Table 2

Agonist efficacies and potencies at hH₂R-G_sαS and gpH₂R-G_sαS expressed in Sf9 cell membranes

Compound	hH₂R-G_sαS		gpH₂R-G_sαS		EC₅₀hH₂R-G_sαS/EC₅₀ gpH₂R-G_sαS
Compound	Efficacy	EC₅₀	Rel. Pot.	Efficacy	EC₅₀hH₂R-G_sαS/EC₅₀ gpH₂R-G_sαS	EC₅₀	Rel. Pot.
		μM			μM
1(HIS)	1.00	1.26 ± 0.25	100	1.00	1.20 ± 0.24	100	1.05
2(DIM)	0.85 ± 0.07	1.94 ± 0.07^2-150	65.0	0.93 ± 0.04	1.20 ± 0.21	100	1.62
3(AMT)	0.90 ± 0.06	0.45 ± 0.04	280	1.04 ± 0.05	0.44 ± 0.04	271	1.02
4(BET)	0.73 ± 0.07	33.6 ± 7.2^2-150	4.00	0.73 ± 0.06	51.4 ± 8.3	2.00	0.65
5 (IMP)	0.84 ± 0.04^2-150	0.20 ± 0.02^2-150	641	1.00 ± 0.12	0.04 ± 0.01	3060	5.00
6(BU-E-42)	0.86 ± 0.04^2-150	0.42 ± 0.07^2-150	303	1.02 ± 0.08	0.07 ± 0.02	1670	6.00
7 (BU-E-43)	0.56 ± 0.05^2-150	0.28 ± 0.07	448	0.93 ± 0.15	0.19 ± 0.06	631	1.47
8(ARP)	0.79 ± 0.07^2-150	0.19 ± 0.04^2-150	659	1.02 ± 0.04	0.07 ± 0.01	1600	2.71
9 (BU-E-47)	0.77 ± 0.17^2-150	0.15 ± 0.06^2-150	844	1.07 ± 0.02	0.04 ± 0.00	3030	3.75
10(BU-E-48)	0.70 ± 0.10^2-150	0.14 ± 0.02^2-150	928	1.00 ± 0.02	0.03 ± 0.00	4080	4.67
11 (BU-E-75)	0.87 ± 0.15	0.33 ± 0.10^2-150	376	1.04 ± 0.03	0.05 ± 0.00	2470	6.60
12(BU-E-96)	0.78 ± 0.04^2-150	0.31 ± 0.14^2-150	404	0.99 ± 0.08	0.08 ± 0.02	1470	3.88
13 (D281)	0.51 ± 0.14^2-150	0.81 ± 0.13^2-150	155	0.87 ± 0.09	0.21 ± 0.09	570	3.86

Potencies and efficacies of ligands at hH₂R-G_sαS and gpH₂R-G_sαS were determined in the GTPase assay. GTP hydrolysis was determined as described under Experimental Procedures. Reaction mixtures contained Sf9 membranes expressing fusion proteins and agonists at concentrations from 1 nM to 1 mM as appropriate to generate saturated concentration/response curves. Curves were analyzed by nonlinear regression. Typical basal GTPase activities ranged between ∼1 and 2 pmol/mg/min, and typical GTPase activities stimulated by HIS (100 μM) ranged between ∼4 and 8 pmol/mg/min. To calculate agonist efficacies, the maximum stimulatory effect of HIS was set at 1.00, and the stimulatory effects of other agonists were referred to this value. Data shown are the means ± SD of four to six experiments performed in duplicate. The relative potency (Rel. Pot.) of HIS was set at 100, and the potencies of other agonists were referred to this value to facilitate comparison of agonist potencies with hH₂R-A271D-G_sαS, NgpChH₂R-G_sαS, and NhCgpH₂R-G_sαS(Table 4). Efficacies and potencies, respectively, of ligands at hH₂R-G_sαS were compared with the corresponding parameters at gpH₂R-G_sαS using the t test.

↵2-150 , p < 0.05.