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Materials and Methods (Conditioning procedure)

Subjects
Male C57BL/6J mice (8 weeks old) were purchased from Jackson Laboratories
(Bar Harbor, Maine). Mice were housed in groups of 5/cage with food and water adlibitum
and were acclimatized to the vivarium for one week before experiments began.
Animal care was in accordance with the Guide for the Care and Use of Laboratory
Animals (National Research Council, National Academy Press, 1996) and was approved
by the University of Miami Animal Care and Use Committee.
Drugs
Cocaine HCl and (+)-MK-801 hydrogen maleate (dizocilpine) were dissolved in
saline (0.9% NaCl). The NR2B antagonist ifenprodil [α-(4-Hydroxyphenyl)-β-methyl-4-
benzyl-1-piperidineethanol(+)-tartrate salt] was dissolved in distilled water. Traxoprodil
[(1S,2S)-1-(4-hydroxy-phenyl)-2-(4-hydroxy-4-phenylpiperidino)-1-propanol], another
NR2B-containing NMDAR antagonist (Chenard et al., 1995), and the nNOS inhibitor 7-
nitroindazole (7-NI) were dissolved in a 1:3:6 mixture of DMSO, polyethylene glycol
and distilled water, respectively. The MEK inhibitor SL327 was dissolved in 40% DMSO
(Atkins et al., 1998). All drugs were purchased from Sigma-Aldrich (St. Louis, MO,
USA). Drugs and vehicles were administered intraperitoneally (i.p.) in a volume of
0.1ml/10g.
Conditioning procedure
Place preference was monitored using custom-designed Plexiglas cages (Opto-
Max Activity Meter v2.16; Columbus Instruments) as previously described (Liddie et al.,
2012). The training context consisted of two compartments. One compartment had black
and white striped walls and a white floor covered with stainless steel grid; the other
compartment had black walls and smooth black floor. Each compartment was scanned by
7 infrared beams. A null zone 8 cm wide was assigned at the interface of the two
compartments to ensure that only full entry into each compartment is registered as ‘real’
time spent in each zone. On the first day, mice were habituated (15min) to the training
context; preconditioning compartment-preference/aversion was determined. Preconditioning
average times spent in the black, striped and null zones during 1200 seconds
were 456±12, 516±13 and 217±9 seconds, respectively. Half the subjects showed slight
preference for one side or the other. Accordingly, mice were paired with cocaine in the
less preferred compartment. Although this may be viewed as a biased design, half of the
mice were paired with cocaine in the black compartment and the other half were paired
with cocaine in the black and white-striped compartment making this design ‘partially
biased’. Following habituation, mice were conditioned over 4 days by a) 11.25mg/kg
(Fix-C) or b) 3, 6, 12 and 24mg/kg; one dose per day (Esc-C) as we previously described
(Itzhak & Anderson, 2012). Doses were chosen to control for total amount of cocaine
administered over 4 days. Post-conditioning average time spent in the null zone during
1200 seconds CPP test was 192±7 seconds. Likewise time spent in the null zone
following pharmacological treatments, pre- and post-CPP, was not significantly different
than vehicle treatment (201+9 seconds).

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