APOPTOSIS AND PATHWAYS INVOLVED IN A PRENATAL ACUTE HYPOXIA MODEL

S.FISZER DE PLAZAS; S.GIUSTI; A.MITRIDATE DE NOVARA; M.VACOTTO

San Diego, California

Congreso; Society for Neuroscience 2004, 34th Annual Meeting; 2004

Society for Neuroscience

JNK activation triggers cell death due to apoptosis, which may be accompanied by activation of the caspase pathways. The aim of this work was to determine the appearance of apoptosis and molecular alterations taking place in the JNK and caspase-3 pathways, following prenatal acute hypoxia in chick embryos (60 in, 8% O2). Accordingly, at embryonic day (ED) 12two methodologies were applied evaluated at different times post-hypoxia (p-h): 1) detection of death due to apoptosis on optic lobe sections, by means of TUNEL fluorescence; and 2) determination of JNK and P-JNK levels, as well as that of active caspase-3, using Western blot. TUNEL assays disclosed that hypoxia leads to a significant increase in apoptotic cells, evidenced at 60 min. and peaking at 6 h p-h (50 and 200%, respect to control). Determinations by western blot showed that at 0 min p-h there was a significant increase in JNK and P-JNK levels. P-h analysis exhibited a drop in JNK over time, whereas peak P-JNK levels were reached at 10 and 30 min, with a P-JNK/JNK ratio of 1.912±0.304, respectively (p<0.01). Active caspase-3 levels at 0, 10 and 30 min p-h remains unchanged vs. control, reaching peak values at 60 and 120 min p-h, of 1.495±0.206 and 1.504±0.273, respectively (p<0.01). To conclude, results at molecular level suggest that at ED 12 one of the possible mechanisms, triggered by hypoxia would be a process of programmed cell death, correlating with prior activation of pro-apoptotic proteins such as JNK and caspase-3.