Growth Hormone Neuroprotection Against Kainate Excitotoxicity in the Retina is Mediated by Notch/PTEN/Akt Signaling

Purpose: In the retina, growth hormone (GH) promotes axonal growth, synaptic restoration, and protective actions against excitotoxicity. Notch signaling pathway is critical for neural development and participates in the retinal neuroregenerative process. We investigated the interaction of GH with Notch signaling pathway during its neuroprotective effect against excitotoxic damage in the chicken retina. Methods: Kainate (KA) was used as excitotoxic agent and changes in the mRNA expression of several signaling markers were determined by qPCR. Also, changes in phosphorylation and immunoreactivity were determined by Western blotting. Histology and immunohistochemistry were performed for morphometric analysis. Overexpression of GH was performed in the quail neuroretinal-derived immortalized cell line (QNR/D) cell line. Exogenous GH was administered to retinal primary cell cultures to study the activation of signaling pathways. Results: KA disrupted the retinal cytoarchitecture and induced significant cell loss in several retinal layers, but the coaddition of GH effectively prevented these adverse effects. We showed that GH upregulates the Notch signaling pathway during neuroprotection leading to phosphorylation of the PI3K/Akt signaling pathways through downregulation of PTEN. In contrast, cotreatment of GH with the Notch signaling inhibitor, DAPT, prevented its neuroprotective effect against KA. We identified binding sites in Notch1 and Notch2 genes for STAT5. Also, GH prevented Muller cell transdifferentiation and downregulated Sox2, FGF2, and PCNA after cotreatment with KA. Additionally, GH modified TNF receptors immunoreactivity suggesting anti-inflammatory actions. Conclusions: Our data indicate that the neuroprotective effects of GH against KA injury in the retina are mediated through the regulation of Notch signaling. Additionally, anti-inflammatory and antiproliferative effects were observed.