Abstracts

Rationale: Brain-derived neurotrophic factor (BDNF) appears to be associated with epileptogenesis. Danio rerio, popular named as zebrafish, has been recognized for epilepsy studies; however, the temporal transcript profile of the BDNF in the zebrafish brain after an acute seizure is unknown. The main aim of this study was to investigate the transcript profile of bdnf, ntrk2a and ntrk2b genes in adult zebrafish brain after a seizure. Methods: Adult zebrafish were separated in control group (CG, n=6) and seizure group (SG, n=6); a pool of two brains were used to compose one sample. Animals from SG were individually exposed to PTZ 15mM for 2-3 minutes and observed the three phases of epileptic behavior as previous described by Baraban et al. 2005. CG was submitted to the same handling condition and time as SG, but in PTZ-free water. Animals were anesthetized and their brains removed for RNA extraction, by TRIzol® protocol at 0.05h, 12h, 24h, 48h and 72h after seizure. Quality of RNA was assessed by spectrophotometry and agarose gel electrophoresis. Reverse transcriptase quantitative-PCR using TaqMan (Applied Biosystems) assays was performed to evaluate the transcript profile of bdnf and its receptors ntrk2a and ntrk2b genes. Runs were carried out in triplicate using 18S as endogenous control (Applied Biosystems). The relative quantification (RQ) was calculated by the equation RQ=2 -ΔΔCT. This study was approved by animal ethical committee of our Institution. Results: Comparisons between CG and SG showed an increase of bdnf mRNA levels at 0.05h after seizure, RQCG = 0.27±0.01 and RQSG = 0.67±0.20. No difference was found for ntrk2b mRNA levels at the same time (RQCG = 0.40±0.03 and RQSG= 0.31±0.06), but a slight down regulation was found to ntrk2a (RQCG=0.26±0.02 and RQSG= 0.17±0.02). At 24h and 48h, an up-regulation of bdnf and ntrk2b was found in SG group (bdnf 24h: RQCG = 0.59±0.10 and RQSG= 1.02±0.27; bdnf 48h: RQCG = 0.43±0.07 and RQSG= 1.03±0.16; ntrk2b 24h: RQCG = 0.60±0.07 and RQSG= 1.16±0.26; ntrk2b 48h: RQCG = 0.35±0.06 and RQSG= 0.83±0.18). There are no difference between groups at 12h and 72h. The relationship between ligand and receptors showed that the transcript of bdnf gene is more abundant than ntrk2a and ntrk2b transcripts only immediately of seizure (bdnf RQSG = 0.67±0.20; ntrk2a RQSG = 0.17±0.02; ntrk2b RQSG = 0.31±0.06). Conclusions: It has been reported that both BDNF transcript and protein levels are increased after seizures in patients and experimental models. In rats, Bdnf transcript levels have returned close to the control levels 12h after seizure. Our results have yielded insights into a different pattern of transcript profile of the bdnf gene and its receptors after seizure insult. Support: CNPq and FAEPEX. Reference: Baraban SC, Taylor MR, Castro PA, Baier H. Pentylenetetrazole-Induced Changes in Zebrafish Behavior, Neural Activity, and c-fos Expression. Neuroscience 2005; 131:759-768.