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Abstract: Contraction of airway smooth muscle (ASM) is regulated by the physiological, structural and mechanical environment in the lung. We review two in vitro techniques, lung slices and airway segment preparations, that enable in situ ASM contraction and airway narrowing to be visualized. Lung slices and airway segment approaches bridge a gap between cell culture and isolated ASM, and whole animal studies. Imaging techniques enable key upstream events involved in airway narrowing, such as ASM cell signalling and structural and mechanical events impinging on ASM, to be investigated. [Copyright &y& Elsevier]

The article presents a study on the mineralized tissue changes on the origin site of metacarpal condylar fracture in young Thoroughbred horses prior to race training. A point-projection digital x-ray imaging visualized bone slices in mediolateral planes through the centre of rotation of the metacarpo-phalangeal joint (MCPJ) of distal third metacarpal bones (Mc3) of the horses. The radiolucency found that the fracture reflects bone and joint development abnormality present before activity.

• Introduction • Nomenclature • Uterine contractions and the ‘functional syncytium theory’ • Interstitial cells of Cajal in the gastrointestinal and urinary tracts • Are there m-ICLC in myometrium? • The effect of c-kit inhibition on myometrial contractility • Electrophysiology of m-ICLC • Imaging of tissue level signals in myometrium • Emerging concepts – the possible role of extracellular ATP in myometrial contractility • Where do m-ICLC fit in with a model for the generation of myometrial contractions? • Future research directions • Conclusion A strict regulation of contractility in the uterus and fallopian tube is essential for various reproductive functions. The uterus contributes, through either increased contractility or periods of relative quiescence, to: ( i) expulsion of menstrual debris, ( ii) sperm transport, ( iii) adequate embryo placement during implantation, ( iv) enlarging its capacity during pregnancy and ( v) parturition. The dominant cell population of the uterine wall consists of smooth muscle cells that contain the contractile apparatus responsible for the generation of contractile force. Recent interest has focused on a new population of cells located throughout the myometrium on the borders of smooth muscle bundles. These cells are similar to interstitial cells of Cajal (ICC) in the gut that are responsible for the generation of electrical slow waves that control peristalsis. A precise role for myometrial Cajal-like interstitial cells (m-ICLC) has not been identified. m-ICLC express the c-kit receptor, involved in creating and maintaining the ICC phenotype in the gastrointestinal tract. However, both acute and prolonged inhibition of this receptor with the c-kit antagonist imatinib mesylate does not appear to affect the spontaneous contractility of myometrium. Calcium imaging of live tissue slices suggests that contractile signalling starts on the borders of smooth muscle bundles where m-ICLC are located and recently the possible role of extracellular ATP signalling from m-ICLC has been studied. This manuscript reviews the evidence regarding tissue-level signalling in the myometrium with a particular emphasis on the anatomical and possible functional aspects of m-ICLC as new elements of the contractile mechanisms in the uterus. [ABSTRACT FROM AUTHOR]

Abstract: Fibroblast growth factor 2 (FGF2) is involved in hippocampus-dependent learning. In this study, the effects of FGF2 on the excitability were investigated in granule cells of rat dentate gyrus. Hippocampal slices were used to perform patch clamp recordings in granule cells. Extracellularly applied FGF2 early quenched the depolarization-induced repetitive firing, suggesting a decreased excitability under these conditions. Consistently, transient and sustained voltage-gated K+ currents decreased in a dose-dependent manner, repolarization phase of action potential was slowed down, afterhyperpolarization was reduced, and membrane resistance was decreased. These effects were not mediated by tyrosine kinase FGF2 receptors. Moreover, an involvement of G protein signaling was ruled out, as well as an intracellular action of FGF2. Considering the relationship between FGF2 and hippocampal functions, the modulation of neuron excitability by activity-driven FGF2 release may be regarded as a part of a homeostatic mechanism of self-regulation of hippocampal activity. [Copyright &y& Elsevier]

The release of neurotransmitters and modulators has been studied mostly using labeled preloaded compounds. For several reasons, however, the estimated release may not reliably reflect the release of endogenous compounds. The basal and K+-evoked release of the neuroactive endogenous amino acids GABA, glycine, taurine, l-glutamate and l-aspartate was now studied in slices from the hippocampus and brain stem from 7-day-old and 3-month-old mice under control and ischemic conditions. The release of synaptically not active l-glutamine, l-alanine, l-threonine and l-serine was assessed for comparison. The estimates for the hippocampus and brainstem were markedly different and also different in developing and adult mice. GABA release was much greater in 3-month-old than in 7-day-old mice, whereas with taurine the situation was the opposite, in the hippocampus in particular. K+ stimulation enhanced glycine release more in the mature than immature brain stem while in the hippocampus the converse was observed. Ischemia enhanced the release of all neuroactive amino acids in both brain regions, the effects being relatively most pronounced in the case of GABA, aspartate and glutamate in the hippocampus in 3-month-old mice, and taurine in 7-day-old and glycine in 3-month-old mice in the brain stem. These results are qualitatively similar to those obtained on earlier experiments with labeled preloaded amino acids. However, the magnitudes of the release cannot be quite correctly estimated using radioactive labels. In developing mice only taurine release may counteract the harmful effects of excitatory amino acids in ischemia in both hippocampus and brain stem. [ABSTRACT FROM AUTHOR]

Abstract: Analysis of the interplay between cell proliferation and death has been greatly advantaged by the development of CNS slice preparations. In slices, interactions between neurons and neurons and the glial cells are fundamentally preserved in a fashion close to the in vivo situation. In parallel, these preparations offer the possibility of an easy experimental manipulation. Two main types of slices are currently in use: the acute slices, which are short living preparations where the major functions of the intact brain (including neurogenesis) are maintained, and the organotypic cultures, where the maturation and plasticity of neuronal circuitries in relation to naturally occurring neuronal death and/or experimental insults can be followed over several weeks in vitro. We will discuss here the main advantages/disadvantages linked to the use of CNS slices for histological analysis of neuronal proliferation and death, as well as the main findings obtained in the most popular types of preparations, i.e. the cortical, hippocampal, cerebellar and retinal slices. [Copyright &y& Elsevier]

Abstract: Although fructose is commonly used as a sweetener, its effects on brain function are unclear. Using rat hippocampal slices, we found that fructose and mannose, like pyruvate, preserve ATP levels during 3-h of glucose deprivation. Similarly, fructose and mannose restored synaptic potentials (excitatory postsynaptic potential, EPSPs) depressed during glucose deprivation. However, restoration of synaptic responses was slow and only partial with fructose. EPSPs supported by mannose were inhibited by cytochalasin B (CCB), a glucose transport inhibitor, but were not inhibited by α-cyano-4-hydroxycinnamate (4-CIN), a monocarboxylate transport inhibitor, indicating that neurons use mannose via glucose transporters. In contrast, both CCB and 4-CIN depressed EPSPs supported by fructose, suggesting that fructose may be taken up by non-neuronal cells through CCB sensitive hexose transporters and metabolized to a monocarboxylate for subsequent use during neuronal respiration. Supporting this possibility, 20 minutes of oxygen deprivation in the presence of fructose resulted in functional and morphological deterioration whereas oxygen deprivation in the presence of glucose or mannose had minimal toxic effects. These results indicate that neuronal fructose utilization differs from glucose and mannose and likely involves release of monocarboxylates from glia. [Copyright &y& Elsevier]

