Methodological aspects of in vitro sensing of L-glutamate in acute brain slices

L-Glutamate is a major amino acid neurotransmitter in the central neuronal system of the mammalian brain and plays a vital role in brain development, synaptic plasticity, neurotoxicity, and neuropathological disorders. Despite technical limitations, progress is being made in sensing L-glutamate in vivo and in vitro. Sophisticated microsensors with the necessary spatial and temporal resolution have recently been emerging, which enable us to discern regional distribution, concentration levels, and temporal changes of L-glutamate in acute brain slices. The L-glutamate sensors for in vitro sensing have different structures and sizes, such as glass capillary-based enzyme sensors, polymer-coated enzyme sensors, and patch sensors based on natural sensing probes. The concentration of L-glutamate released in brain slices by chemical stimulation is markedly dependent on neuronal regions, types of stimulation, and sensing methods. Real- and long-time monitoring of L-glutamate in acute hippocampal slices is beginning to shed light on L-glutamate release related to the molecular mechanisms of long-term potentiation. Progress is also being made toward the visualization of L-glutamate release in acute hippocampal slices. The methodological aspects of in vitro sensing of L-glutamate are discussed.     

A glass capillary microelectrode based on capillarity and its application to the detection of L-glutamate release from mouse brain slices.

A new glass capillary microelectrode for L-glutamate is described using pulled glass capillaries (tip size, approximately 12.5 microm) with a very small volume (approximately 2 microl) of inner solution containing glutamate oxidase (GluOx) and ascorbate oxidase. The operation of the electrode is based on capillary action that samples L-glutamate into the inner solution. The enzyme reaction by GluOx generates hydrogen peroxide that is detected at an Os-gel-HRP polymer modified Pt electrode in a three-electrode configuration. The amperometric response behavior of the electrode was characterized in terms of the capillarity, response time, sensitivity and selectivity for measurements of L-glutamate. The currents at 0 V vs. Ag/AgCl increased linearly with the L-glutamate concentration from 10 to 150 microM for in vitro and in situ calibrations. The response was highly selective to L-glutamate over ascorbate, dopamine, serotonin and other amino acids. The detection of L-glutamate in the extracellular fluids of different regions of mouse hippocampal slices under stimulation of KCl was demonstrated.     

New BATO complexes for brain and myocardial imaging

Two new BATO complexes^99mTcC1 (dmg)₃BC₆H₄CH₃ (Cholor[bis {2,3-butanedionedioxime(1-)-O} {2,3-butanedionedioximato (2-)-N,N',N",N'",N",N"'} (m-tolueneborato) technetium]) and^99m TcCl (4-MCDO)₃MeB, ([bis{4-methyl-1,2-cyclohexanedioximato (1-)-O} {4-methyl-cyclohexane-1, 2-dione-dioximato (2-)-O} {methyl-borato (2-)-N,N', N", N'", N", N"'} chlorotechnetium]), generally called BATO (Boronic Acid Adducts of Technetium Dioximes, had been synthesized and evaluated for potential use in brain and myocardial perfusion imaging. Their labeling conditions were also investigated. In their biodistribution studyies they showed higher radiochemical stability and rapid brain uptake and myocardial uptake in mice. After i.v administration, the first complex had 0.87%ID in the brain and 1.02% ID in the heart at 2 min and it had a longer retention in brain (0.62%ID was maintained at 15 min postinection) but rapidly cleared from heart (0.33%ID postinjection). For the second complex, it showed very rapid blood clearance. The uptake of heart, lung and blood in mice at 2 min respectively were: 1.32%ID, 2.48%ID and 6.66%ID. These two complexes formation were 2rapid, simple and of high yield(91%). The processes were easy to kit formulation.     

Raman spectroscopic imaging for in vivo detection of cerebral brain metastases

We report for the first time a proof-of-concept experiment employing Raman spectroscopy to detect intracerebral tumors in vivo by brain surface mapping. Raman spectroscopy is a non-destructive biophotonic method which probes molecular vibrations. It provides a specific fingerprint of the biochemical composition and structure of tissue without using any labels. Here, the Raman system was coupled to a fiber-optic probe. Metastatic brain tumors were induced by injection of murine melanoma cells into the carotid artery of mice, which led to subcortical and cortical tumor growth within 14 days. Before data acquisition, the cortex was exposed by creating a bony window covered by a calcium fluoride window. Spectral contributions were assigned to proteins, lipids, blood, water, bone, and melanin. Based on the spectral information, Raman images enabled the localization of cortical and subcortical tumor cell aggregates with accuracy of roughly 250 μm. This study demonstrates the prospects of Raman spectroscopy as an intravital tool to detect cerebral pathologies and opens the field for biophotonic imaging of the living brain. Future investigations aim to reduce the exposure time from minutes to seconds and improve the lateral resolution.     

