Sodium Channels Transient Receptor Potential Vanilloid 1 Activation by Dietary Capsaicin Promotes Urinary Sodium Excretion by Inhibiting Epithelial Sodium Channel α Subunit–Mediated Sodium Reabsorption Li Li,* Fei Wang,* Xing Wei, Yi Liang, Yuanting Cui, Feng Gao, Jian Zhong, Yunfei Pu, Yu Zhao, Zhencheng Yan, William J. Arendshorst, Bernd Nilius, Jing Chen, Daoyan Liu, Zhiming Zhu Abstract—High salt (HS) intake contributes to the development of hypertension. Epithelial sodium channels play crucial roles in regulating renal sodium reabsorption and blood pressure. The renal transient receptor potential vanilloid 1 (TRPV1) cation channel can be activated by its agonist capsaicin. However, it is unknown whether dietary factors can act on urinary sodium excretion and renal epithelial sodium channel (ENaC) function. Here, we report that TRPV1 activation by dietary capsaicin increased urinary sodium excretion through reducing sodium reabsorption in wild-type (WT) mice on a HS diet but not in TRPV1–/– mice. The effect of capsaicin on urinary sodium excretion was involved in inhibiting αENaC and its related with-no-lysine kinase 1/serum- and glucocorticoid-inducible protein kinase 1 pathway in renal cortical collecting ducts of WT mice. Dietary capsaicin further reduced the increased αENaC activity in WT mice attributed to the HS diet. In contrast, this capsaicin effect was absent in TRPV1–/– mice. Immunoprecipitation study indicated αENaC specifically coexpressed and functionally interact with TRPV1 in renal cortical collecting ducts of WT mice. Additionally, ENaC activity and expression were suppressed by capsaicin-mediated TRPV1 activation in cultured M1-cortical collecting duct cells. Long-term dietary capsaicin prevented the development of high blood pressure in WT mice on a HS diet. It concludes that TRPV1 activation in the cortical collecting ducts by capsaicin increases urinary sodium excretion and avoids HS diet–induced hypertension through antagonizing αENaC-mediated urinary sodium reabsorption. Dietary capsaicin may represent a promising lifestyle intervention in populations exposed to a high dietary salt intake. (Hypertension. 2014;64:397-404.) Online Data Supplement
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Key Words: capsaicin
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ENaC alpha
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hypertension ■ kidney collecting duct potential cation channel V1
T
he pathogenesis of hypertension is caused by both genetic susceptibility and environmental risk factors.1 One of major environmental factors for hypertension is high-sodium or lowpotassium dietary intake.2 Maintenance of a constant intravascular fluid volume and blood pressure depends on the kidneys’ ability to regulate the urinary sodium excretion (UNaV).3 Several renal sodium transporters, such as the thiazide-sensitive NaCl cotransporter (NCC) and the amiloride-sensitive epithelial sodium channel (ENaC), play crucial roles in regulating renal sodium reabsorption and blood pressure.4 Multigene kinases, including Ste20-related proline–alanine–rich kinase (SPAK), with-no-lysine kinase (WNK) 4 and 1, oxidative stress response kinase 1 (OSR1), and serum- and glucocorticoid-inducible
■
sodium diet
■
transient receptor
protein kinase 1 (SGK1), regulate renal electrolyte transport. Abnormal signaling pathways attributable to aberrations of these kinases can result in renal sodium retention and hypertension.5 Currently, strategies to prevent the development of hypertension include avoidance of a high-salt (HS) diet and increased vegetables intake in general population. In addition, thiazide diuretics are commonly used to treat patients with hypertension through inhibiting renal sodium reabsorption in the distal convoluted tubule, thereby increasing the UNaV and reducing extracellular fluid volume.6 However, nondrug intervention– mediated UNaV is scarcely studied. The transient receptor potential vanilloid 1 (TRPV1) cation channel is a polymodal nonselective cation channel that can
