Synaptic Modifications in Neostriatum Induced by Perinatal Asphyxia: 2-D and 3-Electron Microscopy Study

CAPANI F ; GE SARACENO; M HOLUBIEC; S BADORREY; L GIRALDEZ

Rio de Janeiro

Conferencia; International Congress of Electron Microscopy; 2011

IMC17

ynaptic Modifications in Neostriatum Induced by Perinatal Asphyxia: 2-D and 3-Electron Microscopy Study. F. Capani1,  G. E. Saraceno1,  M. Holubiec1,  S. Badorrey 1, Juan Romero,  D. Madureira de Oliveria 2,  G.  Barreto 3;  L. D. Giraldez Álvarez2;1Instituto de Investigaciones Cardiológicas “Prof. Dr. Alberto C. Taquini” (ININCA),  UBACONICET. 2Laboratório de Neuroquímica e Biología Celular, Instituto de Ciências da Saúde, Universidade. Federal da Bahía (UFBA), Campus do Canela, 40110-100, Salvador, Bahía, Brasil.  3 Department of Anesthesia. Stanford University School of Medicine, Stanford, CA. E-mail: fcapani@fmed.uba.ar Synaptic dysfunction has been associated with neuronal cell death following hypoxia. The lack of knowledge on the mechanisms underlying this dysfunction prompted us to investigate the morphological changes in the postsynaptic densities (PSDs) induced by hypoxia using both two dimensional (2D) and three dimensional (3D) electron microscopic analyses and correlative light and electron microscopy fluorescence. For this study we have used a perinatal asphyxia model (PA) Full-term pregnant rats on gestational day 22, were anesthetized, rapidly decapitated and the uterus horns were isolated through an abdominal incision and placed in a water bath at 37 °C for different time of asphyxia and intraasphytic hypothermia. When their physiological conditions improved, they were given to surrogate mothers Adult male of 1 month old and 6 month old were subjected to an intracardiac perfusion using a mix of paraformaldehyde and glutaraldehyde under deep anaesthesia. Short tem changes for actin in the synapses were studied using photooxidation with phalloidin-eosin as is described in (1). For long term changes we combined a correlative ethanolic phosphotungstic acid (E-PTA) and immunolectron microscopy for ubiquitin staining and electron tomography and 3-D reconstruction techniques (2) Using two dimensional (2D) electron microscopic analyses, an increase in the F-actin staining in neostriatum of 30 days-old asphyctic rats was observed at confocal and electron microscopic level using correlative fluorescent electron microscopy. Mushroom-shaped spines showed the most consistent staining. After six months of PA, PSDs of the rat neostriatum are highly modified and ubiquitinated 6 months after induction of PA., We observed an increment of PSD thickness dependent on the duration and severity of the hypoxic insult. Using 3-d reconstruction and electron tomography we observed showed clear signs of damage in the Asphyctic.  All of these changes were blocked hypothermia. These changes are correlated with intense staining for ubiquitin. Overall results demonstrate that synaptic dysfunction following PA might be produced by early changes in the actin organization and long-term misfolding and aggregation of proteins in the PSDs. We hypothesize that the early changes in the PSDs of the rat neostriatum might be correlated with the important modification and ubiquitization of the PSDs observed late in asphyctic adult rats. In addition hypothermia appears as a potential therapeutic tool (3). (1) Capani F. et al, J Comp Neurol., 435 (2001) 156-170(2) Capani F. et al, J Histochem Cytochem., 49 (2001) 1351-1361.(3) Capani F. et al, Exp Neurol., 219 (2009) 404-413 (4) Supported by UBACYT M407, CONICET 5784, DA0810