Redistribution of aquaporin-2 water channels induced by vasopressin in rat kidney inner medullary collecting duct

Abstract

Aquaporin-2 (AQP2) is the predominant vasopressin-regulated water channel of the renal collecting duct. We tested whether vasopressin induces translocation of AQP2 from intracellular vesicles into the apical plasma membrane. AQP2 was quantitated in plasma membrane and intracellular vesicle fractions prepared from the inner medulla of one kidney from each rat before or 20 min after intravenous 1-desamino-8-D-arginine vasopressin (DDAVP) treatment, using immunoblotting and densitometry. Contralateral kidneys were prepared for immunofluorescence and immunoelectron microscopy. Immunoblotting revealed that, compared with untreated controls, DDAVP treatment significantly increased the fraction of AQP2 protein associated with the plasma membrane fraction relative to intracellular vesicles. This increase averaged 2.0-fold in untreated rats and 2.9-fold in rats water loaded for 12 h. Water loading, presumably by suppressing circulating vasopressin levels, decreased the fraction of AQP2 associated with the plasma membrane by 55%, suggesting retrieval of AQP2 from the plasma membrane. In rats sequentially thirsted for 48 h to increase expression and then water loaded for 72 h to minimize plasma membrane labeling, DDAVP caused a 12-fold increase in the plasma membrane to intracellular vesicle labeling ratio. The accentuation of the DDAVP response seen after water loading is consistent with the observed increase in the fraction of AQP2 in the intracellular pool available for insertion. Immunofluorescence confirmed a marked DDAVP-induced redistribution of AQP2 from intracellular to plasma membrane domains. Furthermore, quantitative immunoelectron microscopy demonstrated a 3.4-fold increase in apical plasma membrane to intracellular vesicle labeling ratio. These results provide a direct in vivo demonstration of vasopressin-induced translocation of AQP2 into the apical plasma membrane.

Acute and chronic metabolic acidosis interferes with aquaporin-2 translocation in the rat kidney collecting ducts

“…[21][22][23] Kidneys from control and acidosis rats were homogenized in an ice-cold isolation solution (10 mM triethanolamine (pH 7.6) and 250 mM sucrose) containing protease inhibitors, leupeptin (1 mg ml À1 ; Peptide Institute, Osaka, Japan) and phenylmethylsulfonyl fluoride (0.1 mg ml À1 ; Wako Pure Chemical, Tokyo, Japan), some of which were collected as whole-kidney homogenates. The remainders were centrifuged initially at 4000 g for 10 min at 4 1C (Himac centrifuge CR5B, Hitachi, Tokyo, Japan) to remove incompletely homogenized fragments and nuclei.…”

Membrane Trafficking of the Cystic Fibrosis Gene Product, Cystic Fibrosis Transmembrane Conductance Regulator, Tagged with Green Fluorescent Protein in Madin-Darby Canine Kidney Cells

“…Agonist-stimulated trafficking of polytopic membrane transport proteins is often cell type-specific. For example, aquaporin-2 traffics from intracellular vesicles to the apical plasma membrane following treatment with cAMP-stimulating agents in collecting duct principal cells (52) but not in Xenopus oocytes (53). Similarly, insulin stimulates GLUT-4 trafficking from an intracellular pool to the plasma membrane in adipocytes and skeletal muscle but not in heterologous expression systems (54).…”

From vasopressin receptor to water channel: intracellular traffic, constraint and by-pass

“…Interestingly, it is not only the general increase in AQP2 expression which is associated with fluid balance status, but also its location within the cells. Thus, water loading is associated with an increased translocation of AQP2 from membrane to intracellular vesicle without any change in total cellular water channels, as indicated by gold particle labelling density (Marples et al 1995a). In contrast, V 2 agonist treatment results in the shift of AQP2 molecules from the intracellular vesicles to the apical plasma membrane (Marples et al 1995a).…”

“…When VP binds to its V 2 receptor in the collecting duct cell both cAMP and calcium mobilization pathways can be activated (Ecelbarger et al 1996). This latter mechanism could by-pass the AC system, perhaps explaining how dDAVP can increase AQP2 expression in the apical membrane during lithium treatment (Marples et al 1995a). How this second postulated V 2 receptor mechanism might increase AQP2 expression when there is a reduction in receptor expression associated with increased circulating VP levels but not when that receptor expression is reduced in the absence of ligand stimulation, is unclear.…”

“…This recently cloned molecule (Fushimi et al 1993) is located in the apical membranes of collecting duct cells, and within the cytoplasm, where they appear to be stored in vesicles called aggraphores (Harris et al 1991, Nielsen et al 1993. In the presence of VP there is an immediate increase in the presence of AQP2 in the apical membranes and an increased movement of aggraphores towards these membranes, followed by a longer-term increase in AQP2 production within the cells, as determined by immunofluorescence (Marples et al 1995a). There is evidence that microtubules might be implicated in VP-stimulated water transport (Phillips & Taylor 1989), perhaps associated with the movement of aggraphores to the apical membrane.…”