Mitochondrial Dynamics in Cortical Neurons

Mitochondrial traffic in cortical neurons in culture was evaluated at 37 °C employing wide-field time-laps fluorescence microscopy as described [17 (link)]. Mitochondrial motility was documented with an inverted microscope Nikon Eclipse TE2000-U utilizing Nikon CFI Plan Apo 100× 1.4 NA objective and Cool SNAP_HQ Photometrics camera (Roper Scientific, Tucson, AZ, USA) managed by MetaMorph software 6.3 (Molecular Devices, Downingtown, PA, USA). The excitation light (480 ± 20 nm) was provided by a Lambda-LS system (Sutter Instruments, Novato, CA, USA), and fluorescence was recorded with a 505 nm dichroic mirror at 535 ± 25 nm. The images were captured at 1 Hz during the whole duration of the experiment (5 min). Mitochondrial traffic was evaluated following construction of kymographs with NIH ImageJ software (version 1.53a).