Resumo

Objectives. Pentylenetetrazole (PTZ) is able to succeed as an aversive unconditioned stimulus (US) in the rat olfactory fear conditioning (OFC) paradigm [Neurobiol Learn Mem. 91: 32-40, 2009; Neurobiol Learn Mem. 92: 512-18, 2009]. As the PTZ half-life lasts 116 ± 25 min [J Pharmacol Exp Ther. 234: 624-628, 1985], a time beyond the duration of experimental condition, we decided to investigate some biological marker that could underlie the best fit for PTZ function as an US for OFC. Neural structures such as the prefrontal cortex (PFC), the hippocampus (HIP) and the periaqueductal gray (PAG) appears to be involved in the mediation of conditioned fear responses, and we sought to register their electrographic (EEG) activity under the influence of PTZ treatment trying to find out the best temporal alignment with the exposure to the olfactory stimulus to be conditioned. Methods. The experimental procedures involved two independent sets of experiments performed in three-month-old male Long-Evans hooded rats: 1) measurement of EEG activity; and 2) OFC task. EEG recordings were performed through electrodes implants directed to PFC, HIP and dorsolateral PAG during 40 min after the systemic injection of PTZ (15 mg/kg). In the OFC paradigm, 2 consecutive phases were outlined: acquisition (2 days) and expression (3 days). The acquisition was performed in a chamber connected to a shock generator. On day 1, subjects were familiarized. On the second day, 10 or 30 min before the conditioning (2.5 min of duration) to the amylacetate (AA)-odor (CS), animals received a systemic injection of PTZ or saline associated with 3 pairings of footshock (0.4 mA/2s; 30 s inter-trial period). The expression phase took place in an odor box comprising an open and an enclosed communicating compartment, and consisted of 3 consecutive sessions (10 min each): familiarization (day 3; neutral odor); CS1-test (day 4; AA-odor) and CS2-test (day 5; neutral odor). The behavioral responses scored were the percentage of CS approach time (%AT) and of enclosed compartment hidden time (%HT). Results. The data analysis showed EEG alterations as high voltage slow waves (non-convulsant) mainly in CPF and HIP, starting at 364 ± 154 s after PTZ infusion. The total incidence of the discharges was 90 ± 15 s and medium duration of each alteration was 3.8 ± 0.4 s during the 40 min observation period. In Experiment 2, ANOVA followed by Duncan’s test showed that only rats paired (PTZ/AA) 10 min after the PTZ administration decreased %AT (control (C) = 22±5; PTZ/AA = 12±3) and increase %HT (C = 49±6; PTZ/AA = 71±8) during CS1-test. In addition, when rats were conditioned receiving three footshocks associated with PTZ, the conditioned responses exhibited was of greater magnitude than before (%AT= 8±2 and %HT= 81±5). OFC (associated or not with 3 footshocks) 30 min after PTZ, failed to express the defensive behavior associated to fear conditioning. Conclusions. The results showed that PTZ is able to succeed as an aversive US in the OFC, restricted to the latency to the first EEG activity. It highlights the usefulness of EEG and behavioral approaches to optimize procedures in the fear conditioning paradigm. Financial support: CNPq, CAPES, FAPESP.