Patrick J. Drew - US grants

DESCRIPTION (provided by applicant): Changes in spontaneous and sensory evoked cerebral blood flow are extensively used to infer neural activity in the brain with functional magnetic resonance imaging (fMRI). However, the relationship of these hemodynamic changes to neural activity, the pathways by which neural activity controls blood flow, and the interaction between sensory-evoked and spontaneous activity are still poorly understood. To address these questions, we will use 2-photon laser-scanning microscopy (2PLSM) and intrinsic optical signal (IOS) to measure spontaneous and sensory evoked hemodynamics in the somatosensory cortex of awake, head-fixed mice. We will measure the relationship between both sensory evoked and spontaneous neural activity and the subsequent blood flow ('neurovascular coupling'). The signaling pathways that underlying this coupling will be dissected by measuring neural activity and blood flow in transgenic mice lacking muscarinic cholinergic receptors on either blood vessels or cortical neurons. This will allow us to disentangle the direct vasodilatory effects of acetylcholine from the indirect effects mediated via increases in neural excitability. Finally, we will determine how sensory evoked activity interacts with ongoing spontaneous activity in the brain. Quantifying how spontaneous and sensory-evoked neural activity couples to blood flow is critical for understanding the function of the healthy brain.

DESCRIPTION (provided by applicant): The goal of this project is to quantify how natural behaviors modulate hemodynamic signals in the cortex, and to measure their coupling to neural activity. We will concurrently measure neural activity and hemodynamic signals, using electrophysiology, intrinsic optical signal imaging and two-photon laser scanning microscopy, in the cortex of awake, head-fixed mice. Mice will be free to alternate between running, grooming or quiescent behaviors on top of a spherical treadmill. We will use pharmacology to dissect the relative contributions of central neural and peripheral cardiovascular mechanisms in controlling cerebral blood flow. The neurovascular impulse response function, which defines the relationship between neural activity and blood flow, will be quantified across behaviors to test the constancy of neurovascular coupling. Lastly, we will test whether functional networks, areas with correlated blood flow during rest, are similarly correlated during locomotion. Understanding if, and how, the behavioral state modulates the cortical hemodynamic response, the coupling of blood flow to neural activity, and functional connectivity, are all critical for the interpretationof hemodynamic signals.

? DESCRIPTION (provided by applicant): Understanding the relationship between neural activity and cerebral blood flow is critical for interpreting hemodynamic signals, such as those measured with fMRI. It has long been assumed that blood flow to a brain region reported the average, or linear summation, of local neural activity. Recent work has cast this simplistic model into doubt. This proposal will use in vivo two-photon imaging, in close coordination with computational analysis methods, to distinguish between two alternative hypotheses of how neural activity is coupled to changes in blood flow. In one model, a 'democracy', blood flow is controlled by a linear sum of all neural activity. Alternatively, in an 'oligarchy', small groups o highly active neurons exert a disproportionate amount of control over blood flow, resulting in non-linear neurovascular coupling. Computational modeling will be used to test if the observed linear or non-linear coupling can be mechanistically explained by the production and diffusion of nitric oxide (NO). The proposed experiments will be performed in the olfactory bulb of rats, where discrete subpopulations of neurons (glomeruli) will be visualized and stimulated with odors. Two-photon microscopy will be used to simultaneously measure neural activity and blood flow in defined neural populations and single blood vessels. Targeted applications of drugs will be made to increase or decrease the neural activity in a single glomerulus. These experiments will be guided by real-time data analysis to determine the optimal stimulus or pharmacological perturbation in order to obtain a more accurate quantification of the linearity or nonlinearity of neurovascular coupling. In parallel, computational models will be constructed to test if the generation and diffusion of NO, a potent vasodilator, can account for the observed neurovascular coupling. This proposal is a collaboration between the labs of Dr. Serge Charpak, who has expertise using two-photon microscopy to simultaneously measure neural activity and blood flow changes in the olfactory bulb, and that of Dr. Patrick Drew, who has a background in computational neuroscience and has developed novel hemodynamic data analysis methods. The combination of these two approaches will yield a quantitative understanding of how blood flow changes relate to neural activity, and a determination of the mechanisms underlying neurovascular coupling. Hemodynamic signals, such as those measured by fMRI, are extensively used in inferring brain activity non-invasively, and being able to convert these hemodynamic signals into neural activity would be invaluable in diagnosing cognitive and neurological disorders. However, what specifically these changes in blood flow tell us about neural activity is not known. This proposal will result in a quantitative understanding of how neural activity is translated into hemodynamic signals, which will have immediate application to the interpretation of human imaging studies. This proposal will support undergraduates in mentored summer research projects, building on Dr. Drew's track record of mentoring women and underrepresented minorities in undergraduate research. The results will be incorporated into an interdisciplinary undergraduate class taught by Dr. Drew, Physical principles of living organisms, which applies physics and engineering principles to the study of biological systems.

