RUI: Cytoskeletal Systems and Calcium in Ooplasmic Movementsand Cytokinesis in the Egg of a Teleost Fish, the Medaka (Oryzias latipes)

这项研究的目标是：1)确定微丝和微管在青冈卵卵质运动中的作用，2)确定细胞质钙脉冲、梯度和波在青冈卵卵质运动中的作用，以及3)确定细胞质分裂的机械事件与微丝、钙脉冲、青竹卵前两次卵裂过程中胚盘中的梯度和波。卵浆运动和胞质分裂将用延时视频显微镜进行研究，适当时使用相位对比、诺马尔斯基DIC和荧光光学技术。细胞骨架系统的作用将通过1)应用或微量注射扰乱这些系统的药物，以及2)分别用荧光鬼臼蛋白和间接免疫荧光来定位微丝和微管来研究。钙的作用将通过1)用钙敏感的光蛋白Aequorin寻找钙脉冲、梯度和波，以及2)将钙缓冲液注射到鸡蛋的细胞质中来研究。以鱼卵为模型，这些研究应该为受精卵细胞质的两个基本活动的生理基础提供新的见解：结构重排和分裂为子细胞。这项研究还将为本科生提供机会，利用最先进的光学显微镜技术参与积极的研究计划。

The goals of the proposed research are: 1) to determine the roles of microfilaments and microtubules in movements of the ooplasm in the medaka egg, 2) to determine the role of cytoplasmic calcium pulses, gradients, and waves in movements of the ooplasm of the medaka egg, and 3) to determine the relationship between the mechanical events of cytokinesis and microfilaments, calcium pulses, gradients and waves in the blastodisc during the first two cleavages of the medaka egg. Ooplasmic movements and cytokinesis will be studied with time-lapse video microscopy, using phase contrast, Nomarski DIC, and epifluorescence optics where appropriate. The role of cytoskeletal systems will be studied by 1) applying or microinjecting drugs that disrupt these systems, and 2) localizing microfilaments and microtubules with fluorescent phalloidins and by indirect immunofluorescence, respectively. The role of calcium will be studied by 1) looking for calcium pulses, gradients, and waves with the calcium-sensitive photoprotein, Aequorin, and 2) injecting calcium buffers into the cytoplasm of the egg. Using the fish egg as a model, these studies should provide new insights into the physiological bases of two fundamental activities of the cytoplasm of the fertilized egg: its structural rearrangement and its division into daughter cells. This research will also provide the opportunity for undergraduates to participate in an active research program using state of the art techniques of light microscopy.