SPATIAL TRANSCRIPTOMIC PROFILING REVEALS MOLECULAR SIGNATURES OF PERINEURONAL NET-ASSOCIATED PARVALBUMIN INTERNEURONS

Perineuronal nets (PNNs) are specialized extracellular matrix structures that preferentially ensheath parvalbumin-expressing (Pvalb) fast-spiking interneurons. PNNs regulate synaptic plasticity, protect neurons from oxidative stress, and their disruption has been implicated in neuropsychiatric disorders including schizophrenia and Alzheimer's disease. However, the transcriptomic signature distinguishing PNN-positive from PNN-negative Pvalb neurons remains poorly understood.

Here, we combined Xenium spatial transcriptomics with immunohistochemical detection of PNNs (WFA) to characterize the molecular signatures of PNN-associated neurons in adult mouse brains using a 297-gene panel and cell type classification via Allen’s MapMyCells, as well as anatomical reference registration. Out of 400,000 cells, we identified over 9,000 Pvalb interneurons across five mice, and quantified PNN intensity at single-cell resolution. 97% of PNN+ cells in isocortex, striatum and a small sample of thalamus were Pvalb GABAergic.

While PNNs are nearly exclusive to PV basket cells in isocortex, PNN presence and intensity varies continuously across the population, rather than marking a discrete transcriptomic subcluster. Differential expression analysis revealed that PNN-associated genes show overlapping distributions between PNN+ and PNN− cells, suggesting PNN presence reflects a maturational continuum rather than a binary transcriptional switch. Correlation analysis revealed a coordinated shift in glutamate receptor expression, including a switch in NMDA receptor subunit composition, that tracks with PNN intensity, linking extracellular matrix maturation to synaptic receptor configuration.

Our integrated spatial approach links extracellular matrix organization to single-cell transcriptomic states and provides new insights into the molecular basis of PNN heterogeneity.