Evidence for cell type dependent mechanisms in agonist-induced down-regulation of beta-adrenoceptors.
Processing of the beta-adrenoceptors by mammalian tumor cells, Chang liver cells and HeLa cells both containing beta-2-adrenoceptors, was studied using 125I-labeled iodocyanopindolol and 3H-labeled CGP12177. No difference could be detected between the results obtained with these two ligands. During desensitization of these cells by the beta-adrenergic agonist isoproterenol a time dependent decrease of intracellular cyclic-AMP was accompanied by loss of beta-adrenoceptors. The loss of receptors could not be prevented by the cytoskeleton disrupting agents colchicine and cytochalasin B. The effect of the lysosomotropic agent chloroquine is dependent upon the cell type. It showed no effect on the agonist-induced down-regulation of beta-adrenoceptors on Chang liver cells in contrast to a prominent inhibiting effect on HeLa cells. These results highly suggest that lysosomal enzymes are involved in the process of down-regulation of beta-adrenoceptors on certain cell types only. Colchicine and cytochalasin B, as well as Gpp(NH)p were able to uncouple the guanine nucleotide binding protein and the beta-adrenoceptor. This uncoupling however, did not inhibit the loss of receptors measured by radioligand binding techniques during challenge with agonists. From experiments performed to separate membrane fractions on non-linear sucrose gradients it cannot be concluded that receptors are redistributed intracellular or in the plane of the membrane. Withdrawal of the agonist during receptor down-regulation resulted in the reappearance of measurable receptors within 20 hr for receptors on HeLa cells and within 60 hr for Chang liver cells. The reappearance of receptors could be totally blocked by cycloheximide, indicating that these receptors were newly synthesized and not internalized receptors. From the results of our experiments it is concluded that beta-adrenoceptors, present on Chang liver cells and HeLa cells are down-regulated during desensitization by isoproterenol without uptake in intracellular structures of the cells. The fate of these down-regulated receptors remains unclear. It is also concluded that no universal mechanism can be proposed for beta-adrenoceptor down-regulation on mammalian cells.
Rademaker
B
B
R&D Pharmaceutical Division, Duphar B.V., Weesp, The Netherlands.
Kramer
K
K
Bast
A
A
Timmerman
H
H
D000318:Adrenergic beta-Agonists / Q000494:pharmacology*; D002478:Cells, Cultured; D000242:Cyclic AMP / Q000378:metabolism; D015536:Down-Regulation / Q000187:drug effects*; D006367:HeLa Cells; D006801:Humans; D007545:Isoproterenol / Q000494:pharmacology; D008099:Liver / Q000166:cytology / Q000187:drug effects; D011869:Radioligand Assay; D011943:Receptors, Adrenergic, beta / Q000187:drug effects*
D000318:Adrenergic beta-Agonists; D011943:Receptors, Adrenergic, beta; D000242:Cyclic AMP; D007545:Isoproterenol