Neuroscience has advanced over the years largely due to animal experiments, particularly in mice. These experiments are generally challenging and require thorough preparation to be successfully carried out. The training required to perform procedures on mice must be rigorous to minimize the risk of errors that could lead to experimental failure and, equally important, to prevent unnecessary suffering of the animals involved. In this study, we present a detailed description of the surgical procedure for intrahippocampal injection in mice using a motorized stereotaxic system equipped with synchronized drilling and microinjection modules. The protocol emphasizes precise anatomical targeting, controlled infusion parameters, and standardized procedural steps designed to enhance reproducibility and minimize tissue trauma. Key aspects of the technique include stereotaxic atlas alignment, skull reference acquisition, controlled drilling to the dura mater, and microinjection of small tracer volumes under physiologically relevant conditions. This methodological framework provides a reliable platform for investigating brain parenchymal transport mechanisms, including intramural periarterial drainage pathways implicated in neurodegenerative disorders such as Alzheimer’s disease.
