Identification and regulation of angiotensin II receptor subtypes on NG108-15 cells.
NG108-15 cells, a neurally derived clonal cell line, express various components of the renin-angiotensin system and thus serve as a model of the cellular action of angiotensin (Ang) II. NG108-15 cells contain a high-affinity binding site for Ang II, with a Kd of 1.1 nM and a Bmax of 6.5 fmol/mg protein. Ang peptides competed for 125I-Ang II binding with an order of potency of Ang II greater than Ang-(2-8) much greater than Ang-(1-7). The subtype 1 (or B)-selective Ang II receptor antagonist DuP 753 as well as [Sar1,Ile8]Ang II and [Sar1,Thr8]Ang II competed for Ang II binding with high affinity, whereas the subtype 2 (or A)-selective Ang receptor antagonist CGP 42112A was partially effective only at a 300-fold higher concentration. When NG108-15 cells were induced to differentiate by treatment with dibutyryl cyclic adenosine 3',5'-monophosphate, the density of Ang II receptors increased dramatically, with little change in affinity (1.1 versus 4.2 nM) or competition by Ang peptides. In marked contrast to undifferentiated cells, CGP 42112A became a potent competitor (IC50, 1 nM) for the majority (90-95%) of Ang II binding, whereas DuP 753 competed for only 5-10% of the binding sites. Ang II caused a dose-dependent mobilization of cytosolic Ca2+ in undifferentiated NG108-15 cells through activation of phospholipase C and the production of inositol 1,4,5-trisphosphate. In these cells, Ca2+ mobilization was blocked by either DuP 753 or the sarcosine Ang II analogues, whereas CGP 42112A was ineffective. Ang II also mobilized intracellular Ca2+ in differentiated NG108-15 cells.(ABSTRACT TRUNCATED AT 250 WORDS)
Tallant
E A
EA
Department of Brain and Vascular Research, Cleveland Clinic Foundation, Ohio 44195-5286.
Diz
D I
DI
Khosla
M C
MC
Ferrario
C M
CM
D015059:1-Sarcosine-8-Isoleucine Angiotensin II / Q000378:metabolism; D000804:Angiotensin II / Q000031:analogs & derivatives / Q000037:antagonists & inhibitors / Q000378:metabolism; D057911:Angiotensin Receptor Antagonists; D001667:Binding, Competitive; D002118:Calcium / Q000378:metabolism; D006822:Hybrid Cells; D007093:Imidazoles / Q000378:metabolism; D019808:Losartan; D009474:Neurons / Q000378:metabolism*; D009842:Oligopeptides / Q000378:metabolism; D011945:Receptors, Angiotensin / Q000378:metabolism*; D015398:Signal Transduction; D013777:Tetrazoles / Q000378:metabolism; D014407:Tumor Cells, Cultured
D057911:Angiotensin Receptor Antagonists; D007093:Imidazoles; D009842:Oligopeptides; D011945:Receptors, Angiotensin; D013777:Tetrazoles; D000804:Angiotensin II; C060894:CGP 42112A; C014805:angiotensin II, Sar(1)-Thr(8)-; D015059:1-Sarcosine-8-Isoleucine Angiotensin II; D019808:Losartan; D002118:Calcium