Mechanoregulation of intracellular Ca²⁺concentration is attenuated in collecting duct of monocilium-impairedorpkmice

Abstract

Autosomal recessive polycystic kidney disease (ARPKD) is characterized by the progressive dilatation of collecting ducts, the nephron segments responsible for the final renal regulation of sodium, potassium, acid-base, and water balance. Murine models of ARPKD possess mutations in genes encoding cilia-associated proteins, including Tg737 in orpk mice. New findings implicate defects in structure/function of primary cilia as central to the development of polycystic kidney disease. Our group (Liu W, Xu S, Woda C, Kim P, Weinbaum S, and Satlin LM, Am J Physiol Renal Physiol 285: F998-F1012, 2003) recently reported that increases in luminal flow rate in rabbit collecting ducts increase intracellular Ca(2+) concentration ([Ca(2+)](i)) in cells therein. We thus hypothesized that fluid shear acting on the apical membrane or hydrodynamic bending moments acting on the cilium increase renal epithelial [Ca(2+)](i). To further explore this, we tested whether flow-induced [Ca(2+)](i) transients in collecting ducts from mutant orpk mice, which possess structurally abnormal cilia, differ from those in controls. Isolated segments from 1- and 2-wk-old mice were microperfused in vitro and loaded with fura 2; [Ca(2+)](i) was measured by digital ratio fluorometry before and after the rate of luminal flow was increased. All collecting ducts responded to an increase in flow with an increase in [Ca(2+)](i), a response that appeared to be dependent on luminal Ca(2+) entry. However, the magnitude of the increase in [Ca(2+)](i) in 2- but not 1-wk-old mutant orpk animals was blunted. We speculate that this defect in mechano-induced Ca(2+) signaling in orpk mice leads to aberrant structure and function of the collecting duct in ARPKD.

Role of Primary Cilia in the Pathogenesis of Polycystic Kidney Disease

“…Furthermore, Nauli et al (53) revealed that this flow response required the function of the polycystins, which localize in the cilium. Similar defects were reported by Liu et al (54) in perfused tubules that were isolated from Tg737 orpk mutants. Although calcium is a widely known second messenger, the downstream effects of this signal and the importance for cyst development are unknown.…”

Human ADPKD primary cyst epithelial cells with a novel, single codon deletion in the PKD1 gene exhibit defective ciliary polycystin localization and loss of flow-induced Ca2+ signaling

“…Nauli et al (30) soon proposed that the PC1/PC2 complex functions as a flow-sensing mechanoreceptor in the primary cilia of primary cultures of mouse embryonic renal epithelial cells and showed that cells from pkd1(Ϫ/Ϫ) mice are deficient in this proposed sensing function. The partially defective flow-sensing in the orpk mouse (21,55) further supported the proposed central role of defective ciliary sensation of and/or response to tubular flow to the cystogenesis of ADPKD (30). The pkd1(Ϫ/Ϫ) mouse embryonic renal epithelial cells in which flow-induced signaling defects were observed completely lacked PC1 and PC2 polypeptides.…”

“…This ciliary phenotype of human cyst epithelial cells in primary culture is intermediate in severity between the severely shortened, dysmorphic cilia of cells cultured from the orpk mouse with a hypomorphic polaris mutation (40) Flow-sensitive Ca 2ϩ -signaling is a property of isolated, perfused rabbit and mouse collecting ducts (22,21) and has also been reported in isolated, perfused mouse medullary thick ascending limb (19). In the orpk mouse model of recessive PKD, flow-evoked Ca 2ϩ signaling in the cortical collecting duct (CCD) remained normal during the first postnatal week but was slightly reduced compared with wild-type CCD at age 2 wk (21), a difference replicated in CCD cells grown in primary culture (55).…”

From streaming‐potentials to shear stress: 25 years of bone cell mechanotransduction

“…59 Malone et al 60 have recently demonstrated that fluid flow deflects the primary cilium in models of osteoblastic and osteocytic cells, and disruption of the cilium via pharmacological or genetic means diminishes the effects of fluid flow on prostaglandin production and osteopontin expression. However, disrupting the primary cilium did not affect the ability of osteoblastic cells to respond to fluid flow with an increase in intracellular calcium, as is the case in kidney cells, 61 suggesting that other mechanosensory mechanisms must work in conjunction with the cilium. Indeed, it is likely that groups of these potential mechanosensors, as well as some yet to be identified, act in concert to allow bone cells to respond to interstitial fluid flow.…”