The Difference Between The Response To Glutamate Excitotoxicity and The Role Of Ca2+ Channel Blockers in Cortical Neuron and SH-SY5Y Cells Cultures

Year 2022, Volume: 7 Issue: 1, 45 - 52, 31.03.2022

Betül Çiçek , Ali Taghizadehghalehjoughi , Ahmet Hacımüftüoğlu , Aysegul Yılmaz

https://doi.org/10.35229/jaes.1003400

Abstract

Cortical neuron and SH-SY5Y cells are widely used in glutamate excitotoxicity studies, but it is unclear which one better reflects this model. Generally, glutamate induces toxicity conditions by leading to L and L/N-Ca2+channels activation and cell death via lethal Ca2+ influx. To evaluate this hypothesis, the effects of L and L/N-Ca2+ channel blockers, lacidipine, and amlodipine under excitotoxic conditions were evaluated. At the same time, in this study, we aimed to determine that these two cell lines better reflect this model. To induce excitotoxicity, cortical neuron and SH-SY5Y cells were incubated with glutamate 10-5 mM. After 30 min incubation with glutamate, different concentration (1, 2 and 4 µg lacidipine and 20, 50 and 100 µM amlodipine) were applied these cells. Possible neuroprotective roles of lacidipine and amlodipine were investigated through cell viability, oxidative stress, and apoptotic alterations. Our results showed that SH-SY5Y cells are the more ideal cell line for oxidative stress-mediated glutamate toxicity. In addition, 4 µg lacidipine and 100 µM amlodipine had significant neuroprotective roles in these cells, but the most protective effect was also determined in SH-SY5Y cells at 100 µM amlodipine. The highest viability rate on cell lines was found at 88,8 % in SH-SY5Y cells treated with 100 μM amlodipine. Results from the TAC, TOS, LDH assays, and flow cytometry analysis were correlated to our MTT results. Taken together, our results indicate that SH-SY5Y cells are more effective at reflecting glutamate-induced excitotoxicity and 100μM amlodipine has a more protective effect in treating this toxicity.

Keywords

cortical neuron, glutamate excitotoxicity, Ca2+, SH-SY5Y, lacidipine, amlodipine

Kortikal Nöron ve SH-SY5Y Hücre Kültürlerinde Glutamat Eksitotoksisitesine Yanıt ile Ca2+ Kanal Blokerlerinin Rolü Arasındaki Fark

Abstract

Kortikal nöron ve SH-SY5Y hücreleri glutamat eksitotoksisite çalışmalarında yaygın olarak kullanılmaktadır, ancak hangisinin bu modeli daha iyi yansıttığı belirsizdir. Genel olarak glutamat, L ve L/N-Ca2+ kanallarının aktivasyonuna ve öldürücü Ca2+ akışı yoluyla hücre ölümüne yol açarak toksisite koşullarını indükler. Bu hipotezi değerlendirmek için eksitotoksik koşullar altında L ve L/N-Ca2+ kanal blokerleri, lasidipin ve amlodipinin etkileri değerlendirildi. Aynı zamanda bu çalışmada bu iki hücre hattının bu modeli daha iyi yansıttığını belirlemeyi amaçladık. Eksitotoksisiteyi indüklemek için kortikal nöron ve SH-SY5Y hücreleri glutamat 10-5 mM mM ile inkübe edildi. Glutamat ile 30 dakikalık inkübasyondan sonra, bu hücrelere farklı konsantrasyonlar (1, 2 ve 4 µg lasidipin ve 20, 50 ve 100 μM amlodipin) uygulandı.Lasidipin ve amlodipinin olası nöroprotektif rolleri, hücre canlılığı, oksidatif stres ve apoptotik değişiklikler yoluyla araştırıldı. Sonuçlarımız SH-SY5Y hücrelerinin oksidatif stres aracılı glutamat toksisitesini daha iyi yansıttığını gösterdi. 4 µg lasidipin ve 100 μM amlodipin bu hücrelerde önemli nöroprotektif rollere sahip olmasına rağmen en yüksek koruyucu etki 100 μM amlodipin'de SH-SY5Y hücrelerinde de belirlendi. En yüksek canlılık oranı, 100 μM amlodipin ile tedavi edilen SH-SY5Y hücrelerinde %88,8 olarak bulundu.TAC, TOS, LDH tahlilleri ve flow sitometrik analizinden elde edilen sonuçlar MTT sonuçlarımızla benzerdi. Tüm sonuçlar birlikte değerlendirildiğinde SH-SY5Y hücrelerinin glutamat kaynaklı eksitotoksisiteyi yansıtmada daha etkili hücre hattı olduğunu ve 100 μM amlodipinin bu toksisitenin tedavisinde daha koruyucu bir etkiye sahip olduğunu göstermektedir.

Keywords

kortikal nöron, SH-SY5Y, glutamat eksitotoksisitesi, Ca2+, lacidipin, amlodipin

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