Al., 1997), but many aspects of this response have not been effectively studied, as an illustration, the degree of neuronal activity required plus the consequences for brain function. Within this study, we cultured astrocytes in serum-free defined medium and examined the effects of development factors (GFs) and cytokines on calcium dynamics. While astrocytes happen to be studied by calcium imaging for 10 years, their reported pharmacological TrxR Storage & Stability properties and response pattern vary (transient or oscillatory), based on the species, brain region, and age on the animal (McCarthy and Salm, 1991; Muller et al., 1997; Cai et al., 2000), and this complexity tends to make it tough to decide the physiological function of those phenomena. We propose that this variability could reflect an important function of astrocytes, namely that they could respond to elements, for example ions, neurotransmitters, bioactive lipid metabolites, GFs, cytokines, and adhesion molecules, in the CNS environment and may alter the atmosphere by altering their morphology and metabolism, including the expression of enzymes and other factors (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 important in maintaining CNS homeostasis but would bring about important variation inside the outcomes of research on cultured astrocytes. Smaller differences in culture conditions, such as cell density, serum, plus the presence of other cell populations, impact astrocyte metabolism by the presence of soluble factors or by cell adhesion, top to further changes in the medium. Since of this synergistic effect, initial small differences can generate markedly various results. To prevent these troubles and to Pim Storage & Stability examine the real properties of astrocytes, it truly is essential to culture them in well defined situations, for which serum-free defined medium is ideal. The usage of defined medium makes it possible for the effect from the environment on the physiological properties of astrocytes to be systematically studied and ought to present essential new data on astrocyte function. Calcium oscillation is reported to create repetitive glutamate release from astrocytes that then impacts surrounding neurons (Pasti et al., 1997, 2001), suggesting a feedback mechanism in the astrocyte for the neuronal network. We consequently studied calcium oscillation in astrocytes cultured in defined medium and compared these responses with these in tissue slice preparations to ascertain irrespective of whether our culture system benefits could possibly be applied to in vivo research.Materials and MethodsCell culture and calcium imaging. Astrocytes had been isolated in the cerebral cortex of postnatal day 1 Wistar rats, applying a modification on the technique described by Levison and McCarthy (1991). Briefly, brain cells had been prepared from the cortices of 10 5 brains by trypsinization, trituration, and filtration, and seeded at 1.3 10 four cells/cm 2 in 75 cm 2 plastic flasks (Sumitomo Bakelite, Tokyo, Japan). Right 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) with a medium adjust every three d, the resulting mixed glia culture was shaken at 260 rpm for 18 hr at 37 after which rinsed with medium to eliminate nonastrocytic cells. The adherent cells were subcultured by trypsinization and seeded at a density of.