| Sequential encoding of motor programs is essential for behavior generation.The neuronal sequential activity has been established in the hippocampus and motor cortex during memory or decision making tasks.However,the neural substrate underlying instinctive behavioral sequences such as that in hunting,is still mostly unknown.Hunting is a well-established instinctive behavior with a specific sequential behavior actions,including the prey search,pursuit,attack,and consumption,which can be readily elicited in a laboratory setting.The hunting behavior highly suits to addressing questions of how neural mechanisms drive behavior as it is both reproducible yet also flexible.Lateral periaqueductal gray is located in the downstream of the neural network.LPAG mainly receives projections from the forebrain and a small part receives projections from the brainstem,which are widely projected to the downstream brain regions,playing the role of connecting the above and the below.During hunting,immunohistochemistry showed that c-Fos expression and the activity of LPAG neurons increased,suggesting that this brain region was involved in hunting.However,it is unclear that how the LPAG is involved in the different phases of predatory hunting,the specific cell type and the mechanism of predation.In addition,the LPAG is widely accepted GABAergic inputs associated with predation from the central amygdala(Ce A),lateral hypothalamus(LH)and Zona incerta(ZI),however,how these inputs affect the formation of the dynamic predatory sequence in the LPAG is largely unknown.In this research,we use genetically engineered mice to mark the specific cell type,multichannel recording,fiber photometry,optogenetics,immunohistochemical,apoptosis and combination of adeno-associated virus specific markers to probe the changes of neuronal activity during different phases of predation and predatory mechanism in the LPAG,clear cell type and the role of distinct input loops in the regulation of different predatory phases.The major research contents are as follows:(1)Investigating the changes of LPAG neuronal activity during predatory hunting: By the multi-channels recording and synchronous recording of predation video,it is proved that a neuronal ensemble sequence was formed in the LPAG across predatory process,which was time-locked to different hunting actions.(2)Identifying dynamic change and function of two subpopulations across predatory hunting: Using optrode recording,fiber photometry and optogenetics,it is explored the dynamic changes and regulatory roles of γ-aminobutyric acid neurons(Vgat)and glutamatergic neurons(Vglut2)in the LPAG during different predatory phases.The results demonstrate that Vgat+neurons mainly control the chase and attack phases,whereas Vglut2+ neurons primarily control the attack phase.(3)Elucidating the mechanisms of predatory hunting in the LPAG: By means of optrode recording,masseter electromyogram recording and optogenetics,Vglut2+ neurons mainly encode the execution of movement to cause predation.Optrode recording shows that Vgat+ neurons are involved in reward seeking.Optogenetic manipulation of Vgat+ neurons confirms the adequacy and necessity of positive reinforcement and regulation of motivational drive,thus proving that Vgat+ neurons drive motivation to induce predation.(4)Revealing the integration of distinct upstream inputs in the LPAG: Using slice electrophysiology to assess functional connectivity of Ce A/LH/ZI-LPAG pathway.Ablation of GABAergic inputs of Ce A,LH and ZI by apoptosis respectively suppresses predation,and indicates that different clusters involved in predation are damaged respectively in the LPAG by using multi-channel recording,and affected the formation of activity pattern dynamics.These results confirm the intact sequential activity pattern is necessary for efficient hunting actionsIn summary,the central nervous system provides stereotypical but flexible functional neuronal ensembles to encode instinctive behaviors.When an instinctive behavior generates,a series of neuronal ensembles are activated sequentially,which chains various behavioral actions into sequences.The behavioral actions are considered to be stereotyped,where one neuronal ensemble activated may induce only one fixed type of behavioral output.In contrast,the behavioral sequence is flexible,where the order of ensembles could be adjusted by earlier experience(motor planning,Top-down)and sensory evidence(close-range interaction,bottom-up).In such cases,behavioral sequences may be hierarchically organized and influenced by internal states or sensory stimuli.We reason that in this manner,the behavior is robustly performed with low cost but still retains flexibility.Thus,we provide a framework of neuronal ensemble sequences across predatory hunting,which may shed light on other instinctive behaviors. |
