Characterization of cultured dorsal root ganglion neuron P2X receptors

Abstract

P2X receptors for adenosine 5'-triphosphate (ATP) comprise a family of ligand-gated cation channels with distinct characteristics which are dependent on the receptor subunits (P2X1-7) expressed, and the homomeric or heteromeric assembly of protein subunits in individual cells. We describe the properties of P2X receptors expressed by cultured adult rat dorsal root ganglion cells on the basis of the time course of responses to ATP, alpha, beta-methylene adenosine 5'-triphosphate (alpha, beta-meATP) and 2-methyl-thioadenosine 5'-triphosphate (2-meSATP), and using the antagonists 2',3'-O-(2,4,6-trinitrophenyl) ATP (TNP-ATP), a novel and highly selective purinoceptor antagonist, suramin and iso-pyridocalphosphate-6-azophenyl-2',5' disulphonic acid (PPADS). ATP (10 microM) evoked inward currents in approximately 95% of neurons tested and > 80% responded with a fast transient inward current that rapidly inactivated during the continued presence of ATP. Of the remaining neurons, approximately 4% showed a sustained response and approximately 10% showed a combination of transient and sustained components. Rapid application of ATP, alpha,beta-meATP and 2meSATP demonstrated these to be full agonists of the rapidly inactivating P2X receptor (pA50 values = 5.83, 5.86 and 5.55, respectively), whilst uridine triphosphate (UTP) and 1-beta,gamma-methyleneadenosine 5'-triphosphate (1-beta,gamma-meATP) were ineffective as agonists. These rapidly inactivating responses could be inhibited by TNP-ATP, suramin and PPADS (pIC50 = 9.5, 6.5, 6.4, respectively). Using inactivation protocols, we demonstrate the presence of homomeric P2X3-like receptors and non-inactivating P2X receptors, which indicates that individual subsets of adult dorsal root ganglion neurons have distinct P2X receptor phenotypes, and that individual DRG neurons may express multiple P2X receptor subtypes.

Neutralization of Nerve Growth Factor Induces Plasticity of ATP-Sensitive P2X3Receptors of Nociceptive Trigeminal Ganglion Neurons

“…The kinetics of P2X 2/3 current decay in heterologous expression systems are primarily influenced by the relative ratio between P2X 3 and P2X 2 injected cDNAs (Liu et al, 2001). Native P2X 2/3 receptors display biphasic currents that reach steady-state conditions at the end of agonist application (Burgard et al, 1999;Grubb and Evans, 1999;Simonetti et al, 2006). We assumed that currents comprising a residual current at the end of a 2-s-long agonist application (I residual ) Ն5% of peak current (I peak ) were suggestive of heteromeric P2X 2/3 receptors (Grubb and Evans, 1999;Simonetti et al, 2006).…”

“…Native P2X 2/3 receptors display biphasic currents that reach steady-state conditions at the end of agonist application (Burgard et al, 1999;Grubb and Evans, 1999;Simonetti et al, 2006). We assumed that currents comprising a residual current at the end of a 2-s-long agonist application (I residual ) Ն5% of peak current (I peak ) were suggestive of heteromeric P2X 2/3 receptors (Grubb and Evans, 1999;Simonetti et al, 2006). This calculation was used to normalize the slow current to the size of the preceding fast response that could vary depending on the experimental protocol.…”

Cutaneous Sensory Neurons Expressing the Mrgprd Receptor Sense Extracellular ATP and Are Putative Nociceptors

“…6A). Both the pharmacology and the current kinetics suggest the presence of homomeric P2X3 receptors (Chen et al 1995;Grubb and Evans 1999;North and Surprenant 2000;Pankratov Yu et al 2001). In MrgprdϪ neurons, 65% of those responding to either 50 M ATP or 50 M ␣␤Me-ATP displayed current that had more prolonged kinetics and was much slower to desensitize (Fig.…”

Effects of A‐317491, a novel and selective P2X3/P2X2/3 receptor antagonist, on neuropathic, inflammatory and chemogenic nociception following intrathecal and intraplantar administration

“…ATP depolarizes primary sensory neurons (Cook et al, 1997;Grubb & Evans, 1999) as well as neurons in the dorsal horn of the spinal cord (Jahr & Jessell, 1985;Li & Perl, 1995). ATP triggers neuronal excitation through several different subtypes of P2 purinoceptors.…”