An image-domain, contrast material extraction method for Dual-Energy CT

PubMed Central

Lambert, Jack W.; Sun, Yuxin; Gould, Robert G.; Ohliger, Michael A.; Li, Zhixi; Yeh, Benjamin M.

2016-01-01

Objectives Conventional material decomposition techniques for dual-energy CT (DECT) assume mass or volume conservation, where the CT number of each voxel is fully assigned to predefined materials. We present an image-domain contrast material extraction process (CMEP) method that preferentially extracts contrast-producing materials while leaving the remaining image intact. Materials and Methods Image processing freeware (Fiji) is used to perform consecutive arithmetic operations on a dual-energy ratio map to generate masks, which are then applied to the original images to generate material-specific images. First, a low-energy image is divided by a high-energy image to generate a ratio map. The ratio map is then split into material-specific masks. Ratio intervals known to correspond to particular materials (e.g. iodine, calcium) are assigned a multiplier of 1, while ratio values in between these intervals are assigned linear gradients from 0 to 1. The masks are then multiplied by an original CT image to produce material-specific images. The method was tested quantitatively at Dual-Source (DSCT) and Rapid kVp-Switching CT (RSCT) with phantoms using pure and mixed formulations of tungsten, calcium and iodine. Errors were evaluated by comparing the known material concentrations with those derived from the CMEP material-specific images. Further qualitative evaluation was performed in vivo at RSCT with a rabbit model using identical CMEP parameters to the phantom. Orally administered tungsten, vascularly administered iodine, and skeletal calcium were used as the three contrast materials. Results All five material combinations; tungsten, iodine and calcium, and mixtures of tungsten-calcium and iodine-calcium, showed distinct dual-energy ratios, largely independent of material concentration at both DSCT and RSCT. The CMEP was successful in both phantoms and in vivo. For pure contrast materials in the phantom, the maximum error between the known and CMEP-derived material concentrations was 0.9 mg/mL, 24.9 mg/mL and 0.4 mg/mL for iodine, calcium and tungsten respectively. Mixtures of iodine and calcium showed the highest discrepancies, which reflected the sensitivity of iodine to the image-type chosen for the extraction of the final material-specific image. The rabbit model was able to clearly show the three extracted material phases, vascular iodine, oral tungsten and skeletal calcium. Some skeletal calcium was misassigned to the extracted iodine image, however this did not impede the depiction of the vasculature. Conclusions The CMEP is a straightforward, image domain approach to extract material signal at dual-energy CT. It has particular value for separation of experimental high-Z contrast elements from conventional iodine contrast or calcium, even when the exact attenuation coefficient profiles of desired contrast materials may be unknown. The CMEP is readily implemented in the image-domain within freeware, and can be adapted for use with images from multiple vendors. PMID:27875338

Dual energy micro CT SkyScan 1173 for the characterization of urinary stone

NASA Astrophysics Data System (ADS)

Fitri, L. A.; Asyana, V.; Ridwan, T.; Anwary, F.; Soekersi, H.; Latief, F. D. E.; Haryanto, F.

2016-03-01

Knowledge of the composition of urinary stones is an essential part to determine suitable treatments for patients. The aim of this research is to characterize the urinary stones by using dual energy micro CT SkyScan 11173. This technique combines high-energy and low- energy scanning during a single acquisition. Six human urinary stones were scanned in vitro using 80 kV and 120 kV micro CT SkyScan 1173. Projected images were produced by micro CT SkyScan 1173 and then reconstructed using NRecon (in-house software from SkyScan) to obtain a complete 3D image. The urinary stone images were analysed using CT analyser to obtain information of internal structure and Hounsfield Unit (HU) values to determine the information regarding the composition of the urinary stones, respectively. HU values obtained from some regions of interest in the same slice are compared to a reference HU. The analysis shows information of the composition of the six scanned stones obtained. The six stones consist of stone number 1 (calcium+cystine), number 2 (calcium+struvite), number 3 (calcium+cystine+struvite), number 4 (calcium), number 5 (calcium+cystine+struvite), and number 6 (calcium+uric acid). This shows that dual energy micro CT SkyScan 1173 was able to characterize the composition of the urinary stone.

Membrane Potential and Calcium Dynamics in Beta Cells from Mouse Pancreas Tissue Slices: Theory, Experimentation, and Analysis.

PubMed

Dolenšek, Jurij; Špelič, Denis; Klemen, Maša Skelin; Žalik, Borut; Gosak, Marko; Rupnik, Marjan Slak; Stožer, Andraž

2015-10-28

Beta cells in the pancreatic islets of Langerhans are precise biological sensors for glucose and play a central role in balancing the organism between catabolic and anabolic needs. A hallmark of the beta cell response to glucose are oscillatory changes of membrane potential that are tightly coupled with oscillatory changes in intracellular calcium concentration which, in turn, elicit oscillations of insulin secretion. Both membrane potential and calcium changes spread from one beta cell to the other in a wave-like manner. In order to assess the properties of the abovementioned responses to physiological and pathological stimuli, the main challenge remains how to effectively measure membrane potential and calcium changes at the same time with high spatial and temporal resolution, and also in as many cells as possible. To date, the most wide-spread approach has employed the electrophysiological patch-clamp method to monitor membrane potential changes. Inherently, this technique has many advantages, such as a direct contact with the cell and a high temporal resolution. However, it allows one to assess information from a single cell only. In some instances, this technique has been used in conjunction with CCD camera-based imaging, offering the opportunity to simultaneously monitor membrane potential and calcium changes, but not in the same cells and not with a reliable cellular or subcellular spatial resolution. Recently, a novel family of highly-sensitive membrane potential reporter dyes in combination with high temporal and spatial confocal calcium imaging allows for simultaneously detecting membrane potential and calcium changes in many cells at a time. Since the signals yielded from both types of reporter dyes are inherently noisy, we have developed complex methods of data denoising that permit for visualization and pixel-wise analysis of signals. Combining the experimental approach of high-resolution imaging with the advanced analysis of noisy data enables novel physiological insights and reassessment of current concepts in unprecedented detail.

Calcification detection of abdominal aorta in CT images and 3D visualization in VR devices.

