Jaishree Jalewa

Neuroprotective and anti-apoptotic effects of liraglutide on SH-SY5Y cells exposed to methylglyoxal stress.

Glucagon‐like peptide 1 (GLP‐1) is a growth factor that has demonstrated neuroprotective properties in a range of studies. In an APPswe/PS1ΔE9 mouse model of Alzheimers disease (AD), we previously found protective effects on memory formation, synaptic plasticity, synapse survival and a reduction of amyloid synthesis and plaque load in the brain. Here, we analyse the neuroprotective properties of the GLP‐1 analogue liraglutide in human neuroblastoma cell line SH‐SY5Y during methyl glyoxal stress. We show for the first time that cell viability was enhanced by liraglutide (XTT assay) in a dose‐dependent way, while cytotoxicity (LDH assay) and apoptosis were reduced. Expression of the pro‐survival Mcl1 signaling protein was increased, as was activation of cell survival kinases Akt, MEK1/2 and the transcription factor p90RSK. Liraglutide also decreased pro‐apoptotic Bax and Bik expression. In addition, the membrane potential and the influx of calcium into the cell were enhanced by liraglutide. GLP‐1 receptor expression was also increased by the drug. The results demonstrate a range of growth factor‐related cytoprotective processes induced by liraglutide, which is currently on the market as a treatment for type 2 diabetes (Victoza®). It is also tested in clinical trials in patients with Alzheimer disease.

Neuroprotective effects of lixisenatide and liraglutide in the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine mouse model of Parkinson's disease.

Glucagon-like peptide 1 (GLP-1) is a growth factor. GLP-1 mimetics are on the market as treatments for type 2 diabetes and are well tolerated. These drugs have shown neuroprotective properties in animal models of neurodegenerative disorders. In addition, the GLP-1 mimetic exendin-4 has shown protective effects in animal models of Parkinsons disease (PD), and a clinical trial in PD patients showed promising first results. Liraglutide and lixisenatide are two newer GLP-1 mimetics which have a longer biological half-life than exendin-4. We previously showed that these drugs have neuroprotective properties in an animal model of Alzheimers disease. Here we demonstrate the neuroprotective effects in the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) mouse model of PD. MPTP was injected once-daily (20mg/kg i.p.) for 7 days, and drugs were injected once-daily for 14 days i.p. When comparing exendin-4 (10 nmol/kg), liraglutide (25 nmol/kg) and lixisenatide (10 nmol/kg), it was found that exendin-4 showed no protective effects at the dose chosen. Both liraglutide and lixisenatide showed effects in preventing the MPTP-induced motor impairment (Rotarod, open-field locomotion, catalepsy test), reduction in tyrosine hydroxylase (TH) levels (dopamine synthesis) in the substantia nigra and basal ganglia, a reduction of the pro-apoptotic signaling molecule BAX and an increase in the anti-apoptotic signaling molecule B-cell lymphoma-2. The results demonstrate that in this study, both liraglutide and lixisenatide are superior to exendin-4, and both drugs show promise as a novel treatment of PD.

Novel incretin analogues improve autophagy and protect from mitochondrial stress induced by rotenone in SH-SY5Y cells

Currently, there is no viable treatment available for Parkinsons disease (PD) that stops or reverses disease progression. Interestingly, studies testing the glucagon‐like‐peptide‐1 (GLP‐1) mimetic Exendin‐4 have shown neuroprotective/neurorestorative properties in pre‐clinical tests and in a pilot clinical study of PD. Incretin analogues were originally developed to treat type 2 diabetes and several are currently on the market. In this study, we tested novel incretin analogues on the dopaminergic SH‐SY5Y neuroblastoma cells against a toxic mitochondrial complex I inhibitor, Rotenone. Here, we investigate for the first time the effects of six different incretin receptor agonists – Liraglutide, D‐Ser2‐Oxyntomodulin, a GLP‐1/GIP Dual receptor agonist, dAla(2)‐GIP‐GluPal, Val(8)GLP‐1‐GluPal and exendin‐4. Post‐treatment with doses of 1, 10 or 100 nM of incretin analogues for 12 h increased the survival of SH‐SY5Y cells treated with 1 μM Rotenone for 12 h. Furthermore, we studied the post‐treatment effect of 100 nM incretin analogues against 1 μM Rotenone stress on apoptosis, mitochondrial stress and autophagy markers. We found significant protective effects of the analogues against Rotenone stress on cell survival and on mitochondrial and autophagy‐associated markers. The novel GLP‐1/GIP Dual receptor agonist was superior and effective at a tenfold lower concentration compared to the other analogues. Using the Phosphatidylinositol 3‐kinase (PI3K) inhibitor, LY294002, we further show that the neuroprotective effects are partially PI3K‐independent. Our data suggest that the neuroprotective properties exhibited by incretin analogues against Rotenone stress involve enhanced autophagy, increased Akt‐mediated cell survival and amelioration of mitochondrial dysfunction. These mechanisms can explain the neuroprotective effects of incretin analogues reported in clinical trials.

