Stable expression of high affinity NK 1 (substance P) and NK 2(neurokinin A) receptors but low affinity NK 3 (neurokinin B) receptors in transfected CHO cells

Abstract

Stable CHO cell clones which selectively express all three rat tachykinin receptors were established by transfection. The binding of radiolabeled substance P and neurokinin A (substance K) to CHO clones expressing the NK 1 and NK 2 receptors, respectively, were saturatable and of high affinity ( K d1=0.17 nM (NK 1); 3.4 nM (NK 2)). Scatchard analysis of the binding data indicated for both receptors binding to a single population of binding sites, and competition binding studies showed that the binding specificities of the receptors corresponded to those of classical NK 1 and NK 2 receptors. In contrast, the binding of eledoisin to the NK 3 receptor expressed in the transfected CHO cells was of low affinity (IC 50=240 nM) compared to the high affinity of the receptor found when it was transiently expressed in COS-7 cells (IC 50=8 nM). However, in both cases the receptor exhibited the specificity of a classical NK 3 receptor. The established cell clones may provide an important tool for further analysis of the molecular mechanisms involved in binding, activation, and coupling of receptors for tachykinin peptides.