The inositol Inpp5k 5-phosphatase affects osmoregulation through the vasopressin-aquaporin 2 pathway in the collecting system

Inositol Inpp5k (or Pps, SKIP) is a member of the inositol polyphosphate 5-phosphatases family with a poorly characterized function in vivo. In this study, we explored the function of this inositol 5-phosphatase in mice and cells overexpressing the 42-kDa mouse Inpp5k protein. Inpp5k transgenic mice present defects in water metabolism characterized by a reduced plasma osmolality at baseline, a delayed urinary water excretion following a water load, and an increased acute response to vasopressin. These defects are associated with the expression of the Inpp5k transgene in renal collecting ducts and with alterations in the arginine vasopressin/aquaporin-2 signalling pathway in this tubular segment. Analysis in a mouse collecting duct mCCD cell line revealed that Inpp5k overexpression leads to increased expression of the arginine vasopressin receptor type 2 and increased cAMP response to arginine vasopressin, providing a basis for increased aquaporin-2 expression and plasma membrane localization with increased osmotically induced water transport. Altogether, our results indicate that Inpp5k 5-phosphatase is important for the control of the arginine vasopressin/aquaporin-2 signalling pathway and water transport in kidney collecting duct

Mice lacking TRPV4 show abnormal thirst regulation in absence of renal defects

TRPV4 is a polymodal cation channel of the transient receptor potential (TRP) family. It is expressed in hypothalamic neurons including the subfornical organ, median preoptic area, and the organum vasculosum of the lamina terminalis, where it could play a role in osmotic sensing. TRPV4 has also been evidenced in the thick ascending limb (TAL) of the rat nephron, where its role remains unclear. To further analyze the role of TRPV4 in osmoregulation, we investigated its distribution pattern and the functional consequences of its disruption in mouse. Using qPCR on microdissected segments and immunohistochemistry, we found that TRPV4 is abundantly expressed in the proximal tubules (apical and basolateral), late distal convoluted tubules and throughout the connecting tubules and collecting ducts (basolateral), including principal and intercalated cells. By contrast, TRPV4 was undetectable in glomeruli and TAL, and weakly abundant in early DCT. Metabolic studies in 12 weeks old TRPV4 knock-out (KO) and wild-type (WT) mice revealed that disruption of TRPV4 does not influence renal function and urinary concentration at baseline. The KO mice did not exhibit anomalies in activity, rearing, food and water intake. Moreover, both WT and KO mice showed a similar renal NaCl and water excretion in response to furosemide injection, acute water loading (100 l/g BW i.p.), overnight water deprivation, and chronic dietary NaCl restriction (17 days, 0.01% NaCl),. However, acute administration of hypertonic saline solution (0.5 M NaCl) resulted in a significantly lower water intake in the TRPV4 KO vs WT mice, suggesting a central defect in osmoregulation. These results demonstrate that TRPV4 is widely expressed in the mouse kidney, but not in the TAL. Disruption of TRPV4 is not reflected by significant alterations in the renal handling of NaCl and water. However, TRPV4 KO mice show a decreased thirst response to hypertonic NaCl, which substantiates the major role of TRPV4 in the central regulation of osmolality

P2Y2 receptor activation inhibits the expression of the sodium-chloride cotransporter NCC in distal convoluted tubule cells

Luminal nucleotide stimulation is known to reduce Na+ transport in the distal nephron. Previous studies suggest that this mechanism may involve the thiazide-sensitive Na+-Cl- cotransporter (NCC), which plays an essential role in NaCl reabsorption in the cells lining the distal convoluted tubule (DCT). Here we show that stimulation of mouse DCT (mDCT) cells with ATP or UTP promoted Ca2+ transients and decreased the expression of NCC at both mRNA and protein levels. Specific siRNA-mediated silencing of P2Y2 receptors almost completely abolished ATP/UTP-induced Ca2+ transients and significantly reduced ATP/UTP-induced decrease of NCC expression. To test whether local variations in the intracellular Ca2+ concentration ([Ca2+]i) may control NCC transcription, we overexpressed the Ca2+-binding protein parvalbumin selectively in the cytosol or in the nucleus of mDCT cells. The decrease in NCC mRNA upon nucleotide stimulation was abolished in cells overexpressing cytosolic PV but not in cells overexpressing either a nuclear-targeted PV or a mutated PV unable to bind Ca2+. Using a firefly luciferase reporter gene strategy, we observed that the activity of NCC promoter region from -1 to -2,200 bp was not regulated by changes in [Ca2+]i. In contrast, high cytosolic calcium level induced instability of NCC mRNA. We conclude that in mDCT cells: (1) P2Y2 receptor is essential for the intracellular Ca2+ signaling induced by ATP/UTP stimulation; (2) P2Y2-mediated increase of cytoplasmic Ca2+ concentration down-regulates the expression of NCC; (3) the decrease of NCC expression occurs, at least in part, via destabilization of its mRNA

A transgenic mouse model for uromodulin-associated kidney diseases shows specific tubulo-interstitial damage, urinary concentrating defect and renal failure

