PROJECT SUMMARY Our knowledge of signal processing in various parts of the human brain has been heavily influenced by non- invasive functional magnetic resonance imaging (fMRI) experiments. FMRI infers the location and selectivity of neural activity from vascular signals. However, brain circuits are much more complex than regional differences in neuronal selectivity. Specifically, the largest part of the brain (neocortex) accounts for up to 80% of the brain volume and is divided into six distinct layers. Specific computations, e.g., local processing vs. feedforward inputs vs. vs. feedback inputs, are done in specific cortical laminae. Thus, if high-resolution layer-specific fMRI is shown to reflect the repertoire of neural computations performed across these cortical layers, it would be an invaluable refinement to non-invasive imaging. However, despite the widespread usage of low-resolution fMRI, a detailed understanding of how neural activity generates vascular responses remains unknown. The goal of this project is to elucidate the link between neural and vascular signals across laminae by combining two-photon imaging of neural and vascular responses with ultra-high-field (UHF) fMRI. Experiments will use sensory visual stimuli that induce layer-specific responses. In cat primary visual cortex (V1), which has a functional architecture (e.g., maps for stimulus orientation) similar to human V1, we will measure neural activity (synaptic and spiking) with single-cell resolution together with vascular signals (blood flow, blood volume, and oxygenation) in individual vessels across the entire cortical thickness. We will also perform UHF lamina-specific fMRI in cat (9.4 and16.4 T) and human (7 and 10.5 T) V1 to relate fMRI signals to the single- vessel responses. Lastly, we will develop a model to relate lamina-specific vascular signals to neural activity. In Aim 1, we test the hypothesis that vascular signals selective for stimulus orientation are present in cortical layers 2/3 (and 5/6) while untuned responses occur in layer 4 and pial vessels. Grating visual stimuli will be used, while varying orientation and eye preference (ocular dominance) systematically. Since binocular integration is stronger outside layer 4, eye preference vascular signals should be most prominent in layer 4. In Aim 2, we will test the hypothesis that in any given cortical lamina, glutamate release in regions around an individual blood vessel best accounts for the selectivity of vascular responses compared to spiking activity—in terms of the preferred stimulus orientation and tuning width. Aim 3 is to build a computational model to determine effective minimum voxel size for BOLD fMRI. The model will be tested against simultaneously measured vascular and neural activity to natural scene stimuli using two-photon imaging. If the source signals at the finest spatial scales have laminar specificity, we can correlate laminar-specific fMRI signals to differences in neural processing. To our knowledge, this is the first study that brings together such a wide repertoire of approaches into a single project to understand the neural and laminar basis of fMRI.

PROJECT NARRATIVE Much of our knowledge of human brain function will continue to come from functional magnetic resonance imaging (fMRI) experiments, which infers the location and selectivity of neural activity from vascular signals. Therefore, a detailed understanding of how neural activity generates selective vascular responses is critical to interpreting these data in healthy subjects and in neurological disease, including Alzheimer's, Parkinson's, diabetes and stroke, where neurovascular coupling is thought to become defective. This BRAIN initiative R01 award will support research on the spatial scale of neurovascular coupling in the living brain across imaging modalities using an animal model system and human subjects.