Urocortin 2 treatment is protective in excitotoxic retinal degeneration

Urocortin 2 (Ucn 2) is a corticotrop releasing factor paralog peptide with many physiological functions and it has widespread distribution. There are some data on the cytoprotective effects of Ucn 2, but less is known about its neuro- and retinoprotective actions. We have previously shown that Ucn 2 is protective in ischemia-induced retinal degeneration. The aim of the present study was to examine the protective potential of Ucn 2 in monosodiumglutamate (MSG)-induced retinal degeneration by routine histology and to investigate cell-type specific effects by immunohistochemistry. Rat pups received MSG applied on postnatal days 1, 5 and 9 and Ucn 2 was injected intravitreally into one eye. Retinas were processed for histology and immunocytochemistry after 3 weeks. Immunolabeling was determined for glial fibrillary acidic protein, vesicular glutamate transporter 1, protein kinase Cα, calbindin, parvalbumin and calretinin. Retinal tissue from animals treated with MSG showed severe degeneration compared to normal retinas, but intravitreal Ucn 2 treatment resulted in a retained retinal structure both at histological and neurochemical levels: distinct inner retinal layers and rescued inner retinal cells (different types of amacrine and rod bipolar cells) could be observed. These findings support the neuroprotective function of Ucn 2 in MSG-induced retinal degeneration

PACAP promotes neuron survival in early experimental diabetic retinopathy

Metabolic changes induced by diabetes lead to a multifactorial progressive disease of the retina with an extremely complex pathogenesis. One of the mechanisms of retinal cell death in diabetes is via apoptosis. Our previous results show that pituitary adenylate cyclase activating polypeptide (PACAP) attenuates the morphological and neurochemical changes in a rat model of diabetic retinopathy. The aim of this study was to investigate the mechanisms of this protective effect. Retinas of streptozotocin-induced diabetic rats were analyzed using apoptosis detection combined with immunolabeling. Western blot was used to measure levels of pro- and anti-apoptotic pathways. Intraocular PACAP injection markedly attenuated diabetic retinal injury: increased levels of the anti-apoptotic p-Akt, p-ERK1, p-ERK2, PKC, Bcl-2, while decreased levels of the pro-apoptotic p-p38MAPK and activated caspases (8, 3, 12) were detected. The number of apoptotic cells increased in all nuclear layers of diabetic retinas, but significantly decreased after PACAP treatment. Our results clearly demonstrate that the protective effects of PACAP are mediated, at least partly, by attenuating apoptosis, including also that of the dopaminergic amacrine cells. Inhibition of apoptosis is one of the PACAP-induced pathways with therapeutic potential in early experimental diabetic retinopathy

PACAP application improves functional outcome of chronic retinal ischemic injury in rats – evidence from electroretinographic measurements

Retinoprotective effects of pituitary adenylate cyclase activating polypeptide (PACAP) are well-known and have been demonstrated in various pathological conditions, such as diabetic retinopathy, excitotoxic retinal injury, UV light-induced degeneration and ischemic retinal lesion. The neuronal degeneration observed in the different retinal layers under the above pathological conditions can be successfully decreased by PACAP, however, whether this morphological improvement is also reflected in functional amelioration remains unknown. Therefore, our purpose was to investigate the protective effect of PACAP on the rat retina after bilateral common carotid artery occlusion (BCCAO) with electroretinography (ERG) to parallel the functional data with the previous morphological and neurochemical observations. Control eyes received saline treatment while PACAP was injected into the vitreous space of the other eye immediately after the induction of ischemia. Retinal damage and protective effects of PACAP were quantified by the changes in the wave forms and amplitudes. On postoperative days 2 and 14 several parameters were assessed with special attention to the changes of b wave. The results confirm that the previously described morphological protection induced by PACAP treatment is reflected in functional improvement in ischemic retinal lesions

