University Of Minnesota

NIH Research Projects · FY 2024 · 2020-08

Research Summary/Abstract: The overall goal of the Grenning research lab is to develop programable and highly diversifiable platforms for accessing complex chemical space of interest to drug discovery. Summarized herein is an overview of work and future directions aimed at transforming the classic Cope rearrangement into a versatile synthetic transformation of high value for complex molecule synthesis. While variants of the Cope rearrangement (e.g. the oxy-Cope rearrangement, aza-Cope rearrangement, and divinylcyclopropane Cope rearrangement) have found extensive use and value in modern chemical synthesis, the classic Cope has not. This transformation is a diamond in the rough that, post-MIRA funding, will have clear and diverse value to chemical synthesis and drug discovery. In our published and unpublished works, we have addressed (or are addressing) fundamental challenges related to thermodynamics and kinetics and proposed potential applications of this transformation in modular complex molecule synthesis. Our current and future directions will involve continuing to improve our understanding of this transformation, develop unique, complexity generating transformations and/or sequences where this transformation plays a key role, introduce a variety of catalytic-asymmetric methods for accessing enantioenriched building blocks, and prepare molecules of modern interest to drug discovery; a well-rounded and diverse research program focusing both on fundamental and applied chemical discoveries. Regarding the latter goal, we currently have on going collaborations with many medicinal chemistry and chemical biology groups and will continue to make new connections allowing for the most impactful discoveries related to synthesis and medicine. Funding of this proposal will result in new and general transformations of value beyond the scope of the proposal and new leads for drug discovery. For example, we have already established a highly modular route to Vorinostat analogs and collaboratively (with the Pflum lab at Wayne State University) will examine their bioactivity as HDAC inhibitors. Beyond or chemistry products (methods, syntheses, and molecules), we are requesting significant funding for the training of students and postdocs to professional synthetic chemists which will be of critical value to a knowledgeable, scientific workforce of value to a variety of technical industries. For example, Ph.D. graduates from my lab are currently continuing their studies as post docs (e.g. Primali Navaratne; Stoltz Lab) or have gone directly into industry (Ehsan Fereyduni; Research Scientist at Intel). ! 1!

NIH Research Projects · FY 2024 · 2020-08

Project Summary Every 5 minutes, a patient is denied an MRI scan because of an active implanted medical device. Therefore, there is a clinical need to improve the safety of MRI scans to allow patients with implants to be imaged. Ability to safely scan patients with implants would have an overall public health impact. If we can develop a safe framework to be used to image these patients, we will be able to use MRI to monitor the stage of their diseases and the efficacy of their treatment. In this project we aim to develop a workflow that utilizes multi-channel and receiver arrays in order to safely image patients with Deep Bran Stimulators (DBS). We will utilize a pre-scan strategy (accelerated by parallel acquisition) to calculate RF current induced on DBS leads and predict electrode tip temperature. We are going to calculate/design implant friendly excitation solutions/pTx pulses and utilize them to reduce RF heating around the DBS electrodes. We will demonstrate the temperature reduction around the DBS leads using fluoroscopic probes and evaluate image quality. We will test the performance of our approach by scanning phantoms, anesthetized animals (swine) and human cadavers. We will finally scan human subjects implanted with full DBS systems (electrode, extension, IPG) using the methods developed in this proposal.

NIH Research Projects · FY 2026 · 2020-08

Project Summary Cells have evolved intricate enzymatic machineries that help them exist and survive redox stresses in their microenvironment. Enzymatic redox sensing, signaling, and response mechanisms are critical for a diverse set of physiological processes in all forms of life ranging from bacteria and plants to humans. Unlike other cellular signaling processes, redox signaling involves highly reactive reagents such as nitric oxide (NO), carbon monoxide (CO), and reactive oxygen species that raise concerns regarding the potential of these reagents for participation in other non-specific reactions. Yet, such side reactions are uncommon under physiological conditions suggesting high specificity and selectivity of enzymatic redox signal transduction pathways. In turn, we ask the following two pertinent questions: a) What makes redox signaling pathways so specific? and b) Can we rationally and systematically reprogram redox signal transduction pathways to re-instate/disrupt cellular redox balance? These questions have been largely overlooked from biological and inorganic chemistry perspective, even though redox imbalances are responsible for a variety of diseases ranging from neurological disorders to cancer. Our lab focuses on these paradigm shifting questions and the long-term goal of our research program is to develop molecular strategies that reprogram sensing/signaling mechanisms of biological redox reagents involved in human health and disease. In this grant period, we focus on heme iron-based DosS and DosT enzymes that sense low O2 or NO or CO in the bacterial microenvironment to signal dormancy as a response. Using our combined expertise in metalloprotein structure-function studies, protein engineering, enzymology and spectroscopy, we will develop structure-guided protein design approaches to reprogram the redox stimuli sensitivity and specificity of these enzymes as well as understand the implications of such modulations on redox signaling and cellular behavior. We will also use cutting-edge structural biology techniques to investigate the structural/molecular basis of signal transduction in these enzymes. Finally, we will develop redox-selective activity-based probes to quantify active/inactive sensor levels in the cell. Overall, our findings will not only provide a fundamental understanding of cellular redox sensing and signaling mechanisms, but also inform on mechanisms to modulate cellular physiology and phenotypic responses. Health Relevance: Maintenance of a normal intracellular redox status is crucial for regulating physiological responses. Any imbalance in this status results in a variety of diseases such as cancer, cardiovascular and neurological disorders. Our research program aims to design molecular approaches that reprogram the ability of cells to sense and signal redox changes in their environment. These approaches could be applied to re-instate cellular redox homeostasis in diseased states.

NIH Research Projects · FY 2024 · 2020-08

Project Summary The mission of the Bridges to Biomedical Research Careers is to provide financial and academic support for community college students emerging from underserved populations in Minnesota and enable their pursuit of a baccalaureate degree by participating in quality research experiences to increase the diversity of future biomedical scientists. The program is a collaboration between the University of Minnesota Duluth (UMD) and the UM Medical School Duluth Campus (UMMSD) to provide financial and academic resources to ten science students enrolled at Fond du Lac Tribal and Community College (FDLTCC) in Cloquet, MN and Lake Superior College (LSC) in Duluth, MN. Additionally, the program will provide career counseling, faculty mentoring, and active learning projects to develop critical thinking and to engage in bench research to learn the craft of a scientist. The Baccalaureate degree awarding institution will be UMD, which provides academic, research, and community outreach to northern and central Minnesota. Program objectives are: 1) Identify candidates with an aptitude for inquiry based science; 2) Provide academic counseling to facilitate the trainees’ development; 3) Provide quality mentoring that nurtures trainee participation in research; 4) Provide active learning experiences to hone critical thinking; and 5) Provide experience in cutting-edge research to learn the tools of biomedical science. The expected outcomes are to complete an AA degree by three years and transfer to a university to complete a baccalaureate degree in a biomedical-related discipline. The goal is to produce graduates who self-identity as a biomedical scientist.

