Growth hormone‐releasing peptide 6 protection of hypothalamic neurons from glutamate excitotoxicity is caspase independent and not mediated by insulin‐like growth factor I

Abstract

Treatment of the fetal hypothalamic neuronal cell line RCA-6 with growth hormone-releasing peptide 6, an agonist of the ghrelin receptor, or insulin-like growth factor I activates intracellular signalling cascades associated with anti-apoptotic actions. Abnormally high concentrations of glutamate provoke over-excitation of neurons leading to cell damage and apoptosis. Thus, the aim of this study was to investigate whether the administration of growth hormone-releasing peptide 6 and insulin-like growth factor I attenuates monosodium glutamate-induced apoptosis in RCA-6 neurons and the mechanisms involved. Two different mechanisms are involved in glutamate-induced cell death, one by means of caspase activation and the second through activation of a caspase-independent pathway of apoptosis mediated by the translocation of apoptosis-inducing factor. Growth hormone-releasing peptide 6 partially reversed glutamate-induced cell death but not the activation of caspases, suggesting blockage of the caspase-independent cell death pathway, which included interference with the translocation of apoptosis-inducing factor to the nucleus associated with the induction of Bcl-2. In contrast, the addition of insulin-like growth factor I to RCA-6 neurons abolished glutamate-induced caspase activation and cell death. These data demonstrate for the first time a neuroprotective role for growth hormone secretagogues in the caspase-independent cell death pathway and indicate that these peptides have neuroprotective effects independent of its induction of insulin-like growth factor I.

Neuroprotective actions of ghrelin and growth hormone secretagogues

“…GHRP-6 is capable of preventing this glutamate-induced cell death in both the hypothalamus and cerebellum (Delgado-Rubín de Célix et al, 2006). Moreover, MSG also induces cell death in the neuronal cell line RCA-6 by activating the intrinsic pathway of apoptosis through caspase-9 and -3/7 (Delgado-Rubín et al, 2009), which is in agreement with previous studies indicating that neurons and the neuroblastoma cell lines B50 and SH-SY5Y use the intrinsic pathway to undergo apoptotic cell death in response to excitotoxicity (Henshall et al, 2001; Mattson, 2003). However, although GHRP-6 partially inhibits MSG-induced cell death in RCA-6 neurons, it does not modify caspase-9 or -3/7 activities (Delgado-Rubín et al, 2009).…”

Involvement of Astrocytes in Mediating the Central Effects of Ghrelin

“…This is not related to an increase in the number of excitatory synapses as shown previously [111]. The glutamate release induced by ghrelin also stimulates the uptake of glutamate by astrocytes in order to prevent excess excitability and excitotoxicity to neighboring cells [127,128,129,130]. …”

“…Other studies report that ghrelin exerts its neuroprotective effects through stimulation of the protein kinase A and C pathways [151]. GHRP-6 is capable of preventing glutamate-induced neuronal death in both the hypothalamus and cerebellum [129] and also in the hypothalamic neuronal cell line RCA-6 [127]. Ghrelin has a neuroprotective role in hippocampal neurons against KA-induced excitotoxicity [161] via activation of the PI3K/AKT pathway and inhibition of the mitochondrial apoptotic pathway.…”

Differential effects of GH and GH-releasing peptide-6 on astrocytes

“…Treatment with GHRP-6 did not increase GFAP levels but stimulated proliferation in the hypothalamus and hippocampus. GHRP-6 and ghrelin stimulate the proliferation of a wide number of cells including osteoblasts (Kim et al 2005), cardiomyocytes (Pettersson et al 2002), somatotrophs (Dieguez & Casanueva 2000), endothelial cells (Rossi et al 2009) and adipocytes (Thompson et al 2004) and may also affect the CNS by protecting neurons from apoptosis (Frago et al 2002, 2011, Delgado-Rubin de Célix et al 2006, Delgado-Rubin et al 2009). We also evaluated the possible effect of GH and GHRP-6 on neuron number and did not observed any changes in the levels of Tuj1 in the hypothalamus or hippocampus.…”

“…GRLN-R1a activation leads to an increase in intracellular levels of Ca 2C via phosphatidylinositol-specific phospholipase C (Camiña et al 2003, Muccioli et al 2007). Ghrelin and GHRP-6 stimulate proliferation of different cell types (Dieguez & Casanueva 2000, Pettersson et al 2002, Thompson et al 2004, Rossi et al 2009) and may also affect the CNS by protecting neurons from apoptosis (Frago et al 2002, 2011, Delgado-Rubin de Célix et al 2006, Delgado-Rubin et al 2009). …”