Fluorescence detection of Europiumdoped very small superparamagnetic iron oxide nanoparticles in murine hippocampal slice cultures

In the late 1980s,superparamagnetic iron oxide nanoparticles（SPIO）moved into focus as contrast agents in magnetic resonance imaging（MRI）,due to their strong relaxivity and resulting higher resolution of images.At the time,no one anticipated their high potential in basic research or for medical diagnostic andtreatment. Since then, SPIO have been evaluated notonly as spe- cific markers for MRI, but also for cell labeling and tracking （Li et al., 2013）.

Propofol enhances the field excitatory postsynaptic potentials in CA1 hippocampal slices of young and aged mice

Background Increasing age was shown to decrease the requirements for propfol.However,the mechanisms of ageing-induced potentiation of anesthetic actions have not been clearly explored.The aim of this study is to compare the effects of propofol on the field excitatory postsynaptic potentials （fEPSPs） in hippocampal slices of young and aging mice.Methods Brain slices were prepared from C57BL6 male young （2 months） and aging （＞12 months） mice.The dendritic field excitatory postsynaptic potential was recorded from the CA1 stratum radiatum using patch clamp electrophysiological methods.A bipolar concentric stimulating electrode was placed along the Schaffer collateral for othodromic stimulation.The effects of clinically-relevant concentrations of propofol were studied in the young and ageing mouse tissues.Results Propofol application increased the orthodromically evoked fEPSP produced in slices taken from young and older animals.A striking feature in the I/O relationship was the decreased enhancement of the fEPSPs by propofol in slices from older mice.A clinically relevant concentration of propofol,10 μmol/L,showed more significant enhancement in amplitude and area under the curve （AUC） of fEPSP in young compared to tissues from older mice （amplitude：young （24.9±3.4）％,old （4.6±1.6）％; AUC young （30.6±5.4）％,old （2.1±1.7）％）.There was no statistically significant difference between the paired-pulse facilitation （PPF） ratios calculated for the responses obtained in tissues from young mice.In slices from older mice,in the presence of 10 μmol/L propofol,PPF was decreased and returned to baseline after washout （baseline 1.21±0.01,propofol：1.16±0.01）.Bicuculline （15 μmol/L） blocked the enhancement of propofol on fEPSP in tissues from young and old mice.Conclusion The fEPSP of slices from aging mice demonstrates diminished sensitivity to the enhancing actions of propofol.

Spatiotemporal dynamics of high-K^+-induced epileptiform discharges in hippocampal slice and the effects of valproate

The epileptic seizure is a dynamic process involving a rapid transition from normal activity to a state of hypersynchronous neuronal discharges. Here we investigated the network properties of epileptiform discharges in hippocampal slices in the presence of high K ＋ concentration （8.5 mmol/L） in the bath, and the effects of the anti-epileptic drug valproate （VPA） on epileptiform discharges, using a microelectrode array. We demonstrated that epileptiform discharges were predominantly initiated from the stratum pyramidale layer of CA3a-b and propagated bi-directionally to CA1 and CA3c. Disconnection of CA3 from CA1 abolished the discharges in CA1 without disrupting the initiation of discharges in CA3. Further pharmacological experiments showed that VPA at a clinically relevant concentration （100 μmol/L） suppressed the propagation speed but not the rate or duration of high-K＋-induced discharges. Our findings suggest that pacemakers exist in the CA3a-b region for the generation of epileptiform discharges in the hippocampus. VPA reduces the conduction of such discharges in the network by reducing the propagation speed.

Inhibition of inflammatory mediator release from microglia can treat ischemic/hypoxic brain injury

Interleukin-1α and interleukin-1β aggravate neuronal injury by mediating the inf1αmmatory reaction following ischemic/hypoxic brain injury. It remains unclear whether interleukin-1α and interleukin-1β are released by microglia or astrocytes. This study prepared hippocampal slices that were subsequently subjected to oxygen and glucose deprivation. Hematoxylin-eosin staining verified that neurons exhibited hypoxic changes. Results of enzyme-linked immunosorbent assay found that interleukin-1α and interleukin-1β participated in this hypoxic process. Moreover, when hypoxic injury occurred in the hippocampus, the release of interleukin-1α and interleukin-1β was mediated by the P2X4 receptor and P2X7 receptor. Immunofluorescence staining revealed that during ischemia/hypoxia, the P2X4 receptor, P2X7 receptor, interleukin-1α and interleukin-1β expression was detectable in rat hippocampal microglia, but only P2X4 receptor and P2X7 receptor expression was detected in astrocytes. Results suggested that the P2X4 receptor and P2X7 receptor, respectively, mediated interleukin-1α and interleukin-1β released by microglia, resulting in hippocampal ischemic/hypoxic injury. Astrocytes were activated, but did not synthesize or release interleukin-1α and interleukin-1β.

