ATP-activated channels in rat and bullfrog sensory neurons: current- voltage relation and single-channel behavior

Abstract

Ionic currents activated by extracellular adenosine 5'-triphosphate (ATP) were studied in voltage-clamped dorsal root ganglion neurons from rats and bullfrogs. Under quasiphysiological ionic conditions, ATP-activated current reversed near 0 mV and showed strong inward rectification. Strong inward rectification was maintained even in symmetric solutions of divalent-free Cs glutamate. Examined with a resolution of 10s of microseconds, the rectification was instantaneous. Inward current was greatly reduced when N-methyl-D-glucamine was substituted for external Na. ATP-activated inward currents could be recorded with Ca as the only external cation; estimated from reversal potentials, the ratio of Ca to Na permeability is about 0.3. Unitary channel activity could be recorded when ATP was applied to outside-out patches. When activated, a single channel flickered rapidly, with a mean current of about 0.5 pA at -100 mV. Large concentrations of ATP put the channel in the activated, flickery condition virtually all the time, while at lower concentrations, periods of flickering were interspersed with closures. Analysis of whole-cell current fluctuations showed precisely the characteristics expected if such channels underlie the macroscopic currents.

Competitive Antagonism of Recombinant P2X2/3Receptors by 2′,3′-O-(2,4,6-Trinitrophenyl) Adenosine 5′-Triphosphate (TNP-ATP)

“…In comparison, the on-kinetics for TNP-ATP at an IC 90 concentration was 10-fold slower, and the corresponding off-kinetics were Ͼ25-fold slower than the agonist. Although channel-gating kinetics have not been described adequately for either P2X 3 or P2X 2/3 receptors, channel opening and closing rates estimated from either recombinant or native P2X receptors indicate that these parameters are also significantly faster than TNP-ATP kinetics (Krishtal et al, 1988;Bean et al, 1990;Cloues, 1995;Wright and Li, 1995;Evans, 1996;Ding and Sachs, 1999). Having met these assumptions, the TNP-ATP kinetics measured in the present studies are a reasonable measure of apparent antagonist binding and unbinding rates.…”

Molecular Physiology of P2X Receptors

“…Dorsal root ganglion cells of the bullfrog were thoroughly studied by Bean et al (17)(18)(19). The currents develop within 8 ms at saturating ATP concentrations (100 M), which was the limit of the solution exchange around an intact neuron.…”

Activation and Regulation of Purinergic P2X Receptor Channels

“…The concentration-dependence of P2XR activation measured by whole-cell (Bean et al, 1990;Jiang et al, 2003) and single-cell (Ding and Sachs, 1999) recording suggested that three molecules of ATP were required for channel gating. Biochemical studies showed that under nondenaturizing conditions, P2XRs migrated as trimers in polyacrylamide gel electrophoresis, therefore suggesting that this was the conformation of native functional channels (Nicke et al, 1998).…”