Hydrogen peroxide mobilizes Ca2+ through two distinct mechanisms in rat hepatocytes |
| |
Authors: | Hirohiko Sato Teruko Takeo Qiang Liu Kyoko Nakano Tomohiro Osanai Sechiko Suga Makoto Wakui Jie Wu |
| |
Institution: | 1.Department of Physiology, Hirosaki University School of Medicine, Hirosaki 036-8562, Japan;2.Department of Medical Technology, Hirosaki University School of Health Sciences, Hirosaki 036-8564, Japan;3.Division of Neurology, Barrow Neurological Institute, Phoenix, AZ, 85013-4496, USA;4.Department of Internal Medicine, Hirosaki University School of Medicine, Hirosaki 036-8562, Japan |
| |
Abstract: | 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 processes.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 blocker 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 {"type":"entrez-nucleotide","attrs":{"text":"U73122","term_id":"4098075","term_text":"U73122"}}U73122 but was not eliminated by removal of extracellular Ca2+. PLC activity was increased by 1 mmol/L H2O2 but not by 400 μmol/L H2O2.Conclusion:H2O2 mobilizes Ca2+ through two distinct mechanisms. In one, 400 μmol/L H2O2-induced sustained Ca2+]i elevation 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. |
| |
Keywords: | hydrogen peroxide Ca2+ dynamics non-selective cation channel intracellular ATP phospholipase C hepatocyte patch-clamp |
|
|