Olfa Masmoudi-Kouki

Role of PACAP and VIP in astroglial functions

Astrocytes represent at least 50% of the volume of the human brain. Besides their roles in various supportive functions, astrocytes are involved in the regulation of stem cell proliferation, synaptic plasticity and neuroprotection. Astrocytes also influence neuronal physiology by responding to neurotransmitters and neuropeptides and by releasing regulatory factors termed gliotransmitters. In particular, astrocytes express the PACAP-specific receptor PAC1-R and the PACAP/VIP mutual receptors VPAC1-R and VPAC2-R during development and/or in the adult. There is now clear evidence that PACAP and VIP modulate a number of astrocyte activities such as proliferation, plasticity, glycogen production, and biosynthesis of neurotrophic factors and gliotransmitters.

Pituitary adenylate cyclase-activating polypeptide protects astroglial cells against oxidative stress-induced apoptosis

J. Neurochem. (2011) 117, 403–411.

The Octadecaneuropeptide ODN Protects Astrocytes against Hydrogen Peroxide-Induced Apoptosis via a PKA/MAPK-Dependent Mechanism

Astrocytes synthesize and release endozepines, a family of regulatory peptides, including the octadecaneuropeptide (ODN) an endogenous ligand of both central-type benzodiazepine (CBR) and metabotropic receptors. We have recently shown that ODN exerts a protective effect against hydrogen peroxide (H2O2)-induced oxidative stress in astrocytes. The purpose of the present study was to determine the type of receptor and the transduction pathways involved in the protective effect of ODN in cultured rat astrocytes. We have first observed a protective activity of ODN at very low concentrations that was abrogated by the metabotropic ODN receptor antagonist cyclo1–8[DLeu5]OP, but not by the CBR antagonist flumazenil. We have also found that the metabotropic ODN receptor is positively coupled to adenylyl cyclase in astrocytes and that the glioprotective action of ODN upon H2O2-induced astrocyte death is PKA- and MEK-dependent, but PLC/PKC-independent. Downstream of PKA, ODN induced ERK phosphorylation, which in turn activated the expression of the anti-apoptotic gene Bcl-2 and blocked the stimulation by H2O2 of the pro-apoptotic gene Bax. The effect of ODN on the Bax/Bcl-2 balance contributed to abolish the deleterious action of H2O2 on mitochondrial membrane integrity and caspase-3 activation. Finally, the inhibitory effect of ODN on caspase-3 activity was shown to be PKA and MEK-dependent. In conclusion, the present results demonstrate that the potent glioprotective action of ODN against oxidative stress involves the metabotropic ODN receptor coupled to the PKA/ERK-kinase pathway to inhibit caspase-3 activation.

Protective effect of the octadecaneuropeptide on hydrogen peroxide‐induced oxidative stress and cell death in cultured rat astrocytes

J. Neurochem. (2011) 118, 416–428.

The octadecaneuropeptide ODN prevents 6-hydroxydopamine-induced apoptosis of cerebellar granule neurons through a PKC-MAPK-dependent pathway.

Oxidative stress, induced by various neurodegenerative diseases, initiates a cascade of events leading to apoptosis, and thus plays a critical role in neuronal injury. In this study, we have investigated the potential neuroprotective effect of the octadecaneuropeptide (ODN) on 6‐hydroxydopamine (6‐OHDA)‐induced oxidative stress and apoptosis in cerebellar granule neurons (CGN). ODN, which is produced by astrocytes, is an endogenous ligand for both central‐type benzodiazepine receptors (CBR) and a metabotropic receptor. Incubation of neurons with subnanomolar concentrations of ODN (10−18 to 10−12 M) inhibited 6‐OHDA‐evoked cell death in a concentration‐dependent manner. The effect of ODN on neuronal survival was abrogated by the metabotropic receptor antagonist, cyclo1–8[DLeu5]OP, but not by a CBR antagonist. ODN stimulated polyphosphoinositide turnover and ERK phosphorylation in CGN. The protective effect of ODN against 6‐OHDA toxicity involved the phospholipase C/ERK MAPK transduction cascade. 6‐OHDA treatment induced an accumulation of reactive oxygen species, an increase of the expression of the pro‐apoptotic gene Bax, a drop of the mitochondrial membrane potential and a stimulation of caspase‐3 activity. Exposure of 6‐OHDA‐treated cells to ODN blocked all the deleterious effects of the toxin. Taken together, these data demonstrate for the first time that ODN is a neuroprotective agent that prevents 6‐OHDA‐induced oxidative stress and apoptotic cell death.

