Brown University

NIH Research Projects · FY 2024 · 2023-09

PROJECT SUMMARY One in three Americans will be diagnosed with Alzheimer’s disease (AD) if no new prevention and intervention strategies are developed. The pathological hallmarks of AD include accumulation extracellular amyloid plaques consisting of amyloid beta (Aβ), processed from amyloid precursor protein (APP), and intracellular neurofibrillary tangles comprised of phospho-tau. These features are proceeded and precipitated by a wide range of intra- and inter-cellular functions, with derailed neuroinflammation being among the earliest manifestations in human brains. Of note, Chitinase-3-like protein 1 (CHI3L1, or YKL-40), an inflammatory protein mainly secreted by reactive astrocytes in the brain, has been documented to be a powerful AD biomarker, and its cerebrospinal fluid (CSF) level has recently been reported to be potentially the first disease indicator. However, how it functions in the brain and influences neuroinflammation and AD pathogenesis remains to be elucidated. In contrast, in peripheral tissues such as in the pulmonary system, CHI3L1 has been well characterized as an immune signaling molecule that controls many aspects of inflammatory processes via specific cell surface receptors and downstream signaling pathways. Teaming up with leading expertise in CHI3L1 biology in lung diseases, the Huang laboratory has gathered evidence showing that CHI3L1 is expressed more abundantly in astrocytes from AD brains and can mediate the inflammatory signaling and cellular responses as in the periphery. Supported by my own preliminary data, my central hypothesis is that CHI3L1 secreted by activated astrocytes engages a neuronal receptor and triggers a signaling cascade in neurons that contributes to the inflammatory neurotoxicity and leads to neurodegeneration and relevant AD features. My overall objective here is to define a neuronal signaling mechanism whereby CHI3L1 regulates glia-derived neuroinflammatory response and the resultant neurodegeneration. By using human neurons generated from pluripotent stem cells in single cultures and a Chi3l1-floxed mouse strain, I propose to pursue two specific aims in order to test my hypothesis and achieve the objective. In Aim 1, I will identify the CHI3L1 receptor and downstream signaling pathway in stem cell-derived human neurons in vitro. In Aim 2, I will determine the effects of CHI3L1 expression on inflammatory neurotoxicity in vivo in transgenic mouse brains with conditional Chi3l1 knockout and expression of AD pathology. My expected outcome is to elucidate an essential signaling function of CHI3L1 in neurons in response to neuroinflammatory stress. My results will reveal a mechanistic role CHI3L1 plays in the brain, departing from being merely a biomarker for AD and neuroinflammation. There will be a positive impact as my findings can be used in targeted therapeutics to combat AD. Finally, the primary responsibility of a F31 awardee is, in lieu of performing experiments, to prepare him/herself for future success in research. With this regard, my major goal of award period is to leverage the rich resources at Brown and execute a rigorous training plan that will impart to me the skills, ingenuity and experimental acumen necessary for growth into an independent investigator.

NIH Research Projects · FY 2025 · 2023-09

PROJECT SUMMARY This R01 application proposes conducting a fully powered randomized controlled trial (RCT) to test the effect of a patient navigation intervention for men who are disproportionately impacted by the HIV/AIDS epidemic in the United States. Pre-exposure prophylaxis (PrEP), a once daily medication, can dramatically reduce HIV acquisition risk. However, social and structural barriers have contributed to suboptimal PrEP initiation, adherence, and retention in care. Our prior NIH-funded pilot study (R34MH109371; MPI: Nunn, Chan, Mena) developed and evaluated an Intervention to Retain and Adhere Men in PrEP (RAMP-IT-UP), a brief strengths-based patient navigation program to enhance PrEP care outcomes among young men. The intervention was found to be highly acceptable among men and demonstrated preliminary effectiveness. Compared to control participants, RAMP-IT-UP participants were statistically more likely to initiate PrEP and adhere to PrEP based on pharmacy fill data and PrEP blood levels. Additionally, RAMP-IT-UP participants were more likely to be retained in PrEP care at the 3-month and 6-month clinical visits. Specific Aim #1 of this study will conduct a fully powered randomized controlled trial (RCT to estimate the effectiveness of RAMP-IT-UP in improving PrEP adherence and care outcomes among men in real-world community health center settings (CHCs). Specific Aim #2 will estimate the cost-effectiveness of RAMP-IT-UP among men attending CHCs compared to standard of care. We will also determine the cost-effectiveness of differing levels of intensity of navigation services to prevent HIV based on data collected in Specific Aim #1. Our goal is to develop a cost-effective intervention that enhances PrEP care outcomes and reduces HIV incidence for men which will be relevant for CHCs across the US. The long-term goal of this work is to decrease HIV incidence among men, which aligns with federal Ending the HIV Epidemic and National HIV/AIDS Strategy goals. This application is led by an experienced team of investigators with a proven track record conducting HIV, PrEP and disparities research in real-world clinical settings.

NIH Research Projects · FY 2025 · 2023-09

HNN U24 DISSEMINATION PROJECT SUMMARY The Human Neocortical Neurosolver (HNN) neural modeling tool was developed with BRAIN Initiative funding (R01EB022889: 09/2016–06/2020) to meet the Initiative’s goal to “develop innovative technologies to understand brain circuits and ensembles of circuits that inform understanding of the human brain and mechanisms for treating its disorder”. HNN is a biophysically principled neocortical circuit model with appropriate physics that allow bridging from macroscale human magneto- and electro-encephalography (M/EEG) signals to their cellular and circuit level generators. HNN is a hypothesis development and testing tool whose design and capabilities are unique compared to other M/EEG neural modeling software. A key value of HNN is to connect functionally- relevant human signals to circuit level dynamics studied in animal models, including data from revolutionary genetic and imaging tools used in mice and monkeys (e.g., Neuropixel recordings, calcium, and voltage imaging). These links are essential to discovering new principles of brain information processing and to developing treatments when this processing is disrupted by neuropathology. There is widespread use of M/EEG, and myriad inferences drawn from these signals about human brain function and health: HNN provides a highly accessible tool for researchers to make principled connections to the detailed neurons and circuits underlying these signals, to test new ideas and ground conclusions in circuit-level reality. The neuroscience community is actively engaged in the use of HNN for basic and clinical research, including studies of Alzheimer’s disease, autism spectrum disorder, pain, depression, and healthy development. While HNN was successfully developed, there remain several challenges for growth and long-term sustainability. The goal of this proposal is to support dissemination of HNN for broadly accessibly use and community-driven development. Identified challenges in maintenance of HNN’s code and documentation that ensure Findability, Accessibility, Interoperability, and Reusability will be addressed (Aim I), and key enhancements necessary to support end-user needs and experimental validation of model-derived predictions will be developed (Aim II). Interpreting the complex multiscale origin of M/EEG signals with HNN’s large-scale neural often requires domain expertise in computational neural modeling, human M/EEG, and neural dynamics. To support this need, we will continue to work with the community to integrate HNN into their projects through workshops, direct collaboration, and consultation with end-user groups, and by enabling HNN simulation on freely accessible supercomputers (Aim III). End-user feedback and documented support needs will be used to develop a Plan for Sustainability. A world-class Steering Committee includes developers of widely-adopted neuroscience software who will share their expertise to help HNN reach its maximal potential as translational neuroscience tool.

