ABSTRACT Series of disappointing clinical trial outcomes have ushered the Alzheimer's field (AD) into an era of preventive interventions. Researchers now recognize that AD is a continuum and that interventions should start in the pre-symptomatic phase. To that end, transcutaneous vagus nerve stimulation (tVNS) is a promising non- invasive approach, as its mechanisms have been attributed to the brain system that is initially affected by AD pathology, the norepinephrine locus coeruleus (LC) system. Animal and preliminary studies in patients and healthy individuals from our group and others demonstrated that tVNS alters locus coeruleus (LC) and nucleus tractus solitarius (NTS) functioning and enhances memory. This indicates that tVNS targets the site of initial AD pathology and might have the potential to delay AD-related cognitive impairment. Thus, tVNS could address an important challenge in the field: non-invasively delaying disease progression prior to onset of cognitive decline or significant accumulation of pathology. The overarching goal of this proposal is to apply single and repeated tVNS in pre-symptomatic older individuals with varying degrees of AD pathology, with the aim to determine the extent of the cognitive effects of tVNS in domains and time, and to relate tVNS outcome to demographics, neurophysiological properties of the LC and NTS as well as burden of AD biomarkers. Our accomplishments in optimizing tVNS and sensitive cognitive measures, ultra-high field brainstem imaging and blood-based biomarkers allow us to examine our central hypothesis: that serial tVNS enhances memory functioning more than single, and in particular in at-risk individuals, in whom AD pathology burden is low to moderate and the NTS-LC system is still responsive to stimulation. To that end, 140 pre-symptomatic older individuals (APOE-E4 enriched) will be enrolled to a double-blind randomized cross-over design of stimulation versus sham tVNS during 7T imaging and blood sampling. This will be followed by randomized allocation to repeated tVNS or sham for 2 weeks and a follow-up cognitive assessment after 2 months. The results of this study will yield important information for future trials assessing tVNS in three important ways: 1) through investigating which cognitive domains are modulated by tVNS on the short and long-term, important for monitoring and determining outcome measures, 2) through identifying demographic characteristics, functional brain and AD-related markers that predict beneficial responses to tVNS, which will be important to identify trial eligibility (Aim 1 and 2) and 3) understanding biological pathways contributing to RAVANS success to confirm target engagement, aid in biomarker stratification or enrichment of the population, and which could serve to monitor progression (Aim 2 and 3). The research proposed is innovative because it aims to define the target population in whom tVNS can be efficacious, based on the known underlying biological p...