- 學位論文
視丘下核放電行為在巴金森症之病態生理角色
The Pathophysiological Role of Subthalamic Discharges in Parkinson’s Disease
摘要
巴金森症是中老年人主要的神經退化性疾病，常引起病患嚴重的運動症狀及日常生活障礙。巴金森症的病態生理尚未完全明瞭，但藉助巴金森症的動物模型的發展，目前已進行有相當廣泛的研究。巴金森症運動症狀的可能病態生理機制，可以用大腦基底核迴路模型在多巴胺缺乏狀態下的變化來的解釋，而視丘下核在此一病態狀況下的迴路中扮演病態生理形成的重要角色。在巴金森症病人或巴金森症模型動物給予視丘下核高頻率電流刺激或甚至破壞此一結構，可以有效地改善實驗動物及病人的巴金森運動症狀，亦說明了視丘下核在巴金森病態生理的重要性。在實驗動物研究中，視丘下核神經元叢集式放電的明顯增加，是目前公認巴金森症的病態電氣生理特徵。但是，視丘下核叢集式放電及巴金森運動症狀之間的因果關係目前尚未建立，而造成此一叢集式放電的離子通道機制目前亦尚未明瞭。本研究藉由活體動物的胞外電生理記錄及動物運動行為測試等實驗，針對實驗動物視丘下核直接注射不同的離子通道阻斷劑或各種方式的電流刺激，並記錄給藥/刺激前、中及後的變化，詳細探討視丘下核叢集式放電行為的形成機制，及視丘下核叢集式放電對巴金森症大鼠運動障礙的影響。我們首先發現可以有效抑制T型鈣離子通道的Ni2+, Mibefradil, NNC55-396 及 Efonidipine能夠有效地降低活體動物視丘下核的叢集式放電數量，但主要抑制L型或其他型鈣離子通道的Cd2+或 Nifedipine則無法降低叢集式放電的現象。直接給予視丘下核上述的T型鈣離子通道抑制劑，可以有效地改善巴金森症大鼠的運動障礙；而Cd2+及 Nifedipine則對動物的行為沒有效果。至此，我們首先作出視丘下核叢集式放電與巴金森症大鼠運動礙症狀有直接關聯的結論，並且發現到視丘下核的神經元細胞膜的性質，尤其是T型鈣離子通道，在視丘下核叢集式放電的生成，以及巴金森運動症狀的出現扮演關鍵的角色。這些發現，更進一步促使我們去探討深腦刺激治療巴金森症的原理，是否與電流改變Ｔ型鈣離子通道的作用有關。接著我們將各種不同形態的電流刺激直接注入視丘下核，包括不同頻率，各種波寬或甚至相反極性的持續性電流，來研究電流刺激對正常及巴金森症大鼠的電生理和行為所產生的改變。我們發現給予視丘下核負極性的持續性電流可以有效地改善巴金森大鼠的運動障礙症狀，而進一步的電生理記錄發現，此電流亦能有效降低的視丘下核神經元叢集式放電的出現，其效果類似Ｔ型鈣離子通道抑制劑。相反地，給予視丘下核正極性的持續性電流可以引起正常大鼠出現類似巴金森運動障礙的症狀。更令我們感到興趣的是，注射相反極性的持續性電流，在電生理記錄時可以發現對視丘下核叢集式放電產生完全相反的作用。依照前面所述的原理，我們認為Ｔ型鈣離子通道的可利用率與巴金森症運動症狀的產生，有著極為密切的關係，我們因此考慮利用先前認為無效的低頻率深腦刺激（頻率<85Hz）來驗證。我們給予低頻率刺激時，將電刺激的波寬大幅延長，原先無效的低頻率電刺激在巴金森實驗動物身上也可以轉變為有效。我們並將此發現進一步應用在臨床上接受深腦刺激治療的病患上。我們蒐集原本裝置深腦刺激治療但卻沒有理想效果的三位病患，將其治療的波寬由傳統的60微秒增加至240微秒，並將刺激的頻率降低至先前認為無效的60Hz以增加病患的耐受度，結果三位病患皆獲得良好的巴金森運動症狀改善的效果。我們總結對於視丘下核的叢集式放電，與造成此一放電形態的Ｔ型鈣離子通道進行調控，可以為未來的巴金森症提供發展全新且更有效治療方式的契機。視丘下核叢集式放電的增加，與動物的巴金森症狀有直接的關聯，此一作用甚至與多巴胺是否正常存在無關。有效的深腦刺激治療巴金森症的機制，很可能與視丘下核神經元是否能夠適當地去極化、調控其離子通道特性、或改變其相關的放電形態有關。本計劃的發現，證明了視丘下核叢集式放電行為與巴金森運動症狀的高度關聯，也為視丘下核深腦刺激治療巴金森症的機制，提出了清楚的說明。
並列摘要
Parkinson’s disease (PD) is a major neurodegenerative disease in the middle-aged and elderly population, causing severe motor symptoms and marked disability in these patients. The pathophysiology of PD is not yet clearly understood and has been studied extensively by animal models of PD. The cortico-basal ganglia loop model in a dopamine deficiency state has been used to explain the possible pathophysiological mechanism underlying these motor symptoms. According to this model, the subthalamic nucleus (STN) exerts great influences on the basal ganglia output nuclei and plays an important role during the PD pathophysiological state. Giving high frequency electrical stimulation to STN or even lesioning this structure, can effectively ameliorate contralateral parkinsonian motor symptoms in both experimental animals and human PD patients, and these evidences further illustrate the importance of STN in PD pathophysiology. In experimental animal studies, an increase of neuronal burst activities in the STN is a well-documented electrophysiological feature in PD. However, the causal relation between subthalamic burst discharges and PD symptoms remains to be established. The ionic basis underlying the pathological bursts is also obscure. In this project, we studied the in vivo single-units extracellular electrophysiology and open field behavior test, before, during and after direct administration of various channel blockers and different protocols of electrical