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Increased expression of renal TRPM6 compensates for Mg(2+) wasting during furosemide treatment.

van Angelen AA, van der Kemp AW, Hoenderop JG, Bindels RJ - Clin Kidney J (2012)

Bottom Line: By diminishing sodium (Na(+)) reabsorption, loop diuretics reduce the lumen-positive transepithelial voltage and consequently diminish paracellular transport of magnesium (Mg(2+)) and calcium (Ca(2+)) in TAL.The present study shows specific renal upregulation of TRPM6, NCC, TRPV5 and calbindin-D28K.During chronic furosemide treatment, enhanced active reabsorption of Mg(2+) via the epithelial channel TRPM6 in DCT compensates for the reduced reabsorption of Mg(2+) in TAL.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology , Radboud University Nijmegen Medical Centre , Nijmegen , the Netherlands.

ABSTRACT

Background: Furosemide is a loop diuretic, which blocks the Na(+), K(+), 2Cl(-) cotransporter (NKCC2) in the thick ascending limb of Henle (TAL). By diminishing sodium (Na(+)) reabsorption, loop diuretics reduce the lumen-positive transepithelial voltage and consequently diminish paracellular transport of magnesium (Mg(2+)) and calcium (Ca(2+)) in TAL. Indeed, furosemide promotes urinary Mg(2+) excretion; however, it is unclear whether this leads, especially during prolonged treatment, to hypomagnesaemia. The aim of the present study was, therefore, to determine the effect of chronic furosemide application on renal Mg(2+) handling in mice.

Methods: Two groups of 10 mice received an osmotic minipump subcutaneously for 7 days with vehicle or 30 mg/kg/day furosemide. Serum and urine electrolyte concentrations were determined. Next, renal mRNA levels of the epithelial Mg(2+) channel (TRPM6), the Na(+), Cl(-) cotransporter (NCC), the epithelial Ca(2+) channel (TRPV5), the cytosolic Ca(2+)-binding protein calbindin-D28K, as well parvalbumin (PV), claudin-7 (CLDN7) and claudin-8 (CLDN8), the epithelial Na(+) channel (ENaC) and the Na(+)-H(+) exchanger 3 (NHE3) were determined by real-time quantitative polymerase chain reaction. Renal protein levels of NCC, TRPV5, calbindin-D28K and ENaC were also measured using semi-quantitative immunohistochemistry and immunoblotting.

Results: The mice chronically treated with 30 mg/kg/day furosemide displayed a significant polyuria (2.1 ± 0.3 and 1.3 ± 0.2 mL/24 h, furosemide versus control respectively, P < 0.05). Furosemide treatment resulted in increased serum concentrations of Na(+) [158 ± 3 (treated) and 147 ± 1 mmol/L (control), P < 0.01], whereas serum K(+), Ca(2+) and Mg(2+) values were not significantly altered in mice treated with furosemide. Urinary excretion of Na(+), K(+), Ca(2+) and Mg(2+) was not affected by chronic furosemide treatment. The present study shows specific renal upregulation of TRPM6, NCC, TRPV5 and calbindin-D28K.

Conclusions: During chronic furosemide treatment, enhanced active reabsorption of Mg(2+) via the epithelial channel TRPM6 in DCT compensates for the reduced reabsorption of Mg(2+) in TAL.

No MeSH data available.


Related in: MedlinePlus

Effect of chronic furosemide treatment on renal TRPM6 mRNA expression level. Real-time qPCR was used to determine the epithelial Mg2+ channel TRPM6 mRNA expression level in the kidneys of mice chronically treated with furosemide (30 mg/kg/day for 7 days). Expression levels are corrected for GAPDH and presented as relative percentage of expression in control mice. Values are presented as means ± SEM (n = 10). *P < 0.05 compared with control.
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SFS140F1: Effect of chronic furosemide treatment on renal TRPM6 mRNA expression level. Real-time qPCR was used to determine the epithelial Mg2+ channel TRPM6 mRNA expression level in the kidneys of mice chronically treated with furosemide (30 mg/kg/day for 7 days). Expression levels are corrected for GAPDH and presented as relative percentage of expression in control mice. Values are presented as means ± SEM (n = 10). *P < 0.05 compared with control.

Mentions: The effect of furosemide treatment on the renal expression level of TRPM6 was determined by real-time quantitative PCR (qPCR). A significant increase in renal TRPM6 mRNA expression was observed in mice chronically treated with furosemide (129 ± 9 and 100 ± 6%, furosemide versus control, P < 0.05) (Figure 1). Next, the renal mRNA expression level and protein abundance of the thiazide-sensitive NCC were determined. Chronic furosemide treatment had no effect on mRNA expression of NCC (98 ± 4 and 100 ± 5%, furosemide versus control, P > 0.2) (Figure 2A). The renal protein abundance of NCC was examined by IHC. In order to semi-quantify the protein expression, the amount of immunopositive tubules in the total kidney cortex was determined for each experimental group. The averaged values of the furosemide-treated group are presented as relative percentage of expression in control mice. In contrast with the mRNA level, the protein abundance of NCC was substantially increased (176 ± 9 and 100 ± 8%, furosemide versus control, P < 0.01) (Figure 2B).Fig. 1.


