral membranes. DCT cells are largely water impermeable, and reabsorb sodium and chloride across the apical membrane via electroneurtral pathways. Prominent among this is the thiazide-sensitive sodium chloride cotransporter, target of widely used diuretic drugs. These cells also play a key role in magnesium reabsorption, which occurs predominantly, via a transient receptor potential channel (TRPM6). Human genetic diseases in which DCT function is perturbed have provided critical insights into the physiological role of the DCT, and how transport is regulated. These include Familial Hyperkalemic Hypertension, the salt-wasting diseases Gitelman syndrome and EAST syndrome, and hereditary hypomagnesemias. The DCT is also established as an important target for the hormones angiotensin II and aldosterone; it also appears to respond to sympathetic-nerve stimulation and changes in plasma potassium. Here, we discuss what is currently known about DCT physiology. Early studies that determined transport rates of ions by the DCT are described, as are the channels and transporters expressed along the DCT with the advent of molecular cloning. Regulation of expression and activity of these channels and transporters is also described; particular emphasis is placed on the contribution of genetic forms of DCT dysregulation to our understanding.

Rene J M Bindels - One of the best experts on this subject based on the ideXlab platform.

  • mechanisms coupling sodium and magnesium reabsorption in the Distal Convoluted Tubule of the kidney
    Acta Physiologica, 2020
    Co-Authors: Gijs Franken, Anastasia Adella, Rene J M Bindels, Jeroen H F De Baaij
    Abstract:

    Hypomagnesaemia is a common feature of renal Na+ wasting disorders such as Gitelman and EAST/SeSAME syndrome. These genetic defects specifically affect Na+ reabsorption in the Distal Convoluted Tubule, where Mg2+ reabsorption is tightly regulated. Apical uptake via TRPM6 Mg2+ channels and basolateral Mg2+ extrusion via a putative Na+ -Mg2+ exchanger determines Mg2+ reabsorption in the Distal Convoluted Tubule. However, the mechanisms that explain the high incidence of hypomagnesaemia in patients with Na+ wasting disorders of the Distal Convoluted Tubule are largely unknown. In this review, we describe three potential mechanisms by which Mg2+ reabsorption in the Distal Convoluted Tubule is linked to Na+ reabsorption. First, decreased activity of the thiazide-sensitive Na+ /Cl- cotransporter (NCC) results in shortening of the segment, reducing the Mg2+ reabsorption capacity. Second, the activity of TRPM6 and NCC are determined by common regulatory pathways. Secondary effects of NCC dysregulation such as hormonal imbalance, therefore, might disturb TRPM6 expression. Third, the basolateral membrane potential, maintained by the K+ permeability and Na+ -K+ -ATPase activity, provides the driving force for Na+ and Mg2+ extrusion. Depolarisation of the basolateral membrane potential in Na+ wasting disorders of the Distal Convoluted Tubule may therefore lead to reduced activity of the putative Na+ -Mg2+ exchanger SLC41A1. Elucidating the interconnections between Mg2+ and Na+ transport in the Distal Convoluted Tubule is hampered by the currently available models. Our analysis indicates that the coupling of Na+ and Mg2+ reabsorption may be multifactorial and that advanced experimental models are required to study the molecular mechanisms.

  • Tubular flow activates magnesium transport in the Distal Convoluted Tubule.
    FASEB journal : official publication of the Federation of American Societies for Experimental Biology, 2018
    Co-Authors: Eric H. J. Verschuren, Joost G. J. Hoenderop, Dorien J.m. Peters, Francisco J. Arjona, Rene J M Bindels
    Abstract:

