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Characterization of constitutive and acid-induced outwardly rectifying chloride currents in immortalized mouse distal tubular cells.


ABSTRACT: Thiazides block Na+ reabsorption while enhancing Ca2+ reabsorption in the kidney. As previously demonstrated in immortalized mouse distal convoluted tubule (MDCT) cells, chlorothiazide application induced a robust plasma membrane hyperpolarization, which increased Ca2+ uptake. This essential thiazide-induced hyperpolarization was prevented by the Cl- channel inhibitor 5-Nitro-2-(3-phenylpropylamino) benzoic acid (NPPB), implicating NPPB-sensitive Cl- channels, however the nature of these Cl- channels has been rarely described in the literature. Here we show that MDCT cells express a dominant, outwardly rectifying Cl- current at extracellular pH7.4. This constitutive Cl- current was more permeable to larger anions (Eisenman sequence I; I->Br-?Cl-) and was substantially inhibited by >100mM [Ca2+]o, which distinguished it from ClC-K2/barttin. Moreover, the constitutive Cl- current was blocked by NPPB, along with other Cl- channel inhibitors (4,4'-diisothiocyanatostilbene-2,2'-disulfonate, DIDS; flufenamic acid, FFA). Subjecting the MDCT cells to an acidic extracellular solution (pH<5.5) induced a substantially larger outwardly rectifying NPPB-sensitive Cl- current. This acid-induced Cl- current was also anion permeable (I->Br->Cl-), but was distinguished from the constitutive Cl- current by its rectification characteristics, ion sensitivities, and response to FFA. In addition, we have identified similar outwardly rectifying and acid-sensitive currents in immortalized cells from the inner medullary collecting duct (mIMCD-3 cells). Expression of an acid-induced Cl- current would be particularly relevant in the acidic IMCD (pH<5.5). To our knowledge, the properties of these Cl- currents are unique and provide the mechanisms to account for the Cl- efflux previously speculated to be present in MDCT cells.

SUBMITTER: Valinsky WC 

PROVIDER: S-EPMC5482324 | biostudies-literature | 2017 Aug

REPOSITORIES: biostudies-literature

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Characterization of constitutive and acid-induced outwardly rectifying chloride currents in immortalized mouse distal tubular cells.

Valinsky William C WC   Touyz Rhian M RM   Shrier Alvin A  

Biochimica et biophysica acta. General subjects 20170505 8


Thiazides block Na<sup>+</sup> reabsorption while enhancing Ca<sup>2+</sup> reabsorption in the kidney. As previously demonstrated in immortalized mouse distal convoluted tubule (MDCT) cells, chlorothiazide application induced a robust plasma membrane hyperpolarization, which increased Ca<sup>2+</sup> uptake. This essential thiazide-induced hyperpolarization was prevented by the Cl<sup>-</sup> channel inhibitor 5-Nitro-2-(3-phenylpropylamino) benzoic acid (NPPB), implicating NPPB-sensitive Cl<su  ...[more]

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