Impact of Alzheimer's disease risk factors and local neuromelanin content on the transcriptomic landscape of the human locus coeruleus

The locus coeruleus (LC) is a small noradrenergic nucleus in the dorsal pons that sends projections widely across the brain to regulate multiple functions including sleep, memory, arousal, and cognition. The LC has several unique cellular features including the presence of neuromelanin (NM) and its accumulation of a precursor to Alzheimer's Disease (AD) pathology, soluble phospho-tau (pTau), in virtually all individuals by adulthood. NM increases in LC during early life, plateauing in middle age, and eventually dissipating, coinciding with LC neuron degeneration, a process which is observed in healthy aging but accelerated in AD. NM-sensitive neuroimaging predicts clinical severity and future progression of AD. While these findings suggest an etiologic role for the LC in AD, data describing the molecular landscape of the LC in middle age, preceding the clinical manifestation of sporadic AD, is lacking. Understanding the molecular state of the LC during this time period is critical to developing interventions that preserve this brain region and the cognitive functions it supports. Here, we performed Visium spatial transcriptomics on 85 tissue sections of human postmortem LC from a cohort of 33 middle-aged, neurotypical donors, balanced for epidemiologic AD risk factors including sex, African or European ancestry, and APOE genotypes (carriers of the E4/risk or E2/protective alleles). We find an unexpected male-bias in LC expression of cholesterol synthesis pathway genes, suggesting that intrinsic neuronal cholesterol synthesis may be protective against AD. Comparing across APOE genotypes, astrocytic gene expression differs in the immediate vicinity of LC neurons. Finally, we leverage histological images captured from the profiled tissue to associate gene expression to NM abundance, finding that APOE expression is associated with reduced NM content, and that NM-associated genes are enriched for aging-related pathways. These findings provide important information about how AD risk factors and NM content impact molecular correlates of resilience/susceptibility of the aging LC to degeneration. Data are freely available, including through visualization browsers to enable wide utilization of these tools for understanding LC biology in AD and other LC-related neurodegenerative disorders, including Parkinson's disease. Overall design: We obtained tissue blocks of postmortem human brain containing locus coeruleus (LC) from 33 neurotypical adult brain donors, balanced for age, sex, APOE2 and APOE4 carrier status (all donors had at least one copy of one of the two alleles), and predominant genomic ancestry (African Ancestry or European Ancestry). We used the 10x Genomics Visium platform to generate spatial transcriptomics from two sequential tissue sections containing LC for each donor, with a second pair of tissue sections collected for a subset of donors. Data were integrated and clustered, defining a robust LC cluster, two mixed non-LC neuron clusters, and several clusters labeling spots with mixed glial types contents. 77 samples from 30 donors with substantial LC content (=10 spots per tissue section) were retained for downstream analyses. Differential expression (DE) analyses were then performed for the 5 clusters (of 9 total) with adequate count data, including LC, comparing expression between groups for each Alzheimer's risk variable of interest (sex, genomic ancestry, and APOE genotype). Additional analyses were performed for: 1) spatially variable expression within spots of the LC cluster; 2) comparing neuromelanin (NM)containing LC spots NM-negative LC spots; 3) modeling spot-level NM relationships to spot-level gene expression; 4) nonnegative matrix factorization of LC cluster gene expression to identify discrete expression programs; 5) gene set enrichment analyses of DE genes, NM-associated genes, and factorization-identfied LC gene groups.