Agonist‐evoked Ca2+ transients in primary astroglial cultures–modulatory effects of valproic acid

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Abstract
Cytosolic Ca2+ ([Ca2+]i) activity was measured in individual type 1 astroglial cells in primary culture after exposure to glutamate (Glu), quisqualate (QA), γ‐aminobutyric acid (GABA), 5‐hydroxytryptamine (5HT), and noradrenaline (NA) by using the Ca2+ indicator dye fura‐2/AM in a computerized microspectrofluorimetric system. Various patterns of Ca2+ transients were observed, but the most common was biphasic, having an initial sharp peak, rising immediately after stimulation, and then declining to a lower but sustained Ca2+ level. The only substance that diverged from this pattern was GABA, which induced a Ca2+ response with longer latency and a single‐phase curve. The effects of the anticonvulsive drug Na+‐valproate (VPA) were also investigated. After both acute and chronic (5–7 days) exposure to 10−4 M VPA, the GABA‐evoked rises in [Ca2+]i were completely inhibited. VPA also had acute effects on the 5HT‐ and Glu‐evoked Ca2+ spikes. The Ca2+ responses after 5HT stimulation were greatly reduced after exposure to 10‐4 M VPA. The responses after glutamate stimulation were, on the contrary, increased after a similar exposure. No VPA effects were seen on the curve patterns of QA and NA stimulations. The most frequent agonist‐evoked responses were seen after stimulation with 5HT and NA, where over 80% of the tested cells responded. For QA and Glu, the response frequencies were about 40% each, while for GABA it was 20%. The responses after 5HT and NA stimulation were blocked to baseline levels after exposure to ketanserin (5HT2 receptor antagonist) and a combination of prazosin, yohimbine, and propranolol (α1, α2, and β adrenoceptor antagonists, respectively). The rises in [Ca2+]i after stimulation with QA, Glu, and GABA were partly antagonized by 6‐nitro‐7‐cyano‐quinoxaline‐2,3‐dione (CNQX; a QA receptor antagonist, also tested on the glutamate responses) and bicuculline (GABA receptor antagonists). The biphasic curve forms, after stimulation with Glu, QA, 5HT, and NA, were reduced to single‐phase curves with unaffected or slightly reduced initial peaks when the stimulations were performed in Ca2+‐free medium. Interestingly, in some cells Glu induced a curve form that, different from the other Glu‐evoked responses, consisted of an immediate rise in [Ca2+]i, which persisted at the level of maximal response. Furthermore, this response was completely inhibited in Ca2+‐free buffer. The GABA response was also totally inhibited in Ca2+ ‐free buffer.The results suggest that astrocytes in primary culture respond differently to various receptor agonists, with regard to both the intracellular Ca2+ spiking behavior and the response frequency. Furthermore, the data also show that certain agonist‐evoked rises in [Ca2+]i can be significantly modulated by the neuroactive agent VPA.