Roles of endothelin receptors in astrocytic activation on brain pathologies

1. 内皮素诱导神经营养因子的产生：内皮素（ET）刺激大鼠培养的星形胶质细胞中已知的神经营养因子GDNF和BDNF的产生。通过去除细胞内Ca^<2+>和抑制ERK信号通路，et诱导的GDNF产生受到抑制。NFkB抑制剂（pyrrolidin dithiocarbamate）和糖皮质激素预处理对et诱导的GDNF的产生没有影响，而H_2O_2对GDNF的产生有抑制作用。脑室连续输注ETs 7 d后，海马和纹状体GDNF和BDNF分别增加。免疫组化检查显示这些神经营养因子是由星形胶质细胞产生的。这些发现提示星形细胞ET受体的激活可能是神经营养药物刺激神经营养因子的一个靶点。内皮素诱导星形细胞增殖：内皮素诱导星形细胞增殖的机制是通过聚焦局灶黏附激酶（FAK）的作用，FAK是一种与局灶黏附相关的酪氨酸激酶。ET刺激星形胶质细胞中FAK和ERK的磷酸化。细胞松弛素D抑制et诱导的FAK磷酸化，但对ERK无抑制作用。细胞松弛素D减少了ET-1诱导的溴脱氧尿嘧啶（BrdU）的掺入。瞬时转染野生型FAK后，星形细胞BrdU掺入增加。在FAK显性阴性突变体转染的星形胶质细胞中，ET-1对BrdU掺入的影响减弱。ET-1增加培养星形胶质细胞中cyclin D1和D3蛋白的表达。转染野生型FAK后，星形胶质细胞中cyclin D3的表达增加，而cyclin D1的表达不受影响。显性阴性FAK突变体可阻止et诱导的细胞周期蛋白D3表达升高，而D1表达不升高。这些结果提示FAK参与了ETs诱导的星形细胞增殖。

1. Endothelin-induced production of neurotrophic factors : Endothelin (ET) stimulated production of GDNF and BDNF, well-known neurotrophic factors, in rat cultured astorcytes. ET-induced GDNF production was inhibited by removal of intracellular Ca^<2+> and inhibition of ERK signal pathway. An inhibitor of NFkB (pyrrolidin dithiocarbamate) and pre-treatment with glucocorticoid had no effect on the ET-induced GDNF production, while GDNF production induced by H_2O_2 was prevented by them. Continuousinfusion of ETs into cerebral ventricle for 7 days increased GDNF and BDNF in hippocampus and striatum, respectively. Immunohistechemical examinations showed that these neurotrophic factors were produced by astrocytes. These findings suggest that activation of astrocytic ET receptors can be one target of neurotrophic drugs to stimulate neurotrophic factors.2. Endothelin-induced astrocytic proliferation : The mechanisms of ET-indeuced astrocytic proliferation by focusing roles of focal adhesion kinase (FAK), a tyrosine kinase associated with focal adhesions. ET stimulated phosphorylation of both FAK and ERK in astrocytes. Cytochalasin D inhibited ET-induced phosphorylation of FAK, but not that of ERK. Cytochalasin D reduced the incorporation of bromdeoxyuridine (BrdU) induced by ET-1. A transient transfection with wild-type FAK was followed byan increase in astrocytic BrdU incorporation. The effect of ET-1 on BrdU incorporation was diminished in astrocytes transfected with dominant-negative FAK mutants. ET-1 increased expression of cyclin D1 and D3 proteins in cultured astrocytes. Transfection with wild-type FAK increased expression of cyclin D3 in astrocytes, while that of cyclin D1 was not affected. The ET-induced increase in cyclin D3 expression, but not D1, was prevented by dominant-negative FAK mutants. These results suggest an involvement of FAK in astrocytic proliferation induced by ETs.