Chondroitin sulfate (CS) is a type of glycosaminoglycan (GAG) that is abundant in cartilage and perineural networks (PNNs). Changes in the CS signature of PNNs have been implicated in several neurological diseases. Most CS-GAGs contain labile sulfate groups, which can be lost during ionization events that deposit large amounts of internal energy. Infrared matrix-assisted laser desorption electrospray ionization (IR-MALDESI) is a soft ionization technique used for mass spectrometry imaging. In this work, we determine the spatial distribution of CS-GAG disaccharides within rodent brain using IR-MALDESI MSI. Non-, mono-, and disulfated disaccharides were detected with various adducts. All disaccharides colocalized to the PNNs, which are most abundant in the cortex and hippocampus regions of the brain. This is the first MSI study to spatially resolve CS-GAG disaccharides within brain, paving the way for IR-MALDESI to measure GAGs in neurological diseases.