Abstract: Hemodynamic signals allow us to non-invasively assay neural activity and could provide great insight into neural changes during postnatal development. However, it is not clearly understood how neural activity is related to changes in blood flow and oxygenation in the neonatal and juvenile brain. Previous studies in anesthetized animals and sedated humans have come to conflicting results as to the sign and magnitude of neurovascular coupling, and this unresolved issue has stalled the use of hemodynamic imaging in infants and children. In this proposal, we will determine how neurovascular coupling changes during postnatal development in the somatosensory cortex of the awake mouse brain, how this change in neurovascular coupling impacts BOLD fMRI signals, and how behavioral state can alter neurovascular coupling. We will use a multimodal approach, combining optical imaging (intrinsic optical signal imaging and 2-photon microscopy), electrophysiology, and fMRI to elucidate the relationship of hemodynamic signals to neural activity from the levels of single vessels up to the whole brain. We will mechanistically dissect the roles of local neural activity and cardiovascular effects on hemodynamic signals. The research proposed here will enable the use of hemodynamic imaging to study neural activity, plasticity, and neurodevelopmental disorders in infants and juvenile humans and animals.

In the course of its normal function, the brain produces toxic substances that accumulate and are transported from the space between brain cells. If these substances are not cleared, their accumulation is thought to yield crippling results such as Alzheimer's disease and migraines. The mechanics of this clearance is poorly understood, so this research project aim to study and characterize this process. Experimental techniques and computational approaches are being combined to produce a predictive clearance model based on fundamental mechanics principles of fluid flow and diffusion. The experimental study is being conducted in vivo, which will allow for a physiologically-relevant match between brain function and the corresponding deformation of brain tissue and the associated flow of the fluid in-between cells. This study is relevant for advancing the state of the art in neurophysiology and for future development of therapeutic interventions, both pharmacological and surgical, for addressing pathologies including Alzheimer's disease, hydrocephalus, and migraine. This project has an educational component aiming at training graduate and undergraduate students in advanced neuroscience research and in biomedical engineering. Specifically, the researchers and developing and offering a level-appropriate laboratory and computational projects for undergraduates with a focus on the merging of experimental techniques and mechanics in neuroscience.

This project focuses on delivering the first mechanics-based model of the effects of neurovasculature coupling on transport in the brain. A theoretical and computational framework is being created to model multiple concurrent transport mechanisms in a computational framework that integrates empirical in vivo observations of the brain micromechanical neurovascular response to chosen stimuli. The biomedical problem motivating the proposed research is the comparative assessment of convective and diffusive mechanisms for toxic metabolite clearance from the brain interstitium. Buildup of these compounds can be strongly neurotoxic and can trigger neuronal functional instabilities with severe, if not lethal, consequences---from spreading depolarization to epilepsy to Alzheimer's disease to mental illnesses. While vital for brain function, metabolite transport and clearance remains poorly understood. The specific project goals are: 1) To model brain tissue as a deformable porous medium with embedded vasculature, and to apply a numerical scheme developed by the PIs for predicting transport driven by blood vasodilation; 2) To identify sets of relevant physiological conditions from the experiments, and, from these, to define corresponding metabolite transport boundary value problems. Pulsation (heart-gated blood vessel dilation) and functional hyperemia (neurovascular coupling driven vessel dilation) will be considered. Anatomical, material, and loading parameters will be inferred using in vivo two-photon microscopy in the brains of living mice with cranial windows. Fluorescence-based digital image correlation will deliver microscale deformation maps of brain tissue. Fluid flow in the brain will be visualized by infusing fluorescent dyes; 3) To numerically solve the problems in goal 2 to determine interstitial fluid flow and metabolite transport through deformable tissue with convection and diffusion as concurrent mechanisms. Ranges of physiological conditions and constitutive parameters are being tested, and fluid-structure interaction between tissue and fluid-filled paravascular space are being explicitly modeled. The high selectivity of the blood-brain barrier remains a major challenge in developing effective drug delivery methods for brain cancer, dementia, spreading depolarization, and epilepsy. By focusing on metabolite transport in brain, this research project will contribute to advancing pharmacological and surgical therapies for many brain pathologies.