Neuroglobin protects astroglial cells from hydrogen peroxide-induced oxidative stress and apoptotic cell death

Oxidative stress, resulting from accumulation of reactive oxygen species, plays a critical role in astroglial cell death occurring in diverse neuropathological conditions. Numerous studies indicate that neuroglobin (Ngb) promotes neuron survival, but nothing is known regarding the action of Ngb in astroglial cell survival. Thus, the purpose of this study was to investigate the potential glioprotective effect of Ngb on hydrogen peroxide (H2O2)‐induced oxidative stress and apoptosis in cultured mouse astrocytes. Incubation of cells with subnanomolar concentrations of Ngb (10−14–10−10 M) was found to prevent both H2O2‐evoked reduction in surviving cells number and accumulation of reactive oxygen species in a concentration‐dependent manner. Furthermore, Ngb treatment abolishes H2O2‐induced increase in mitochondrial oxygen consumption rates. Concomitantly, Ngb treatment rescues H2O2‐associated reduced expression of endogenous antioxidant enzymes (superoxide dismutases and catalase) and prevents the stimulation of the expression of pro‐inflammatory genes (inducible nitric oxide synthase, cyclooxygenase‐2, and interleukin (IL) IL‐6 and IL‐33). Moreover, Ngb blocks the stimulation of Bax (pro‐apoptotic) and the inhibition of Bcl‐2 (anti‐apoptotic) gene expression induced by H2O2, which in turn abolishes caspase 3 activation. The protective effect of Ngb upon H2O2 induced activation of caspase 3 activity and cell death can be accounted for by activation of protein kinase A and mitogen‐activated protein kinase transduction cascade. Finally, we demonstrate that Ngb increases Akt phosphorylation and prevents H2O2‐provoked inhibition of ERK and Akt phosphorylation. Taken together, these data demonstrate for the first time that Ngb is a glioprotective agent that prevents H2O2‐induced oxidative stress and apoptotic astroglial cell death. Protection of astrocytes from oxidative insult may thus contribute to the neuroprotective effect of Ngb.

Octadecaneuropeptide ODN prevents hydrogen peroxide-induced oxidative damage of biomolecules in cultured rat astrocytes

Oxidative stress, associated with a variety of disorders including neurodegenerative diseases, is a major cause of cellular dysfunction and biomolecule damages which play a crucial role in neuronal apoptosis. Astrocytes specifically synthesize and release endozepines, a family of regulatory peptides, including the octadecaneuropeptide ODN. We have recently shown that ODN is a potent glioprotective agent that prevents hydrogen peroxide (H2O2)-induced oxidative stress and apoptosis. The purpose of the present study was to investigate the potential protective effect of ODN on oxidative-generated damage of biomolecules in cultured rat astrocytes. Incubation of cells with subnanomolar concentrations of ODN (0.1fM-0.1nM) inhibited H2O2-evoked reactive oxygen species accumulation and cell death in a concentration-dependent manner. Exposure of H2O2-treated cells to 0.1nM ODN inhibited superoxide anion generation and blocked oxidative damage of cell molecules caused by H2O2i.e. formation and accumulation of lipid oxidation products, malondialdehydes and conjugated dienes, and protein carbonyl compounds. Taken together, these data demonstrate for the first time that ODN prevents oxidative stress-induced alteration of cellular constituents. ODN is thus a potential candidate to reduce neuronal damage in various pathological conditions involving oxidative neurodegeneration.

