Microdialysis

Approximately 12–18 hours prior to the microdialysis experiment, rats were lightly anesthetized with isoflurane to implant the microdialysis probe through the guide cannula and to secure the animal to the tethering apparatus. The probes (1.5 mm active membrane length, 270 μm OD, 13,000 MWCO) were constructed in the laboratory according to the procedures described by Pettit & Justice (1991). Probes were continuously perfused with artificial cerebrospinal fluid (ACSF; 149 mM NaCl, 2.8 mM KCl, 1.2 mM CaCl₂, 1.2 mM MgCl₂, 0.2 mM ascorbic acid, and 5.4 mM D-glucose) overnight at 0.2 μL/min. The flow rates were increased to 2.0 μL/min at least 2 hours prior to dialysate sample collection and remained at 2.0 μL/min for the duration of the experiment.

Animals were awake and freely moving during the microdialysis experiments. In all experiments, the sample collection interval was 5 minutes and four baseline samples were collected per animal prior to any infusions. Baseline dopamine concentrations were required to have a relative standard deviation <0.16 for data inclusion. To confirm that the dopamine in the dialysate samples was due to calcium-dependent exocytotic release, probes were perfused with calcium-free ACSF for approximately 2 hours at the conclusion of all experiments and additional samples were collected. All dialysate samples were immediately frozen on dry ice upon collection.