NIH Research Projects · FY 2024 · 2020-08

Modified Project Summary/Abstract Section Abstract Children with substantial African ancestry have long been known to have half or less the rate of B-cell acute lymphoblastic leukemia (B-ALL) than do children with other continental ancestries. This is true both in international comparisons of rates of ALL in African nations to those elsewhere, and in comparing rate of B-ALL in African-American (AA) children to that in European-American (EA) children in the United States. The inverse association of African ancestry with incidence of B-ALL is independent of established perinatal risk factors for the disease. Moreover, AA children have lower incidence despite having greater exposure to many putatively causal environmental risk factors for B-ALL than do EA children. Common genetic variants established by genomewide association studies incompletely explain the deficit of B-ALL in AA children, suggesting undiscovered contributing genetic factors may be detected by admixture mapping. We have assembled existing DNA samples and data for 930 B-ALL patients with AA ancestry and will additionally accrue ~590 over the life of the project. We will conduct admixture mapping in the assembled group of patients to detect new genetic loci and new variants at established loci associated with occurrence of B-ALL. In addition, we will examine admixture in association with clinical characteristics at diagnosis and survival. Candidate genes/variants will be functionally evaluated through both in silico and in vitro techniques. The proposed research will potentially answer a long- standing mystery by revealing critical genes or loci that explain the comparative deficit of B-ALL in AA compared to EA children. In addition, we may uncover genes or variants associated with the worse characteristics at presentation in AA patients as well as with worse survival, which will indicate avenues for improving outcome among AA children.

NIH Research Projects · FY 2024 · 2020-08

Weight-related problems are of great public health concern given their high prevalence, health consequences, and unequal distribution across ethnic/racial backgrounds and income levels. The Healthy Weight Promotion in Youth and Families from Diverse Communities Applied Epidemiology Training Program is designed to train future scientists for successful research careers aimed at ensuring positive weight-related health among youth and families from diverse communities. This program is unique in its combined focus on five areas: 1) comprehensively addressing the broad spectrum of weight-related problems, including obesity, poor dietary intake, low physical activity, body dissatisfaction, and disordered eating; 2) promoting positive weight-related health among children and families during critical periods of the life course; 3) achieving greater health by reducing disparities in weight-related problems across ethnic/racial and income levels; 4) engaging in both observational epidemiologic population-based studies and intervention/evaluation research (clinical, community, and policy-oriented); and 5) translating research into action within different settings. This program is designed to provide rigorous, innovative, and pragmatic research training. As a new program, the number of trainees will gradually increase: Year 1: 3 trainees (1 predoc and 2 postdoc); Year 2: 5 trainees (2 predoc and 3 postdoc); Years 3-5: 7 trainees (3 predoc and 4 postdoc). Predoctoral trainees will be funded for up to four years and will complete a PhD in Social and Behavioral Epidemiology. Postdoctoral trainees will be funded for up to three years. Training will promote interdisciplinary learning, collaborative experiences, innovative thinking, rigorous research skills, and leadership capacity to ensure successful and impactful research careers. Training includes: 1) individualized mentoring by experienced interdisciplinary faculty; 2) experiential learning through active involvement in observational and intervention research projects; 3) interdisciplinary professional training for leadership development; 4) formal class work; 5) peer group work; 6) deep dive workshops specifically designed to meet the goals of this training program; and 7) other rich learning opportunities across the University of Minnesota. Benchmarks of success include engagement in research on the broad spectrum of eating and weight-related health in youth and families from diverse communities; dissemination of findings to professional and public audiences; grant writing, successful course work, mentoring of others, and obtaining a research position. The 22 faculty mentors, come from varied disciplines, are well-funded, and have extensive experience conducting innovative and impactful research aimed at healthy weight promotion during critical stages of the life course in diverse populations. Faculty have mentored many successful predoctoral and postdoctoral trainees. The Program Director has an Outstanding Investigator Award from NHLBI that will provide relevant learning and research opportunities for trainees. This training program addresses a crucial need for preparing future scientists to reduce the high prevalence and disparities in weight-related problems.

NIH Research Projects · FY 2024 · 2020-07

PROJECT SUMMARY While tremendous progress has been made to understand mechanisms of epithelial cell injury and fibroblast activation in fibrosis, significant challenges remain in fully understanding the mechanisms of epithelial– mesenchymal crosstalk in normal lung homeostasis and during injury, repair and fibrosis. As in normal lung development, lung repair requires epithelial cells and mesenchymal cells to coordinate with each other to tune cell proliferation, migration, differentiation and apoptosis. Similarly, extracellular matrix (ECM) remodeling is mediated by a myriad of pro-fibrotic and anti-fibrotic factors that are precisely orchestrated by epithelium and mesenchyme during injury, repair and resolution. Therefore, aberrant epithelial mesenchymal interactions can lead to non-healing processes in lung repair with pathological scar formation due to myofibroblast accumulation and ECM deposition, ultimately contributing to pulmonary fibrosis. The long-term goal of this proposal is to restore epithelial-mesenchymal homeostasis in pulmonary fibrosis with transcriptional regulation of critical anti-fibrotic factors. Targeting mechanisms that augment the endogenous anti-fibrotic or fibrosis resolution signals during disease progression may serve as an attractive therapeutic strategy to alleviate lung fibrosis and restore lung function. Recently, single-cell and bulk RNA sequencing identified loss of normal epithelial cell identities and gain of abnormal indeterminate states of differentiation in IPF, with CEBPA identified as a key transcriptional regulator diminished in IPF that was common to multiple independent studies. Our central hypothesis is that CEBPA deficiency in lung epithelial cells promotes lung fibroblast activation and fibrosis, that it normally increases during repair to resolve fibrosis, and that it can be therapeutically restored using non-genome editing CRISPR gene activation to promote fibrosis resolution. This hypothesis will be tested with three specific aims: First, we will evaluate the role of Cebpa signaling from epithelial cells in maintaining lung epithelial-mesenchymal homeostasis and its protective role in pulmonary fibrosis. Second, we will determine whether increasing Cebpa expression with CRISPR gene activation is able to restore epithelial-mesenchymal homeostasis and attenuate fibroblast activation and fibrosis. We will develop and optimize an AAV-mediated approach to enhance Cebpa expression using non-genome editing CRISPR activation in the lung of aged mice that display non-resolving lung fibrosis after bleomycin injury. Lastly, We will test whether the anti-fibrotic effect of epithelial Cebpa expression is mediated by BMP4 and perform RNA-seq- based gene expression analysis of sorted epithelial cells from Cebpa gain and loss of function studies to identify additional novel candidate mediators of epithelial-mesenchymal homeostasis. Taken together, the proposed research studies will reveal critical anti-fibrotic targets for therapeutic interventions aimed at restoring epithelial-mesenchymal homeostasis and alleviating pulmonary fibrosis.

