Gross Eric R, Hsu Anna K, Gross Garrett J
Medical College of Wisconsin, Department of Pharmacology and Toxicology, 8701 Watertown Plank Rd, Milwaukee, WI 53226, USA.
Circ Res. 2004 Apr 16;94(7):960-6. doi: 10.1161/01.RES.0000122392.33172.09. Epub 2004 Feb 19.
Glycogen synthase kinase (GSK) inhibition produced by ischemic preconditioning has been previously shown to be regulated through phosphatidylinositol-3 kinase (PI3K). Therefore, we determined whether opioid-induced cardioprotection (OIC) occurs during reperfusion by altering GSK phosphorylation through PI3K and target of rapamycin (TOR). Furthermore, we determined if selective GSK inhibitors, SB216763(SB21) or SB415286(SB41), emulate OIC. Rats were treated with the nonselective opioid agonist, morphine (MOR, 0.3 mg/kg), the delta-selective opioid agonist BW373U86 (BW, 1 mg/kg), or the GSK inhibitors, SB21 (0.6 mg/kg) or SB41(1.0 mg/kg), either 10 minutes before ischemia or 5 minutes before reperfusion. Five minutes before opioid or SB21 treatment, some rats received either the PI3K inhibitor wortmannin (15 microg/kg) or LY294002 (0.3 mg/kg) or the TOR inhibitor rapamycin (3 microg/kg). After 30 minutes of ischemia followed by 2 hours of reperfusion, infarct size was assessed. MOR, BW, SB41, and SB21 reduced infarct size compared with vehicle when administered before ischemia (42.9+/-2.6, 40.3+/-2.3, 46.6+/-1.6, 42.2+/-1.8 versus 60.0+/-1.1%, respectively; P<0.001) and showed similar protection when administered 5 minutes before reperfusion (43.6+/-2.3, 40.2+/-2.6, 44.8+/-2.8, 39.4+/-0.8%, respectively; P<0.001). Wortmannin, LY294002, and rapamycin were found to inhibit OIC; however, they did not abrogate SB21-induced infarct size reduction. At 5 minutes of reperfusion, both MOR and BW increased P-GSKbeta at Ser9 in the ischemic zone compared with vehicle (181+/-20, 178+/-15 versus 75+/-17 DU, respectively; P<0.05), and this effect was abrogated by prior administration of wortmannin or rapamycin in MOR-treated rats. Furthermore, no differences were seen in phosphorylation of GSKalpha (Ser21 or Tyr279) or phosphorylation of GSKbeta (Tyr216). These data indicate that OIC occurs via the phosphorylation of GSKbeta at Ser9 during reperfusion.
先前已表明，缺血预处理产生的糖原合酶激酶（GSK）抑制作用是通过磷脂酰肌醇-3激酶（PI3K）来调节的。因此，我们确定阿片类药物诱导的心脏保护作用（OIC）在再灌注期间是否通过PI3K和雷帕霉素靶蛋白（TOR）改变GSK磷酸化而发生。此外，我们确定选择性GSK抑制剂SB216763（SB21）或SB415286（SB41）是否能模拟OIC。在缺血前10分钟或再灌注前5分钟，用非选择性阿片类激动剂吗啡（MOR，0.3mg/kg）、δ-选择性阿片类激动剂BW373U86（BW，1mg/kg）或GSK抑制剂SB21（0.6mg/kg）或SB41（1.0mg/kg）对大鼠进行处理。在给予阿片类药物或SB21前5分钟，一些大鼠接受PI3K抑制剂渥曼青霉素（15μg/kg）或LY294002（0.3mg/kg）或TOR抑制剂雷帕霉素（3μg/kg）。缺血30分钟后再灌注2小时，评估梗死面积。与给予溶媒相比，缺血前给予MOR、BW、SB41和SB21可减小梗死面积（分别为42.9±2.6%、40.3±2.3%、46.6±1.6%、42.2±1.8%对60.0±1.1%；P<0.001），再灌注前5分钟给予时也显示出类似的保护作用（分别为43.6±2.3%、40.2±2.6%、44.8±2.8%、39.4±0.8%；P<0.001）。发现渥曼青霉素、LY294002和雷帕霉素可抑制OIC；然而，它们并未消除SB21诱导的梗死面积减小。在再灌注5分钟时，与溶媒相比，缺血区MOR和BW均增加了Ser9位点的磷酸化GSKβ（分别为181±20、178±15对75±17光密度单位；P<0.05），在MOR处理的大鼠中，预先给予渥曼青霉素或雷帕霉素可消除这种作用。此外，在GSKα（Ser21或Tyr279）的磷酸化或GSKβ（Tyr216）的磷酸化方面未观察到差异。这些数据表明，OIC在再灌注期间通过Ser9位点的GSKβ磷酸化而发生。
