Neuromethods: Basic Techniques for Evaluating the Nervous System in Nonclinical Studies.

The first session of the 2025 European Society of Toxicologic Pathology (ESTP) Congress reviewed routine and specialized methods for microscopic evaluation of neural tissues during nonclinical studies. Three longer presentations reviewed brain sampling approaches in safety assessments, including an example to accentuate topographical analysis and integration of toxicology data; specific brain and spinal cord sampling for molecular and protein analyses; and an overview of technical aspects of intraparenchymal drug delivery. Four shorter talks discussed the uses, advantages, disadvantages, and interpretation of several special neurohistological techniques (stains and immunohistochemical markers) for assessing test item-associated responses. Common special methods used (when warranted) for nonclinical studies include Fluoro-Jade or silver stains for detecting neuronal death, Luxol fast blue (LFB) for examining myelin, anti-glial fibrillary acidic protein (GFAP) to demonstrate reactive astrocytes, and anti-ionized calcium-binding adaptor molecule 1 (IBA1) to highlight reactive microglia and macrophages, though alternatives methods were described. The last presentation discussed artificial intelligence as an aid in detecting subtle toxicant-induced lesions during digital pathology analyses (using the Olney lesion [acute neuronal vacuolation and necrosis in the cerebral cortex] as an example). Taken together, talks in this session provided a cohesive overview of traditional and innovative approaches to facilitate microscopic evaluation for potential neurotoxicity in nonclinical studies.