Brief test-pulse stimulation (0.2–0.05 Hz) of naïve (previously nonstimulated) developing hippocampal CA3–CA1 synapses leads to a substantial synaptic depression, explained by -amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) silencing. Using field recordings in hippocampal slices from P8 to P12 rats, we examined this depression of naïve synapses using more prolonged test-pulse stimulation as well as low-frequency (1 Hz) stimulation (LFS). We found that 900 stimuli produced depression during stimulation to ∼40% of the naïve level independent of whether test-pulse stimulation or LFS was used. This result was also observed during combined blockade of N-methyl-D-aspartate/metabotropic glutamate receptors (NMDAR/mGluRs) although the depression was smaller (to ∼55% of naïve level). Using separate blockade of either NMDARs or mGluRs, we found that this impairment of the depression resulted from the NMDAR, and not from the mGluR, blockade. In fact, during NMDAR blockade alone, depression was smaller even than that observed during combined blockade. We also found that mGluR blockade alone facilitated the LFS-induced depression. In conclusion, test-pulse stimulation produced as much depression as LFS when applied to naïve synapses even when allowing for NMDAR and mGluR activation. Our results seem in line with the notion that NMDARs and mGluRs may exert a bidirectional control on AMPA receptor recruitment to synapses. [ABSTRACT FROM AUTHOR]

Abstract: The horizontal third segment (V3h) of the vertebral artery (VA) in 7 cadavers (14 sides) was dissected and the anatomical measurements recorded. Measurements from 24 healthy individuals (48 sides) were taken for comparison using multislice CT scanning. The distance between the medial tip of the VA V3h and the line passing through the mid point of the posterior tuberculum of the atlas was marked as length A. The distance between the medial tip of the VA V3h and the point penetrating the dura mater was classified as length B. The angle between these lines was the alpha (α) angle. Measurements were taken when the head was in a neutral position, as well as in maximum right and left rotation, extension and flexion. In cadavers, the mean α angle (±S.D.) was 82.42±10.34° and 83.21±10.81° on the right and left side, respectively. On multislice CT scanning, the mean α angle was 81.64±10.15° on the right and 83.77±10.65° on the left. These angles varied with the position of the head. [Copyright &y& Elsevier]

Abstract: Drug-induced organ injury is a multifaceted process, involving numerous cell types and mediators, and remains a significant safety issue in pharmaceutical development and clinical therapy. Organotypic in vitro models, including precision-cut tissue slices, possess the multi-cellular, structural and functional features of in vivo tissue to facilitate the elucidation of mechanisms of drug-induced organ injury and to characterize species susceptibilities. This study reviews diclofenac-induced hepatotoxicity and presents data comparing the metabolism, specific binding of diclofenac products to cellular proteins and the effects on liver function in rat, monkey and human liver slices. Concentration- and time-dependent increases in specific protein binding demonstrate the progressive nature of the toxicity. Altered liver function correlated with the species differences in the extent of diclofenac metabolism (rat>monkey or human). Liver injury was not detectable within 24h, unlike specific protein binding, yet it developed by 48h and lower concentrations of diclofenac exhibited effects by 72h, demonstrating that continued metabolism and the accumulation of specific protein binding could lead to altered cell function. The decline of liver slice ATP levels at concentrations not affecting GSH levels implicates mitochondrial dysfunction as a primary indicator of hepatotoxicity, of which oxidative stress may be a contributing cause. Diclofenac affected monkey liver slices function at similar concentrations as rat liver slices, while human liver slices exhibited less extensive specific protein binding and required higher diclofenac concentrations to alter cell function. [Copyright &y& Elsevier]

Organotypic brain slice culture was used to study the direct actions of relaxin on neural cells under ischemic stress. Cortical brain slices from neonatal rats were cultured for 14 days. Experimental slices were placed in a deoxygenated, glucose-free balanced salt solution (BSS) for 1 h, one group with 10−7 M H2 relaxin in the medium and the other without. Control slices were transferred to oxygenated BSS with glucose. Slices were returned to normal culture conditions and analyzed 1, 4, 8, and 24 h post-treatment using propidium iodide (PI) fluorescence to highlight cellular damage or death. The percentages of dead and dying cells in a slice were compared between groups. Relaxin treatment attenuated PI staining at 4 and 8 h postischemia, suggesting a direct action(s) on cells that improves their chance of survival during ischemia. [ABSTRACT FROM AUTHOR]

Abstract: Excessive formation of reactive oxygen species (ROS) and disruption of glutamate uptake have been pointed as two key mechanisms in methylmercury-toxicity. Thus, here we investigate the involvement of glutamatergic system in methylmercury (MeHg) neurotoxicity and whether diphenyl diselenide, ebselen and guanosine could protect cortical rat brain slices from MeHg-induced ROS generation. MeHg (100 and 200μM) increased 2′,7′-dichlorodihydrofluorescin (DCFH) oxidation after 2h of exposure. At 50μM, MeHg increased DCFH oxidation only after 5h of exposure. Guanosine (1 and 5μM) did not caused any effect per se; however, it blocked the increase in DCFH caused by 200 or 50μM MeHg. Ebselen (5 and 10μM) decreased significantly the DCFH oxidation after 2 and 5h of exposure to MeHg. Diphenyl diselenide (5μM) did not change the basal DCFH oxidation, but abolished the pro-oxidant effect of MeHg. MK-801 also abolished the pro-oxidant effect of MeHg. These results demonstrate for the first time the potential antioxidant properties of organoseleniun compounds and guanosine against MeHg-induced ROS generation after short-term exposure in a simple in vitro model. In conclusion, endogenous purine (guanosine) and two synthetic organoselenium compounds can modulate the pro-oxidant effect of MeHg in cortical brain slices. [Copyright &y& Elsevier]

Abstract: In rat hippocampal CA1 pyramidal neurons, γ-aminobutyric acid (GABA) A receptor-mediated inhibitory postsynaptic currents (IPSCs) undergo a paired-pulse depression (PPD) by the second of two pulses, with inter-pulse intervals of 100–2000 ms, applied to the stratum radiatum. While GABA-C receptors are described in the CA1 area, their functional significance is unknown. In this study, the involvement of GABA-C receptors in PPD was examined using an in vitro hippocampal slice preparation. IPSCs evoked by stimulations in stratum radiatum were recorded with patch pipettes from CA1 pyramidal cells. PPD, when induced in the above fashion, was blocked by the GABA-C receptor antagonist (1,2,5,6-Tetrahydropyridin-4-yl) methylphosphinic acid (TPMPA, 10 μM, applied in the superfusing medium). GABA-A and GABA-B receptor-mediated IPSCs, as well as the baclofen-induced suppression of the GABA-A receptor mediated IPSC, were not antagonized by TPMPA (10–20 μM). These results indicate that PPD of the IPSC is mediated by the activation of GABA-C receptors. [Copyright &y& Elsevier]

