臺灣博碩士論文加值系統

拉莫三嗪，是一種廣為人知的電壓門控之鈉離子通道阻斷劑，它可抑制神經元的興奮反應。然而，最近的研究發現，拉莫三嗪亦可促進過極化啟動之環核苷酸門控離子通道的開啟，進而活化分泌伽碼胺基丁酸的聯絡神經元。在海馬迴之齒狀迴區域中，有一種細胞體抑制型聯絡神經元，主要為高頻動作電位的籃狀細胞。它能強而有力地抑制顆粒細胞的活性。值得注意的是，籃狀細胞同時具有相當多的電壓門控之鈉離子通道與過極化啟動之環核苷酸門控離子通道，而這兩種通道皆會促進籃狀細胞產生持續的動作電位。我在大鼠的新鮮腦切片下，使用膜箝制全細胞紀錄，想要研究拉莫三嗪對於籃狀細胞的活性及其伽碼胺基丁酸的輸出的淨效果究竟為何 ? 我們發現，施予拉莫三嗪可明顯抑制顆粒細胞上記錄到的總合突觸後抑制型電流大小。然而，透過同時配對記錄藍狀細胞與顆粒細胞發現: 拉莫三嗪並不影響顆粒細胞上記錄到的單一突觸後抑制型電流的大小及其多重脈衝比率，這意味著: 拉莫三嗪不會影響籃狀細胞軸突末梢伽碼胺基丁酸釋放的機率。所以，拉莫三嗪抑制突觸後抑制型電流的原因，乃是降低突觸前細胞的興奮反應所造成的。與上述結果一致地，拉莫三嗪亦會顯著降低顆粒細胞上自發性突觸後抑制型電流之頻率但不影響微細突觸後抑制型電流的頻率。當再回到海馬迴CA1區域做驗證時，我們發現拉莫三嗪仍會降低在初始層樹突抑制型聯絡神經元的興奮反應，同時其下游的椎狀神經元所記錄到的自發性突觸後抑制型電流也受到明顯的抑制，然而過極化啟動之離子通道電流卻不受影響。總結來說，拉莫三嗪不會增強過極化啟動之離子通道電流也不會促進伽碼胺基丁酸抑制訊息的釋出。拉莫三嗪抗癲癇效果，可能仍來自於鈉離子通道的阻斷作用與整體神經網路抑制的結果。

Lamotrigine (LTG) is generally considered as a voltage-gated sodium (Nav) channel blocker, which can decrease neuronal excitability. However, recent studies suggest that LTG can also serve as a hyperpolarization-activated cyclic nucleotide-gated (HCN) channel enhancer and can increase the excitability of GABAergic interneurons (INs). Perisomatic inhibitory INs, predominantly fast-spiking basket cells (BCs), powerfully inhibit granule cells (GCs) in the hippocampal dentate gyrus. Notably, BCs express abundant Nav channels and HCN channels, both of which are able to support sustained action potential generation. Using patch-clamp whole-cell recording in acute rat hippocampal slices, we investigated the net LTG effect on BC excitability and its GABAergic output. We showed that bath application of LTG significantly decreased the amplitude of evoked compound inhibitory postsynaptic currents (IPSCs) in GCs. In contrast, simultaneous paired recordings from BCs to GCs showed that LTG had no effect on both the amplitude and the multiple-pulse ratio of the unitary IPSCs, suggesting that LTG did not affect GABA release probability at BC axonal terminal, though it suppressed cell excitability. In line with this, LTG decreased spontaneous IPSC (sIPSC) frequency, but not miniature IPSC frequency. When re-examining the LTG effect on GABAergic transmission in the cornus ammonis region 1 (CA1) area, we found that LTG markedly inhibited both the excitability of dendrite-targeting INs in the stratum oriens and the concurrent sIPSCs recorded on their targeting pyramidal cells (PCs) without significant hyperpolarization-activated current (Ih) enhancement. In summary, LTG has no effect on augmenting Ih in GABAergic INs and does not promote GABAergic inhibitory output. The antiepileptic effect of LTG is likely through Nav channel inhibition and the suppression of global neuronal network activity.

目錄
誌謝 ................................................................................................................... i
目錄 ....................................................................................................................... iii
ABSTRACT ............................................................................................................... v
中文摘要 ................................................................................................................. vii
ABBREVIATIONS .......................................................................................... viii
1. INTRODUCTION ....................................................................................... 1
1.1. Lamotrigine, a sodium channel blocker and clinical implications …..... 1
1.2. HCN channel may be a novel LTG target ………………………………. 3
1.2.1. HCN or Ih channel …………………………………………………... 3
1.2.2. Physiological significance of Ih channel …………………………... 4
1.2.3. Pathological significance of HCN channel………………………... 5
1.2.4. LTG may act on HCN channel …………………………………….. 6
1.3. Hippocampal GABAergic interneurons and HCN channel expression
1.3.1. Hippocampus ……………………………………………………...... 6
1.3.2. Diversity of GABAergic interneurons ……………………………... 7
1.3.3. GABAergic transmission………………………………………........ 8
1.3.4. HCN expression on INs ……………………………………………. 9
1.4. The rationale and aims of this study.................................................... 11
1.5. Rapid understanding of the IPSC measurement................................. 12
2. MATERIALS AND METHODS................................................................... 14
2.1. Acute Brain Slice Preparation............................................................. 14
2.2. Electrophysiology................................................................................ 14
2.3. Morphological Identification................................................................. 16
2.4. Solutions and Drugs ......... .................................................................. 16
2.5. Data Analysis and Statistics................................................................ 17
3. RESULTS
3.1. LTG reduced Compound IPSCs ........................................................... 18
3.2. LTG had no effects on Unitary IPSCs………………………………….... 19
3.3. LTG decreased sIPSCs, but not mIPSCs…...………………………...... 20
3.4. LTG Inhibited the axonal and cellular excitability of BCs in the DG…... 20
3.5. LTG suppressed sIPSCs rather than mIPSCs in CA1 PCs………….... 22
3.6. LTG inhibited the excitability of CA1 O-LM INs……………………….... 22
3.7. LTG had no effect on Ih in CA1 O-LM INs, CA1 PCs and DG
fast-spiking BCs …………………………………………………...............
23
3.8. LTG preferentially inhibited excitatory inputs than inhibitory inputs……............................................................................................
24



4. DISCUSSION
4.1 Summary of our results …………………………………………………… 26
4.2 LTG shows no effect on Ih in our evidence……………………………... 27
4.3 LTG on GABAergic neurotransmissions…………………………………. 28
4.4 Miscellaneous Issues……………………………………………………… 30
4.5 Conclusion and perspective………………………………………………. 32

REFERENCES.............................................................................................. 33
FIGURES
Figure 1. ......................................................................................................... 40
Figure 2. ......................................................................................................... 42
Figure 3. ......................................................................................................... 43
Figure 4. ......................................................................................................... 45
Figure 5. ......................................................................................................... 47
Figure 6. ......................................................................................................... 49
Figure 7. ......................................................................................................... 51
Figure 8. ......................................................................................................... 52
Figure 9. ......................................................................................................... 53
Figure 10. ....................................................................................................... 54
Figure 11. ....................................................................................................... 55
Figure 12. ....................................................................................................... 56
Figure 13. ....................................................................................................... 57
Figure 14. ....................................................................................................... 58
Figure 15......................................................................................................... 59

PUBLICATIONS.............................................................................................. 60

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