MERCURY IN CHLORIDE TRANSPORT IN SHARK RECTAL GLAND & RABBIT THICK ASCENDING LIMB

本提案的目的是研究 汞对氯离子转运上皮细胞的毒性作用 汞会抑制鲨鱼直肠腺和 哺乳动物粗的上升肢，但其作用机制 行动是未知的。 在所提出的一组实验中，我们将研究 有机汞和无机汞对氯离子迁移的影响 并将其与厚的升 亨利氏环的一部分 汞、汞离子（无机汞和 有机）或任何化合物对直肠腺的氯转运的影响，或 粗升支可参照我们目前的 了解这些细胞的运输。 抑制作用是 可能在五个不同的网站：1）进入氯化物的网站， 细胞，Na-K-2Cl载体; 2）用Na-K-2Cl载体抑制Na-K-ATP酶。 导致钠、氯化物进入的驱动力降低 3）抑制细胞代谢 导致ATP不足以为泵提供动力; 4）抑制 通过减少顶端膜中的氯离子， 氯离子的膜电导率; 5）抑制钾循环 通过钾传导途径。 这些可能性可以是 使用特异性转运抑制剂与 汞 汞抑制氯离子转运的累加效应 在这些转运步骤的特异性抑制剂存在下， 表明作用位点不同于特异性抑制剂的作用位点。 因为我们发现了意想不到的和惊人的差异， 这两种上皮细胞对无机汞和有机汞的反应， 比较方法将特别富有成效。 我们将 研究汞的作用机制和细胞部位， 测定其在离体灌注直肠腺中的作用，离体 直肠腺小管，直肠腺膜泡，直肠腺浆 膜，厚的升肢细胞，厚的升肢囊泡， 上升肢质膜厚。 具体来说，我们将研究 汞与质膜和细胞溶质蛋白质的结合; 汞对氯协同转运蛋白的作用，钾的流出， 氯离子电导及其对运输相关酶的影响。

The intent of this proposal is to study the mechanism of action of the toxic effect of mercury on transport by chloride transporting epithelia. Mercury inhibits chloride transport in both the shark rectal gland and the mammalian thick ascending limb but its mechanism of action mechanism of action is unknown. In the proposed set of experiments we will study the effect of organic and inorganic mercury on the transport of chloride by the shark rectal gland and compare it with that in the thick ascending limb of the loop of Henle. The mechanism of the inhibition of mercury, mercurial (both inorganic and organic) or of any compound on chloride transport by the rectal gland or the thick ascending limb can be evaluated with reference to our present understanding of transport by these cells. Inhibitory effects are possible at five different sites: 1) The site of entry of chloride into the cell, the Na-K-2CI carrier; 2) Inhibition of Na-K-ATPase with the resulting reduction in the driving force for entry of sodium, chloride and potassium via the cotransporter; 3) Inhibition of cellular metabolism resulting in insufficient ATP to power the pump; 4) Inhibition of the exit of chloride at the apical membrane by a reduction in the apical membrane conductance for chloride; 5) inhibition of potassium recycling through the potassium conductive pathway. These possibilities can be distinguished using specific transport inhibitors in combination with mercury. An additive effect of mercurial to inhibit chloride transport in the presence of specific inhibitors of these transport steps will suggest a site of action of distinct from that of the specific inhibitor. Because we have found unexpected and striking differences in the response of these two epithelia to inorganic and organic mercurial, we believe that the comparative approach will be particularly fruitful. We will examine the mechanism and the cellular site of action of mercury by determining its effect in the isolated perfused rectal gland, isolated rectal gland tubules, rectal gland membrane vesicles, rectal gland plasma membranes, thick ascending limb cells, thick ascending limb vesicles, thick ascending limb plasma membranes. Specifically we will examine the binding of mercury by plasma membranes and cytosolic proteins; the effect of mercury on the chloride cotransporter, the efflux of potassium, the chloride conductance, and its effect on transport related enzymes.