Abstract  Accumulation of the neurotoxic amyloid β-peptide (Aβ) in the brain is a hallmark of Alzheimer’s disease (AD). Several synthetic Aβ peptides have been used to study the mechanisms of toxicity. Here, we sought to establish comparability between two commonly used Aβ peptides Aβ1-42 and Aβ25-35 on an in vitro model of Aβ toxicity. For this purpose we used organotypic slice cultures of rat hippocampus and observed that both Aβ peptides caused similar toxic effects regarding to propidium iodide uptake and caspase-3 activation. In addition, we also did not observe any effect of both peptides on Akt and PTEN phosphorylation; otherwise the phosphorylation of GSK-3β was increased. Although further studies are necessary for understanding mechanisms underlying Aβ peptide toxicity, our results provide strong evidence that Aβ1-42 and the Aβ25-35 peptides induce neural injury in a similar pattern and that Aβ25-35 is a convenient tool for the investigation of neurotoxic mechanisms involved in AD. [ABSTRACT FROM AUTHOR]

Abstract  In the brain stem glycine is associated with multiple sensory and visceral regulations, being involved in, for instance, cardiovascular, respiratory and auditory functions. We here studied the mechanisms of the release of preloaded [3H]glycine from mouse brain stem slices in a superfusion system. A depolarizing concentration of K+ ions (50 mM) evoked glycine release, but in the absence of Ca2+ the effect was attenuated, indicating that a part of the evoked release represents Ca2+-dependent exocytosis. The Ca2+-independent release was enhanced by omission of Na+ and Cl−. The stimulatory effect of extracellular glycine confirmed the involvement of transporters functioning in a reverse direction. A part of the release is mediated by Na+ and Cl− channels, since it was inhibited by the inhibitors of these, riluzole and 4-acetamido-4′-isothiocyanostilbene-2,2′-disulphonate, respectively. Glycine release was potentiated by the activation of protein kinase C and diminished by increasing cyclic guanosine monophosphate levels with a phosphodiesterase inhibitor, zaprinast. The release was also modulated by the phospholipase inhibitor quinacrine and the tyrosine kinase inhibitor genistein. Adenosine A1 receptors likewise regulate glycine release, since it was enhanced by their agonist R(−)N6-(2-phenylisopropyl)adenosine, which effect was blocked by the antagonist 8-cyclopentyl-1,3-dipropylxanthine. The ionotropic glutamate receptor agonists N-methyl-d-aspartate, kainate and 2-amino-3-hydroxy-5-methyl-4-isoxazolepropionate failed to have any effects contrary to their effects in higher brain regions, e.g., in the hippocampus. The group I and III metabotropic glutamate receptor agonists (S)-3,5-dihydroxyphenylglycine and O-phospho-l-serine, respectively, increased the release in a receptor-mediated manner. Glycine release in the brain stem was also markedly enhanced by cell-damaging conditions, including hypoxia, hypoglycemia and ischemia. [ABSTRACT FROM AUTHOR]

Background: In order to investigate whether 1-mm thin slices and multiplanar reconstructions (MPRs) of multi-detector computed tomography (CT) datasets interpreted in addition to isotropic 5-mm thick slices in one session improve the detection of peritoneal carcinomatosis. Methods: The abdominal CT datasets of 44 patients with histologically proven tumors of the abdomen or pelvis were retrospectively evaluated for peritoneal carcinomatosis by four radiologists with variable experience (radiologist 1: >or=10 years, radiologists 2 and 3: 1.5 years, radiologist 4: 0.5 years). In three successive steps, the radiologists evaluated first the axial 5-mm slices, second the 1-mm slices, and third the MPRs and rated their diagnostic confidence. Results: Specificity was nearly unchanged for all the four radiologists. Sensitivity improved for the most experienced and the least experienced radiologists and was unchanged for the two readers with intermediate skills. Except for the third step of radiologist 4, no statistically significant differences in diagnostic performance were detected. The diagnostic confidence of all the four readers benefited to variable degrees from interpretation of the 1-mm slices and MPRs. Conclusions: While 5-mm slices are sufficient for the detection of peritoneal carcinomatosis, 1-mm slices and MPRs can improve sensitivity and diagnostic confidence. [ABSTRACT FROM AUTHOR]

The elastic and hyperelastic properties of biological soft tissues have been of interest to the medical community as there are several applications where parameters characterizing these properties are critical for a reliable outcome. This includes applications such as surgery planning, needle biopsy and cancer diagnosis using medical imaging. While there has been considerable research on the measurement of the linear elastic modulus of small tissue samples, little research has been conducted for measuring parameters that characterize nonlinear elasticity of tissues included in slice specimens. In this paper, we present a method of measuring the hyperelastic parameters of tissue slice samples with tumours. In this method, to measure the hyperelastic properties of a tumour within a slice sample, the tumour was indented to acquire its force-displacement response while the slice remained intact. To calculate the hyperelastic parameters from the acquired data, we developed two inversion techniques that use the slice nonlinear finite element model as their forward problem solver. One of these techniques was based on nonlinear optimization while the other is a novel iterative technique that processes the variable slopes of the force-displacement response to calculate the hyperelastic parameters. The latter was developed specifically for the Yeoh and the second-order polynomial hyperelastic models, since we found that the other optimization-based inversion technique did not perform well with these models. To validate the proposed techniques, we performed numerical and phantom experiments. While we were able to achieve convergence with wide ranges of parameters of initial guesses to within 1% error with the numerical simulation experiments, we achieved convergence to within errors of around 5% with the tissue mimicking phantoms. Moreover, we successfully applied these techniques to data we acquired from nine pathological breast tissue slice specimens where the goal was to determine the hyperelastic properties of the tumour within the breast tissue slices. [ABSTRACT FROM AUTHOR]

Abstract: Non classical bioisosters of modafinil featuring interesting biological profile have been easily produced through replacement of the sulfoxide function with a carbonyl group and modification of the carboxylic acid amide functionality. [Copyright &y& Elsevier]

Ionic conductances that generate membrane potential oscillations in neurons of layer II of the parasubiculum were studied using whole cell current-clamp recordings in horizontal slices from the rat brain. Blockade of ionotropic glutamate and GABA synaptic transmission did not reduce the power of the oscillations, indicating that oscillations are not dependent on synaptic inputs. Oscillations were eliminated when cells were hyperpolarized 6–10 mV below spike threshold, indicating that they are mediated by voltage-dependent conductances. Application of TTX completely eliminated oscillations, suggesting that Na+currents are required for the generation of the oscillations. Oscillations were not reduced by blocking Ca2+currents with Cd2+or Ca2+-free artificial cerebrospinal fluid, or by blocking K+conductances with either 50 µM or 5 mM 4-aminopyridine (4-AP), 30 mM tetraethylammonium (TEA), or Ba2+(1–2 mM). Oscillations also persisted during blockade of the muscarinic-dependent K+current, IM, using the selective antagonist XE-991 (10 µM). However, oscillations were significantly attenuated by blocking the hyperpolarization-activated cationic current Ihwith Cs+and were almost completely blocked by the more potent Ihblocker ZD7288 (100 µM). Intrinsic membrane potential oscillations in neurons of layer II of the parasubiculum are therefore likely driven by an interaction between an inward persistent Na+current and time-dependent deactivation of Ih. These voltage-dependent conductances provide a mechanism for the generation of membrane potential oscillations that can help support rhythmic network activity within the parasubiculum during theta-related behaviors. [ABSTRACT FROM AUTHOR]