PubMed

Garcia-Berna, Jose A; Sanchez-Gomez, Juan M; Hermanns, Judith; Garcia-Mateos, Gines; Fernandez-Aleman, Jose L

2016-08-01

Automatic calcification detection in abdominal aorta consists of a set of computer vision techniques to quantify the amount of calcium that is found around this artery. Knowing that information, it is possible to perform statistical studies that relate vascular diseases with the presence of calcium in these structures. To facilitate the detection in CT images, a contrast is usually injected into the circulatory system of the patients to distinguish the aorta from other body tissues and organs. This contrast increases the absorption of X-rays by human blood, making it easier the measurement of calcifications. Based on this idea, a new system capable of detecting and tracking the aorta artery has been developed with an estimation of the calcium found surrounding the aorta. Besides, the system is complemented with a 3D visualization mode of the image set which is designed for the new generation of immersive VR devices.

Imaging with organic indicators and high-speed charge-coupled device cameras in neurons: some applications where these classic techniques have advantages.

PubMed

Ross, William N; Miyazaki, Kenichi; Popovic, Marko A; Zecevic, Dejan

2015-04-01

Dynamic calcium and voltage imaging is a major tool in modern cellular neuroscience. Since the beginning of their use over 40 years ago, there have been major improvements in indicators, microscopes, imaging systems, and computers. While cutting edge research has trended toward the use of genetically encoded calcium or voltage indicators, two-photon microscopes, and in vivo preparations, it is worth noting that some questions still may be best approached using more classical methodologies and preparations. In this review, we highlight a few examples in neurons where the combination of charge-coupled device (CCD) imaging and classical organic indicators has revealed information that has so far been more informative than results using the more modern systems. These experiments take advantage of the high frame rates, sensitivity, and spatial integration of the best CCD cameras. These cameras can respond to the faster kinetics of organic voltage and calcium indicators, which closely reflect the fast dynamics of the underlying cellular events.

Lead in calcium supplements.

PubMed Central

Scelfo, G M; Flegal, A R

2000-01-01

Intercalibrated measurements of lead in calcium supplements indicate the importance of rigorous analytical techniques to accurately quantify contaminant exposures in complex matrices. Without such techniques, measurements of lead concentrations in calcium supplements may be either erroneously low, by as much as 50%, or below the detection limit needed for new public health criteria. In this study, we determined the lead content of 136 brands of supplements that were purchased in 1996. The calcium in the products was derived from natural sources (bonemeal, dolomite, or oyster shell) or was synthesized and/or refined (chelated and nonchelated calcium). The dried products were acid digested and analyzed for lead by high resolution-inductively coupled plasma-mass spectrometry. The method's limit of quantitation averaged 0.06 microg/g, with a coefficient of variation of 1.7% and a 90-100% lead recovery of a bonemeal standard reference material. Two-thirds of those calcium supplements failed to meet the 1999 California criteria for acceptable lead levels (1.5 microg/daily dose of calcium) in consumer products. The nonchelated synthesized and/or refined calcium products, specifically antacids and infant formulas, had the lowest lead concentrations, ranging from nondetectable to 2.9 microg Pb/g calcium, and had the largest proportion of brands meeting the new criteria (85% of the antacids and 100% of the infant formulas). Images Figure 1 Figure 2 PMID:10753088

In vivo imaging of neural activity

PubMed Central

Yang, Weijian; Yuste, Rafael

2017-01-01

Since the introduction of calcium imaging to monitor neuronal activity with single-cell resolution, optical imaging methods have revolutionized neuroscience by enabling systematic recordings of neuronal circuits in living animals. The plethora of methods for functional neural imaging can be daunting to the nonexpert to navigate. Here we review advanced microscopy techniques for in vivo functional imaging and offer guidelines for which technologies are best suited for particular applications. PMID:28362436

Skin suturing and cortical surface viral infusion improves imaging of neuronal ensemble activity with head-mounted miniature microscopes.

PubMed

Li, Xinjian; Cao, Vania Y; Zhang, Wenyu; Mastwal, Surjeet S; Liu, Qing; Otte, Stephani; Wang, Kuan Hong

2017-11-01

In vivo optical imaging of neural activity provides important insights into brain functions at the single-cell level. Cranial windows and virally delivered calcium indicators are commonly used for imaging cortical activity through two-photon microscopes in head-fixed animals. Recently, head-mounted one-photon microscopes have been developed for freely behaving animals. However, minimizing tissue damage from the virus injection procedure and maintaining window clarity for imaging can be technically challenging. We used a wide-diameter glass pipette at the cortical surface for infusing the viral calcium reporter AAV-GCaMP6 into the cortex. After infusion, the scalp skin over the implanted optical window was sutured to facilitate postoperative recovery. The sutured scalp was removed approximately two weeks later and a miniature microscope was attached above the window to image neuronal activity in freely moving mice. We found that cortical surface virus infusion efficiently labeled neurons in superficial layers, and scalp skin suturing helped to maintain the long-term clarity of optical windows. As a result, several hundred neurons could be recorded in freely moving animals. Compared to intracortical virus injection and open-scalp postoperative recovery, our methods minimized tissue damage and dura overgrowth underneath the optical window, and significantly increased the experimental success rate and the yield of identified neurons. Our improved cranial surgery technique allows for high-yield calcium imaging of cortical neurons with head-mounted microscopes in freely behaving animals. This technique may be beneficial for other optical applications such as two-photon microscopy, multi-site imaging, and optogenetic modulation. Published by Elsevier B.V.

NeuroCa: integrated framework for systematic analysis of spatiotemporal neuronal activity patterns from large-scale optical recording data

PubMed Central

Jang, Min Jee; Nam, Yoonkey

2015-01-01

Abstract. Optical recording facilitates monitoring the activity of a large neural network at the cellular scale, but the analysis and interpretation of the collected data remain challenging. Here, we present a MATLAB-based toolbox, named NeuroCa, for the automated processing and quantitative analysis of large-scale calcium imaging data. Our tool includes several computational algorithms to extract the calcium spike trains of individual neurons from the calcium imaging data in an automatic fashion. Two algorithms were developed to decompose the imaging data into the activity of individual cells and subsequently detect calcium spikes from each neuronal signal. Applying our method to dense networks in dissociated cultures, we were able to obtain the calcium spike trains of ∼1000 neurons in a few minutes. Further analyses using these data permitted the quantification of neuronal responses to chemical stimuli as well as functional mapping of spatiotemporal patterns in neuronal firing within the spontaneous, synchronous activity of a large network. These results demonstrate that our method not only automates time-consuming, labor-intensive tasks in the analysis of neural data obtained using optical recording techniques but also provides a systematic way to visualize and quantify the collective dynamics of a network in terms of its cellular elements. PMID:26229973

Monitoring changes in the intracellular calcium concentration and synaptic efficacy in the mollusc Aplysia.

