Internalization of V2-vasopressin receptors in LLC-PK1-cells: evidence for receptor-mediated endocytosis.
The mechanism of internalization of the vasopressin-receptor (V2-subtype) of LLC-PK1-cells, a pig renal tubular cell line, is unknown. We studied internalization utilizing a novel, highly specific vasopressin analogue ((125I)-[8-p(OH)-phenylpropionyl]-LVP, 2000 Ci/mmol). Scatchard analysis performed with membranes of LLC-PK1-cells revealed a Kd of 0.8 +/- 0.2 nM and a Bmax of 366 +/- 41 fmol/mg of protein. Degradation of the ligand was excluded by RP-HPLC-analysis. Internalization was proven by the acid-wash technique, quantitative light-microscopic autoradiography and electron microscopy. The ligand was internalized in a time- and temperature-dependent manner. At 4 degrees C, no uptake was found; at 22 degrees C, after 30 min of incubation, more than 50% of the radioligand was found inside the cell. Electron microscopy demonstrated that plasma-membrane bound vasopressin receptors are internalized by receptor-mediated endocytosis via coated pits.
Hocher
B
B
Abteilungen für Innere Medizin, Freie Universität Berlin, Germany.
Merker
H J
HJ
Dürr
J A
JA
Schiller
S
S
Gross
P
P
Hensen
J
J
D000818:Animals; D001345:Autoradiography; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism / Q000648:ultrastructure; D004705:Endocytosis; D007457:Iodine Radioisotopes; D007700:Kinetics; D008854:Microscopy, Electron; D011945:Receptors, Angiotensin / Q000378:metabolism*; D017483:Receptors, Vasopressin; D013696:Temperature; D014667:Vasopressins / Q000378:metabolism*
D007457:Iodine Radioisotopes; D011945:Receptors, Angiotensin; D017483:Receptors, Vasopressin; D014667:Vasopressins