Received January 2, 2014; first decision January 23, 2014; revision accepted April 27, 2014. From the Department of Hypertension and Endocrinology, Center for Hypertension and Metabolic Diseases, Daping Hospital, Third Military Medical University, Chongqing Institute of Hypertension, Chongqing, China (L.L., F.W., X.W., Y.L., Y.C., F.G., J.Z., Y.P., Y.Z., Z.Y., J.C., D.L., Z.Z.); Department of Cell Biology and Physiology, School of Medicine, University of North Carolina at Chapel Hill (W.J.A.); and Department of Cell Molecular Medicine, Laboratory Ion Channel Research, Campus Gasthuisberg, Leuven, Belgium (B.N.). *These authors contributed equally to this work. The online-only Data Supplement is available with this article at http://hyper.ahajournals.org/lookup/suppl/doi:10.1161/HYPERTENSIONAHA. 114.03105/-/DC1. Correspondence to Zhiming Zhu or Daoyan Liu, Department of Hypertension and Endocrinology, Center for Hypertension and Metabolic Diseases, Daping Hospital, Third Military Medical University, Chongqing Institute of Hypertension, Changjiang Zhilu, No. 10, Chongqing 400042, China. E-mail [email protected] or [email protected] © 2014 American Heart Association, Inc. Hypertension is available at http://hyper.ahajournals.org
DOI: 10.1161/HYPERTENSIONAHA.114.03105
Downloaded from http://hyper.ahajournals.org/ at 397 CONS CALIFORNIA DIG LIB on February 24, 2015
398 Hypertension August 2014 be specifically activated by heat or capsaicin, a major pungent ingredient in hot peppers.7 We recently reported that TRPV1 activation by chronic dietary capsaicin improves endothelial function and lowers blood pressure in genetically hypertensive rats.8 Several studies showed that TRPV1 is expressed in the kidneys and is involved in renal sodium handling.9 Li and Wang9 showed that NCC but not ENaC was functionally upregulated in the kidneys of Wistar rats subjected to acute capsaicin-sensitive sensory nerve degeneration plus HS intake. It is unknown how chronic dietary capsaicin affects renal sodium excretion and blood pressure during long-term HS diet. In this study, we hypothesized that TRPV1 activation by dietary capsaicin contributes to renal benefits through inhibiting sodium transport. Thus, we examined this hypothesis in vitro and in vivo.
Materials and Methods Detailed methods and reagents used can be found in the online-only Data Supplement. Culture of the M1 cortical collecting duct (CCD) cell line, intracellular free calcium and sodium measurement, immunoprecipitation, immunoblotting analysis, blood pressure and urinay sodium excretion measurement, and standardized techniques were performed as described previously. All mice (C57BL/6 wild-type [WT] mice and TRPV1–/– mice) were purchased from the Jackson Laboratory (Bar Harbor, Maine). Procedures were performed in accordance with protocols approved by the Institutional Animal Care and Research Advisory Committee. The data are expressed as the mean±SEM from 5 to 8 independent experiments or mice. Comparisons between groups were analyzed using Kruskal–Wallis tests followed by the Mann–Whitney U test for multiple comparisons as appropriate with the Bonferroni correction (SPSS15.0 software; SPSS Inc, Chicago, IL). For all analyses, 2-sided P values
•
Key Words: capsaicin
■
ENaC alpha
■
hypertension ■ kidney collecting duct potential cation channel V1
T
he pathogenesis of hypertension is caused by both genetic susceptibility and environmental risk factors.1 One of major environmental factors for hypertension is high-sodium or lowpotassium dietary intake.2 Maintenance of a constant intravascular fluid volume and blood pressure depends on the kidneys’ ability to regulate the urinary sodium excretion (UNaV).3 Several renal sodium transporters, such as the thiazide-sensitive NaCl cotransporter (NCC) and the amiloride-sensitive epithelial sodium channel (ENaC), play crucial roles in regulating renal sodium reabsorption and blood pressure.4 Multigene kinases, including Ste20-related proline–alanine–rich kinase (SPAK), with-no-lysine kinase (WNK) 4 and 1, oxidative stress response kinase 1 (OSR1), and serum- and glucocorticoid-inducible