PubMed

Ludwar, Bjoern Ch; Evans, Colin G; Cropper, Elizabeth C

2012-07-15

It has been suggested that changes in intracellular calcium mediate the induction of a number of important forms of synaptic plasticity (e.g., homosynaptic facilitation). These hypotheses can be tested by simultaneously monitoring changes in intracellular calcium and alterations in synaptic efficacy. We demonstrate how this can be accomplished by combining calcium imaging with intracellular recording techniques. Our experiments are conducted in a buccal ganglion of the mollusc Aplysia californica. This preparation has a number of experimentally advantageous features: Ganglia can be easily removed from Aplysia and experiments use adult neurons that make normal synaptic connections and have a normal ion channel distribution. Due to the low metabolic rate of the animal and the relatively low temperatures (14-16 °C) that are natural for Aplysia, preparations are stable for long periods of time. To detect changes in intracellular free calcium we will use the cell impermeant version of Calcium Orange which is easily 'loaded' into a neuron via iontophoresis. When this long wavelength fluorescent dye binds to calcium, fluorescence intensity increases. Calcium Orange has fast kinetic properties and, unlike ratiometric dyes (e.g., Fura 2), requires no filter wheel for imaging. It is fairly photo stable and less phototoxic than other dyes (e.g., fluo-3). Like all non-ratiometric dyes, Calcium Orange indicates relative changes in calcium concentration. But, because it is not possible to account for changes in dye concentration due to loading and diffusion, it can not be calibrated to provide absolute calcium concentrations. An upright, fixed stage, compound microscope was used to image neurons with a CCD camera capable of recording around 30 frames per second. In Aplysia this temporal resolution is more than adequate to detect even a single spike induced alteration in the intracellular calcium concentration. Sharp electrodes are simultaneously used to induce and record synaptic transmission in identified pre- and postsynaptic neurons. At the conclusion of each trial, a custom script combines electrophysiology and imaging data. To ensure proper synchronization we use a light pulse from a LED mounted in the camera port of the microscope. Manipulation of presynaptic calcium levels (e.g. via intracellular EGTA injection) allows us to test specific hypotheses, concerning the role of intracellular calcium in mediating various forms of plasticity.

The metabolic response to excitotoxicity - lessons from single-cell imaging.

PubMed

Connolly, Niamh M C; Prehn, Jochen H M

2015-04-01

Excitotoxicity is a pathological process implicated in neuronal death during ischaemia, traumatic brain injuries and neurodegenerative diseases. Excitotoxicity is caused by excess levels of glutamate and over-activation of NMDA or calcium-permeable AMPA receptors on neuronal membranes, leading to ionic influx, energetic stress and potential neuronal death. The metabolic response of neurons to excitotoxicity is complex and plays a key role in the ability of the neuron to adapt and recover from such an insult. Single-cell imaging is a powerful experimental technique that can be used to study the neuronal metabolic response to excitotoxicity in vitro and, increasingly, in vivo. Here, we review some of the knowledge of the neuronal metabolic response to excitotoxicity gained from in vitro single-cell imaging, including calcium and ATP dynamics and their effects on mitochondrial function, along with the contribution of glucose metabolism, oxidative stress and additional neuroprotective signalling mechanisms. Future work will combine knowledge gained from single-cell imaging with data from biochemical and computational techniques to garner holistic information about the metabolic response to excitotoxicity at the whole brain level and transfer this knowledge to a clinical setting.

The feasibility of using microwave-induced thermoacoustic tomography for detection and evaluation of renal calculi.

PubMed

Cao, Caijun; Nie, Liming; Lou, Cunguang; Xing, Da

2010-09-07

Imaging of renal calculi is important for patients who suffered a urinary calculus prior to treatment. The available imaging techniques include plain x-ray, ultrasound scan, intravenous urogram, computed tomography, etc. However, the visualization of a uric acid calculus (radiolucent calculi) is difficult and often impossible by the above imaging methods. In this paper, a new detection method based on microwave-induced thermoacoustic tomography was developed to detect the renal calculi. Thermoacoustic images of calcium oxalate and uric acid calculus were compared with their x-ray images. The microwave absorption differences among the calcium oxalate calculus, uric acid calculus and normal kidney tissue could be evaluated by the amplitude of the thermoacoustic signals. The calculi hidden in the swine kidney were clearly imaged with excellent contrast and resolution in the three orthogonal thermoacoustic images. The results indicate that thermoacoustic imaging may be developed as a complementary method for detecting renal calculi, and its low cost and effective feature shows high potential for clinical applications.

Prostaglandin E2 activates channel-mediated calcium entry in human erythrocytes: an indication for a blood clot formation supporting process.

PubMed

Kaestner, Lars; Tabellion, Wiebke; Lipp, Peter; Bernhardt, Ingolf

2004-12-01

Prostaglandin E(2) (PGE(2)) is released from platelets when they are activated. Using fluorescence imaging and the patch-clamp technique, we provide evidence that PGE(2) at physiological concentrations (10(-10) M) activates calcium rises mediated by calcium influx through a non-selective cation-channel in human red blood cells. The extent of calcium increase varied between cells with a total of 45% of the cells responding. It is well known that calcium increases elicited the calcium-activated potassium channel (Gardos channel) in the red cell membrane. Previously, it was shown that the Gardos channel activation results in potassium efflux and shrinkage of the cells. Therefore, we conclude that the PGE(2) responses of red blood cells described here reveal a direct and active participation of erythrocytes in blood clot formation.