Abstract: Precision-cut liver slices and primary hepatocytes constitute suitable model systems for studying liver function. Frequently, urea cycle activity is used as a parameter to determine hepatocyte viability. Liver cells contain high levels of the urea cycle enzyme arginase, which converts arginine into urea and ornithine. Arginase can leak from the cells into the supernatants, converting arginine directly to urea and in this way circumventing the urea cycle. In this study, a hepatocellular cell line (HepG2 cells), a primary rat hepatocyte culture, and precision-cut rat liver slices were compared with respect to arginase leakage in the media by determining arginine conversion into urea. HepG2 cells did not show arginine conversion to urea during 24h incubations. In contrast, in both precision-cut liver slices and primary rat hepatocytes all arginine was converted to urea. Arginase activity was confirmed by showing that freshly added arginine to the cell-free supernatants again was converted to urea. In conclusion, when choosing urea production of primary hepatocytes cultures as a viability indicator, one has to take into account that arginase can leak from the cells into the supernatant. This can lead to an overestimation of the viability of the cells, since arginase converts arginine into urea without involvement of the urea cycle. We suggest using an extra incubation in an arginine-free buffer supplemented with ornithine and NH4Cl. In addition, arginase leakage can lead to depletion of the supernatant of arginine in primary hepatocytes cell cultures. This might have implications for studying cellular activities where arginine is involved, like, e.g. nitric oxide (NO) production. [Copyright &y& Elsevier]

The imaging of the brachial plexus (BP) cannot easily be achieved because of the extension of the region to be studied, the different tissue compositions of the adjacent anatomic structures and the necessity to work with thin tissue slices. Here is a description of the brachial plexus MRI (bpMRI) protocol that we use for the pre-operative evaluation of patients affected by the following brachial plexus (BP) pathologies: neoplastic, post-traumatic, inflammatory. The surgical implications are discussed. A survey was done on 115 patients (66 males; 49 females; mean age 46.4 years old), which underwent a bpMRI over a 32-month period (32 post-traumatic, 26 neoplastic, 25 affected by inflammatory diseases). Our bpMRI protocol is evaluated according to each mechanism of disease. The bpMRI provides a topographic localization of the tumour showing the nerve dislocations or infiltration by the neoplasm. In traumatic patients we suggest that the bpMRI could be useful to detect the degree of axonal damage when the nerve lesion is in continuity. We also consider the diagnosis and the possible surgical implications for the inflammatory plexopathies. In conclusion, a standardized bpMRI protocol has an invaluable diagnostic value for the surgeons and neurologists. It is only limited by its duration (at least 45 min). [ABSTRACT FROM AUTHOR]

Background/Aims: Cardiac function is increasingly studied using murine models. However, current multicellular preparations to investigate contractile properties have substantial technical and biological limitations and are especially difficult to apply to the developing murine heart. Methods: Newborn murine hearts were cut with a vibratome into viable tissue slices. The structural and functional integrity of the tissue was shown by histology, ATP content and sharp electrode recordings. Results: Within the first 48 hours after slicing structure remained intact without induction of apoptosis. ATP concentrations and action potential parameters were comparable to those of physiological tissue. Isometric force measurements demonstrated a physiological force-frequency relationship with a ‘primary-phase’ negative force-frequency relationship up to 1-2 Hz and a ‘secondary-phase’ positive force-frequency relationship up to 8 Hz. (-)-Isoproterenol (10-6 mol/l) increased active force to 251±35% (n=15) of baseline values and shortened relaxation times indicating a preserved beta-adrenergic regulation of contraction. Changes of the force-frequency relationship after application of ryanodine and nifedipine indicated functionality of calcium release from the sarcoplasmic reticulum and of L-type calcium channels. Conclusion: Generation of viable, physiological intact ventricular slices from neonatal hearts is feasible and provides a robust model to study loaded contractions. Copyright © 2007 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Recently Fourier Transform Infrared (FTIR) spectroscopic imaging has been used as a tool to detect the changes in cellular composition that may reflect the onset of a disease. This approach has been investigated as a mean of monitoring the change of the biochemical composition of cells and providing a diagnostic tool for various human cancers and other diseases. The discrimination between different types of tissue based upon spectroscopic data is often achieved using various multivariate clustering techniques. However, the number of clusters is a common unknown feature for the clustering methods, such as hierarchical cluster analysis, k-means and fuzzy c-means. In this study, we apply a FCM bused clustering algorithm to obtain the best number of clusters as given by the minimum validity index value. This often results in an excessive number of clusters being created due to the complexity of this biochemical system. A novel method to automatically merge clusters was developed to try to address this problem. Three lymph node tissue sections were examined to evaluate our new method. These results showed that this approach can merge the clusters which have similar biochemistry. Consequently, the overall algorithm automatically identifies clusters match accurately match the main tissue types that are independently determined by the clinician. [ABSTRACT FROM AUTHOR]

Cortical computations are an emergent property of neural dynamics. To understand how neural dynamics emerges within local cortical networks, we characterized the development and underlying mechanisms of spontaneous dynamics in cortical organotypic slices. We observed not only a quantitative increase in the levels of spontaneous dynamics, but a qualitative transition from brief bursts of activity to well defined Up states during the first 4 weeks in vitro. Analysis of cellular and synaptic properties indicates that these changes are driven by increasing excitatory drive accompanied by changes in the balance of excitation and inhibition. Examination of the structure of spontaneous dynamics revealed no evidence of precisely repeating patterns. Slices exposed to chronic patterned stimulation exhibited decreased levels of spontaneous activity, suggesting homeostatic control of the levels of network activity. Together, these results suggest that Up states reflect a fundamental mode of network dynamics that emerges through the orchestrated regulation of multiple cellular and synaptic properties in parallel. [ABSTRACT FROM AUTHOR]

The group I metabotropic glutamate receptor agonist ( S)-3,5-dihydroxyphenylglycine (DHPG) elicited two phases of synchronized neuronal (epileptiform) discharges in hippocampal slices: an initial phase of short duration discharges followed by a phase of prolonged discharges. We assessed the involvement of transient receptor potential canonical (TRPC) channels in these responses. Pre-treatment of hippocampal slices with TRPC channel blockers, 1-[beta-[3-(4-methoxyphenyl)propoxy]-4-methoxyphenethyl]-1H-imidazole hydrochloride (SKF96365) or 2-aminoethoxydiphenyl borate, did not affect the short epileptiform discharges but blocked the prolonged epileptiform discharges. SKF96365 suppressed ongoing DHPG-induced prolonged epileptiform discharges. Western blot analysis showed that the total TRPC4 or TRPC5 proteins in hippocampal slices were unchanged following DHPG. DHPG increased TRPC4 and TRPC5 in the cytoplasmic compartment and decreased these proteins in the plasma membrane. Translocation of TRPC4 and TRPC5 was suppressed when the epileptiform discharges were blocked by ionotropic glutamate receptor blockers. Translocation of TRPC4 and TRPC5 was also prevented in slices from phospholipase C (PLC) β1 knockout mice, even when synchronized discharges were elicited by the convulsant 4-aminopyridine. The results suggest that TRPC channels are involved in generating DHPG-induced prolonged epileptiform discharges. This function of TRPC channels is associated with a neuronal activity- and PLCβ1-dependent translocation of TRPC4 and TRPC5 proteins from the plasmalemma to the cytoplasmic compartment. [ABSTRACT FROM AUTHOR]

