CHARACTERIZATION OF THE ANGIOTENSIN-II AT(1) RECEPTOR SUBTYPE INVOLVED IN DNA-SYNTHESIS IN CULTURED VASCULAR SMOOTH-MUSCLE CELLS

1 This study was undertaken in cultured vascular smooth muscle cells t o characterize the angiotensin II (AII) AT(1) receptor subtype involve d in DNA synthesis because (i) the AII receptor involved in vascular p roliferation has previously been characterized in vitro in rat aortic cells and identified as an AT(1) subtype and (ii) molecular cloning an d biochemical studies have provided evidence for the existence of diff erent AT(1) receptor subtypes. 2 In cultured rat aortic vascular smoot h muscle (VSMC), exposure to AII (0.1 to 100 nM) resulted in a concent ration-dependent increase in [[H-3]-thymidine incorporation with an EC (50) of 1.41 +/- 0.51 nM. Maximal stimulation was observed in the pres ence of 100 nM AII and corresponded to 271 +/- 40% of basal [H-3]-thym idine incorporation. 3 To characterize the AII AT(1) receptor subtype involved in this effect, cells were exposed to AII (3 nM) in the absen ce or presence of increasing concentrations of various AII receptor an tagonists. The stimulatory effect of AII (3 nM) on [[H-3]-thymidine in corporation in VSMC was antagonized by the non-selective AT(1)/AT(2), receptor antagonist, [Sar(1),Ile(8)]-AII (IC50 = 5.6 nM), by the AT(1A )/AT(1B) receptor antagonist, losartan (IC50 = 10.5 nM) and the AT(1) receptor antagonist, L-158809 (IC50= 0.20 nM). The selective AT(2) rec eptor ligand, CGP 421121A, antagonized AII-induced [H-3]-thymidine inc orporation with an IC50 of 6.3 +/- 1.3 mu M while the AT(2)/AT(1B), re ceptor antagonist, PD 123319, was found to be almost inactive (IC50> 1 0 mu M). 4 Under the same experimental conditions, angiotensin III (AI II) was found to be at least 50 times less potent than AII with an app arent EC(50) of 81.6 +/- 7.7 nM. At the highest concentration tested ( 10 mu M), the effect of AIII corresponded to 327 +/- 61% of basal [H-3 ]-thymidine incorporation. 5 These results confirm that AII can stimul ate DNA synthesis in VSMC through an AT(1) receptor. Furthermore, the pharmacological characterization of this AT(1), receptor is compatible with the AT(1A) receptor subtype recently described on cultured mesan gial cells since (i) the AT(1A)/AT(1B) receptor antagonist losartan is active at nanomolar concentrations, (ii) micromolar concentrations of the AT(2)/ AT(1B) receptor antagonist PD 123319 are ineffective at an tagonizing the AII-induced [H-3]-thymidine incorporation and (iii) AII is at least 50 times more potent than AIII in stimulating DNA synthes is.