Transparent, Flexible, Low Noise Graphene Electrodes for Simultaneous Electrophysiology and Neuroimaging

PubMed Central

Kuzum, Duygu; Takano, Hajime; Shim, Euijae; Reed, Jason C; Juul, Halvor; Richardson, Andrew G.; de Vries, Julius; Bink, Hank; Dichter, Marc A.; Lucas, Timothy H.; Coulter, Douglas A.; Cubukcu, Ertugrul; Litt, Brian

2014-01-01

Calcium imaging is a versatile experimental approach capable of resolving single neurons with single-cell spatial resolution in the brain. Electrophysiological recordings provide high temporal, but limited spatial resolution, due to the geometrical inaccessibility of the brain. An approach that integrates the advantages of both techniques could provide new insights into functions of neural circuits. Here, we report a transparent, flexible neural electrode technology based on graphene, which enables simultaneous optical imaging and electrophysiological recording. We demonstrate that hippocampal slices can be imaged through transparent graphene electrodes by both confocal and two-photon microscopy without causing any light-induced artifacts in the electrical recordings. Graphene electrodes record high frequency bursting activity and slow synaptic potentials that are hard to resolve by multi-cellular calcium imaging. This transparent electrode technology may pave the way for high spatio-temporal resolution electrooptic mapping of the dynamic neuronal activity. PMID:25327632

Particle Tracking Facilitates Real Time Capable Motion Correction in 2D or 3D Two-Photon Imaging of Neuronal Activity.

PubMed

Aghayee, Samira; Winkowski, Daniel E; Bowen, Zachary; Marshall, Erin E; Harrington, Matt J; Kanold, Patrick O; Losert, Wolfgang

2017-01-01

The application of 2-photon laser scanning microscopy (TPLSM) techniques to measure the dynamics of cellular calcium signals in populations of neurons is an extremely powerful technique for characterizing neural activity within the central nervous system. The use of TPLSM on awake and behaving subjects promises new insights into how neural circuit elements cooperatively interact to form sensory perceptions and generate behavior. A major challenge in imaging such preparations is unavoidable animal and tissue movement, which leads to shifts in the imaging location (jitter). The presence of image motion can lead to artifacts, especially since quantification of TPLSM images involves analysis of fluctuations in fluorescence intensities for each neuron, determined from small regions of interest (ROIs). Here, we validate a new motion correction approach to compensate for motion of TPLSM images in the superficial layers of auditory cortex of awake mice. We use a nominally uniform fluorescent signal as a secondary signal to complement the dynamic signals from genetically encoded calcium indicators. We tested motion correction for single plane time lapse imaging as well as multiplane (i.e., volume) time lapse imaging of cortical tissue. Our procedure of motion correction relies on locating the brightest neurons and tracking their positions over time using established techniques of particle finding and tracking. We show that our tracking based approach provides subpixel resolution without compromising speed. Unlike most established methods, our algorithm also captures deformations of the field of view and thus can compensate e.g., for rotations. Object tracking based motion correction thus offers an alternative approach for motion correction, one that is well suited for real time spike inference analysis and feedback control, and for correcting for tissue distortions.

Particle Tracking Facilitates Real Time Capable Motion Correction in 2D or 3D Two-Photon Imaging of Neuronal Activity

PubMed Central

Aghayee, Samira; Winkowski, Daniel E.; Bowen, Zachary; Marshall, Erin E.; Harrington, Matt J.; Kanold, Patrick O.; Losert, Wolfgang

2017-01-01

The application of 2-photon laser scanning microscopy (TPLSM) techniques to measure the dynamics of cellular calcium signals in populations of neurons is an extremely powerful technique for characterizing neural activity within the central nervous system. The use of TPLSM on awake and behaving subjects promises new insights into how neural circuit elements cooperatively interact to form sensory perceptions and generate behavior. A major challenge in imaging such preparations is unavoidable animal and tissue movement, which leads to shifts in the imaging location (jitter). The presence of image motion can lead to artifacts, especially since quantification of TPLSM images involves analysis of fluctuations in fluorescence intensities for each neuron, determined from small regions of interest (ROIs). Here, we validate a new motion correction approach to compensate for motion of TPLSM images in the superficial layers of auditory cortex of awake mice. We use a nominally uniform fluorescent signal as a secondary signal to complement the dynamic signals from genetically encoded calcium indicators. We tested motion correction for single plane time lapse imaging as well as multiplane (i.e., volume) time lapse imaging of cortical tissue. Our procedure of motion correction relies on locating the brightest neurons and tracking their positions over time using established techniques of particle finding and tracking. We show that our tracking based approach provides subpixel resolution without compromising speed. Unlike most established methods, our algorithm also captures deformations of the field of view and thus can compensate e.g., for rotations. Object tracking based motion correction thus offers an alternative approach for motion correction, one that is well suited for real time spike inference analysis and feedback control, and for correcting for tissue distortions. PMID:28860973

Emerging MRI Methods in Translational Cardiovascular Research

PubMed Central

Vandsburger, Moriel H; Epstein, Frederick H

2011-01-01

Cardiac magnetic resonance imaging (CMR) has become a reference standard modality for imaging of left ventricular (LV) structure and function, and, using late gadolinium enhancement, for imaging myocardial infarction. Emerging CMR techniques enable a more comprehensive examination of the heart, making CMR an excellent tool for use in translational cardiovascular research. Specifically, emerging CMR methods have been developed to measure the extent of myocardial edema, changes in ventricular mechanics, changes in tissue composition as a result of fibrosis, and changes in myocardial perfusion as a function of both disease and infarct healing. New CMR techniques also enable the tracking of labeled cells, molecular imaging of biomarkers of disease, and changes in calcium flux in cardiomyocytes. In addition, MRI can quantify blood flow velocity and wall shear stress in large blood vessels. Almost all of these techniques can be applied in both pre-clinical and clinical settings, enabling both the techniques themselves and the knowledge gained using such techniques in pre-clinical research to be translated from the lab bench to the patient bedside. PMID:21452060

Selective dopamine receptor 4 activation mediates the hippocampal neuronal calcium response via IP3 and ryanodine receptors.

PubMed

Wang, Ya-Li; Wang, Jian-Gang; Guo, Fang-Li; Gao, Xia-Huan; Zhao, Dan-Dan; Zhang, Lin; Wang, Jian-Zhi; Lu, Cheng-Biao

2017-09-01

Intracellular calcium is a key factor in most cellular processes, including cell growth, differentiation, proliferation and neurotransmitter release. Dopamine (DA) mediates synaptic transmission by regulating the intracellular calcium content. It is not clear, however, which specific subunit of the DA receptor contributes to DA modulation of intracellular calcium content changes. Through the traditional technique of Fura-2 calcium imaging, this study demonstrated that the DA can induce transient calcium in cultured hippocampal neurons and that this response can be mimicked by a selective dopamine receptor 4 (DR4) agonist PD168077 (PD). PD-induced calcium transience can be blocked by a calcium chelator, such as BAPTA-AM, or by pre-treatment of neurons with thapsigargin, a IP 3 receptor antagonist, or a micromolar concentration of ryanodine, a ryanodine receptor (RyR) antagonist. However PD-induced calcium transience cannot be blocked by pre-treatment of neurons with a free-calcium medium or a cocktail of NMDA receptor, L-type calcium channel and alpha7 nicotinic acetylcholine receptor blockers. These results indicate that the calcium response induced by DR4 activation is mainly through activation of IP 3 receptor in internal stores, which is likely to contribute to the DA modulation of synaptic transmission and cognitive function. Copyright © 2017. Published by Elsevier B.V.

