EFFECTS OF ERYTHROPOIETIN ON NITRIC OXIDE SYNTHASE TYPES IN THE HIPPOCAMPUS AND FRONTAL CORTEX IN PTZ-INDUCED SEIZURES IN RATS

Öz

Objective: Nitric oxide, an important mediator in the dysfunctions of learning-memory and epileptogenesis, is formed by nitric oxide synthase (NOS) in neuronal cells in many areas of the brain, primarily the hippocampus. It is has been shown that erythropoietin (EPO) has neuroprotective and antiepileptic effects. The aims of this study was to investigate the effects of EPO pre-treatment on convulsions and NOS species in the hippocampus and frontal cortex in pentylenetetrazole (PTZ)-induced generalized seizure model. Material and Method: Forty adult male Wistar albino rats were divided into four groups: Control-saline; PTZ-single dose 60 mg/ kg; EPO-3000 IU/kg; and EPO+PTZ- EPO pretreatment 24 hours before PTZ administration. After the administration of PTZ, the seizure severity (stage) was observed and scored. The levels of neuronal NOS (nNOS), endothelial NOS (eNOS), inducible NOS (iNOS) levels in the plasma, hippocampus and frontal cortex specimens of animals and expressions of NOS species in the hippocampus tissues were examined. Results: EPO pretreatment decreased seizure stage (p<0.01). EPO administration alone reduced nNOS levels in plasma (p<0.001), frontal cortex (p<0.05), hippocampus (p<0.001). Hipocampus nNOS level and expression increased in the EPO+PTZ group compared to the PTZ group (p<0.001). eNOS (p<0.01) and iNOS (p<0.05) levels of hippocampus decreased in the PTZ group; EPO pretreatment before PTZ application improved nNOS level. Conclusion: The EPO pre-treatment demonstrated anticonvulsive effects by increasing eNOS and nNOS levels. EPO pre-treatment can reduce the hyperexcitability of neurons by increasing blood flow in the hypocampus via eNOS. In addition, EPO may contribute to its anticonvulsive effects by inhibiting the decrease of eNOS and nNOS by acting as antioxidant.

Anahtar Kelimeler

• nitric oxide synthase,hippocampus,seizure,erythropoietin,seizure

SIÇANLARDA PTZ-KAYNAKLI NÖBETLERDE ERİTROPOİETİNİN HİPOKAMPUS VE FRONTAL KORTEKSTE NİTRİK OKSİT SENTAZ TÜRLERİ ÜZERİNE ETKİLERİ

Öz

Amaç: Öğrenme-hafıza bozukluklarında ve epileptogenezde önemli bir medyatör olan nitrik oksit, hipokampus başta olmak üzere beynin birçok bölgesinde nöronal hücrelerde nitrik oksit sentaz (NOS) tarafından oluşturulur. Eritropoietin (EPO)’nun nöroprotektif ve antiepileptik etkileri olduğu gösterilmiştir. EPO ön-uygulamasının, pentilentetrazol (PTZ) ile indüklenen jeneralize nöbet modelinde konvulsiyonlar ve hipokampus ve frontal kortekste NOS türleri üzerine etkilerini araştırmayı amaçladık. Gereç ve Yöntem: Wistar albino 40 erkek sıçan 4 gruba ayrıldı: kontrol, PTZ-tek doz 60 mg/kg; EPO-3000 IU/kg; EPO+PTZ grubu-PTZ uygulanmasından 24 saat önce EPO ön-uygulaması yapılan grup. PTZ uygulamasından sonra, nöbet şiddeti gözlendi ve skorlandı. Sıçanların plazma, hipokampus ve frontal korteks örneklerinde nöronal NOS (nNOS), endotelyal NOS (eNOS), indüklenebilir NOS (iNOS) düzeyleri ve hipokampusde NOS türlerinin anlatımları değerlendirildi. Bulgular: EPO ön-uygulaması nöbet şiddetini azalttı. Tek başına EPO uygulaması nNOS düzeyini plazma (p<0,001), frontal korteks (p<0,05) ve hipokampus (p<0,001) örneklerinde azalttı. Hipokampus nNOS düzeyi ve anlatımı, EPO+PTZ grubunda, PTZ grubuna göre arttı (p<0,001). PTZ uygulamasıyla hipokampusde eNOS (p<0,01) ve iNOS (p<0,05) azalırken; EPO ön-uygulamasıyla kontrole yakın olduğu gözlendi. Sonuç: EPO ön-uygulaması antikonvulsif etkilerini, eNOS ve nNOS düzeylerini artırarak gösterdi. EPO ön-uygulaması, eNOS aracılığıyla hipokapusde kan akımını artırarak, nöronların hipereksitabilitesini azaltabilir. Ayrıca EPO, antioksidan görevi görerek eNOS ve nNOS’un azalmasını engelleyerek antikonvulzif etkilerine katkıda bulunabilir.

