Replication Data for: Differential axonal trafficking of Neuropeptide Y-, LAMP1- and RAB7-tagged organelles in vivo

Different organelles traveling through neurons exhibit distinct properties in vitro, but this has not been investigated in the intact brain. We established simultaneous dual color 2- photon microscopy to visualize the trafficking of Neuropeptide Y-, LAMP1- and RAB7- tagged organelles in thalamocortical axons imaged in mouse cortex in vivo. This revealed that LAMP1- and RAB7-tagged organelles move significantly faster than NPY tagged organelles in both anterograde and retrograde direction. NPY travelled more selectively in anterograde direction than LAMP1 and RAB7. By using a synapse marker and a calcium sensor we further investigated the transport dynamics of NPY-tagged organelles. We found that these organelles slow down and pause at synapses. In contrast to previous in vitro studies, a significant increase of transport speed was observed after spontaneous activity and elevated calcium levels in vivo as well as electrically stimulated activity in acute brain slices. Together, we show a remarkable diversity in speeds and properties of three axonal organelle marker in vivo that differ from properties previously observed in vitro.