Endocrine Abstracts

Corticotropin-releasing hormone (CRH), a 41-aminoacid peptide that is mainly produced in the hypothalamic paraventricular nucleus (PVN), plays a crucial role in stress response and is considered to be the central driving force in the activity of hypothalamic–pituitary–adrenal (HPA) axis. Previous studies of neuronal input to the PVN were limited by experimental techniques and could not be restricted by a single neuron type. In the present study, by using a novel deficient rabies virus combined with the CRF-Cre transgenic mouse, which not only limits the start neurons within CRF neuron in PVN, but also restrict the input neurons to monosynaptic scale, we investigated the input projections of CRH neurons in the PVN. We observed the monosynaptic input projections received by CRH neurons in the whole brain and discovered projection neurons in forebrain, midbrain, hindbrain and other brain regions except in olfactory bulb. Basal forebrain, preoptic area, hypothalamus, brainstem reticular nucleus all had a large number of fluorescence-labeled projection neurons. Remarkably, the direct projections from cortex have not been reported before. These neurons distributed in layer 5 and 6 of orbital cortex, motor cortex and piriform cortex, and other sub-regions. Benefit from the good neuronal morphology revealed by virus tracing, we could observe that most of the labeled neurons obviously had the morphological characteristics of pyramidal neurons, such as the apical dendrites and branches, basal dendrites and the axon structures extending downward in cortical surface. Dendritic spines could also be clearly visible. We further obtained the input projection imaging data of CRH neurons with single cell resolution in the whole brain range using fluorescence micro-optical sectioning tomography system (fMOST). After data processing, we obtained the 3D distribution pattern of input projection neurons in the entire brain which demonstrated the detail morphologies of projection neurons inside mPFC, BST, amygdala and PVN, etc. as well as their local connections. We also dissected the entire morphologies of labeled pyramidal neurons within mPFC and traced their axons to the region near the injection site. The above neuronal tracing and reconstruction experiments have demonstrated the CRH-neuron-specific monosynaptic inputs to PVN at the whole brain scale for the first time.