Role Of P2X Receptor In Neuropathic Pain And The Rapid Modulation Of P2X Receptors By Glucocorticoid In Rat Dorsal Root Ganglion Neurons

Neuropathic pain is a kind of chronic pain that is complicate and difficult to cure . ATP as an excitatory neurotransmitter or neuromodulator is implicated in peripheral sensory transduction of noxious stimuli by activating P2X receptors. Previous evidence indicates that dorsal root ganglion (DRG) neurons, the primary sensory neurons express multiple P2 receptors, including P2X1–6. Among them, P2X3 receptors were present predominantly in a subpopulation of small-diameter sensory neurons related with nociceptive reception. P2X receptors belong to a superfamily of ligand-gated, non-selective cation channels. Activation of P2X receptors results in the generation of an inward cation current with a subsequent depolarization. Depolarization of the peripheral nerve terminals produces a sensation of pain. Therefore, depression of the excitability of DRG neurons becomes a kind of methods for pain therapy.Glucocorticoid (GC) affects virtually all organs that regulate metabolism, hydromineral balance, growth, development, and neuronal functions. Classically, GC is thought to penetrate into the cytoplasm of neurons, bind to intracellular receptors, and induce genomic effects through new protein synthesis. In addition to the classical genomic pathways, GC has been found to generate multiple rapid effects on various tissues and cells through non-genomic pathways. Recent researcth found that GC is involved in pain transduction. It has been demonstrated that neuropeptide content in rat DRG neurons is altered by GC. ATP is implicated in peripheral sensory transduction of noxious stimuli by activating P2X receptors. It is clear that many noxious stimuli can lead to an increase in extracellular levels of GC. Whether there exists a rapid modulatory role of GC on the actions of extracellular ATP in DRG neurons remains unclear.The present study is aimed to explore the following questions:1. Whether P2X receptors play a role in modulating generateion and developmention of neuropathic pain;2. Whether a nongenomic action of GC on ATP induced whol cell currents is present in primary afferent neurons ;3. Whether a nongenomic action of GC on ATP induced intracellular calcium concentration ( [Ca2+]i ) increase is present in primary afferent neurons .Results1. Alteration of P2X3 receptor expression and electrophysiological character in neuropathic pain(1) After CCI, animals developed clear behaviors indicative of neuropathic pain such as thermal and mechanical hyperalgesia compared with sham animals. These pain behaviors can be reversed by TNP-ATP (P2X receptor antagonist), indicating an involvement of P2X receptors.(2) A significant increasion in P2X3 immunoreactivity was observed in the ipsilateral (injured) L4-6 DRG and spinal cord in CCI 7,14 d rats.(3) In small to medium DRG neurons, a significant increasion in the number of cells exhibiting a transient current to ATP was observed in CCI 7,14 d rats. Moreover, amplitude of these currents were increased.2. GC inhibited ATP-induced transient current in DRG neurons(1) Three types of currents (transient, sustained and biphasic) were evoked by ATP(100μmol/L) in cultured DRG neurons.(2) When DRG neurons were pretreatment with dexamethasone (Dex, 0.01–10μmol/L) for 30s, an inhibition of the transient current and the transient component of the biphasic current evoked by ATP in DRG neurons was observed . The inhibitory effect of Dex was dose-dependent.(3) The inhibitory effect of Dex on ATP-induced currents was blocked by glucocorticoid receptor antagonist RU38486(10μmol/L), but not by G protein inhibitor GDP-β-S (0.2 mmol/L)(4) The inhibitory effect of Dex on ATP-induced currents was blocked by protein kinase A inhibitor H-89(10μmol/L), but not by protein kinase C inhibitor Chelerythrine chloride (10μmol/L).(5) corticosterone (CORT) had a similar inhibitory effect on ATP-induced currents to Dex. Membrane-impermeable bovine serum albumin-conjugated corticosterone (CORT-BSA) failed to mimic the effects of CORT.3. GC inhibited ATP-induced [Ca2+]i increase in DRG neurons(1). ATP(100μmol/L) caused increased [Ca2+]i transients in small to medium DRG neurons. ATP induced [Ca2+]i changes via activation of P2X receptors and extracellular calcium influx .(2). When DRG neurons were pretreatment with Dex (0.01–10μmol/L) for 5 min, an inhibition of ATP-induced [Ca2+]i increase in DRG neurons was observed . The inhibitory effect of Dex was dose-dependent.(3). The inhibitory effect of Dex on ATP-induced [Ca2+]i increase was blocked by glucocorticoid receptor antagonist RU38486(10μmol/L), but not by G protein inhibitor GDP-β-S (0.2 mmol/L)(4). The inhibitory effect of Dex on ATP-induced [Ca2+]i increase was blocked by protein kinase A inhibitor H-89(10μmol/L), but not by protein kinase C inhibitor Chelerythrine chloride (10μmol/L).(5). CORT had a similar inhibitory effect on ATP-induced [Ca2+]i increase to Dex. CORT-BSA failed to mimic the effects of CORT.Conclusion1. P2X receptors play an important role in modulating generation and developmention of neuropathic pain by increasing the expression and role of P2X3 receptor in DRG neurons.2. GC can inhibit ATP-induced transient current and ATP-induced [Ca2+]i increase in DRG neurons that mediated by glucocorticoid receptor through activing PKA signal pathway. These results suggest that GC might participate in control of pain by modulating the actions of extracellular ATP in sensory neurons.