Table 1

Receptor binding properties of TIPP, DPDPE, and ICI-174864 in the absence or presence of GppNHp/NaCl in GH₃DORT membranes or in digitonin-permeabilized cells

GH₃DORT	TIPP		DPDPE		ICI-174864
GH₃DORT	K_i	Uncoupled/Coupled Ratio	K_i	Uncoupled/Coupled Ratio	K_i	Uncoupled/Coupled Ratio
	nM		nM		nM
Membranes
−GppNHp/NaCl (coupled)	3.02 ± 0.16		14.5 ± 2.22		304 ± 73.2
⁺GppNHp/NaCl (uncoupled)	1.16 ± 0.11^1-150	0.384	418 ± 51.2^1-150	28.8	18.8 ± 3.8^1-150	0.062
Permeabilized
−GppNHp/NaCl (coupled)	4.77 ± 1.39		13.2 ± 5.6		680 ± 46.1
⁺GppNHp/NaCl (uncoupled)	0.42 ± 0.09^1-150	0.088	379 ± 61.1^1-150	28.7	83.8 ± 5.46^1-150	0.123

Competitive inhibition of [³H]diprenorphine binding (1.0 nM) in GH₃DORT membranes or in permeabilized cells by TIPP, DPDPE, or ICI-174864 (10⁻¹²–10⁻⁵ M) in the presence or absence of GppNHp (25 μM) and NaCl (100 mM) was performed as described under Experimental Procedures. Affinity (K_i) values are expressed as mean ± S.E.M. and were calculated from IC₅₀ values derived from complete concentration-effect curves using the Cheng-Prusoff equation (see Figs.1 and 5). The K_d for diprenorphine used to calculate the K_i values in membranes was 0.71 ± 0.04 nM (−GppNHp/NaCl) and 1.06 ± 0.14 nM (⁺GppNHp/NaCl). TheK_d for diprenorphine in permeabilized cells was 1.29 ± 0.11 nM (−GppNHp/NaCl) and 0.93 ± 0.10 nM (⁺GppNHp/NaCl). Data for each value presented represent three separate experiments performed in duplicate.

↵1-150 Significantly different from corresponding −GppNHp/NaCl (coupled)(P < 0.05, Student's t test).