The Formation And Regulation Of Auditory Steady State Response In Thalamo-cortical Neural Network

Objective: Auditory Steady State Response(ASSR)is an electrophysiological response to rapid periodic auditory stimuli with phase-locked frequency and phase.ASSR is a commonly used biological marker to evaluate neurological function in clinical electroencephalogram examination,which can effectively evaluate the degree of neurological diseases such as depression and schizophrenia,and is also widely used in animal model experiments of neuropharmacology.Previous studies have shown that sound stimulation at the Gamma frequency band(40Hz)induces the strongest ASSR response,so 40 Hz sound stimulation is commonly used in neuroscience studies to evaluate the integrity of sensory pathways and the synchronization of neuronal activity.In previous studies,we have found that ASSR response intensity of Auditory Cortex(AC)is stronger than other cortical regions such as prefrontal cortex,suggesting that AC region may be the origin of ASSR signals observed in EEG.However,it is not clear which layer of the AC Cortex is ASSR signal’s origination.In addition,the sensory cortex,including the AC,receives neural projections from the thalamus.The thalamus is the largest ovoid gray matter mass in the diencephalon.It receives information projection from the ascending sensory pathways of the brain stem and then passes to the cerebral cortex after integrated processing.There are two pathways of projection between auditory thalamus and cortex.One is the specific projection pathway from medial geniculate body(MGB)to AC,which is responsible for transmitting specific auditory information such as frequency and intensity of sound.The other is a non-specific projection pathway from the dorsal nucleus of the medial thalamus(MD)to the posterior parietal cortex(PPC)and prefrontal cortex(PFC),which can deliver stimulus signals that can cause brain arousal and changes in attention state.These two pathways coexist in the auditory system,but it is not clear how ASSR signals are transmitted in these two pathways,and whether there is a crossover or circuit between the two pathways.Another concern is that the intensity of ASSR signal can be affected by different states of consciousness,and EEG can even be used to monitor ASSR signal in clinical practice as a reference indicator to judge the degree of anesthesia.However,the regulation mechanism of anesthesia on ASSR response is not clear,so it is necessary to explore the correlation between ASSR and conscious state and its internal mechanism from the two levels of local network between cortical sublayers and the network between thalamus and cortex.To solve the above problems,we first recorded the Local Field Potential(LFP)and Single Unit Activity(SUA)of each sub-layer in AC region of awaked mice using multi-channel silicon electrode.By analyzing their response to the sound stimulation of40 Hz Click-Trains,the specific origin position of ASSR in AC region and its transmission mechanism among different sub-layers were discussed.Secondly,we used chronic microelectrode implantation to synchronously record LFP signals in each brain region and analyze the functional connectivity of each brain region to construct a pattern of ASSR information transmission between thalamus and cortex.Finally,the effects of anesthesia on ASSR in mice were analyzed by the above two electrode recording methods.These research results are helpful to reveal the ASSR signal in the formation and transmission mechanism of AC,parsing the thalamus-cortex function of neural network to connect the working principle of the clear consciousness of ASSR regulatory mechanism,and can provide a reference for analysis of potential causes of abnormal ASSR,solid foundation for clinical effective reading ASSR eeg test result.Methods: 1.Surgery and electrode implantation: During head surgery under isoflurane anesthesia,the target brain region of AC region was marked according to the coordinates of mouse brain localization map,and a customized metal ring was embedded in the temporal side of the mouse skull and fixed with dental cement.After two weeks of recovery,the mice were fixed on the silent running plate with a special fixed rod and a metal ring buried in the skull in advance to ensure that the mice could run at will.At the same time,a multi-channel silicon electrode was inserted into the AC region of mice,and70 db sound stimulation was given to 40 Hz clicks(40Hz clicks)waves with a frequency of 0.5 seconds and a pulse width of 0.2 ms.LFP and SUA were recorded.2.Analyze the collected LFP of AC region: Local Field Potential(LFP)was obtained by 0.1-300 Hz filtering of original EEG data.Mean Trail Power(MTP)and Phase Lock Factor(PLF)were calculated by time-frequency