Uncoupling of ATP-depletion and cell death in human dopaminergic neurons

Abstract

The mitochondrial inhibitor 1-methyl-4-phenylpyridinium (MPP(+)) is the toxicologically relevant metabolite of 1-methyl-4-phenyltetrahydropyridine (MPTP), which causes relatively selective degeneration of dopaminergic neurons in the substantia nigra. Dopaminergic LUHMES cells were used to investigate whether ATP-depletion can be uncoupled from cell death as a downstream event in these fully post-mitotic human neurons. Biochemical assays indicated that in the homogeneously differentiated cell cultures, MPP(+) was taken up by the dopamine transporter (DAT). MPP(+) then triggered oxidative stress and caspase activation, as well as ATP-depletion followed by cell death. Enhanced survival of the neurons in the presence of agents interfering with mitochondrial pathology, such as the fission inhibitor Mdivi-1 or a Bax channel blocker suggested a pivotal role of mitochondria in this model. However, these compounds did not prevent cellular ATP-depletion. To further investigate whether cells could be rescued despite respiratory chain inhibition by MPP(+), we have chosen a diverse set of pharmacological inhibitors well-known to interfere with MPP(+) toxicity. The antioxidant ascorbate, the iron chelator desferoxamine, the stress kinase inhibitor CEP1347, and different caspase inhibitors reduced cell death, but allowed ATP-depletion in protected cells. None of these compounds interfered with MPP(+) accumulation in the cells. These findings suggest that ATP-depletion, as the initial mitochondrial effect of MPP(+), requires further downstream processes to result in neuronal death. These processes may form self-enhancing signaling loops, that aggravate an initial energetic impairment and eventually determine cell fate.

Effects of Partial Inhibition of Respiratory Complex I on H2O2 Production by Isolated Brain Mitochondria in Different Respiratory States

“…The mechanisms involved in dopaminergic neurodegeneration linked to complex I inhibition probably involve increased mitochondrial reactive oxygen species (ROS) generation and/or cellular energy crisis due to reduced respiratory capacity [10,[13][14][15][16][17][18][19]. Indeed, marked stimulation of H 2 O 2 production by isolated brain mitochondria has been reported when complex I is extensively inhibited by rotenone [20][21][22][23] through a mechanism linked to an increased NADH/NAD ?…”

Generation of genetically-modified human differentiated cells for toxicological tests and the study of neurodegenerative diseases

“…Following the observation of the early MPP + -mediated effect on mitochondrial movement, we addressed the question of a potential mechanism operating independent of ATP depletion (Pöltl et al, 2012). In addition to its inhibitory action on mitochondrial complex I, MPP + promotes oxidative stress conditions (Seaton et al, 1997).…”

Activation of SK2 channels preserves ER Ca2+ homeostasis and protects against ER stress-induced cell death

“…Earlier studies indicate that neuronal apoptosis can be diminished by lowered mitochondrial respiration and ATP production. [43][44][45][46] Here, the inhibitory effects on mitochondrial respiration were accompanied by an initial boost in glycolysis (Supplementary Figure S6), which was previously associated with the protective properties of CyPPA pretreatment in oxidative damage. 29 However, further investigations are needed to clarify whether changes in mitochondrial respiration by SK channel activation contribute to the protective effects under conditions of ER stress.…”