Neuroanatomical organization and functional roles of PVN MC4R pathways in physiological and behavioral regulations

Objective The paraventricular nucleus of hypothalamus (PVN) is an integrative center in the brain orchestrating a wide range of physiological and behavioral responses. While the PVN melanocortin 4 receptor (MC4R) signaling (PVNMC4R+) is undoubtedly involved in feeding regulation, the neuroanatomical organization of PVNMC4R+ pathway and its role in diverse physiological and behavioral regulations have not been fully understood. Here we aimed to better characterize the input-output organization of PVNMC4R+ neurons and further test their potential functional roles beyond feeding. Methods Using a combination of viral tools, we performed a comprehensive mapping of PVNMC4R+ circuits and tested the effects of chemogenetic activation of PVNMC4R+ neurons on thermogenesis, cardiovascular control and other behavioral regulations beyond feeding. Results We found that PVNMC4R+ neurons broadly innervate many different brain regions known to be important not only for feeding but also for neuroendocrine and autonomic control of thermogenesis and cardiovascular function, including but not limited to preoptic area, median eminence, parabrachial nucleus, locus coeruleus, nucleus of solitary tract, ventrolateral medulla and thoracic spinal cord. Contrary to broad efferent projections, PVNMC4R+ neurons receive monosynaptic inputs from limited brain regions, including medial preoptic nucleus, arcuate and dorsomedial hypothalamic nuclei, and supraoptic nucleus. Consistent with broad efferent projections, chemogenetic activation of PVNMC4R+ neurons not only suppressed feeding but also led to an apparent increase in heart rate, blood pressure and brown adipose tissue thermogenesis. Strikingly, these physiological changes accompanied an unexpected repetitive bedding-removing behavior followed by hypoactivity and resting-like behavior. Conclusions Our results clarify the neuroanatomical organization of PVNMC4R+ circuits and shed new light on the roles of PVNMC4R+ pathways in autonomic control of thermogenesis, cardiovascular function and other behavioral regulations. ### Competing Interest Statement The authors have declared no competing interest. * AP : Area postrema ARC : Arcuate nucleus BNST : Bed nucleus of the stria terminalis BP Blood pressure ChR2 : Channelrhodopsin-2 CNO : Clozapine-N-oxide CVLM : Caudal ventrolateral medulla DAP Diastolic arterial pressure DMH : Dorsomedial nucleus of hypothalamus DMV Dorsal motor nucleus of the vagus DREADD : Designer Receptors Exclusively Activated by Designer Drugs Flp Flippase HR : Heart rate IHC : Immunohistochemistry LC : Locus coeruleus LHA : Lateral hypothalamic area MAP : Mean arterial pressure MC4R : Melanocortin 4 receptor ME : Median eminence NTS : Nucleus of solitary tract PAG : Periaqueductal gray PBN : Parabrachial nucleus POA : Preoptic area PVN : Paraventricular nucleus of hypothalamus PVT Paraventricular nucleus of thalamus RV : Rabies virus RV-EnvA-ΔG : EnvA-pseudotyped G-deleted Rabies viruses RVLM Rostral ventrolateral medulla SAP : Systolic arterial pressure TSC : Thoracic spinal cord VMH : Ventromedial nucleus of hypothalamus vSUB Ventral subiculum