Chemical Genetic Regulation Of CA3 Pyramidal Neurons Affects Hippocampal High-frequency Oscillations In Epilepsy

Temporal lobe epilepsy(TLE)is a type of focal epilepsy that recurs,and its onset is related to transient abnormal excitation of neurons in the brain,mainly manifested as hypersynchronous activity of brain neurons.Traditional EEG studies of epilepsy typically focus on low-frequency signals in the 0.5-80 Hz frequency range,and in recent years,high-frequency oscillations(HFOs)have become widely recognized.Studies suggest that the appearance of HFOs may be closely related to paroxysmal activity in the epileptogenic or seizure-generating area,and may be an important biological marker of epilepsy onset.At the same time,studies have shown that the dynamic coupling between HFOs and lowfrequency EEG signals may reflect the occurrence and development process of epilepsy,but the neural mechanism of this process is currently unclear.The main lesion of TLE occurs in the hippocampal region,manifested as hippocampal tissue atrophy sclerosis and a large number of lost neurons.Among them,CA3 is considered to be an important "relay station" for the propagation of epilepsy in the hippocampal network.The excitatory projection of its pyramidal neurons plays an important role in the occurrence and development of epilepsy,but how CA3 pyramidal neurons regulate the onset of epilepsy is not fully understood.In this study,based on the lithium chloride-pilocarpine-induced epileptic mouse model,hippocampal CA3 pyramidal neurons were inhibited by chemical genetic means,and the mouse’s epileptic behavior,local field potential and neuronal discharge of CA3 and CA1 areas in different states were recorded.The characteristics of the mouse’s epileptic behavior,HFOs,highlow frequency signal coupling,and the relationship between HFOs and pyramidal neuron discharge were analyzed before and after chemical genetic intervention,in order to explore the significance of HFOs and high-low frequency signal coupling in epileptic seizures.The main research results are as follows:1.Statistical analysis of epileptic behavior in mice found that inhibiting hippocampal CA3 pyramidal neurons significantly prolonged the latency period of systemic seizures and sustained seizures,and significantly reduced the number of systemic seizures and the total duration of seizures,suggesting that inhibiting hippocampal CA3 pyramidal neurons can significantly alleviate epileptic seizures in mice.2.Statistical analysis of the characteristics of HFOs found that the number,duration,and amplitude of ripples(80-250 Hz)and fast ripples(FRs,250-500 Hz)in HFOs varied in different states.The number of FRs significantly increased during epileptic seizures,and significantly decreased when CA3 pyramidal cell discharge was inhibited.This suggests that the roles of the two in epileptic seizures are different,and compared to ripples,FRs can better characterize the state of epileptic seizures.3.The results of the high-low frequency phase locking analysis showed that ripples were significantly coupled with some low-frequency signals in different conditions.Inhibiting the excitability of CA3 pyramidal neurons resulted in significant coupling between ripples and low-frequency signals in all frequency bands,indicating that altering the activity of CA3 pyramidal neurons can affect the coupling between ripples and lowfrequency signals during epileptic seizures.In the baseline state,FRs had very few significant coupling relationships with low-frequency signals,but as the seizures developed,the significant coupling relationships increased.Inhibiting CA3 pyramidal neurons did not change the significant coupling relationship between FRs and lowfrequency signals during large-scale seizures.This suggests that the coupling relationship between FRs and low-frequency signals may indicate the onset of epilepsy rather than the degree of epileptic seizures.4.The results of the phase relationship between FRs and pyramidal neuron discharge showed that during large-scale epileptic seizures,the discharge rate of neurons within the duration of FRs was significantly increased.Inhibiting CA3 pyramidal neurons significantly reduced the discharge rate of neurons within FRs,and neurons were more likely to discharge at the troughs of FRs.This suggests that the generation of FRs is directly related to the excitatory discharge of pyramidal neurons.In summary,this study used chemical genetic methods to inhibit the activity of CA3 pyramidal neurons in the hippocampus,and investigated the changes in HFOs and highlow frequency coupling during epileptic seizures.The relationship between ripples/FRs and the state of epilepsy was explored.The results suggest that FRs are a better indicator of the state of epileptic seizures and may serve as an important biological marker for the onset of epilepsy.This study provides new insights for further exploring and understanding the neural mechanisms underlying the onset of epilepsy.