Figure 3.

[³H]DPCPX association and dissociation kinetics at 25°C to human A₁ receptors stably expressed in CHO cell membranes. (a) Membranes were pretreated with 10 U ml⁻¹ ADA during preparation (as indicated in the Methods) and the assays were subsequently performed in the presence of 0.1 U ml⁻¹ ADA. (b) Membranes were pretreated with 7 M urea during preparation (as indicated in Methods). [³H]DPCPX concentrations were 0.02 nM for association and 1 nM for dissociation experiments, in 50 mM Tris(HCl) buffer. Nonspecific binding was defined using 10 μM R-PIA. For the dissociation kinetic experiments, membranes were initially equilibrated with radioligand for 90 min prior to addition of 10 μM R-PIA at various time points. Data points represent the mean±s.e.m. obtained from three separate experiments conducted in triplicate.