Histology Core

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Summary/Abstract, Core C: Histology The overall goal of this U19 application is to determine the brainwide neural basis and behavioral consequences of internal states, such as task engagement, as mice perform a standardized decision-making task. Registration of electrophysiological recording sites into a common anatomical framework represents a critical step in comparing these experiments across projects and synthesizing the results into a theoretical model, so an accurate and robust histological and image analysis methodology is essential for the development of high-quality, reproducible datasets. Histology Core C therefore aims to deliver two key outcomes. First, this core will centralize and standardize histology data acquisition and registration to a reference atlas across experimental Projects 1, 2, and 4. Through previous work by groups participating in this proposal, the anatomical reconstruction and registration pipeline for recordings with Neuropixels probes using serial-section two-photon imaging has been established, optimized, and validated. This core will develop an open-source software package for sample registration to the Allen mouse brain common coordinate framework, version 3 (CCF). This package will include functions for assessing registration quality, and will be extended to handle new experimental recording methods as needed for the proposed experiments, including the reconstruction and registration of labeled cells investigated with calcium imaging, the registration of functional ultrasound imaging data onto the Allen CCF, and the reconstruction of cells, pathways, and optic fiber insertion sites in fiber photometry and optogenetic stimulation experiments. Second, to support the additional experimental aims proposed in this application, particularly optogenetic perturbations in Project 2 and calcium imaging in Project 4, this core will develop clearing and histological labeling protocols to interrogate nervous system structure. Many of the technical hurdles to delivering these aims have already been overcome. The centralizing of histological processing in the International Brain Laboratory consortium is well established, and a first version of this open-source software is in its final stages of development. A whole-brain tissue clearing and lightsheet imaging methodology has been demonstrated, and its reliable registration to the common framework is under assessment. A range of histological procedures that can immunohistochemically label large blocks of tissue are available, and will form the basis for further histological interrogation. In conclusion, Histology Core C will perform the essential tasks of processing mouse brains for histological reconstruction, generating structural datasets from this tissue in close collaboration with U19 projects, and supplying the resulting protocols, software tools, and datasets to the wider community.