P2X4 receptor and chronic pain - developing novel pharmacological tools to target P2X4 in the central nervous system

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Venoms from snakes, spiders, scorpions and marine animals contain a staggering array of peptides and small molecules. Purification of peptide toxins have revolutionised the understanding of the structure and function of ion channels in vertebrates. These highly specific peptide toxins mediate their actions through binding to ion channels and either blocking or enhancing their activity. Typically these peptides are highly potent and selective for individual ion channels, even individual isoforms, and as such these have the potential to be modified to develop new pharmacological tools for research and drug discovery. P2X4 is an important ligand-gated ion channel activated by extracellular ATP and is thought to play a critical role in the development of neuropathic pain. P2X4 regulates Brain Derived Neurotrophic Factor (BDNF) secretion from activated spinal cord microglia and causes alteration of dorsal horn neuronal excitability culminating in pathological pain. This project will identify and characterise a series of peptides isolated from spider venoms acting on the human P2X4 ligand-gated ion channel. The student will learn how to fractionate venoms using reverse phase HPLC to purify active peptide fractions and will characterise their effects using plate reader methods and patch clamp electrophysiology. We will use our novel modulators of P2X4 to investigate the role of this ion channel in microglial responses, in particular the role of P2X4 in secretion of BDNF from microglia using ELISA techniques.