Materials and Reagents

1. Custom made T-maze, made of Acrylic glass GS black (Bilek + Schüll, Vienna, Austria)
   
2. Male Sprague-Dawley rats (12-13 weeks old)
   Note: They were bred and maintained in the Core Unit of Biomedical Research, Division of Laboratory Animal Science and Genetics, Medical University of Vienna.
   
3. Pharmacological agents
   Note: They were applied in the experimental room since taking the animals to a different location causes novelty stress, that may potentiate the brief stress of intraperitoneal injection that we conducted.
   
4. Standard maintenance food (ssniff, R/M-H, Soest, Germany)
5. Food reward: e.g., food pellets were provided (Dustless Precision Pellets^®, Rodent purified diet, 45 mg; Bio-Serv, catalog number: F0021 )
6. Incidin^® Extra N (Ecolab, catalog number: 30 125 30 , PZN 002 357 95)
   
7. 1% incidin (see Recipes)
   

Equipment

1. Three lamps (LED chip, 20 W) for indirect illumination (mounted on a stand placed 1.3-1.5 m above the floor directed to the ceiling); Illumination within arms: 40-50 lux
   
2. Recording camera (video camera, ABUS AUGUST BREMICKER SÖHNE KG, model: EyseoEcoLine, catalog number: TV7003 )
   
3. T-maze
   For the apparatus see Figure 1. Reward food pellets were placed outside the T-maze scattered over the table to mask olfactory cues during training. Visual cues (equipment, walls and doors, Figure 1B) were identifiable around the maze. Additional cues like paper printouts of black and white figures were placed on room walls two meters above the floor (Sánchez-Santed et al., 1997). The maze was cleaned with 1% Incidin after the training of each animal to remove olfactory cues. Indirect illumination (from floor to ceiling) provides equal light intensities (40-50 lux) in each arm. Trials were monitored with a camera (mounted on the ceiling directly above the maze- and videos stored on a PC. A freely available video capture program (Debut Video Capture) was used to store the videos on a computer. Paper printouts of figures (210 mm x 279 mm) placed at room walls and equipment served as visual cues.
   
   
   Figure 1. T-maze placed on a desk (A) and printouts of external cues placed on experimental room walls (B). Two-goal arms (50 cm long, 10 cm in width, with walls of 25 cm height) and the starting arm (70 cm) could be separated by a guillotine door (A). The maze was placed on a table with a height of 80 cm. The start arm was equipped with a starting box (20 cm in length) separated from the maze by a guillotine door. At the end of each goal arm, reward food pellets were provided in a small plastic cup (30 mm in diameter and 12 mm in height) to mask the food pellet.
   

Procedure