O'Dowd N A, Canny M J
Biology Department, Carleton University, 1125 Colonel By Drive, Ottawa, Canada KIS 5B6.
New Phytol. 1993 Dec;125(4):743-748. doi: 10.1111/j.1469-8137.1993.tb03923.x.
The identification of sites in leaves where transpiration water crosses cell membranes and enters the symplast has previously been made using freeze-substitution to locate concentrations of dye [e.g. sulphorhodamine (SR)] moving with the transpiration stream, and left outside the membranes where the water passes through. These concentrations were called sumps, and the sites of entry to the symplast were called flumes. A simple method of locating sumps, and therefore flumes, is described. Fresh leaves, fed SR solution through their cut petioles for pulse periods of 0.5 h or more, followed or not by a chase of water, were sectioned by hand under paraffin oil, and the sections mounted in the same fluid. Observation of the sections by simple bright-field microscopy revealed sumps of SR at the same sites, and of the same crystalline nature as found in the freeze-substituted preparations. The saving in preparation time is of the order of > 100-fold, at the sacrifice of resolution (5-10 μm compared with 0.2 μm). A limited survey of grass, sedge and dicotyledon leaves by this method confirmed in all essentials the results found by freeze-substitution, and in addition, revealed flumes at the fusoid cells on the flanks of the veins of bamboo leaves, and at the same position next to the water tissue of Cyperus leaves. The rate of accumulation of crystalline SR in the sumps inside tracheary elements suggests that the concentration of this non-permeating solute in the xylem sap increased by about 1000-fold in the finest veins during 1-2 h of transpiration in the dye solution.
此前，人们利用冷冻置换法来确定蒸腾水流穿过细胞膜并进入共质体的叶片部位，具体做法是定位随蒸腾流移动且留在水流经过的膜外的染料（如磺基罗丹明，SR）浓度。这些浓度区域被称为“汇”，进入共质体的部位则被称为“槽”。本文描述了一种定位“汇”进而确定“槽”的简单方法。将新鲜叶片通过其叶柄切口饲喂SR溶液0.5小时或更长时间的脉冲期，之后或不进行水的追踪冲洗，然后在石蜡油下手工切片，并将切片置于相同液体中。通过简单的明场显微镜观察切片，发现SR的“汇”位于与冷冻置换制剂中相同的部位，且具有相同的晶体性质。制备时间节省了100倍以上，但分辨率有所牺牲（从0.2μm降至5 - 10μm）。用这种方法对禾本科、莎草科和双子叶植物叶片进行的有限调查，在所有要点上都证实了冷冻置换法的结果，此外，还揭示了竹叶叶脉两侧的纺锤状细胞处以及莎草叶水组织旁边相同位置存在“槽”。管状分子内“汇”中结晶SR的积累速率表明，在染料溶液中蒸腾1 - 2小时期间，木质部汁液中这种非渗透性溶质的浓度在最细的叶脉中增加了约1000倍。