NIH Research Projects · FY 2024 · 2020-07

Summary With the recent advances in high-throughput biotechnologies and associated flooding of Big Data, there is an urgent demand and shortage of the next-generation PhD-level scientists who have strong biostatistical and computational skills to handle Big Data while understanding biology, especially genetics and genomics. However, traditional training in biostatistics, which emphasizes almost exclusively on statistical theory and methods, no longer meets the need; similarly, to meet Big Data challenge, traditional training in genetics could be expanded and strengthened with more training in biostatistics/data science. As a response, this application continues the development and refinement of an innovative and interdisciplinary pre-doctoral T32 “Biostatistics in Genetics and Genomics” (BiG2) training program. The primary mission of this training grant is to prepare Biostatistics, Genetics and other predoctoral trainees for leadership roles in biomedical research through excellent training and mentorship in both biostatistics and genetics/genomics. To meet the overarching objective specified by the NIGMS's RFA, our training program requires completion by each trainee of required and elective coursework in an affiliated PhD program with extra training in Biostatistics and/or Genetics, mentored learning through interdisciplinary research projects through lab rotations, development of critical thinking, communication and networking skills, attendance at and participation in journal clubs, seminars and national meetings, successful completion of a program of interdisciplinary research through their dissertation, and future career development, all of which give our trainees broad training in a rapidly expanding field. The most distinctive feature of the proposed training program is its interdisciplinary nature integrating both a biostatistical (or methodological/computational) aspect and a genetic/biomedical aspect. A key is to ensure each trainee learns how to carry out critical, reproducible, cohesive and interdisciplinary research program, including collaborating with various biomedical researchers. We request support for 4-5-6-6-6 slots in years 1-5 for predoctoral trainees from Biostatistics, Genetics and other affiliated PhD programs. Each trainee is to be funded for two years at the early stage of his/her PhD study, after which his/her support will switch to projects funded by the faculty mentors' research grants. As measurable outcomes, in a short term, we will examine the success rates of the trainee's timely completion of their PhD studies and their job placements; in a long term, it will be determined by the success and impact of their future careers in biomedical research; in between, it will be their publications, awards and career transitions around the beginning of their careers.

NIH Research Projects · FY 2025 · 2020-07

The Life Course Center (LCC) at the University of Minnesota (UMN) develops, supports, and coordinates innovative, high-quality, and transformative interdisciplinary research on the demography and economics of aging. LCC research addresses four research themes. (1) Later life population trends in context: the impact of both persistent and shifting macrosocial contexts for explaining changes across historical time and place in aging processes and outcomes, including physical and cognitive functioning, disability, morbidity, mortality, general health, and well-being. (2) Life course dynamics as disparity mechanisms: the processes by which social change and contexts—especially the impacts of early and cumulative life experiences—play out in the lives of population groups to reduce or exacerbate disparities in health outcomes. (3) Interrelationships among work, family, community participation, and health: the consequences of changing economic, familial, social, and institutional environments on life course engagement in paid work, family responsibilities (e.g., caregiving, living arrangements, engagement), and volunteering, as well as the implications for life chances, health, and life quality in the mid- and later-life course. (4) Health care services and supports for an aging population: the health and financial implications of the care infrastructure for older adults, which includes healthcare organizations, the healthcare workforce, payment models for care, and other policies for the care and support of aging populations. LCC research addressing these research themes focuses on four health disparity priority populations: (1) socioeconomically disadvantaged populations; (2) racial minority populations; (3) rural populations; and (4) sexual minority populations. The central goal of LCC is to provide an unparalleled environment for researchers to conduct impactful research to improve the health and well-being of older adults. This will be accomplished through four infrastructure Cores. Core A: Administrative and Research Support Core. This Core will provide leadership and vision, as well as logistical support and oversight for the activities of the other three Cores. This Core will also be responsible for meeting all reporting requirements to the Coordinating Center. Core B: Program Development (Pilot) Core. The Pilot Core will administer two pilot grant programs: the Emerging Scholar Pilot Projects and Mentoring Program and the Highly Innovative/Time Sensitive Pilot Program. Both will support small-scale, innovative interdisciplinary research projects. Core C: Communication and Dissemination Core. This Core will work with the other three Cores to promote the activities and achievements of LCC and its members. Core D: External Network Core. The Network for Data-Intensive Research on Aging (NDIRA) will foster an interdisciplinary community of scholars across disciplines, career stages, and institutions and introduce them to novel sources of data that are fundamental to understanding health and aging. NDIRA will address challenges associated with big population data and stimulate use of the novel data sources LCC members build for life course research.

NIH Research Projects · FY 2024 · 2020-07

DESCRIPTION (provided by applicant): The mission of the Midwest Center for Occupational Health and Safety (MCOHS) Education and Research Center (ERC), as a center of excellence, is to provide: 1) cutting-edge interdisciplinary academic and research training to prepare exceptional leaders who make significant contributions to occupational safety and health (OSH), and 2) continuing education (CE) to prepare professionals to address current and emerging threats to the nation's workforce. This proposal requests continued funding for the period, July 1, 2015 to June 30, 2020. Objective: To address the need for an adequate supply of qualified personnel to carry out the purposes of the Occupational Safety and Health Act and reduce the national burden of work-related injury and illness in the Midwest region served by the MCOHS, and beyond. Rationale: A previous conclusion in an Institute of Medicine report, that remains true, stated, "...the continuing burden of largely preventable occupational diseases and injuries and the lack of adequate occupational safety and health (OSH) services ...indicate a clear need for more OSH professionals at all levels." Further confirmation is noted in the recent NIOSH-commissioned report, "National Assessment of the Occupational Safety and Health Workforce," identifying needs that greatly exceed available trained OSH professionals. Design: An innovative administrative structure, guided by a strategic plan and committed advisory board, supports enhanced efforts in interdisciplinary research, education, and outreach, including research-to practice, and strengthens diversity recruitment. Rigorous graduate academic and research programs enable quality training in (degrees and projected numbers): Industrial Hygiene (PhD-11, MS-12, MPH-18); Occupational and Environmental Medicine (MPH-17); Occupational and Environmental Health Nursing (PhD- 3, MS-0, MPH-13); Occupational Health Services Research and Policy (PhD-10); Occupational and Environmental Epidemiology (PhD-8, MPH-2); and Occupational Injury Prevention Research (PhD-9); while program duration varies, program expectations are: masters' (~2 years); PhD (~ 4 years) In addition, a major CE Program offers novel courses in-person and through distance learning (projected =18,000) to meet the needs of a diverse workforce.

