Discrepancies in characterization of σ sites in the mouse central nervous system

The characteristics of [³H](+)-pentazocine and [³H]1,3-di(2-tolyl)guanidine (DTG) binding to mouse whole brain, cortex, cerebellum and spinal cord membranes were investigated in radioreceptor assays. [³H](+)-Pentazocine bound to a single, high affinity site (K_d = 1.2 - 1.6 nM) with increasing density along the neuraxis from the cortex (B_max = 543 fmol/mg protein) to the spinal cord (B_max = 886 fmol/mg protein). Hot saturation studies resolved the presence of one binding site for [³H]DTG showing no tissue variations in terms of density (B_max = 1075-1264 fmol/mg protein) or affinity (K_d = 16.6-22.3 nM). Incubation with 100 nM (+)-pentazocine revealed two classes of high affinity [³H]DTG labeled binding sites corresponding to σ₁ and σ₂ subtypes. A preponderance of σ₂ sites was revealed in all investigated tissues. Different pharmacological profiles were demonstrated for the σ₂ sites in mouse whole brain compared to mouse spinal cord. However, competition studies indicated that the whole brain and spinal [³H](+)-pentazocine labeled σ₁ binding sites exhibited similar pharmacological properties. The density of [³H](+)-pentazocine labeled σ₁ population was found not to match that of [³H]DTG labeled σ₁ site throughout the mouse central nervous system. The presence of low affinity [³H]DTG labeled sites was demonstrated in cold saturation experiments. Equilibrium binding data for the low affinity [³H]DTG binding site resulted in an increasing density (B_max = 1973-11369 fmol/mg protein) with a decreasing affinity (K_d = 242-943 nM) in mouse cortex through the spinal cord.