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|Title:||Immunohistochemical protocol to identify glial fibrillary acid protein (GFAP) in the dorsal horn of the spinal cord|
|Author:||Silva, Greice Anne|
Machado, Nathalia Leilane
Fazan, Valeria Paula
|Abstract:||Pain in neonates can induce long‐term changes in areas responsible for processing nociceptive stimulus . Astrocytes are active in supporting and controlling neuronal signaling and in response to noxious stimuli they may undergo morphological changes, such as hypertrophy and increased expression of glial fibrillary acid protein (GFAP). We showed that rats receiving painful stimuli in the neonatal period presented changes in sensory, motor and limbic cortical areas related to nociception even after 180 days after the stimulus . Alterations on the dorsal horn of the spinal cord in this model of neonatal pain were not yet investigated and to do so, we are testing a GFAP immunostaining protocol. Four male Wistar rats at 15 and 180 days of age were anesthetized and perfused with 4% buffered paraformaldehyde. Transverse sections of the spinal cord were cut at L4‐L6 levels and frozen. Cryosections (20 µm) were incubated in primary antibody polyclonal rabbit anti‐GFAP for 4 hours (1:1.500) followed by polyclonal donkey anti‐rabbit IgG for 45 minutes (1:250). Slides were washed in PBST, mounted in a mixture of glycerol/PBS (3:1) and observed with a light microscope (TS‐100, Nikon, Tokyo, Japan) equipped with a high resolution camera. Our results show that the method is reliable and reproducible for GFAP identification and quantification in the spinal cord of normal rats. Future studies will use this method to better understand the spinal cord response to noxious stimulus in the neonatal period.  Sanada et al 2014, Int J Dev Neurosci 35,55|
|Editor:||Federation of American Societies for Experimental Biology|
|Appears in Collections:||IB - Artigos e Outros Documentos|
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