Mechanism of extracellular ATP-induced inhibition of store-operated Ca^<2+> entry

在大多数细胞中，激动剂诱导的磷脂酶C/Gq激活产生IP_3，打开内质网中的Ca^<2+>通道。Ca^<2+>储存器的排空甚至减少触发储存器操作的Ca^<2+>穿过质膜进入，这被认为是Ca^<2+>在非电可兴奋细胞中的主要内流。我们已经报道了细胞外ATP完全抑制thapsigargin诱导的储存操作的Ca^<2+>进入大鼠原代培养的棕色脂肪细胞。在这个项目中，我们研究了atp诱导的抑制储存操作的Ca^<2+>入口的机制，得到了以下结果。在没有细胞外Mg^<2+>的情况下，细胞外ATP抑制了储存操作的Ca^<2+>的进入，这表明ect-酶的激活不起作用。P2受体拮抗剂苏拉明。PPADS抑制了thapsigargin诱导的Ca^<2+>内流，并抑制了Ca^<2+>的瞬时升高。细胞外ATP刺激肌动蛋白丝在细胞外周区域（靠近质膜）的重新分配。细胞外ATP增强了thapsigarin预处理细胞中肌动蛋白的外周聚合5。P2受体拮抗剂抑制atp诱导的外周肌动蛋白聚合6。肌动蛋白聚合抑制剂细胞松弛素D阻断ATP的作用，此外，细胞外ATP不抑制储存操作的Ca^<2+>进入细胞松弛素D预处理的细胞。这些观察结果表明，细胞外ATP通过P2受体介导刺激皮层肌动蛋白层的形成，从而阻断Ca^<2+>在大鼠棕色脂肪细胞中的内流。

Agonist-induced activation of phospholipase C/Gq produces IP_3 which opens the Ca^<2+> channels in ER in most of the cells. The emptying or even decrease of Ca^<2+> stores triggers the store-operated Ca^<2+> entry across the plasma membrane, which has been recognized to be the major Ca^<2+> influx in non-electric excitable cells. We have reported that the extracellular ATP completely Inhibits the thapsigargin-induced store-operated Ca^<2+> entry in rat primary cultured brown adipocytes. In this project, we investigated the mechanism of the ATP-induced inhibition of the store-operated Ca^<2+> entry and obtained the following results.1. Extracellular ATP inhibited the store-operated Ca^<2+> entry in the absence of extracellular Mg^<2+> suggesting that the activation of ect-enzymes does not play a role.2. P2 receptor antagonists suramin. and PPADS suppressed the thapsigargin-induced Ca^<2+> influx as well as the inhibition of the transient Ca^<2+> elevation.3. Extracellular ATP stimulated the redistribution of actin filaments in peripheral region of the cell (near the plasma membrane).4. Extracellular ATP enhanced peripheral polymerization of actin in thapsigargin-pretreated cells5. P2 receptor antagonists inhibited the ATP-lnduced peripheral actin polymerization6. Actin polymerization inhibitor cytochalasin D blocked the effect of ATP, and furthermore, extracellular ATP did not inhibit store-operated Ca^<2+> entry in cytochalasin D-pretreated cells.These observations suggest that extracellular ATP stimulates the formation of the cortical actin layer mediated through P2 receptors, resulting in blocking the influx of Ca^<2+> in rat brown adipocytes.