# Nitric oxide mediates ET-1-induced-inhibition of NPPB-sensitive Cl− currents in the early distal convoluted tubule of the mouse kidney

## TL;DR

The paper shows that nitric oxide helps endothelin-1 reduce chloride currents in mouse kidney cells, affecting salt absorption.

## Contribution

The study reveals a new pathway involving nitric oxide and phosphodiesterase-2 in regulating chloride channels in the kidney's distal convoluted tubule.

## Key findings

- ET-1 inhibits ClC-K2 channels in the DCT via a nitric oxide and sGMP-dependent pathway.
- Phosphodiesterase-2 inhibition prevents ET-1's effect on NCC and ClC-K2.
- NO-donor or cGMP alone inhibits ClC-K2 activity without additional ET-1 effects.

## Abstract

Endothelin-1 (ET-1) from renal-tubule-epithelial-cells inhibits NaCl reabsorption via ETB receptor in an autocrine manner, and inhibition of ETB receptors leads to salt-sensitive-hypertension. In the distal convoluted tubule (DCT), NaCl enters the cell via NaCl-cotransporter (NCC) and Cl− exits the cell in part by ClC-K2 channels, which play a role in regulating With-No-lysine kinase 4 (WNK4). The study aims to explore whether ET-1-induced inhibition of NaCl absorption is also achieved by inhibiting the basolateral Cl− channels in the DCT. Patch-clamp and immunoblotting assessed ET-1 effects on DCT Cl− channels and NCC. Immunofluorescence images detected ETB-receptor expression in parvalbumin-positive DCT. Application of ET-1 decreased NPPB-sensitive Cl− currents and reduced 10-pS Cl− channel activity (ClC-K2), defined by NPo (A product of channel number and open probability); this effect was absent in the presence of ETB receptor inhibitor. Application of L-NAME (nitric oxide synthase inhibitor), ODQ (soluble guanosine cyclase inhibitor) or Bay-60-7550 (phosphodiesterase-2-inhibitor) perse did not affect Cl− channels, but it abolished the inhibitory effect of ET-1. In contrast, application of NO-donor or cGMP inhibited ClC-K2 channel activity of the DCT. Moreover, ET-1 had no additional inhibitory effect of ET-1 on ClC-K2 in the presence of NO-donor or cGMP. Immunoblotting demonstrated that ET-1 treatment (200 nM) of renal cortex decreased NCC phosphorylation and total NCC expression, an effect that was abolished by inhibiting phosphodiesterase-2 but not by KT-5823 (PKG-inhibitor). In conclusion, ET-1 inhibits NCC and ClC-K2 in DCT by NO-sGMP-phosphodiesterase-2-dependent pathway.