NIH Research Projects · FY 2026 · 2023-09

Alcohol-related risks are highest in young adulthood and this age group is most likely to be exposed to the alcohol use of their peers. Young adults are also highly influenced by their peers, both for high-risk behaviors and for prosocial actions. Bystander helping is one way that peer influence in potentially dangerous alcohol consumption circumstances can reduce harm to those at risk. Bystander helping is the phenomenon when witnesses to a problematic event step in to intervene in some way, but bystander helping has been under-investigated when hazardous alcohol use is the target behavior. In order to develop interventions that effectively leverage peer influence for harm reduction in the young adult community, research must be conducted on the circumstances of alcohol-related events, and specifically the experiences of those who could be bystander helpers. Drawing from bystander intervention research on sexual assault and bullying prevention, we propose to investigate the contexts and conditions under which bystander helping for hazardous alcohol use occurs naturalistically. The aims of this research are to investigate the event-level (Aim 1) and individual difference (Aim 2) predictors of bystander helping in response to peer alcohol-related risk, and to investigate the relationship between helping approaches and bystander and peer outcomes (Aim 3). Following community engagement with an advisory group and a pilot phase in which we will finalize procedures and assessments, we will conduct an assessment study with a sample of young adults (N = 200; ages 18-25) balanced on sex and reflecting the US in race/ethnicity, who will report on their exposure to the hazardous drinking of others in their environment for 28 days using ecological momentary assessment (EMA) methods. Participants will complete random and morning reports on their own smartphones that will collect information about alcohol-related behavior witnessed, the social context, the characteristics of the person showing the risk behavior, the nature of the relationship between that person and the bystander, the bystander’s own state including their own level of intoxication, perceived barriers to intervention, bystander strategies used, outcomes of any bystander helping, and the outcomes to the person showing the hazardous behavior. Each evening survey will pull forward information from the previous survey to allow for assessment of subsequent observations, and morning surveys will assess further the risks observed, behavioral responses, and outcomes. Methods used during the baseline assessment and orientation, including a social network interview, will facilitate brief, accurate and private EMA reports. We expect the information derived from this investigation will contribute in a substantive way to the development of effective trainings for individuals who are exposed to the hazardous drinking of others.

NIH Research Projects · FY 2025 · 2023-08

PROJECT SUMMARY Pneumonia is an important global health problem 1. Influenza A virus (IAV) leads to an estimated 500,000 deaths annually, and these numbers can be even higher during IAV pandemic years 2. A complication following infection with IAV is bacterial pneumonia, and this can lead to a more severe disease 3- 21. To survive a given infection hosts must be able to not only clear the pathogen, but also tolerate to the effects of the pathogen or the host response. The latter processes are referred to as disease tolerance. Normally, a balance between these two processes is reached, and the infection resolves 2223. My research lab has been at the forefront of exploring the concept of host disease tolerance in deadly complex respiratory infections 242322 • 25 • 26. We have shown that during pulmonary IAV/bacterial coinfection this balance between resistance and tolerance is disrupted causing increased lethality 22,24. Regulated cell death (RCD) plays important roles in both resistance and tolerance to infection. Cell death can be broadly categorized as membrane permeable (e.g. necroptosis and pyroptosis) or membrane impermeable (e.g. apoptosis). During infection several different types of cell death can be induced each inducing a unique response. Macrophages are responsible for both resistance mechanisms such as coordination of the immune response, and they contribute to tolerance in the form of the tissue repair responses after damage. This proposal aims to bridge gaps in our current understanding of complex respiratory infections by investigating the role of Caspase-8. a key regulator of RCD. in relevant in vitro and in vivo models of IAV/bacterial coinfection. We will focus on understanding how Caspase-8 pathways in lung macrophages during IAV/bacterial coinfection contributes to resistance and tolerance. Our preliminary data has demonstrated that Caspase-8 deficiency in macrophages leads to increased host resistance at the expense of host tolerance in the early stages of coinfection. This is most likely due to increased necroptosis and anti-microbial responses. We hypothesize that during severe lung infections such as those that occur with bacterial infection following IAV infection macrophage cell death controlled by Caspase-8 plays an important role in regulating the balance between resistance and tolerance. In this study we will determine the role that Caspase-8 in macrophages plays throughout the course of infection. We will use relevant in vitro models using human primary macrophages, as well as in vivo models. These studies will lay the foundation for future studies on understanding the impact of RCD in macrophages during complex pulmonary infections, which may ultimately lead to improved treatment options for patients with complex and severe lung infections.

NIH Research Projects · FY 2025 · 2023-08

Project Summary The long-term objective of this project is to develop a revolutionary quantum mechanical solid-state magnetometer designed to non-invasively detect femtoTesla (fT) scale magnetic fields derived from the brain’s electrical activities during natural human experiences. The project has been designed to address the vision of the NIH Brain Initiative: Transformative Brain Non-invasive Imaging Technology Development. The core component of the magnetometer is a quantum-based magnetic tunnel junction (MTJ), a nanoscale sensing device with potentially unprecedented sensitivity and performance. Through a series of steps and interdisciplinary collaboration, this project is expected to increase the sensitivity of the current MTJs by several orders of magnitude and to develop triaxial MTJ sensors capable of recording the brain's magnetic fields with the highest information density. Once the desired sensitivity is achieved, the project will build a whole-head 300-channel magnetoencephalographic (MEG) system based on MTJs, which can operate fully untethered, without the need for an expensive magnetically shielded room or nulling coils. By design, the sensors will be immune to natural head motion, further enabling the system to function in natural environment. The project will address several challenges in designing and producing sensors that can detect magnetic fields as low as 50 fT. First, the project will improve the detectability of current prototype sensors by several orders of magnitude by using a series of innovative approaches in sensor design, atomic engineering, fabrication, and noise reduction. Second, the project will design and package triaxial sensors that can simultaneously measure tangential and radial magnetic fields. Third, the project will reduce the size of the sensors by more than five times compared to other technologies, so that more sensors can be implemented on a full-head helmet to improve spatial resolution and localization. Finally, and importantly, the project will increase the field dynamic range of the sensors over the technologies based on OPM (optically pumped magnetometer) and SQUID (superconducting quantum interference device), so that the MTJ-MEG system can operate without the need of magnetic shielding to allow real-world applications. Once these groundbreaking MTJ sensors are developed, the project will integrate the MTJ-MEG system architecture and make it available for verifying performance versus competing technologies. The project will use the MTJ-MEG system to assess brain related signals during sensory stimulation, cognitive processing, and motor actions. If successful, this project will develop a transformative MTJ-MEG system with unpresented levels of performance to produce a dynamic picture of the brain under natural settings covering the whole lifespan.