NIH Research Projects · FY 2024 · 2020-07

PROJECT SUMMARY Increasing evidence indicates that three-dimensional (3D) genome organization is required to regulate gene function and its alterations are associated with many diseases. The genome is organized into compartments that align with the temporal order of DNA replication (replication timing – RT). However, little is known about the mechanisms underlying 3D genome organization. Recently, we identified cis elements of RT and 3D genome organization control (early replicating control elements – ERCEs) in murine embryonic stem cells. ERCEs are enriched in enhancer epigenetic marks, form strong chromatin interactions and are bound by pluripotency-specific transcription factors. Moreover, I developed an integrative model of gene regulatory networks that predicts that co-occupancy of cell type-specific transcription factors regulate RT. Here, we will study what are the regulatory elements of genome organization in human differentiated cell types, investigate how trans-acting factors control these elements, and define how 3D genome organization is remodeled during development and evolution. Our central hypothesis is that co-occupancy of cell type-specific transcription factors at ERCEs is required to regulate the 3D genome organization. To test this hypothesis, we will delete and insert candidate ERCEs into the genome of human differentiated cell types and test their effect on RT, 3D genome organization and gene expression. We will track ERCE activation using highly-synchronous human embryonic stem cells differentiation systems. Finally, we will analyze 3D genome organization evolution using primary cells derived from different species. My laboratory is uniquely positioned to perform the proposed research with broad expertise in the experimental design and analysis of 3D genome organization during human cell fate commitment. Moreover, numerous resources available at the University of Minnesota will facilitate the success of this project, including state-of-the-art technologies for genome editing (Genome Engineering Shared Resource), next-generation sequencing (Genomics Center), stem cells (Stem Cell Institute), imaging (Imaging Center) and bioinformatic tools (Minnesota Supercomputing Institute). We expect that our work will contribute significantly to understand the fundamental principles of genome organization and its relationship to gene function. The proposed research combines several innovative aspects such as integrative computational models to predict regulatory elements of large-scale chromatin organization, genome engineering technologies and optimized differentiation protocols of human embryonic stem cells to dissect the mechanisms that control 3D genome organization during development and evolution.

NIH Research Projects · FY 2025 · 2020-07

Project Summary The goal of this project is to provide biostatistical leadership and data management infrastructure for the BLOODSAFE initiative, an NHLBI initiative which seeks to increase the supply of safe blood available for transfusion in Sub-Saharan Africa. This leadership includes coordination of activities across multiple projects, collaboration with scientists at NHLBI on program oversight and provision of expertise regarding all aspects of study design and implementation. This expertise includes but is not limited to coordination with local experts, provision of training to local experts, assistance with study design, harmonization of data elements across multiple studies, assistance with implementation of interventions, data management, data quality assurance, data analysis and study finding dissemination to ensure that discoveries are communicated to the scientific community and translated into local best practices in a timely manner. These goals will be fulfilled by completion of the following aims: provide study coordination activities and oversight across all studies at all stages of study development, including regulatory oversight, logistical support and training opportunities in conjunction with NHLBI; provide expertise in study design, initiation and implementation; provide expertise in data management including standardization of data elements, design of data collection instruments, development of data quality assurance systems and systems for specimen tracking; and provide expertise in all aspects of study reporting, data analysis and result dissemination and development of novel analysis strategies as required for fulfilment of study objectives.

NIH Research Projects · FY 2024 · 2020-07

Project Summary Age-related macular degeneration (AMD) typically produces extensive central vision loss, leading to difficulty in reading, navigating, and other critical tasks. Current assistive devices for vision with central scotomas, while promising, are limited in effectiveness. This proposal aims to fill this gap by advancing understanding of remapping, a method that improves the effectiveness of patients’ residual vision by shifting information from inside the central blind spot (scotoma) to intact locations in the visual field. Only a handful of past studies have examined remapping, including just a single study of patients. Our group recently demonstrated the method’s promise, showing that it can improve reading substantially in observers with simulated field loss. Critically, patients differ widely in the shape of their scotomas, and the quality of their vision across the visual field. This variability imposes severe limitations on the “one size fits all” remapping approaches used in prior work, that simply shift the image away from the scotoma center. If, for example, a patient’s preferred retinal location (used for high acuity tasks) is near the lower left edge of their scotoma, shifting text upward or rightward may not aid reading as much as shifting it downward, as the latter will place the text in regions of best visual acuity. This proposal will test the value of personalized remappings, which shift the image in a way that is optimized for each patient’s residual vision. The remappings are constructed using a novel letter recognition perimetry task, which measures performance across the visual field: The personalized remappings are constructed to shift text to maximize observers’ total letter recognition ability. Proposed work will first test the hypothesis that personalized remapping can improve single word recognition. Single word reading will be measured with a variety of scotoma sizes, shapes, and PRL locations. Performance with personalized remapping will be compared to that with traditional remapping and no remapping in both control observers with simulated scotomas and people with macular degeneration. Proposed work will also test the hypothesis that personalized remappings can improve free reading. Reading speed, error rates, and eye movement patterns will be measured in a sentence reading task, and in free reading of natural images containing text. Preliminary data support the value of remapping generally, and the potential of the letter recognition perimetry task for building personalized remappings. Results of this proposal will provide the first thorough testing of remapping, and will also inform models of peripheral reading in patients and controls. The proposed studies will develop and evaluate a novel method for assisting people with low-visions, tailored to their individual residual performance. Personlaized remapping may also be incorporated into practical visual aids to improve daily visual function and quality of life.

