All experiments were performed in triplicate and the data shown are representative

e for Complex II and Ascorbate Near-Infrared Light Rescues a Parkinson Fly Model and TMPD for Complex IV. To inhibit upstream Complexes rotenone was added for Complex IIdriven oxygen consumption measurements, antimycin A for Complex IV-driven oxygen consumption and potassium cyanide to block Complex IV. To test if laser light can stimulate oxygen consumption, we applied 808 nm laser light of 25 mW/cm2 for 100 s directly onto the mitochon- drial suspension and measured the slope of ADP-stimulated oxygen consumption before and after the light stimulus. We calculated the ratio of oxygen consumption after versus before light stimulus. As a control we performed the same treatment but did not switch on the light and calculated the ratio of the slope after versus before `mock treatment’. 3 Near-Infrared Light Rescues a Parkinson Fly Model Statistical Analysis All experiments were evaluated using the appropriate ANOVA test followed by post hoc Dunnett test using GraphPad Prism 6. Results 808 nm Light Stimulation Rescues pink1 Mutant Flight Defects Complex IV of the ETC shows absorption maxima around 670 nm and 800840 nm , but the higher wavelength penetrates tissues more effectively. We used an 808 nm laser fitted with a frontal light distributor to simultaneously irradiate groups of control and pink1B9 null mutant SCH58261 site animals using different conditions and tested 25617690 for their ability to fly at different time points following light exposure. We find that pink1B9 mutant flies irradiated for 100 s with 25 mW/cm2 808 nm light show rescue of their ability to fly which is most pronounced 5 h following the light stimulation, while no effect was observed in control flies. The rescue is only partial but is very similar to the level obtained previously when testing various genetic and pharmacological conditions suppressing pink1 mutant flight defects. The rescue appears specific to 808 nm light because irradiating pink1B9 mutants using 100 s of 25 mW/cm2 730 nm light that is less absorbed by Complex IV in cells in vitro, does not rescue the inability of the mutants to fly and has no effect on control flies. In most of the following experiments we thus used 100 s of 25 mW/cm2 808 nm light to irradiate the flies. 808 nm Light Rescues Functional Defects in pink1 Mutant Mitochondria Pink1B9 mutant flies suffer from mitochondrial dysfunction, including lower ATP levels and reduced mitochondrial membrane potential . We irradiated adult mutant and control flies and measured ATP levels 5 h later in neuron-enriched and muscle-enriched material using a luciferase based assay. Irradiation results in increased ATP levels. Similarly, we assessed ym at larval motor neuron endplates 5 h after irradiation using JC-1, a potentiometric dye. JC-1 reports on the ym at Drosophila motor endplates because in control boutons, red JC-1 fluorescence is clearly visible while in samples treated with the Complex I inhibitor 24900262 rotenone, red labeling is significantly reduced. Similarly, in samples treated with FCCP, an uncoupler, red labeling is lost. As reported before, red JC-1 fluorescence in pink1B9 larvae is reduced compared to controls. While JC-1 red/green fluorescence ratio between irradiated and non-irradiated pink1RV larvae is not appreciably different, red JC-1 fluorescence in irradiated pink1B9 larvae is significantly rescued compared to nonirradiated pink1B9 animals. These data suggest that 808 nm light improves the mitochondrial membrane potential and ATP levels in pink1B9 mutan