NIH Research Projects · FY 2025 · 2023-08

Project Summary: Mitochondria are critical for cells with high energy demand. Mitochondrial ATP synthase completes the final step of ATP production. Mutations in ATP synthase cause a range of diseases, including Schizophrenia, and heart failure. Thus, the basic biology of ATP synthase is important for health and disease. The subunits of the ATP synthase are encoded by the nuclear and mitochondrial genome. A unique challenge of assembling the ATP synthase is to coordinate the gene expressions from dual genetic origins. Recent studies showed that cytosolic and mitochondrial translation are temporally synchronized to maintain the stoichiometry of the ATP synthase subunits (Couvillion, M.T., et al. Nature, 2016; Soto, I, et al. Genome Biology, 2022). However, the underlying mechanism of the cross-compartment communication remains largely unknown. I aim to fill in the knowledge gap in this proposed study. Using single-molecule fluorescent in situ hybridization (smFISH) in yeast, I imaged the mRNAs of a nuclear-encoded subunit, ATP2, and a mitochondrial-encoded subunit, ATP6/8. I discovered that the ATP2 and ATP6/8 mRNA co-localized on the mitochondrial network. Based on this observation, I hypothesize that the cytosolic and mitochondrial translation co-localize on opposite sides of the mitochondrial double membrane, thereby promoting the assembly of the ATP synthase. In Aim 1, I propose to determine how the ATP2 mRNAs co-translationally associate with mitochondria. In Aim 2, I will define the spatial coordination between the cytosolic and mitochondrial mRNAs in yeast and cultured neurons. In Aim 3, I shall identify the regulating proteins and dissect the underlying mechanism of the cross-compartment co-localization. This study will uncover a novel mechanism by which cells spatially coordinate the nuclear and mitochondrial gene expressions during mitochondrial biogenesis. My PhD training with Dr. Peter Walter prepared me with the skills in organelle biology, yeast cell biology, and biochemistry. My postdoctoral training with Dr. Robert Singer equipped me with the expertise in single-molecule imaging. These complementary skills give me the unique opportunity to conduct this proposed study. During the K99 phase, Dr. Singer will help me develop the skills to image mRNAs in yeast and cultured neurons. Extending my research from yeast to neurons will increase the impact of my future research. Dr. Michael Rout (Co-Mentor) will provide the expertise to identify the proteins that bind to the co-localizing mRNAs. Dr. Liza Pon (Co-Mentor), Dr. Thomas Fox (Consultant), and Dr. Christof Osman (Collaborator) form my mitochondrial mentoring team. Their diverse background will allow me to acquire the knowledge and skills of different aspects of mitochondria. This study will open opportunities to study the spatial regulation of mitochondrial gene expression. It will serve as a foundation for an independent research program in my future laboratory.

NIH Research Projects · FY 2025 · 2023-08

PROJECT SUMMARY/ABSTRACT With expanding cannabis legalization and concurrent significant increases in cannabis-related traffic fatalities, driving under the influence of cannabis (DUIC) has become a major public health concern. Despite the impairment in driving skills and increased crash risk associated with cannabis intoxication, little is known about how individuals make the decision to drive (or not) after using cannabis. The proposed project advances our team’s work on cannabis and DUIC risk factors to identify person-level (e.g., DUIC cognitions), within-person processes (e.g., subjective intoxication, affective state, cannabinoid concentration, quantity, route of administration), and contextual influences (e.g., social context, environmental conditions) that predict DUIC behaviors in daily life. The proposed multi-method project will provide unique qualitative and quantitative information about individual-level DUIC behavior using geographically explicit ecological momentary assessment (EMA), which combines event-level data on cannabis use with spatio-temporal data on vehicle movement. Our preliminary data has demonstrated the feasibility of these methods by integrating passively collected continuous location data using a vehicle-based GPS tracking device with EMA data on driving and cannabis use. Frequent and less frequent cannabis users (N = 260) will complete smartphone measures of cannabis use, concurrent substance use, affect, momentary impulsivity, perceived DUIC dangerousness, driving intentions, driving motives and destinations/location, and context during a 4-week EMA period. DUIC (i.e., driving within a pharmacologically-relevant timeframe) will be identified by integrating geospatial data (i.e., latitude/longitude/time and vehicle’s movement) passively and continuously collected in the field with EMA data on cannabis use. Weekly testing of participant cannabis for Δ⁹-tetrahydrocannabinol (THC) and cannabidiol (CBD) concentration will be done using a near-infrared spectroscopy device. This will be the first study to prospectively examine the influence of within-person and contextual predictors on DUIC likelihood and distance traveled (Aim 1). We will explore driving-related cognitions, indices of working memory capacity, and user characteristics in relation to cannabis use and driving experience as potential moderators of the effects of event-level predictors on DUIC (Aim 2). To provide insights on contextual factors and decision-making processes related to DUIC, participants will complete a narrative qualitative interview focused on annotating maps of representative DUIC and non-DUIC trips generated by GPS data during one of the four weeks of the EMA period (Aim 3). Exploratory machine learning analyses will be used to characterize and distinguish DUIC episodes from non-DUIC driving episodes (Aim 4). The current project will be the first full-scale study to integrate such real-time individual-level exposure data with DUIC outcomes of interest. Findings can have important implications for cannabis regulatory science and DUIC prevention efforts, which is critically needed to protect public safety.

NIH Research Projects · FY 2025 · 2023-08

The long-term goal of the COBRE Center for Central Nervous System Function is to develop a neuroscience research center that provides research support to a wide array of neuroscience researchers at Brown University and its affiliated hospitals and the Rhode Island neuroscience research community. Since we began our COBRE in 2013, we have supported 13 Project Leaders and nine Pilot Project Leaders as they developed their research and academic careers. Collectively, our supported cohort has been awarded nearly 20 R01 or equivalent research grants, which along with other awarded grants has yielded more than $50 million in external funding. Grants relevant to our COBRE received by members of our Research Cores have totaled another $4 million. Of the 11 Project Leaders who served two or more years, nine have successfully graduated, with a tenth having received positive reviews of an R01 grant proposal. Collectively, our COBRE supported scientists have published nearly 130 peer-reviewed papers citing COBRE support and more than 400 papers overall. These papers have appeared in prestigious journals, including Nature Human Behavior, Current Biology, Neuron, eLife, PLoS Computational Biology, Journal of Neuroscience and other field-specific journals. We have developed a research core to assist our community in best practices for experimental design and data analysis. The main objective for Phase 3 COBRE support relates to building-out our research core to serve the needs of the neuroscience community at Brown and Rhode Island and also to expand and strengthen our Pilot Project Program. We will enhance the scope of our Research Core by integrating two existing neuroscience core facilities, the MRI Research Facility and the Rodent Behavior Phenotyping Core, into one administrative entity. Additionally, we will partner with service cores of other local COBREs and those at Brown-affiliated hospitals to broaden our service outreach. Our vision is that by sustaining and transitioning the established research infrastructure into an integrated entity, we will enable neuroscientists to conduct their work more effectively and efficiently.

NIH Research Projects · FY 2025 · 2023-08

PROJECT SUMMARY: Methamphetamine is a highly addictive drug that has major effects on the brain’s learning and memory systems. Methamphetamine use is commonly predicted by high use of legal psychostimulants such nicotine or alcohol, but the mechanisms underlying sequential use of alcohol, nicotine and methamphetamine are not well understood. Both nicotine and alcohol alter the function of brain circuits required for adaptive decisions, but the cellular and molecular mechanisms through which this occurs largely remain a mystery. Identifying the gene regulatory networks within select circuits in high alcohol or high nicotine seeking animals and deciphering how methamphetamine alters these networks is necessary to decode the fundamental molecular underpinnings through which methamphetamine influences maladaptive choice. However, the diversity of circuits and cells involved in these responses has confounded our ability to identify key molecular pathways. Our goal is to use a genes-circuits-behavior approach in Drosophila melanogaster to provide an innovative and holistic understanding of the core fundamental principles through which sequential use of psychostimulants alter the molecular landscape and neural dynamics of memory circuits to alter behavioral decisions. Transcriptional regulation within memory circuits is a fundamental process through which psychostimulants drive maladaptive changes in the brain function. This work provides the causal gene regulatory mechanisms through which high preference for alcohol or nicotine affects methamphetamine response in memory circuits. This has major implications for how co-use of psychostimulants alters the molecular landscape that drives drug-seeking behavior.

