Desensitization masks nanomolar potency of ATP for the P2X1 receptor

ATP-gated P2X(1) receptors feature fast activation and fast desensitization combined with slow recovery from desensitized states. Here, we exploited a non-desensitizing P2X(2)/P2X(1), chimera that includes the entire P2X(1) ectodomain (Werner, P., Seward, E. P., Buell, G. N., and North, M A. (1996) Proc. Natl. Acad. Sci. U. S. A. 93, 1548515490) to obtain a macroscopic representation of intrinsic receptor kinetics without bias arising from the overlap of channel activation and desensitization. From the stationary currents made amenable to analysis by this chimera, an EC50 for ATP of 3.3 nm was derived, representing a >200- and >7000-fold higher ATP potency than observed for the parental P2X(1) and P2X(2A) receptors, respectively. Also, other agonists activated the P2X(2)/P2X(1), chimera with nanomolar EC50 values ranging from 3.5 to 73 nm in the following rank order: 2-methylthio-ATP, 2',3'-O-(4-benzoylbenzoyl)-ATP, alpha,beta-methylene-ATP, adenosine 5'-0-(3-thiotriphosphate). Upon washout, the P2X(2)/P2X(1) chimera deactivated slowly with a time constant (ranging from 63 to 2.5 s) that is inversely related to the EC50 value for the corresponding agonist. This suggests that deactivation time courses reflect unbinding rates, which by themselves define agonist potencies. The P2X(2)/P2X(1) chimera and the P2X(1) receptor possess virtually identical sensitivity to inhibition by the P2X(1) receptor-selective antagonist NF279, a suramin analog. These results suggest that the P2X(1), ectodomain confers nanomolar ATP sensitivity, which, within the wild-type P2X(1) receptor, is obscured by desensitization such that only a micromolar ATP potency can be deduced from peak current measurements, representing an amalgam of activation and desensitization.