Nitric oxide synthesis stimulated by arachidonic acid accumulation via PPARα in acetylcholine-stimulated gastric mucous cells

Saori Tanaka; Shigenori Ito; Chikao Shimamoto; Hitoshi Matsumura; Toshio Inui; Yoshinori Marunaka; Takashi Nakahari

doi:10.1113/EP089517

Nitric oxide synthesis stimulated by arachidonic acid accumulation via PPARα in acetylcholine-stimulated gastric mucous cells

Exp Physiol. 2021 Sep;106(9):1939-1949. doi: 10.1113/EP089517. Epub 2021 Jul 23.

Authors

Saori Tanaka^{1

2}, Shigenori Ito³, Chikao Shimamoto², Hitoshi Matsumura², Toshio Inui^{1

4}, Yoshinori Marunaka^{1

5

6}, Takashi Nakahari¹

Affiliations

¹ Research Unit for Epithelial Physiology, Research Organization of Science and Technology, Biwako Kusatsu Campus, Ritsumeikan University, Kusatsu, Japan.
² Laboratory of Pharmacotherapy, Faculty of Pharmacy, Osaka Medical and Pharmaceutical University, Takatsuki, Japan.
³ Department of Chemistry, Faculty of Medicine, Osaka Medical and Pharmaceutical University, Takatsuki, Japan.
⁴ Saisei Mirai Clinics, Moriguchi, Japan.
⁵ Medical Research Institute, Kyoto Industrial Health Association, Kyoto, Japan.
⁶ Department of Molecular Cell Physiology, Graduate School of Medical Science, Kyoto Prefectural University of Medicine, Kyoto, Japan.

PMID: 34216172
DOI: 10.1113/EP089517

Abstract

New findings: What is the central question of this study? Arachidonic acid (AA) stimulates NO production in antral mucous cells without any increase in [Ca²⁺ ]_i . Given that the intracellular AA concentration is too low to measure, the relationship between AA accumulation and NO production remains uncertain. Is AA accumulation a key step for NO production? What is the main finding and its importance? We demonstrated that AA accumulation is a key step for NO production. The amount of AA released could be measured using fluorescence-HPLC. The intracellular AA concentration was maintained at < 1 μM. Nitric oxide is produced by AA accumulation in antral mucous cells, not as a direct effect of [Ca²⁺ ]_i .

Abstract: In the present study, we demonstrate that NO production is stimulated by an accumulation of arachidonic acid (AA) mediated via peroxisome proliferation-activated receptor α (PPARα) and that the NO produced enhances Ca²⁺ -regulated exocytosis in ACh-stimulated antral mucous cells. The amount of AA released from the antral mucosa, measured by fluorescence high-performance liquid chromatography (F-HPLC), was increased by addition of ionomycin (10 μM) or ACh, suggesting that AA accumulation is stimulated by an increase in [Ca²⁺ ]_i . The AA production was inhibited by an inhibitor of cytosolic phospholipase A2 (cPLA2-inhα). GW6471 (a PPARα inhibitor) and cPLA2-inhα inhibited NO synthesis stimulated by ACh. Moreover, indomethacin, an inhibitor of cyclooxygenase, stimulated AA accumulation and NO production. However, acetylsalicylic acid did not stimulate AA production and NO synthesis. An analogue of AA (AACOCF3) alone stimulated NO synthesis, which was inhibited by GW6471. In antral mucous cells, indomethacin enhanced Ca²⁺ -regulated exocytosis by increasing NO via PPARα, and the enhancement was abolished by GW6471 and cPLA2-inhα. Thus, AA produced via PLA2 activation is the key step for NO synthesis in ACh-stimulated antral mucous cells and plays important roles in maintaining antral mucous secretion, especially in Ca²⁺ -regulated exocytosis.

Keywords: indomethacin; intracellular Ca2+ concentration; mucin exocytosis; nitric oxide; peroxisome proliferation-activated receptor α; phospholipase A2.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Acetylcholine* / pharmacology
Arachidonic Acid / pharmacology
Calcium
Gastric Mucosa
Nitric Oxide*
PPAR alpha / pharmacology
Pyloric Antrum

Substances

PPAR alpha
Arachidonic Acid
Nitric Oxide
Acetylcholine
Calcium