Electrophysiological characterization of N-methyl- d-aspartate receptors in rat dorsal root ganglia neurons

Abstract

In the peripheral nervous system, N-methyl- d-aspartate receptors (NMDAR) expressed on the central and peripheral terminals of primary afferent neurons are involved in nociception. We used single cell imaging of intracellular calcium concentration ([Ca 2+] i) and patch clamp techniques to characterize the functional properties of NMDARs on adult rat dorsal root ganglia (DRG) neurons in primary culture and selectively on those innervating the distal colon. In Mg 2+-free extracellular solution, rapid perfusion of DRG neurons with 250 μM NMDA and 10 μM glycine caused a significant increase in [Ca 2+] i, and elicited inward currents in whole cell patch clamp recordings when the holding potential was −60 mV. Both effects were reversibly inhibited by 200 μM ketamine in a use-dependent manner. The EC 50 values for NMDA and glycine were 64 and 1.9 μM with Hill slope coefficients of 1.4 and 1.3, respectively. At negative potentials, extracellular Mg 2+ blocked currents in a concentration- and voltage-dependent manner. The IC 50 for Mg 2+ at a holding potential of −100 mV was 2.0 μM. The NMDAR subtype-selective antagonist, ifenprodil, inhibited 94% of the NMDA and glycine-induced current with an IC 50 of 2.6 μM. There was no evidence of multiple binding sites for ifenprodil. There was no significant difference in the NMDAR current density on DRG neurons that had innervated the colon, nor was there a difference in the EC 50 for ifenprodil. These results demonstrate that functional NMDARs expressed by DRG neurons innervating both somatic and visceral tissues of adult rats are composed predominantly of NR2B subunits.