Influx In Hippocampal Neurons Research Articles

Rapid effect of bisphenol A on glutamate-induced Ca2+ influx in hippocampal neurons of rats

Intracellular Ca2+ signaling plays an essential role in synaptic plasticity. This study examined the effect of BPA on concentration of intracellular Ca2+ ([Ca2+]i) by measuring fluorescence intensity of Ca2+ in hippocampal neurons in vitro. The results showed that BPA for 30 min exerted dose-dependently dual effects on glutamate-elevated [Ca2+]i: BPA at 1–10 μM suppressed but at 1–100 nM enhanced glutamate-raised [Ca2+]i. BPA-potentiated [Ca2+]i was blocked by the antagonist of NMDA receptor and was eliminated by an estrogen-related receptor gamma (ERRγ) antagonist rather than an AR antagonist. Both inhibitors of MAPK/ERKs and MAPK/p38 blocked BPA-enhanced [Ca2+]i. Co-treatment of BPA with 17β-E2 or DHT eliminated the enhancement of 17β-E2, DHT, and BPA in glutamate-elevated [Ca2+]i. These results suggest that BPA at nanomole level rapidly enhances Ca2+ influx through NMDA receptor by ERRγ-mediated MAPK/ERKs and MAPK/p38 signaling pathways. However, BPA antagonizes both estrogen and androgen enhancing NMDA receptor-mediated Ca2+ influx in hippocampal neurons.

VOLTAGE‐GATED SODIUM CHANNEL ACTIVATION TRIGGERS CALCIUM INFLUX AND HIPPOCAMPAL NEURON COMPLEXITY

Background and significanceAfter stroke, the molecular mechanisms underlying recovery are similar to the development of the nervous system. During development, neuronal activity regulates both the morphology and connectivity of neurons. Neuronal activity primarily involves voltage‐gated sodium channel (VGNaC) activation. Calcium signaling through the N‐methyl‐D‐aspartate receptor (NMDAR) drives morphological changes such as, dendritic arborization, synaptogenesis, and spinogenesis. VGNaC activation influences NMDAR function making this pathway an attractive therapeutic target for stroke recovery.HypothesisActivation of VGNaCs by PbTx‐2 augments NMDAR signaling resulting in and increased Ca2+ influx leading to dendritic arborization and spinogenesis in hippocampal neurons.Experimental design and MethodsWe used the VGNaC gating modifier, brevetoxin (PbTx‐2), to assess the effects of VGNaC activation on neuronal morphology. We defined the calcium‐signaling pathway triggered by PbTx‐2 in a population of hippocampal neurons (HNs), and evaluated PbTx‐2‐induced effects on dendritic arborization and spinogenesis in organotypic hippocampal slice culture (OHSC).HNscells were prepared from E18 Swiss Webster mouse embryos and plated in 96‐well plates. All experiments were run on (10 Days in vitro). A FlexStation2 was used to monitor PbTx‐2‐induced Ca2+ influx.OHSCSlices were prepared from P5, Thy‐1 YFP transgenic, mouse pups. Slices were treated with Pbtx‐2 18H after plating. A Leica SP8 confocal microscope was used to generate Z‐stack images of neurons or dendrites. Imaris‐XT software was used to reconstruct 3D‐images of the dendritic arbor, run Sholl analysis, and measure spine density.Results and ConclusionIn HNs, the EC50 for PbTx‐2‐induced Ca2+ influx was 376 nM (95% CI, 281–504 nM). The PbTx‐2 triggered Ca2+ influx in HNs was through the reverse mode of operation of Na+/Ca2+ exchanger, the NMDAR and the L‐type Ca2+ channel pathways. In OHSC, four‐day treatment with PbTx‐2 (100 nM) produced significant increases in dendritic arbor complexity as determined by Sholl analysis (p<0.005, ANOVA). PbTx‐2 treatment (300 nM) produced significant increases in spine density (p<0.001, ANOVA). Modifying the gating properties of VGNaC using PbTx‐2 enhanced the morphology of hippocampal neurons. Overall, these studies are consistent with the hypothesis that VGNaC activation may represent a novel pharmacological strategy to regulate NMDAR function and aid in recovery post‐stroke.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

MagR Alone Is Insufficient to Confer Cellular Calcium Responses to Magnetic Stimulation

Magnetic manipulation of cell activity offers advantages over optical manipulation but an ideal tool remains elusive. The MagR protein was found through its interaction with cryptochrome (Cry) and the protein in solution appeared to respond to magnetic stimulation (MS). After we initiated an investigation on the specific role of MagR in cellular response to MS, a subsequent study claimed that MagR expression alone could achieve cellular activation by MS. Here we report that despite systematically testing different ways of measuring intracellular calcium and different MS protocols, it was not possible to detect any cellular or neuronal responses to MS in MagR-expressing HEK cells or primary neurons from the dorsal root ganglion and the hippocampus. By contrast, in neurons co-expressing MagR and channelrhodopin, optical but not MS increased calcium influx in hippocampal neurons. Our results indicate that MagR alone is not sufficient to confer cellular magnetic responses.

MicroRNA-223 is neuroprotective by targeting glutamate receptors

Stroke is a major cause of mortality and morbidity worldwide. Extracellular glutamate accumulation leading to overstimulation of the ionotropic glutamate receptors mediates neuronal injury in stroke and in neurodegenerative disorders. Here we show that miR-223 controls the response to neuronal injury by regulating the functional expression of the glutamate receptor subunits GluR2 and NR2B in brain. Overexpression of miR-223 lowers the levels of GluR2 and NR2B by targeting 3'-UTR target sites (TSs) in GluR2 and NR2B, inhibits NMDA-induced calcium influx in hippocampal neurons, and protects the brain from neuronal cell death following transient global ischemia and excitotoxic injury. MiR-223 deficiency results in higher levels of NR2B and GluR2, enhanced NMDA-induced calcium influx, and increased miniature excitatory postsynaptic currents in hippocampal neurons. In addition, the absence of MiR-223 leads to contextual, but not cued memory deficits and increased neuronal cell death following transient global ischemia and excitotoxicity. These data identify miR-223 as a major regulator of the expression of GluR2 and NR2B, and suggest a therapeutic role for miR-223 in stroke and other excitotoxic neuronal disorders.