    Magnesium (Mg2+) is an important cofactor of many enzymes crucial for life; therefore, maintaining a Mg2+ balance in the body is essential. In the kidney, the Distal Convoluted Tubule (DCT) determines the final urinary Mg2+ excretion. The nephron is subjected to variable urinary flow, but little is known about the influence of flow on Mg2+ transport. Primary cilia, which are mechanosensory organelles that sense changes in flow, are expressed on tubular epithelial cells. This study aimed to elucidate whether urinary flow facilitates DCT Mg2+ transport. To this end, mouse DCT15 cells, with and without primary cilia, were exposed to physiologic fluid flow generating 0.3, 0.6, and 1.2 dyn/cm2 fluid shear stress (FSS). FSS stimulated Mg2+ uptake significantly. Net Mg2+ uptake ( i.e., the difference between static and FSS) followed a single component saturable first-order transport function and was independent of FSS magnitude and primary cilia. FSS did not affect the expression of magnesiotropic genes, including Cnnm2, Kcna1, Proegf, Trpm6, and Trpm7. Transient receptor potential cation channel subfamily melastatin (TRPM) member 7 (Trmp7) inhibition by 2-aminoethyl diphenyl borinate or knockout of TRPM6 did not alter net Mg2+ uptake, suggesting that TRPM6/TRPM7 homo/heterodimeric channels are not involved in FSS-activated Mg2+ transport. In summary, FSS generated by physiologic fluid flow is a new factor activating Mg2+ transport in DCT independent of primary cilia.-Verschuren, E. H. J., Hoenderop, J. G. J., Peters, D. J. M., Arjona, F. J., Bindels, R. J. M. Tubular flow activates magnesium transport in the Distal Convoluted Tubule.

  • Elucidation of the Distal Convoluted Tubule transcriptome identifies new candidate genes involved in renal Mg2+ handling
    American journal of physiology. Renal physiology, 2013
    Co-Authors: Jeroen H F De Baaij, Rene J M Bindels, Marian J. A. Groot Koerkamp, Marla Lavrijsen, Femke Van Zeeland, Hans Meijer, Frank C. P. Holstege, Joost G. J. Hoenderop
    Abstract:

    The kidney plays a key role in the maintenance of Mg2+ homeostasis. Specifically, the Distal Convoluted Tubule (DCT) is instrumental in the fine-tuning of renal Mg2+ handling. In recent years, here...

  • cisplatin induced injury of the renal Distal Convoluted Tubule is associated with hypomagnesaemia in mice
    Nephrology Dialysis Transplantation, 2013
    Co-Authors: Annelies A. Van Angelen, Joost G. J. Hoenderop, Bob Glaudemans, Annemiete W.c.m. Van Der Kemp, Rene J M Bindels
    Abstract:

    Background. Cisplatin is an effective anti-neoplastic drug, but its clinical use is limited due to dose-dependent nephrotoxicity. The majority of cisplatin-treated patients develop hypomagnesaemia, often associated with a reduced glomerular filtration rate (GFR), polyuria and other electrolyte disturbances. The aim of this study is to unravel the molecular mechanism responsible for these particular electrolyte disturbances. Methods. Two groups of 10 mice were injected intraperitoneally three times, once every 4 days, with cisplatin (5 mg/kg body weight,) or vehicle. Serum and urine electrolyte concentrations were determined. Next, renal mRNA levels of Distal Convoluted Tubule (DCT) genes epithelial Mg 2+ channel TRPM6, the Na + -Cl − cotransporter (NCC), and parvalbumin (PV), as well as marker genes for other tubular segments were measured by real-time qPCR. Subsequently, renal protein levels of NCC, PV, aquaporin 1 and aquaporin 2 were determined using immunoblotting and immunohistochemistry (IHC). Results. The cisplatin-treated mice developed significant polyuria (2.5± 0.3 and 0.9± 0.1 mL/24 h, cisplatin versus control, P < 0.05), reduced creatinine clearance rate (CCr) (0.18±0.02 and 0.26±0.02 mL/min, cisplatin versus control, P <0.05) and a substantially reduced serum level of Mg 2+ (1.23 ± 0.03 and 1.58 ± 0.03 mmol/L, cisplatin versus control, P<0.05), whereas serum Ca 2+ ,N a + and K + values were not altered. Measurements of 24 h urinary excretion demonstrated markedly increased Mg 2+ ,C a 2+ , Na + and K + levels in the cisplatin-treated group, whereas Pi levels were not changed. The mRNA levels of TRPM6, NCC and PV were significantly reduced in the cisplatin group. The expression levels of the marker genes for other tubular segments were unaltered, except for claudin-16, which was significantly up-regulated by the cisplatin treatment. The observed DCT-specific down-regulation was confirmed at the protein level. Conclusions. The present study identified the DCT as an important cisplatin-affected renal segment, explaining the high prevalence of hypomagnesaemia following treatment.