Accelerated retinal aging in PACAP knock-out mice

Pituitary adenylate cyclase activating polypeptide (PACAP) is a neurotrophic and neuroprotective peptide. PACAP and its receptors are widely distributed in the retina. A number of reports provided evidence that PACAP is neuroprotective in retinal degenerations. The current study compared retina cell type-specific differences in young (3-4months) and aged adults (14-16months), of wild-type (WT) mice and knock-out (KO) mice lacking endogenous PACAP production during the course of aging. Histological, immunocytochemical and Western blot examinations were performed. The staining for standard neurochemical markers (tyrosine hydroxylase for dopaminergic cells, calbindin 28 kDa for horizontal cells, protein kinase Calpha for rod bipolar cells) of young adult PACAP KO retinas showed no substantial alterations compared to young adult WT retinas, except for the specific PACAP receptor (PAC1-R) staining. We could not detect PAC1-R immunoreactivity in bipolar and horizontal cells in young adult PACAP KO animals. Some other age-related changes were observed only in the PACAP KO mice only. These alterations included horizontal and rod bipolar cell dendritic sprouting into the photoreceptor layer and decreased ganglion cell number. Also, Muller glial cells showed elevated GFAP expression compared to the aging WT retinas. Furthermore, Western blot analyses revealed significant differences between the phosphorylation state of ERK1/2 and JNK in KO mice, indicating alterations in the MAPK signaling pathway. These results support the conclusion that endogenous PACAP contributes to protection against aging of the nervous system

Novel Neuroprotective Strategies in Ischemic Retinal Lesions

Retinal ischemia can be effectively modeled by permanent bilateral common carotid artery occlusion, which leads to chronic hypoperfusion-induced degeneration in the entire rat retina. The complex pathways leading to retinal cell death offer a complex approach of neuroprotective strategies. In the present review we summarize recent findings with different neuroprotective candidate molecules. We describe the protective effects of intravitreal treatment with: (i) urocortin 2; (ii) a mitochondrial ATP-sensitive K+ channel opener, diazoxide; (iii) a neurotrophic factor, pituitary adenylate cyclase activating polypeptide; and (iv) a novel poly(ADP-ribose) polymerase inhibitor (HO3089). The retinoprotective effects are demonstrated with morphological description and effects on apoptotic pathways using molecular biological techniques

Protective effects of the neuropeptides PACAP, substance P and the somatostatin analogue octreotide in retinal ischemia: a metabolomic analysis.

Ischemia is a primary cause of neuronal death in retinal diseases and the somatostatin subtype receptor 2 agonist octreotide (OCT) is known to decrease ischemia-induced retinal cell death. Using a recently optimized ex vivo mouse model of retinal ischemia, we tested the anti-ischemic potential of two additional neuropeptides, pituitary adenylate cyclase activating peptide (PACAP) and substance P (SP), and monitored the major changes occurring at the metabolic level. Metabolomics analyses were performed via fast HPLC online using a microTOF-Q MS instrument, a workflow that is increasingly becoming the gold standard in the field of metabolomics. The metabolomic approach allowed detection of the most significant alterations induced in the retina by ischemia and of the significance of the protective effects exerted by OCT, PACAP or SP. All treatments were shown to reduce ischemia-induced cell death, vascular endothelial growth factor over-expression and glutamate release. The metabolomic analysis showed that OCT and, to a lesser extent, also PACAP or SP, were able to counteract the ischemia-induced oxidative stress and to promote, with various efficacies, (i) decreased accumulation of glutamate and normalization of glutathione homeostasis; (ii) reduced build-up of a-ketoglutarate, which might serve as a substrate for the enhanced biosynthesis of glutamate in response to ischemia; (iii) reduced accumulation of peroxidized lipids and inflammatory mediators; (iv) the normalization of glycolytic fluxes and thus preventing the over-accumulation of lactate or either promoting the down-regulation of the glyoxalate anti-oxidant system; (v) a reduced metabolic shift from glycolysis towards the PPP or either a blockade at the non-oxidative phase of the PPP; and (vi) tuning down of purine metabolism. In addition, OCT seemed to stimulate nitric oxide production. None of the treatments was able to restore ATP production, although ATP reservoirs were partly replenished by OCT, PACAP or SP. These data indicate that, in addition to that of somatostatin, peptidergic systems such as those of PACAP and SP deserve attention in view of peptide-based therapies to treat ischemic retinal disorders

