Electrophysiology studies in vitro have generally focused on forms of plasticity which are rapidly induced and last for minutes to hours. However, it is well known that plasticity at some cellular and synaptic loci are induced and expressed over many hours or days. One limitation in examining these forms of plasticity is the lack of preparations that allow stimulation and recording of the same tissue over a 24h period or more. Here we describe a simple method for repeated recordings and stimulating the same organotypic slices (different neurons) over a 24h window. We use the conventional interface organotypic culture method together with a custom chamber, which allows recordings on the intact filter, and DiI to mark the stimulation sites. We show that the health of the neurons, as defined by intrinsic excitability, excitatory and inhibitory input-output curves, and morphology remains unchanged over the 24h period. This simple technique provides a means to investigate long-term forms of plasticity that may be induced under conditions similar to those observed in vivo. Additionally, it provides the opportunity to perform long-term morphological and pharmacological studies.