Hydrogen peroxide mobilizes Ca2+ through two distinct mechanisms in rat hepatocytes

• 来源期刊: 中国药理学报（英文版）
• 关键词: hydrogen peroxide Ca2+ dynamics non-selective cation channel intracellular ATP phospholipase C hepatocyte patch-clamp
• 年，卷（期）: 2009,（1）
• 页码范围: 78-89
• 页数: 12页
• 语种: 英文
• DOI: 10.1038/aps.2008.4

摘要

Aim: Hydrogen peroxide (H2O2) is produced during liver transplantation. Ischemia/reperfusion induces oxidation and causes intracellular Ca2+ overload, which harms liver cells. Our goal was to determine the precise mechanisms of these pro-cesses. Methods: Hepatocytes were extracted from rats. Intracellular Ca2+ concentrations ([Ca2+]i), inner mitochondrial membrane potentials and NAD(P)H levels were measured using fluorescence imaging. Phospholipase C (PLC) activity was detected using exogenous PIP2. ATP concentrations were measured using the luciferin-luciferase method. Patch-clamp recordings were performed to evaluate membrane currents. Results: H2O2 increased intracellular Ca2+ concentrations ([Ca2+]I) across two kinetic phases. A low concentration (400 μmol/L) of H2O2 induced a sustained elevation of [Ca2+]i that was reversed by removing extracellular Ca2+. H2O2 increased membrane currents consistent with intracellular ATP concentrations. The non-selective ATP-sensitive cation channel block-er amiloride inhibited H2O2-induced membrane current increases and [Ca2+]i elevation. A high concentration (1 mmol/L)of H2O2 induced an additional transient elevation of [Ca2+]i, which was abolished by the specific PLC blocker U73122 but was not eliminated by removal of extracellular Ca2+. PLC activity was increased by 1 mmol/L H2O2but not by 400 μmol/L H2O2. Conclusions: H2O2 mobilizes Ca2+ through two distinct mechanisms. In one, 400 μmol/L H2O2-induced sustained [Ca2+]ielevation is mediated via a Ca2+ influx mechanism, under which H2O2 impairs mitochondrial function via oxidative stress, reduces intracellular ATP production, and in turn opens ATP-sensitive, non-specific cation channels, leading to Ca2+ influx.In contrast, 1 mmol/L H2O2-induced transient elevation of [Ca2+]i is mediated via activation of the PLC signaling pathway and subsequently, by mobilization of Ca2+ from intracellular Ca2+ stores.