Gas-phase acidity of D-glucose. A density functional theory study

The gas-phase acidity of D-glucopyranose was studied by means of B3LYP calculations combined with 6-31G(d,p) or 6-31+G(d,p) standard basis sets. For each anomer, deprotonation of the various primary and secondary hydroxyl groups was considered. As in solution, the anomeric hydroxyl is found to be the most acidic for both anomers, but only when the 6-31+G(d,p) basis set is used for geometry optimization. Deprotonation of the anomeric hydroxyl induces an important C(1)--O endocyclic bond elongation and subsequently promotes an energetically favored ring-opening process as attested by the very small calculated activation barriers. The results also suggest that interconversion between the various deprotonated alpha- and beta-anomers may easily occur under slightly energetic conditions. B3LYP/6-311+G(2df,2p) calculations led to the an absolute gas-phase acidity of deltaacidGo(298)(alpha-D-glucose) = 1398 kJ mol(-1). This estimate matches well the only experimental value available to date. Finally, this study again confirms that the use of diffuse functions on heavy atoms is necessary to describe anionic systems properly and to achieve good relative and absolute gas-phase acidities.     

Development of 99mTc-labelled complexes for imaging dopamine transporters in the brain

Technetium-99m (T_1/2=6h, 140keV) is the most commonly used short-lived radionuclide for diagnostic nuclear medicine imaging. It is important from an inorganic chemistry point of view to develop novel ligands and chelation chemistry associated with this radionuclide, because many patients could potentially benefit from advances in technetium chemistry. Recent studies showed that formation of tropane derivatives containing a neutral [Tc^VO]³⁺N₂S₂ complex are useful as dopamine transporter imaging agents. These agents may be important for the imaging of patients with Parkinson's and other neurodegenerative diseases.     

Recent development in selective Tau tracers for PET imaging in the brain

Abnormal Tau deposition is a crucial pathological hallmark of various neurodegenerative disorders defined as tauopathies, of which Alzheimer's disease is the most prominent one. To date, a large number of chemical entities with different structures have been developed as Tau imaging tracers for the early diagnosis of tauopathies. Several of them with excellent bio-properties are currently being assessed in clinical trials, and more recently, the Tauvid? ([¹⁸F]Flortaucipir, also known as [¹⁸F]AV1451 or [¹⁸F]T807) as the first Tau tracer was approved by the U.S. Food and Drug Administration in 2020. This review summarized the latest development of Tau tracers and analyzed their chemical structures, with particular attention to the effects of chemical structures on biological properties. In addition, we also discuss the limitations of current Tau imaging tracers, issues that need attention in the development of new tracers, and possible future directions.     

壳聚糖-Cu^2＋复合物膜固定葡萄糖氧化酶用于葡萄糖传感

利用壳聚糖(CS)对Cu2 的配位吸附作用,制得壳聚糖-Cu2 复合膜修饰电极,进一步通过Cu2 与蛋白质的配位作用,再将葡萄糖氧化酶(GOD)固定在金电极上制得GOD-Cu2 -CS/Au电极.该酶电极检测葡萄糖的线性范围为0.03～2.5 mmol·L-1,线性相关系数为0.997,检出限(S/N=3)为1 μmol·L-1.以电化学石英晶体微天平(EQCM)技术监测了各修饰过程.     

Enantiospecific synthesis of S-quinuclidinol from D-glucose: A strategy for the synthesis of chiral quinuclidines.

Two alternative strategies for the synthesis of quinuclidines from hexoses are outlined. One of these is employed in an enantiospecific synthesis of S-quinuclidinol whereby a two carbon fragment is introduced at C-4 of D-glucose.     