analysis of LFP based on wavelet transform to investigate the reaction intensity at a specific frequency range of35-45 Hz.Current Source Density(CSD)analysis method was used to remove the influence of volume conduction effect on LFP results,and ASSR characteristics of different layers in AC region were analyzed.Event-Related Coherence(ERCOH)and Granger causality tests were used to analyze the correlation and causality of the layers to determine where ASSR originated in the AC region.3.Analyze the spike activity collected from AC region: The raw EEG data was filtered at 300-5000 Hz to obtain Single Unit Activity(SUA).The neurons were classified by the characteristics of spike waveform and the discharge rate was analyzed.Spike Field Coherence(SFC)and interneuronal discharge correlations were used to investigate the involvement of all layers of cell discharge in ASSR.4.Tubes were embedded in MGB,MD,AC,PFC and PPC brain regions of mice:Electrodes were embedded at the outlet of microtubules and fixed with dental cement.After recovery from surgery,the sober head was fixed and ASSR responses were induced by 40 Hz Click-Trains to explore the characteristics of ASSR responses in various brain regions,and the correlation between brain regions and the direction of information flow were explored by ERCOH and Granger causality tests.5.Comparison of LFP before and after anesthesia: LFP was recorded in each layer of the AC region or in five brain regions such as MGB for one hour in the awake state of mice,followed by intraperitoneal injection of Dexmedetomidine(80 μ g/kg),followed by four hours of LFP.ASSR,ERCOH and directivity of LFP were compared before and after anesthesia.Results: 1.We found that LFP of each layer in AC region showed stable ASSR under the sound stimulation of Click-Trains at 40 Hz,and ASSR of G layer was the strongest.The time-frequency analysis results also show that there is a strong correlation between ASSR signals from G layer and P layer.Granger causality test results show that information from layer G flows to layer S and layer I,and information from layer S then flows to layer P.The cell results also showed that the G layer cells responded most strongly to Cick-Trains,among which the interneurons were the most involved.2.It was found that ASSR response was strongest in AC region,followed by MGB,and weak in MD region.However,there was almost no ASSR response in PFC and PPC region,and only a weak response at the beginning of sound.From the correlation analysis,MGB has the strongest correlation with AC,MD has a strong correlation with AC region,and correlations of other brain regions are weak.3.LFP in each brain region of thalamus and cortex showed ASSR’s amplitude,MTP and PLF were the strongest in AC,followed by MGB,MD,and PPC and PFC region showed the weakest response.4.ERCOH results showed high ASSR Coherence between MGB,MD and AC,low ASSR Coherence between MGB and MD with PPC and PFC,and the lowest ASSR Coherence between AC,PPC and PFC.5.Anesthesia reduced the intensity of ASSR response in each layer of AC region and the correlation of ERCOH between layers.ASSR of G layer decreased the most,and the information-driving ability of G layer to other layers weakened,while the information-driving ability of I layer increased.6.ASSR response intensity and ERCOH correlation between MGB,MD and AC were significantly reduced under anesthesia.However,ASSR response intensity in PFC region increased,and its correlation with other brain regions also increased.Conclusions: 1.The ASSR response induced by Click-Trains sound stimulation in the AC region originates from the G layer,and is first transmitted to the S layer and I layer,and then from the S layer to the P layer.Inhibitory neurons in G layer that can generate synchronous discharge activity with 40 Hz sound stimulation may be the main participants in the formation of ASSR signal.2.The response of ASSR induced by 40 Hz Click-Trains is strong in MGB and AC region,but weak in MD,PPC and PFC region.MGB and AC had strong ASSR signal transmission,while MD and PPC and PFC had weak ASSR signal transmission.The ASSR signal transmission between cortical regions and to thalamus MGB and MD was also weak.Thus,the specific auditory pathway projected from MGB to AC is the main origin of cortical ASSR signals.3.Anesthesia significantly reduced the response of ASSR in all layers of AC region,with the greatest decrease in G layer.ASSR origination was transformed from G layer to I layer.Anesthesia decreased the response intensity of ASSR in MGB,MD and AC regions,but increased the response intensity of ASSR in PFC regions.The upward ASSR information transmission from thalamus to cortex was inhibited,while the downward information transmission from PFC to PPC,AC and thalamus was enhanced.