NIH Research Projects · FY 2025 · 2023-08

Project Summary Redox imbalances of reactive sulfur species (RSS) in cardiac cells contribute to cardiovascular diseases such as myocardial ischemia-reperfusion (MI/R) injury and heart failure. Applications of sulfane sulfurs have been found to cause cardioprotective effects, and sulfane sulfur bioavailability in plasma has even been recently suggested to be a biomarker for cardiovascular disease (CVD). However, this field currently lacks clear understandings as to how endogenous RSS, particularly sulfane sulfurs, work. Our long-term goal is to elucidate the complex sulfane sulfur pathways at various health stages of cardiac cells and use this information to drive the development of early diagnostic tools and therapies for CVD. Specifically, this project will meet the critical need of having effective methods to study these pathways by developing chemical tools that: 1) quantifiably ‘see into cells/tissues’ despite the presence of biological fluids such as blood and 2) turn-on only after subcellular localization. These highly sensitive, specific, and targetable/triggerable next generation fluorescent sensors will allow us to probe the cardioprotective roles of sulfane sulfurs in cardiac cell models of MI/R injury even to subcellular extents and non-invasively decipher complex cardiovascular sulfur-mediated redox pathways. We expect that: 1) promising sensors will be identified for future evaluations in animal models of MI/R and 2) our tools and studies will establish a strong basis for advancing the clinical potential of sulfane sulfurs through a greater understanding of the mechanisms by which sulfane sulfurs regulate redox environments in CVD. The long-term goal for this fellowship award is to develop essential skills for a successful career as a chemical biology professor studying human diseases at an R1 institution. A team consisting of the sponsor and collaborators has been assembled with expertise in organic chemistry and synthesis, redox biology, chemical biology, molecular and cell biology, computational/data sciences, and cardiovascular diseases. Further training will be obtained from conference attendance and presentations in conjunction with teaching certificate programs, outreach, and leadership positions at Brown University.

NIH Research Projects · FY 2026 · 2023-08

PROJECT SUMMARY Aging is a complex process where perturbation of multiple molecular pathways contributes to organ deterioration and aging-related morbidity and mortality. One component of the aging process is severe dysregulation of gene expression that contributes to changes in the proteome of aged cells, which can compromise cell function. Understanding the mechanisms responsible for aging-induced perturbation of gene expression will be key to develop the necessary medical therapies. Although significant progress has been made in understanding the transcriptional changes occurring with aging, very little is known about how post- transcriptional events, such as RNA modifications, control protein synthesis during aging. In this proposal we will examine the role of METTL3-mediated m6A mRNA methylation in the context of aging using muscle aging as a model system. We hypothesize that METTL3-dependent mRNA methylation regulates the process of aging by controlling the translation of specific pro-hypertrophic mRNAs. For the first time, utilizing gain- and loss-of-function approaches we will characterize this novel regulatory program by establishing the molecular mechanisms by which m6A regulates the life of select mRNAs and assess the global aging- and METTL3- dependent translation dynamics (aim 1), examining the therapeutic benefit of enhancing m6A content to counteract sarcopenia in clinically relevant animal models (aim 2), and define the role of m6A binding proteins in muscle aging (aim 3). Completion of the proposed aims will allow the uncovering of a novel mechanism responsible for post-transcriptional regulation of aging with significant therapeutics ramifications.

NIH Research Projects · FY 2025 · 2023-08

Abstract Immune checkpoints play an important role in restraining the immune response, maintaining self- tolerance and preventing excessive collateral damage. Immune checkpoint blockade has revolutionized cancer immunotherapy but can lead to immune related adverse events (IrAEs). Patients receiving programmed cell death protein 1 (PD-1) blockade can develop IrAEs including vitiligo, colitis, endocrinopathies, and Sicca Syndrome, an autoimmune disease characterized by the accumulation of immune cells in the salivary and lacrimal glands, leading to deterioration and dysfunction. After immune cell activation, PD-1 is upregulated, and upon interacting with its ligand PD-L1, PD-1 signaling results in reduced proliferation, cytokine production, and cytotoxicity. PD-1 is expressed on infiltrating lymphocytes of the salivary gland including natural killer (NK) and T cells. Hyporesponsive immune cells in the salivary gland are thought to retain the integrity of this delicate tissue, while inadvertently contributing to viral latency. While trying to understand why lymphocytes in this organ are hyporesponsive, I unexpectedly found that genetic loss of PD-1 results in increased CD8+ T cell number and frequency in the salivary glands of naïve animals. Additional preliminary data suggests that NK cells may control CD8+ T cell expansion in the salivary gland. Based on these data, I hypothesize that in order to preserve the integrity of the salivary gland, NK cells may control T cell proliferation via the PD-1/PD-L1 axis in this organ. Therefore, in Specific Aim 1, I will elucidate the molecular mechanism by which PD-1 regulates CD8+ T cells in the salivary gland, and characterize the transcriptome and effector functions of the expanded CD8+ T cell population. In Specific Aim 2, I will characterize the potential immunopathological consequences of CD8+ T cell expansion in the PD-1 KO salivary gland by assessing histopathology and saliva secretions. These studies of PD-1 on salivary gland immune cells should provide insights for prevention and treatments of IrAEs. In preparation for the proposed work, my training has taken place in the outstanding immunology laboratory of Dr. Laurent Brossay, as well as within the supportive Pathobiology Graduate Program. My training experience will be enriched by attending and presenting at national and international conferences, and thoughtful mentoring by my sponsor. Completion of this proposal will prepare me with a repertoire of skills and the key foundational knowledge required for a successful career as an independent researcher.