current stimulation into STN, in normal rats and in the 6-hydroxydopamine (6-OHDA)-induced parkinsonian rat model. We first showed that Ni2+, mibefradil, NNC55-0396, and efonidipine which can evidently inhibit T-type Ca2+ currents, but not Cd2+ or nifedipine that preferentially inhibits L-type or the other Ca2+ currents, effectively diminish STN burst activities in single-units extracellular recordings in vivo. More interestingly, topical administration of T channel inhibitors also dramatically remedies the locomotor deficits in 6-OHDA-lesioned parkinsonian rats. Cd2+ and nifedipine show no such behavioral effects. We first concluded that STN burst discharges is directly correlated with locomotor deficit in 6-OHDA-lesioned parkinsonian rats. The knowledge that intrinsic membrane properties, especially T-type Ca2+ channels, play a key role in the genesis of burst discharges in STN and parkinsonian locomotor symptoms, promoted us to further explore whether DBS exerts its clinical benefits on PD with changes in T-currents or other conductances. We applied different stimulation protocols, including different frequencies, various pulse widths and even constant currents of opposite polarity, to STN in vivo and documented the electrophysiological and behavioral effects of the stimulation in normal and parkinsonian rodents. We found that delivery of negative constant current into STN dramatically ameliorated locomotor deficits in parkinsonian rats. It also decreased burst discharges effectively in STN neurons, similar to T channel inhibitors. In contrast, delivery of positive constant currents to STN induced PD-like locomotor deficits in normal rats. Interestingly, injection of constant currents of opposite polarity altered STN burst discharges in exactly opposite ways. According to the rationale that the availability of T-type Ca2+ currents is critically involved in the genesis of parkinsonian locomotor deficits, we showed that low-frequency (frequency <85 Hz) deep brain stimulation (DBS) which has been considered ineffective in PD can be readily turned into an effective treatment if only the depolarizing pulse is lengthened. The effect of correlatively adjusted DBS protocols was also explored clinically. The therapeutic effect of DBS was greatly improved in three PD patients simply by increasing the pulse width from 60 to 240 μs, even at a lower stimulation frequency of 60 Hz which was considered in non-effective stimulation range. We concluded that modulation of subthalamic T-type Ca2+ currents and consequently burst discharges may provide novel and promising strategies for the treatment of Parkinson’s disease. The increased tendency of STN burst discharges may by itself serve as a direct cause of parkinsonian locomotor deficits, even in the absence of deranged dopaminergic innervation. Effective DBS therapy in PD very likely relies on adequate depolarization, and consequent modification of the relevant ionic currents and discharge patterns, of STN neurons. The findings of this study well illustrate the role of subthalamic discharges in the pathophysiology of PD motor symptoms and further demonstrate the underlying mechanism of STN DBS in Parkinson’s disease.