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© Copyright 2020 Adiya Rakymzhan © Copyright 2020 Adiya Rakymzhan Studying Cerebral Blood Flow in Mouse Cortex with Optical Microangiography Adiya Rakymzhan A thesis submitted in partial fulfillment of the requirements for the degree of Master of Science University of Washington 2020 Committee: Ruikang K. Wang Azadeh Yazdan-Shahmorad Program Authorized to Offer Degree: Bioengineering University of Washington Abstract Studying Cerebral Blood Flow in Mouse Cortex with Optical Microangiography Adiya Rakymzhan Chair of the Supervisory Committee: Professor Ruikang K. Wang Bioengineering Department Optical coherence tomography (OCT) is a non-contact three-dimensional (3D) imaging modality which enables fast in vivo volumetric visualization of internal structures of biological tissues with high resolution. In this thesis, first, the application of OCT advanced extension - optical microangiography (OMAG) - was used to reveal the temporal profile of hemodynamic response to neural activation and the variation of depth-resolved blood flow changes in mouse cerebral cortex. The results showed that the largest hemodynamic response occurred in cortical layer IV. Second, OMAG and Doppler OMAG (DOMAG) techniques were applied to quantitatively evaluate the anesthesia effect on cerebral vessel morphology and blood flow in mouse model. According to the results, isoflurane and ketamine-xylazine caused vasodilation both in large and capillary vessels, which led to increased CBF. Overall, this thesis demonstrated that OMAG is an effective tool for studying depth-resolved microvasculature and blood flow in animal cortex. TABLE OF CONTENTS Chapter 1. INTRODUCTION ......................................................................................................... 1 1.1 Significance and research motivation ................................................................................... 1 1.2 Current methods and scientific gap ....................................................................................... 1 1.3 The advantages of optical coherence tomography (OCT) .................................................... 2 1.4 The scope of the thesis .......................................................................................................... 2 Chapter 2. METHODS.................................................................................................................... 4 2.1 ISOI ....................................................................................................................................... 4 2.2 OCT ....................................................................................................................................... 5 2.2.1 OMAG ............................................................................................................................ 8 2.2.2 DOMAG ......................................................................................................................... 8 Chapter 3. OMAG REVEALS TEMPORAL AND DEPTH-RESOLVED HEMODYNAMIC CHANGE IN MOUSE BARREL CORTEX DURING WHISKER STIMULATION ................ 10 3.1 Background and motivation ................................................................................................ 10 3.2 Materials and methods ........................................................................................................ 11 3.2.1 Animal preparation ....................................................................................................... 11 3.2.2 IOSI .............................................................................................................................. 12 3.2.3 OMAG .......................................................................................................................... 12 3.2.4 Stimulation.................................................................................................................... 13 3.2 Results ................................................................................................................................. 13 i 3.3.1 Reflectance signal change upon functional activation ................................................. 13 3.3.2 OMAG signal change upon functional activation ........................................................ 15 3.3.3 OMAG signal variation over depth .............................................................................. 16 3.4 Discussion ........................................................................................................................... 18 3.5 Conclusion ........................................................................................................................... 21 Chapter 4. OMAG REVEALS HEMODYNAMIC DIFFERENCE AT THREE DIFFERENT REGIMES: AWAKE, ISOFLURANE, KETAMINE/XYLAZINE ............................................. 23 4.1 Background and motivation ................................................................................................ 23 4.2 Materials and methods ........................................................................................................ 24 4.2.1 Animal preparation ....................................................................................................... 24 4.2.2 Awake imaging ............................................................................................................. 24 4.2.3 Anesthetized imaging ................................................................................................... 25 4.2.4 Imaging timeline ........................................................................................................... 25 4.2.5 OMAG .......................................................................................................................... 25 4.2.6 DOMAG ....................................................................................................................... 26 4.3 Data analysis ....................................................................................................................... 26 4.3.1 Vein and artery diameter measurement ........................................................................ 26 4.3.2 Capillary density and flux quantification ..................................................................... 27 4.3.3 CBF analysis ................................................................................................................. 28 4.3.4 Statistical analysis......................................................................................................... 28 ii 4.4 Results ................................................................................................................................. 28 4.4.1 Vessel dilation .............................................................................................................. 28 4.4.2 Capillary density and flux ............................................................................................. 30 4.4.3 CBF measurements from penetrating vessels ............................................................... 32 4.5 Discussion ........................................................................................................................... 35 4.6 Conclusion ........................................................................................................................... 37 Chapter 5. SUMMARY ................................................................................................................ 39 Bibliography ................................................................................................................................. 40 iii Chapter 1. INTRODUCTION 1.1 Significance and research motivation A regional increase in cerebral blood flow (CBF) during local neural activation is associated with neurovascular coupling (NVC), which is also referred to as functional hyperaemia.[1] NVC is a vital process, which regulates oxygen delivery through blood stream to supply the activated neurons for normal brain functioning.[1] Therefore, studying the mechanisms of CBF regulation at NVC is crucial for the improved understanding of brain activity under normal and neurodegenerative conditions. 1.2 Current methods and scientific gap Number of imaging modalities have been applied to study CBF. Traditional imaging methods such as functional magnetic resonance imaging (fMRI) [2] and positron emission tomography (PET) [3] are limited in spatial and temporal resolution to detect hemodynamic responses. Optical imaging techniques offer a broad range of contrast mechanisms and can be readily applied to animal studies to measure CBF parameters. However, assessing all these parameters with a single technique is still challenging. For instance, laser Doppler imaging (LDI) is great at evaluating blood flow, but has to compromise between spatial and temporal resolution. [4] Photoacoustic microscopy (PAM) can provide information about vascular morphology and flow, but requires physical contact with the object. [5] Intrinsic optical signal imaging (IOSI) [6] and laser speckle imaging (LSI) [7] are excellent full-field imaging tools for measuring CBF and blood oxygenation. Although the given technologies are widely used, their detection is two-dimensional (2D) with no 1 access to deep-layer structures. Three-dimensional (3D) imaging methods, such as two-photon laser scanning microscopy (TPM) [8-10] and confocal laser scanning microscopy (CLSM) [11] have been proven effective in studying neurovascular dynamics. However, they both cannot have sufficient field of view (FOV)
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