Prophylactic liraglutide treatment prevents amyloid plaque deposition, chronic inflammation and memory impairment in APP/PS1 mice

Type 2 diabetes is a risk factor for Alzheimers disease (AD). Previously, we have shown that the diabetes drug liraglutide is protective in middle aged and in old APP/PS1 mice. Here, we show that liraglutide has prophylactic properties. When injecting liraglutide once-daily ip. in two months old mice for 8 months, the main hallmarks of AD were much reduced. Memory formation in object recognition and Morris water maze were normalised and synapse loss and the loss of synaptic plasticity was prevented. In addition, amyloid plaque load, including dense core congophilic plaques, was much reduced. Chronic inflammation (activated microglia) was also reduced in the cortex, and neurogenesis was enhanced in the dentate gyrus. The results demonstrate that liraglutide may protect from progressive neurodegeneration that develops in AD. The drug is currently in clinical trials in patients with AD.

Increased number of orexin/hypocretin neurons with high and prolonged external stress-induced depression.

It has been found that dysregulation in the orexin/hypocretin (Ox/HCRT) neuropeptide system in the lateral hypothalamus (LHA) is known to affect sleep disorder, depression and motor activities. However, to date there is no common agreement regarding the resulting specific changes induced in the Ox system. In this study, we inject corticosterone to produce stress-induced depressed mice and investigate the Ox neuronal and corresponding behavioural changes. Different doses (10, 20, 50mg/kgbw) of corticosterone were injected in adult mice, and then were tested in the open field test, forced swim test, tail suspension test, elevated plus maze test and motor activity measurements to validate the depressed animal model. Significant dose-dependent behavioural changes were observed in correlation with the doses of corticosterone. The effect is most significant and robust in the high 50mg/kgbw dose group five weeks after injection. Interestingly, we found on average a reduction in motor activity during the 12-hour dark phase (awake) of the depressed mice and no significant change during the light phase (asleep). Finally, using confocal microscopy, immunofluorescence (IF) analysis shows a significant increase (∼20%) in the number of Ox neurons in the LHA of the depressed mice as compared to the age-matched controls. This study suggests that an increase in Ox neuronal signaling may be functionally linked to high and prolonged external stress-induced depression.

Neural Circuit Interactions between the Dorsal Raphe Nucleus and the Lateral Hypothalamus: An Experimental and Computational Study

Orexinergic/hypocretinergic (Ox) neurotransmission plays an important role in regulating sleep, as well as in anxiety and depression, for which the serotonergic (5-HT) system is also involved in. However, little is known regarding the direct and indirect interactions between 5-HT in the dorsal raphe nucleus (DRN) and Ox neurons in the lateral hypothalamus (LHA). In this study, we report the additional presence of 5-HT1BR, 5-HT2AR, 5-HT2CR and fast ligand-gated 5-HT3AR subtypes on the Ox neurons of transgenic Ox-enhanced green fluorescent protein (Ox-EGFP) and wild type C57Bl/6 mice using single and double immunofluorescence (IF) staining, respectively, and quantify the colocalization for each 5-HT receptor subtype. We further reveal the presence of 5-HT3AR and 5-HT1AR on GABAergic neurons in LHA. We also identify NMDAR1, OX1R and OX2R on Ox neurons, but none on adjacent GABAergic neurons. This suggests a one-way relationship between LHA’s GABAergic and Ox neurons, wherein GABAergic neurons exerts an inhibitory effect on Ox neurons under partial DRN’s 5-HT control. We also show that Ox axonal projections receive glutamatergic (PSD-95 immunopositive) and GABAergic (Gephyrin immunopositive) inputs in the DRN. We consider these and other available findings into our computational model to explore possible effects of neural circuit connection types and timescales on the DRN-LHA system’s dynamics. We find that if the connections from 5-HT to LHA’s GABAergic neurons are weakly excitatory or inhibitory, the network exhibits slow oscillations; not observed when the connection is strongly excitatory. Furthermore, if Ox directly excites 5-HT neurons at a fast timescale, phasic Ox activation can lead to an increase in 5-HT activity; no significant effect with slower timescale. Overall, our experimental and computational approaches provide insights towards a more complete understanding of the complex relationship between 5-HT in the DRN and Ox in the LHA.