  • Cisplatin-induced injury of the renal Distal Convoluted Tubule is associated with hypomagnesaemia in mice
    Nephrology dialysis transplantation : official publication of the European Dialysis and Transplant Association - European Renal Association, 2012
    Co-Authors: Annelies A. Van Angelen, Joost G. J. Hoenderop, Bob Glaudemans, Annemiete W.c.m. Van Der Kemp, Rene J M Bindels
    Abstract:

    Background. Cisplatin is an effective anti-neoplastic drug, but its clinical use is limited due to dose-dependent nephrotoxicity. The majority of cisplatin-treated patients develop hypomagnesaemia, often associated with a reduced glomerular filtration rate (GFR), polyuria and other electrolyte disturbances. The aim of this study is to unravel the molecular mechanism responsible for these particular electrolyte disturbances. Methods. Two groups of 10 mice were injected intraperitoneally three times, once every 4 days, with cisplatin (5 mg/kg body weight,) or vehicle. Serum and urine electrolyte concentrations were determined. Next, renal mRNA levels of Distal Convoluted Tubule (DCT) genes epithelial Mg 2+ channel TRPM6, the Na + -Cl − cotransporter (NCC), and parvalbumin (PV), as well as marker genes for other tubular segments were measured by real-time qPCR. Subsequently, renal protein levels of NCC, PV, aquaporin 1 and aquaporin 2 were determined using immunoblotting and immunohistochemistry (IHC). Results. The cisplatin-treated mice developed significant polyuria (2.5± 0.3 and 0.9± 0.1 mL/24 h, cisplatin versus control, P < 0.05), reduced creatinine clearance rate (CCr) (0.18±0.02 and 0.26±0.02 mL/min, cisplatin versus control, P

Gordon Ritchie - One of the best experts on this subject based on the ideXlab platform.

  • Adenosine modulates Mg2+ uptake in Distal Convoluted Tubule cells via A1 and A2purinoceptors
    American journal of physiology. Renal physiology, 2001
    Co-Authors: Hyung Sub Kang, Long-jun Dai, Gordon Ritchie, Dirk Kerstan, Gary A. Quamme
    Abstract:

    tk;1Adenosine plays a role in the control of water and electrolyte reabsorption in the Distal Tubule. As the Distal Convoluted Tubule is important in the regulation of renal Mg2+ balance, we determ...

  • β-Adrenergic agonists stimulate Mg2+ uptake in mouse Distal Convoluted Tubule cells
    American journal of physiology. Renal physiology, 2000
    Co-Authors: Hyung Sub Kang, Long-jun Dai, Gordon Ritchie, Dirk Kerstan, Gary A. Quamme
    Abstract:

    β-Adrenergic agonists influence electrolyte reabsorption in the proximal Tubule, loop of Henle, and Distal Tubule. Although isoproterenol enhances magnesium absorption in the thick ascending limb, it is unclear what effect, if any, β-adrenergic agonists have on tubular magnesium handling. The effects of isoproterenol were studied in immortalized mouse Distal Convoluted Tubule (MDCT) cells by measuring cellular cAMP formation with radioimmunoassays and Mg2+ uptake with fluorescence techniques. Intracellular free Mg2+ concentration ([Mg2+]i) was measured in single MDCT cells by using microfluorescence with mag-fura-2. To assess Mg2+uptake, MDCT cells were first Mg2+ depleted to 0.22 ± 0.01 mM by culturing in Mg2+-free media for 16 h and then placed in 1.5 mM MgCl2, and the changes in [Mg2+]i were determined. [Mg2+]i returned to basal levels, 0.53 ± 0.02 mM, with a mean refill rate, d([Mg2+]i)/dt, of 168 ± 11 nM/s. Isoproterenol stimulated Mg2+ entry in a concentration-dependent manner, with a maximal respon...

