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Kidney Week

Abstract: PO1359

TRPV4 Calcium Channel Activity Is Increased by With-No-Lysine Kinase 1 in the Collecting Duct Cells

Session Information

Category: Genetic Diseases of the Kidneys

  • 1002 Genetic Diseases of the Kidneys: Non-Cystic

Authors

  • Tomilin, Victor N., The University of Texas Health Science Center at Houston John P and Katherine G McGovern Medical School, Houston, Texas, United States
  • Pyrshev, Kyrylo, The University of Texas Health Science Center at Houston John P and Katherine G McGovern Medical School, Houston, Texas, United States
  • Hassanzadeh Khayyat, Naghmeh, The University of Texas Health Science Center at Houston John P and Katherine G McGovern Medical School, Houston, Texas, United States
  • Zaika, Oleg L., The University of Texas Health Science Center at Houston John P and Katherine G McGovern Medical School, Houston, Texas, United States
  • Pochynyuk, Oleh, The University of Texas Health Science Center at Houston John P and Katherine G McGovern Medical School, Houston, Texas, United States
Background

Kidneys play a central role in regulation of potassium homeostasis and maintaining plasma K+ levels within a narrow physiological range. Dietary K+ load increases circulating levels of the mineralocorticoid aldosterone leading to kaliuresis via stimulation calcium-activated large conductive maxi-K+ (BK) channel dependent K+ secretion in the collecting duct cells. WNK1 and WNK4 (With-no-lysine) kinases, have been recognized to regulate K+ balance, in part, by orchestrating BK-dependent K+ secretion in the ASDN (aldosterone sensitive renal nephron). Ca2+-permeable TRPV4 channel is essential for BK activation in the distal nephron, as we have recently demonstrated. Also of note, high K+ diet increases TRPV4 activity and expression largely in an aldosterone-dependent manner.

Methods

Patch-Clamp; [Ca2+]i imaging; Western blotting;

Results

In the current study, we aimed to test whether WNK1/4 contribute to regulation of TRPV4 by aldosterone. First we treatment of mpkCCDc14 cells with 1 µM aldosterone for 24 h it as expected, increased TRPV4-dependent Ca2+ influx approximately 2 fold. Inhibition of WNK1/4 with the pan-blocker of WNK, WNK463 (100 nM for 24h) decreased basal TRPV4 activity by approximately 30% and virtually abolished stimulation of TRPV4 by aldosterone. Similarly, WNK1 blockade with WNK-in-11 (400 nM, 24 h) produced comparable inhibitory effects on the basal and aldosterone-dependent TRPV4 activity as WNK463. Western blots performed on apical plasma membrane fraction from the same cells showed a complete block of the stimulatory effect of aldosterone after treatment with WNK-in-11 (400 nM, 24 h). Co-expression of TRPV4 and WNK1 into Chinese hamster ovary (CHO) cells increased the macroscopic TRPV4-dependent cation currents from 160±10 pA/pF to 250±22 pA/pF. In contrast, overexpression of TRPV4 with a dominant negative WNK1 variant (K233M) decreased the whole cell currents to 60±5 pA/pF suggesting both stimulatory and permissive roles of WNK1 in regulation of TRPV4.

Conclusion

Overall, we show that WNK1 is essential in controling of basal TRPV4 activity and its activation by aldosterone in collecting duct cells. We propose that this new mechanism is likely contributing to regulation of urinary K+ levels to maintain systemic homeostasis.

Funding

  • Other NIH Support