Desensitization of LLC-PK1 cells by vasopressin results in receptor down-regulation.
The molecular mechanism of desensitization of antidiuretic hormone receptors is not well understood. Preincubation of LLC-PK1 cells with lysine vasopressin (LVP) (10(-6) M, 5 h) decreased subsequent LVP-stimulated cAMP accumulation in cells by 83% and reduced the Vmax of LVP-stimulated adenylate cyclase by 81%. Such preincubation also reduced by 90% the binding of [3H]LVP to both intact cells and isolated plasma membranes, suggesting a loss of vasopressin receptors. Both the reduction in cAMP response and the apparent loss of receptors showed similar dose and time dependence. Monensin (33 microM) did not alter [3H]LVP binding or stimulation of cAMP by LVP, nor did it prevent desensitization. However, membranes prepared from cells preincubated with LVP in the presence of monensin did not show a decrease in [3H]LVP binding. Forskolin preincubation, at 0.1, 1, 10 and 100 microM, did not alter [3H]LVP binding or accumulation of cellular cAMP by LVP, nor did it induce desensitization to LVP. Cells desensitized with varying LVP concentrations in the presence of 10 microM forskolin displayed the same loss of [3H]LVP binding and LVP responsiveness as observed in the absence of forskolin. LVP-desensitized cells, upon removal from LVP-containing medium, recovered cAMP responsiveness to LVP and specific binding of [3H]LVP at the same rate, achieving control levels after 50 h. Recovery was prevented by cycloheximide (25 micrograms/ml). These findings are consistent with a desensitization process involving LVP-mediated receptor internalization, and a recovery process requiring protein synthesis.
Lester
B R
BR
Sheppard
J R
JR
Burman
M
M
Somkuti
S B
SB
Stassen
F L
FL
D000262:Adenylyl Cyclases / Q000378:metabolism; D000818:Animals; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D005576:Colforsin; D000242:Cyclic AMP / Q000096:biosynthesis; D003513:Cycloheximide / Q000494:pharmacology; D004224:Diterpenes / Q000494:pharmacology; D004361:Drug Tolerance; D007700:Kinetics; D008236:Lypressin / Q000378:metabolism / Q000494:pharmacology*; D008985:Monensin / Q000494:pharmacology; D011945:Receptors, Angiotensin / Q000187:drug effects / Q000378:metabolism*; D011956:Receptors, Cell Surface / Q000378:metabolism*; D017483:Receptors, Vasopressin; D014316:Tritium
D004224:Diterpenes; D011945:Receptors, Angiotensin; D011956:Receptors, Cell Surface; D017483:Receptors, Vasopressin; C029150:lysine-vasopressin receptors; D014316:Tritium; D005576:Colforsin; D008236:Lypressin; D008985:Monensin; D003513:Cycloheximide; D000242:Cyclic AMP; D000262:Adenylyl Cyclases