The blood-brain barrier (BBB) is a structural and functional barrier that regulates the passage of molecules into and out of the brain to maintain the neural microenvironment. We have previously developed the in vitro BBB model with human brain microvascular endothelial cells (HBMEC). However, in vivo HBMEC are shown to interact with astrocytes and also exposed to shear stress through blood flow. In an attempt to develop the BBB model to mimic the in vivo condition we constructed the flow-based in vitro BBB model using HBMEC and human fetal astrocytes (HFA). We also examined the effect of astrocyte conditioned medium (ACM) in lieu of HFA to study the role of secreted factor(s) on the BBB properties. The tightness of HBMEC monolayer was assessed by the permeability of dextran and propidium iodide as well as by measuring the transendothelial electrical resistance (TEER). We showed that the HBMEC permeability was reduced and TEER was increased by non-contact, co-cultivation with HFA and ACM. The exposure of HBMEC to shear stress also exhibited decreased permeability. Moreover, HFA/ACM and shear flow exhibited additive effect of decreasing the permeability of HBMEC monolayer. In addition, we showed that the HBMEC expression of ZO-1 (tight junction protein) was increased by co-cultivation with ACM and in response to shear stress. These findings suggest that the non-contact co-cultivation with HFA helps maintain the barrier properties of HBMEC by secreting factor(s) into the medium. Our in vitro flow model system with the cells of human origin should be useful for studying the interactions between endothelial cells, glial cells, and secreted factor(s) as well as the role of shear stress in the barrier property of HBMEC.

Background and Purpose-We characterized the differential effects of glycine at different levels in the induction of postischemic long-term potentiation, as well as in the neuronal damage induced by focal ischemia. Methods-Whole-cell patch clamp recordings were obtained from rat hippocampal slice preparations. In vitro ischemia and postischemic long-term potentiation were induced by oxygen and glucose deprivation. In vivo ischemia was induced by transient middle cerebral artery occlusion. Results-In both in vitro and in vivo ischemia models, glycine at low level exerts deleterious effects in postischemic long-term potentiation and ischemic neuronal injury by modulation of the N-methyl-D-aspartate receptor coagonist site; whereas glycine at high level exerts neuroprotective effects by activation of glycine receptor and subsequent differential regulation of N-methyl-D-aspartate receptor subunit components. Conclusions-Our results provide a molecular basis for the dual roles of glycine in ischemic injury through distinct mechanisms, and they suggest that glycine receptors could be a potential target for clinical treatment of stroke. (Stroke. 2012;43:2212-2220.)Key Words: glycine Ⅲ ischemia Ⅲ electrophysiology Ⅲ middle cerebral artery occlusion Ⅲ N-methyl-D Ⅲ aspartate receptor A pathological form of plasticity, named postischemic long-term potentiation (i-LTP), was observed in glutamatereceptor-mediated neurotransmission after stroke. [1][2][3] There is evidence that i-LTP in the hippocampus may exert a detrimental effect via facilitation of excitotoxic damage. 3 This long-term enhancement in AMPA-and NMDA-receptor-mediated excitatory responses was mainly attributable to overstimulation of glutamatergic neurotransmission by excessively released extracellular glutamate in the postischemic brain. Given that overexcitation of neurons caused by stroke disturbs the balance between excitation and inhibition, restoring this balance via intervention with additional inhibition seems to be a potential and practical strategy.Ischemia elicits the rapid release of various amino acid neurotransmitters, including glycine. 4,5 Glycine is a 2-faceted bioactive molecule in the central nervous system. 6 Glycine is a strychnine-insensitive coagonist for N-methyl-D-aspartate receptors (NMDAR) and is essential for activation of NMDARs. [7][8][9] And yet, glycine is one of the main inhibitory neurotransmitters in the central nervous system. 10 Over the past decade, accumulating evidence has suggested that functional glycine receptors are present throughout all regions in the hippocampus, and they play an important role in regulating excitability and plasticity. These strychnine-sensitive glycine receptors (GlyR), if located postsynaptically, are mostly in extrasynaptic sites. 11 Because GlyRs activation induces Cl Ϫ flux and neuronal hyperpolarization, and thus suppresses neuronal excitability, we sought to determine whether and how activation of extrasynaptic GlyRs could help restore excitation-inhibition balance and activity-dependent p...

In vivo experience induces changes in synaptic NMDA receptor (NMDAR) subunit components, which are correlated with subsequent modifications of synaptic plasticity. However, little is known about how these subunit changes regulate the induction threshold of subsequent plasticity. At hippocampal Schaffer collateral-CA1 synapses, we first examined whether a recent history of neuronal activity could affect subsequent synaptic plasticity through its actions on NMDAR subunit components. We found that prior activity history produced by priming stimulations (PSs) across a wide range of frequencies (1-100 Hz) could induce bidirectional changes in the NR2A/NR2B ratio, which governs the threshold for subsequent long-term potentiation/long-term depression (LTP/LTD). Manipulating the NR2A/NR2B ratio through partial NR2 subunit blockade mimicked the PS regulation of the LTP/LTD threshold. Our results demonstrate that activity-dependent changes in the NR2A/NR2B ratio can be critical factors in metaplastic regulation of the LTP/LTD threshold.

A new fluoropolymer is proposed as the basis of a novel class of sensors. The devices are based on selective chromogenic reactions and in situ long-path optical absorbance measurement. The polymer is transparent from 200 to 2000 nm and has the lowest known refractive index (RI) of any synthetic polymer. The RI is less than that of water. A tube of this material, filled with an aqueous solution (or virtually any other liquid), behaves as a liquid core optical fiber. As a result, long-path length optical cells are possible without significant loss of light. The material is highly permeable to a number of trace gases of environmental interest. Relative to common poly-(tetrafluoroethylene) (PTFE)-type Teflon, the new amorphous fluoropolymer (Teflon AF 2400) is more than 3 orders of magnitude more permeable to many gases. If a Teflon AF tube is filled with a reagent that responds to a gaseous analyte by undergoing a change that is spectroscopically detectable, an unusually versatile, sensitive, and inexpensive gas sensor can be made with conventional optical fibers at each end, connected respectively to an inexpensive light source such as a light-emitting diode and a photodiode detector. A capillary hollow fiber structure allows a high surface-to-volume ratio, allowing high sensitivity, and supports a thin wall, with response times down to subsecond periods. The potential for similar sensors for volatile organic compounds dissolved in water is also demonstrated.

Glycine in the hippocampus can exert its effect on both synaptic NMDA receptors (NMDARs) and extrasynaptic functional glycine receptors (GlyRs) via distinct binding sites. Previous studies have reported that glycine induces long-term potentiation (LTP) through the activation of synaptic NMDARs. However, little is known about the potential role of the activated GlyRs that are largely located in extrasynaptic regions. We report here that relatively high levels of glycine achieved either by exogenous glycine application or by the elevation of endogenous glycine accumulation with an antagonist of the glycine transporter induced long-term depression (LTD) of excitatory postsynaptic currents (EPSCs) in hippocampal CA1 pyramidal neurons. The co-application of glycine with the selective GlyR antagonist strychnine changed glycine-induced LTD (Gly-LTD) to LTP. Blocking the postsynaptic GlyR-gated net chloride flux by manipulating intracellular chloride concentrations failed to elicit any changes in EPSCs. These results suggest that GlyRs are involved in Gly-LTD. Furthermore, this new form of chemical LTD was accompanied by the internalization of postsynaptic AMPA receptors and required the activation of NMDARs. Therefore, our present findings reveal an important function of GlyR activation and modulation in gating the direction of synaptic plasticity.