Background The accumulation of amyloid beta is a diagnostic hallmark of Alzheimer’s disease. However, the cell types that are most sensitive to its accumulation, and the transcriptional pathways that are induced by amyloid beta in specific cell types, remain incompletely understood. Method We performed single nucleus RNAseq analysis (n = 1.1 million profiles) on frontal cortical biopsies (n = 58) from individuals undergoing shunt placement for idiopathic normal pressure hydrocephalus (iNPH). Of these 58 biopsies, 21 were positive of amyloid beta, and 8 were positive for both amyloid beta and hyperphosphorylated tau. We clustered the profiles into 92 distinct clusters, and performed an integrative analysis with 30 published human and mouse datasets. We tested for proportional loss or gain of specific populations in the presence of amyloid or amyloid + tau, and identified differentially expressed genes in each population. Results Activated microglial and astrocytic populations were proportionally over‐represented in the pathology‐containing biopsies. Compared with postmortem datasets, the biopsy‐derived profiles showed dramatically lower activation of immediate‐early gene signatures, suggesting stronger fidelity of measured transcriptional programs to the in vivo human cell states. Comparative analysis of microglial populations with published datasets of other diseases revealed transcriptional pathways specifically induced by amyloid accumulation. Specific subsets of layer 1 interneurons were under‐represented both in amyloid‐positive biopsies, and in published datasets generated from early (Braak stage < 4) postmortem tissue. Conclusions The use of fresh cortical biopsies enabled a faithful measurement and analysis of glial states induced by the accumulation of amyloid beta. Specific subsets of layer 1 interneurons are particularly sensitive to depletion by amyloid beta accumulation in frontal cortex.