Early life influenza infection and glial dysregulation

SUMMARY Millions of children in the United States contract respiratory infections such as influenza, RSV, and COVID-19 every year, with those under the age of five affected the most severely1,2. Respiratory infections can have long- term effects on the central nervous system in both adults and children3-11. In fact, currently, 6 million children are suffering from long COVID, and up to 44% of these children experience cognitive impairment3. Childhood is a critical period for synaptic formation and myelination, making the brain particularly vulnerable to insults that can lead to permanent and long-lasting neurocognitive changes12. In rodents, postnatal day (P)14 is a crucial period of peak synaptogenesis and myelination12-14. As a result, during this time, a respiratory immune challenge such as influenza can result in persistent neurodevelopmental dysfunction. While many studies have investigated the neurological effects of systemic immune challenges during the prenatal and adult periods15-18, the impact of influenza infection during early postnatal life remains understudied. The proposed study aims to investigate the effect of influenza infection at P14 on glial dysregulation in mice. The overarching hypothesis is that influenza infection results in multicellular dysfunction driven by aberrant microglia. My preliminary data show that influenza infection at P14 leads to an increase in reactive microglia one-week post-infection in the cingulum and dentate gyrus, along with elevated expression of inflammatory genes in microglia as revealed by single-nuclei RNA sequencing. I will employ advanced transcriptomic analysis to examine changes in glial sub-states, and to identify alterations in synapse/pruning-associated genes (Aim 1A). Additionally, changes in microglia-mediated synaptic pruning will be assessed using Imaris 3D reconstructions to quantify microglial engulfment of synapses following infection. My preliminary findings indicate that influenza infection results in a decrease in OPCs and oligodendrocytes in the cingulum and dentate gyrus one-week post-infection. I will use an established optogenetics paradigm19,20 to stimulate excitatory dentate gyrus neurons and test for changes in activity- dependent myelination following infection (Aim 2). Finally, based on the hypothesis that reactive microglia dysregulate oligodendroglial dynamics, microglia will be depleted between P7-P21 to determine if this rescues the observed loss of oligodendroglial cells (Aim 3). Given my experiences investigating the impact of prenatal environmental immune challenges on microglia and behavior, I am well-prepared to execute these experiments. This work will be conducted under the sponsorship of Michelle Monje, MD/PhD, a leading expert in glial-neuron interactions and myelin plasticity. The co-sponsorship of Catherine Blish, MD/PhD, will ensure comprehensive guidance on virologic aspects, and collaborations with Karl Deisseroth, MD/PhD, Akiko Iwasaki, PhD, and Beth Stevens, PhD, will provide the necessary support to successfully achieve the aims of this project. Stanford's rigorous training environment and the collective support from these experts will ensure the project's success and my growth into an innovative physician-scientist capable of developing therapies for neuroimmune diseases.