P2-159: Melatonin meliorates calyculin A-induced axonal transport defects and spatial memory retention impairment in rats

Abstract

We have recently found that melatonin protects SH-SY5Y neuroblastoma cells from calyculin A-induced neurofilament impairment and neurotoxicity. In the present study, we are further investigating the in vivo effect of melatonin on spatial memory retention in rats by bilateral injection of calyculin A (CA), a potent and specific inhibitor of protein phosphatase-2A (PP-2A) and protein phosphatase-1 (PP-1) into hippocampus and exploring its underling mechanisms. We examined memory retention of the rats by the Morris Water-Maze Test, analyzed the axonal transport by a time-lapse recording of pEGFP-labeled neurofilament-M (pEGFP-NF-M) subunit in live neuroblastoma N2a cells. In addition, we explored the effect of melatonin on the morphological alteration of the cells during inhibition of the phosphatases by establishing a cell model showing steady outgrowth of axon-like cell processes and employed a stereological system to analyze the retraction of the processes. In the present study,we found that the supplementation of melatonin by intraperitoneal injection for 1 week before injecting CA not only significantly improves CA-induced memory retention deficits of the rats in the Morris Water-Maze Test, but also decreases hyperphosphorylation of neuronal cytoskeletal protein tau and neurofilaments. To further explore the underlying mechanisms by which melatonin exerted the protective effect on spatial memory deficit in rats induced by inhibition of PP-2A and PP-1, we analyzed the axonal transport in live neuroblastoma N2a cells and found that melatonin partially reversed CA-induced impairment of transport of pEGFP-NF-M in the axon-like processes of the cells. In the meantime, we found by western blot that melatonin also decreased CA-induced hyperphosphorylation of cytoskeletal proteins. Furthermore, we found CA-treatment inhibited outgrowth of the cell processes and prolonged treatment with CA caused retraction of the processes and meanwhile, the early neurodegenerative varicosities were also obvious in the CA-treated cells, and the administration of melatonin significantly arrested these pathological alterations. These data suggest that melatonin not only meliorated impaired intracellular transport induced by suppression of PP-2A and PP-1, but also cell degeneration, which may underlie the basis of the effects of protection of melatonin on memory deficits induced by suppression of the phosphatases.