NIH Research Projects · FY 2025 · 2023-07

PROJECT SUMMARY The goal of this treatment development project is to develop and pilot an adaptive mobile health (mHealth) intervention that targets the co-use of alcohol and opioids in young adults with opioid use disorder (OUD). Alcohol is an under-recognized contributor to the opioid crisis that, when used with opioids, substantially increases the risk of opioid overdose and other use-related problems. Despite clear public health significance, however, no existing intervention targets alcohol-opioid co-use. This gap is especially detrimental to young adults aged 18-25, who have the highest rates of alcohol and opioid (i.e., nonmedical prescription opioid or heroin) use and use disorders of any age group. Over half of young adults who use nonprescription opioids report past-year alcohol-opioid co-use, yet few young adults receive treatment for either substance. MHealth interventions, which use technology (e.g., smartphone applications) to intervene in daily life are highly accessible and acceptable to young adults. MHealth thus offers a means of targeting alcohol-opioid co-use that can reach young adults not currently in treatment. However, developing an effective mHealth intervention requires an ecologically valid understanding of the target behavior, which does not exist for alcohol-opioid co- use. Thus, the first step of the proposed project is to use ecological momentary assessment (EMA), wherein 60 young adults (ages 18-25) with mild to moderate OUD who report regular alcohol-opioid co-use will complete EMA multiple times daily via smartphone for 3 weeks to examine how, why, and when young adults co-use alcohol and opioids in real time in their daily lives. Using established guidelines for developing mHealth interventions, we will then use this EMA knowledge base to develop a smartphone-based mHealth platform targeting alcohol-opioid co-use. The intervention will be founded on principles of motivational enhancement therapy and cognitive behavioral therapy and deliver tailored micro-interventions based on participants’ responses to EMA assessments. We will refine the intervention based on feedback from two iterative usability studies with 10 participants. Finally, in a new sample of 60 young adults, we will conduct a Stage 1 randomized clinical trial, comparing the mHealth intervention to EMA-only and remote assessment-only control conditions. We will evaluate the intervention’s feasibility and acceptability and putative mechanisms of action. Sustained benefit will be evaluated at 3-month follow-up. This study will address national priorities to respond to the accelerating opioid crisis. It will provide vital new information on the nature of alcohol-opioid co-use in daily life and develop and pilot an innovative adaptive mHealth intervention to address this behavior in young adults.

NIH Research Projects · FY 2024 · 2023-07

Project Summary/Abstract Multiple arrhythmia conditions manifest in the heart due to conduction disorder, a failure of conduction between local islands of cardiomyocytes that are separated physically by millimeter (mm) to centimeter (cm) distances of non- or poorly conductive tissue. While electronic devices such as implantable cardioverter-defibrillators provide life-saving support for patients, their complications and lack of biological integration for long-term conduction restoration limit their success. A novel therapeutic approach is to provide cell-based physical connections between electrically active cardiomyocytes that could resynchronize cardiac electrophysiology to reduce arrhythmia risk and promote efficient cardiac pumping. Our long-term goal is to re-engineer electromechanical function of diseased hearts and specifically to address the critical need in clinical cardiac electrophysiology practice for long-lasting, anatomical electrical connections with biological responsiveness between disparate islands of cardiomyocytes in the heart. The objective of this proposal is to explore efficacy of a novel “bioelectric thread” we are developing that is made of natural biomaterials and hiPSC-derived cardiomyocytes (hiPSC-CMs). This technology is intended for cardiomyocyte-based coupling across mm to cm distances via formation of a continuous bridge of hiPSC-CMs. Our central hypotheses are that delivery of a confluent layer of cardiomyocytes along microthreads will create an electrical bridge via cellular gap junctions with known conduction velocity, and that this bioelectric cell bridge will be established within one week to enable electrophysiological synchronization and ameliorate conduction problems. Our preliminary data show that hiPSC-CM conduction along microthreads transmits action potential signals and calcium transients across at least 1.5 cm at 2.7 cm/s conduction velocity between two engineered cardiac tissues within 1 day in vitro. We propose to advance the biomanufacturing of bioelectric threads using 3D bioprinting and develop an injection-based device for precise implantation in the heart in Aim 1. We will assess our hypotheses in Aim 2 by evaluating electrical coupling and efficacy of cardiac synchrony in two different models of conduction anomalies after implantation of bioelectric threads. The parallel aims develop critically important technologies in tissue engineering to advance regeneration of cardiac conduction. The development of novel therapies for durable, biologically responsive conduction is significant because failure of current approaches in patients are associated with increased arrhythmia and mortality risk, necessitating novel solutions. This project is innovative in its use of 3D bioprinting for biomanufacturing of bioelectric threads, development of a delivery system for precise local implant in the heart, and evaluation of efficacy in diverse models of conduction disorder. The successful development of this technology requires investment in this early phase, and in doing so, it is likely that bioelectric threads will move the field of cardiac conduction repair into a new era, where regeneration of native-like anatomy and function becomes an attractive strategy for patients with cardiac conduction defects.

NIH Research Projects · FY 2024 · 2023-07

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative agent of coronavirus disease 19 (COVID-19), representing a global health threat. The viral spike protein, anchored on the surface of the viral envelope as homotrimers, binds to angiotensin-converting enzyme 2 (ACE2) and mediates the cellular entry of this virus. The receptor binding domain (RBD) of spike directly binds to ACE2, which induces a conformational change that facilitates virus fusion. This fusion event releases the SARS-CoV-2 genome into the cytoplasm. Spike protein, specifically the RBD, is the primary antigenic target for COVID vaccines in the market and interfering with the interface between RBD and ACE2 is the mechanism of action for the majority of existing therapeutic antibodies, indicating the importance of RBD and its binding to the cellular receptor for controlling SARS-CoV-2. Thus far, several cellular factors have been identified to facilitate cellular entry of SARS-CoV-2 (neuropilin-1, heparan sulfate, and C-type lectins). However, it is unclear whether there are any cellular proteins that inhibit viral entry. Our new preliminary data reveal that cellular entry of SARS-CoV-2 is inhibited by a novel inhibitory cellular protein, Leucin-rich repeat containing 15 (LRRC15). We performed binding assays using recombinant proteins in cells and in cell-free models that show LRRC15 directly interacts with the RBD of spike with a moderate affinity (KD = 43~148 nM, depending on domain and variant). Although ACE2 also interacts with the spike via the RBD, the interaction of LRRC15-RBD does not compete or stabilize ACE2-RBD interactions, suggesting non- overlapping binding sites. Further analysis of human lung single cell RNA sequencing dataset reveals that expression of LRRC15 is primarily detected in fibroblasts and particularly enriched in pathological fibroblasts in COVID-19 patients. ACE2 and LRRC15 are not co-expressed in the same cell types in the lung. Strikingly, LRRC15 inhibits spike-mediated viral entry not only in the same cells, but also in neighboring cells in trans. Expression of LRRC15 in ACE2+ cells blocked spike-mediated viral entry in ACE2+LRRC15- cells, providing a unique concept of viral entry inhibition by an inhibitory factor. This result suggests a protective role of LRRC15 in a physiological context. Our central hypothesis is that human LRRC15 acts as an inhibitory entry factor for SARS-CoV-2, acting in trans as a decoy receptor expressed in non-susceptible pathological fibroblasts in the lung. This proposal will explore by which LRRC15 inhibits entry of SARS-CoV-2 in trans through two specific aims. Aim 1. Determine molecular mechanisms of entry inhibition by LRRC15. Aim 2. Evaluate pathological fibroblasts providing the inhibitory function ex vivo. This study provides an insight into therapeutic development and a better understanding of COVID-19.

