Neurosphere Culture From Adult Guinea Pig Superficial Cortex

Objective:The aims of this study are to determine whether cells isolated from adult guinea pig superficial cortex may form neurospheres in vitro and to determine their proliferation and differentiation ability.Methods:Adult male guinea pigs were used. The superficial cortex (approximately layers Ⅰ and Ⅲ), deep cortex (approximately layers IV-WM) and hippocampus were disassociated in DMEM/F12solution. Cells were then cultured, propagated and passaged in the media without serum but composed of EGF and bFGF. Then neurospheres were induced to differentiate in culture medium with serum and NGF. Neurospheres and differentiated cells were identificated by immunofluorescence. The electrophysiological properties of cells differentiated from neural stem/progenitor cells were characterized by patch-clamp recording.Results:Cells isolated from superficial cortex, deep cortex and hippocampus of adult guinea pig have the potency to form neurosphere-like cell clusters. Cloning formation rate of cells in hippocampus decreased with ages of guinea pigs, and there is no obvious effect of age on the cloning formation rate of cortex. Immunohistochemistry revealed that cells of neurospheres expressed Nestin, and some coexpressed GFAP. Neurosphere cells were found to differentiate into neurons (MAP-2positive or (3-III Tubulin positive) and astrocytes (GFAP positive). Mature Na+-K+currents and continuous action potential could be recorded in most of neurons by patch-clamp.Conclusion:Cells from superficial cortex of adult guinea pig have the ability to self-renew and to develop into astrocytes and functional neurons of Central Nervous System. Superficial cortex may be a putative neurogenic niche in adult guinea pig cerebrum.Cloning formation rate of cells in hippocampus decreased with ages of guinea pigs, and there is no obvious effect of age on the cloning formation rate of cortex.