Neurodegenerative proteinopathies characterized by intracellular aggregates of tau proteins, termed tauopathies, include Alzheimer's disease (AD), frontotemporal lobar degeneration (FTLD) with tau pathology (FTLD-tau), and related disorders. Pathological tau proteins derived from human AD brains (AD-tau) act as proteopathic seeds that initiate the templated aggregation of soluble tau upon intracerebral injection into tau transgenic (Tg) and wild-type mice, thereby modeling human tau pathology. In this study, we found that aged Tg mice of both sexes expressing human tau proteins harboring a pathogenic P301LMAPTmutation labeled with green fluorescent protein (T40PL-GFP Tg mouse line) exhibited hyperphosphorylated tau mislocalized to the somatodentritic domain of neurons, but these mice did not developde novoinsoluble tau aggregates, which are characteristic of human AD and related tauopathies. However, intracerebral injections of either T40PL preformed fibrils (PFFs) or AD-tau seeds into T40PL-GFP mice induced abundant intraneuronal pathological inclusions of hyperphosphorylated T40PL-GFP. These injections of pathological tau resulted in the propagation of tau pathology from the injection site to neuroanatomically connected brain regions, and these tau inclusions consisted of both T40PL-GFP and WT endogenous mouse tau. Primary neurons cultured from the brains of neonatal T40PL-GFP mice provided an informativein vitromodel for examining the uptake and localization of tau PFFs. These findings demonstrate the seeded aggregation of T40PL-GFPin vivoby synthetic PFFs and human AD-tau and the utility of this system to study the neuropathological spread of tau aggregates.SIGNIFICANCE STATEMENTThe stereotypical spread of pathological tau protein aggregates have recently been attributed to the transmission of proteopathic seeds. Despite the extensive use of transgenic mouse models to investigate the propagation of tau pathologyin vivo, details of the aggregation process such as the early seeding events leading to new tau pathology have remained elusive. This study validates the use of GFP-labeled tau expressed by neuronsin vivoandin vitroas models for investigating mechanisms underlying the seeded transmission of tau pathology as well as tau-focused drug discovery to identify disease-modifying therapies for AD and related tauopathies.