Anahtar Kelimeler

• eritropoietin,nitrik oksit sentaz,hipokampus,nöbet

Kaynakça

1. 1. Palabıyık M. Kronik levetirasetam uygulanan normal ve temporal lob epilepsi modeli oluşturulmuş sıçanlarda aromataz ekspresyonu değişiklikleri. 2015 Ankara Hacettepe Üniversitesi Sağlık Bilimleri Enstitüsü Doktora Tezi.
2. 2. Shorvon SD. Drug treatment of epilepsy in the century of the ILAE: the second 50 years, 1959–2009. Epilepsia 2009:50(s3);93-130.
3. 3. Watanabe M, Miyai A, Danjo S, Nakamura Y, Itoh K. The threshold of pentylenetetrazole-induced convulsive seizures, but not that of nonconvulsive seizures, is controlled by the nitric oxide levels in murine brains. Exp Neurol 2013:247;645-52.
4. 4. Brown GC. Nitric oxide and neuronal death. Nitric oxide 2010: 23(3);153-65.
5. 5. Samland H, Huitron-Resendiz S, Masliah E, Criado J, Henriksen SJ, Campbell IL. Profound increase in sensitivity to glutamatergic-but not cholinergic agonist-induced seizures in transgenic mice with astrocyte production of IL-6. J Neurosci Res 2003:73(2);176-87.
6. 6. Toda N, Ayajiki K, Okamura T. Cerebral blood flow regulation by nitric oxide: recent advances. Pharmacol Rev 2009:61(1);62-97.
7. 7. Toda N, Okamura T. The pharmacology of nitric oxide in the peripheral nervous system of blood vessels. Pharmacol Rev 2003:55(2);271-324.
8. 8. Kim SY, Buckwalter M, Soreq H, Vezzani A, Kaufer D. Blood–brain barrier dysfunction–induced inflammatory signaling in brain pathology and epileptogenesis. Epilepsia 2012:53(s6);37-44.