■
sodium diet
■
transient receptor
protein kinase 1 (SGK1), regulate renal electrolyte transport. Abnormal signaling pathways attributable to aberrations of these kinases can result in renal sodium retention and hypertension.5 Currently, strategies to prevent the development of hypertension include avoidance of a high-salt (HS) diet and increased vegetables intake in general population. In addition, thiazide diuretics are commonly used to treat patients with hypertension through inhibiting renal sodium reabsorption in the distal convoluted tubule, thereby increasing the UNaV and reducing extracellular fluid volume.6 However, nondrug intervention– mediated UNaV is scarcely studied. The transient receptor potential vanilloid 1 (TRPV1) cation channel is a polymodal nonselective cation channel that can
Received January 2, 2014; first decision January 23, 2014; revision accepted April 27, 2014. From the Department of Hypertension and Endocrinology, Center for Hypertension and Metabolic Diseases, Daping Hospital, Third Military Medical University, Chongqing Institute of Hypertension, Chongqing, China (L.L., F.W., X.W., Y.L., Y.C., F.G., J.Z., Y.P., Y.Z., Z.Y., J.C., D.L., Z.Z.); Department of Cell Biology and Physiology, School of Medicine, University of North Carolina at Chapel Hill (W.J.A.); and Department of Cell Molecular Medicine, Laboratory Ion Channel Research, Campus Gasthuisberg, Leuven, Belgium (B.N.). *These authors contributed equally to this work. The online-only Data Supplement is available with this article at http://hyper.ahajournals.org/lookup/suppl/doi:10.1161/HYPERTENSIONAHA. 114.03105/-/DC1. Correspondence to Zhiming Zhu or Daoyan Liu, Department of Hypertension and Endocrinology, Center for Hypertension and Metabolic Diseases, Daping Hospital, Third Military Medical University, Chongqing Institute of Hypertension, Changjiang Zhilu, No. 10, Chongqing 400042, China. E-mail [email protected] or [email protected] © 2014 American Heart Association, Inc. Hypertension is available at http://hyper.ahajournals.org
DOI: 10.1161/HYPERTENSIONAHA.114.03105
Downloaded from http://hyper.ahajournals.org/ at 397 CONS CALIFORNIA DIG LIB on February 24, 2015
398 Hypertension August 2014 be specifically activated by heat or capsaicin, a major pungent ingredient in hot peppers.7 We recently reported that TRPV1 activation by chronic dietary capsaicin improves endothelial function and lowers blood pressure in genetically hypertensive rats.8 Several studies showed that TRPV1 is expressed in the kidneys and is involved in renal sodium handling.9 Li and Wang9 showed that NCC but not ENaC was functionally upregulated in the kidneys of Wistar rats subjected to acute capsaicin-sensitive sensory nerve degeneration plus HS intake. It is unknown how chronic dietary capsaicin affects renal sodium excretion and blood pressure during long-term HS diet. In this study, we hypothesized that TRPV1 activation by dietary capsaicin contributes to renal benefits through inhibiting sodium transport. Thus, we examined this hypothesis in vitro and in vivo.
Materials and Methods Detailed methods and reagents used can be found in the online-only Data Supplement. Culture of the M1 cortical collecting duct (CCD) cell line, intracellular free calcium and sodium measurement, immunoprecipitation, immunoblotting analysis, blood pressure and urinay sodium excretion measurement, and standardized techniques were performed as described previously. All mice (C57BL/6 wild-type [WT] mice and TRPV1–/– mice) were purchased from the Jackson Laboratory (Bar Harbor, Maine). Procedures were performed in accordance with protocols approved by the Institutional Animal Care and Research Advisory Committee. The data are expressed as the mean±SEM from 5 to 8 independent experiments or mice. Comparisons between groups were analyzed using Kruskal–Wallis tests followed by the Mann–Whitney U test for multiple comparisons as appropriate with the Bonferroni correction (SPSS15.0 software; SPSS Inc, Chicago, IL). For all analyses, 2-sided P values