Paeonol attenuates inflammation-mediated neurotoxicity and microglial activation

Chronic activation of microglial cells endangers neuronal survival through the release of various proinflammatory and neurotoxic factors. The root of Paeonia lactiflora Pall has been considered useful for the treatment of various disorders in traditional oriental medicine. Paeonol, found in the root of Paeonia lactiflora Pall, has a wide range of pharmacological functions, including anti-oxidative, anti-inflammatory and neuroprotective activities. The objective of this study was to examine the efficacy of paeonol in the repression of inflammation-induced neurotoxicity and microglial cell activation. Organotypic hippocampal slice cultures and primary microglial cells from rat brain were stimulated with bacterial lipopolysaccharide. Paeonol pretreatment was performed for 30 minutes prior to lipopolysaccharide addition. Cell viability and nitrite （the production of nitric oxide）, tumor necrosis factor-alpha and interleukin-lbeta products were measured after lipopolysaccharide treatment. In organotypic hippocampal slice cultures, paeonol blocked lipopolysaccharide-related hippocampal cell death and inhibited the release of nitrite and interleukin-lbeta. Paeonol was effective in inhibiting nitric oxide release from primary microglial cells. It also reduced the lipopolysaccharide-stimulated release of tumor necrosis factor-alpha and intefleukin-1β from microglial cells. Paeonol possesses neuroprotective activity in a model of inflammation-induced neurotoxicity and reduces the release of neurotoxic and proinflammatory factors in activated microglial cells.

Neuroprotective effects of bovine colostrum on intracerebral hemorrhage-induced apoptotic neuronal cell death in rats

Brain cell death after intracerebral hemorrhage may be mediated in part by an apoptotic mechanism Colostrum is the first milk produced by mammals for their young. It plays an important role in protection and development by providing various antibodies, growth factors and nutrients, and has been used for various diseases in many countries. In the present study, we investigated the anti-apoptotic effects of bovine colostrum using organotypic hippocampal slice cultures and an intracerebral hemorrhage animal model. We performed densitometric measurements of propidium iodide uptake, a step-down avoidance task, Nissl staining, and caspase-3 immunohistochemistry. The present results revealed that colostrum treatment significantly suppressed N-methyI-D-aspartic acid-induced neuronal cell death in the rat hippocampus. Moreover, colostrum treatment improved short-term memory by suppressing hemorrhage-induced apoptotic neuronal cell death and decreasing the volume of the lesion induced by intracerebral hemorrhage in the rat hippocampus. These results suggest that colostrum may have a beneficial role in recovering brain function following hemorrhagic stroke by suppressing apoptotic cell death.

Neuropharmacological Characterization of Extracts from Rhodiola rosea, Oenothera paradoxa and Paullinia cupana in Comparison to Caffeine

To find possible therapeutic applications involving the Central Nervous System (CNS) for herbals is a major challenge during functional food and drug discovery and development programmes. Despite the availability of numerous in vitro and in vivo tests, there is no single agreed screening procedure for pharmacological testing of herbal extracts with anticipated CNS activity. Experience gained from more than 25 years of testing has shown that two models give reasonably reliable orientation for future CNS applications: construction of an electropharmacogram based on wireless recording of field potentials from the depth of the brain of freely moving rats (Tele-Stereo-EEG) and recording of the population spike produced by pyramidal cells from hippocampal slices in vitro. A combination of these two methods has now been used to characterize the pharmacological profile of extracts from Rhodiola rosea root, Oenothera paradoxa seeds and Paullinia cupana seeds. Spectral analysis of field potentials revealed attenuation of alpha2 and beta1 waves was common for all extracts. According to previous studies, this is interpreted as activation of the dopaminergic and glutamatergic transmission. In addition, Oenothera and Rhodiola extracts attenuated delta and theta power, probably related to interference with the cholinergic and norepinephrinergic transmission, respectively. Using discriminant analysis for comparison with reference pharmaceutical and botanical drugs, Rhodiola projected near the position of Ginkgo extract, whereas Oenothera extract was projected near the position of Tramadol, an analgesic drug. Physical motion was increased only in the presence of Paullinia extract and caffeine. Increases of long-term potentiation were observed in the presence of Rhodiola extract, Paullinia extract and caffeine. The combined information predicts stimulant and cognitive function-enhancing activities in humans for the Rhodiola extract, which could also be used as a possible caffeine-replacement, and antidepressant and analgesic activity for the Oenothera extract.

