Features of ligand binding in homogenate and section preparations

The binding of [3H]N-methyl scopolamine (NMS) to muscarinic binding sites in rat forebrain was compared in two types of preparations: homogenates and slide-mounted sections. Under standard assay conditions, [3H]NMS bound to the muscarinic binding site with an apparent Kd that was almost one order of magnitude lower in homogenates (Kd = 0.15 nM) than sections (Kd = 1.25 nM). In addition, the muscarinic agonist carbachol inhibited [3H]NMS binding more effectively in homogenates (Ki = 5 microM) than sections (Ki = 100 microM). The higher Kd observed in sections was dependent upon the thickness of the section since the apparent Kd decreased as the thickness of the section was reduced. A slower equilibration rate may account, in part, for the higher apparent Kd seen in thicker sections. The results support previous evidence that certain ligands exhibit different binding profiles in homogenate and section preparations.