Morphological and functional analysis of protease-activated receptor 2 on intracellular calcium dynamics in rat parotid gland acinar cells

蛋白酶激活受体（PARs）是一类新型的第七类跨膜结构域g蛋白偶联受体，可通过蛋白水解裂解激活。最近的研究表明，PARs存在于多种细胞中，并在许多重要功能的调节中发挥着重要作用。研究了PARs刺激是否会引起腮腺反应的问题；特别参考PARs刺激时细胞内Ca^<2+>([Ca^<2+>]_i)动力学。在本研究中，PAR2 mRNA在腮腺中强烈表达。在腮腺腺泡细胞中，PAR2-AP激活肽（PAR2-AP） SLIGRL-NH_2诱导[Ca^<2+>]_i升高。清除细胞外Ca^<2+>和使用Ca^<2+>通道阻滞剂均不能抑制par2 - ap诱导的[Ca^<2+>]_i升高。对PAR-2激活的反应主要是由细胞内Ca^<2+>储存的Ca^<2+>动员引起的。该肽诱导的Ca^<2+>释放和进入被一氧化氮合酶（NOS）抑制剂L-NAME部分抑制。NO供体GEA 3162，而不是8-溴- cgmp，模拟了PAR2激活非容性钙进入（NCCE）的作用。KN93（一种CAM激酶II抑制剂）和W7（一种钙调素抑制剂）都能完全阻断Ca^<2+>从细胞内Ca2+储存中释放和Ca^<2+>从细胞外空间内流。丁卡因和DHAB部分阻断了这些增加。这些结果表明PAR2-AP激活NCCE通路。我们提出par2在腮腺中的作用依赖于CAMKII， PAR2-AP激活红嘌呤样受体。

Protease-activated receptors(PARs) represent a novel class seven transmembrane domain G-protein coupled receptors, which are activated by proteolytic cleavage. Recent studies have reported that PARs are present in a variety of cells and have been prominently implicated in the regulation of a number of vital functions. The issue of whether the stimulation of PARs induces responses in parotid glands was examined ; with special reference to intracellular Ca^<2+>([Ca^<2+>]_i) dynamics during PARs stimulation. In the present study, PAR2 mRNA was expressed strongly in the parotid glands. In parotid acinar cells, PAR2-activating peptide(PAR2-AP), SLIGRL-NH_2, induced an increase in[Ca^<2+>]_i. Both removing of extracellular Ca^<2+> and using of Ca^<2+> channel blockers did not inhibit the PAR2-AP-induced[Ca^<2+>]_i increase. The response to PAR-2 activation was mainly caused by Ca^<2+> mobilization from intracellular Ca^<2+> stores. This peptide induced Ca^<2+> release and entry were partially inhibited by the nitric oxide synthase(NOS) inhibitor, L-NAME. The NO donor, GEA 3162, but not 8-bromo-cGMP, mimicked the effects of PAR2 in activating non capacitative calcium entry(NCCE). Both KN93(a CAM kinase II inhibitor) and W7(a calmodulin inhibitor) completely blocked a Ca^<2+> release from intracellular Ca2+store and a Ca^<2+> influx from extracellular spaces. Tetracaine and DHAB partially blocked these increases. These results indicate that PAR2-AP activates NCCE pathway. And we proposed that an effect of PAR-2 in parotid gland is dependent on CAMKII and a PAR2-AP activates the ryanodine-like receptors.