NIH Research Projects · FY 2026 · 2020-07

Abstract A living kidney donor (LD) undergoes a major operation with potential risks of living with a single kidney. Studies comparing LDs with matched general population controls have found no differences in long-term outcomes between groups. However, recent studies comparing LDs to healthy matched controls have reported that LDs are at increased risk for end-stage kidney disease (ESKD), cardiovascular disease (CVD) and death. However, in these studies: the controls were not contemporaneous [born close to the same year]; not from the same geographic area; not known to be healthy on the date matching the donation date; and not matched for family history of ESKD. Each of these factors can impact the relative risk of developing kidney disease and its consequences. There are also concerns about the short follow-up and the statistical analyses of these studies. Some commentaries have suggested that the LD risk was under-estimated; others, overestimated. Having a complete and accurate understanding of true long-term LD risks is important to provide data to inform future LD candidates (informed consent), as well as to inform the design of long-term health maintenance of donors. Government agencies, providers, other stakeholders, and LDs themselves are asking for better quality long-term data. LDs at the University of Minnesota (U of MN) (1st transplant 1963), have been surveyed every 3 years, and data entered into a LD-specific database. Using the Rochester Epidemiology Project (REP), we optimally matched the LDs with healthy controls. The REP includes data as early as the 1950 and established a linked medical records system that has followed the medical history of residents of Olmsted County (same geographic area as the U of MN and Mayo Clinic). LDs were matched with contemporaneous healthy REP controls on age, gender, race/ethnicity. Matched controls then had medical chart review to ensure health at the time corresponding to the donation date. Chart-validated controls had data entered into a designated database; and were sent a survey, similar to the U of MN LD survey, asking about current health. In addition, data has been collected on Mayo Clinic LDs and matched controls. For all LDs and matched controls, data has been supplemented by information from the NDI (death), Minnesota State (death) and USRDS (ESKD). With this grant, we will use our comprehensive dataset to compare long-term (>40 yrs) outcomes of LDs and controls. Because of the quality of the data in the LD and REP datasets, we will be able to study and provide the best data to date on: LD risk of ESKD and death; risk of the more common intermediate events that precede ESKD or death (e.g., CVD); the impact of known risk factors such as family history or smoking on donation risk; the impact of post-donation new onset disease (e.g., diabetes) on risk; and the impact of donation on subsequent pregnancies. We will provide the most comprehensive data, to date, to inform prospective LDs about long-term risk, and to inform the follow-up and care of current LDs.

NIH Research Projects · FY 2024 · 2020-06

Clinical trials testing type 1 insulin-like growth factor receptor (IGF-1R) inhibitors failed in endocrine- sensitive and resistant breast cancer. These trials failed to include targeting of the insulin receptor (IR) and essential component of the IGF signaling system. Further, data from patients with endocrine resistant breast cancer showed that insulin receptor (IR) is more highly expressed in breast cancer cells than IGF- 1R. While it may seem futile to target IR, data show the fetal isoform of IR, IR-A, is more highly expressed than the adult isoform IR-B in cancers. Thus, it may be possible to create a cancer specific inhibitor of a highly expressed receptor for breast cancer treatment. We hypothesize that targeting of IR alone and in combination with other breast cancer therapeutics will be an effective therapy. Moreover, specifically targeting IR-A will be cancer specific with little impact on glucose homeostasis. To test this hypothesis, we propose three specific aims: 1) Engineer IR-A antagonists using small synthetic protein ligands via directed evolution; 2) Demonstrate IR-A regulation of the breast cancer malignant phenotype compared to IR-B and define a mechanism; and 3) Evaluate the efficacy of an IR-A specific antagonist, our existing IR-A and IR-B antagonists, and IGF-1R antagonists alone and in combination in breast cancer model systems Major advances in breast cancer have been the direct result of understanding and targeting key growth regulatory signals. Based on the failure of trials targeting the IGF-1R, we now have clear evidence that IR play a critical role in breast cancer development. Just as we were at the among the first to develop IGF-1R inhibitors, we have shown that IR inhibitors also may be used to target breast cancer. Completion of this proposal will further the development of new targeted breast cancer therapies. Cancer specific IR may be accomplished by development of an IR-A specific inhibitor. Given the growing number of women with hyperinsulinemia, creating a cancer specific inhibitor of IR could have significant impact.

NIH Research Projects · FY 2026 · 2020-06

PROJECT SUMMARY Decades ago, the cerebellum was explored as a potential target for the epilepsies. Mixed results ultimately reduced enthusiasm, but we have now shown that with the correct stimulation parameters, electrical stimulation of the cerebellar cortex can provide robust seizure control in a mouse model of temporal lobe epilepsy. During the previous funding period, we also demonstrated that optogenetic excitation of excitatory neurons in the cerebellar fastigial nucleus with projections to the central lateral thalamus (but not other populations of fastigial neurons) was able to inhibit hippocampal seizures. The fastigial nucleus may make a better target for deep brain stimulation than the cerebellar cortex. Unfortunately, it is not yet known if optimization of stimulation parameters can allow for robust seizure control when targeting the cerebellar nucleus (rather than cerebellar cortex) with on-demand electrical stimulation. In healthy animals, we further demonstrated that optogenetic stimulation of the cerebellum can cause a transient inhibition of CA1 pyramidal cells, and mixed, but structured, effects on CA1 interneurons. How cerebellar intervention in epilepsy impacts CA1 excitatory and inhibitory neuronal populations is unknown. Importantly, not only can the cerebellum impact seizures, but also seizures can impact the cerebellum, suggesting bidirectional functional modulation. In this proposal, we therefore examine 1) the impact of on-demand cerebellar stimulation on CA1 principle and inhibitory interneuron populations in chronically epileptic animals, 2) the impact of seizures on cerebellar fastigial and central lateral thalamic neurons, and the impact of on-demand optogenetic intervention targeting the fastigial nucleus on the central lateral nucleus, and 3) if, with Bayesian optimization, electrical stimulation of the fastigial nucleus can be an effective strategy, if this outperforms targeting of a different cerebellar nucleus (the dentate nucleus), and if targeting the downstream central lateral nucleus can also be an effective approach. Collectively, this data will significantly improve our understanding of temporal lobe seizure networks (extending far beyond the temporal lobe), improve our understanding of how cerebellar modulation may result in seizure inhibition, and ultimately identify potential new intervention strategies.

