A recent study demonstrated, LY294002 solubility dmso for example, that a long noncoding RNA that is anti-sense to a K+ channel subunit (Kcna2) is upregulated following peripheral nerve injury, leading to a downregulation of the K+ channel and a resulting increase in the excitability of
DRG neurons, increasing neuropathic pain (Zhao et al., 2013). In the early years, the study of local translation was hampered by the technical difficulty of obtaining pure and sufficient quantities of dendrites and axons for analysis. Pioneering studies used metabolic labeling to demonstrate the synthesis of specific proteins such as tubulin in axons (Giuditta et al., 1968 and Koenig, 2009), but the possibility that the signal arose from cell-body contamination could not be eliminated due to these technical limitations. Localized translation was convincingly demonstrated by surgically severing the soma from its processes (Aakalu et al., 2001, Campbell and Holt, 2001 and Kang and Schuman, 1996) and,
more recently, by the use of chambers in which the processes (dendrites or axons) are fluidically isolated from cell bodies (Eng et al., 1999 and Taylor et al., 2010). Other methods for isolating neuronal processes include substrates with limited pore size that allow axons to penetrate but not cell bodies (Torre and Steward, 1992 and Zheng et al., 2001) and laser capture microdissection (Zivraj et al., 2010). Apoptosis inhibitor These methods combined with the rapid increase in the sensitivity of profiling techniques have enabled genome-wide transcriptome analyses to be performed on axons and dendrites in a variety of neurons (see below). The visualization and identification of newly synthesized proteins has also been a hurdle due to issues of sensitivity (detecting low levels of newly synthesized proteins) as well as difficulties in distinguishing between the movement of existing proteins and the synthesis of new proteins. Puromycin, a tRNA analog, can be used together with fluorescent
tags (Smith et al., 2005) or antibodies (Schmidt et al., 2009) to label sites of protein synthesis. Fluorescent reporters, such as photo-switchable Kaede, fused to the Vasopressin Receptor 3′UTR regulatory region of mRNAs of interest have enabled de novo protein synthesis to be monitored live in neuronal processes (Aakalu et al., 2001, Brittis et al., 2002 and Leung et al., 2006). In addition, new methods have been developed to selectively label the pool of newly synthesized proteins, to ascertain a given cell type or cellular compartment as the site of synthesis, and to visualize the newly synthesized proteins. These methods make use of noncanonical amino acids that cross cell membranes and get charged onto tRNAs by the cell’s own tRNA synthetases and then incorporated into new protein during protein synthesis.