Arterial spin labeling (ASL) magnetically inverts or saturates the spins in arterial blood and uses them as endogenous tracers. Conventionally, the tagging band is upstream or nonselective to the target slices. In the brain, ASL-based functional magnetic resonance imaging (fMRI) has been shown to detect activation better localized in gray matter than blood oxygenation level dependent contrast. More recently, velocity selective-ASL (VS-ASL) was proposed to tag spins according to their flow velocity. One desirable characteristic of VS-ASL is its capability to generate tags sufficiently close to the target slices and thereby circumvent the complication of non-zero transit delay. In this study, we investigate the feasibility of VS-ASL in fMRI by comparing it with a conventional ASL method (PICORE). The results from the visual cortex of healthy volunteers show that VS-ASL and PICORE have comparable spatial specificity in detecting the flow change induced by neuronal activity. Velocity selective-arterial spin labeling can further distinguish the contribution from different flow directions but spurious elevation of fractional signal change may occur when the VS tagging is applied off the direction of blood supply. The flow reaches the vicinity of perfusion at a cutoff velocity (Vc) of 2 cm/sec whereas the activation exclusively detected by Vc=4 cm/sec implies the arteriolar response to the neuronal activity and a respondent vessel diameter up to 240 μm. Velocity selective imaging can remove intravascular signal from the vessels where the flow velocity is above Vc.Journal of Cerebral Blood Flow & Metabolism (2007) 27, 831–838. doi:10.1038/sj.jcbfm.9600386; published online 16 August 2006 [ABSTRACT FROM AUTHOR]

Abstract??Experiments with washing of carrot slices using distilled water and 0.9% (w/w) NaCl in combination with varying number and pulses of high intensity white were carried out. Slices of carrot 3.5?cm in diameter and 2?mm thick were inoculated using diluted dispersions of yeasts (Saccharomyces cerevisiae) for varying time (0?240?min) and then the slices were treated with high intensity pulsed white light (one pulse = 0.7?J/cm2) using from none to 24 pulses. The major part of the yeast cells were killed using two pulses of light. The studies included washing of inoculated slices up to five times using salt 0.9% (w/w) NaCl and distilled water and inoculation at 22??C for between 5 and 120?min. It was concluded that treatment of carrot slices with high intensity white light may reduce the load of yeast cells with up to 6?log cycles. Inoculation time at 22??C had no effect on the maximum load of yeast cells. [ABSTRACT FROM AUTHOR]

Abstract: The contribution of platelet-activating factor (PAF) to the induction of neocortical LTP was examined in rat brain slices containing the primary somatosensory cortex (SI). Field potentials evoked by single pulse stimulation in cortical layer IV were recorded from layer II/III. In control experiments, tetanic high frequency stimulation (HFS) resulted in input-specific, NMDA receptor-dependent LTP (21.1±3.2%; mean±SEM; n =15; P <0.001). BN-52021 (5 μM), an antagonist at the extracellular PAF receptor, weakened the HFS-induced LTP to 12.4±2.7% (n =11; P <0.05). In contrast, HFS-induced LTP was significantly enhanced to 29.4±2.3% (n =11; P <0.05) when brain slices were superfused with ACSF containing the PAF receptor-agonist C-PAF (1.5 μM). The difference between LTP weakened by BN-52021 and LTP enhanced by C-PAF was highly significant (P <0.0005). These results suggest a physiological contribution of PAF to the induction of LTP in neocortical area SI. This contribution of PAF is mediated by an action at extracellular receptor sites. [Copyright &y& Elsevier]

Type I collagen, the major organic component of bone matrix, undergoes a series of post-translational modifications that occur with aging, such as the non-enzymatic glycation. This spontaneous reaction leads to the formation of advanced glycation end products (AGEs), which accumulate in bone tissue and affect its structural and mechanical properties. We have investigated the role of matrix AGEs on bone resorption mediated by mature osteoclasts and the effects of exogenous AGEs on osteoclastogenesis. Using in vitro resorption assays performed on control- and AGE-modified bone and ivory slices, we showed that the resorption process was markedly inhibited when mature osteoclasts were seeded on slices containing matrix pentosidine, a well characterized AGE. More specifically, the total area resorbed per slice, and the area degraded per resorption lacuna created by osteoclasts, were significantly decreased in AGE-containing slices. This inhibition of bone resorption was confirmed by a marked reduction of the release of type I collagen fragments generated by the collagenolytic enzymes secreted by osteoclasts in the culture medium of AGE-modified mineralized matrices. This effect is likely to result from decreased solubility of collagen molecules in the presence of AGEs, as documented by the reduction of pepsin-mediated digestion of AGE-containing collagen. We found that AGE-modified BSA totally inhibited osteoclastogenesis in vitro, most likely by impairing the commitment of osteoclast progenitors into pre-osteoclastic cells. Although the mechanisms remain unknown, AGEs might interfere with osteoclastic differentiation and activity through their interaction with specific cell-surface receptors, because we showed that both osteoclast progenitors and mature osteoclasts expressed different AGEs receptors, including receptor for AGEs (RAGEs). These results suggest that AGEs decreased osteoclast-induced bone resorption, by altering not only the structural integrity of bone matrix proteins but also the osteoclastic differentiation process. We suggest that AGEs may play a role in the alterations of bone remodeling associated with aging and diabetes. [ABSTRACT FROM AUTHOR]

What regulates the spread of activity through cortical circuits? We present here data indicating a pivotal role for a vetoing inhibition restraining modules of pyramidal neurons. We combined fast calcium imaging of network activity with whole-cell recordings to examine epileptiform propagation in mouse neocortical slices. Epileptiform activity was induced by washing Mg2+ ions out of the slice. Pyramidal cells receive barrages of inhibitory inputs in advance of the epileptiform wave. The inhibitory barrages are effectively nullified at low doses of picrotoxin (2.5-5 µM).When present, however, these inhibitory barrages occlude an intense excitatory synaptic drive that would normally exceed action potential threshold by approximately a factor of 10. Despite this level of excitation, the inhibitory barrages suppress firing, thereby limiting further neuronal recruitment to the ictal event. Pyramidal neurons are recruited to the epileptiform event once the inhibitory restraint fails and are recruited in spatially clustered populations (150-250 µm diameter). The recruitment of the cells within a given module is virtually simultaneous, and thus epileptiform events progress in intermittent (0.5-1 Hz) steps across the cortical network. We propose that the interneurons that supply the vetoing inhibition define these modular circuit territories. [ABSTRACT FROM AUTHOR]

GABA is the inhibitory neurotransmitter in most brain stem nuclei. The properties of release of preloaded [3H]GABA were now investigated with slices from the mouse brain stem under normal and ischemic (oxygen and glucose deprivation) conditions, using a superfusion system. The ischemic GABA release increased about fourfold in comparison with normal conditions. The tyrosine kinase inhibitor genistein had no effect on GABA release, while the phospholipase inhibitor quinacrine reduced both the basal and K+-evoked release in normoxia and ischemia. The activator of protein kinase C (PKC) 4β-phorbol 12-myristate 13-acetate had no effects on the releases, whereas the PKC inhibitor chelerythrine reduced the basal release in ischemia. When the cyclic guanosine monophosphate (cGMP) levels were increased by superfusion with zaprinast and other phosphodiesterase inhibitors, GABA release was reduced under normal conditions. The NO donors S-nitroso- N-acetylpenicillamine (SNAP) and hydroxylamine (HA) enhanced the basal and K+-stimulated release by acting directly on presynaptic terminals. Under ischemic conditions GABA release was enhanced when cGMP levels were increased by zaprinast. This effect was confirmed by inhibition of the release by the guanylate cyclase inhibitor 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ). The NO-producing agents SNAP, HA, and sodium nitroprusside potentiated GABA release in ischemia. These effects were reduced by the NO synthase inhibitor NG-nitro- l-arginine, but not by ODQ. The results show that particularly NO and cGMP regulate both normal and ischemic GABA release in the brain stem. Their effects are however complex. [ABSTRACT FROM AUTHOR]

