Department of Pharmacology, Temple University School of Medicine, Philadelphia, Pennslyvania 19140, USA.
Am J Physiol Cell Physiol. 2011 Sep;301(3):C559-65. doi: 10.1152/ajpcell.00123.2011. Epub 2011 May 25.
Angiotensin II is a modulator of myometrial activity; both AT(1) and AT(2) receptors are expressed in myometrium. Since in other tissues angiotensin II has been reported to activate intracellular receptors, we assessed the effects of intracellular administration of angiotensin II via microinjection on myometrium, using calcium imaging. Intracellular injection of angiotensin II increased cytosolic Ca(2+) concentration (Ca(2+)) in myometrial cells in a dose-dependent manner. The effect was abolished by the AT(1) receptor antagonist losartan but not by the AT(2) receptor antagonist PD-123319. Disruption of the endo-lysosomal system, but not that of Golgi apparatus, prevented the angiotensin II-induced increase in Ca(2+). Blockade of AT(1) receptor internalization had no effect, whereas blockade of microautophagy abolished the increase in Ca(2+) produced by intracellular injection of angiotensin II; this indicates that microautophagy is a critical step in transporting the peptide into the endo-lysosomes lumenum. The response to angiotensin II was slightly reduced in Ca(2+)-free saline, indicating a major involvement of Ca(2+) release from internal stores. Blockade of inositol 1,4,5-trisphosphate (IP(3)) receptors with heparin and xestospongin C or inhibition of phospholipase C (PLC) with U-73122 abolished the response to angiotensin II, supporting the involvement of PLC-IP(3) pathway. Angiotensin II-induced increase in Ca(2+) was slightly reduced by antagonism of ryanodine receptors. Taken together, our results indicate for the first time that in myometrial cells, intracellular angiotensin II activates AT(1)-like receptors on lysosomes and activates PLC-IP(3)-dependent Ca(2+) release from endoplasmic reticulum; the response is further augmented by a Ca(2+)-induced Ca(2+) release mechanism via ryanodine receptors activation.
血管紧张素 II 是子宫肌活动的调节剂；在子宫肌中均表达 AT(1)和 AT(2)受体。由于在其他组织中已报道血管紧张素 II 激活细胞内受体，我们通过钙成像评估了通过微注射将血管紧张素 II 细胞内给药对子宫肌的影响。细胞内注射血管紧张素 II 以剂量依赖性方式增加子宫肌细胞中的细胞浆 Ca(2+)浓度 (Ca(2+))。该作用被 AT(1)受体拮抗剂 losartan 但不是 AT(2)受体拮抗剂 PD-123319 所阻断。破坏内溶酶体系统，但不是高尔基体，可防止血管紧张素 II 诱导的 Ca(2+)增加。阻断 AT(1)受体内化没有效果，而阻断微自噬则消除了细胞内注射血管紧张素 II 引起的 Ca(2+)增加；这表明微自噬是将肽转运到内溶酶体腔中的关键步骤。在无钙盐水中，对血管紧张素 II 的反应略有降低，表明 Ca(2+)从内部储存中释放的主要参与。肝素和 xestospongin C 阻断肌醇 1,4,5-三磷酸 (IP(3))受体或 U-73122 抑制磷脂酶 C (PLC) 均消除了对血管紧张素 II 的反应，支持 PLC-IP(3)途径的参与。血管紧张素 II 诱导的 Ca(2+)增加通过拮抗肌醇 1,4,5-三磷酸 (IP(3))受体而略有降低。总之，我们的结果首次表明，在子宫肌细胞中，细胞内血管紧张素 II 在溶酶体上激活 AT(1)样受体，并激活 PLC-IP(3)-依赖性内质网 Ca(2+)释放；通过肌醇 1,4,5-三磷酸 (IP(3))受体激活诱导的 Ca(2+)释放机制进一步增强了该反应。
