Neural Regeneration Research ›› 2020, Vol. 15 ›› Issue (4): 712-723.doi: 10.4103/1673-5374.266916

Previous Articles     Next Articles

Long-term adenosine A1 receptor activation-induced sortilin expression promotes α-synuclein upregulation in dopaminergic neurons

Yun-Cheng Lv1, 2, An-Bo Gao1, 3, Jing Yang4, Li-Yuan Zhong1, Bo Jia1, Shu-Hui Ouyang1, Le Gui2, Tian-Hong Peng1, Sha Sun1,Francisco S. Cayabyab2#br#   

  1. 1 Clinical Anatomy & Reproductive Medicine Application Institute, Hengyang Medical College, University of South China, Hengyang, Hunan Province, China
    2 Department of Surgery, College of Medicine, University of Saskatchewan, Saskatoon, Canada
    3 Institute of Pharmacy and Pharmacology, School of Pharmaceutical Science, University of South China, Hengyang, Hunan Province, China
    4 Department of Metabolism & Endocrinology, the First Affiliated Hospital, Hengyang Medical College, University of South China, Hengyang, Hunan Province, China
  • Online:2020-04-15 Published:2020-05-29
  • Contact: Tian-Hong Peng, PhD,thpeng67@163.com; Sha Sun, BS,1060565208@qq.com; Francisco S. Cayabyab, PhD,frank.cayabyab@usash.ca.
  • Supported by:
    This study was supported by the National Natural Sciences Foundation of China, No. 81770460 (to YCL); the Postdoctoral Research Fellowship of the Saskatchewan Health Research Foundation, No. SHRF, 4144 (to YCL); the third level of the Chuanshan Talent project of the University of South China, No. 2017CST20 (to YCL); the Aid Program, No. 2017KJ268 and the Key Lab for Clinical Anatomy & Reproductive Medicine, No. 2017KJ182 from the Science and Technology Bureau of Hengyang City, China (to YCL and XC); the Postgraduate Student Research Innovation Projects of Hunan Province, China, No. CX2018B62 (to ABG).

PDF

192

Abstract:

Prolonged activation of adenosine A1 receptor likely leads to damage of dopaminergic neurons and subsequent development of neurodegenerative diseases. However, the pathogenesis underlying long-term adenosine A1 receptor activation-induced neurodegeneration remains unclear. In this study, rats were intraperitoneally injected with 5 mg/kg of the adenosine A1 receptor agonist N6-cyclopentyladenosine (CPA) for five weeks. The mobility of rats was evaluated by forced swimming test, while their cognitive capabilities were evaluated by Y-maze test. Expression of sortilin, α-synuclein, p-JUN, and c-JUN proteins in the substantia nigra were detected by western blot analysis. In addition, immunofluorescence staining of sortilin and α-synuclein was performed to detect expression in the substantia nigra. The results showed that, compared with adenosine A1 receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine (5 mg/kg) + CPA co-treated rats, motor and memory abilities were reduced, surface expression of sortin and α-synuclein in dopaminergic neurons was reduced, and total sortilin and total α-synuclein were increased in CPA-treated rats. MN9D cells were incubated with 500 nM CPA alone or in combination with 10 μM SP600125 (JNK inhibitor) for 48 hours. Quantitative real-time polymerase chain reaction analysis of sortilin and α-synuclein mRNA levels in MN9D cells revealed upregulated sortilin expression in MN9D cells cultured with CPA alone, but the combination of CPA and SP600125 could inhibit this expression. Predictions made using Jasper, PROMO, and Alibaba online databases identified a highly conserved sequence in the sortilin promoter that was predicted to bind JUN in both humans and rodents. A luciferase reporter assay of sortilin promoter plasmid-transfected HEK293T cells confirmed this prediction. After sortilin expression was inhibited by sh-SORT1, expression of p-JUN and c-JUN was detected by western blot analysis. Long-term adenosine A1 receptor activation levels upregulated α-synuclein expression at the post-transcriptional level by affecting sortilin expression. The online tool Raptor-X-Binding and Discovery Studio 4.5 prediction software predicted that sortilin can bind to α-synuclein. Co-immunoprecipitation revealed an interaction between sortilin and α-synuclein in MN9D cells. Our findings indicate that suppression of prolonged adenosine A1 receptor activation potently inhibited sortilin expression and α-synuclein accumulation, and dramatically improved host cognition and kineticism. This study was approved by the University Committee of Animal Care and Supply at the University of Saskatchewan (approval No. AUP#20070090) in March 2007, and the Animals Ethics Committee of University of South China (approval No. LL0387-USC) in June 2017.

Key words: cognitive dysfunction, dopaminergic neuron, dyskinesia, JNK/c-JUN pathway, long-term adenosine A1 receptor activation, neural regeneration, neurodegenerative diseases, sortilin, α-synuclein