Protective Intestinal Effects of Pituitary Adenylate Cyclase Activating Polypeptide

Pituitary adenylate cyclase activating polypeptide (PACAP) is an endogenous neuropeptide widely distributed throughout the body, including the gastrointestinal tract. Several effects have been described in human and animal intestines. Among others, PACAP infl uences secretion of intestinal glands, blood fl ow, and smooth muscle contraction. PACAP is a well-known cytoprotective peptide with strong anti-apoptotic, anti-infl ammatory, and antioxidant effects. The present review gives an overview of the intestinal protective actions of this neuropeptide. Exogenous PACAP treatment was protective in a rat model of small bowel autotransplantation. Radioimmunoassay (RIA) analysis of the intestinal tissue showed that endogenous PACAP levels gradually decreased with longer-lasting ischemic periods, prevented by PACAP addition. PACAP counteracted deleterious effects of ischemia on oxidative stress markers and cytokines. Another series of experiments investigated the role of endogenous PACAP in intestines in PACAP knockout (KO) mice. Warm ischemia–reperfusion injury and cold preservation models showed that the lack of PACAP caused a higher vulnerability against ischemic periods. Changes were more severe in PACAP KO mice at all examined time points. This fi nding was supported by increased levels of oxidative stress markers and decreased expression of antioxidant molecules. PACAP was proven to be protective not only in ischemic but also in infl ammatory bowel diseases. A recent study showed that PACAP treatment prolonged survival of Toxoplasma gondii infected mice suffering from acute ileitis and was able to reduce the ileal expression of proinfl ammatory cytokines. We completed the present review with recent clinical results obtained in patients suffering from infl ammatory bowel diseases. It was found that PACAP levels were altered depending on the activity, type of the disease, and antibiotic therapy, suggesting its probable role in infl ammatory events of the intestine

Histological Evaluation of Diabetic Neurodegeneration in the Retina of Zucker Diabetic Fatty (ZDF) Rats

In diabetes, retinal dysfunctions exist prior to clinically detectable vasculopathy, however the pathology behind these functional deficits is still not fully established. Previously, our group published a detailed study on the retinal histopathology of type 1 diabetic (T1D) rat model, where specific alterations were detected. Although the majority of human diabetic patients have type 2 diabetes (T2D), similar studies on T2D models are practically absent. To fill this gap, we examined Zucker Diabetic Fatty (ZDF) rats - a model for T2D - by immunohistochemistry at the age of 32 weeks. Glial reactivity was observed in all diabetic specimens, accompanied by an increase in the number of microglia cells. Prominent outer segment degeneration was detectable with changes in cone opsin expression pattern, without a decrease in the number of labelled elements. The immunoreactivity of AII amacrine cells was markedly decreased and changes were detectable in the number and staining of some other amacrine cell subtypes, while most other cells examined did not show any major alterations. Overall, the retinal histology of ZDF rats shows a surprising similarity to T1D rats indicating that despite the different evolution of the disease, the neuroretinal cells affected are the same in both subtypes of diabetes

Neuropeptides, Trophic Factors, and Other Substances Providing Morphofunctional and Metabolic Protection in Experimental Models of Diabetic Retinopathy

Vision is the most important sensory modality for many species, including humans. Damage to the retina results in vision loss or even blindness. One of the most serious complications of diabetes, a disease that has seen a worldwide increase in prevalence, is diabetic retinopathy. This condition stems from consequences of pathological metabolism and develops in 75% of patients with type 1 and 50% with type 2 diabetes. The development of novel protective drugs is essential. In this review we provide a description of the disease and conclude that type 1 diabetes and type 2 diabetes lead to the same retinopathy. We evaluate existing experimental models and recent developments in finding effective compounds against this disorder. In our opinion, the best models are the long-term streptozotocin-induced diabetes and Otsuka Long-Evans Tokushima Fatty and spontaneously diabetic Torii rats, while the most promising substances are topically administered somatostatin and pigment epithelium-derived factor analogs, antivasculogenic substances, and systemic antioxidants. Future drug development should focus on these