Resolving Contributions of Oxygen-Consuming and ROS-Generating Enzymes at the Synapse

作者：Abdel Rahman Engy A; Mahmoud Ali M; Aaliya Abdullah; Radwan Yasmine; Yasseen Basma; Al Okda Abdelrahman; Atwa Ahmed; Elhanafy Eslam; Habashy Moaaz; Ali Sameh S

来源：Oxidative Medicine and Cellular Longevity, 2016, 2016: 1089364.

DOI：10.1155/2016/1089364

<jats:p>Disruption of cellular redox homeostasis is implicated in a wide variety of pathologic conditions and aging. A fundamental factor that dictates such balance is the ratio between mitochondria-mediated complete oxygen reduction into water and incomplete reduction into superoxide radical by mitochondria and NADPH oxidase (NOX) enzymatic activity. Here we determined mitochondrial as well as NOX-dependent rates of oxygen consumption in parallel with H<jats:sub>2</jats:sub>O<jats:sub>2</jats:sub>generation in freshly isolated synaptosomes using high resolution respirometry combined with fluorescence or electrochemical sensory. Our results indicate that although synaptic mitochondria exhibit substantially higher respiratory activities (8–82-fold greater than NOX oxygen consumption depending on mitochondrial respiratory state), NADPH-dependent oxygen consumption is associated with greater H<jats:sub>2</jats:sub>O<jats:sub>2</jats:sub>production (6-7-fold higher NOX-H<jats:sub>2</jats:sub>O<jats:sub>2</jats:sub>). We also show that, in terms of the consumed oxygen, while synaptic mitochondria “leaked”<mml:math xmlns:mml="http://www.w3.org/1998/Math/MathML" id="M1"><mml:mn fontstyle="italic">0.71</mml:mn><mml:mi>%</mml:mi><mml:mo>±</mml:mo><mml:mn fontstyle="italic">0.12</mml:mn></mml:math>H<jats:sub>2</jats:sub>O<jats:sub>2</jats:sub>during NAD<jats:sup>+</jats:sup>-linked resting,<mml:math xmlns:mml="http://www.w3.org/1998/Math/MathML" id="M2"><mml:mn fontstyle="italic">0.21</mml:mn><mml:mi>%</mml:mi><mml:mo>±</mml:mo><mml:mn fontstyle="italic">0.04</mml:mn></mml:math>during NAD<jats:sup>+</jats:sup>-linked active respiration, and<mml:math xmlns:mml="http://www.w3.org/1998/Math/MathML" id="M3"><mml:mn fontstyle="italic">0.07</mml:mn><mml:mi>%</mml:mi><mml:mo>±</mml:mo><mml:mn fontstyle="italic">0.02</mml:mn></mml:math>during FAD<jats:sup>+</jats:sup>-linked active respiration, NOX converted<mml:math xmlns:mml="http://www.w3.org/1998/Math/MathML" id="M4"><mml:mn fontstyle="italic">38</mml:mn><mml:mi>%</mml:mi><mml:mo>±</mml:mo><mml:mn fontstyle="italic">13</mml:mn></mml:math>of O<jats:sub>2</jats:sub>into H<jats:sub>2</jats:sub>O<jats:sub>2</jats:sub>. Our results indicate that NOX rather than mitochondria is the major source of synaptic H<jats:sub>2</jats:sub>O<jats:sub>2</jats:sub>. The present approach may assist in the identification of redox-modulating synaptic factors that underlie a variety of physiological and pathological processes in neurons.</jats:p>

• 出版日期2016