NIH Research Projects · FY 2024 · 2023-07

PROJECT SUMMARY/ABSTRACT Career Goal: My long-time career goal is to become an independent investigator studying the effects of health care policies on access and use of maternal and infant health care services among low-income populations. With additional skills in claims data, subject matter expertise, and causal inference methods, I plan to build a career focused on generating evidence to improve maternal and child health outcomes and reduce disparities in the United States. Career Development: I will pursue the following training aims during the mentored (K99) phase of the award: (1) clinical maternal and infant health content expertise; (2) claims data analysis skills; (3) advanced causal methods, and; (4) professional development. Research Project: Extending postpartum Medicaid has the potential to address poor maternal health in the United States, with additional implications for infant care use and health.1 Approximately 63% of maternal deaths in the U.S. occur postpartum, and more than half are considered to be preventable.2 However, despite high postpartum healthcare needs, pregnancy- related Medicaid ends after 60 days postpartum. Medicaid pays for almost half of all births in the US and disproportionately covers births to the low-income population and people of color.3–5 There has recently been increased interest in postpartum Medicaid, including an American Rescue Plan (ARP) option to extend Medicaid one-year postpartum.6 Prior to this, the March 2020 Families First Coronavirus Response Act (FFCRA) prevented Medicaid disenrollment during the pandemic, thereby extending postpartum Medicaid eligibility, which can inform current postpartum Medicaid policies under consideration. Using the Rhode Island All-Payer claims data, this proposal's Specific Aims are: K99/1) To evaluate the effects of the FFCRA on coverage and maternal and infant care in the postpartum period, R00/2) To evaluate the heterogeneous effects of the FFCRA on outcomes by race and ethnicity, R00/3) To evaluate how the COVID-19 pandemic affected the effects of extended postpartum Medicaid eligibility on care use. Mentorship: I have assembled a highly accomplished team of experts at Brown University to provide mentorship and guidance as I transition to research independence, comprised of Dr. Trivedi, Professor in the Department of Health Services, Policy and Practice and Department of Medicine; Dr. Steenland, Research Assistant Professor of Population Studies in the Population Studies and Training Center; Dr. Vivier, Professor of Health Services, Policy and Practice and Professor of Pediatrics and Emergency Medicine; Dr. Wilson, Professor and Chair of the Department of Health Services, Policy and Practice, and Professor of Medicine; Dr. Savitz, Professor of Epidemiology and Professor of Obstetrics and Gynecology and Pediatrics; and Dr. Tuuli, Chace-Joukowsky Professor and Chair of Obstetrics & Gynecology. Future Directions: The training and research in this proposal will provide me with a unique set of expertise and skills to prepare me to reach research independence and submit an R01 application to examine the effects of state variation in postpartum Medicaid policies using claims data.

NIH Research Projects · FY 2025 · 2023-07

The preschool years are a critical time for shaping food preferences and eating behaviors which, in turn, affect dietary behaviors in adults and life-long risks for obesity, diabetes, cardiovascular disease and other chronic conditions. Unfortunately, many US children, especially low-income and ethnic minorities, have dietary patterns associated with obesity and increased morbidity. Thus, to improve child diet quality, it is essential to develop effective targeted interventions in settings where children (especially higher risk children) spend time. There is a national call for dietary interventions that span multiple settings including the childcare and home environments. In response to PAS-20-160, we build upon our efficacious Healthy Start intervention with family childcare home (FCCH) providers (FCCP) to pilot a novel, 8-month multilevel tailored intervention to reach families through FCCH. This would be the first study to incorporate family-based intervention components into FCCH. FCCH are a promising intervention setting as parents trust FCCPs as extended family members and FCCP feel comfortable talking to parents about children’s diet, but want more training to do so effectively. The 8-month intervention will include an adapted version of the Healthy Start intervention for English and Spanish-speaking FCCP as well as FCCP training to deliver nutrition messages to parents using an existing childcare App, complemented with FCCH environmental cues and tailored print and videos for parents. Specific Aims are to: SA.1. Conduct formative research with FCCP and parents to inform refinement of the multi-level intervention. SA.2. Conduct a pilot trial with 40 FCCP and 80 parents with 18-54-month-old children to evaluate feasibility and acceptability of intervention and study protocols, and preliminary efficacy of the nutrition intervention compared to an attention-matched control on: a). children’s dietary quality at FCCH and home, b). social/physical FCCH and home food environments, c). children’s dermal carotenoid levels and z-BMI scores. SA3. Conduct post qualitative interviews with parents, FCCP, coaches, staff & partners to assess the intervention’s acceptability and suggestions for improvement. In sum, feasible and effective interventions to improve young children’s diets are urgently needed. The FCCH is a novel and untapped setting to intervene with both FCCP and parents simultaneously to affect children’s diets in both the childcare and home settings, which has the potential to more fully impact the child’s overall diet and weight status. This pilot feasibility research of a novel, multi-level intervention will inform a future full-scale cluster RCT, which will fill important research gaps and move the frontier of nutrition research forward.

NIH Research Projects · FY 2025 · 2023-07

Project Summary Metastasis remains a major obstacle in the treatment of breast cancer. A key contributor to metastasis is the epithelial-to-mesenchymal transition (EMT) program. During EMT, cancer cells gain mesenchymal, migratory, invasive, and stem cell properties. Cancer stem cells (CSCs) play crucial roles in metastatic progression, cancer recurrence, and chemoresistance. There is an urgent need for CSC-targeting therapies to combat recurrence and prevent metastasis. The premiere strategy to exploit the vulnerabilities of CSCs is to identify essential factors that support the stemness phenotype. Recent research has shown that the EMT process does not move cells between the binary “epithelial” and “mesenchymal” states. Instead, it is a continuum, and the cells can maintain both epithelial and mesenchymal characteristics (hybrid E/M) and possess high cellular plasticity and stemness. During EMT, changes in intermediate filaments, particularly upregulation of the mesenchymal-associated filament protein vimentin, have been documented. However, the role of vimentin in EMT and stemness and the consequence of co-expression of vimentin and cytokeratin (CK) in hybrid E/M cells with high stemness are not known. Besides, the function of vimentin is regulated through dynamic phosphorylation and dephosphorylation on critical sites, also known as “vimentin phospho-malleability. Preliminary data show that inhibiting phospho- malleability of vimentin serine-56 (S56) inhibits stemness and induces multinucleation in triple-negative breast cancer (TNBC) cells. It does not affect cancer cells with epithelial phenotypes, which do not express vimentin, or fibroblasts, which only express vimentin. Among the various phosphorylation sites, the phospho-malleability of S56 alone can induce multinucleation and inhibit stemness. Therefore, we hypothesize that vimentin S56 phospho-malleability is critical for maintaining the stemness of carcinoma cells expressing vimentin, and it is a driver of metastasis. Interfering with vimentin S56 phosphorylation will result in selective elimination of CSCs, inhibition of metastasis, and increased chemo-sensitivity. We propose the following aims to test this hypothesis: (1) Characterize how vimentin S56 phospho-malleability influences EMT and stem cell properties. (2) Define how the interaction between phospho-malleable vimentin and CK affects stemness in hybrid E/M cells with plasticity. (3) Determine the importance of vimentin S56 phospho-malleability on metastasis. Through innovative experiments using hybrid E/M cell lines, we will identify signaling pathways and vimentin-interacting proteins in the context of vimentin S56 phospho-malleability. Using patient TNBC samples, syngeneic tumor xenografts, and human patient-derived xenografts, we will demonstrate the biological importance of vimentin S56 phospho-malleability for stemness in vitro and metastasis in vivo. Understanding the importance of vimentin S56 phospho-malleability and the impact of impairing this will aid in developing novel EMT and CSC-targeting therapies to treat metastasis-prone TNBCs.

