Arita J, Hashi A, Hoshi K, Mazawa S, Suzuki S
Department of Physiology, Yamanashi Medical University, Yamanashi, Japan.
Neuroendocrinology. 1998 Sep;68(3):163-71. doi: 10.1159/000054362.
Dopaminergic agonists are effective in vivo in inhibiting lactotrope proliferation and prolactin (PRL)-secreting pituitary tumors. The purpose of the present study was to demonstrate in vitro actions of dopaminergic agents on proliferation and cell shape of rat lactotropes. Anterior pituitary cells cultured with serum-free, chemically defined medium were treated with dopaminergic agents and were labeled with 5-bromo-2'-deoxyuridine (BrdU) for 3 h before the end of culture. BrdU-labeling indices indicative of the proliferation rate of lactotropes were determined by double immunofluorescence staining for BrdU and PRL. Treatment with dopamine for 21 h decreased BrdU-labeling indices of lactotropes in a dose-dependent manner with a nadir at 3 x 10(-7) M. The inhibitory action of 10(-5) M dopamine appeared 15 h after the initiation of treatment and became pronounced with time up to 33 h. The dopamine action was mimicked by treatment with the D2 receptor agonist bromocriptine at concentrations over 10(-9) M. Phase-contrast microscopy revealed that the flat polygonal cell shape of cultured lactotropes had changed to a round refractive cell shape after treatment with dopamine or bromocriptine, and that these changes in cell shape exactly paralleled those in the BrdU-labeling index. The changes in cell shape of lactotropes were accompanied by changes in subcellular distribution of actin filaments. Pretreatment with 10(-7) M eticlopride, a D2 receptor antagonist, blocked the dopamine- or bromocriptine-induced changes in both BrdU-labeling index and cell shape. These results suggest that (1) the in vitro experimental system established in the present study is a good model for studying the mechanism of the antiproliferative action of dopamine and (2) D2-receptor-mediated inhibition of proliferation of lactotropes in serum-free culture is closely related to changes in actin organization and cell shape.
多巴胺能激动剂在体内可有效抑制催乳素细胞增殖及分泌催乳素（PRL）的垂体肿瘤。本研究的目的是证明多巴胺能药物对大鼠催乳素细胞增殖和细胞形态的体外作用。用无血清、化学成分明确的培养基培养的垂体前叶细胞，用多巴胺能药物处理，并在培养结束前3小时用5-溴-2'-脱氧尿苷（BrdU）标记。通过对BrdU和PRL进行双重免疫荧光染色来测定指示催乳素细胞增殖率的BrdU标记指数。用多巴胺处理21小时可使催乳素细胞的BrdU标记指数呈剂量依赖性降低，在3×10⁻⁷ M时达到最低点。10⁻⁵ M多巴胺的抑制作用在处理开始后15小时出现，并随时间延长至33小时而变得明显。D2受体激动剂溴隐亭在浓度超过10⁻⁹ M时可模拟多巴胺的作用。相差显微镜显示，用多巴胺或溴隐亭处理后，培养的催乳素细胞扁平多边形的细胞形态已变为圆形折光细胞形态，且这些细胞形态的变化与BrdU标记指数的变化完全平行。催乳素细胞形态的变化伴随着肌动蛋白丝亚细胞分布的变化。用D2受体拮抗剂10⁻⁷ M甲氧氯普胺预处理可阻断多巴胺或溴隐亭诱导的BrdU标记指数和细胞形态的变化。这些结果表明：（1）本研究建立的体外实验系统是研究多巴胺抗增殖作用机制的良好模型；（2）在无血清培养中，D2受体介导的催乳素细胞增殖抑制与肌动蛋白组织和细胞形态的变化密切相关。
