Contribution of bradykinin and nitric oxide to AT₂ receptor‐mediated differentiation in PC12 W cells

Abstract

We investigated the effect of angiotensin II on intracellular cyclic GMP content and neurite outgrowth as an indicator of cell differentiation in PC12 W cells. Neurite outgrowth was examined by phase-contrast microscopy. Outgrown neurites were classified as small, medium and large, and were expressed as neurites per 100 cells. Angiotensin II (10 )7 M) increased the outgrowth of medium and large neurites by mean ± SEM 20.2 ± 2.3 and 6.6 ± 1.4 compared with 1.66 ± 0.5 and 0.1 ± 0.06 neurites per 100 cells in control. Cellular cyclic GMP content increased by 50-250% with angiotensin II at concentrations of 10 )6 -10 )4 M. Both blockade of AT 2 receptors and of nitric oxide synthase markedly reduced angiotensin II-induced neurite outgrowth and cyclic GMP production. In contrast, B 2 receptor blockade had no effect or even increased these angiotensin II effects. Sodium nitroprusside and 8-bromo-cyclic GMP both mimicked the effects of angiotensin II on cell differentiation. The protein kinase G inhibitor KT-5823 inhibited the neurite outgrowth induced by both angiotensin II and 8-bromo-cyclic GMP. Our results demonstrate that angiotensin II can stimulate cell differentiation in PC12 W cells by nitric oxide-related and cyclic GMP-dependent mechanisms. The effects of angiotensin II on cell differentiation and cyclic GMP production were mediated via the AT 2 receptor and further enhanced by bradykinin B 2 receptor blockade. Keywords: angiotensin II, AT 2 receptor, cell differentiation, bradykinin, cyclic GMP, nitric oxide. Two major angiotensin II (Ang II) receptor subtypes, AT 1 and AT 2 , have been identified using specific receptor antagonists such as losartan and PD 123319. Most of the established physiological actions of Ang II, such as regulation of blood pressure and electrolyte homoeostasis, as well as its growth promoting actions, are mediated by the AT 1 receptor. However, results of recent studies in various tissues and cell lines have highlighted a number of actions of Ang II that are mediated by activation of AT 2 receptors, including growth inhibition, differentiation, apoptosis and nerve regeneration (Nakajima et al. 1995;Stoll et al. 1995;Yamada et al. 1996;Lucius et al. 1998).Two major strategies have been developed to block the renin-angiotensin system, first, the blockade of angiotensinconverting enzyme (ACE) by the action of ACE inhibitors and, second, the selective blockade of the AT 1 receptor by AT 1 receptor antagonists. ACE inhibitors inhibit the generation of Ang II and block the effects of the peptide on both the AT 1 and the AT 2 receptor. In addition, ACE inhibition in part prevents the degradation of bradykinin, leading to a stimulation of bradykinin B 2 receptor-mediated release of paracrine mediators, such as nitric oxide (NO) and/or prostaglandins. Bradykinin-mediated actions appear to be of major importance for the organ protective effects of ACE inhibitors . On the other hand, AT 1 receptor antagonists selectively block the AT 1 receptor, leave the AT 2 receptor unopposed and do...

Angiotensin II Type 2 Receptor–Bradykinin B 2 Receptor Functional Heterodimerization

“…effect of activated AT 2 R on cGMP and NO production. 38 Both B 2 R agonist and antagonist were observed to have a similar influence on NO production when combined with AT 2 R agonist. However, the mechanism of NO production because of B 2 R agonist is different from that of the antagonist.…”

Cardiovascular and renal function of angiotensin II type-2 receptors

“…Accordingly, in the rat aorta, ANG increases cGMP levels by stimulation of bradykinin and subsequent NO production via AT 2 receptors [33] and, in hypertrophied rat hearts, blockade of AT 2 receptors results in reduced cGMP levels that augments the growth promoting effect of AT 1 receptor stimulation [34]. Recent data also suggest the participation of NO/cGMP-and bradykininpathways in AT 2 receptor-mediated PC12W cell differentiation and in the inhibition of ANG-induced contraction of rat uterine arteries by AT 2 receptors [35,36].…”

Correlation between Neuritogenic Action of Nitric Oxide and the Rate of Nitric Oxide Production in PC12h Cells

“…In PC12 cells, it has been further shown that NO donors also enhanced the neuritogenic effects of NGF, however, NO donors did not induce PC12 cell neurites in the absence of NGF. 3,4) Meanwhile, we and other investigators have previously reported that several NO donors induced neurites in PC12h cells 5) and PC12W, 6) a subclone of PC12 cells, and in NG108-15. 7) NO plays an important role in intracellular and intercellular signaling events such as neural development, synaptic plasticity, and neuroendocrine secretion by low level NO production through neuronal and/or endothelial NOS activation.…”