Metabolic mapping of rat brain activity associated with conditioned fear extinction and renewal, and improvement of extinction memory by the metabolic enhancer methylene blue

Changes in brain metabolism associated with the consolidation, extinction and recall of fear memories were investigated in rats using two complementary brain metabolic mapping approaches. First, fluorodeoxyglucose (FDG) metabolic mapping technique was used to track the stimulus-evoked changes in brain glucose uptake that mostly occur during the first ten minutes following the FDG administration. Second, cytochrome oxidase (CO) histochemistry, which is wellsuited for tracking long-term changes in brain metabolic capacity, was utilized. By combining these two techniques, brain structures involved in fear extinction memory consolidation and retention were compared to brain regions that displayed altered metabolic activity during conditioned fear memory recall. Additionally, since memory consolidation requires expenditure of energy, enhancement of brain oxidative phosphorylation through CO activity increase was tested as a possible way for improving extinction memory retention in rats. Low doses of the metabolic enhancer methylene blue (MB) were used to enhance CO activity in the post-extinction training period, to test the hypothesis that neurons with high metabolic demand which are engaged in consolidation and retention of the extinction memory would benefit most from the presence of a metabolic-enhancing drug. The results suggest that during conditioned fear renewal, the auditory conditioned stimulus activates the neural representation of the footshock unconditioned stimulus, thus supporting Pavlov’s stimulussubstitution model of classical conditioning. Quantitative CO histochemistry revealed that Pavlovian fear acquisition training increased metabolic capacity in several brain regions, including medial prefrontal cortex (mPFC) and septum, while extinction training reduced CO activity to levels comparable to the pseudorandom group. A functional neural network model of extinction explored how the direct influences on regions such as mPFC and amygdala might change between fear extinction recall and fear renewal. Finally, the third experiment illustrated that MB might be a useful adjunct to exposure therapy, since it improved consolidation and retention of fear extinction in our animal model of specific phobias.