9. 9. Vezzani A, Balosso S, Ravizza T. Inflammation and epilepsy. Handb Clin Neurol. 2012:107;163-75.
10. 10. Vezzani A, Auvin S, Ravizza T, Aronica E. Glia-neuronal interactions in ictogenesis and epileptogenesis: role of inflammatory mediators. In: Noebels JL, Avoli M, Rogawski MA, et al, editors. Jasper’s Basic Mechanisms of the Epilepsies [Internet]. 4th edition. Bethesda (MD): National Center for Biotechnology Information (US); 2012.
11. 11. İzci Y, Erbaş YC. Hippocampus: Its Structure and Functions, Türk Nöroşir Derg, 2015:25:3;287-95.
12. 12. Nakatomi H, Kuriu T, Okabe S, Yamamoto SI, Hatano O, Kawahara N, Nakafuku M. Regeneration of hippocampal pyramidal neurons after ischemic brain injury by recruitment of endogenous neural progenitors. Cell 2002:110(4);429-41.
13. 13. Nadler JV, Perry BW, Cotman CW. Intraventricular kainic acid preferentially destroys hippocampal pyramidal cells. Nature 1978:271(5646);676.
14. 14. Lehmann TN, Gabriel S, Kovacs R, Eilers A, Kivi A, Schulze K et al. Alterations of neuronal connectivity in area CA1 of hippocampal slices from temporal lobe epilepsy patients and from pilocarpine-treated epileptic rats. Epilepsia 2000:41(6);S190-4.
15. 15. Suzuki M, Hagino H, Nohara S, Zhou SY, Kawasaki Y, Takahashi T, Kurachi M. Male-specific volume expansion of the human hippocampus during adolescence. Cereb Cortex 2004:15(2);187-93.
16. 16. Stretton J, Thompson PJ. Frontal lobe function in temporal lobe epilepsy. Epilepsy Res 2012:98(1);1-13.
17. 17. Cave CB, Squire LR. Intact verbal and nonverbal short-term memory following damage to the human hippocampus. Hippocampus 1992:2(2);151-63.
18. 18. Chapados C, Petrides M. Ventrolateral and dorsomedial frontal cortex lesions impair mnemonic context retrieval. Proc Biol Sci 2015:282(1801);20142555.
19. 19. Erbaş O, Çınar BP, Solmaz V, Çavuşoğlu T, Ateş U. The neuroprotective effect of erythropoietin on experimental Parkinson model in rats. Neuropeptides 2015:49;1-5.
20. 20. Gu L, Xu H, Wang F, Xu G, Sinha D, Wang J, Lu L. Erythropoietin exerts a neuroprotective function against glutamate neurotoxicity in experimental diabetic retina. Invest Ophthalmol Vis Sci 2014:55(12);8208-22.
21. 21. Bahçekapılı N, Akgün-Dar K, Albeniz I, Kapucu A., Kandil A., Yağız O, Üzüm G. Erythropoietin pretreatment suppresses seizures and prevents the increase in inflammatory mediators during pentylenetetrazole-induced generalized seizures. Int J Neurosci 2014:124(10);762-70.
22. 22. Genc S, Kuralay F, Genc K, Akhisaroglu M, Fadiloglu S, Yorukoglu K, Gure A. Erythropoietin exerts neuroprotection in 1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine-treated C57/BL mice via increasing nitric oxide production. Neurosci Lett 2001:298(2);139-41.
23. 23. Chen ZY, Wang L, Asavaritkrai P, Noguchi CT. Up-regulation of erythropoietin receptor by nitric oxide mediates hypoxia preconditioning. J Neurosci Res 2010:88(14);3180-8.
24. 24. Tian J, Kim SF, Hester L, Snyder SH. S-nitrosylation/ activation of COX-2 mediates NMDA neurotoxicity. Proc Natl Acad Sci 2008:105(30);10537-40.
25. 25. Keswani SC, Bosch-Marcé M, Reed N, Fischer A, Semenza GL, Höke A. Nitric oxide prevents axonal degeneration by inducing HIF-1–dependent expression of erythropoietin. Proc Natl Acad Sci 2011:108(12);4986-90.
26. 26. Homayoun H, Khavandgar S, Dehpour AR. Anticonvulsant effects of cyclosporin A on pentylentetrazol-induced seizure and kindling: modulation by nitricoxidergic system. Brain Res 2002;939:1-10.
27. 27. Lüscher C, Malenka RC. NMDA receptor-dependent long-term potentiation and long-term depression (LTP/ LTD). Cold Spring Harb Perspect Biol 2012:4(6);a005710.
28. 28. Itoh K, Watanabe M. Paradoxical facilitation of pentylenetetrazole-induced convulsion susceptibility in mice lacking neuronal nitric oxide synthase. Neuroscience 2009:159;735-43.
29. 29. Gotti S, Sica M, Viglietti-Panzica C, Panzica G. Distribution of nitric oxide synthase immunoreactivity in the mouse brain. Microsc Res Tech 2005:68(1);13-35.
30. 30. Sandoval R, Gonzalez A, Caviedes A, Pancetti F, Smalla KH, Kaehne T, Michea L, Gundelfinger ED, Wyneken U. Homeostatic NMDA receptor down-regulation via brain derived neurotrophic factor and nitric oxide-dependent signalling in cortical but not in hippocampal neurons. J Neurochem 2011:118;760-72.
31. 31. Chuang YC, Chen SD, Lin TK, Liou CW, Chang WN, Chan SH, Chang AY. Upregulation of nitric oxide synthase II contributes to apoptotic cell death in the hippocampal CA3 subfield via a cytochrome c/caspase-3 signaling cascade following induction of experimental temporal lobe status epilepticus in the rat. Neuropharmacology 2007:52;126373.
32. 32. Kovacs R, Rabanus A, Otahal J, Patzak A, Kardos J, Albus K, Heinemann U, Kann O. Endogenous nitric oxide is a key promoting factor for initiation of seizure-like events in hippocampal and entorhinal cortex slices. J Neurosci 2009:29;8565-77.
33. 33. Garry PS, Ezra M, Rowland MJ, Westbrook J, Pattinson KTS. The role of the nitric oxide pathway in brain injury and its treatment—from bench to bedside. Exp Neurol 2015:263;235-43.
34. 34. Standaert DG. NMDA receptors and nitric oxide synthase. Nature 1999:4;13-14.
35. 35. Brown GC, Borutaite V. Nitric oxide, mitochondria, and cell death. IUBMB life 2001:52(3-5);189-95.
36. 36. Kilic U, Kilic E, Soliz J, Bassetti CI, Gassmann M, Hermann DM. Erythropoietin protects from axotomy-induced degeneration of retinal ganglion cells by activating ERK-1/2. FASEB J 2005:19(2);249-51.
37. 37. Abdallah DM. Genetics- Anticonvulsant potential of the peroxisome proliferator-activated receptor gamma agonist pioglitazone in pentylenetetrazole-induced acute seizures and kindling in mice. In: Acton QA, editor. Epilepsy: New Insights for the Healthcare Professional. Atlanta; 2011. p.72.
38. 38. Nguyen AQ, Cherry BH, Scott GF, Ryou MG, Mallet RT. Erythropoietin: powerful protection of ischemic and post-ischemic brain. Exp Biol Med 2014:239(11);1461-75.