Hypoxia-sensitive fluorescent probes for in vivo real-time fluorescence imaging of acute ischemia

Based on the findings that the azo functional group has excellent properties as the hypoxia-sensor moiety, we developed hypoxia-sensitive near-infrared fluorescent probes in which a large fluorescence increase is triggered by the cleavage of an azo bond. The probes were used for fluorescence imaging of hypoxic cells and real-time monitoring of ischemia in the liver and kidney of live mice.     

Solution of the Percus-Yevick equation for hard disks

The authors solve the Percus-Yevick equation in two dimensions by reducing it to a set of simple integral equations. They numerically obtain both the pair correlation function and the equation of state for a hard disk fluid and find good agreement with available Monte Carlo results. The present method of resolution may be generalized to any even dimension.     

Novel radioiodinated sibutramine and fluoxetine as models for brain imaging

Brain imaging is a process which allows scientists and physicians to view and monitor the areas of the brain which allow diagnosis and following up different abnormalities in the brain. The aim of this study was to develop potential radiopharmaceuticals for the non-invasive brain imaging. Sibutramine and fluoxetine (two drugs that have the ability to cross blood–brain barrier) were successfully labeled with ¹²⁵I via direct electrophilic substitution reaction at ambient temperature. The reaction parameters studied were substrate concentration, oxidizing agent concentration, pH of the reaction mixture, reaction temperature, reaction time and in vitro stability of the iodocompounds. The iodocompounds gave maximum labeling yield of 92 ± 2.77 and 93 ± 2.1%, respectively, and maintained stability throughout working period (24 h). Biodistribution studies showed that maximum in vivo uptake of the iodocompounds in the brain was 5.7 ± 0.19 and 6.14 ± 0.26% injected activity/g tissue organ, respectively, at 15 and 5 min post-injection, whereas the clearance from the mice appeared to proceed via the hepatobiliary pathway. Brain uptake of ¹²⁵I-sibutramine and ¹²⁵I-fluoxetine is higher than that of ^99mTc-ECD and ^99mTc-HMPAO (currently used radiopharmaceuticals for brain imaging) and so radioiodinated sibutramine and fluoxetine could be used instead of ^99mTc-ECD and ^99mTc-HMPAO for brain SPECT.     

High-accuracy assessment of finiteness effects for hard disks

In a recent investigation the comparison between theoretical and computer-experimental results for the finiteness effects in hard disk systems has not been entirely conclusive. Thus, we present new Monte Carlo data of high accuracy. We also discuss some aspects of experimental design as well as refined methods of error analysis. The quantity considered is the compressibility factor Z, depending on the shape of the periodic box (for given density and particle number). The reported discrepancies in V dependence are stringently substantiated, and a semi-empirical correction of the theoretical approach is proposed. The new data also revealed an unexpected discrepancy in N dependence. For this second kind of finiteness effects, we replace the usual calculation via the virial coefficients by a computer-experimental estimate.     

Radioiodinated phenoxyacetic acid derivatives as potential brain imaging agents. II. Structure-biodistribution relationship

In developing new brain imaging agents for single photon emission computed tomography (SPECT), we synthesized eleven radioiodinated phenoxyacetic acid derivatives and investigated the relationship between the chemical structure and in vivo characteristics. Biodistribution studies in mice revealed high initial brain uptake for all the compounds. Blood radioactivity level depended markedly upon the chemical stability of the compound. The alpha, alpha-dimethylester derivative (1e), amide derivatives (2a--c) and diamine derivatives (3a, b, 4), which were stable to hydrolysis, showed low blood activity levels following i.v. administration. Disappearance of the ester and amide compounds from the brain was rapid. However, the diamine derivatives displayed improved retention in the brain. Compounds 3a and 4 possessed the best combination of high brain uptake and sufficient retention to be useful as potential brain imaging radiopharmaceuticals with SPECT devices.     

MALDI-ion mobility-TOFMS imaging of lipids in rat brain tissue

While maintaining anatomical integrity, matrix assisted laser desorption/ionization mass spectrometry (MALDI-MS) has allowed researchers to directly probe tissue, map the distribution of analytes and elucidate molecular structure with minimal preparation. MALDI-ion mobility (IM)-orthogonal time-of-flight mass spectrometry (oTOFMS) provides an advantage by initially separating different classes of biomolecules such as lipids, peptides, and nucleotides by their IM drift times prior to mass analysis. In the present work the distribution of phosphatidlycholine and cerebroside species was mapped from 16 microm thick coronal rat brain sections using MALDI-IM-oTOFMS. Furthermore, the use of gold nanoparticles as a matrix enables detection of cerebrosides, which although highly concentrated in brain tissue, are not easily observed as positive ions because of intense signals from lipids such as phosphatidlycholines and sphingomyelins.     

