5. A-beta is produced by neurons and cleared through the vasculature cleared through the vasculature

6. Blood flow deficits to the brain also observed in Alzheimer’s patients

9. Image is formed by scanning laser focus through the sample and recording fluorescence intensity

10. Image is formed by scanning laser focus through the sample and recording fluorescence intensity

11. Image fluorescently-labeled features in brain of anesthetized rodent with glass-covered craniotomy

12. Imaging fluorescent blood plasma yields a 3-D cortical “micro-angiography”

13. In vivo imaging of vasculature and amyloid plaques in AD mouse models amyloid plaques in AD mouse models plaques labeled by methoxy-X04 vasculature with intravenous dye injection Aged APPswe/PS1 mice with craniotomies

14. Dye labels blood plasma, but not blood cells, allowing identification of flowing vs. stalled vessels

15. Fraction of capillaries with stalled blood flow increased to ~2% in mouse models of AD Temporary stalls that shift between capillary segments, rather than permanent occlusions 2843 capillaries in 6 AD mice 2475 capillaries in 4 WT mice

16. Labeling to distinguish red blood cells, leukocytes, and thrombi as potential cause of stalls leukocytes: rhodamine-6G and Hoechst thrombi: rhodamine-6G red blood cells: unlabeled

17. In vivo imaging of capillary stalls with rhodamine-6G and Hoechst labeling red - Texas-Red dextran green - rhodamine 6G blue - methoxy-X04 and Hoescht

18. Majority of capillary stalls in AD mouse models are caused by leukocyte plugs

19. How does a single stalled capillary affect blood flow? affect blood flow? N. Nishimura, et al., Nature Methods 3, 99 (2006) Laser injury to vessel triggers clotting Map changes in flow after capillary clot baseline flow post-clot flow mm/s

20. A single stalled capillary causes reduced blood flow in multiple downstream vessel branches N. Nishimura, et al., Nature Methods 3, 99 (2006) Post-clot blood flow speed (fraction of baseline)

21. Map of blood vessels and amyloid plaques from AD mouse plaques from AD mouse

22. Location of stalled capillaries

23. Simulate blood flow changes in capillaries downstream from plugged vessels

24. Simulate blood flow changes in capillaries downstream from plugged vessels

25. Simulate blood flow changes in capillaries downstream from plugged vessels

26. Increases in number of stalled capillaries causes decreases in average cerebral blood flow 2% of capillaries stalled predicts a 30% decrease in flow compared to controls

29. Cyclic relationship between vascular stalls and Aβ aggregates as a driver of AD progression leukocyte plugs Aβ aggregates

31. Acknowledgements Thanks to William Klunk for donation of methoxy-X04

33. To occlude a small blood vessel, we optically injure the vessel to initiate endogenous clotting cascade N. Nishimura, et al., Nature Methods 3, 99 (2006)

34. Femtosecond laser irradiation to produce a clot in a targeted, sub-surface brain capillary