Uromodulin-associated kidney diseases (UAKD) are autosomal-dominant disorders characterized by alteration of urinary concentrating ability, tubulo-interstitial fibrosis, hyperuricaemia and renal cysts at the cortico-medullary junction. UAKD are caused by mutations in UMOD, the gene encoding uromodulin. Although uromodulin is the most abundant protein secreted in urine, its physiological role remains elusive. Several in vitro studies demonstrated that mutations in uromodulin lead to endoplasmic reticulum (ER) retention of mutant protein, but their relevance in vivo has not been studied. We here report on the generation and characterization of the first transgenic mouse model for UAKD. Transgenic mice that express the C147W mutant uromodulin (Tg(UmodC147W)), corresponding to the well-established patient mutation C148W, were compared with expression-matched transgenic mice expressing the wild-type protein (Tg(Umodwt)). Tg(UmodC147W) mice recapitulate most of the UAKD features, with urinary concentrating defect of renal origin and progressive renal injury, i.e. tubulo-interstitial fibrosis with inflammatory cell infiltration, tubule dilation and specific damage of the thick ascending limb of Henle's loop, leading to mild renal failure. As observed in patients, Tg(UmodC147W) mice show a marked reduction of urinary uromodulin excretion. Mutant uromodulin trafficking to the plasma membrane is indeed impaired as it is retained in the ER of expressing cells leading to ER hyperplasia. The Tg(UmodC147W) mice represent a unique model that recapitulates most of the features associated with UAKD. Our data clearly demonstrate a gain-of-toxic function of uromodulin mutations providing insights into the pathogenetic mechanism of the disease. These findings may also be relevant for other tubulo-interstitial or ER-storage disorders

TRPV4 is associated with central rather than nephrogenic osmoregulation.

TRPV4 is a polymodal cation channel expressed in osmosensitive neurons of the hypothalamus and in the mammalian nephron. The segmental distribution and role(s) of TRPV4 in osmoregulation remain debated. We investigated the renal distribution pattern of TRPV4 and the functional consequences of its disruption in mouse models. Using qPCR on microdissected segments, immunohistochemistry, and a LacZ reporter mouse, we found that TRPV4 is abundantly expressed in the proximal tubule, the late distal convoluted tubule, and throughout the connecting tubule and collecting duct, including principal and intercalated cells. TRPV4 was undetectable in the glomeruli and thick ascending limb and weakly abundant in the early distal convoluted tubule. Metabolic studies in Trpv4 (+/+) and Trpv4 (-/-) littermates revealed that the lack of TRPV4 did not influence activity, food and water intake, renal function, and urinary concentration at baseline. The mice showed a similar response to furosemide, water loading and deprivation, acid loading, and dietary NaCl restriction. However, Trpv4 (-/-) mice showed a significantly lower vasopressin synthesis and release after water deprivation, with a loss of the positive correlation between plasma osmolality and plasma vasopressin levels, and a delayed water intake upon acute administration of hypertonic saline. Specific activation of TRPV4 in primary cultures of proximal tubule cells increased albumin uptake, whereas no effect of TRPV4 deletion could be observed at baseline. These data reveal that, despite its abundant expression in tubular segments, TRPV4 does not play a major role in the kidney or is efficiently compensated when deleted. Instead, TRPV4 is critical for the release of vasopressin, the sensation of thirst, and the central osmoregulation

TRPV4 is associated with central rather than nephrogenic osmoregulation

TRPV4 is a polymodal cation channel expressed in osmosensitive neurons of the hypothalamus and in the mammalian nephron. The segmental distribution and role(s) of TRPV4 in osmoregulation remain debated. We investigated the renal distribution pattern of TRPV4 and the functional consequences of its disruption in mouse models. Using qPCR on microdissected segments, immunohistochemistry, and a LacZ reporter mouse, we found that TRPV4 is abundantly expressed in the proximal tubule, the late distal convoluted tubule, and throughout the connecting tubule and collecting duct, including principal and intercalated cells. TRPV4 was undetectable in the glomeruli and thick ascending limb and weakly abundant in the early distal convoluted tubule. Metabolic studies in Trpv4 (+/+) and Trpv4 (-/-) littermates revealed that the lack of TRPV4 did not influence activity, food and water intake, renal function, and urinary concentration at baseline. The mice showed a similar response to furosemide, water loading and deprivation, acid loading, and dietary NaCl restriction. However, Trpv4 (-/-) mice showed a significantly lower vasopressin synthesis and release after water deprivation, with a loss of the positive correlation between plasma osmolality and plasma vasopressin levels, and a delayed water intake upon acute administration of hypertonic saline. Specific activation of TRPV4 in primary cultures of proximal tubule cells increased albumin uptake, whereas no effect of TRPV4 deletion could be observed at baseline. These data reveal that, despite its abundant expression in tubular segments, TRPV4 does not play a major role in the kidney or is efficiently compensated when deleted. Instead, TRPV4 is critical for the release of vasopressin, the sensation of thirst, and the central osmoregulation.status: publishe

Dietary behavior: Developing a taxonomy diet, eating and nutrition

Dietary behavior: Developing a taxonomy diet, eating and nutrition. Final dedipac symposiu