NIH Research Projects · FY 2025 · 2023-07

Medical diagnosis is a critical component of effective health care but misdiagnosis, delayed diagnosis and incorrect triage is common especially in urgent or emergency care settings, and a major contributor to adverse clinical events. While improvements in health care are important in addressing misdiagnosis, patients have a key role by recognizing potentially serious symptoms and seeking care in a timely manner. These concerns are of particular importance for patients requiring urgent or emergency care with potentially life-threatening diseases, such as transient ischemic attacks (TIA), stroke or myocardial infarction. They often fail to recognize the seriousness of their symptoms and may fail to seek care promptly, resulting in missed treatments and poorer outcomes. Patients with stroke typically must be treated within 4 hours to achieve a good response, and public education campaigns have not significantly helped. Smartphone apps for medical diagnosis termed Symptom Checkers (SCs) are widely available to patients in the US and worldwide. They have been shown to be usable by patients, and can affect patient decision making and care seeking behavior. Evidence from our work and others has shown that, if used correctly, SCs can achieve accuracy of diagnosis and triage close to that of physicians (relying on symptom data). However, most studies are based on case summaries created by physicians, SC apps are not used by patients, and lack evidence on the effects of patient characteristics, or SCs influence on patient decision making. To address these gaps we will evaluate the safety, usability, diagnostic and triage accuracy of a leading symptom checker in use by patients in an emergency department or urgent primary care, and the effect of SC outputs on patients’ decisions to seek care. In aim 1 we will recruit 700 patients to use a SC app from Ada Health when they are seen in urgent primary care or the emergency department at Rhode Island Hospital (RIH), including those with possible symptoms of TIA or stroke. This builds on our previous studies of the Ada SC with 241 patients recruited in these locations. The level of urgency of care they intend to seek will be assessed before and after use of Ada, along with a questionnaire on app usability. Diagnostic and triage accuracy will compared to the assessment of the physician who saw the patient, and the Ada results both compared with, and critiqued by, independent physicians viewing the symptom data collected by Ada. We will also evaluate the effects of different presentations of diagnosis and triage data on patient decision making. In aim 2 we will utilize 2 unique data sets of 2300 patients with possible TIA or stroke seen in the ED at RIH, and use machine learning techniques to create new algorithms to improve early diagnosis and risk stratification. Performance will be compared with existing algorithms and guidelines on accurate diagnosis and effective management of these conditions. We will also analyze a data set of 158,000 patients with possible TIA or stroke who used the Ada app in a community setting, and evaluate the influence of patient characteristics, including age, sex, race, ethnicity, country, and socio- economic group on their symptoms, comparing this to the RIH data set results. The results of these studies will improve our understanding of symptom checker safety, ability to recognize high risk patients and direct them to seek care, and the potential impact on health service use, for a broad range of patients including those with TIA or stroke.

NIH Research Projects · FY 2025 · 2023-07

Survival of an organism relies on its ability to navigate through a complex environment. During navigation, the nervous system integrates spatial information from a wide variety of sensory modalities into a stable heading direction. In insects, such as the fruit fly Drosophila melanogaster, this process takes place in the central complex, a series of neuropil structures in the brain. A hallmark of the central complex is its columnar organization, in which neuronal activity appears as stable bumps that correlate with changes in the organism’s heading direction. The Drosophila central complex has been the subject of extensive anatomical and functional characterization, which has revealed how sensory percepts are transformed through its various compartments. However, the understanding of how neuronal activity in the central complex is translated into behavior is strictly correlational. To establish a causal relationship between central complex activity and heading direction, a thorough behavioral dissection is required. Further, the circuits that relay central complex activity to motor command centers are still poorly understood. Our proposal details a comprehensive research plan to optogenetically manipulate the inputs and outputs of the Drosophila fan-shaped body (FB), a key structure in the central complex. These manipulations will reveal the contributions of the FB input and output neurons to generating a heading direction. To achieve this, we developed a behavior chamber for tracking locomotion of walking flies during optogenetic manipulations. We use this chamber to activate FB neurons in different sensory contexts. Additionally, we will map how outputs from the central complex are routed to command centers in the brain. To this end, we will employ various new configurations of trans-Tango, the transsynaptic circuit mapping and manipulation technique developed by our laboratory. The first of these is trans-Tango(behavior), which allows selective optogenetic manipulation of the postsynaptic partners of a chosen starting population. The second is ds-Tango, a method for mapping neurons mono- and di-synaptic to a chosen starter population. The final is retro-Tango, a transsynaptic mapping tool that works in the retrograde direction. Our studies will reveal how spatial information of sensory cues is translated through multiple layers of circuitry to elicit the navigational drive. Our results will, therefore, represent a key step forward in the knowledge of how the nervous system transforms sensory information into behavior. Finally, our studies will reveal circuit motifs in insects that may be conserved in mammals to perform analogous functions. Therefore, the circuits we describe in insects may contribute to the understanding of spatial perception in humans, one of the first cognitive faculties impacted in Alzheimer's disease.

NIH Research Projects · FY 2026 · 2023-06

Abstract PFAS are a class of man-made organofluorine compounds whose presence in the environment is an emerging, worldwide public health concern and a priority in environmental and human health research. Many PFAS are environmentally persistent, bioaccumulative, and have long half-lives in humans. Of particular concern is that environmental exposure to PFAS may increase the risk of kidney cancer, whose incidence has shown unexplained sustained and rapid increases in the US during the past decades. Several lines of emerging evidence strongly support that PFAS exposures may increase the risk of kidney cancer among humans: First, the kidney is a target organ of PFAS, which cause serious damage to the kidney at the levels commonly experienced by US population; Second, PFAS exposures have been associated with kidney damage that is known to increase kidney cancer risk; Third, a recent population-based case-control study of 324 RCC cases and 324 controls from NCI supports the findings of early occupational mortality studies that PFAS exposure increases kidney cancer risk. Much larger prospective studies of diverse, non-occupationally exposed populations are urgently needed to either confirm or refute these preliminary findings. Here, we propose a prospective nested case-control study to test the association between PFAS exposure and renal cell carcinoma (RCC, more than 85% of all cases of kidney cancer). We will utilize the extensive resources from five large and well-characterized US prospective cohorts: The Southern Community Cohort Study; Hispanic Community Health Study/Study of Latinos Cohort; Nurses’ Health Study; Health Professionals Follow-Up Study; and Physicians' Health Study. We will measure pre-diagnostic plasma levels of 40 PFAS from ~650 incident RCC cases and their individually matched controls (~1,300 controls in 1:2 ratio). Specifically, we will examine the following hypotheses: 1. Environmental exposure to PFAS increases the risk of RCC. In particular, environmental exposure to PFOA, PFOS and PFHxS increases RCC risk; 2. Risk of RCC associated with environmental PFAS exposure varies by mixture patterns, and varies by PFAS chemical structure, branched vs linear isomers, and short vs. long chain length; and 3. Established baseline risk factors of RCC (hypertension, obesity, diabetes and smoking) moderate the relationship between PFAS exposure and risk of RCC. In exploratory aims, we will test if the risk of RCC associated with individual PFAS exposures varies by histologic type of RCC or by race, and thus may contribute to racial disparities in kidney cancer risk. This proposed study fills a critical knowledge gap and represents the first large prospective cohort study to investigate the alleged association between PFAS exposure and kidney cancer risk in the US among diverse cohorts of non-occupationally exposed individuals. The results of the study will inform the medical and public health communities about the potential health effects of PFAS exposure and can help regulators make sound, science-based decisions.

