Neuropeptide Y stimulates retinal neural cell proliferation – involvement of nitric oxide

Abstract

Neuropeptide Y (NPY) is a 36 amino acid peptide widely present in the CNS, including the retina. Previous studies have demonstrated that NPY promotes cell proliferation of rat post‐natal hippocampal and olfactory epithelium precursor cells. The aim of this work was to investigate the role of NPY on cell proliferation of rat retinal neural cells. For this purpose, primary retinal cell cultures expressing NPY, and NPY Y1, Y2, Y4 and Y5 receptors [Álvaro et al., (2007) Neurochem. Int., 50, 757] were used. NPY (10–1000 nM) stimulated cell proliferation through the activation of NPY Y1, Y2 and Y5 receptors. NPY also increased the number of proliferating neuronal progenitor cells (BrdU+/nestin+ cells). The intracellular mechanisms coupled to NPY receptors activation that mediate the increase in cell proliferation were also investigated. The stimulatory effect of NPY on cell proliferation was reduced by l‐nitroarginine‐methyl‐esther (l‐NAME; 500 μM), a nitric oxide synthase inhibitor, 1H‐[1,2,4]oxadiazolo‐[4, 3‐a]quinoxalin‐1‐one (ODQ; 20 μM), a soluble guanylyl cyclase inhibitor or U0126 (1 μM), an inhibitor of the extracellular signal‐regulated kinase 1/2 (ERK 1/2). In conclusion, NPY stimulates retinal neural cell proliferation, and this effect is mediated through nitric oxide–cyclic GMP and ERK 1/2 pathways.

Intracellular Nitric Oxide Mediates Neuroproliferative Effect of Neuropeptide Y on Postnatal Hippocampal Precursor Cells

“…In fact, a link between NO and the physiological effects of NPY was suggested by Morley et al (35), who showed that a low dose of NPY was able to increase NOS expression in the hypothalamus by 147% and that an NPY-induced increase in food intake was mediated via NOS. More recently, Alvaro et al (23) showed that NOS was involved in mediating the NPY-induced proliferation of retinal neural cells, further supporting our hypothesis. Through the use of pharmacological, immunocytochemical, and live cell imaging techniques, we describe the involvement of NO signaling in mediating the neuroproliferative effect of NPY on cultures derived from the postnatal rat hippocampus.…”

“…Recently, Lou et al (42) have shown that the effects of NO on neural stem/precursor cells differ significantly depending on whether it was released extracellularly (antiproliferative) or intracellularly (proliferative). Although the involvement of NO mechanisms was thought to underlie the proliferative effect of NPY in retinal neural cells (23), our results are the first to demonstrate that intracellular NO produced by nNOS signals the complete Y 1 receptor-mediated proliferative effect of NPY on hippocampal nestin ϩ precursors, whereas extracellular NO had the opposite antiproliferative effect. We also show that ERK1/2 activation by NPY is solely mediated via NOS-NO mechanisms.…”

“…It has been proposed that NPY-releasing interneurons in the dentate hilus release NPY onto progenitors within the SGZ to regulate their proliferation and adult hippocampal neurogenesis (22). The signaling mechanisms underlying the proliferative effect of NPY are not fully understood, although the extracellular signalregulated kinases (ERK) 1/2 of the mitogen-activated protein kinases (MAPK) signaling pathway have been shown to play a major role (14,20,23).…”

“…Exposure to Pharmacological Agonists and Antagonists and BrdU Incorporation-The concentrations of agonists or antagonists were chosen based on published literature and included NPY (1 M; Sigma-Aldrich) (15), the nonselective NOS inhibitor N()-nitro-L-arginine methyl ester (L-NAME) (500 M; Sigma-Aldrich) (23), the inactive enantiomer N()-nitro-D-arginine methyl ester (D-NAME) (500 M; Sigma), the selective Y 1 agonist [F7, P34]NPY (1 M; courtesy of Prof. Dr. A. G. BeckSickinger, Universität Leipzig, Germany) (15), the NOS substrate L-arginine (500 M; Sigma-Aldrich), the inactive enantiomer D-arginine (500 M; Sigma-Aldrich), the cGMP analog 8-Br-cGMP (20 M; Tocris Bioscience) (23) (42), and the NO scavenger carboxy-PTIO (CPTIO, 10 M; Tocris Bioscience) (42,43). Agonists or antagonists were added to cultures for 3 or 5 DIV or for 6 h on the final day before cells were rinsed once in 0.1 M phosphate-buffered saline (PBS; Sigma-Aldrich) and fixed in 4% paraformaldehyde (Sigma-Aldrich) for 20 min.…”

“…The signaling mechanisms initiated by the NPY Y receptors are diverse, and activation of ERK1/2 may be through an NOindependent pathway (13,23). The levels of ERK1/2 phosphorylation were assessed by Western blotting in response to NOS inhibition (L-NAME) to determine the importance of the NOcGMP pathway in mediating ERK1/2 activation and the proliferative effects of NPY (Fig.…”