Quantitative evaluation of extrinsic factors influencing electrical excitability in neuronal networks： Voltage Threshold Measurement Method(VTMM)

The electrical excitability of neural networks is influenced by different environmental factors. Effective and simple methods are required to objectively and quantitatively evaluate the influence of such factors, including variations in temperature and pharmaceutical dosage. The aim of this paper was to introduce ‘the voltage threshold measurement method＇, which is a new method using microelectrode arrays that can quantitatively evaluate the influence of different factors on the electrical excitability of neural networks. We sought to verify the feasibility and efficacy of the method by studying the effects of acetylcholine, ethanol, and temperature on hippocampal neuronal networks and hippocampal brain slices. First, we determined the voltage of the stimulation pulse signal that elicited action potentials in the two types of neural networks under normal conditions. Second, we obtained the voltage thresholds for the two types of neural networks under different concentrations of acetylcholine, ethanol, and different temperatures. Finally, we obtained the relationship between voltage threshold and the three influential factors. Our results indicated that the normal voltage thresholds of the hippocampal neuronal network and hippocampal slice preparation were 56 and 31 m V, respectively. The voltage thresholds of the two types of neural networks were inversely proportional to acetylcholine concentration, and had an exponential dependency on ethanol concentration. The curves of the voltage threshold and the temperature of the medium for the two types of neural networks were U-shaped. The hippocampal neuronal network and hippocampal slice preparations lost their excitability when the temperature of the medium decreased below 34 and 33°C or increased above 42 and 43°C, respectively. These results demonstrate that the voltage threshold measurement method is effective and simple for examining the performance/excitability of neuronal networks.

GABA_A Receptor Activity Suppresses the Transition from Interictal to Ictal Epileptiform Discharges in Juvenile Mouse Hippocampus

Exploring the transition from inter-ictal to ictal epileptiform discharges(IDs) and how GABAAreceptormediated action affects the onset of IDs will enrich our understanding of epileptogenesis and epilepsy treatment.We used Mg2+-free artificial cerebrospinal fluid(ACSF) to induce epileptiform discharges in juvenile mouse hippocampal slices and used a micro-electrode array to record the discharges. After the slices were exposed to Mg2+-free ACSF for 10 min–20 min, synchronous recurrent seizurelike events were recorded across the slices, and each event evolved from inter-ictal epileptiform discharges(IIDs) to pre-ictal epileptiform discharges(PIDs), and then to IDs.During the transition from IIDs to PIDs, the duration of discharges increased and the inter-discharge interval decreased. After adding 3 lmol/L of the GABAAreceptor agonist muscimol, PIDs and IDs disappeared, and IIDs remained. Further, the application of 10 lmol/L muscimol abolished all the epileptiform discharges. When the GABAAreceptor antagonist bicuculline was applied at 10 lmol/L, IIDs and PIDs disappeared, and IDs remained at decreased intervals. These results indicated that there are dynamic changes in the hippocampal network preceding the onset of IDs, and GABAAreceptor activity suppresses the transition from IIDs to IDs in juvenile mouse hippocampus.

Use of multi-electrode array recordings in studies of network synaptic plasticity in both time and space

Simultaneous multisite recording using multi-electrode arrays（MEAs） in cultured and acutely-dissociated brain slices and other tissues is an emerging technique in the field of network electrophysiology.Over the past 40 years,great efforts have been made by both scientists and commercial concerns,to advance this technique.The MEA technique has been widely applied to many regions of the brain,retina,heart and smooth muscle in various studies at the network level.The present review starts from the development of MEA techniques and their uses in brain preparations,and then specifically concentrates on the use of MEA recordings in studies of synaptic plasticity at the network level in both the temporal and spatial domains.Because the MEA technique helps bridge the gap between single-cell recordings and behavioral assays,its wide application will undoubtedly shed light on the mechanisms underlying brain functions and dysfunctions at the network level that remained largely unknown due to the technical difficulties before it matured.