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

Abstract

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 σ 1 and σ 2 subtypes. A preponderance of σ 2 sites was revealed in all investigated tissues. Different pharmacological profiles were demonstrated for the σ 2 sites in mouse whole brain compared to mouse spinal cord. However, competition studies indicated that the whole brain and spinal [ 3H](+)-pentazocine labeled σ 1 binding sites exhibited similar pharmacological properties. The density of [ 3H](+)-pentazocine labeled σ 1 population was found not to match that of [ 3H]DTG labeled σ 1 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.