Application of dried-droplets deposited on pre-cut filter paper disks for quantitative LA-ICP-MS imaging of biologically relevant minor and trace elements in tissue samples

In this work, a novel calibration approach for minor and trace element quantification in LA-ICP-MS imaging of biological tissues is presented. Droplets of aqueous standard solutions are deposited onto pre-cut pieces of filter paper, allowed to dry, and sputtered with a thin gold layer for use as pseudo-internal standard. Analysis of the standards using LA-ICP-MS is performed using radial line-scans across the filters. In contrast to conventionally used preparation of matrix-matched tissue standards, the dried-droplet approach offers a variety of advantages: The standards are easy to prepare, no characterization of the standards using acid digestion is required, no handling of biological materials is necessary, and the concentration range, as well the number of investigated analytes is almost unlimited. The proposed quantification method has been verified using homogenized tissue standards with known analyte concentrations before being applied to a human malignant mesothelioma biopsy from a patient who had not received any chemotherapeutic treatment. Elemental distribution images were acquired at a lateral resolution of 40 μm per pixel, limits of detection ranging from 0.1 μg g⁻¹ (Mn, Ni, Cu, Zn) to 13.2 μg g⁻¹ (K) were reached.     

Subcellular imaging mass spectrometry of brain tissue

Imaging mass spectrometry provides both chemical information and the spatial distribution of each analyte detected. Here it is demonstrated how imaging mass spectrometry of tissue at subcellular resolution can be achieved by combining the high spatial resolution of secondary ion mass spectrometry (SIMS) with the sample preparation protocols of matrix-assisted laser desorption/ionization (MALDI). Despite mechanistic differences and sampling 10(5) times less material, matrix-enhanced (ME)-SIMS of tissue samples yields similar results to MALDI (up to m/z 2500), in agreement with previous studies on standard compounds. In this regard ME-SIMS represents an attractive alternative to polyatomic primary ions for increasing the molecular ion yield. ME-SIMS of whole organs and thin sections of the cerebral ganglia of Lymnaea stagnalis demonstrate the advantages of ME-SIMS for chemical imaging mass spectrometry. Subcellular distributions of cellular analytes are clearly obtained, and the matrix provides an in situ height map of the tissue, allowing the user to identify rapidly regions prone to topographical artifacts and to deconvolute topographical losses in mass resolution and signal-to-noise ratio.     

Fluorescence response mechanism of D-glucose selectivity for supramolecular probes composed of phenylboronic-acid-modified beta-cyclodextrin and styrylpyridinium dyes.

Supramolecular complex formation of phenylboronic-acid-modified beta-cyclodextrin (1) with 1-methyl-4-(4-dimethylaminostyryl)pyridinium (C1SP) in aqueous solutions containing saccharides was fully clarified to gain an insight into the observed D-glucose (D-glc) selectivity of a supramolecular fluorescent probe composed of 1 and the 1-heptyl analogue of C1SP (Chem. Commun., 2006, 4319). At pH 9.6, where 1 was in its anionic form, both the stability and the fluorescence of the 1/C1SP complex were reduced by the formation of boronate esters of 1 with saccharides. Among the saccharides, D-glc had the smallest effect on destabilization of the 1/C1SP complex, almost completely retaining the fluorescence of the 1/C1SP complex that was reduced by other saccharides by approximately 2/3. Under neutral conditions, D-glc enhanced the fluorescence of the 1/C1SP complex by increasing the fraction of anionic 1 while minimally decreasing the stability and fluorescence of the 1/C1SP complex. Although other saccharides also increased the fraction of the anionic 1, their relatively large effects on the destabilization and reduction of fluorescence of the 1/C1SP complex limited the enhancement of the fluorescence of the 1-C1SP system under neutral conditions.     

Recent progress in base-catalyzed isomerization of D-glucose into D-fructose

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