Background: The study of viruses is inseparable from their host, and the relevance of in vitro models is dependent on their ability to mimic an aspect of the in vivo environment. Technological, ethical, and legislative developments have led to the emergence of organoid models in many species. The study of equine herpesviruses could in turn benefit from this new approach. Objectives: To validate a cerebral organoid model derived from equine reprogrammed pluripotent stem cells (ePSCs) to obtain a new relevant in vitro 3D model to study host-pathogen interactions, in particular neurotropic viruses. Study design: Development of new 3D models and proof of concepts. Methods: Equine cerebral organoids (eCOs) were generated from ePSCs in neural induction medium and grown individually in wells. At D50, the eCOs were infected with one of the three EHV-1 strains (A2254, G2254, and C2254) respectively or with a reference EHV-4 strain at 10000 or 1000 pfu. After 5 days, nucleic acid extraction from organoids and their culture supernatants was performed and viral load of EHV-1 or EHV-4 was determined as previously described.[1] Results: After 5 days post-contact with 10 000 pfu/ml, all organoids are positive for EHV-1 and the viral load increased by more than 3 Log. An increase of the viral load in the culture supernatant was also measured. The same observations were made after infection with 1000 pfu/ml. Organoids appear to be also permissive for EHV-4 even if the strain seems to replicate less than the 3 EHV-1 strains. Main limitations: Lack of specific antibodies for immunocytochemistry of eCOs. Conclusions: These results show for the first time the capacity of the eCOs model to replicate three different strains of EHV-1 and one EHV-4 strain, opening the way for further studies on cell-virus interactions in a neural environment. Ethical animal research: Not applicable. Informed consent: Not applicable. Competing interests: None declared. Funding: Eperon HVE4 IRCP Fund N49-2019, CRB-ANIM-ANR-11-INBS-0003 to BP. Reference: [1]: Thieulent CJ, Hue ES, Fortier CI, Dallemagne P, Zientara S, Munier-Lehmann H, Hans A, Fortier GD, Pitel P-H, Vidalain P-O, et al. Screening and Evaluation of Antiviral Compounds against Equid Alpha-Herpesviruses Using an Impedance-Based Cellular Assay. Virology 2019;526:105–116.