3H]ZM241385—an antagonist radioligand for adenosine A 2A receptors in rat brain

Abstract

Binding of the novel adenosine A 2A receptor-selective antagonist radioligand [2- 3H]-4-(2-[7-amino-2-{2-furyl}{1,2,4}triazolo{2,3- a}{1,3,5,}triazin-5-yl amino]ethyl)phenol ([ 3H]ZM241385) was examined using particulate preparations and frozen sections of rat brain. In membranes from the rat striatum, binding was saturable, reversible and temperature-dependent. Analysis of saturation isotherms indicated that [ 3H]ZM241385 bound with high affinity ( K d of 0.84 nM), high density (1680 fmol mg protein −1) and with a high proportion of specific binding (93% at 1 nM radioligand). Examination of competition profiles indicated that [ 3H]ZM241385 bound to sites with an A 2A adenosine receptor-like rank order. The presence of guanosine 5′-(3-thio)-triphosphate failed to alter either [ 3H]ZM241385 binding or agonist competition for [ 3H]ZM241385 binding. Autoradiographic analysis of [ 3H]ZM241385 binding to frozen sections of rat brain indicated specific binding to the rat striatum of similar affinity ( K d of 0.43 nM) and susceptibility to adenosine receptor ligands. At 2 nM [ 3H]ZM241385, specific binding comprised 95±1% total binding. In the hippocampus and frontal cortex, binding of [ 3H]ZM241385 failed to saturate and was of lower density. Taken together, these results indicate that [ 3H]ZM241385 should prove to be a useful radioligand in the characterisation of adenosine A 2A receptors.