Non-serotonergic dorsal and median raphe projection onto parvalbumin- and calbindin-containing neurons in hippocampus and septum

The median raphe nucleus is involved in controlling and maintaining hippocampal activity through its projection to inhibitory neurons in medial septum and hippocampus. It has been shown that anterogradely axonal-traced fibers originating in the median raphe nucleus project onto calbindin-containing neurons in hippocampus and parvalbumin-containing neurons in medial septum. Parallel immunohistochemistry studies showing serotonin fibers contacting calbindin- and parvalbumin-positive neurons have led to the assumption that raphe fibers projecting on these types of neurons are mainly serotonergic. However, in both dorsal and median raphe nucleus there is a large amount of non-serotonergic neurons which also are projecting neurons, indicating that a part of the raphe fibers projecting to hippocampus and septum may be non-serotonergic. Our aim was to determine whether there is a non-serotonergic projection from the raphe nucleus onto calbindin- and parvalbumin-containing neurons in hippocampus and septum. Biotin dextran amine was used as the anterograde neuronal tracer and injected into either dorsal or median raphe nucleus. By use of triple immunofluorescence-labeling we analyzed the serotonergic content of the biotin dextran amine-labeled fibers contacting parvalbumin- and calbindin-positive neurons. Surprisingly, we found a significant non-serotonergic projection from both dorsal and median raphe nuclei onto calbindin- and parvalbumin-containing interneurons in septum and hippocampus, with a preference in hippocampus for projecting onto calbindin-positive neurons. These results indicate that the raphe nuclei may exert their control on hippocampal and septal activity not only through a serotonergic projection, but also through a significant non-serotonergic pathway.