TABLE 3.

Potency for substrates and inhibitors in competing for [³H]nisoxetine binding in hNET mutants

Data were calculated as a percent of total [³H]nisoxetine binding in the absence of competing substrate or inhibitor and analyzed by nonlinear regression. Data displayed in the table are IC₅₀ values with confidence intervals (CI) given within parenthesis. The number of experiments (n) is given under the ligand. Statistical significance was determined by an F test by comparing fits in which selected values were constrained to be equal or were allowed to differ. The null hypothesis was that the best fit parameter for the value did not differ. A conclusion of statistical significance represents a rejection of the null hypothesis and indicates a difference between designated values. F test values and degrees of freedom are given only in cases of statistical differences.

Ligands	IC50 (95% CI)
	Room Temperature			4°C
	hNET	T58A-hNET	T58D-hNET	hNET	T58A-hNET	T58D-hNET
	nM
Substrates
DA	490	370	35*	384	89*†	44*
(n = 3)	(230–1000)	(140–1000)	(3–430)	(170–850)	(52–160)	(3–60)
			F1,23 = 7.474*		F1,30 = 6.845*	F1,20 = 4.814**
					F1,26 = 4.958†
NE	308	745*	141	527	427	N.S.
(RT, n = 3)	(139–681)	(527–1051)	(28–697)	(182–1524)	(177–1031)
(4°C, n = 6)		F1,30 = 5.287*
AMPH	182	102	16*	2.7†††	4.6†††	5.3
(RT, n = 3)	(58–565)	(39–270)	(5.2–47)	(1.5–4.7)	(3.4–6.3)	(2.2–12)
(4°C, n = 6)			F1,30 = 6.703*	F1,63 = 31.64†††	F1,63 = 39.03†††
Inhibitors
Cocaine	1122	1680	1340	858	1208	1171
	(383–3280)	(793–3960)	(416–4340)	(678–1080)	(496–2943)	(200–6900)
Desipramine	10	12	6	18	12	18
	(5.9–18)	(6.2–23)	(1.2–27)	(14–23)	(9–16)	(5–67)
Benztropine	3290	4088	4990	N.D.	N.D.	N.D.
	(2280–4760)	(2626–6365)	(1848–13,480)

N.S., no significant binding.

^*P < 0.05 compared with wild-type value.

^**P < 0.01 compared with wild-type value.

***P < 0.001 compared with wild-type value.

^†P < 0.05 compared with values at RT.

^††P < 0.01 compared with values at RT.

^†††P < 0.001 compared with values at RT.