Chaki S, Inagami T
Department of Biochemistry, Vanderbilt University School of Medicine, Nashville, Tennessee 37232.
Mol Pharmacol. 1993 Apr;43(4):603-8.
We previously reported that angiotensin II (Ang II) increases cGMP content through a new Ang II receptor subtype that is distinct from both the AT1 and AT2 subtypes in differentiated Neuro-2A cells. In this study, the mechanism of the Ang II-stimulated cGMP increase was investigated in comparison with bradykinin- and atrial natriuretic factor (ANF)-stimulated cGMP increases in differentiated Neuro-2A cells. Ang II increased cGMP in differentiated Neuro-2A cells rapidly, with a maximal effect in 30 sec and a return to basal levels in 60 sec. Removal of extracellular Ca2+ or pretreatment with a membrane-permeable Ca2+ chelator [1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid tetraacetoxymethyl ester] attenuated Ang II-stimulated cGMP accumulation. Both the time course and Ca2+ dependency of the effect of Ang II were similar to those of the effect of bradykinin, which activates soluble guanylyl cyclase, but distinct from those of the effect of ANF, which activates particulate guanylyl cyclase. Methylene blue, an inhibitor of soluble guanylyl cyclase, attenuated the effects of Ang II and bradykinin but not that of ANF. LaCl3, a nonspecific Ca2+ blocker, prevented Ang II-stimulated cGMP accumulation. L-type Ca2+ channel blockers, nifedipine and diltiazem, or an N-type Ca2+ channel blocker, omega-conotoxin, failed to inhibit the effect of Ang II. Ang II had no effect on formation of 1,4,5-inositol trisphosphate or cAMP content, whereas bradykinin stimulated 1,4,5-inositol trisphosphate formation in differentiated Neuro-2A cells. Further, the nitric oxide synthase inhibitors NG-monomethyl-L-arginine and NG-nitro-L-arginine attenuated Ang II- and bradykinin-stimulated elevation of cGMP content but not that stimulated by ANF. The Ca2+ ionophore A23187 also stimulated cGMP formation and the effect was inhibited by the nitric oxide synthase inhibitors. These results indicate that the newly found Ang II receptor mediates cGMP formation through activation of soluble guanylyl cyclase and that the activation is mediated by nitric oxide, which is increased by Ca2+ influx via an ion channel distinct from the L-type and N-type Ca2+ channels.
我们之前报道过，在分化的Neuro-2A细胞中，血管紧张素II（Ang II）通过一种新的Ang II受体亚型增加cGMP含量，该亚型与AT1和AT2亚型均不同。在本研究中，我们比较了在分化的Neuro-2A细胞中，Ang II刺激cGMP增加的机制与缓激肽和心房钠尿肽（ANF）刺激cGMP增加的机制。Ang II能迅速增加分化的Neuro-2A细胞中的cGMP，30秒时达到最大效应，60秒时恢复到基础水平。去除细胞外Ca2+或用膜通透性Ca2+螯合剂[1,2-双（2-氨基苯氧基）乙烷-N,N,N',N'-四乙酸四乙酰甲酯]预处理可减弱Ang II刺激的cGMP积累。Ang II作用的时间进程和Ca2+依赖性与缓激肽的作用相似，缓激肽激活可溶性鸟苷酸环化酶，但与ANF的作用不同，ANF激活颗粒性鸟苷酸环化酶。可溶性鸟苷酸环化酶抑制剂亚甲蓝减弱了Ang II和缓激肽的作用，但不影响ANF的作用。非特异性Ca2+阻滞剂LaCl3可阻止Ang II刺激的cGMP积累。L型Ca2+通道阻滞剂硝苯地平和地尔硫䓬，或N型Ca2+通道阻滞剂ω-芋螺毒素未能抑制Ang II的作用。Ang II对1,4,5-肌醇三磷酸的形成或cAMP含量无影响，而缓激肽可刺激分化的Neuro-2A细胞中1,4,5-肌醇三磷酸的形成。此外，一氧化氮合酶抑制剂NG-单甲基-L-精氨酸和NG-硝基-L-精氨酸减弱了Ang II和缓激肽刺激的cGMP含量升高，但不影响ANF刺激的升高。Ca2+离子载体A23187也刺激cGMP形成，且该作用被一氧化氮合酶抑制剂抑制。这些结果表明，新发现的Ang II受体通过激活可溶性鸟苷酸环化酶介导cGMP形成，且该激活由一氧化氮介导，一氧化氮通过与L型和N型Ca2+通道不同的离子通道内流增加。