NIH Research Projects · FY 2026 · 2023-06

Feeding is a fundamental behavior that is tightly regulated to precisely meet the metabolic needs of the animal. The primary decision that an animal must make regarding food is whether to ingest it or reject it. Substances with high nutritional value are ingested, while toxins and harmful substances are rejected. To make this decision, the animal relies on its sense of taste to evaluate the quality of the food. Most animals respond to sweet and bitter tastants with different stereotyped behaviors: sweet substances, often calorie rich, are appetitive and accepted, while bitter compounds, usually harmful, are rejected and avoided. Another important part of the decision whether to ingest or reject potential food is the metabolic need of the animal that is manifested by the balance between hunger and satiety. This aspect of the internal state of the animals is evaluated through an intricate balance between various hormones and neuromodulators, some signal hunger while the others signal satiety. How the concerted action of the various hormones and neuromodulators affects feeding is an area of significant interest as dysregulation of feeding behavior results in obesity or in malnutrition and their associated morbidities. In this proposal, we focus on one network of neuromodulatory neurons in Drosophila in which a neuromodulator termed leucokinin is secreted by certain subsets of neurons within the network and detected by others. We propose to test the hypothesis that the leucokinin network of neuromodulatory neurons integrates information about taste quality and the internal state of the fly to modulates feeding behavior. In this network, one set of neurons receives inputs from two others: taste information from one, and information about the internal state of the animal from the other. The recipient neurons integrate the two streams of information and in turn modulate feeding behavior by secreting other neuropeptides that regulate feeding. To test this hypothesis, we use a multipronged approach that includes anatomical, functional and behavioral analyses. In our proposed study, we use state of the art techniques to label neuronal connectivity and to manipulate the activity of selective subsets of neurons to examine the behavioral and functional effects of these manipulations. We also develop a new technique for studying sites of neuromodulation. This technique enables selective, unbiased, brain-wide examination of sites of neuromodulation by specific modulators with cellular resolution. Thus, our studies will deepen our understanding of the regulation of feeding and provide new tools to study neuromodulation, a research area that will increase in importance as neural connectivity maps of more model organisms become available.

NIH Research Projects · FY 2025 · 2023-06

Scientific Abstract: SLC13A5 epilepsy is a newly recognized form of Developmental Epileptic Encephalopathy 25 (DEE25) with seizures beginning within the first days of life along with subsequent intellectual and motor symptoms. In these patients, mutations in the SLC13A5 gene, which encodes a plasma membrane citrate transporter, result in a severe, early onset multi-focal epilepsy and cognitive and behavioral symptoms. How disruption of SLC13A5 function results in dysfunction of neural circuitry is unknown. In patients and in rodent loss of function models of SLC13A5, plasma citrate concentrations are elevated and cytoplasmic citrate is decreased. Since citrate is a precursor to neurotransmitters, diminished glial and neuronal citrate may result in abnormal neuro-transmitter metabolism, contributing to functional defects. However, SLC13A5 loss of function may not account for the full severity of the disorder, and truncations of SLC13A5 are rarely observed in patients. Instead, human genetics show that certain mutations are over-represented as known causative mutations; SLC13A5 G219R (DNA G655A) and T227M (DNA C680T) are the most common recurrent mutations found in approximately two-thirds of all known patients. While the epilepsy is associated with bi-alleleic mutations, the presence of recurrent missense mutations suggests mechanisms more complex than simple autosomal recessive genetics. However, these have not been fully investigated, and no specific treatments for these patients exist. In order to better understand the genetics of SLC13A5 epilepsy, we have developed novel experimental systems. In fly, the entire Drosophila Slc13A5 gene was replaced with the human SLC13A5 coding region. This results in expression of only the human SLC13A5 expressed in the central nervous system. In the humanized line, the G219R mutation causes lethality in contrast to the null, suggesting gain of function mechanisms. In rodents, we show that the equivalent mutation to G219R in mouse SLC13A5 causes more severe epilepsy in direct comparison to the null, again suggesting the over-arching hypothesis of this proposal: that pathogenic mutations in SLC13A5 have gain of function effects, as well as, loss of function effects. However, the understanding of the mechanisms underlying these effects is incomplete, and the determination of both genetic and functional mechanisms are highly important for developing treatments for SLC13A5 epilepsy. We will determine in three Aims to determine:1) what is the normal function of SLC13A5 in brain physiology 2) how do pathogenic mutations in SLC13A5 result in neural dysfunction 3) if novel therapeutic strategies may ameliorate symptoms in SLC13A5 syndrome.

NIH Research Projects · FY 2026 · 2023-05

PROJECT SUMMARY Cannabis use disorder (CUD) is a significant public health concern with origins in adolescence. Cannabis use escalates during the teenage years, and early cannabis use predicts the development of later problems. Leading etiological theories suggest that repeated cannabis use as the brain develops produces changes in reward systems. Over time, these instrumental brain changes alter cannabis effects and increase the incentive salience of cannabis cues, ultimately conferring risk for CUD. This longitudinal study pairs ecological momentary assessment (EMA) in the natural environment and a gold-standard human laboratory (HLAB) paradigm to monitor changes in subjective cannabis effects, cue reactivity, and CUD symptoms across the formative adolescent years. Leveraging smartphone (EMA) reports in natural settings allows for studying adolescents’ reactions to the typically higher potency THC products and varied formulations (e.g., oils, edibles) favored by teenagers. The use of an accelerated longitudinal design allows for charting development from ages 13 to 19 through multicohort assessments completed in a shorter timeframe. We aim to recruit 224 adolescents (ages 13 to 16 at study outset, n = 56 per age cohort) who endorse cannabis use in the past month. Adolescents will complete a baseline HLAB protocol with follow-ups at 1, 2, and 3 years. Each yearly assessment point will also include a 28-day measurement burst of EMA in daily life. Multiple domain latent growth curve modeling will: (Aim 1) characterize age-related changes in sensitivity to rewarding cannabis effects over adolescence (ages 13 to 19); (Aim 2) test prospective relations of CUD symptom progression with change in sensitivity to rewarding cannabis effects; and (Aim 3) test prospective relations of CUD progression with responses to cannabis cues in the natural environment and HLAB. The proposed longitudinal study extends the investigative team’s prior research showing cross-sectional associations of CUD severity with subjective cannabis effects and cue reactivity among adolescents. Our proposal is highly innovative, as well-studied etiological CUD constructs are assessed across adolescence in real-world and laboratory settings using well-operationalized, multidimensional assessments. Further, disaggregating individual differences in change from overall age trends through an accelerated longitudinal design is a sensitive approach that is distinctively innovative. Our proposal addresses a key priority identified by the NIDA Epidemiology Research Branch by efficiently combining the advantages of longitudinal research with behavioral and laboratory-based measures to inform understanding of CUD etiology (NOT-DA-19-066). This proposal will support efforts to prevent the progression of an incubating or emerging CUD by enhancing scientific understanding of the trajectory to more severe harms. Providing new empirical evidence of malleable processes that can serve as targets of preventative interventions has power to reduce the magnitude of societal ramifications of CUD.