Gram-negative bacteria-induced infections result in fever, arrhythmia, and even death. Lipopolysaccharide (LPS), a constituent of bacteria, leads to an inflammatory response under sepsis and increase arrhythmogenesis. This study analyzed the effects on human embryonic stem cell-differentiated cardiomyocytes (HIPSC-CMs) exposed to LPS. A whole cell patch clamp was used to record the action potential (AP) and ionic currents with or without different doses of LPS in HIPSC-CMs. Compared with the control, a different dose (0.04, 0.2, 1, and 5 mu g/ml) of LPS-treated HIPSC-CMs resulted in a longer AP duration. The IC50 of sodium channel current was 1.254 mu g/ml, L-type calcium channel current was 5 mu g/ml, and I-k channel currents were 1.254 mu g/ml. LPS-treated HIPSC-CMs showed a lower sodium channel current, L-type calcium channel current, and I-k channel currents. Furthermore, the expressions of Nav1.5 were decreased, and L-Ca, Kv11.1, and Kv7.1 were increased in LPS-treated HIPSC-CMs. LPS-induced arrhythmogenesis was related to the electrophysiological characteristics of sodium channel current, L-type calcium channel current, and I-k channel currents. These results suggest a potential mechanism of cardiomyocyte injury in endotoxemia.