In vitro studies often use bicarbonate-buffered saline solutions to mimic the normal extracellular environment of tissues. These solutions are typically equilibrated with gaseous O2 and CO2, the latter interacting with bicarbonate ions to maintain a physiological pH. In vitro tissue chambers, like those used for electrophysiology, are usually continually perfused with the gassed buffer, but stopping the perfusion to add expensive chemicals or acquire imaging data is a common practice. The present study demonstrates that this procedure leads to rapid (< 30 s) increases in pH and decreases in PO2 of the detained solution in the tissue chamber. During the first 200 s, pH increased by 0.4 units and resulted in a 25% PO2 reduction of the detained solution. The rates of these changes were dependent on the volume of solution in the chamber. In experiments using acute transverse slices from the lumbar spinal cord of neonatal (postnatal day 0–10) mice, perfusion stoppage of the same duration was accompanied by a 34.7% enhancement of the peak voltage-gated calcium current recorded from ventral horn neurons. In these cells both low voltage-activated and high voltage-activated currents were affected. These currents were unaffected by decreasing PO2 when a CO2-independent buffer was used, suggesting that changes in pH were responsible for the observed effects. It is concluded that the procedure of stopping a bicarbonate/CO2-buffered perfusate results in rapid changes in pH and PO2 of the solution detained in the tissue chamber, and that these changes have the potential to covertly influence experimental results. [ABSTRACT FROM AUTHOR]

We report on the recording of electrical activity in cultured hippocampal slices by a multitransistor array (MTA) with 16,384 elements. Time-resolved imaging is achieved with a resolution of 7.8 µm on an area of 1 mm2 at 2 kHz. A read-out of fewer elements allows an enhanced time resolution. Individual transistor signals are caused by local evoked field potentials. They agree with micropipette measurements in amplitude and shape. The spatial continuity of the records provides time-resolved images of evoked field potentials and allows the detection of functional correlations over large distances. As examples, fast propagating waves of presynaptic action potentials are recorded as well as patterns of excitatory postsynaptic potentials across and along cornu ammonis. [ABSTRACT FROM AUTHOR]

Abstract: The objective of the present study was to evaluate the stability of cytochrome P450 enzymes and of the conjugation enzyme systems epoxide hydrolase, glucuronosyl transferase, sulphotransferase and glutathione S-transferase in precision-cut rat lung slices incubated in RPMI media for different time periods up to 72h. Moreover, the effect of culturing of lung slices on total glutathione levels and glutathione reductase was also investigated. Monitoring of cytochrome P450 activity was achieved using established diagnostic probes, but when activity in the lung was low the maintenance of the various enzymes in culture was determined immunologically using Western blotting. The dealkylation of pentoxyresorufin declined markedly during the first 4h of incubation but in the case of ethoxyresorufin loss of activity was more gradual and less severe. Western blot analysis revealed that the rate of decrease in cytochrome P450 apoprotein levels was isoform-specific with CYP2E1 being the most stable and CYP3A the least stable. Generally, phase II activities, especially cytosolic sulphotransferase, were relatively more stable throughout the incubation period compared with cytochromes P450. Finally, glutathione reductase activity and total glutathione levels were maintained throughout the 72h incubation. The present studies indicate that xenobiotic-metabolising enzymes in precision-cut rat lung slices decline in culture, but the rate of loss differs and depends on the nature of the enzyme. [Copyright &y& Elsevier]

An integrated system is proposed for real-time imaging of tissue deformation in response to externally applied mechanical compression. The purpose of the system is to noninvasively detect tissue lesions based on their stiffness. The method includes applying external compression to the examined subject and acquiring magnetic resonance strain-encoded images using a real-time pulse sequence. In the proposed system, only a single compression is necessary to obtain a two-dimensional map of tissue deformation in a slice, eliminating the need for a complicated compressing device. The system was built and validated using phantom experiments. [ABSTRACT FROM AUTHOR]

Abstract: Cell-attached recording provides a way to record the activity of – and to stimulate – neurons in brain slices without rupturing the cell membrane. This review uses theory and experimental data to address the proper application of this technique and the correct interpretation of the data. Voltage-clamp mode is best-suited for recording cell firing activity, and current-clamp mode is best-suited for recording resting membrane potential and synaptic potentials. The magnitude of the seal resistance determines what types of experiments can be accomplished with a cell-attached recording: a loose seal is adequate for recording action potential currents, and a tight seal is required for evoking action potentials in the attached cell and for recording resting and synaptic potentials. When recording action potential currents, if the researcher does not want to change the firing activity of the cell, then it is important that no current passes from the amplifier through the patch resistance. In order to accomplish this condition, the recording pipette should be held at the potential that gives a holding current of 0. An advantage of cell-attached current-clamp over whole-cell recording is that it accurately depicts whether a synaptic potential is hyperpolarizing or depolarizing without the risk of changing its polarity. [Copyright &y& Elsevier]

Abstract: Cerebrocortical neurons that store and release zinc synaptically are widely recognized as critical in maintenance of cortical excitability and in certain forms of brain injury and disease. Through the last 20 years, this synaptic release has been observed directly or indirectly and reported in more than a score of publications from over a dozen laboratories in eight countries. However, the concentration of zinc released synaptically has not been established with final certainty. In the present work we have considered six aspects of the methods for studying release that can affect the magnitude of zinc release, the imaging of the release, and the calculated concentration of released zinc. We present original data on four of the issues and review published data on two others. We show that common errors can cause up to a 3000-fold underestimation of the concentration of released zinc. The results should help bring consistency to the study of synaptic release of zinc. [Copyright &y& Elsevier]

Abstract: Cytochrome P-450s (CYPs) detoxify a wide variety of xenobiotics and environmental contaminants, but can also bioactivate carcinogenic polycyclic aromatic hydrocarbons, such as benzo(a)pyrene (BaP), to DNA-reactive species. The primary CYPs involved in the metabolism and bioactivation of BaP are CYP1A1 and CYP1B1. Furthermore, BaP can induce expression of CYP1A1 and CYP1B1 via the aryl hydrocarbon receptor. Induction of CYP1A1 and CYP1B1 by BaP in target (lung) and non-target (liver) tissues was investigated utilizing precision-cut rat liver and lung slices exposed to BaP in vitro. Tissue slices were also prepared from rats pretreated in vivo with 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) to induce expression of CYP1A1 and CYP1B1. In addition, in vivo exposure studies were performed with BaP to characterize and validate the use of the in vitro tissue slice model. In vitro exposure of liver and lung slices to BaP resulted in a concentration-dependent increase in CYP1A1 and CYP1B1 mRNA and protein levels, which correlated directly with the exposure-related increase in BaP-DNA adduct levels observed previously in the tissue slices [Harrigan, J.A., Vezina, C.M., McGarrigle, B.P., Ersing, N., Box, H.C., Maccubbin, A.E., Olson, J.R., 2004. DNA adduct formation in precision-cut rat liver and lung slices exposed to benzo(a)pyrene. Toxicological Sciences 77, 307–314]. Pretreatment of animals in vivo with TCDD produced a marked induction of CYP1A1 and CYP1B1 expression in the tissue slices, which was similar to the levels of CYP1A1 and CYP1B1 mRNA achieved in liver and lung following in vivo treatment with BaP. Following in vitro exposure to BaP, the levels of CYP1A1 were greater in the lung than the liver, while following all exposures (in vitro and in vivo), the levels of CYP1B1 mRNA were greater in lung tissue compared to liver. The higher expression of CYP1A1 and CYP1B1 in the lung was associated with higher levels of BaP-DNA adducts in the lung slices (Harrigan et al.’s work) and together, these results may contribute to the tissue specificity of BaP-mediated carcinogenesis. [Copyright &y& Elsevier]