NIH Research Projects · FY 2025 · 2020-05

Project Summary/Abstract Our long-term goals are to understand intracellular active Ca transport in cardiac muscle and to develop drugs that target Ca dysregulation in heart failure. We focus on the cardiac sarcoplasmic reticulum (SR) Ca- ATPase (SERCA2a), the large membrane enzyme that pumps Ca into the SR to relax myocytes after contraction, and on SERCA’s regulatory proteins (regulins, RLN): phospholamban (PLB), sarcolipin (SLN) and dwarf open reading frame peptide (DWORF). Ample experimental and clinical data indicates that activation of muscle Ca transport is a powerful approach to numerous severe and widely spread disorders, especially heart disease. Over decades, we have developed and used spectroscopic techniques to resolve the structural mechanism of SERCA regulation by RLN. The challenge is great due to the complexity of the Ca transport mechanism, which involves dynamic protein-protein interactions, and RLN mechanisms of action are unclear or controversial. We have recently demonstrated fluorescence lifetime (FLT) methods with exquisite precision and resolution of protein structural changes, with high-throughput acquisition enabled by rapid scanning in a microplate reader. In screens of chemical libraries of ≤50,000 small-molecules, we have identified multiple types of SERCA activators. We are poised to implement early-stage drug discovery campaigns targeting the SERCA-RLN interaction, to identify compounds with therapeutic potential to treat Ca dysregulation in heart failure. Our central hypotheses: (1) compounds that shift SERCA-RLN structural state or binding have tissue-specific effects (2) compounds that enhance Ca-transport enhance myocyte contraction, and (3) have antiarrhythmic properties. Aim 1 will accelerate discovery of compounds that target SERCA-RLN interaction. Aim 2 will test effects on SERCA function in cardiac SR, HEK cells expressing human SERCA2a, and in cardiomyocytes. These novel chemical probes will open new avenues to elucidate structure-function mechanisms characteristic of SERCA-RLN. Aim 3 will enable lead discovery, using medicinal chemistry to develop and test analogues of promising Hits from Aim 2. Outcomes of this program will be new lead-like compounds, and a demonstrated systematic process targeting cardiac Ca-transport for drug discovery and development. This process will be ready for implementation in large-scale discovery and development campaigns to be pursued in future academic-industrial partnerships. For impact, we bring together an innovative combination of techniques, technologies, and experts focused on cardiac Ca transport regulation. This project is designed to enable future translation, while also developing tools for mechanistic understanding. SERCA has emerged as a high-value therapeutic target for some of the greatest Public Health challenges, not only in the heart (heart failure, arrhythmia) but also in skeletal muscle (muscular dystrophy, sarcopenia) and non-muscle cells (Alzheimer’s, diabetes, obesity, cancer), so the significance of our proposed research program is great and extends well beyond cardiology. 1

NIH Research Projects · FY 2026 · 2020-05

PROJECT SUMMARY This MIRA renewal application builds upon the work that my laboratory has conducted under the initial MIRA to develop and apply foundational methods for analyzing and engineering cell signaling in human health. The overall vision of our research program is to create enabling tools to uncover more detailed mechanistic understanding of signaling processes that can further facilitate the design of more effective therapies. The broad goals for the next five years are to develop new experimental and computational methodologies that can provide novel insights into the fundamental molecular mechanisms of receptor-ligand interactions, with an emphasis on proteins/interactions that are difficult to study due to their poor biophysical properties, combinatorial binding possibilities arising from multivalency, or complexities arising from cell-bound ligands (i.e., cell-cell interactions). For receptors with poor biophysical properties, such as integral membrane proteins, there are limited methods for high-throughput characterization of the ligands for such receptors since these receptors often require the native cell machinery and/or plasma membrane for proper synthesis, folding, and function. We propose to develop a methodology that leverages the power of directed evolution to rapidly map sequence/function relationships for natural ligands while also enabling engineering of novel binders to these complex yet clinically important targets. For multivalent and multispecific ligands, a key challenge is to enumerate the full combinatorial binding possibilities to multivalent targets and predict binding dynamics, which we incorporated into a new mechanistic model of multivalent binding during our prior support. Here, we propose to extend these efforts to understand how multivalent and multispecific ligand binding dynamics are further altered by binding to cell- surface targets, which will be clinically important for maximizing the efficacy and selectivity of promising new classes of multivalent and multispecific therapeutics. Extending the study to binding on cells requires additional consideration of cell-surface receptor dynamics such as mobility and internalization, which we will incorporate into our mechanistic modeling framework and also test experimentally. Finally, we propose to elucidate design principles for engineering ligand-receptor signaling when the ligand and the receptor exist on two distinct cell types. The nature of this cell-cell dose-response profile is essential for basic developmental processes (and their dysregulation) as well as the design of new therapeutic cells with improved efficacy and specificity. Collectively, advances in these areas of cell signaling will provide essential new tools for answering fundamental biological questions relevant to human health while also facilitating the design of novel molecular and cellular therapeutics for the treatment of a range of diseases.

NIH Research Projects · FY 2024 · 2020-05

Project Summary/Abstract Aging is a risk factor for cardiovascular disease (CVD), and women who experience premature or early menopause have a 50% greater risk for CVD-related death compared with women who experience menopause at the typical age. Indeed, CVD increases aggressively after menopause and is the leading cause of mortality in women in the US. Autonomic and blood pressure (BP) dysregulation, often demonstrated in postmenopausal women, is associated with CVD. Although literature suggests that estrogen is cardioprotective for premenopausal women, little is known regarding how the early loss of sex hormones in premature and early menopause effect BP regulation. This application for a Mentored Research Scientist Development Award (K01) is designed to advance knowledge important for understanding mechanisms contributing to increased CVD in aging women and support the career of Dr. Manda L Keller-Ross, DPT, PhD, an Assistant Professor in the Department of Rehabilitation Medicine, in the Medical School at the University of Minnesota. Dr. Keller- Ross is the PI of the Cardiovascular Research and Rehabilitation Laboratory, where the proposed research will take place. The long-term objectives of this proposal are to determine mechanisms that contribute to greater risk of CVD in premature and early menopausal women. Specifically, Dr. Keller-Ross aims to determine mechanisms driving autonomic BP regulation in premature and early menopausal women near the age of menopausal onset (Aim 1). She will then determine mechanisms driving autonomic BP regulation in women who have lived ≥10 years without functioning ovaries to determine the long-term effects (Aim 2). This proposal is in line with the mission of the NIA, to understand the nature of the aging process in women and how the loss of sex hormones contributes to the number one killer in women, CVD. Dr. Keller-Ross has a clinical research background and seeks mentored training and skill development to enhance her knowledge on biology of aging, women’s health specific to the cardiovascular system and menopause and advanced techniques to measure autonomic function. She will accomplish her career development goals through a combination of formal coursework, mentored skill and technique development for microneurography to measure muscle sympathetic nerve activity and noninvasive measures of baroreflex function and the empirical research described above. Dr. Keller-Ross has established an interdisciplinary mentorship team to guide her in these research and training activities. This committee has senior-level expertise in aging research in females, autonomic regulation of BP research and biostatistical analysis. They are eminently qualified and fully committed to assisting Dr. Keller-Ross to further her training, research and career path and achieve her career objectives. With this training, Dr. Keller-Ross is poised to become a leading scientist in cardiovascular health in aging women, producing research that is directly translatable to clinical practice devising strategic preventative and rehabilitative therapies to improve cardiovascular health, specific to aging women.