A novel GLP-1/GIP dual receptor agonist protects from 6-OHDA lesion in a rat model of Parkinson's disease.

&NA; The incretins glucagon‐like peptide 1 (GLP‐1) and glucose dependent insulinotropic polypeptide (GIP) are growth factors that have shown neuroprotective effects in animal models of Parkinsons and Alzheimers disease. In addition, the GLP‐1 mimetic exendin‐4 has shown protective effects in a clinical trial in Parkinsons disease (PD) patients. GLP‐1 analogues are currently on the market as treatments for type II diabetes. We previously showed that the novel dual agonist (DA‐JC1) was effective in the 1‐methyl‐4‐phenyl‐1,2,3,6‐tetrahydropyridine (MPTP) mouse model of PD. Here we demonstrate that DA‐JC1 is neuroprotective in the 6‐OHDA brain lesion rat model of PD. When treating rats for 6 weeks with DA‐JC1 at 25 nmol/kg ip once‐daily, motor activity as tested in the Rotarod and in the open field was much improved. In the amphetamine and apomorphine circling behaviour tests, the 6‐OHDA induced impairments were much reduced by the DA‐JC1 treatment. The number of TH positive dopaminergic neurons in the substantia nigra was decreased by 6‐OHDA lesion and was increased by DA‐JC1 treatment. Dopamine levels in the basal ganglia were reduced by 6‐OHDA lesion and increased by DA‐JC1. In western blot analysis, levels of the growth factor GDNF and pAkt/CREB cell signaling was enhanced by DA‐JC1. The autophagy marker Beclin1 was also activated by the drug. The results demonstrate that dual GLP‐1/GIP receptor agonists show promise as a novel treatment for PD. HighlightsGLP‐1 incretin analogues are on the market as treatments for diabetes.First clinical trials show good effects in patients with Parkinsons disease.We show that a novel GLP‐1/GIP analogue shows good effects in a rat model of PD.Motor activity is protected, GDNF levels and Akt/CREB signaling is increased by the drug.Autophagy was also enhanced by the novel dual agonist.

Neuroprotective effects of an oxyntomodulin analogue in the MPTP mouse model of Parkinson's disease

Oxyntomodulin is a hormone and a growth factor. It activates two receptors, the Glucagon-like peptide 1 (GLP-1) and the glucagon receptor. GLP-1 mimetics are on the market as treatments for type 2 diabetes and are well tolerated. These drugs have shown neuroprotective properties in animal models of neurodegenerative disorders. In addition, the GLP-1 mimetic exendin-4 has shown protective effects in animal models of Parkinsons disease (PD), and a clinical trial in PD patients showed promising first positive results. D-Ser2-oxyntomodulin (Oxy) is a protease resistant oxyntomodulin analogue that has been developed to treat diabetes. Here we demonstrate for the first time that such analogues have neuroprotective effects. The drug showed protective effects in the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) mouse model of PD. MPTP was injected daily (20 mg/kg i.p.) for 7 days, and Oxy injected once-daily for 14 days i.p. Oxy treatment prevented or reversed the MPTP- induced motor impairment (Rotarod, spontaneous locomotion, swim activity, muscle strength test), the MPTP-induced reduction in Tyrosine Hydroxylase (TH) levels (dopamine synthesis) in the substantia nigra and basal ganglia, the reduction of the synaptic marker synapstophysin, the inactivation of the growth factor kinase Akt/PKB and of the anti-apoptotic signaling molecule Bcl-2, and the increase of levels of the pro-inflammatory cytokine TNF-α. The results demonstrate that oxyntomodulin analogues show promise as a novel treatment of PD.