| Abstract: | 作用在電位依賴型鉀離子通道Kv7 (KCNQ)家族的鉀離子通道開啟劑具有穩定神經元細胞膜電位的作用，近年來Kv7鉀離子通道開啟劑是藥物開發的一個新的領域，藉由活化Kv7蛋白，可以有效的來治療涉及神經元的過度興奮相關疾病，如癲癇、心律不整和神經性疼痛等，目前已知Kv7鉀離子通道開啟劑有ML213與Retigabine。
本研究希望評估已知之鉀離子通道開啟劑ML213、Retigabine與本實驗室新篩檢出之鉀離子通道開啟劑Retigabine的乙苯胺醯衍生物#5540和#8692在於肌強直症離體模式的保護作用，以及在癲癇和心肌缺血再灌流活體模式中的保護效果。
本研究分為肌強直症、癲癇及心肌缺血再灌流傷害三個部分:
在肌強直症的研究利用小白鼠膈神經－橫膈膜肌模式，評估藥物對神經傳遞的影響，以確認其對離子通道的調控作用。
研究使用遠交系(ICR)小鼠，進行膈神經-橫膈膜肌之製備，以電刺激橫膈膜肌，紀錄其張力的變化。實驗結果發現，不論是藉由間接刺激還是直接刺激，在低頻率的電刺激下，9-AC (9-Anthracenecarboxylic acid) (30 μM)誘發之肌強直現象都會受Retigabine、#8692拮抗；在高頻的刺激下，Retigabine可以拮抗給予9-AC後誘發的肌強直現象所造成的TCM (tetanic contraction maintenance) Index下降，#8692無法拮抗9-AC (30 μM) 誘發之肌強直現象。
在癲癇的研究探討 #5540與 #8692是否對於癲癇有較Retigabine和ML213更好的保護效果，實驗利用最大電休克發作實驗 (maximal electroshock seizure, MES) 去模擬癲癇全身性的強直-陣攣性發作（tonic-clonic seizure）評估#5540與#8692是否可以有效地降低癲癇的發作頻率與強度，對於癲癇相較於Retigabine和ML213是否有更好的效果。
研究使用遠交系(ICR)小鼠，進行電刺激誘發癲癇發生，於誘發癲癇前15分鐘經由腹腔注射，投予藥物，統計發生癲癇時的tonic duration和clonic duration，評估 #5540和 #8692的癲癇保護作用。實驗結果發現Retigabine具有減少tonic duration的效果。
在心肌缺血再灌流方面，利用大鼠心肌缺血再灌流傷害動物模式，評估是否可以藉由Kv7鉀離子通道開啟劑活化鉀離子通道，減少心肌缺血再灌流傷害所引起的死亡率，心律不整 (Arrhythmia)，並減少心肌梗塞體積 (Infarct size)，進而達到心臟保護作用。
研究使用Sprague-Dawley (SD) 大鼠，進行六十分鐘左冠狀動脈前降支 (the left anterior descending, LAD) 結紮及一百八十分鐘的再灌流，造成心肌缺血再灌流傷害，在LAD結紮前十五分鐘經靜脈投予ML213、Retigabine或是 #8692，計算動物心肌缺血再灌流傷害誘發之心肌梗塞體積、心律不整及死亡率，評估Kv7蛋白鉀離子通道開啟劑ML213、Retigabine或是 #8692是否具有心臟保護作用可以減少心肌缺血再灌流傷害。實驗初步結果發現相較於投與溶劑的對照組，投予Retigabine或ML213可以減少心律不整發生時間的長短，並顯著減少心肌缺血再灌流誘發的心肌梗塞大小。
本研究結果證明Retigabine、#8692及ML213可能具有治療肌強直疾病的潛力。Retigabine可以減緩癲癇強直性發作。而Retigabine及ML213可能是具有潛力之心臟保護劑，可以保護心臟對抗心肌缺血再灌流傷害。
The Kv7 (KCNQ) potassium channel opener acts on the voltage-gated potassium channel to stabilize membrane potential. Recently, Kv7 potassium channel opener is a new target of drug development. In clinical, Kv7 potassium channel opener is used on the treatment of epilepsy, arrhythmia and neuropathic pain. ML213 and Retigabine are known Kv7 potassium channel opener at present.
In this study, we investigated the protective effect of Kv7 potassium channel opener, ML213, Retigabine and the derivatives of Retigabine #8692 and #5540, on in vitro phrenic nerve-diaphragm model of myotonia and in vivo models of epilepsy and myocardial ischemia/reperfusion injury.
There are three part in our study: 1. myotonia, 2. epilepsy, 3. myocardial ischemia/reperfusion injury.
Myotonia:
We investigated the effect of ML213, Retigabine, #8692 and #5540 on neuron transmission and ion channels regulation by phrenic nerve-diaphragm modelin ICR mice. In our study, Retigabine and #8692 inhibited the myotonia induced by 9-AC (9-Anthracenecarboxylic acid) on low frequency directly and indirectly electrical stimulation. Retigabine also improved the myotonia induced by 9-AC on high frequency directly and indirectly electrical stimulation, but not #8692.
Epilepsy:
In this study, we induced tonic-clonic seizure by maximal electroshock seizure (MES) model to compare the protective effect of #5542 and #8692 with Retigabine and ML213 in ICR mice. The ML213, Retigabine, #8692 and #5540 were administered by intraperitoneal injection15 min before maximal electroshock. We found that there are no significantly differences between Vehicle and ML213, Retigabine, #8692 and #5540 groups on clonic duration. However, administration of Retigabine had protective effect on tonic duration.
Myocardial ischemia/reperfusion injury:
In this study, we examined the cardioprotective effect of ML213, Retigabine, and #8692 on rats subjected to myocardial ischemia/reperfusion injury. We compared mortality, arrhythmias and infarct size induced by myocardial ischemia/reperfusion injury with or without ML213, Retigabine, and #8692 administration.
Myocardial ischemia/reperfusion injury was induced by one hour occlusion of the left anterior descending (LAD) coronary artery and followed by three hours reperfusion in Sprague-Dawley (SD) rats. Vehicle or ML213, Retigabine, and #8692 were administered intravenously 15 minutes before LAD occlusion. We found that the administration of Retigabine and ML213 decreased arrhythmias induced by myocardial ischemia/reperfusion injury, and significantly decreased infarct size compared to the Vehicle-treated group.
Our results suggested that Retigabine and #8692 has the potential to treat myotonia. Retigabine prevent tonic happen. In addition, Retigabine and ML213 have the potential to protect heart against myocardial ischemia/reperfusion injury. |