Photoacoustic imaging of teeth for dentine imaging and enamel characterization

NASA Astrophysics Data System (ADS)

Periyasamy, Vijitha; Rangaraj, Mani; Pramanik, Manojit

2018-02-01

Early detection of dental caries, cracks and lesions is needed to prevent complicated root canal treatment and tooth extraction procedures. Resolution of clinically used x-ray imaging is low, hence optical imaging techniques such as optical coherence tomography, fluorescence imaging, and Raman imaging are widely experimented for imaging dental structures. Photoacoustic effect is used in photon induced photoacoustic streaming technique to debride the root canal. In this study, the extracted teeth were imaged using photoacoustic tomography system at 1064 nm. The degradation of enamel and dentine is an indicator of onset of dental caries. Photoacoustic microscopy (PAM) was used to study the tooth enamel. Images were acquired using acoustic resolution PAM system. This was done to identify microscopic cracks and dental lesion at different anatomical sites (crown and cementum). The PAM tooth profile is an indicator of calcium distribution which is essential for demineralization studies.

A method to investigate the diffusion properties of nuclear calcium.

PubMed

Queisser, Gillian; Wittum, Gabriel

2011-10-01

Modeling biophysical processes in general requires knowledge about underlying biological parameters. The quality of simulation results is strongly influenced by the accuracy of these parameters, hence the identification of parameter values that the model includes is a major part of simulating biophysical processes. In many cases, secondary data can be gathered by experimental setups, which are exploitable by mathematical inverse modeling techniques. Here we describe a method for parameter identification of diffusion properties of calcium in the nuclei of rat hippocampal neurons. The method is based on a Gauss-Newton method for solving a least-squares minimization problem and was formulated in such a way that it is ideally implementable in the simulation platform uG. Making use of independently published space- and time-dependent calcium imaging data, generated from laser-assisted calcium uncaging experiments, here we could identify the diffusion properties of nuclear calcium and were able to validate a previously published model that describes nuclear calcium dynamics as a diffusion process.

Intact calcium signaling in adrenergic-deficient embryonic mouse hearts.

PubMed

Peoples, Jessica N; Taylor, David G; Katchman, Alexander N; Ebert, Steven N

2018-01-22

Mouse embryos that lack the ability to produce the adrenergic hormones, norepinephrine (NE) and epinephrine (EPI), due to disruption of the dopamine beta-hydroxylase (Dbh -/- ) gene inevitably perish from heart failure during mid-gestation. Since adrenergic stimulation is well-known to enhance calcium signaling in developing as well as adult myocardium, and impairments in calcium signaling are typically associated with heart failure, we hypothesized that adrenergic-deficient embryonic hearts would display deficiencies in cardiac calcium signaling relative to adrenergic-competent controls at a developmental stage immediately preceding the onset of heart failure, which first appears beginning or shortly after mouse embryonic day 10.5 (E10.5). To test this hypothesis, we used ratiometric fluorescent calcium imaging techniques to measure cytosolic calcium transients, [Ca 2+ ] i in isolated E10.5 mouse hearts. Our results show that spontaneous [Ca 2+ ] i oscillations were intact and robustly responded to a variety of stimuli including extracellular calcium (5 mM), caffeine (5 mM), and NE (100 nM) in a manner that was indistinguishable from controls. Further, we show similar patterns of distribution (via immunofluorescent histochemical staining) and activity (via patch-clamp recording techniques) for the major voltage-gated plasma membrane calcium channel responsible for the L-type calcium current, I Ca,L , in adrenergic-deficient and control embryonic cardiac cells. These results demonstrate that despite the absence of vital adrenergic hormones that consistently leads to embryonic lethality in vivo, intracellular and extracellular calcium signaling remain essentially intact and functional in embryonic mouse hearts through E10.5. These findings suggest that adrenergic stimulation is not required for the development of intracellular calcium oscillations or extracellular calcium signaling through I Ca,L and that aberrant calcium signaling does not likely contribute to the onset of heart failure in this model. Copyright © 2017. Published by Elsevier Inc.

Physiological Role of Gap-Junctional Hemichannels

PubMed Central

Quist, Arjan Pieter; Rhee, Seung Keun; Lin, Hai; Lal, Ratneshwar

2000-01-01

Hemichannels in the overlapping regions of apposing cells plasma membranes join to form gap junctions and provide an intercellular communication pathway. Hemichannels are also present in the nonjunctional regions of individual cells and their activity is gated by several agents, including calcium. However, their physiological roles are unknown. Using techniques of atomic force microscopy (AFM), fluorescent dye uptake assay, and laser confocal immunofluorescence imaging, we have examined the extracellular calcium-dependent modulation of cell volume. In response to a change in the extracellular physiological calcium concentration (1.8 to ≤1.6 mM) in an otherwise isosmotic condition, real-time AFM imaging revealed a significant and reversible increase in the volume of cells expressing gap-junctional proteins (connexins). Volume change did not occur in cells that were not expressing connexins. However, after the transient or stable transfection of connexin43, volume change did occur. The volume increase was accompanied by cytochalasin D-sensitive higher cell stiffness, which helped maintain cell integrity. These cellular physical changes were prevented by gap-junctional blockers, oleamide and β-glycyrrhetinic acid, or were reversed by returning extracellular calcium to the normal level. We conclude that nongap-junctional hemichannels regulate cell volume in response to the change in extracellular physiological calcium in an otherwise isosmotic situation. PMID:10704454

Production of scandium-44 m and scandium-44 g with deuterons on calcium-44: cross section measurements and production yield calculations.