Estrogens directly potentiate neuronal L-type Ca 2+ channels

L-type voltage-gated Ca(2+)channels (VGCC) play an important role in dendritic development, neuronal survival, and synaptic plasticity. Recent studies have demonstrated that the gonadal steroid estrogen rapidly induces Ca(2+) influx in hippocampal neurons, which is required for neuroprotection and potentiation of LTP. The mechanism by which estrogen rapidly induces this Ca(2+) influx is not clearly understood. We show by electrophysiological studies that extremely low concentrations of estrogens acutely potentiate VGCC in hippocampal neurons, hippocampal slices, and HEK-293 cells transfected with neuronal L-type VGCC, in a manner that was estrogen receptor (ER)-independent. Equilibrium, competitive, and whole-cell binding assays indicate that estrogen directly interacts with the VGCC. Furthermore, a L-type VGCC antagonist to the dihydropyridine site displaced estrogen binding to neuronal membranes, and the effects of estrogen were markedly attenuated in a mutant, dihydropyridine-insensitive L-type VGCC, demonstrating a direct interaction of estrogens with L-type VGCC. Thus, estrogen-induced potentiation of calcium influx via L-type VGCC may link electrical events with rapid intracellular signaling seen with estrogen exposure leading to modulation of synaptic plasticity, neuroprotection, and memory formation.

17β-estradiol induced Ca 2+ influx via L-type calcium channels activates the Src/ERK/cyclic-AMP response element binding protein signal pathway and BCL-2 expression in rat hippocampal neurons: A potential initiation mechanism for estrogen-induced neuroprotection

Our group and others have demonstrated that 17β-estradiol (E2) induces neurotrophic and neuroprotective responses in hippocampal and cortical neurons which are dependent upon the Src/extracellular signal-regulated kinase (ERK) signaling pathways. The purpose of this study was to determine the upstream mechanism(s) that initiates the signaling cascade leading to E2-inducible neuroprotection. We tested the hypothesis that E2 activates rapid Ca 2+ influx in hippocampal neurons, which would lead to activation of the Src/ERK signaling cascade and up-regulation of Bcl-2 protein expression. Using fura-2 ratiometric Ca 2+ imaging, we demonstrated that E2 induced a rapid rise of intracellular Ca 2+ concentration ([Ca 2+] i) within minutes of exposure which was blocked by an L-type Ca 2+ channel antagonist. Inhibition of L-type Ca 2+ channels resulted in a loss of E2 activation of the Src/ERK cascade, activation of cyclic-AMP response element binding protein (CREB) and subsequent increase in Bcl-2. Real-time intracellular Ca 2+ imaging combined with pERK immunofluorescence, demonstrated that E2 induced [Ca 2+] i was coincident with ERK activation in the same neuron. Small interfering RNA knockdown of CREB resulted in a loss of E2 activation of CREB and subsequent E2-induced increase of Bcl-2 expression. We further demonstrated the presence of specific membrane E2 binding sites in hippocampal neurons. Together, these data indicate that E2-induced Ca 2+ influx via the L-type Ca 2+ channel is required for E2 activation of the Src/ERK/CREB/Bcl-2 signaling. Implications of these data for understanding estrogen action in brain and use of estrogen therapy for prevention of neurodegenerative disease are discussed.

Depression of synaptic transmission and evoked NMDA Ca2+ influx in hippocampal neurons by adenosine and its blockade by LTP or ischemia

AbstractNeuronal Ca2+ influx in response to repetitive synaptic activation was determined in rat hippocampal slices by measuring the evoked decreases of the extracellular Ca2+ concentration with ion‐sensitive electrodes in the synaptic layer (ΔCaSpad) and in the CA1 pyramidal cell layer (ΔCaSpyr). The generation of NMDA receptor‐mediated Ca2+ fluxes (NMDA Ca2+ fluxes) was uncovered by measuring the proportion of ΔCa blocked by the NMDA receptor antagonist 2‐amino‐5‐phosphonovalerate (APV); 1 μmol adenosine increased the critical input frequency required to generate synaptic MNDA Ca2+ influx. This effect is apparently exerted by limiting the postsynaptic membrane depolarization and is no longer seen, if the NMDA Ca2+ channels have lost their Mg‐dependent voltage sensitivity. It has been recently reported by Ben‐Ari and co‐workers [(1992) Trends Neurosci 15:333–339] that such a loss of voltage sensitivity, leading to an “upregulation” and persistent generation of NMDA Ca2+ currents, Can be elicited by an activation of the protein kinase C (PKC) and may be responsible for the initiation of synaptic long‐term potentiation (LTP) and for ischemia‐induced nerve cell damage. Consistent with this hypothesis, we found that the depressive effect of 1 μmol adenosine on the generation of NMDA Ca2+ influx or synaptic transmission can be no longer elicited in hippocampal slices after LTP and after preceding transient brain ischemia in vivo. These findings suggest that the mosaic of different adenosine actions includes some which are related to PKC activation. Accordingly, we observed a synaptic modulation by adenosine which was characterized by a reduced Mg‐sensitivity and blocked by phorbol ester treatment. © 1993 Wiley‐Liss, Inc.