Inactivation of aconitase during the apoptosis of mouse cerebellar granule neurons induced by a deprivation of membrane depolarization.
During the excitotoxic neuronal cell death which accompanies an overflow of extracellular Ca(2+) into neurons, aconitase, an oxidative stress-sensitive enzyme of the tricarboxylic acid (TCA)-cycle in mitochondria, is inactivated due to the generation of oxidative stress (Patel et al. [1996] Neuron 16:345-355). In this study, we investigated whether aconitase could be inactivated during the apoptosis of mouse cerebellar granule cells (CGCs), which was caused by a deprivation of membrane depolarization followed by a stoppage of Ca(2+) influx into CGCs. Upon lowering the potassium (K(+)) concentration in medium from 25 to 5 mM (low K(+)), aconitase was inactivated in accordance with the decrease in methylthiazoletetrazolium (MTT)-reducing activity although its mRNA expression did not change. The blockade of Ca(2+) influx into CGCs mediated by nicardipine at 25 mM KCl also caused the inactivation of aconitase, accompanying induction of the apoptosis of CGCs. Suppression of the apoptosis of CGCs mediated by the Ca(2+) influx or neurotrophic factors such as brain-derived neurotrophic factor (BDNF) and adenylate cyclase activating polypeptide-38 (PACAP-38) attenuated the aconitase inactivation as well as the lactate dehydrogenase (LDH)-release and the decrease in MTT reduction. On the other hand, the levels of intracellular glutathione and manganese superoxide dismutase-2 mRNA decreased under the low K(+) condition, supporting a cause for oxidative stress at low K(+) due to a loss of anti-oxidant activity. Thus, the inactivation of aconitase is also caused by a deprivation of Ca(2+) influx into neurons, suggesting that aconitase is a key mitochondrial enzyme influencing the viability of neurons in response to oxidative stress.
Tabuchi
Akiko
A
Toyama Medical and Pharmaceutical University, Department of Biological Chemistry, Faculty of Pharmaceutical Sciences, Toyama, Japan. tsuda@ms.toyama-mpu.ac.jp
Funaji
Kouhei
K
Nakatsubo
Jun
J
Fukuchi
Mamoru
M
Tsuchiya
Tomofusa
T
Tsuda
Masaaki
M
D000154:Aconitate Hydratase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D002478:Cells, Cultured; D002531:Cerebellum / Q000187:drug effects / Q000201:enzymology*; D004789:Enzyme Activation / Q000187:drug effects / Q000502:physiology; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D051379:Mice; D009474:Neurons / Q000187:drug effects / Q000201:enzymology*
D000154:Aconitate Hydratase