Investigating the mechanisms of action of VGF-derived peptides in the nervous system

Abstract

The VGF neurosecretory protein, first identified as a nerve growth factor (NGF) inducible gene product, is selectively synthesised predominantly in neuronal and neuroendocrine cells. The ~68 kDa VGF protein sequence is rich in paired basic amino acids, and thus the protein undergoes endoproteolytic cleavage to produce smaller peptides, which are stored in dense core vesicles and released upon stimulation via the regulated secretory pathway both in vitro and in vivo. Several of these VGF-derived peptides have been characterised and are involved in energy homeostasis, reproductive processes, synaptic plasticity as well as pain modulation. A number of studies have observed an increase in VGF gene expression in various pain models and more recently the VGF-derived peptides, TLQP-21, LQEQ-19 and TLQP-62 showed direct modulation of inflammatory and neuropathic pain when applied in vivo. The molecular mechanisms of action of VGF-derived peptides are not well understood and were investigated in this study. The TLQP-21 peptide, but not LQEQ-19, was shown to dose-dependently induce an increase in intracellular Ca2+ levels from cellular internal stores in brain- and spinal cord-derived primary microglia, in >65 % of the cell population in vitro. Three hour treatment of primary microglia with TLQP-21 (100 nM) induced a 2.78 fold increase in Ccl11 and a 2.28 fold decrease in Cxcl9 gene expression levels relative to the vehicle control (Student's t-test; p ≤ 0.05). Biochemical analysis using affinity chromatography and LC-MS/MS techniques identified the gC1q-R protein as a potential binding partner / receptor for TLQP-21. The gC1q-R protein is a ubiquitously expressed, multi-compartmental protein involved in complement activation, inflammatory processes and the plasma bradykinin formation pathway. These results tentatively suggest that TLQP-21 may contribute to the modulation of pain through activation of primary microglia and potentially involve interactions with components of the complement system. The findings highlight the importance of VGF-derived peptides in pain research and could lead to new perspectives and targets for pain therapeutics.