The rodent olfactory system comprises at least two complementary central pathways: the main and the accessory olfactory pathway. Chemosensory signals that are detected by nasal olfactory sensory neurons are projected to the main olfactory bulb. However, many semiochemicals and other social chemosignals are detected by vomeronasal sensory neurons and relayed to the accessory olfactory bulb. As principal projection neurons, both main and accessory olfactory bulb mitral cells integrate chemosensory input and forward this information to distinct downstream target regions. Despite separate anatomical investigations of the main or accessory olfactory circuitry, parallel comparative analysis of individual projection paths using modern tracing techniques and largely intact brain samples is lacking. Here, we implemented state-of-the-art viral tracing and confocal imaging to allow for the detailed anatomical depiction of both main and accessory olfactory bulb mitral / tufted cell projections. Cre-dependent expression of adeno-associated viral genomes enabled optical tracing in transgenic mice that selectively express Cre recombinase in olfactory bulb projection neurons under the t-box protein 21 (tbx21) promoter. Three-dimensional reconstruction of olfactory bulb principal neuron projections is achieved by utilizing whole-brain slice preparations and cleared tissue samples.
Altogether, this study provides detailed anatomical insight into unique and common target areas along each olfactory pathway, thus, laying a solid foundation for future investigations into parallel olfactory information processing by the main and accessory olfactory systems.