Binding and internalization of a fluorescent vasopressin analogue by collecting duct cells.
Binding and internalization of a fluorescent vasopressin analogue by collecting duct cells.
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通过集合管细胞结合和内化荧光加压素类似物。
DOI：
10.1152/ajpcell.1988.255.5.c622
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发表时间：
1988
期刊：
影响因子：
--
通讯作者：
Kirk,KL
中科院分区：
文献类型：
--
作者：
Kirk,KL
用计算机辅助视频显微镜定量评价了微灌注兔皮质集合小管内主细胞对荧光加压素类似物[1-脱氨基-8-罗丹明赖氨酸加压素（rhoda LVP）]的结合和内化动力学。在25 ℃下，rhoda LVP的结合表现出饱和动力学，在2 nM下具有半最大结合，在大于5 nM的浓度下具有最大结合。Rhoda LVP结合可以通过同时加入10倍高浓度的精氨酸加压素（AVP）或V2受体激动剂1-脱氨基-8-D-精氨酸加压素（去氨加压素）来阻止。在25 ℃下没有检测到明显的内化或rhoda LVP，即，在加入RhodaLVP后，RhodaLVP荧光保持定位于每个主细胞的基底极至少100分钟，并且可以通过随后加入AVP而在很大程度上逆转。相反，在结合开始后将细胞加热至38摄氏度，导致荧光迅速（不到30分钟）迁移到细胞内部，并失去细胞表面的AVP可置换结合。这些结果证明了这种非侵入性光学策略的实用性，用于定量监测激素与完整的集合小管细胞的结合，并证明rhoda LVP与集合小管中主细胞上的V2受体可逆且具有高亲和力地结合。在38摄氏度下，受体复合物的内化（和假定的失活）可能有助于收集小管细胞在生理温度下对加压素的脱敏。
The kinetics of binding and internalization of a fluorescent vasopressin analogue [1-desamino-8-rhodamine lysine vasopressin (rhoda LVP)] by principal cells within the microperfused rabbit cortical collecting tubule were quantitatively assessed with computer-assisted video microscopy. At 25 degrees C, binding of rhoda LVP exhibited saturation kinetics with half-maximal binding at 2 nM and maximal binding at concentrations greater than 5 nM. Rhoda LVP binding could be prevented by the simultaneous addition of a 10-fold higher concentration of arginine vasopressin (AVP) or the V2-receptor agonist, 1-desamino-8-D-arginine vasopressin (desmopressin). No obvious internalization or rhoda LVP was detected at 25 degrees C, i.e., the rhoda LVP fluorescence remained localized to the basal pole of each principal cell for at least 100 min after rhoda LVP addition and could be largely reversed by the subsequent addition of AVP. Conversely, warming the cells to 38 degrees C after binding was initiated resulted in a rapid (less than 30 min) migration of the fluorescence into the cell interior and a loss of AVP-displaceable binding from the cell surface. These results document the utility of this noninvasive optical strategy for quantitatively monitoring hormone binding to intact collecting tubule cells and demonstrate that rhoda LVP binds reversibly and with high affinity to V2 receptors on principal cells in the collecting tubule. The internalization (and presumed inactivation) of the hormone-receptor complex at 38 degrees C may contribute to the desensitization of collecting tubule cells to vasopressin at physiological temperature.