Increased expression of renal TRPM6 compensates for Mg(2+) wasting during furosemide treatment.

van Angelen AA, van der Kemp AW, Hoenderop JG, Bindels RJ - Clin Kidney J (2012)

Effect of chronic furosemide treatment on renal TRPM6 mRNA expression level. Real-time qPCR was used to determine the epithelial Mg2+ channel TRPM6 mRNA expression level in the kidneys of mice chronically treated with furosemide (30 mg/kg/day for 7 days). Expression levels are corrected for GAPDH and presented as relative percentage of expression in control mice. Values are presented as means ± SEM (n = 10). *P < 0.05 compared with control.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4400563&req=5

SFS140F1: Effect of chronic furosemide treatment on renal TRPM6 mRNA expression level. Real-time qPCR was used to determine the epithelial Mg2+ channel TRPM6 mRNA expression level in the kidneys of mice chronically treated with furosemide (30 mg/kg/day for 7 days). Expression levels are corrected for GAPDH and presented as relative percentage of expression in control mice. Values are presented as means ± SEM (n = 10). *P < 0.05 compared with control.
Mentions: The effect of furosemide treatment on the renal expression level of TRPM6 was determined by real-time quantitative PCR (qPCR). A significant increase in renal TRPM6 mRNA expression was observed in mice chronically treated with furosemide (129 ± 9 and 100 ± 6%, furosemide versus control, P < 0.05) (Figure 1). Next, the renal mRNA expression level and protein abundance of the thiazide-sensitive NCC were determined. Chronic furosemide treatment had no effect on mRNA expression of NCC (98 ± 4 and 100 ± 5%, furosemide versus control, P > 0.2) (Figure 2A). The renal protein abundance of NCC was examined by IHC. In order to semi-quantify the protein expression, the amount of immunopositive tubules in the total kidney cortex was determined for each experimental group. The averaged values of the furosemide-treated group are presented as relative percentage of expression in control mice. In contrast with the mRNA level, the protein abundance of NCC was substantially increased (176 ± 9 and 100 ± 8%, furosemide versus control, P < 0.01) (Figure 2B).Fig. 1.

Bottom Line: By diminishing sodium (Na(+)) reabsorption, loop diuretics reduce the lumen-positive transepithelial voltage and consequently diminish paracellular transport of magnesium (Mg(2+)) and calcium (Ca(2+)) in TAL.The present study shows specific renal upregulation of TRPM6, NCC, TRPV5 and calbindin-D28K.During chronic furosemide treatment, enhanced active reabsorption of Mg(2+) via the epithelial channel TRPM6 in DCT compensates for the reduced reabsorption of Mg(2+) in TAL.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology , Radboud University Nijmegen Medical Centre , Nijmegen , the Netherlands.

ABSTRACT

Background: Furosemide is a loop diuretic, which blocks the Na(+), K(+), 2Cl(-) cotransporter (NKCC2) in the thick ascending limb of Henle (TAL). By diminishing sodium (Na(+)) reabsorption, loop diuretics reduce the lumen-positive transepithelial voltage and consequently diminish paracellular transport of magnesium (Mg(2+)) and calcium (Ca(2+)) in TAL. Indeed, furosemide promotes urinary Mg(2+) excretion; however, it is unclear whether this leads, especially during prolonged treatment, to hypomagnesaemia. The aim of the present study was, therefore, to determine the effect of chronic furosemide application on renal Mg(2+) handling in mice.

Methods: Two groups of 10 mice received an osmotic minipump subcutaneously for 7 days with vehicle or 30 mg/kg/day furosemide. Serum and urine electrolyte concentrations were determined. Next, renal mRNA levels of the epithelial Mg(2+) channel (TRPM6), the Na(+), Cl(-) cotransporter (NCC), the epithelial Ca(2+) channel (TRPV5), the cytosolic Ca(2+)-binding protein calbindin-D28K, as well parvalbumin (PV), claudin-7 (CLDN7) and claudin-8 (CLDN8), the epithelial Na(+) channel (ENaC) and the Na(+)-H(+) exchanger 3 (NHE3) were determined by real-time quantitative polymerase chain reaction. Renal protein levels of NCC, TRPV5, calbindin-D28K and ENaC were also measured using semi-quantitative immunohistochemistry and immunoblotting.

Results: The mice chronically treated with 30 mg/kg/day furosemide displayed a significant polyuria (2.1 ± 0.3 and 1.3 ± 0.2 mL/24 h, furosemide versus control respectively, P < 0.05). Furosemide treatment resulted in increased serum concentrations of Na(+) [158 ± 3 (treated) and 147 ± 1 mmol/L (control), P < 0.01], whereas serum K(+), Ca(2+) and Mg(2+) values were not significantly altered in mice treated with furosemide. Urinary excretion of Na(+), K(+), Ca(2+) and Mg(2+) was not affected by chronic furosemide treatment. The present study shows specific renal upregulation of TRPM6, NCC, TRPV5 and calbindin-D28K.

Conclusions: During chronic furosemide treatment, enhanced active reabsorption of Mg(2+) via the epithelial channel TRPM6 in DCT compensates for the reduced reabsorption of Mg(2+) in TAL.

No MeSH data available.


Related in: MedlinePlus