Involvement of endogenous antioxidant systems in the protective activity of pituitary adenylate cyclase-activating polypeptide against hydrogen peroxide-induced oxidative damages in cultured rat astrocytes

Astroglial cells possess an array of cellular defense mechanisms, including superoxide dismutase (SOD) and catalase antioxidant enzymes, to prevent damages caused by oxidative stress. Nevertheless, astroglial cell viability and functionality can be affected by significant oxidative stress. We have previously shown that pituitary adenylate cyclase‐activating polypeptide (PACAP) is a potent glioprotective agent that prevents hydrogen peroxide (H2O2)‐induced apoptosis in cultured astrocytes. The purpose of this study was to investigate the potential protective effect of PACAP against oxidative‐generated alteration of astrocytic antioxidant systems. Incubation of cells with subnanomolar concentrations of PACAP inhibited H2O2‐evoked reactive oxygen species accumulation, mitochondrial respiratory burst, and caspase‐3 mRNA level increase. PACAP also stimulated SOD and catalase activities in a concentration‐dependent manner, and counteracted the inhibitory effect of H2O2 on the activity of these two antioxidant enzymes. The protective action of PACAP against H2O2‐evoked inhibition of antioxidant systems in astrocytes was protein kinase A, PKC, and MAP‐kinase dependent. In the presence of H2O2, the SOD blocker NaCN and the catalase inhibitor 3‐aminotriazole, both suppressed the protective effects of PACAP on SOD and catalase activities, mitochondrial function, and cell survival. Taken together, these results indicate that the anti‐apoptotic effect of PACAP on astroglial cells can account for the activation of endogenous antioxidant enzymes and reduction in respiration rate, thus preserving mitochondrial integrity and preventing caspase‐3 expression provoked by oxidative stress. Considering its powerful anti‐apoptotic and anti‐oxidative properties, the PACAPergic signaling system should thus be considered for the development of new therapeutical approaches to cure various pathologies involving oxidative neurodegeneration.

Beta-amyloid peptide stimulates endozepine release in cultured rat astrocytes through activation of N-formyl peptide receptors

Astroglial cells synthesize and release endozepines, a family of neuropeptides derived from diazepam‐binding inhibitor (DBI). The authors have recently shown that β‐amyloid peptide (Aβ) stimulates DBI gene expression and endozepine release. The purpose of this study was to determine the mechanism of action of Aβ in cultured rat astrocytes. Aβ25–35 and the N‐formyl peptide receptor (FPR) agonist N‐formyl‐Met‐Leu‐Phe (fMLF) increased the secretion of endozepines in a dose‐dependent manner with EC50 value of ≈2 μM. The stimulatory effects of Aβ25–35 and the FPR agonists fMLF and N‐formyl‐Met‐Met‐Met (fMMM) on endozepine release were abrogated by the FPR antagonist N‐t‐Boc‐Phe‐Leu‐Phe‐Leu‐Phe. In contrast, Aβ25–35 increased DBI mRNA expression through a FPR‐independent mechanism. Aβ25–35 induced a transient stimulation of cAMP formation and a sustained activation of polyphosphoinositide turnover. The stimulatory effect of Aβ25–35 on endozepine release was blocked by the adenylyl cyclase inhibitor somatostatin, the protein kinase A (PKA) inhibitor H89, the phospholipase C inhibitor U73122, the protein kinase C (PKC) inhibitor chelerythrine and the ATP binding cassette transporter blocker glyburide. Taken together, these data demonstrate for the first time that Aβ25–35 stimulates endozepine release from rat astrocytes through a FPR receptor positively coupled to PKA and PKC.

PACAP Stimulates Biosynthesis and Release of Endozepines from Rat Astrocytes

Abstract:  Astrocytes synthesize and release endozepines, a family of neuropeptides related to diazepam‐binding inhibitor (DBI). Astroglial cells also express the receptors of pituitary adenylate cyclase‐activating polypeptide (PACAP) and vasoactive intestinal polypeptide (VIP). In the present article, we show that PACAP dose dependently increases DBI gene expression and stimulates endozepine release through activation of PAC1‐R. PACAP increases cAMP formation, enhances polyphosphoinositide turnover, and evokes calcium mobilization from intracellular Ca2+ pools. The effect of PACAP on endozepine release is mediated through the adenylyl cyclase/PKA pathway while the downregulation of astrocyte response to PACAP can be ascribed to activation of the PLC/PKC pathway.