Mon. May 20th, 2024

Al., 1997), but quite a few elements of this response have not been properly studied, for example, the degree of neuronal activity expected and also the consequences for brain function. In this study, we PTEN Species cultured astrocytes in serum-free defined medium and examined the effects of growth elements (GFs) and cytokines on calcium dynamics. Despite the fact that astrocytes have already been studied by calcium imaging for 10 years, their reported pharmacological properties and response pattern vary (transient or oscillatory), depending on the species, brain area, and age on the animal (McCarthy and Salm, 1991; Muller et al., 1997; Cai et al., 2000), and this complexity makes it difficult to ascertain the physiological role of these phenomena. We propose that this variability could reflect an essential function of astrocytes, namely that they are able to respond to things, such as ions, neurotransmitters, bioactive lipid metabolites, GFs, cytokines, and adhesion molecules, in the CNS environment and can alter the atmosphere by altering their morphology and metabolism, like the expression of enzymes as well as other components (Rostworowski et al., 1997;Morita et al. Dual Regulation of Astrocytic Calcium OscillationJ. Neurosci., November 26, 2003 23(34):10944 0952 Stachowiak et al., 1997; Krushel et al., 1998; Verkhratsky et al., 1998; Xian and Zhou, 1999; Bezzi et al., 2001). These responses are critical in keeping CNS homeostasis but would cause significant variation inside the outcomes of research on cultured astrocytes. Modest differences in culture situations, which include cell density, serum, and also the presence of other cell populations, influence astrocyte metabolism by the presence of soluble things or by cell adhesion, leading to further alterations in the medium. Due to the fact of this synergistic impact, initial small differences can generate markedly different outcomes. To avoid these troubles and to examine the true properties of astrocytes, it really is essential to culture them in well defined situations, for which serum-free defined medium is perfect. The use of defined medium enables the effect from the environment on the physiological properties of astrocytes to become systematically studied and should really offer crucial new details on astrocyte function. Calcium oscillation is reported to produce repetitive glutamate release from astrocytes that then affects surrounding neurons (Pasti et al., 1997, 2001), suggesting a feedback mechanism in the astrocyte towards the neuronal network. We for that reason studied calcium oscillation in astrocytes cultured in defined medium and compared these responses with those in tissue slice preparations to figure out no matter whether our culture technique final results might be applied to in vivo studies.Materials and MethodsCell culture and calcium imaging. Astrocytes were isolated from the cerebral cortex of Hexokinase Purity & Documentation postnatal day 1 Wistar rats, applying a modification of the method described by Levison and McCarthy (1991). Briefly, brain cells had been ready in the cortices of 10 5 brains by trypsinization, trituration, and filtration, and seeded at 1.3 ten 4 cells/cm 2 in 75 cm two plastic flasks (Sumitomo Bakelite, Tokyo, Japan). Soon after culture for 12 d at 37 in 5 CO2 humidified air in basal Eagle’s medium containing 10 fetal calf serum (FCS) (Equitech-Bio, Ingram, TX) having a medium alter every 3 d, the resulting mixed glia culture was shaken at 260 rpm for 18 hr at 37 after which rinsed with medium to remove nonastrocytic cells. The adherent cells were subcultured by trypsinization and seeded at a density of.