Binding and internalization of iodinated neurotensin in neuronal cultures from embryonic mouse brain.
The binding and internalization of labeled neurotensin were studied by means of biochemical and light microscopic radioautography techniques in primary cultures of neurons from whole cerebral hemispheres of mouse embryos. Saturable, high affinity neurotensin binding was detected 5-7 days postplating in cells incubated with 0.1 nM 125I-Tyr3-neurotensin at 37 degrees C or 10 degrees C. The binding capacity at equilibrium was 3 times higher at 37 degrees C than at 10 degrees C. Moreover, whereas virtually all the radioactivity bound at 10 degrees C was membrane-bound (i.e. was readily washable by a hypertonic, high pH, NaCl solution), more than 70% of the radioactivity bound at 37 degrees C was intracellular (i.e. resisted the same treatment). Light microscopic radioautograms of whole cells revealed that approximately 16% of neurons were labeled with 125I-Tyr3-neurotensin at either 37 degrees C or 10 degrees C. The labeling was observed over cell bodies and processes, and the density of silver grains associated with perikarya, as compared to processes, was proportionally higher at 37 degrees C than at 10 degrees C. Semi-thin (1 micron thick) sections through cells incubated at 37 degrees C confirmed that a major fraction of the radioactivity was intracellular and showed that it was mainly confined to the cytoplasm. These results indicate that 125I-Tyr3-neurotensin binds to a distinct subset of primary cultured neurons and that a large proportion of the bound radioactivity undergoes rapid internalization in a temperature-dependent manner. It is proposed that this internalization is ligand-induced and that it may play a role in the modulation of central neurotensin receptor levels.
Mazella
J
J
Institut de Pharmacologie Moléculaire et Cellulaire du CNRS, Valbonne, France.
Leonard
K
K
Chabry
J
J
Kitabgi
P
P
Vincent
J P
JP
Beaudet
A
A
D000818:Animals; D001345:Autoradiography; D001921:Brain / Q000166:cytology / Q000196:embryology / Q000378:metabolism*; D002478:Cells, Cultured; D002851:Chromatography, High Pressure Liquid; D005260:Female; D007457:Iodine Radioisotopes; D051379:Mice; D009474:Neurons / Q000378:metabolism*; D009496:Neurotensin / Q000378:metabolism*; D011247:Pregnancy; D012462:Saline Solution, Hypertonic; D013696:Temperature
D007457:Iodine Radioisotopes; D012462:Saline Solution, Hypertonic; D009496:Neurotensin