in-vivo-monitoring-circadian-clock-gene-expression-mouse

<em>In Vivo</em> Monitoring of Circadian Clock Gene Expression in the Mouse Suprachiasmatic Nucleus Using Fluorescence Reporters

This technique combines optical fiber mediated fluorescence recordings with the precise delivery of recombinant adeno-associated virus based gene ...

National Institute of Biological Sciences, Beijing

POMC-derived melanocortins inhibit food intake. In the adult rodent brain, POMC-expressing neurons are located in the arcuate nucleus (ARC) and the nucleus tractus solitarius (NTS), but it remains unclear how POMC neurons in these two brain nuclei regulate feeding behavior and metabolism differentially. Using pharmacogenetic methods to activate or deplete neuron groups in separate brain areas, in the present study, we show that POMC neurons in the ARC and NTS suppress feeding behavior at different time scales. Neurons were activated using the DREADD (designer receptors exclusively activated by designer drugs) method. The evolved human M3-muscarinic receptor was expressed in a selective population of POMC neurons by stereotaxic infusion of Cre-recombinase-dependent, adeno-associated virus vectors into the ARC or NTS of POMC-Cre mice. After injection of the human M3-muscarinic receptor ligand clozapine-N-oxide (1 mg/kg, i.p.), acute activation of NTS POMC neurons produced an immediate inhibition of feeding behavior. In contrast, chronic stimulation was required for ARC POMC neurons to suppress food intake. Using adeno-associated virus delivery of the diphtheria toxin receptor gene, we found that diphtheria toxin-induced ablation of POMC neurons in the ARC but not the NTS, increased food intake, reduced energy expenditure, and ultimately resulted in obesity and metabolic and endocrine disorders. Our results reveal different behavioral functions of POMC neurons in the ARC and NTS, suggesting that POMC neurons regulate feeding and energy homeostasis by integrating long-term adiposity signals from the hypothalamus and short-term satiety signals from the brainstem.

Acute and long-term suppression of feeding behavior by POMC neurons in the brainstem and hypothalamus, respectively.

Stress-induced hyperglycemia is a fundamental adaptive response that mobilizes energy stores in response to threats. Here, our examination of the contributions of the central catecholaminergic (CA) neuronal system to this adaptive response revealed that CA neurons in the ventrolateral medulla (VLM) control stress-induced hyperglycemia. Ablation of VLM CA neurons abolished the hyperglycemic response to both physical and psychological stress, whereas chemogenetic activation of these neurons was sufficient to induce hyperglycemia. We further found that CA neurons in the rostral VLM, but not those in the caudal VLM, cause hyperglycemia via descending projections to the spinal cord. Monosynaptic tracing experiments showed that VLM CA neurons receive direct inputs from multiple stress-responsive brain areas. Optogenetic studies identified an excitatory PVN-VLM circuit that induces hyperglycemia. This study establishes the central role of VLM CA neurons in stress-induced hyperglycemia and substantially expands our understanding of the central mechanism that controls glucose metabolism.

A Central Catecholaminergic Circuit Controls Blood Glucose Levels during Stress.

Endogenous circadian clocks control 24-h physiological and behavioral rhythms in mammals. Here, we report a real-time in vivo fluorescence recording system that enables long-term monitoring of circadian rhythms in the brains of freely moving mice. With a designed reporter of circadian clock gene expression, we tracked robust  transcription reporter rhythms in the suprachiasmatic nucleus (SCN) of WT,  , and  mice in LD (12 h light, 12 h dark) and DD (constant darkness) conditions and verified that signals remained stable for over 6 mo. Further, we recorded  transcriptional rhythms in the subparaventricular zone (SPZ) and hippocampal CA1/2 regions of WT mice housed under LD and DD conditions. By using a Cre-loxP system, we recorded  and  transcription rhythms specifically in vasoactive intestinal peptide (VIP) neurons of the SCN. Finally, we demonstrated the dynamics of  and  transcriptional rhythms in SCN VIP neurons following an 8-h phase advance in the light/dark cycle.

Cheng Zhan

National Institute of Biological Sciences, Beijing

In Vivo Monitoring of Circadian Clock Gene Expression in the Mouse Suprachiasmatic Nucleus Using Fluorescence Reporters