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The effect of hydrogen peroxide (H202) on the intracellular Ca2+ concentration ([Ca 2+]i) of

The effects of mercury and cadmium on the intracellular level of nonproteinaceous thiols in a unicellular green alga Tetraselmis tetrathele (West) Butcher (Prasinophyceae) were investigated by using a fluorescent dye, 5‐chloromethylfluorescein (5CMF), as a probe for nonproteinaceous thiols. The 5CMF fluorescence was observed in cytoplasm, and the intensity of the fluorescence was decreased by exposure of the cells to HgCl2. Analysis of the fluorescent intensity of 5CMF by flow cytometry made it possible to distinguish cells in three states during the dying process caused by HgCl2: a normal state, a thiol‐depleted state, and a dead state. Depletion of nonproteinaceous thiols began within 30 min, and they were completely depleted at 2 h. Most cells died after 24 h of exposure to more than 3.0 μM HgCl2, whereas exposure up to 1.0 mM CdCl2 did not cause depletion of nonproteinaceous thiols or cell death within 48 h.  HPLC analyses revealed that glutathione was a major nonprotein thiol in T. tetrathele and that it was oxidized by exposing the cells to HgCl2. Phytochelatins, which play a great role in the tolerance to heavy metals of higher plants and many algae, could not be found in T. tetrathele. However, a tripeptide, Arg‐Arg‐Glu, was found to be abundant, and it showed ability to bind Hg2+, suggesting that it functions to scavenge heavy metals as well as thiol molecules.

Effects of FTY720 (2-amino-2-(2-[4-octylphenyl]ethyl)-1,3-propanediol HCl), a novel immunosuppressant, were examined on neurons and thymocytes respectively dissociated from rat brains and thymus glands using a flow cytometer to see if FTY720 exerts cytotoxic actions not only on spleen cells as previously reported but also on the other cells. FTY720 at a concentration of 10 microM deteriorated almost all of the thymocytes, while it was not the case for brain neurons. FTY720 increased the intracellular concentration of Ca2+ ([Ca2+]i) of thymocytes in both the presence and absence of external Ca2+, although the [Ca2+]i increased by FTY720 in the presence of external Ca2+ was much greater than that in the absence of external Ca2+. Thus, FTY720 may increase the membrane permeability of Ca2+ and release Ca2+ from intracellular Ca2+ stores in thymocytes. Furthermore, the number of thymocytes stained with ethidium, a dye impermeant to intact membranes, time-dependently increased after drug application. Therefore, FTY720 at concentrations of 3 - 10 microM non-specifically increases the membrane permeability of thymocytes, resulting in necrotic cell death, although FTY720 at micromolar concentrations was reported to induce apoptosis of spleen cells.