KMUP-1對於大鼠蜘蛛膜下腔出血模式腦血管平滑肌細胞鉀離子通道活性之影響

KMUP-1 Prevents the Cerebrovascular K+-Channel Dysfunction After Experimental Subarachnoid Hemorrhage

蔡易霖(Yi-Lin Tsai)

指導教授 ： 吳炳男

高雄醫學大學/醫學院/藥理學科研究所/碩士(2006年)

摘要

蜘蛛膜下腔出血所導致的腦血管痙攣為一種具擴散性且長時間持續性的血管收縮的現象。直至目前研究已發現，大傳導性鈣離子活化型鉀離子通道在血管張力調節方面扮演了一負回饋的角色，因此可預防血管痙攣的產生。KMUP-1為一個化學合成之嘌呤類衍生物，先前研究發現在大鼠胸主動脈血管平滑肌及天竺鼠氣管平滑肌， KMUP-1皆具有鉀離子通道開啟作用，進而使平滑肌產生擴張現象。本研究主要探討蜘蛛膜下腔出血所誘導之血管痙攣的現象是否因血管平滑肌上大傳導性鈣離子活化型鉀離子通道功能不正常所造成，並且探討KMUP-1對於蜘蛛膜下腔出血模式大鼠的保護作用。

本實驗中將大鼠分為四組，控制組、蜘蛛膜下腔出血組、KMUP-1組和pinacidil組。將大鼠自體動脈血注入腦池中，以建立大鼠蜘蛛膜下腔出血模式。當出血後1小時，利用腹腔注射的方式給予藥物KMUP-1或pinacidil且劑量皆為 1 mg/kg。本實驗中，所有動物將於24小時後犧牲。

大鼠基底動脈血管平滑肌細胞則是利用酵素新鮮分離而得。利用全細胞膜片箝制技術紀錄電流發現，於SAH組所測得之iberiotoxin敏感型大傳導性鈣離子活化型鉀離子電流與正常組比較起來有一減弱趨勢，而KMUP-1及Pinacidil組所測之電流則有一恢復趨勢。利用inside-out patches技術來紀錄發現，CTL組、SAH組、KMUP-1及Pinacidil組其離子通道傳導性及電位敏感性並無顯著差異，然而其大傳導性鈣離子活化型鉀離子通道開啟機率 (NPo) 及鈣離子敏感性於SAH 組則有減小現象產生。

由實驗結果推測，當大鼠於蜘蛛膜下腔出血模式後，腦血管平滑肌細胞上大傳導性鈣離子活化性鉀離子通道的活性下降的現象，可能是由於其離子通道對於鈣離子敏感性產生變化所造成。因此，KMUP-1及Pinacidil也許能提供一保護性作用，進而防止SAH所誘導血管痙攣的現象產生。

關鍵字

蜘蛛膜下腔出血 ； 電生理 ； 鉀離子通道

並列摘要

Cerebral vasospasm after aneurismal subarachnoid hemorrhage (SAH) is characterized by diffuse and long-lasting narrowing of arteries. The large conductance Ca2+-activated K+ (BKCa) channel plays a negative feedback role to prevent vasospasm. KMUP-1, a xanthine-based derivative, has been demonstrated to activate K+ channels resulting in the relaxation of aortic and tracheal smooth muscles. In this study, we tried to investigate whether SAH-induced vasospasm could be a defect in the function of vascular BKCa channels, and therefore to explore the protective activity of KMUP-1 in SAH rats.

Rats were divided into four groups including control, SAH, KMUP-1-treated and pinacidil-treated groups. Rats were subjected to experimental SAH by injecting autologous blood into the cisterna magna, then KMUP-1 (1 mg/kg) and pinacidil (1 mg/kg) were administered intraperitoneally at 1 h after SAH in this study. All rats were sacrificed at 24 h after SAH in this study.

Cerebrovascular smooth muscle cells (CVSMCs) were enzymatically isolated from rat basilar arteries. Using whole-cell patch-clamp techniques, the iberiotoxin (IbTX)-sensitive BKCa currents attenuated in SAH groups compared with control groups. Pretreatment with KMUP-1 or pinacidil, the IbTX-sensitive currents were restored. In inside-out patches, the unitary conductance and voltage sensitivity of SAH, KMUP-1 and pinacidil groups showed effects similar to the results of control groups. However, the BKCa channel open probability (NPo) and calcium sensitivity were decreased in SAH groups compared with control groups.

In conclusion, we suggest that the down-regulation of the BKCa channel activity could be due to the modification of the BKCa channel’s calcium sensitivity. From the results indicated that KMUP-1 and pinacidil could protect against SAH-induced vasospasm.

並列關鍵字

Subarachnoid hemorrhage ； Electrophysiology ； Potassium channel

參考文獻

Aihara Y, Jahromi BS, Yassari R, Nikitina E, Agbaje-Williams M, Macdonald RL., 2004, Molecular profile of vascular ion channels after experimental subarachnoid hemorrhage. J Cereb Blood Flow Metab 24(1):75-83.

Alioua A, Tanaka Y, Wallner M, Hofmann F, Ruth P, Meera P, Toro L., 1998, The large conductance, voltage-dependent, and calcium-sensitive K+ channel, Hslo, is a target of cGMP-dependent protein kinase phosphorylation in vivo. J Biol Chem 273(49):32950-6.

Arai H, Hori S, Aramori I, Ohkubo H, Nakanishi S., 1990, Cloning and expression of a cDNA encoding an endothelin receptor. Nature 348(6303):730-2

Biller J, Godersky JC, Adams HP Jr., 1988, Management of aneurysmal subarachnoid hemorrhage. Stroke 19(10):1300-5.

Bolton TB, Lang RJ, Takewaki T., 1984, Mechanisms of action of noradrenaline and carbachol on smooth muscle of guinea-pig anterior mesenteric artery. J Physiol 351:549-72.