Perineuronal nets (PNNs) are specialized extracellular matrix structures observed in the adult brain and play important roles in regulating synaptic plasticity. In this study, we examined the formation of PNNs in the mouse hypothalamus with Wisteria floribunda agglutinin (WFA), a broad PNN marker, and unexpectedly found a previously-unidentified region locating between the fornix and paraventricular nucleus (PVN). To characterize this new region, we performed studies of histochemistry, DNA microarray, neuronal tracing, and neuronal activation analysis by investigating c-Fos expression. Nissl staining could discriminate this region from the neighboring PVN and the central part of the anterior hypothalamic area by differences in neuronal density. We designatted this new region the “delta area” (DA) referring to its characteristic triangular morphology. Microarray and histochemical experiments identified three distinct types of neurons in the DA, enkephalin- and calretinin-positive neurons, both of which were gamma-aminobutyric acid (GABA) -negative, and GABA-positive neurons. Furthermore, the DA was shown to have bidirectional neural connections with the lateral septum (LS) and some of intra-hypothalamic nuclei; ventromedial hypothalamus and dorsal part of the premammillary nucleus. We also confirmed enkephalinergic projections from DA neurons to the LS, and inversely, calbindin-positive LS neurons were identified as an afferent source of the DA. Finally, c-Fos expression analysis revealed that activity of DA neurons tended to be increased by psychological stressors such as the open filed, restraint, and aggressive behaviors, but not by metabolic stimuli of fasting/refeeding and dehydration/rehydration. These results suggest that the DA is a new hypothalamic area with specific functions, which links the hypothalamus to the limbic system.

This work was supported by MEXT to MN and NH.