Alteration of the Expression Levels of Rab3A Affects the Extent of Transcytosis of HRP-Labeled Marker Proteins in Rat CNS NeuronsReport as inadecuate




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It has been hypothesized that Rab3A, a small GTPase, may be closely involved in the process of dense core vesicle exocytosis in various cell types. This possibility was investigated by disrupting the expression levels of Rab3A-mRNA using a small interfering RNA of the Rab3A GTPase Rab3A-siRNA and examining the effect of this on transcytosis of wheat germ agglutinin conjugated with horseradish peroxidase WGA-HRP. Rab3A-siRNA and WGA-HRP were injected into the right vagus nerves of adult rats which were killed 12, 24 or 48 hours later. In some animals, portions of the brain stem containing the nucleus of solitary tract NST were prepared for electron microscopy. In other animals, the nodose ganglion of the vagus nerve was used to determine the levels of expression of Rab3A-mRNA using RT-PCR techniques. It was found that the expression of Rab3A-mRNA was markedly depressed in animals at 12 h after the Rab3A-siRNA injection. In the NST, there was an accumulation of HRP-reaction product RP, recognized as electron dense lysosomal-like structures, in both axons and terminals in the NST 12 h after injection. Some HRP-RP was found in membrane bound vesicles in close proximity to cell membranes and appeared to be in the process of transcytosis. This neuronal transcytosis of HRP-RP appeared to occur at random locations over the axodendritic membranes. These findings indicate that inhibiting the expression of Rab3A-mRNA using Rab3A-siRNA can modulate the level of transcytosis of proteins across neuronal membranes confirming the potentially important role of this GTPase in the process of transcytosis.

KEYWORDS

Rab3A-siRNA, WGA-HRP, Transcytosis, Synapse, Vagus Nerve, Nucleus of Solitary Tract, Vesicle Docking, Quantitative Real Time RT-PCR

Cite this paper

Y. Takeuchi, Y. Matsumoto, T. Miki, K. Warita, Z. Wang, K. Bedi, T. Yakura and J. Liu -Alteration of the Expression Levels of Rab3A Affects the Extent of Transcytosis of HRP-Labeled Marker Proteins in Rat CNS Neurons,- Neuroscience and Medicine, Vol. 2 No. 3, 2011, pp. 282-287. doi: 10.4236-nm.2011.23036.





Author: Yoshiki Takeuchi, Yoshiki Matsumoto, Takanori Miki, Katsuhiko Warita, Zhi-Yu Wang, Kuldip S. Bedi, Tomiko Yakura, Jun-Qian Liu

Source: http://www.scirp.org/



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