Detection of Mitochondrial Reactive Oxygen Species in Living Rat Trigeminal Caudal Neurons.
10.11620/IJOB.2015.40.2.103
- Author:
Hae In LEE
1
;
Sang Woo CHUN
Author Information
1. Department of Oral Physiology, College of Dentistry, Institute of Wonkwang Biomaterial and Implant, Wonkwang University, Iksan 570-749, Korea. physio1@wonkwang.ac.kr
- Publication Type:Original Article
- Keywords:
trigeminal caudal neurons;
electron transport complex;
mitochondrial ROS;
confocal microscopy
- MeSH:
Animals;
Electron Transport;
Fluorescence;
Microscopy, Confocal;
Mitochondria;
Neurons*;
Perfusion;
Rats*;
Reactive Oxygen Species*
- From:International Journal of Oral Biology
2015;40(2):103-109
- CountryRepublic of Korea
- Language:Korean
-
Abstract:
Growing evidence suggests that mitochondrial reactive oxygen species (ROS) are involved in various pain states. This study was performed to investigate whether ROS-induced changes in neuronal excitability in trigeminal subnucleus caudalis are related to ROS generation in mitochondria. Confocal scanning laser microscopy was used to measure ROS-induced fluorescence intensity in live rat trigeminal caudalis slices. The ROS level increased during the perfusion of malate, a mitochondrial substrate, after loading of 2',7'-dichlorofluorescin diacetate (H2DCF-DA), an indicator of the intracellular ROS; the ROS level recovered to the control condition after washout. When pre-treated with phenyl N-tert-butylnitrone (PBN) and 4-hydroxy-2,2,6,6-tetramethylpiperidene-1-oxyl (TEMPOL), malate-induced increase of ROS level was suppressed. To identify the direct relation between elevated ROS levels and mitochondria, we applied the malate after double-loading of H2DCF-DA and chloromethyl-X-rosamine (CMXRos; MitoTracker Red), which is a mitochondria-specific fluorescent probe. As a result, increase of both intracellular ROS and mitochondrial ROS were observed simultaneously. This study demonstrated that elevated ROS in trigeminal subnucleus caudalis neuron can be induced through mitochondrial-ROS pathway, primarily by the leakage of ROS from the mitochondrial electron transport chain.