Project Summary The primary goal of this application is to elucidate the neural basis of resting-state functional magnetic resonance imaging (rsfMRI) signal using multi-modal approaches including multi-echo (ME)- rsfMRI, MR-compatible calcium signal recording, optogenetics and multi-laminar electrophysiology in awake rats. Despite the prominent role of rsfMRI in studying brain network function in health and disease, the neural basis of rsfMRI signal remains poorly understood. In particular, cellular and circuit-level mechanisms underlying resting-state functional connectivity (RSFC) are unknown. This critical knowledge gap has hampered the interpretation of rsfMRI signal in light of underlying neuronal activity. Here, we propose an integrated strategy to mechanistically dissect the neural basis of RSFC by combining ME-rsfMRI, cell-type specific calcium-based fiber photometry, optogenetics and electrophysiology. Specifically, ME-rsfMRI can differentiate neural and non-neural components in rsfMRI signal. In Aim 1, we will apply ME-rsfMRI to obtain precise RSFC quantification by eliminating non-neural artifacts. Second, calcium-based fiber photometry is an optical method that directly measures spiking activity from a defined neuron population (e.g. excitatory neurons) based on a genetically encoded calcium indicator (GCaMP). Thus, combining calcium-based fiber photometry with rsfMRI offers simultaneous neuronal and rsfMRI signal measurement with neuron-type specificity. In Aim 2, we will use this technique to dissect the contributions of spiking activity from individual neuron populations to rsfMRI signal. This study will provide critical insight into the cellular mechanism underlying rsfMRI signal. We will also determine how cell-type specific calcium signals link to the local field potential and spiking activity measured by electrophysiology. In Aim 3, we will integrate optogenetics, rsfMRI and multi-laminar electrophysiology to determine the causal relationship between RSFC modulation and layer-specific neural activity change. We will modulate RSFC by optogenetically increasing neuronal excitability using Stabilized Step-Function Opsin (SSFO), and examine the resulting RSFC and layer-specific electrophysiology signal changes. Linking multi-laminar electrophysiology and rsfMRI data will bridge mesoscopic scale neuronal activity at each cortical layer and large- scale cortico-cortical connectivity. Therefore, this study will help reveal the circuit-level mechanism of RSFC. Critically, to avoid influences of anesthesia, all multi-modal data will be measured in awake rats, which also enables direct translation of our results to human rsfMRI studies. Together, the proposed research will provide critically needed knowledge that can bridge neuronal and hemodynamic activities at rest across wide spatiotemporal scales. Such knowledge so far is impossible to gain from human studies, but will directly impact our understanding of human rsfMRI research. Finally, given the high clinical relevance of rsfMRI, the proposed research will be an essential step toward our long-term goal of establishing rsfMRI as a noninvasive tool for aiding diagnosis and/or evaluating treatment options for brain disorders.

Abstract Changes in neural activity drive changes in local blood flow in the brain. These changes in cerebral hemodynamics are important for maintaining normal brain health, and are used for non-invasively inferring neural activity. However, hemodynamic signals can decouple from local neural activity, making the understanding of the mechanism of this decoupling critical for interpreting and decoding hemodynamic signals. We hypothesize that hemodynamic signals in a given brain area are controlled by both vasodilatory signals released by local neurons, and by vasoconstrictory neuromodulation, specifically noradrenaline, both of which will vary with behavioral state, such locomotion. We will mechanistically test this hypothesis using optical imaging (intrinsic optical signal imaging and 2-photon microscopy), local pharmacological infusions, chemogenetics, oxygen polarography, and electrophysiology in awake mice. These experiments will elucidate how noradrenergic modulation, which is involved in alertness and attention, interacts with local neural activity to generate changes in blood flow and oxygenation. The end result of these experiments will be a unified understanding of how neural activity and neuromodulation control hemodynamic signals during behavior. 1