Changes In Local Field Potential Due To Targeted Ischemia In The Hippocampus And Amygdala And Its Effect On Associated Fear Memory

Stroke is a disease which has high morbidity, mortality, and can cause a high instance of disabilities. These derived factors from stroke have become a serious threat to both human health and society in general. Stroke usually refers to:neuronal cell death, functional deficits of neurons, and nervous system dysfunction-all are main symptoms of a class of blood circulation disorders of the brain. Stroke can be classified as either ischemic stroke or hemorrhagic stroke, where ischemic stroke accounts for 60-70% of all stroke patients. Basic science and clinical research have developed a variety of treatments dedicated to facilitating functional recovery in damaged brain areas after stroke. These include:thrombolytic therapy, rehabilitation therapy and transcranial magnetic stimulation. In clinical practice, thrombolytic therapy provides the best results, but only within a 4.5 hour window immediately after the stroke. After this timeframe, no other treatments are able to produce such a desirable effect. The lack of proper treatment may be attributed to the lack of specific treatment targets and the complexity as well as redundancy of neural networks. However, emerging optogenetic techniques are able to accurately regulate specific aspects of neuronal activity in the brain, thus allowing for precise control of neural circuits. We hope to gain insight on the mechanisms involved in the recovery of damaged areas in the post-ischemic brain. By studying this subject from a fundamental prospective, we hope to provide a new theoretical basis for the potential in using optogenetics as a clinical treatment for stroke.The main symptoms of stroke in addition to motor dysfunction also include defects in learning and memory, language, emotion, and other higher brain functions, thereby indicating that strokes may not in fact be limited to just the somatosensory and motor cortices. Rather, strokes may also occur in the hippocampus and amygdala, among other parts of the limbic system. Today, studies of ischemia occurring in the motor cortex ischemia are common, while research on ischemic stroke of the deep nuclei and related areas are remarkably few. By using multi-channel technology recording in vivo, we can study the effect of ischemia on different brain regions related to fear learning such as the hippocampus and amygdala. These techniques, combined with optogenetics to regulate upstream excitability, allow further exploration of the effect of ischemic stroke on fear memory relevant areas (such as the lateral amygdala), as well as functional recovery from stroke.The results are as follows:(1) Endothelin (ET-1) and (MCAO) are the main methods used to model ischemia in rats. However, MCAO-induced ischemia occurs mostly in the striatum and the cerebral cortex, and therefore is not applicable to deep brain research. Furthermore, ET-1-induced ischemia lasts only three days before recovery, and is therefore unable to effectively simulate the effect of long-term stroke damage on learning and memory. We advanced the traditional photothrombosis parameters to a 532nm laser with an intensity of 30mW/mm2 and a local irradiation of 1 hour to achieve a targeted ischemia in the dorsal hippocampus and the lateral amygdala. From this method, the recovery occurs after longer period of time (seven days) and therefore can be applied to long-term functional and behavioral tests on ischemic animals. Furthermore, no additional abnormal behaviours manifest as a result of this method. (2) The dorsal hippocampus and lateral amygdala affects contextual fear memory and tone-related fear memory respectively, and inducing ischemia in these areas will affect brain function correspondingly. (3) On this basis, we aim to further explore the effect of ischemia on the dorsal hippocampus and lateral amygdala on local field potential (LFP) and changes within sub-bands of the power spectrum. We found that during the tone-related fear memory training under physiological conditions, total power in the lateral amygdala increased significantly when stimulated by the conditioned stimulus (CS, tone), with the increase mainly related to the theta and gamma sub-bands. However, during ischemic conditions of the lateral amygdala, power density recorded in the initial CS segment and Inter-Trial Interval (ITI) segment increased. When the power density began to show a gradual downward trend, it was due mainly to the theta and gamma sub-bands. When testing tone-related fear memory of the amygdala under physiological conditions, if the CS (tone) was not paired with the unconditioned stimulus (US, foot shock), then the total energy only initially increased as well as the energy in the theta and gamma band. As the CS (tone) and US (shock) were not paired, stimulating the CS continuously weakened the tone-related fear memory. When testing tone-related fear memory of the amygdala under ischemic conditions, The power of each sub-band did not change significantly,including the theta and gamma sub-bands.These results suggest that ischemia of the dorsal hippocampus and lateral amygdala significantly inhibited LFP power and delta, theta, beta, and gamma sub-band power. The reason may be that under an ischemic state, synaptic contact and fiber projections between brain regions become interrupted. (4) After the optogenetic model was successfully constructed, through behavioral tests, we assumed that optogenetic regulation of the ventral part of the secondary auditory cortex (AuV) neuronal activity upstream to ischemic brain regions can promote functional recovery of the downstream LA area. By regulating activity of specified brain areas on a circuit level, optogenetic techniques may be a possibility in establishing new clinical therapies for stroke.To summarize, the study successfully establishes a method to induce deep brain ischemia in targeted areas lasting seven days, which is then suitable for long-term behavioral tests. The induced ischemia in specific brain regions correspond to impaired higher brain functions and its main symptoms are shown to accurately simulate ischemic stroke. We will follow up by using a combination of optogenetics and multi-channel recording techniques in the upstream region of the lateral amygdala which is the ventral part of the secondary auditory cortex (AuV), by giving a light stimulus simultaneously to both upstream and downstream regions and record its oscillations. Furthermore, we look to study the following two aspects of the AuV and LA regions:EEG between the two areas, and correlation between theta and gamma oscillations in single neurons or within a population; thereby providing experimental evidence for elucidating circuit-level mechanisms of the functional recovery of fear memory and learning.