NIH Research Projects · FY 2024 · 2020-04

PROJECT SUMMARY A common pathological state strongly associated with both obesity and aging is insulin resistance (IR) in which cells become resistant to the effects of insulin. IR is a hallmark of prediabetes, affecting a third of Americans. It also represents a major risk factor for type 2 diabetes mellitus, physical dysfunction, heart disease, and dementia. Other than exercise and diet, limited mechanism-based strategies exist to improve IR. Another shared feature of obesity and aging is accumulation of p21Cip1–highly-expressing (p21high) cells in various tissues. However, the roles of p21high cells in IR and physical dysfunction remain largely unknown. To examine the relationship between p21high cells and IR, we have generated and validated a novel “p21-Cre” transgenic mouse model containing a p21 promoter driving a Cre fused to a tamoxifen-inducible estrogen receptor (ER) element. This novel model enables us to monitor, kill or modulate p21high cells in vivo without affecting other cells. In our preliminary studies, we find that intermittent clearance of p21high cells in obese mice significantly alleviates IR, indicating that strategies targeting these cells could result in novel approaches for managing IR and metabolic dysfunction. Based on these findings, we will test our central hypothesis that targeting p21high cells will alleviate metabolic and physical dysfunction associated with obesity. We will use p21-Cre mouse models to examine the role (Aim 1) and underlying mechanism (Aim 2) of p21high cells in IR and physical dysfunction. We will also leverage powerful single cell transcriptomics (SCT) technology to reveal the heterogeneity and conserved transcriptomic features of these p21high cells in tissues with obesity. This project will have a broad impact on both aging and obesity research by determining how p21high cells contribute to IR. Using multiple in vivo models, coupled with the powerful approach of single cell transcriptomics, we expect to gain a comprehensive understanding of p21high cells (at both functional and expression levels) in vivo. Results from this work will also enable future testing of pharmacological interventions that eliminate these cells to treat not only metabolic dysfunction, but also a wide range of age-related diseases.

NIH Research Projects · FY 2025 · 2020-04

OVERALL - PROJECT SUMMARY The purpose of our P50 Conte Center is to develop, test, and apply computational models of disrupted state representation processes to behavioral and neural measures relevant to psychosis spectrum illness across mouse, macaque, and human experiments. Our premise is that, in order to respond adaptively to the environment, the brain must process information to develop accurate and stable representations of the current state of the world; this requires neural processes that: 1) estimate the current state of the world; 2) use reward-based feedback to learn to recognize new states; and 3) maintain state representation across time gaps. State representation processes rely on precise neural activity including timing synchrony between prefrontal and sensory systems and across prefrontal and subcortical networks, including striatal systems. Our overarching hypothesis is that a range of computationally identifiable microcognitive processes contribute to state representation disruptions in psychosis, that these vary dimensionally across individuals (potentially manifesting as computational subprofiles), and that their underlying neural contributors, once understood, could serve as targets for precision treatments. Informed by our Center’s recent findings from NMDAR-antagonism in macaques, from NMDAR ablation and dopamine (ant)agonism in mice, and from EEG+fMRI studies in humans with and without psychosis, we will examine variable patterns of behavior and neurophysiology during task performance, examining the interplay between state estimation, state maintenance, and state learning computations. Aim 1: Computational science. Informed by biologically-grounded theoretical models and successes in the previous cycle, we will continue to develop methods to measure computational parameters of state representation processes, integrating across two tasks. Each task probes distinct neurocognitive and neural system operations– cognitive control and reward-based decision-making– highly relevant to psychosis. Aim 2: Non-human animal experimental science. Applying these computational methods, and drawing on findings from our human studies, we will analyze the behavior and neurophysiology of state representation processes relevant to psychosis, in mice and in macaques, under normal conditions and after manipulations of NMDAR and dopaminergic systems. Aim 3: Clinical science. Applying these computational methods, and drawing on findings from our non-human animal studies, we will identify the computational dimensions relevant to psychosis and their neural system correlates in humans (via EEG studies and innovative within-subject Precision Functional Mapping) with a focus on interactions among CEN, SN, and DMN network dynamics; we will identify how computational subprofiles in humans can be altered in response to manipulations of NMDAR and dopaminergic systems.

NIH Research Projects · FY 2026 · 2020-03

ABSTRACT Central nervous system (CNS) relapse is a major cause of treatment failure among patients with acute lymphoblastic leukemia (ALL). Notably, isolated CNS relapse occurs in ~3-8% of children with ALL and accounts for 30–40% of initial relapses in some clinical trials. Furthermore, current CNS-directed therapies are associated with significant toxicities. As a result, novel CNS-directed leukemia therapies are urgently needed to improve long-term outcomes while decreasing treatment-related morbidity. Although extensive research has demonstrated a critical role of the bone marrow microenvironment in leukemia biology, the impact of the CNS microenvironment on leukemia cell survival and chemoresistance is largely unknown. We developed a novel ex vivo co-culture system and an in vivo xenotransplantation approach to investigate the effects of the CNS niche on leukemia biology and chemoresistance. We then used these model systems to identify that 1) leukemia cells cultured in cerebral spinal fluid (CSF) in vitro and in vivo have diminished survival relative to serum or media, 2) leukemia cells predominantly localize to the meninges within the CNS, and 3) leukemia cells co-cultured with meningeal cells, or associated with the meninges of mice, exhibit enhanced survival and chemoresistance. We then identified that direct meningeal-leukemia interactions promote leukemia cell survival by modulating apoptosis balance, cell cycle progression, and quiescence. Importantly, leukemia chemoresistance was reversible and overcome by detaching the leukemia cells from the meninges. We then used a co-culture adhesion assay to identify drugs that disrupt the interaction between leukemia and meningeal cells. In addition to identifying several drugs that inhibit canonical cell adhesion targets and pathways, including the CXCR4 antagonist AMD3100, we found that Me6TREN, a novel small-molecule hematopoietic stem cell (HSC) mobilizing compound, also disrupts the interaction between leukemia and meningeal cells. This work demonstrates that the meninges exert a unique and critical influence on leukemia chemoresistance and defines novel mechanisms of CNS relapse beyond the well-described role of the blood- brain barrier. Based on this work, our central hypothesis is that the leukemia-meningeal cell interaction is a critical regulator of leukemia cell survival and chemoresistance in the CNS. Moreover, from a therapeutic standpoint, we hypothesize that niche disruption may be more efficacious in the CNS than in the bone marrow because of the less supportive environment of the CSF relative to the blood or serum. The objectives in this proposal are to use our in vitro and in vivo model systems for CNS leukemia to dissect the molecular mechanisms that mediate leukemia adhesion (Aim 1) and chemoresistance (Aim 2) in the CNS and test novel, clinically translatable therapies for CNS leukemia including Me6TREN and AMD3100 (Aim 3).

