Rapid agonist-induced internalization of the 5-hydroxytryptamine2A receptor occurs via the endosome pathway in vitro.
The mechanism by which agonists induce 5-hydroxytryptamine2A (5-HT2A) receptor internalization was investigated in a clonal cell line stably transfected with the 5-HT2A receptor cDNA. Confocal laser microscopy of immunolabeled 5-HT2A receptors in control (untreated) cells demonstrated that most of the immunoreactivity was associated with the cell surface. After quipazine administration, a significant increase in intracellular immunofluorescence was measured. Time course studies demonstrated rapid agonist-dependent internalization of 5-HT2A receptors, with significant internalization occurring as early as 5 min after agonist administration at 37 degrees. In GF-62 cells, agonist-induced internalization was blocked by preincubation with the 5-HT2A receptor antagonist ketanserin. Internalization was also temperature sensitive because agonist-induced internalization did not occur at 4 degrees. Dual-label experiments disclosed that 5-HT2A and transferrin receptors were internalized via the same endocytotic vesicles. These results suggest that 5-HT2A receptors and transferrin receptors are internalized via the endosomal pathway in GF-62 cells. Although 5-HT2A receptors were internalized, down-regulation, or loss of radioligand binding sites, did not occur. Our results demonstrate that agonists rapidly induce 5-HT2A receptor internalization via the endosomal pathway and that internalization can be dissociated from down-regulation.
Berry
S A
SA
Department of Biochemistry Case Western Reserve University School of Medicine, Cleveland, Ohio 44106, USA.
Shah
M C
MC
Khan
N
N
Roth
B L
BL
D016475:3T3 Cells; D000818:Animals; D015536:Down-Regulation; D011992:Endosomes / Q000378:metabolism*; D007650:Ketanserin / Q000494:pharmacology; D051379:Mice; D011985:Receptors, Serotonin / Q000032:analysis / Q000378:metabolism*; D011990:Receptors, Transferrin / Q000032:analysis; D017366:Serotonin Receptor Agonists / Q000494:pharmacology*; D013696:Temperature
D011985:Receptors, Serotonin; D011990:Receptors, Transferrin; D017366:Serotonin Receptor Agonists; D007650:Ketanserin