ToolBox: Live Imaging of intracellular organelle transport in induced pluripotent stem cell‐derived neurons

Induced pluripotent stem cells (iPSCs) hold promise to revolutionize studies of intracellular transport in live human neurons and to shed new light on the role of dysfunctional transport in neurodegenerative disorders. Here, we describe an approach for live imaging of axonal and dendritic transport in iPSC-derived cortical neurons. We use transfection and transient expression of genetically-encoded fluorescent markers to characterize the motility of Rab-positive vesicles, including early, late and recycling endosomes, as well as autophagosomes and mitochondria in iPSC-derived neurons. Comparing transport parameters of these organelles with data from primary rat hippocampal neurons, we uncover remarkable similarities. In addition, we generated lysosomal-associated membrane protein 1 (LAMP1)-enhanced green fluorescent protein (EGFP) knock-in iPSCs and show that knock-in neurons can be used to study the transport of endogenously labeled vesicles, as a parallel approach to the transient overexpression of fluorescently labeled organelle markers.