Inhibition of binding of the labelled antagonist (−)[ 3 H]CGP 12177 by the full agonist (−)isoproterenol results in shallow competition curves, characteristic of the presence of both high- and low-affinity states of β-adrenoceptors (βAR). When in excess, the GTP analog 5′-guanylylimidodiphosphate (GppNHp) is expected to convert all receptors in the high-affinity state to the low-affinity state. However, in the rat cortex and cerebellum synaptosomes, a proportion of the βAR in the high-affinity state was GppNHp-insensitive. This apparent GppNHp-insensitivity decreased with decreasing temperature of incubation. Moreover, it was totally abolished by the gentle treatment of membranes with 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate (CHAPS). We propose that a protein factor interacts with the βAR/G s protein complex and that it induces the GppNHp-insensitivity. This factor would be released by CHAPS in a functional form because it may regenerate the GppNHp-insensitivity after concentration and reconstitution with CHAPS-treated membranes. It is likely that the factor acts as a stabiliser of βAR in the high-affinity state.