Heterogeneity of binding sites for N-ethylcarboxamido[3H]adenosine in rat brain: effects of N-ethylmaleimide.
Binding sites for N-ethylcarboxamido[3H]adenosine (NECA) in rat brain membranes and cryostat sections were examined in relation to their sensitivities to displacement by unlabeled NECA and R(-)-phenylisopropyladenosine (R-PIA). In membrane fractions from cerebral cortex, cerebellum, hippocampus and striatum, nanomolar concentrations of these adenosine receptor agonists displaced [3H]NECA such that R-PIA was more effective than NECA, consistent with the presence of an A1-adenosine receptor. At concentrations of displacing agent higher than 1 microM, R-PIA was unable to displace [3H]NECA further in cerebral cortex, cerebellum and hippocampus. In striatum, a second R-PIA-sensitive component of [3H]NECA binding was evident which was more sensitive to NECA than to R-PIA, i.e. it showed the characteristics of an A2-adenosine receptor. In striatum, however, R-PIA was also unable to displace [3H]NECA binding completely. Similar results were obtained in quantitative autoradiographic studies. Preincubation of cryostat sections with N-ethylmaleimide (NEM) abolished both the A1- and R-PIA-insensitive binding components such that both NECA and R-PIA were able to displace [3H]NECA binding completely. The remaining sites showed IC50 values of 0.13 and 3.68 microM for NECA and R-PIA, respectively. These A2-like [3H]NECA binding sites had a highly specific distribution in the brain, being concentrated in the striatum, nucleus accumbens and olfactory tubercle. The results indicate the presence in brain tissue of at least 3 classes of [3H]NECA binding sites, an A1-site, an A2-site and a third, unidentified R-PIA-insensitive site.
D015762:2-Chloroadenosine; D000241:Adenosine / Q000031:analogs & derivatives* / Q000378:metabolism / Q000494:pharmacology; D019830:Adenosine-5'-(N-ethylcarboxamide); D000818:Animals; D001345:Autoradiography; D001665:Binding Sites / Q000187:drug effects; D001921:Brain / Q000187:drug effects / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism; D005033:Ethylmaleimide / Q000494:pharmacology*; D011869:Radioligand Assay; D051381:Rats; D013347:Subcellular Fractions / Q000378:metabolism
D015762:2-Chloroadenosine; D019830:Adenosine-5'-(N-ethylcarboxamide); D000241:Adenosine; D005033:Ethylmaleimide