title: Overexpression of melatonin membrane receptors increases calcium-binding proteins and protects VSC4.1 motoneurons from glutamate toxicity through multiple mechanisms.
abstract: Melatonin has shown particular promise as a neuroprotective agent to prevent motoneuron death in animal models of both amyotrophic lateral sclerosis (ALS) and spinal cord injuries (SCI). However, an understanding of the roles of endogenous melatonin receptors including MT1, MT2, and orphan G-protein receptor 50 (GPR50) in neuroprotection is lacking. To address this deficiency, we utilized plasmids for transfection and overexpression of individual melatonin receptors in the ventral spinal cord 4.1 (VSC4.1) motoneuron cell line. Receptor-mediated cytoprotection following exposure to glutamate at a toxic level (25 μm) was determined by assessing cell viability, apoptosis, and intracellular free Ca(2+) levels. Our findings indicate a novel role for MT1 and MT2 for increasing expression of the calcium-binding proteins calbindin D28K and parvalbumin. Increased levels of calbindin D28K and parvalbumin in VSC4.1 cells overexpressing MT1 and MT2 were associated with cytoprotective effects including inhibition of proapoptotic signaling, downregulation of inflammatory factors, and expression of prosurvival markers. Interestingly, the neuroprotective effects conferred by overexpression of MT1 and/or MT2 were also associated with increases in the estrogen receptor β (ERβ): estrogen receptor α (ERα) ratio and upregulation of angiogenic factors. GPR50 did not exhibit cytoprotective effects. To further confirm the involvement of the melatonin receptors, we silenced both MT1 and MT2 in VSC4.1 cells using RNA interference technology. Knockdown of MT1 and MT2 led to an increase in glutamate toxicity, which was only partially reversed by melatonin treatment. Taken together, our findings suggest that the neuroprotection against glutamate toxicity exhibited by melatonin may depend on MT1 and MT2 but not GPR50.
authors: Division of Neurology, Department of Neurosciences, Medical University of South Carolina, Charleston, SC, USA Department of Cellular and Structural Biology, University of Texas, San Antonio, TX, USA Department of Pathology, Microbiology, and Immunology, University of South Carolina School of Medicine, Columbia, SC, USA.
authors: Arabinda
authors: A
authors: Das
authors: Gerald
authors: G
authors: Wallace
authors: Russel J
authors: RJ
authors: Reiter
authors: Abhay K
authors: AK
authors: Varma
authors: Swapan K
authors: SK
authors: Ray
authors: Naren L
authors: NL
authors: Banik
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D064026:Calbindins / Q000096:biosynthesis; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D055785:Gene Knockdown Techniques; D018698:Glutamic Acid / Q000633:toxicity*; D051379:Mice; D009046:Motor Neurons / Q000187:drug effects* / Q000378:metabolism; D009419:Nerve Tissue Proteins / Q000187:drug effects; D018696:Neuroprotective Agents / Q000494:pharmacology; D010320:Parvalbumins / Q000096:biosynthesis; D034741:RNA, Small Interfering / Q000494:pharmacology; D044122:Receptor, Melatonin, MT1 / Q000235:genetics; D044123:Receptor, Melatonin, MT2 / Q000235:genetics; D043562:Receptors, G-Protein-Coupled / Q000187:drug effects; D044094:Receptors, Melatonin / Q000096:biosynthesis*
chemical_list: D064026:Calbindins; C509755:Gpr50 receptor, mouse; D009419:Nerve Tissue Proteins; D018696:Neuroprotective Agents; D010320:Parvalbumins; D034741:RNA, Small Interfering; D044122:Receptor, Melatonin, MT1; D044123:Receptor, Melatonin, MT2; D043562:Receptors, G-Protein-Coupled; D044094:Receptors, Melatonin; D018698:Glutamic Acid