Endogenous activation of metabotropic glutamate receptors in neocortical development causes neuronal calcium oscillations

Abstract

] i increases in local cell clusters, and synchronous transients in cell pairs undergoing mitotic division (8). In the postnatal neocortex, coordinated [Ca 2ϩ ] i rises can occur in 3-100 adjacent neurons, termed neuronal domains (9). These spontaneous events are mediated by gap junctional communication (10) and appear to involve cell-to-cell diffusion of the second messenger inositol trisphosphate (9, 11). Neocortical domains apparently can occur independently of action potential activity, but their frequency can be increased by activation of metabotropic glutamate receptors (mGluR) (11) (19,20). Recent work has also shown that the frequency and duration of [Ca 2ϩ ] i oscillations can differentially influence the expression of specific genes (21-23). One known mechanism for the generation of [Ca 2ϩ ] i oscillations by cell-to-cell signaling involves activation of the mGluR5 subtype of the group I mGluRs, which are highly expressed in the immature neocortex (16, 24, 25).Activation of mGluR5 leads to the production of inositol trisphosphate by phospholipase C, which, in turn, leads to increases in [Ca 2ϩ ] i . Little is known, however, about the role that mGluR signaling plays in the physiology of developing neocortical neurons.To investigate the potential role of mGluR activation in generating spontaneous [Ca 2ϩ ] i oscillations in developing neocortical neurons, we performed a series of calcium imaging experiments using embryonic and postnatal mouse neocortical slices. We found that activation of the mGluR5 subtype of mGluRs leads to [Ca 2ϩ ] i oscillations in immature neurons, and that endogenous glutamate acting on group I mGluRs is responsible for spontaneous [Ca 2ϩ ] i oscillations in developing neocortex. MethodsTissue Preparation. Coronal slices of neocortex (300-400 m) were prepared from neonatal or embryonic Swiss-Webster mice (Taconic Farms) by using a Vibratome (Pelco, Redding, CA) as previously described (13). For embryonic slices, brains were embedded in 4% low-melting agarose (Fisher Scientific) in PBS with glucose (Life Technologies, Grand Island, NY) prior to slicing. Slices were prepared and incubated in standard artificial cerebrospinal fluid (ACSF), which contains 125 mM NaCl, 2.5 mM KCl, 1.25 mM NaH 2 PO 4 , 1 mM MgSO 4 , 2 mM CaCl 2 , 25 mM NaHCO 3 , and 20 mM glucose (pH 7.4), oxygenated with 95% O 2 ͞5% CO 2 . Following slice preparation, slices were incubated in ACSF at room temperature for 15 min prior to loading with the indicator dye.Fluo-3 Loading. Cells were loaded with the calcium indicator dye fluo-3 by bath application of fluo-3 acetoxymethyl ester (fluo-3-AM; Molecular Probes). Loading was performed for 1-4 hr at room temperature in ACSF containing 15 M fluo-3-AM, 0.0067% pluronic F-127 (Molecular Probes), and 0.33% DMSO (Sigma). The loading chamber was continuously oxygenated with 95% O 2 ͞5% CO 2 .Calcium Imaging. Measurements of relative changes in [Ca 2ϩ ] i were made using vital epifluorescence microscopy of mouse neocortical slices loaded with fluo-3. Fo...

Brain-derived Neurotrophic Factor-induced Potentiation of Ca2+ Oscillations in Developing Cortical Neurons

“…The spontaneous cortical activities are assumed to differ. They are mediated by activation of metabotropic glutamate (20) or GABA A (21) receptors or by Ca 2ϩ mediated by IP 3 through gap junctions (23). In this study, we found that synchronized spontaneous Ca 2ϩ oscillations occurred in cortical neurons obtained from postnatal 2-3-day-old rats after 5 days of in vitro culture (DIV 5).…”

“…In the cortex, it was reported that spontaneous oscillations were seen only in individual cells (20,21) from small groups consisting of 5-50 neurons (37) or in larger populations (24). The spontaneous cortical activities are assumed to differ.…”

“…In the cerebral cortex, several patterns of Ca 2ϩ activity have been observed during cortical development, and the metabotropic glutamate receptors (mGluR), GABA A receptors, gap junctions, and ionotropic glutamate receptors are all able to mediate them. The Ca 2ϩ oscillations mediated by mGluR are triggered by mGluR agonists in neonatal or embryonic cortical mouse slices, and this Ca 2ϩ activity is not synchronized in any population of cells (20). In the early postnatal stage, GABA A receptor activation has been shown to be involved in Ca 2ϩ oscillations, and again this Ca 2ϩ activity is not synchronized (21).…”

Activation of Metabotropic Glutamate Receptor mGlu5 on Nuclear Membranes Mediates Intranuclear Ca2+ Changes in Heterologous Cell Types and Neurons

“…Although mGlu1 and mGlu5 utilize similar signal transduction pathways, activation of mGlu1 receptors elicits a single Ca 2ϩ peak (30) whereas mGlu5 activation leads to a marked oscillatory response in heterologous cells (30 -32) as well as neocortical, hippocampal (33), and spinal cord neurons (34). It has been estimated that 90% of the Ca 2ϩ oscillations that occur spontaneously in immature cortical neurons can be ascribed to mGlu5 receptors (33).…”

“…It has been estimated that 90% of the Ca 2ϩ oscillations that occur spontaneously in immature cortical neurons can be ascribed to mGlu5 receptors (33). Inasmuch as Ca 2ϩ oscillations can differentially affect gene expression (35), they play an important role in the control of developmental events.…”