Whereas virotherapy has emerged as a novel and promising approach for neoplastic diseases, appropriate model systems have hampered preclinical evaluation of candidate conditionally replicative adenovirus agents (CRAds) with respect to liver toxicity. This is due to the inability of human viral agents to cross species. We have recently shown the human liver tissue slice model to be a facile means to validate adenoviral replication. On this basis, we sought to determine whether our ex vivo liver tissue slice model could be used to assess CRAd-mediated liver toxicity. We analyzed and compared the toxicity of a conditionally replicative adenovirus (AdΔ24) to that of a replication incompetent adenovirus (Adnull [E1−]) in mouse and human liver tissue slices. To accomplish this, we examined the hepatic apoptosis expression profile by DNA microarray analyses, and compared these results to extracellular release of aminotransferase enzymes, along with direct evidence of apoptosis by caspase-3 immunhistochemical staining and TUNEL assays. Human and mouse liver tissue slices demonstrated a marked increase in extracellular release of aminotransferase enzymes on infection with AdΔ24 compared to Adnull. AdΔ24-mediated liver toxicity was further demonstrated by apoptosis induction, as detected by caspase-3 immunohistochemical staining, TUNEL assay and microarray analysis. In conclusion, concordance of CRAd-mediated apoptosis in both the human and the mouse liver tissue slice models was demonstrated, despite the limited replication ability of CRAds in mouse liver slices. The results of this study, defining the CRAd-mediated apoptosis gene expression profiles in human and mouse liver, may lay a foundation for preclinical liver toxicity analysis of CRAd agents.Cancer Gene Therapy (2006) 13, 606–618. doi:10.1038/sj.cgt.7700934; published online 6 January 2006 [ABSTRACT FROM AUTHOR]

The aim of this study was to investigate whether synaptic plasticity and metaplasticity in slice cultures of the young rat hippocampus were comparable to previously reported synaptic plasticity and metaplasticity in acute adult hippocampal slices. This is relevant since differences do exist between the preparations as a result of age and the ex vivo maintenance. We prepared and maintained slice cultures from 5- to 6-day-old rats according to the porous membrane method. After 12–16 days in vitro, extracellular low-frequency stimulation (LFS) and high-frequency stimulation (HFS) protocols were applied to the Schaffer collaterals, and extracellular field potentials were recorded in area CA1. LFS and HFS induced long-term depression (LTD) and long-term potentiation (LTP), respectively. LTP could be reversed by LFS, as could LTD by HFS 60 min after induction. Plotting the amount of LTD and LTP versus stimulation protocol demonstrated frequency-dependence of the sign and extent of plasticity. Priming activation of group 1 metabotropic glutamate receptors (mGluRs) with DHPG facilitated subsequent LTP, revealing a metaplastic effect similar to that observed in acute slices. Immunohistochemistry for group 1 mGluR subtypes mGluR1α and mGluR5 showed both receptors to be present in these cultures. We conclude that synaptic plasticity and mGluR-mediated metaplasticity are largely comparable to those effects found in acute in vitro techniques. [ABSTRACT FROM AUTHOR]

Abstract: Although much interest has attended the cryopreservation of immature neurons for subsequent therapeutic intracerebral transplantation, there are no reports on the cryopreservation of organized adult cerebral tissue slices of potential interest for pharmaceutical drug development. We report here the first experiments on cryopreservation of mature rat transverse hippocampal slices. Freezing at 1.2°C/min to −20°C or below using 10 or 30% v/v glycerol or 20% v/v dimethyl sulfoxide yielded extremely poor results. Hippocampal slices were also rapidly inactivated by simple exposure to a temperature of 0°C in artificial cerebrospinal fluid (aCSF). This effect was mitigated somewhat by 0.8mM vitamin C, the use of a more “intracellular” version of aCSF having reduced sodium and calcium levels and higher potassium levels, and the presence of a 25% w/v mixture of dimethyl sulfoxide, formamide, and ethylene glycol (“VEG solutes”; Cryobiology 48, pp. 22–35, 2004). It was not mitigated by glycerol, aspirin, indomethacin, or mannitol addition to aCSF. When RPS-2 (Cryobiology 21, pp. 260–273, 1984) was used as a carrier solution for up to 50% w/v VEG solutes, 0°C was more protective than 10°C. Raising VEG concentration to 53% w/v allowed slice vitrification without injury from vitrification and rewarming per se, but was much more damaging than exposure to 50% w/v VEG. This problem was overcome by using the analogous 61% w/v VM3 vitrification solution (Cryobiology 48, pp. 157–178, 2004) containing polyvinylpyrrolidone and two extracellular “ice blockers.” With VM3, it was possible to attain a tissue K+/Na+ ratio after vitrification ranging from 91 to 108% of that obtained with untreated control slices. Microscopic examination showed severe damage in frozen–thawed slices, but generally good to excellent ultrastructural and histological preservation after vitrification. Our results provide the first demonstration that both the viability and the structure of mature organized, complex neural networks can be well preserved by vitrification. These results may assist neuropsychiatric drug evaluation and development and the transplantation of integrated brain regions to correct brain disease or injury. [Copyright &y& Elsevier]

Abstract: Cryopreservation offers the potential to maximize the use and availability of biological materials that have a limited supply. This study demonstrates an enhanced technique for the parallel cryopreservation of a series of liver tissue slices using a tray modeled from aluminium foil and low concentrations of a cryoprotectant. Cooling and warming rates of approximately 2000 and 3900°Cmin−1, respectively, were achieved as the thermal capacity of the foil-tray was significantly reduced compared to the aluminium sandwich device introduced by Day et al. [S.H. Day, D.A. Nicoll-Griffith, J.M. Silva, Cryopreservation of rat and human liver slices by rapid freezing, Cryobiology 38 (1999) 154–159]. Additionally, the two critical steps involved in the sandwich approach, i.e., clamping the plates and complete filling of the entire space between the plates with liquid, can be omitted using the foil tray. The viability of the slices was verified by measuring tetrazolium salt reduction capacity, cytosolic enzyme lactate dehydrogenase leakage, and ethoxycoumarin metabolism. [Copyright &y& Elsevier]

This paper describes the effect of ohmic heating on juice yield from potato and apple tissues. Two different types of experiments involving different treatment chambers were carried out: textural and conductivity study of cylindrical samples and juice yield tests of tissue slices. The investigations show how tissue disintegration degree and juice yield depend on the field intensity, temperature, treatment duration and type of plant tissue. The best juice extraction was observed when the plant tissue was treated electrically at a moderate temperature of 50°C. It was explained by the combined effect, related to electroporation of cell membranes and thermal softening of tissue. [Copyright &y& Elsevier]

