Stable expression of the mouse levocabastine-sensitive neurotensin receptor in HEK 293 cell line: binding properties, photoaffinity labeling, and internalization mechanism

Abstract

The recently cloned new subtype of G protein-coupled neurotensin receptor (NTRL) was stably expressed in the HEK 293 cell line in order to investigate its binding and internalization properties. The expressed receptor exhibited the typical binding characteristics of the low affinity, levocabastine-sensitive binding site previously described in rat and mouse brain and was detected as a protein with an apparent MW of 45 kDa by photoaffinity labeling. Although intracellular modulation of adenylate cyclase, guanylate cyclase and phospholipase C was not detected after application of neurotensin or levocabastine on NTRL-transfected cells, this receptor was able to internalize iodinated neurotensin. The internalization process was followed by recycling of receptors to the cell membrane. By contrast, no recycling was observed with the high affinity neurotensin receptor (NTRH). The differential intracellular routing of NTRH and NTRL after internalization is most probably the consequence of their divergent carboxy-terminal sequences.