Cell Deformation at the Air-Liquid Interface Evokes Intracellular Ca2+ Increase and ATP Release in Cultured Rat Urothelial Cells

Jiliang Wen; Zhenghao Chen; Mengmeng Zhao; Shulu Zu; Shengtian Zhao; Shaoyong Wang; Xiulin Zhang

doi:10.3389/fphys.2021.631022

Cell Deformation at the Air-Liquid Interface Evokes Intracellular Ca²⁺ Increase and ATP Release in Cultured Rat Urothelial Cells

Front Physiol. 2021 Feb 3:12:631022. doi: 10.3389/fphys.2021.631022. eCollection 2021.

Authors

Jiliang Wen^{1

2}, Zhenghao Chen¹, Mengmeng Zhao¹, Shulu Zu¹, Shengtian Zhao², Shaoyong Wang¹, Xiulin Zhang¹

Affiliations

¹ Department of Urology, The Second Hospital, Cheeloo College of Medicine, Shandong University, Jinan, China.
² Department of Urology, Shandong Provincial Hospital, Cheeloo College of Medicine, Shandong University, Jinan, China.

Abstract

Urothelial cells have been implicated in bladder mechanosensory transduction, and thus, initiation of the micturition reflex. Cell deformation caused by tension forces at an air-liquid interface (ALI) can induce an increase in intracellular Ca²⁺ concentration ([Ca²⁺]_i) and ATP release in some epithelial cells. In this study, we aimed to examine the cellular mechanisms underlying ALI-induced [Ca²⁺]_i increase in cultured urothelial cells. The ALI was created by stopping the influx of the perfusion but maintaining efflux. The [Ca²⁺]_i increase was measured using the Ca²⁺ imaging method. The ALI evoked a reversible [Ca²⁺]_i increase and ATP release in urothelial cells, which was almost abolished by GdCl₃. The specific antagonist of the transient receptor potential vanilloid (TRPV4) channel (HC0674) and the antagonist of the pannexin 1 channel (¹⁰panx) both diminished the [Ca²⁺]_i increase. The blocker of Ca²⁺-ATPase pumps on the endoplasmic reticulum (thapsigargin), the IP3 receptor antagonist (Xest-C), and the ryanodine receptor antagonist (ryanodine) all attenuated the [Ca²⁺]_i increase. Degrading extracellular ATP with apyrase or blocking ATP receptors (P2X or P2Y) with pyridoxalphosphate-6-azophenyl-2',4'-disulfonic acid (PPADS) significantly attenuated the [Ca²⁺]_i increase. Our results suggest that both Ca²⁺ influx via TRPV4 or pannexin 1 and Ca²⁺ release from intracellular Ca²⁺ stores via IP3 or ryanodine receptors contribute to the mechanical responses of urothelial cells. The release of ATP further enhances the [Ca²⁺]_i increase by activating P2X and P2Y receptors via autocrine or paracrine mechanisms.

Keywords: ATP; TRPV4 channel; mechanosensory transduction; pannaxin-1 channel; urothelial cells.