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Biomedical Imaging and Instrumentation

Biomedical Imaging and Instrumentation - Poster Session E

Poster F7 - Two photon imaging of red blood cell velocity in small brain vessels of Alzheimer's mice

Saturday, October 26, 2024
10:00 AM - 11:00 AM EST
Location: Exhibit Hall E, F & G

Introduction:  Alzheimer’s disease (AD) has complex pathological mechanisms that start silently on cellular level and present decades later as cognitive impairment and dementia. Approximately 6.9 million Americans over the age of 65 are living with AD in 2024 [1]. Although AD is the most common form of dementia, we still have incomplete understanding of its origins. The classic hallmarks of AD are amyloid plagues and tau tangles, however recent studies show that the origins of AD are combination of vulnerability factors such as old age, female sex, poor vascular and immune health, genetic susceptibilities and socioeconomic factors [1]. Due to the complexity of the disease progression, analyses of multiple factors such as cerebral blood flow within the brain capillaries, can help us detect early AD signs and understand how those subtle functional changes relate to the brain’s decline. Cerebral blood flow (CBF) decreases are an early pathological feature in AD and in this study we aimed to establish whether those CBF changes are evident in the smallest blood vessels in the brain [2]. We used in vivo two-photon microscopy in mouse models of AD to track the velocity of red blood cells (RBC) within the brain small vessels and correlated those velocities with the vessel diameters.

Materials and

Methods:  The AD mouse model (N=5) was B6C3 mice expressing human amyloid precursor protein and presenilin (APP/PS1) genes cloned from humans with AD. The controls were the wild-type B6C3 (N=3), age (5-16 months) and sex matched. We installed chronic cortical windows in the mice for optical imaging. The mice were trained for 2 weeks to be imaged awake and head-fixed on a treadmill using a multiphoton Ultima Laser Scanner. An injection of a 1x phosphate buffered saline solution of 2mg/mL Sulforhodamine (SR101) was used to image the brain vasculature with the laser tuned at 920 nm. Line-scan data was obtained by repetitive scanning on the centerline of capillaries in the animal brain. As the RBC flowed through the capillaries during the imaging, bright and dark streaks were produced due to the alternating plasma and cells and the angle of those streaks is a function of the RBC velocity. Velocities and diameters were computed in MATLAB using standard functions. The raw strips of data were limited to blocks of 250 ms, gaussian smoothed and background was subtracted. To obtain the velocity for the imaged blocks, we applied a Radon transform to get the angle of maximum variance [3]. At the line-scanned vessel we manually selected part of the vessel cross section forming a rectangular region of interest (ROI) (Fig 1a) and averaged the area into a line profile. Full width at half maximum (FWHM) of this profile was used to compute the diameters of the vessels (Fig 1b).

Results, Conclusions, and Discussions: Our results showed a decrease in RBC velocity for AD mice as the diameter of the vessels increased (Fig 2). We applied also linear regression to the wild type mice data set which produced an equation of y = 0.021083x+0.74806 indicating a trend for increase in RBC velocity, however, this relationship proved insignificant (p=0.268). In contrast, the linear regression of the AD mice data gave us an equation of y = -0.035345 + 1.4311 with a significant negative relationship (p=0.0108). Within the wild type dataset, we excluded two RBC velocity points two standard deviations above the mean of the distribution. Even with these outliers, the p value for the wild type mice was not significant (p = 0.654). Preliminary analysis of age dependance of RBC velocities showed negative trend in AD mice with no statistical significance likely due to the small sample size and no change in wild type mice. We are currently analyzing more animals to increase the dataset size to improve the statistical power. One explanation for our findings shown on Fig 2 could be pericyte impairment since they control the capillary vascular tone and diameter. Pericytes were shown to be impaired in AD and this affects the CBF and small vessel contractility [4]. The lower velocity at the small vessel segments with slightly larger diameter may be due to loss of pericyte activity. Neuronal activity and immune responses to changes in RBC velocity can be also studied to show its effects on neurodegeneration and the link between neurovascular coupling and the immune system. In ischemic conditions where blood flow is limited and RBC velocity decreases, immune responses of microglia reduce their homeostatic functions [5]. Findings from this work will serve as a basis for our further analyses to include other vulnerability factors such as sex, age and microglial activity around the vessels. Since multiple factors contribute to the degeneration of the brain in Alzheimer’s, quantitative image analysis of vascular function such as RBC flow and capillary diameter allow for identifying subtle early brain changes and can help AD detection and therapy development.

Acknowledgements (Optional):  1. Alzheimer's Association (2024). Alzheimer’s disease facts and figures.

2. N. Korte, et al., (2020) "Cerebral blood flow decrease as an early pathological mechanism in Alzheimer's disease," Acta Neuropathol 140, 793-810

3. P. Drew, et al., (2010). Rapid determination of particle velocity from space-time images using the Radon transform. Journal of comp neuroscience, 29(1-2), 5–11.

4. C. Hall, et al., (2014). Capillary pericytes regulate cerebral blood flow in health and disease. Nature, 508(7494), 55–60.

5. T. Masuda, et al., (2011). Capillary blood flow around microglial somata determines dynamics of microglial processes in ischemic conditions. Glia, 59(11), 1744–1753.