PubMed

Duchemin, C; Guertin, A; Haddad, F; Michel, N; Métivier, V

2015-09-07

HIGHLIGHTS • Production of Sc-44 m, Sc-44 g and contaminants. • Experimental values determined using the stacked-foil technique. • Thick-Target production Yield (TTY) calculations. • Comparison with the TALYS code version 1.6.Among the large number of radionuclides of medical interest, Sc-44 is promising for PET imaging. Either the ground-state Sc-44 g or the metastable-state Sc-44 m can be used for such applications, depending on the molecule used as vector. This study compares the production rates of both Sc-44 states, when protons or deuterons are used as projectiles on an enriched Calcium-44 target. This work presents the first set of data for the deuteron route. The results are compared with the TALYS code. The Thick-Target production Yields of Sc-44 m and Sc-44 g are calculated and compared with those for the proton route for three different scenarios: the production of Sc-44 g for conventional PET imaging, its production for the new 3 γ imaging technique developed at the SUBATECH laboratory and the production of a Sc-44 m/Sc-44 g in vivo generator for antibody labelling.

Vascularization of bioprosthetic valve material

NASA Astrophysics Data System (ADS)

Boughner, Derek R.; Dunmore-Buyze, Joy; Heenatigala, Dino; Lohmann, Tara; Ellis, Chris G.

1999-04-01

Cell membrane remnants represent a probable nucleation site for calcium deposition in bioprosthetic heart valves. Calcification is a primary failure mode of both bovine pericardial and porcine aortic heterograft bioprosthesis but the nonuniform pattern of calcium distribution within the tissue remains unexplained. Searching for a likely cellular source, we considered the possibility of a previously overlooked small blood vessel network. Using a videomicroscopy technique, we examined 5 matched pairs of porcine aortic and pulmonary valves and 14 samples from 6 bovine pericardia. Tissue was placed on a Leitz Metallux microscope and transilluminated with a 75 watt mercury lamp. Video images were obtained using a silicon intensified target camera equipped with a 431 nm interference filter to maximize contrast of red cells trapped in a capillary microvasculature. Video images were recorded for analysis on a Silicon Graphics Image Analysis work station equipped with a video frame grabber. For porcine valves, the technique demonstrated a vascular bed in the central spongiosa at cusp bases with vessel sizes from 6-80 micrometers . Bovine pericardium differed with a more uniform distribution of 7-100 micrometers vessels residing centrally. Thus, small blood vessel endothelial cells provide a potential explanation patterns of bioprosthetic calcification.

Studying dyadic structure-function relationships: a review of current modeling approaches and new insights into Ca2+ (mis)handling.

PubMed

Maleckar, Mary M; Edwards, Andrew G; Louch, William E; Lines, Glenn T

2017-01-01

Excitation-contraction coupling in cardiac myocytes requires calcium influx through L-type calcium channels in the sarcolemma, which gates calcium release through sarcoplasmic reticulum ryanodine receptors in a process known as calcium-induced calcium release, producing a myoplasmic calcium transient and enabling cardiomyocyte contraction. The spatio-temporal dynamics of calcium release, buffering, and reuptake into the sarcoplasmic reticulum play a central role in excitation-contraction coupling in both normal and diseased cardiac myocytes. However, further quantitative understanding of these cells' calcium machinery and the study of mechanisms that underlie both normal cardiac function and calcium-dependent etiologies in heart disease requires accurate knowledge of cardiac ultrastructure, protein distribution and subcellular function. As current imaging techniques are limited in spatial resolution, limiting insight into changes in calcium handling, computational models of excitation-contraction coupling have been increasingly employed to probe these structure-function relationships. This review will focus on the development of structural models of cardiac calcium dynamics at the subcellular level, orienting the reader broadly towards the development of models of subcellular calcium handling in cardiomyocytes. Specific focus will be given to progress in recent years in terms of multi-scale modeling employing resolved spatial models of subcellular calcium machinery. A review of the state-of-the-art will be followed by a review of emergent insights into calcium-dependent etiologies in heart disease and, finally, we will offer a perspective on future directions for related computational modeling and simulation efforts.

Two-photon imaging of neuronal activity in motor cortex of marmosets during upper-limb movement tasks.

PubMed

Ebina, Teppei; Masamizu, Yoshito; Tanaka, Yasuhiro R; Watakabe, Akiya; Hirakawa, Reiko; Hirayama, Yuka; Hira, Riichiro; Terada, Shin-Ichiro; Koketsu, Daisuke; Hikosaka, Kazuo; Mizukami, Hiroaki; Nambu, Atsushi; Sasaki, Erika; Yamamori, Tetsuo; Matsuzaki, Masanori

2018-05-14

Two-photon imaging in behaving animals has revealed neuronal activities related to behavioral and cognitive function at single-cell resolution. However, marmosets have posed a challenge due to limited success in training on motor tasks. Here we report the development of protocols to train head-fixed common marmosets to perform upper-limb movement tasks and simultaneously perform two-photon imaging. After 2-5 months of training sessions, head-fixed marmosets can control a manipulandum to move a cursor to a target on a screen. We conduct two-photon calcium imaging of layer 2/3 neurons in the motor cortex during this motor task performance, and detect task-relevant activity from multiple neurons at cellular and subcellular resolutions. In a two-target reaching task, some neurons show direction-selective activity over the training days. In a short-term force-field adaptation task, some neurons change their activity when the force field is on. Two-photon calcium imaging in behaving marmosets may become a fundamental technique for determining the spatial organization of the cortical dynamics underlying action and cognition.

Calcium Chloride in Neonatal Parenteral Nutrition Solutions with and without Added Cysteine: Compatibility Studies Using Laser and Micro-Flow Imaging Methodology.

PubMed

Huston, Robert K; Christensen, J Mark; Alshahrani, Sultan M; Mohamed, Sumeia M; Clark, Sara M; Nason, Jeffrey A; Wu, Ying Xing

