Characterization of pulmonary sigma receptors by radioligand binding

Abstract This study establishes the expression of appreciable populations of sites on mouse lung membranes that exhibit radioligand binding properties and pharmacology consistent with assignment as sigma₁ and sigma₂ receptors. Specific binding of the sigma₁ receptor radioligand [³H](+)-pentazocine reached steady state within 6 h at 37 °C. Saturation studies revealed high affinity binding to a single class of sites (K_d 1.36±0.04 nM; B_max 967±11 fmol/mg protein). Inhibition studies showed appropriate sigma₁ receptor pharmacology, including higher affinity for (+)-N-allylnormetazocine with respect to the (-)-enantiomer, and positive allosteric modulation of dextromethorphan binding by phenytoin. Using [³H]1,3-di(2-tolyl)guanidine in the presence of (+)-pentazocine to assess sigma₂ receptor binding, steady state was achieved within 2 min at 25 °C. Cold saturation studies revealed one high affinity, low capacity binding site (K_d 31.8±8.3 nM; B_max 921±228 fmol/mg protein) that displayed sigma₂ receptor pharmacology. A very low affinity, high capacity interaction also was observed that represents saturable, but not sigma receptor specific, binding. A panel of ligands showed rank order inhibition of radioligand binding appropriate for the sigma₂ receptor, with ifenprodil displaying the highest apparent affinity. In vivo, dextromethorphan inhibited the specific binding of a radioiodinated sigma₁ receptor ligand in lung with an ED₅₀ of 1.2 μmol/kg, a value near the recommended dosage for the drug as a cough suppressant. Overall, the present work provides a foundation for studies of drug interactions with pulmonary sigma₁ and sigma₂ receptors in vitro and in vivo.