AbstractNeuronal dense core vesicles (DCVs) transport many cargo molecules like neuropeptides and neurotrophins to their release sites in dendrites or axons. The transport properties of DCVs in axons of the intact mammalian brain are unknown. We used viral expression of a DCV cargo reporter (NPY-Venus/Cherry) in the thalamus and two-photon in vivo imaging to visualize axonal DCV trafficking in thalamo-cortical projections of anesthetized and awake mice. We found an average speed of 1 μm/s, maximal speeds of up to 5 μm/s and a pausing fraction of ~11%. Directionality of transport differed between anesthetized and awake mice. In vivo microtubule +-end extension imaging using Macf18-GFP revealed microtubular growth at 0.12 μm/s and provided positive identification of antero- and retrograde axonal transport. Consistent with previous reports, anterograde transport was faster (~2.1 μm/s) than retrograde transport (~1.4 μm/s). In summary, DCVs are transported with faster maximal speeds and lower pausing fraction in vivo compared to previous results obtained in vitro. Finally, we found that DCVs slowed down upon presynaptic bouton approach. We propose that this mechanism promotes synaptic localization and cargo release.Key pointsDespite their immense physiological and pathophysiological importance, we know very little about the biology of dense core vesicle (DCV) trafficking in the intact mammalian brain.DCVs are transported at similar average speeds in the anesthetized and awake mouse brain compared to neurons in culture, yet maximal speed and pausing fraction of transport were higher.Microtubule +-end extension imaging visualized microtubular growth at 0.12 μm/s and revealed that DCVs were transported faster in the anterograde direction.DCV transport slowed down upon presynaptic bouton approach, possibly promoting synaptic localization and cargo release.Our work provides a basis to extrapolate DCV transport properties determined in cultured neurons to the intact mouse brain and reveal novel features such as slowing upon bouton approach and brain state-dependent trafficking directionality.
In vivo three-photon microscopy of subcortical structures within an intact mouse brain