Competitive interaction of pirenzepine with rat brain muscarinic acetylcholine receptors

In the present work, we studied the details of the interaction of the nonclassical muscarinic receptor antagonist pirenzepine with [³H]quinuclidinyl benzilate binding sites in rat brain homogenates. Pirenzepine showed biphasic competition curves with a Hill coefficient lower than unity, and these curves were better described according to a two-site receptor model. The affinities and the relative preponderance of these sites were constant at different ligand concentrations, in accordance with a competitive type of interaction. Similarly, pirenzepine did not influence the rate of dissociation of the [³H]quinuclidinyl benzilate-receptor complex, even at relatively high concentrations. However, although low concentrations of pirenzepine decreased the affinity of [³H]quinuclidinyl benzilate for the receptor without affecting the density of the binding sites, higher concentrations of the antagonist decreased the receptor number in a reversible fashion. Schild plots of these data indicated an apparent deviation from simple competition in this experimental design, an observation which can be attributed to the selectivity of pirenzepine for different receptor subtypes. Furthermore, pirenzepine, at concentrations high enough to saturate both its high- and low-affinity sites protected [³H]quiniclidinyl benzilate binding sites in the brain against irreversible alkylation by propylbenzilylcholine mustard. Therefore, our data support a competitive nature of interaction of pirenzepine with rat brain muscarinic receptors.