The Effect And Mechanism Of Regulating Feeding Behavior During Nocturnal Wakefulness In The Lateral Hypothalamic Astrocytes

Objective:Obesity is a serious medical and social problem,which seriously interferes with people’s life and work,and even leads to a series of complications,bringing a heavy burden to the family and society.Although a large number of literatures have shown that sleep disorders can promote obesity,however the neural mechanisms regulating sleep-wake behavior and feeding behavior and their interactions are still unclear.This project applies the chemogenetics technology to activate astrocytes in the lateral hypothalamus specifically.We find astrocytes have an important effect on sleep-wake behavior and feeding behavior through activating gamma-aminobutyric acid neurons with the help of the EEG/EMG record,electrophysiology,calcium imaging techniques.This project is helpful to clarify the neurobiological mechanism of the lateral hypothalamus regulating sleep-wake behavior and feeding behavior,and to improve the regulatory theory that sleep deficiency contribute to obesity because of the increase of feeding behavior.It is expected to provide new therapeutic targets and ideas for the clinical treatment of obesity patients caused by sleep disorders.Methods:1.Experimental animal grouping: C57BL/6J mice and VGAT-Cre mice were randomly divided into experimental group and control group.2.Animal surgery: virus was injected into LHA through stereotactic technology,and electrodes or optical fibers are embedded over the LHA at the same time.3.Optical fiber recording experiment:The activity changes of LHA astrocytes during spontaneous sleep-wake and feeding behaviors were recorded using a monochromatic multi-channel optical fiber recording system.The activity changes of and astrocytes and GABAergic neurons after activation of LHA astrocytes were recorded using a dual-color fluorescence signal system.4.Polysomnography and data analysis: The changes of sleep-wake behavior were recorded in active and inactive period after activation or inhibition of LHA astrocytes by CNO.5.Behavioral experiment: LHA astrocytes were activated to observe the changes of feeding behavior and predation behavior of mice after CNO injection.6.Gene identification: Tissue of the hybridized mice(10-15g)was taken,and the supernatant of the tissues was cleaved for PCR and electrophoresis,and VGAT-Cre mice were screened.7.Immunofluorescence fluorescence technique: 2h after the injection of CNO,the brain was collected by heart perfusion,and then the expression of virus was observed by immunofluorescence histochemistry technique after frozen sections.Result:1.Astrocytes in the lateral hypothalamus promote arousal behavior.(1)Immunohistochemical results showed that most e GFP virus expressed cells were GFAP positive,while only a few Neu N-positive cells were GFAP expressed cells.(2)In the spontaneous sleep-wake behavior,the calcium signal activity during the transition from wakefulness to non-REM sleep in the experimental group decreased,while the calcium signal activity during the transition from NREM sleep to wakefulness increased.(3)Activation of astrocytes in the lateral hypothalamus of the experimental group promoted long-term arousal.The total duration of arousal 5 hours after injection of CNO in the inactive period increased(P<0.001),the total duration of NREM sleep decreased(P<0.001),and the total duration of REM sleep decreased(P<0.001).After injection of CNO in the active phase,the total duration of wakefulness increased(P<0.001),the total duration of wakefulness in non-REM sleep decreased(P<0.001),and the total duration of REM sleep decreased(P<0.001).(4)Astrocytes in the lateral hypothalamus of the experimental group had a tendency to inhibit arousal,but there were no statistical differences in arousal,NREM sleep and REM sleep per hour after injection,but the total duration of arousal 5 hours after injection of CNO decreased(P<0.01),and the total duration of NREM sleep increased(P<0.01)in the active phase.2.Astrocytes in the lateral hypothalamic promote feeding behavior during nocturnal wakefulness.(1)The activity of astrocytes in lateral hypothalamus was significantly increased during feeding(P<0.001).(2)Activation of lateral hypothalamic astrocytes promoted feeding behavior in inactive stage,and the feeding amount(P<0.001),cumulative feeding time(P<0.001)and feeding times were significantly increased after injection of CNO(P<0.001).(3)Activation of lateral hypothalamic astrocytes promoted the increase of exercise(P<0.001).(4)Activation of lateral hypothalamus astrocytes could promote predation behavior,and predation amount(P<0.05)and attack times were significantly increased(P<0.001).3.Activation of astrocytes in the lateral hypothalamus promotes the activation ofγ-aminobutyric acid neurons in this region.(1)Activation of astrocytes in the lateral hypothalamus significantly increased the activity of neurons in this region(P<0.001).(2)Activation of astrocytes in the lateral hypothalamus significantly increased the activity of GABAergic neurons in this region(P<0.001).Conclusion:Lateral hypothalamic astrocytes promote feeding behavior during nocturnal wakefulness,and there is an underlying mechanism that the lateral hypothalamic astrocytes promote feeding behavior during nocturnal wakefulness through activation of lateral hypothalamic GABAergic neurons.