NIH Research Projects · FY 2026 · 2020-03

Project Summary As the population of the United States ages, the health burden imposed by diseases of aging is expected to increase concomitantly. Social factors, including low socioeconomic status, social isolation, and low social support, are among the best predictors of susceptibility to diseases of aging, as well as lifespan itself. Nevertheless, key questions about the causal relationship and the biological mechanisms that link social experiences to health and aging remain unanswered. Animal models are a powerful tool to address these questions. Like humans, other social mammals exhibit strong associations between social adversity, health, and mortality. Unlike humans, though, they experience less complex environments, have shorter generation times, and can be subjected to experimental manipulation in controlled environments. However, the use of animal models to identify mechanisms underlying social determinants of aging and aging-associated diseases is still limited. To grow and support the relevant research community, in 2020 we founded an R24 High-Priority Behavioral and Social Research Network on Animal Models for the Social Dimensions of Health and Aging (SDoHA). In less than four years, we have successfully grown the Network by supporting career development fellowships, and pilot grants as well as organizing meetings and scientific publications. These successes highlight three areas central to accelerating research on animal models for SDoHA, which together inform the Aims for this renewal application. Specifically, we will: Aim 1. Grow the animal models for SDoHA Network and support members at crucial career stages. Aim 2. Diversify the animal taxa used in the study of SDoHA. Aim 3. Support the development of animal models for translational geroscience and foster collaborations with researchers studying social gradients in humans. The renewal of our Network will continue to transform a weakly connected community into a self-sustaining field of researchers equipped to conduct impactful research on the social dimensions of aging. Together, this work will lay the foundation for establishing models and experimental approaches that directly inform our understanding the social determinants of health and lifespan in animals and humans, and are expected to have direct translational application to human health and well-being during aging.

NIH Research Projects · FY 2025 · 2020-03

Class V dental restorations, those on the lower third of the tooth, are the least durable type of dental restoration and are quickly increasing in placement due to a rapidly aging society. Water and bacteria mediated destruction of the dentin/restoration interface is the main cause of failure. Maintenance of integrity between the restorative material and surrounding tissue is an unexplored route to increase lifespans of Class V restorations to shield the vulnerable adhesive interface from bacteria-mediated degradation, promote restoration stability, and increase restoration aesthetics. Tissue attachment to current Class V restorative surfaces does not occur as current restorative materials lack any activity designed to influence tissue responses. This lack of attachment reveals the vulnerable dentin/restoration interface and exacerbates Class V failure. Functional tissues near Class V restorations may be able to naturally prevent subgingival plaque and lead to longer lifespans by exploiting the tissue. Our central goal is to develop new materials and apply experimental approaches toward reducing Class V failure. In Aim 1, we aim to understand oral cellular activation responsible for signaling and test candidate molecules for their ability to survive enzymatic degradation. In Aim 2, we will demonstrate our system enables biocompatible, facile, rapid, and robust delivery of biofunctional molecules to the material surface and retains molecules through a series of challenges meant to simulate the oral environment. The results will potentially extend lifespans of Class V dental restorative materials and reduce longterm healthcare costs. This training grant will provide pivotal opportunities to learn techniques including flow cytometry, high resolution X-ray photoelectron spectroscopy, and molecular biology techniques. Improved understanding of these techniques will further my progress toward becoming an independent academic researcher at a dental school.

NIH Research Projects · FY 2026 · 2020-02

Project Summary/Abstract Sensory perception requires the coordinated activity of tens of thousands of neurons functioning together in large-scale networks. The capabilities of these networks are defined and constrained by their development, yet the mechanisms underlying their formation remain incompletely understood. In the visual cortex of primates and carnivores, modular networks consisting of nearby neurons sharing coordinated activity and similar tuning properties extend across millimeters of cortical surface. Prior to eye-opening, such large-scale networks are already evident in correlated spontaneous activity, whose structure can predict future visually-evoked responses. We have previously shown that local recurrent mechanisms utilizing local excitation / lateral inhibition (LELI) interactions account for these modular functional networks in the week prior to eye-opening. However, major gaps remain in our ability to relate early network structure to mature sensory function. All previous work occurred at ages when spontaneous activity is already modular, meaning that its developmental origins remain unclear. Furthermore, early modular networks undergo refinement prior to eye-opening, but the role of changes in recurrent interactions versus changes in inputs is unknown. The experiments in this proposal will address these gaps, first by imaging spontaneous activity earlier in development than prior work, beginning at an age when cortical neurons are still completing their migration. This will allow us to capture the onset of modular activity in the cortex and investigate whether the same mechanisms that underlie modular activity later in development are also involved from the earliest stages. In order to determine if intracortical circuits are responsible for generating dominant modes of activity that refine in the week prior to eye-opening and serve as a dynamic scaffold for future visual representations, we will utilize our ability to directly stimulate the cortex with spatially-patterned optogenetics. These experiments will allow us to separate the contributions of changes in input from those of changes within recurrent circuits to both the developmental refinement of cortical networks and their potential role as a template for the future structure of visually-evoked representations. Finally, in order to understand whether a modular organization generated through recurrent LELI mechanisms exists elsewhere in visual cortices, we will image spontaneous activity in higher visual areas throughout development, and use patterned optogenetic stimulation to directly test the role of LELI mechanisms outside V1. Understanding the mechanisms through which modular network structure first emerges and whether recurrent circuits serve to generate dominant modes of activity that function as templates for sensory representations is critical for understanding how functional visual circuits are constructed during development. Determining if such mechanisms operate throughout higher visual areas is key for understanding the function organization of these regions. Collectively, the studies in this proposal will provide critical new insights into both the developmental origins of millimeter- scale functional networks and their role in shaping future functional organization throughout the visual cortex.