帕金森氏症的成因與神經化學傳遞物質多巴胺缺乏有著密切的關係，而多巴胺在精神相關的疾病與運動調節的功能上也扮演重要的角色，因此建立一個能急性期或長期監測動物腦中多巴胺濃度的平台，可以幫助我們針對一些新穎的帕金森症治療方法評估其療效。在本研究中，應用電化學的方法進行量測與評估，發展新的感測電極達到高靈敏度與選擇性以克服腦中複雜的干擾物質，以及進行急性期動物實驗之驗證。我們利用金奈米粒子(Au-NPs)的吸附來增加電極的表面積，再經由11-mercaptoundecanoic acid (MUA)和3-mercaptopropionic acid (MPA)分子改質以加強靈敏性與選擇性。經由水溶液實驗得知，新式感測電極對多巴胺的氧化峰電位大約在0.28 V、且檢測極限低於100 nM，更能有效排除電化學活性干擾物的影響(例如抗壞血酸，ascorbic acid)。在動物體內實驗的部分則在麻醉下進行，感測電極被植入大鼠的紋狀體(striatum)進行記錄，電刺激電極則植入中前腦束 (medial forebrain bundle，MFB)，成功地利用電刺激與藥物的方式誘發多巴胺釋放。電刺激的參數為 100 Hz的刺激頻率、300 μA的電流強度持續電刺激1秒，每1秒的刺激則間隔30秒休息。在電刺激持續的期間可以明顯觀察到電流訊號上升，當電刺激結束後約需5 ~ 10秒回到基準電流值，且反應的電流大小與電刺激的頻率、電流強度有著正向的關係。此外，我們也利用了多巴胺的前驅物左旋多巴胺(L-DOPA)進行體內測試，也同樣引起訊號的上升但長達3個小時。總結來說，利用具有高靈敏性與選擇性的Au-NPs-MPA修飾電極進行量測，經由電刺激或是藥物誘發的處理都已經成功量測到訊號，適合將此平台應用於新型的帕金森氏症治療的動物實驗模型，例如重複性經顱磁刺激(repetitive transcranial magnetic stimulation , rTMS)的評估。

Parkinson’s disease (PD) is known related to dopamine (DA) depletion which is an important neurotransmitter linking to psychosis and locomotion modulation in the biological system. Knowing the DA level of animal’s brain could provide a direct evidence for evaluating novel treatments for PD, especially in the acute or long-term studies. The aims of this study are to develop DA sensing microelectrodes and their in-vitro and acute in-vivo validation. Among various DA sensing techniques, there are many advantages for applying electrochemical techniques to DA detection, including good temporal resolution and easy to minimize but they also have some critical hinders mainly the sensitivity to low DA level and specificity to various other electrochemical substances in the brain.
In this study, the designed sensing microelectrode was coated gold nanoparticles (Au-NPs) by electrodeposition and self-assembled the 11-mercaptoundecanoic acid (MUA) and 3-mercaptopropionic acid (MPA) to enhance its sensitivity and selectivity. For in vitro verification test, the Au-NPs-MPA modified microelectrodes exhibited an oxidation potential of DA at 0.28 V with a low detection limit below 100 nM. Moreover, the modified microelectrodes have effectively eliminated electronegative species, such as ascorbic acid, AA, the major interference to DA in the brain. For in-vivo validation, the DA secretion can be evoked by pharmacological and electrical stimulation. The recording microelectrodes were implanted in striatum of the rat brain under anesthesia condition and the stimulation electrodes were applied with bipolar waveforms to stimulate in medial forebrain bundle (MFB). Stimulation parameters were set at 100 Hz frequency, 300 μA intensity, 1 sec stimulating period and 30 sec resting interval. Following the stimulation, the recorded signal rose abruptly and fell gradually, lasting about 5~10 sec before returned to the baseline. Moreover, responsive currents exhibited a positive trend correlated to the higher stimulating intensity and frequency. In addition, the in-vivo sensing of DA was also validated by administration of L-DOPA which also increased the DA release in anesthesia rat but required 3 hours to response.
In conclusion, the platform for monitoring the DA level in anesthesia rat using Au-NPs-MPA modified microelectrodes with high sensitivity and selectivity has been successfully verified in electrical or L-DOPA intervention. The established platform and animal model are quite suitable for the implantation electrochemical sensing microelectrode for acute or long-term morning of DA level to evaluate the effect of various novel therapies, in our case repetitive transcranial magnetic stimulation (rTMS) for PD rat.

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