2015-01-01

Previous studies of compatibility of calcium chloride (CaCl2) and phosphates have not included particle counts in the range specified by the United States Pharmacopeia. Micro-flow imaging techniques have been shown to be comparable to light obscuration when determining particle count and size in pharmaceutical solutions. The purpose of this study was to do compatibility testing for parenteral nutrition (PN) solutions containing CaCl2 using dynamic light scattering and micro-flow imaging techniques. Solutions containing TrophAmine (Braun Medical Inc, Irvine, CA), CaCl2, and sodium phosphate (NaPhos) were compounded with and without cysteine. All solutions contained standard additives to neonatal PN solutions including dextrose, trace metals, and electrolytes. Control solutions contained no calcium or phosphate. Solutions were analyzed for particle size and particle count. Means of Z-average particle size and particle counts of controls were determined. Study solutions were compared to controls and United States Pharmacopeia (USP) Chapter 788 guidelines. The maximum amount of Phos that was compatible in solutions that contained at least 10 mmol/L of Ca in 2.5% amino acids (AA) was determined. Compatibility of these solutions was verified by performing analyses of 5 repeats of these solutions. Microscopic analyses of the repeats were also performed. Amounts of CaCl2 and NaPhos that were compatible in solutions containing 1.5%, 2%, 2.5%, and 3% AA were determined. The maximum amount of NaPhos that could be added to TrophAmine solutions of > = 2.5% AA containing at least 10 mmol/L of CaCl2 was 7.5 mmol/L. Adding 50 mg/dL of cysteine increased the amount of NaPhos that could be added to solutions containing 10 mmol/L of CaCl2 to 10 mmol/L. Calcium chloride can be added to neonatal PN solutions containing NaPhos in concentrations that can potentially provide an intravenous intake of adequate amounts of calcium and phosphorus.

Measuring coronary calcium on CT images adjusted for attenuation differences.

PubMed

Nelson, Jennifer Clark; Kronmal, Richard A; Carr, J Jeffrey; McNitt-Gray, Michael F; Wong, Nathan D; Loria, Catherine M; Goldin, Jonathan G; Williams, O Dale; Detrano, Robert

2005-05-01

To quantify scanner and participant variability in attenuation values for computed tomographic (CT) images assessed for coronary calcium and define a method for standardizing attenuation values and calibrating calcium measurements. Institutional review board approval and participant informed consent were obtained at all study sites. An image attenuation adjustment method involving the use of available calibration phantom data to define standard attenuation values was developed. The method was applied to images from two population-based multicenter studies: the Coronary Artery Risk Development in Young Adults study (3041 participants) and the Multi-Ethnic Study of Atherosclerosis (6814 participants). To quantify the variability in attenuation, analysis of variance techniques were used to compare the CT numbers of standardized torso phantom regions across study sites, and multivariate linear regression models of participant-specific calibration phantom attenuation values that included participant age, race, sex, body mass index (BMI), smoking status, and site as covariates were developed. To assess the effect of the calibration method on calcium measurements, Pearson correlation coefficients between unadjusted and attenuation-adjusted calcium measurements were computed. Multivariate models were used to examine the effect of sex, race, BMI, smoking status, unadjusted score, and site on Agatston score adjustments. Mean attenuation values (CT numbers) of a standard calibration phantom scanned beneath participants varied significantly according to scanner and participant BMI (P < .001 for both). Values were lowest for Siemens multi-detector row CT scanners (110.0 HU), followed by GE-Imatron electron-beam (116.0 HU) and GE LightSpeed multi-detector row scanners (121.5 HU). Values were also lower for morbidly obese (BMI, > or =40.0 kg/m(2)) participants (108.9 HU), followed by obese (BMI, 30.0-39.9 kg/m(2)) (114.8 HU), overweight (BMI, 25.0-29.9 kg/m(2)) (118.5 HU), and normal-weight or underweight (BMI, <25.0 kg/m(2)) (120.1 HU) participants. Agatston score calibration adjustments ranged from -650 to 1071 (mean, -8 +/- 50 [standard deviation]) and increased with Agatston score (P < .001). The direction and magnitude of adjustment varied significantly according to scanner and BMI (P < .001 for both) and were consistent with phantom attenuation results in that calibration resulted in score decreases for images with higher phantom attenuation values. Image attenuation values vary by scanner and participant body size, producing calcium score differences that are not due to true calcium burden disparities. Use of calibration phantoms to adjust attenuation values and calibrate calcium measurements in research studies and clinical practice may improve the comparability of such measurements between persons scanned with different scanners and within persons over time.

CALCIUM BINDING TO INTESTINAL MEMBRANES

PubMed Central

Oschman, James L.; Wall, Betty J.

1972-01-01

Flame photometry reveals that glutaraldehyde and buffer solutions in routine use for electron microscopy contain varying amounts of calcium. The presence of electron-opaque deposits adjacent to membranes in a variety of tissues can be correlated with the presence of calcium in the fixative. In insect intestine (midgut), deposits occur adjacent to apical and lateral plasma membranes. The deposits are particularly evident in tissues fixed in glutaraldehyde without postosmication. They are also observed in osmicated tissue if calcium is added to wash and osmium solutions. Deposits are absent when calcium-free fixatives are used, but are present when traces of CaCl2 (as low as 5 x 10-5 M) are added. The deposits occur at regular intervals along junctional membranes, providing images strikingly similar to those obtained by other workers who have used pyroantimonate in an effort to localize sodium. Other divalent cations (Mg++, Sr++, Ba++, Mn++, Fe++) appear to substitute for calcium, while sodium, potassium, lanthanum, and mercury do not. After postfixing with osmium with calcium added, the deposits can be resolved as patches along the inner leaflet of apical and lateral plasma membranes. The dense regions may thus localize membrane constituents that bind calcium. The results are discussed in relation to the role of calcium in control of cell-to-cell communication, intestinal calcium uptake, and the pyroantimonate technique for ion localization. PMID:4569411

Evaluation of the 3D spatial distribution of the Calcium/Phosphorus ratio in bone using computed-tomography dual-energy analysis.

PubMed

Hadjipanteli, A; Kourkoumelis, N; Fromme, P; Huang, J; Speller, R D

2016-01-01

The Calcium/Phosphorus (Ca/P) ratio was shown to vary between healthy bones and bones with osteoporotic symptoms. The relation of the Ca/P ratio to bone quality remains under investigation. To study this relation and determine if the ratio can be used to predict bone fractures, a non-invasive 3D imaging technique is required. The first aim of this study was to test the effectiveness of a computed-tomography dual-energy analysis (CT-DEA) technique developed to assess the Ca/P ratio in bone apatite (collagen-free bone) in identifying differences between healthy and inflammation-mediated osteoporotic (IMO) bones. The second aim was to extend the above technique for its application to a more complex structure, intact bone, that could potentially lead to clinical use. For the first aim, healthy and IMO rabbit cortical bone apatite samples were assessed. For the second aim, some changes were made to the technique, which was applied to healthy and IMO intact bone samples. Statistically significant differences between healthy and IMO bone apatite were found for the bulk Ca/P ratio, low Ca/P ratio proportion and interconnected low Ca/P ratio proportion. For the intact bone samples, the bulk Ca/P ratio was found to be significantly different between healthy and IMO. Results show that the CT-DEA technique can be used to identify differences in the Ca/P ratio between healthy and osteoporotic, in both bone apatite and intact bone. With quantitative imaging becoming an increasingly important advancement in medical imaging, CT-DEA for bone decomposition could potentially have several applications. Copyright © 2015. Published by Elsevier Ltd.

