The Calcium-Sensing Receptor Increases Activity of the Renal NCC through the WNK4-SPAK Pathway

Abstract

Hypercalciuria can result from activation of the basolateral calcium-sensing receptor (CaSR), which in the thick ascending limb of Henle's loop controls Ca excretion and NaCl reabsorption in response to extracellular Ca However, the function of CaSR in the regulation of NaCl reabsorption in the distal convoluted tubule (DCT) is unknown. We hypothesized that CaSR in this location is involved in activating the thiazide-sensitive NaCl cotransporter (NCC) to prevent NaCl loss. We used a combination of and models to examine the effects of CaSR on NCC activity. Because the KLHL3-WNK4-SPAK pathway is involved in regulating NaCl reabsorption in the DCT, we assessed the involvement of this pathway as well. Thiazide-sensitive Na uptake assays in oocytes revealed that NCC activity increased in a WNK4-dependent manner upon activation of CaSR with Gd In HEK293 cells, treatment with the calcimimetic R-568 stimulated SPAK phosphorylation only in the presence of WNK4. The WNK4 inhibitor WNK463 also prevented this effect. Furthermore, CaSR activation in HEK293 cells led to phosphorylation of KLHL3 and WNK4 and increased WNK4 abundance and activity. Finally, acute oral administration of R-568 in mice led to the phosphorylation of NCC. Activation of CaSR can increase NCC activity the WNK4-SPAK pathway. It is possible that activation of CaSR by Ca in the apical membrane of the DCT increases NaCl reabsorption by NCC, with the consequent, well known decrease of Ca reabsorption, further promoting hypercalciuria.

WNK4 limits distal calcium losses following acute furosemide treatment

“…In the present study, we have demonstrated that knockout animals have exaggerated calcium wasting in response to acute furosemide treatment. Bazua‐Valenti et al recently proposed that increased calcium delivery to the DCT, as occurs following furosemide treatment, stimulates the apical calcium‐sensing receptor (CsR) to activate WNK4 and NCC, further promoting hypercalciuria (Bazúa‐Valenti et al ). However, we found increased hypercalciuria following acute furosemide administration in the absence of WNK4, which is the opposite of which this mechanism would indicate.…”

Calcium-Sensing Receptor and Regulation of WNK Kinases in the Kidney

“…In contrast to TAL, where CaSR signaling inhibits the NaCl reabsorption, activation of CaSR in DCT cells has been suggested to stimulate NCC in order to compensate for NaCl loss in the preceding TAL [13]. CaSR-induced NCC activation is mediated by WNK4, at least in cell culture and Xenopus laevis oocytes transfected with the two products [13].…”

“…In contrast to TAL, where CaSR signaling inhibits the NaCl reabsorption, activation of CaSR in DCT cells has been suggested to stimulate NCC in order to compensate for NaCl loss in the preceding TAL [13]. CaSR-induced NCC activation is mediated by WNK4, at least in cell culture and Xenopus laevis oocytes transfected with the two products [13]. In these ex vivo systems, CaSR activation has been shown to promote PKC-dependent phosphorylation of WNK4 and KLHL3 resulting in enhanced WNK4 kinase activity and reduced degradation of the kinase via the KLHL3-dependent pathway [13,91].…”

“…Via these mechanisms, CaSR modulates expression, surface abundance, and phosphorylation of transport proteins mediating calcium reabsorption in kidney epithelia [9][10][11]. Since calcium handling in the distal nephron is functionally coupled to sodium transport, CaSR signaling has been also implicated in the volume homeostasis and blood pressure control [12][13][14][15]. The distal nephron comprises the thick ascending limb (TAL), the distal convoluted tubule (DCT), and the connecting tubule (CNT).…”

“…The ensuing cortical collecting duct (CD) is functionally associated with the distal nephron, although it is derived from the ureteral bud like the other CD portions and does not belong to the nephron from the anatomical point of view [16][17][18]. There is growing evidence for CaSR-dependent regulation of the Na + -K + -2Cl − cotransporter (NKCC2) in TAL and the Na + -Cl − cotransporter (NCC) in DCT [12,13] (for review, see [19]). NKCC2 is critical to the urinary concentration and volume homeostasis [20,21].…”