古典恐懼制約（classical fear conditioning）中，有兩個刺激參與其中，分別是制約刺激（conditional stimulus，如：一段聲音）與非制約刺激（unconditional stimulus，如：微電擊）。其中常見的兩種安排為延宕制約（delay conditioning）與痕跡制約（trace conditioning）。延宕制約的非制約刺激接在制約刺激後出現，兩刺激重疊且同時結束；痕跡制約中兩刺激間有一個空白的時間間隔，稱作痕跡間隔。在此研究中，我們討論了制約刺激與非制約刺激之間的連結較弱是由於時間間隔變長所導致，或是因為動物將制約刺激當成安全訊號（safety signal），另外也分析動物是否可能準確預測電擊該出現的時間點。動物依據組別分別進行延宕制約或不同痕跡間隔（30秒、60秒或120秒）之痕跡制約，試驗間隔（inter-trial interval）為240秒。進行恐懼制約後，動物進入另一個情境測試對聲音的恐懼程度（實驗1）或在能夠按壓壓桿得到食物以維持基本活動量的情況下測試對聲音的恐懼程度（實驗2）。行為結果顯示，隨著痕跡間隔越長，動物對聲音的恐懼表現越低，且痕跡組別（30秒與60秒）於制約刺激結束後表現出更高的恐懼（實驗1）。但在能夠按壓壓桿以維持基本活動量下，痕跡組別對於聲音接表現出低的凍結（freezing）反應與低的壓桿抑制比（suppression ratio），且一次試驗（trial）中最高的恐懼表現仍為聲音出現時（實驗2）。神經生理方面，我們利用c-fos蛋白測量於實驗2中，動物提取恐懼記憶時腦區的神經活化程度，並發現基底外側杏仁核（basolateral complex of amygdala）與內側前額葉皮質（medial prefrontal cortex）的活化程度與對聲音的凍結程度呈現正相關的結果。基底外側杏仁核為儲存制約刺激與非制約刺激之間的連結，我們的結果支持了動物於痕跡制約中，對於制約刺激表現出較低的恐懼反應的原因較可能是因為制約刺激與非制約刺激的連結較弱所導致。

In classical fear conditioning, the conditional stimulus (CS, e.g. a tone) and the unconditional stimulus (US, e.g. a foot shock) are presented. Different from delay conditioning, in which the US is presented after the CS onset and the two stimuli overlap and end together, in trace conditioning, the CS and the US are separated by a stimulus-free temporal gap called “trace interval”. In this study, we aimed to examine whether the association between the CS and the US become weaker simply because the trace intervals become longer, or alternatively, the rats treat the CS as safety signals and fear to tones surge after CS offset. The animals underwent delay conditioning or trace conditioning with different trace intervals (30s, 60s, or 120s) and with inter-trial intervals (ITI) of 240s. After fear conditioning, their fear to tones were assessed in a shifted context (Experiment 1) or on top of the operant behavior of lever-pressing for food reward that maintained the baseline activities (Experiment 2). Behaviorally, our data showed that the fear to tones became lower as the trace interval became longer, and TRACE groups (30s and 60s) showed slightly increased fear after CS offset (Experiment 1). However, when operant behavior of lever-pressing was implemented to maintain baseline activities, the TRACE groups showed low freezing and suppression ratio to tones, and CS presentation was the time the rats showed highest fear in all of the experimental groups (Experiment 2). Neurobiologically, c-Fos activation levels were assessed in fear-related brain regions during fear retrieval in Experiment 2, and we found positive correlations between the freezing levels to tones and activation levels in basolateral complex of the amygdala and medial prefrontal cortex. Since the basolateral complex is where the CS-US association occurred and stored, our results supported the idea that the low fear response to the CS in animals trained with trace procedures was more likely a result of the weaker CS-US association.

中文摘要 i
Abstract ii
Chapter 1 Introduction 1
1.1 Pavlovian fear conditioning 1
1.2 Neurobiology of trace fear conditioning 1
1.3 Trace fear conditioning with long trace intervals 3
1.4 Specific aims 4
Chapter 2 Materials and methods 6
2.1 Subjects 6
2.2 Behavioral apparatus 6
2.3 Behavioral procedures 7
2.4 Immunohistochemistry 8
2.5 Data analysis and Statistics 9
2.5.1 Behavioral results 9
2.5.2 c-Fos quantification 11
Chapter 3 Results 12
3.1 Experiment 1: Fear conditioning under different trace intervals 12
3.1.1 Behavioral results 12
3.2 Experiment 2: Fear retrieval on top of lever-pressing behavior 14
3.2.1 Behavioral results 15
3.2.2 c-Fos quantifications 18
Chapter 4 Discussion 21
Table 27
Figures 28
Reference 49

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