Two non-stoichiometric binding sites had previously been characterized for the NK-1 receptor using two different types of radio-labelled analogues of substance P. However, the question remained on their eventual conformational interconversion induced or not by the ligand. In this study, kinetic, saturation, and competition studies using [H-3]propionyl[Pro(9)]SP demonstrate the existence of two independent binding components in CHO cells transfected with the human NK-1 receptor, with K-d values of 0.040 nM (approximate to-20% of total sites) and 5.9 nM (approximate to80% of total sites) that correspond to those of the two previously described binding sites. These two binding sites do not seem to interconvert since the minor one can be selectively extinguished in saturation studies in the presence of a SP analogue specific of this binding site. (C) 2003 Elsevier Science (USA). All rights reserved.