Bioanalytic Core

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PROJECT SUMMARY (Bioanalytic Core, Sanchez) The Bioanalytic Core is a multifunctional core that will support the research efforts of each of the Projects by providing histological services, developing new innovative techniques and assays, and by training members of the Center to improve their own technical skills. The Aims of the Bioanalytic Core are: (1) To Provide basic neurohistology services for verification of probe placement or injection sites for Projects 1-4, including brain sectioning, histological staining, and immunohistochemistry for viral vector detection, and microscopy for reconstruction of recording and injection sites. (2) To provide assay services and to develop new techniques for molecular characterization of specific cell types, and for validating novel CRISPR viral vectors for Projects 1-4. These services include OXTR autoradiography, and the highly sensitive RNAScope mRNA in situ hybridization (ISH) technique to co-localize mRNA in cells for the purpose of molecular phenotyping. This technique will allow for the co-localization of mRNA for OXTR with other neuromodulator systems such as dopamine D1 and D2 receptors or with choline acetyltransferase (ChAT). We will also develop PCR/sequencing or Western assays to validate viral vector mediated CRISPR editing of OXTR or OT gene for Projects 1 and 3. Finally, the core will perform genotyping for OXTR polymorphisms in prairie voles in support of Projects 1 and 2. (3) To provide training in the techniques used in the Core to make sure that students, postdoctoral fellows, staff and trainees of the Center gain experience in basic histology and staining, autoradiography, ISH, IHC, neuroanatomy and fluorescent microscopy, which will be useful for their future career development. The services of the Bioanalytic Core will facilitate progress in every project and will serve as the center memory for techniques as trainees come and go. We will also develop new assays as needed and will perform basic neurohistology with standardization across projects while freeing up time for trainees to make progress in their own projects. The Bioanalytic Core is supported by the strong neuroanatomical expertise of the PI (Sanchez) in mapping neuropeptide and steroid receptors in the primate brain, studies performed in collaboration with the Center PI (Young), who has additional expertise with OT receptor systems both in rodent and primate brain. Co-investigator Kiyoshi Inoue brings critical extensive expertise in situ hybridization using RNAScope, OXTR autoradiography, general histology for probe anatomical verification, immunohistochemistry, and genotyping the prairie vole OXTR gene. These services will facilitate the integrated research programs of the Conte Center by providing unified and standardized services for each of the Projects, and allow investigators for the first time to identify the molecular phenotype of neurons activated by the OT system, and facilitate validation of new techniques such as optogenetic transgene expression and CRISPR editing of the OT system. These services will streamline the research activity of the Projects to maximize their productivity, while also providing valuable training on each of the techniques used to all Conte personnel who wish to learn these techniques.