Endoplasmic reticulum stress (ER stress) is involved in the development and progression of various forms of heart disease and may lead to myocardial apoptosis. Sphingosine-1-phosphate (S1P) possesses cardioprotective properties, including anti-apoptosis. However, little is known about the link between S1P and ER stress-induced myocardial apoptosis. This study investigated the regulatory role of S1P in ER stress-induced apoptosis in cardiomyocytes.ER stress and myocardial apoptosis were induced by transverse aortic constriction (TAC) or tunicamycin in mice, which were then treated with 2-acetyl-5-tetrahydroxybutyl imidazole (THI) or S1P. AC16 cells were treated with tunicamycin or thapsigargin, or pretreated with S1P, sphingosine-1-phosphate receptor (S1PR) subtype antagonists, S1PR1 agonist, and PI3K and MEK inhibitors. Cardiac function, the level of S1P in plasma and heart, ER stress markers, cell viability, and apoptosis were detected.S1P reduced the expression of ER stress-related molecules and ER stress-induced myocardial apoptosis in mice subjected to TAC or an injection of tunicamycin. Furthermore, in AC16 cells exposed to thapsigargin or tunicamycin, S1P decreased the expression of ER stress-related molecules, promoting cell viability and survival. Nevertheless, the S1PR1 antagonist abrogated the protection of S1P. Subsequently, in TAC S1PR1 heterozygous (S1PR1+/-) mice, S1P had no effect on ER stress and apoptosis in cardiomyocytes. Notably, in vitro, the impact of anti-ER stress-induced myocardial apoptosis by the S1PR1 agonist was reversed by PI3K and MEK inhibitors.This study is the first to demonstrate that S1P relieves ER stress-induced myocardial apoptosis via S1PR1/AKT and S1PR1/ERK1/2, which are potential therapeutic targets for heart disease.Copyright © 2022. Published by Elsevier Inc.