Figure 4: Neuron-to-neuron transfer of rTg4510 mouse brain-derived human tau species in a three-chambered microfluidic device.

(a) PBS-soluble extract from rTg4510 brain (12-month-old, 500 ng ml⁻¹ human tau) was added to the 1st chamber of a 3-chamber microfluidic device. Diffusion of brain extract from the 1st to the 2nd chamber was blocked by a hydrostatic pressure barrier. (b) Immunostaining for human tau (green) and total (human and mouse) tau (red) at day 5. Human tau positive neurons were detected in the 2nd chamber (white arrow). Neurons in the side reservoir of the 2nd chamber were negative for human tau staining (bottom). (c) A human tau positive axon (arrow) and dendrite (arrow head) extending from the 2nd chamber neuron. (d) Concentration dependency of tau uptake and propagation. rTg4510 brain extract (PBS-3,000g) was diluted in culture medium to obtain three different concentrations (6, 60 and 600 ng ml⁻¹) of human tau and added into the 1st chamber. Neurons were immunostained for human tau and total (human and mouse) tau at day 5. (d, right) Quantification of fluorescence intensity of human tau staining in the 2nd chamber. (n=4–7). One-way ANOVA. (e) Time course of neuron-to-neuron transfer of rTg4510 brain-derived human tau. The rTg4510 brain extract (PBS-3,000g, 500 ng ml⁻¹ human tau) was added to the 1st chamber and incubated for up to 14 days. Neurons were immunostained at different time points. Human tau positive 2nd chamber neurons (arrow head) and axons from the 1st chamber neuron (arrow) were detected after 5 days of incubation. Human tau positive axons were detected in the 3rd chamber after 8 days (arrow). Scale bar, 50 μm. *P<0.05.