Optogenetic Interrogation and Manipulation of Vascular Blood Flow in Cortex

Understanding blood flow regulatory mechanisms that correlate the regional blood flow with the level of local neuronal activity in brain is an ongoing research. Discerning different aspects of this coupling is of substantial importance in interpretation of functional imaging results, such as functional magnetic resonance imaging (fMRI), that rely on hemodynamic recordings to detect and image brain neuronal activity. Moreover, this understanding can provide insight into blood flow disorders under different pathophysiological conditions and possible treatments for such disorders. The blood regulatory mechanisms can be studied at two different; however, complementary levels: at the cellular level or at the vascular level. To fully understand the regulatory mechanisms in brain, it is essential to discern details of the coupling mechanism in each level. While, the cellular pathways of the coupling mechanism has been studied extensively in the past few decades, our understanding of the vascular response to brain activity is fairly basic. The main objective of this dissertation is to develop proper methods and instrumentation to interrogate regional cortical vasodynamics in response to local brain stimulation. For this purpose we offer the design of a custom-made OCT scanner and the necessary lens mechanisms to integrate the OCT system, fluorescence imaging, and optogenetic stimulation technologies in a single system. The design uses off-the-shelf components for a cost-effective design. The modular design of the device allows scientists to modify it in accordance with their research needs. With this multi-modal system we are able to monitor blood flow, blood velocity, and lumen diameter of pial vessels, simultaneously. Additionally, the system design provides the possibility of generating arbitrary spatial stimulation light pattern on brain. These abilities enables researchers to capture more diverse datasets and, eventually, obtain a more comprehensive picture of the vasodynamics in the brain. Along with the device we also proposed new biological experiments that are tailored to investigate the spatio-temporal properties of the vascular response to optical neurostimulation of the excitatory neurons. We demonstrate the ability of the proposed methods to investigate the effect of length and amplitude of stimulation on the temporal pattern of response in the blood flow, blood velocity, and diameter of the pial vessels. Moreover, we offer systemic approaches to investigate the spatial characteristics of the response in a vascular network. In these methods we apply arbitrary spatial patterns of optical stimulation to the cortex of transgenic mice and monitor the attributes of surrounding vessels. With this flexibility we were able to image the brain region that is influenced by a pial artery. After characterizing the spatio-temporal properties of the vascular blood flow response to optical neuro-modulation, we demonstrate the design and application of an optogenetic-based closed-loop controller mechanism in the brain. This controller, uses a proportional–integral–derivative (PID) compensator to engineer temporal optogenetic stimulation light pulses and maintain the flow of blood at various user defined levels in a set of selected arteries. Upon tuning the gain values of the PID controller we obtained a near to critically-damped response in the blood flow of selected arterial vessels

Design and Implementation Guidelines for a Modular Spectral-Domain Optical Coherence Tomography Scanner

In the past decades, spectral-domain optical coherence tomography (SD-OCT) has transformed into a widely popular imaging technology which is used in many research and clinical applications. Despite such fast growth in the field, the technology has not been readily accessible to many research laboratories either due to the cost or inflexibility of the commercially available systems or due to the lack of essential knowledge in the field of optics to develop custom-made scanners that suit specific applications. This paper aims to provide a detailed discussion on the design and development process of a typical SD-OCT scanner. The effects of multiple design parameters, for the main optical and optomechanical components, on the overall performance of the imaging system are analyzed and discussions are provided to serve as a guideline for the development of a custom SD-OCT system. While this article can be generalized for different applications, we will demonstrate the design of a SD-OCT system and representative results for in vivo brain imaging. We explain procedures to measure the axial and transversal resolutions and field of view of the system and to understand the discrepancies between the experimental and theoretical values. The specific aim of this piece is to facilitate the process of constructing custom-made SD-OCT scanners for research groups with minimum understanding of concepts in optical design and medical imaging

Electrical Neural Stimulation and Simultaneous <i>in Vivo</i> Monitoring with Transparent Graphene Electrode Arrays Implanted in GCaMP6f Mice

Electrical stimulation using implantable electrodes is widely used to treat various neuronal disorders such as Parkinson’s disease and epilepsy and is a widely used research tool in neuroscience studies. However, to date, devices that help better understand the mechanisms of electrical stimulation in neural tissues have been limited to opaque neural electrodes. Imaging spatiotemporal neural responses to electrical stimulation with minimal artifact could allow for various studies that are impossible with existing opaque electrodes. Here, we demonstrate electrical brain stimulation and simultaneous optical monitoring of the underlying neural tissues using carbon-based, fully transparent graphene electrodes implanted in GCaMP6f mice. Fluorescence imaging of neural activity for varying electrical stimulation parameters was conducted with minimal image artifact through transparent graphene electrodes. In addition, full-field imaging of electrical stimulation verified more efficient neural activation with cathode leading stimulation compared to anode leading stimulation. We have characterized the charge density limitation of capacitive four-layer graphene electrodes as 116.07–174.10 μC/cm² based on electrochemical impedance spectroscopy, cyclic voltammetry, failure bench testing, and <i>in vivo</i> testing. This study demonstrates the transparent ability of graphene neural electrodes and provides a method to further increase understanding and potentially improve therapeutic electrical stimulation in the central and peripheral nervous systems