  • aminoglycosides inhibit hormone stimulated mg2 uptake in mouse Distal Convoluted Tubule cells
    Canadian Journal of Physiology and Pharmacology, 2000
    Co-Authors: Hyung Sub Kang, Long-jun Dai, Gordon Ritchie, Dirk Kerstan, Gary A. Quamme
    Abstract:

    The clinical use of aminoglycosides often leads to renal magnesium wasting and hypomagnesemia. Of the nephron segments, both the thick ascending limb of Henle's loop and the Distal Tubule play significant roles in renal magnesium conservation but the Distal Convoluted Tubule exerts the final control of urinary excretion. An immortalized mouse Distal Convoluted Tubule (MDCT) cell line has been extensively used to study the cellular mechanisms of magnesium transport in this nephron segment. Peptide hormones, such as parathyroid hormone (PTH), glucagon, calcitonin, and arginine vasopressin (AVP) stimulate Mg2+ uptake in MDCT cells that is modulated by extracellular polyvalent cations, Ca2+ and Mg2+. The present studies determined the effect of aminoglycosides on parathyroid hormone (PTH)-mediated cAMP formation and Mg2+ uptake in MDCT cells. Gentamicin, a prototypic aminoglycoside, elicited transient increases in intracellular Ca2+ from basal levels of 102 +/- 13 nM to 713 +/- 125 nM, suggesting a receptor-mediated response. In order to determine Mg2+ transport, MDCT cells were Mg(2+)-depleted by culturing in Mg(2+)-free media for 16 h and Mg2+ uptake was measured by microfluorescence after placing the depleted cells in 1.0 mM MgCl2. The mean rate of Mg2+ uptake, d([Mg2+]i)/dt, was 138 +/- 24 nM/s in control MDCT cells. Gentamicin (50 microM) did not affect basal Mg2+ uptake (105 +/- 29 nM/s), but inhibited PTH stimulated Mg2+ entry, decreasing it from 257 +/- 36 nM/s to 108 +/- 42 nM/s. This was associated with diminished PTH-stimulated cAMP formation, from 80 +/- 2.5 to 23 +/- 1 pmol/mg protein x 5 min. Other aminoglycosides such as tobramycin, streptomycin, and neomycin also inhibited PTH-stimulated Mg2+ entry and cAMP formation. As these antibiotics are positively charged, the data suggest that aminoglycosides act through an extracellular polyvalent cation-sensing receptor present in Distal Convoluted Tubule cells. We infer from these studies that aminoglycosides inhibit hormone-stimulated Mg2+ absorption in the Distal Convoluted Tubule that may contribute to the renal magnesium wasting frequently observed with the clinical use of these antibiotics.

  • Aminoglycosides inhibit hormone-stimulated Mg 2+ uptake in mouse Distal Convoluted Tubule cells
    Canadian journal of physiology and pharmacology, 2000
    Co-Authors: Hyung Sub Kang, Long-jun Dai, Gordon Ritchie, Dirk Kerstan, Gary A. Quamme
    Abstract:

    The clinical use of aminoglycosides often leads to renal magnesium wasting and hypomagnesemia. Of the nephron segments, both the thick ascending limb of Henle's loop and the Distal Tubule play significant roles in renal magnesium conservation but the Distal Convoluted Tubule exerts the final control of urinary excretion. An immortalized mouse Distal Convoluted Tubule (MDCT) cell line has been extensively used to study the cellular mechanisms of magnesium transport in this nephron segment. Peptide hormones, such as parathyroid hormone (PTH), glucagon, calcitonin, and arginine vasopressin (AVP) stimulate Mg 2+ uptake in MDCT cells that is modulated by extracellular polyvalent cations, Ca 2+ and Mg 2+ . The present studies determined the effect of aminoglycosides on parathyroid hormone (PTH)-mediated cAMP formation and Mg 2+ uptake in MDCT cells. Gentamicin, a prototypic aminoglycoside, illicited transient increases in intracellular Ca 2+ from basal levels of 102 ± 13 nM to 713 ± 125 nM, suggesting a receptor-mediated response. In order to determine Mg 2+ transport, MDCT cells were Mg 2+ -depleted by culturing in Mg 2+ -free media for 16 h and Mg 2+ uptake was measured by microfluorescence after placing the depleted cells in 1.0 mM MgCl2. The mean rate of Mg 2+ uptake, d((Mg 2+ )i)/dt, was 138 ± 24 nM/s in control MDCT cells. Gentamicin (50 μM) did not affect basal Mg 2+ uptake (105 ± 29 nM/s), but inhibited PTH stimulated Mg 2+ entry, decreasing it from 257 ± 36 nM/s to 108 ± 42 nM/s. This was associated with diminished PTH-stimulated cAMP formation, from 80 ±2 .5 to 23 ±1p mol/mgprotein·5 min. Other aminoglycosides such as tobramycin, streptomycin, and neomycin also inhibited PTH-stimulated Mg 2+ entry and cAMP formation. As these antibiotics are positively charged, the data suggest that aminoglycosides act through an extracellular polyvalent cation-sensing receptor present in Distal Convoluted Tubule cells. We infer from these studies that aminoglycosides inhibit hormone-stimulated Mg 2+ absorption in the Distal Convoluted Tubule that may contribute to the renal magnesium wasting frequently observed with the clinical use of

  • b-Adrenergic agonists stimulate Mg 21 uptake in mouse Distal Convoluted Tubule cells
    2000
    Co-Authors: Hyung Sub Kang, Long-jun Dai, Gordon Ritchie, Dirk Kerstan, Gary A. Quamme
    Abstract:

    Kang, Hyung Sub, Dirk Kerstan, Long-Jun Dai, Gordon Ritchie, and Gary A. Quamme. b-Adrenergic agonists stimulate Mg uptake in mouse Distal Convoluted Tubule cells. Am J Physiol Renal Physiol 279: F1116–F1123, 2000.—b-Adrenergic agonists influence electrolyte reabsorption in the proximal Tubule, loop of Henle, and Distal Tubule. Although isoproterenol enhances magnesium absorption in the thick ascending limb, it is unclear what effect, if any, b-adrenergic agonists have on tubular magnesium handling. The effects of isoproterenol were studied in immortalized mouse Distal Convoluted Tubule (MDCT) cells by measuring cellular cAMP formation with radioimmunoassays and Mg uptake with fluorescence techniques. Intracellular free Mg concentration ([Mg]i) was measured in single MDCT cells by using microfluorescence with mag-fura-2. To assess Mg uptake, MDCT cells were first Mg depleted to 0.22 6 0.01 mM by culturing in Mg-free media for 16 h and then placed in 1.5 mM MgCl2, and the changes in [Mg ]i were determined. [Mg]i returned to basal levels, 0.53 6 0.02 mM, with a mean refill rate, d([Mg]i)/dt, of 168 6 11 nM/s. Isoproterenol stimulated Mg entry in a concentration-dependent manner, with a maximal response of 252 6 11 nM/s, at a concentration of 10 M, that represented a 50 6 7% increase in uptake rate above control values. This was associated with a sixfold increase in intracellular cAMP generation. Isoproterenol-stimulated Mg uptake was completely inhibited with RpcAMPS, a protein kinase A inhibitor, and U-73122, a phospholipase C inhibitor, and partially blocked by RO 31– 822, a protein kinase C inhibitor. Accordingly, isoproterenolmediated Mg entry rates involve multiple intracellular signaling pathways. Aldosterone potentiated isoproterenolstimulated Mg uptake (326 6 31 nM/s), whereas elevation of extracellular Ca inhibited isoproterenol-mediated cAMP accumulation and Mg uptake, 117 6 37 nM/s. These studies demonstrate that isoproterenol stimulates Mg uptake in a cell line of mouse Distal Convoluted Tubules that is modulated by hormonal and extracellular influences.