Abstract: The differential toxicity of two anticancer agents is described using the in vitro rat liver slice culture model. Liver slices from F-344 rats were cultured for 5days in Waymouth’s-based medium with exposure to a range of geldanamycin (GEL) or 17-allylaminogeldanamycin (17-AAG) concentrations. GEL induced concentration-dependent reduction of alkaline phosphatase and of γ-glutamyl transferase levels, which are indicators of biliary epithelial cell(s) (BEC) viability, and exhibited hepatocellular toxicity at higher concentrations. Histologically, BEC cell injury was evident at the lowest GEL concentration (0.1μM) and progressed to overt bile duct necrosis at 5μM, a level at which hepatocellular damage was also more prominent. Slices exposed to the same concentrations were more sensitive to toxic effects of GEL than of 17-AAG. 17-AAG at the lowest concentration had more slice biomarker retention than GEL, and histological analysis revealed minimal toxic effect on BEC. With increasing concentration, BEC were progressively lost, and BEC proliferation was completely inhibited at 5μM 17-AAG. Hepatocellular injury was evident only at high dose exposures. This is believed to be the first use of an in vitro liver tissue model to accurately predict the differential and concentration-dependent toxicities of these compounds. [Copyright &y& Elsevier]

Maximum intensity projections reconstructions from 2.5 mm unenhanced multidetector computed tomography axial slices were obtained from 49 patients within the first 6 h of anterior-circulation cerebral strokes to identify different patterns of the dense artery sign and their prognostic implications for location and extent of the infarcted areas. The dense artery sign was found in 67.3% of cases. Increased density of the whole M1 segment with extension to M2 of the middle cerebral artery was associated with a wider extension of cerebral infarcts in comparison to M1 segment alone or distal M1 and M2. A dense sylvian branch of the middle cerebral artery pattern was associated with a more restricted extension of infarct territory. We found 62.5% of patients without a demonstrable dense artery to have a limited peripheral cortical or capsulonuclear lesion. In patients with a 7-10 points on the Alberta Stroke Early Programme Computed Tomography Score and a dense proximal MCA in the first hours of ictus the mean decrease in the score between baseline and follow-up was 5.09+/-1.92 points. In conclusion, maximum intensity projections from thin-slice images can be quickly obtained from standard computed tomography datasets using a multidetector scanner and are useful in identifying and correctly localizing the dense artery sign, with prognostic implications for the entity of cerebral damage. [ABSTRACT FROM AUTHOR]

We have reported that long-term potentiation (LTP) can be reliably induced in motor cortex of adult, freely moving rats by the application of spaced and repeated high frequency stimulating trains to the white matter. In the present study, we monitored field potentials (FPs) and LTP in both layer II/III and V in coronal slices of motor cortex taken from implanted control and previously potentiated Long–Evans rats. The baseline FP amplitudes were decreased in layer II/III, and the amplitude of small spikes was significantly increased in layer V in slices from previously potentiated rats compared to unpotentiated control rats. In response to high frequency stimulation applied to the slice itself, both implanted control and previously potentiated rats showed similar levels of LTP in layer II/III. LTP could not be induced in layer V. These results show that layer II/III and V respond differently to high frequency stimulation in vitro. In addition, layer II/III responds very differently in slice compared to chronic preparations. [ABSTRACT FROM AUTHOR]

Acetabular angles (AAs) and dorsal acetabular rim angles acquired by computed tomographic (CT) imaging have been used to assess patient response to juvenile pubic symphysiodesis surgery. The purpose of this study was to evaluate the effects of patient positioning and slice selection on these angles, and an attempt was made to devise a repeatable method of measuring these angles that would eliminate positioning effects. We found significant variation in AAs with small differences in pelvic tilt and slice selection. Dorsal acetabular rim angles were not affected. As a result of positioning differences from one CT study to the next, every attempt should be made to standardize pelvic tilt, or eliminate its effect on AAs by standardizing gantry angle in relation to an anatomic landmark that will not change over time. The floor of the sacral vertebral canal may be a reasonable landmark for this purpose and deserves further study. [ABSTRACT FROM AUTHOR]

We have found that two-photon fluorescence imaging of nicotinamide adenine dinucleotide (NADH) provides the sensitivity and spatial three-dimensional resolution to resolve metabolic signatures in processes of astrocytes and neurons deep in highly scattering brain tissue slices. This functional imaging reveals spatiotemporal partitioning of glycolytic and oxidative metabolism between astrocytes and neurons during focal neural activity that establishes a unifying hypothesis for neurometabolic coupling in which early oxidative metabolism in neurons is eventually sustained by late activation of the astrocyte-neuron lactate shuttle. Our model integrates existing views of brain energy metabolism and is in accord with known macroscopic physiological changes in vivo. [ABSTRACT FROM AUTHOR]

Taking tissue slices of the embryonic and newborn pancreas is a novel approach for the study of the perinatal development of this gland. The aim of this study was to describe the morphology and physiology of in vivo and in vitro developing β-cells. In addition, we wanted to lay a foundation for the functional analysis of other pancreatic cells, either alone or as part of an integrative pancreatic physiology approach. We used cytochemistry and light microscopy to detect specific markers and the whole-cell patch-clamp to assess the function of single β-cells. The insulin signal in the embryonic β-cells was condensed to a subcellular compartment and redistributed throughout the cytosol during the first 2 days after birth. The hormone distribution correlated well with the development of membrane excitability and hormone release competence in β-cells. Endocrine cells survived in the organotypic tissue culture and maintained their physiological properties for weeks. We conclude that our preparation fulfills the criteria for a method of choice to characterize the function of developing pancreas in wild-type and genetically modified mice that die at birth. We suggest organotypic culture for in vitro studies of the development and regeneration of β-cells. [ABSTRACT FROM AUTHOR]

Nitric oxide generation from porcine kidney slices is assessed using a new planar NO-selective amperometric sensor. The planar shape of the sensor allows for direct NO measurements near the surface (10 μm) of renal tissue slices in real time. Renal NO production may be modulated by the addition of L-arginine, arginine homopolymers (R2, R6, R10), and protamine, all of which can potentially transport across cellular membranes and provide a substrate for nitric oxide synthase within kidney parenchyma. Real-time amperometric measurements demonstrate that most L-arginine species can translocate across the cell membrane and rapidly increase NO production. However, no increase in NO generation is observed when the dimer of L-arginine (R2) is added to the solution bathing the tissue, suggesting that this species cannot permeate cell membranes. The degree of enhancement in NO generation observed for L-arginine and the larger peptides depends on the structure and follows the following sequence: R10 (decanter) > protamine > R6 (hexamer) > L-arginine. Protamine and the R10 decamer, especially, induce the largest increases in NO generation owing to theft apparent rapid translocation into cells and subsequent cleavage by proteases to create high intracellular levels of L-arginine. The effect of sensor size (for sensor dimensions of 0.15- and 1-mm outer diameters) on the measured surface NO levels is also examined. The larger sensor traps more NO but hinders access of the L-arginine species to the tissue area between the flat distal plane of the sensor and the surface of the kidney slice. The use of such NO-generating peptides may be important in numerous biological systems that depend on NO production, such as ischemia-reperfusion injury and thrombogenesis. [ABSTRACT FROM AUTHOR]