Sanches Glauco, de Alencar Laura Sá, Ventura Ana Lucia Marques
Departament of Neurobiology, Institute of Biology, Federal Fluminense University, Cx. Postal 100180, Niterói, RJ 24001-970, Brazil.
Int J Dev Neurosci. 2002 Feb;20(1):21-7. doi: 10.1016/s0736-5748(02)00004-7.
Both ATP and acetylcholine can induce the mobilization of intracellular calcium in the early developing chick embryo retina, a response that decreases during retinal development. In this study, the effects of these transmitters on the turnover of phosphoinositides and proliferation of developing retinal cells in culture were characterized. While ATP, UTP or carbachol were able to induce a >400% accumulation of phosphoinositides in retinal cell cultures, only ATP promoted a dose-dependent increase in [(3)H]-thymidine incorporation in cultured cells (EC(50)=8.6 microM), a response that was inhibited by the P2 receptor antagonist pyridoxal-phosphate-6-azophenyl-2',4'-disulfonic acid (PPADS) (0.1 or 0.25 mM). ADP, but not UTP or adenosine, also stimulated the proliferation of retinal cells (EC(50)=5.8 microM), indicating that activation of P2Y1 receptors mediates the proliferative response of retinal cells to ATP. The mitogenic effect of ATP was completely prevented by the PKC inhibitor chelerythrine chloride (0.5 microM) and the phospholipase C (PLC) inhibitor U73122 (0.5 microM). PD 98059 (25 or 50 microM), an inhibitor of the activation of extracellular signal-regulated kinases (ERKs) also blocked the increase in [(3)H]-thymidine incorporation induced by ATP. Moreover, the effect of ATP was pronounced in cultures obtained from retinas at embryonic days 6-8, but not at day 9. Since Müller and bipolar cells are the predominant cell types that proliferate at these embryonic stages, our data suggest that ATP, through activation of P2Y1 receptors coupled to phospholipase C, PKC and MAP kinases, affects DNA synthesis in one or both of these cell types in culture.
ATP和乙酰胆碱均可诱导早期发育的鸡胚视网膜中细胞内钙的动员，这种反应在视网膜发育过程中会减弱。在本研究中，对这些递质对培养的视网膜细胞中磷酸肌醇周转和发育中视网膜细胞增殖的影响进行了表征。虽然ATP、UTP或卡巴胆碱能够在视网膜细胞培养物中诱导磷酸肌醇积累超过400%，但只有ATP能促进培养细胞中[³H] - 胸腺嘧啶核苷掺入量呈剂量依赖性增加（EC₅₀ = 8.6 μM），P2受体拮抗剂磷酸吡哆醛 - 6 - 偶氮苯 - 2',4'-二磺酸（PPADS）（0.1或0.25 mM）可抑制这种反应。ADP能刺激视网膜细胞增殖（EC₅₀ = 5.8 μM），而UTP或腺苷则不能，这表明P2Y1受体的激活介导了视网膜细胞对ATP的增殖反应。ATP的促有丝分裂作用被PKC抑制剂氯化白屈菜红碱（0.5 μM）和磷脂酶C（PLC）抑制剂U73122（0.5 μM）完全阻断。细胞外信号调节激酶（ERK）激活抑制剂PD 98059（25或50 μM）也能阻断ATP诱导的[³H] - 胸腺嘧啶核苷掺入量增加。此外，ATP的作用在胚胎第6 - 8天视网膜获得的培养物中明显，但在第9天则不明显。由于穆勒细胞和双极细胞是这些胚胎阶段增殖的主要细胞类型，我们的数据表明，ATP通过激活与磷脂酶C、PKC和丝裂原活化蛋白激酶偶联的P2Y1受体，影响培养中这一种或两种细胞类型的DNA合成。
