Graphic state 4

The attentional set-shift task (AST) was conducted in rat operant boxes (Coulbourn Instruments, Whitehall, PA, USA), containing a food magazine port between two levers, which were located below two lights on the front panel. A house light was positioned on the rear wall. Operant boxes were individually stored within a sound-attenuating chamber with a fan for aeration and noise dampening. The input and output from each box were transmitted to Graphic State 4 software (Coulbourn Instruments) on an Optiplex 9020 computer.

Behavioral testing was conducted in eight standard operant chambers (Coulbourn Instruments). Chambers were contained in sound-attenuating cubicles, and were computer controlled through Graphic State 4.0 software (Coulbourn Instruments). Locomotor activity was monitored via infrared motion detectors installed on the ceilings of the chambers. Each operant chamber was equipped with a food trough containing a photobeam sensor to detect nosepokes into the trough, two retractable levers (one on each side of the food trough), a feeder installed on the outside wall of the chamber and connected to the food trough to deliver 45 mg purified ingredient rodent food pellets (Test Diet 5TUL), and a stainless steel floor grate connected to a shock generator that could deliver scrambled footshocks. Each sound-attenuating cubicle included a house light mounted on the rear wall (outside of the operant chamber).

Behavioral testing was conducted in 12 operant test chambers (Coulbourn Instruments), housed in sound-attenuating cabinets. Each chamber contained a food pellet delivery trough equipped with a photobeam to detect entries and a 1.12 W lamp to illuminate the trough, into which 45 mg soy-free food pellets (TestDiet; 5TUM) could be dispensed. The trough was centrally located in the front of the chamber and ∼2 cm above the floor. Each chamber contained two retractable levers 11 cm above the floor on both the left and right sides of the food trough. An additional 1.12 W house light was mounted to the top of the rear wall of the cabinets. The floor of each chamber was composed of a row of stainless steel bars coupled to a shock generator (Coulbourn Instruments), which was used to administer scrambled footshocks. An activity monitor was mounted on the ceiling of the chamber to detect locomotor activity through the use of an array of infrared detectors. The test chambers were controlled via Graphic State 4.0 software (Coulbourn Instruments), which allowed programmable steps for each task protocol and collection of data from the operant chambers. In each cohort, the order in which young and aged rats were tested in each chamber was counterbalanced, and chambers were cleaned with dilute chlorhexidine between successive rats.

Animals of the second cohort were tested for their performance on both Barnes maze and passive avoidance tasks. To assess spatial reference memory, mice were trained on Barnes maze for 5 days before surgery, as previously described [19 (link)], then tested again on days 3 and 7 after reperfusion for the time needed to escape into the hole, the number of error pokes, and the length of the animal’s path prior to escape. An automated passive avoidance apparatus (Coulbourn Instruments) was used to assess avoidance learning with automated sensing and shock systems (GraphicState® 4, Coulbourn Instruments). The apparatus included a double compartment chamber with one lit and one dark compartment. Mice were allowed to explore the chamber for 5 min on habituation phase. Following habituation, the mice were given one trial where a shock is associated with the dark side, allowed 48 h of rest, and then tested for retention measured as latency to enter the dark side. Testing was repeated on days 3 and 7 post-reperfusion with no shock delivered during test phase.