Na ⁺-K ⁺-2Cl ^– cotransporter 2 located in the human and murine gastric mucosa is involved in secretagogue-induced gastric acid secretion and is downregulated in lipopolysaccharide-treated mice

Li-Fei Zheng  ¹ , Tuo Ji  ¹ , Zi-Hao Guo  ² , Tao Wang  ³ , Xiao-Lin Xiu  ¹ , Xiao-Yu Liu  ¹ , Shi-Chao Li  ¹ , Lu Sun  ¹ , Hong Xue  ¹ , Yue Zhang  ¹ , Jin-Xia Zhu  ⁴ Affiliations

• PMID: 32423868
• DOI: 10.1016/j.ejphar.2020.173162

Abstract

Na⁺-K⁺-2Cl^– cotransporter (NKCC) is expressed at exceptionally high levels in gastric parietal cells. Bumetanide, a potent loop diuretic that blocks NKCC, usually causes a decrease in gastric acid secretion. Endotoxaemia causes hypochlorhydria in vivo, in which lipopolysaccharide (LPS) plays an important role. This study aimed to investigate the effect of NKCC2 on gastric acid secretion and its alteration in LPS-treated mice. The scanning ion-selective electrode technique and real-time pH titration combined with RNA interference were used to determine the effects of bumetanide on gastric acid secretion. Immunochemistry and Western blotting were performed to investigate the changes in NKCC2 expression in LPS-treated mice. Immunoreactivity of NKCC1 and NKCC2 was mainly observed near the basolateral and apical membranes of parietal cells, respectively. Pretreatment with bumetanide reduced the histamine-stimulated H⁺ flux in the mouse gastric mucosa. The apical, but not the basolateral, addition of bumetanide inhibited forskolin- or histamine/3-isobutyl-1-methylxanthine(IBMX)-induced gastric acid secretion. In vivo treatment with NKCC2 siRNA inhibited forskolin-induced acid secretion. Upon histamine stimulation, the majority of NKCC2 was targeted to the apical membrane in the gastric mucosa and in primary cultured parietal cells. The expression of NKCC2 and vesicle-associated membrane protein-2 (VAMP2), but not that of H⁺/K⁺-ATPase, was decreased in the gastric mucosa of LPS-treated mice. Blocking apical NKCC2, but not basolateral NKCC1, by bumetanide inhibited secretagogue-induced gastric acid secretion, during which the membrane trafficking of NKCC2 may be necessary. The downregulation of NKCC2 and VAMP2 may be related to the reduced gastric acid secretion induced by LPS.

Keywords: Bumetanide; Gastric acid secretion; Lipopolysaccharide; Na(+)-k(+)-2Cl(-) cotransporter; Parietal cell.