Use of Genetically-encoded Calcium Indicators for Live Cell Calcium Imaging and Localization in Virus-infected Cells

PubMed Central

Perry, Jacob L.; Ramachandran, Nina K.; Utama, Budi; Hyser, Joseph M.

2015-01-01

Calcium signaling is a ubiquitous and versatile process involved in nearly every cellular process, and exploitation of host calcium signals is a common strategy used by viruses to facilitate replication and cause disease. Small molecule fluorescent calcium dyes have been used by many to examine changes in host cell calcium signaling and calcium channel activation during virus infections, but disadvantages of these dyes, including poor loading and poor long-term retention, complicate analysis of calcium imaging in virus-infected cells due to changes in cell physiology and membrane integrity. The recent expansion of genetically-encoded calcium indicators (GECIs), including blue and red-shifted color variants and variants with calcium affinities appropriate for calcium storage organelles like the endoplasmic reticulum (ER), make the use of GECIs an attractive alternative for calcium imaging in the context of virus infections. Here we describe the development and testing of cell lines stably expressing both green cytoplasmic (GCaMP5G and GCaMP6s) and red ER-targeted (RCEPIAer) GECIs. Using three viruses (rotavirus, poliovirus and respiratory syncytial virus) previously shown to disrupt host calcium homeostasis, we show the GECI cell lines can be used to detect simultaneous cytoplasmic and ER calcium signals. Further, we demonstrate the GECI expression has sufficient stability to enable long-term confocal imaging of both cytoplasmic and ER calcium during the course of virus infections. PMID:26344758

Calcium (Ca2+) waves data calibration and analysis using image processing techniques

PubMed Central

2013-01-01

Background Calcium (Ca2+) propagates within tissues serving as an important information carrier. In particular, cilia beat frequency in oviduct cells is partially regulated by Ca2+ changes. Thus, measuring the calcium density and characterizing the traveling wave plays a key role in understanding biological phenomena. However, current methods to measure propagation velocities and other wave characteristics involve several manual or time-consuming procedures. This limits the amount of information that can be extracted, and the statistical quality of the analysis. Results Our work provides a framework based on image processing procedures that enables a fast, automatic and robust characterization of data from two-filter fluorescence Ca2+ experiments. We calculate the mean velocity of the wave-front, and use theoretical models to extract meaningful parameters like wave amplitude, decay rate and time of excitation. Conclusions Measurements done by different operators showed a high degree of reproducibility. This framework is also extended to a single filter fluorescence experiments, allowing higher sampling rates, and thus an increased accuracy in velocity measurements. PMID:23679062

Breast Microcalcification Detection Using Super-Resolution Ultrasound Image Reconstruction

DTIC Science & Technology

2010-09-01

microcalcifications often occur as one of two types: calcium oxalate dihydrate or calcium hydroxyapatite. Their sizes range approximately from 0.1 mm to 0.5 mm...super-resolution imaging, ultrasound imaging, wave equation. 1. INTRODUCTION Microcalcifications, tiny specks of mineral deposits ( calcium ), are the

Characterization of Biogenic Gas and Mineral Formation Process by Denitrification in Porous Media

NASA Astrophysics Data System (ADS)

Hall, C. A.; Kim, D.; Mahabadi, N.; van Paassen, L. A.

2017-12-01

Biologically mediated processes have been regarded and developed as an alternative approach to traditional ground improvement techniques. Denitrification has been investigated as a potential ground improvement process towards liquefaction hazard mitigation. During denitrification, microorganisms reduce nitrate to dinitrogen gas and facilitate calcium carbonate precipitation as a by-product under adequate environmental conditions. The formation of dinitrogen gas desaturates soils and allows for potential pore pressure dampening during earthquake events. While, precipitation of calcium carbonate can improve the mechanical properties by filling the voids and cementing soil particles. As a result of small changes in gas and mineral phases, the mechanical properties of soils can be significantly affected. Prior research has primarily focused on quantitative analysis of overall residual calcium carbonate mineral and biogenic gas products in lab-scale porous media. However, the distribution of these products at the pore-scale has not been well-investigated. In this research, denitrification is activated in a microfluidic chip simulating a homogenous pore structure. The denitrification process is monitored by sequential image capture, where gas and mineral phase changes are evaluated by image processing. Analysis of these images correspond with previous findings, which demonstrate that biogenic gas behaviour at the pore scale is affected by the balance between reaction, diffusion, and convection rates.

Calcium-based biomaterials for diagnosis, treatment, and theranostics.

PubMed

Qi, Chao; Lin, Jing; Fu, Lian-Hua; Huang, Peng

2018-01-22

Calcium-based (CaXs) biomaterials including calcium phosphates, calcium carbonates, calcium silicate and calcium fluoride have been widely utilized in the biomedical field owing to their excellent biocompatibility and biodegradability. In recent years, CaXs biomaterials have been strategically integrated with imaging contrast agents and therapeutic agents for various molecular imaging modalities including fluorescence imaging, magnetic resonance imaging, ultrasound imaging or multimodal imaging, as well as for various therapeutic approaches including chemotherapy, gene therapy, hyperthermia therapy, photodynamic therapy, radiation therapy, or combination therapy, even imaging-guided therapy. Compared with other inorganic biomaterials such as silica-, carbon-, and gold-based biomaterials, CaXs biomaterials can dissolve into nontoxic ions and participate in the normal metabolism of organisms. Thus, they offer safer clinical solutions for disease theranostics. This review focuses on the state-of-the-art progress in CaXs biomaterials, which covers from their categories, characteristics and preparation methods to their bioapplications including diagnosis, treatment, and theranostics. Moreover, the current trends and key problems as well as the future prospects and challenges of CaXs biomaterials are also discussed at the end.

Functional neuroanatomy of the rhinophore of Archidoris pseudoargus

NASA Astrophysics Data System (ADS)

Wertz, Adrian; Rössler, Wolfgang; Obermayer, Malu; Bickmeyer, Ulf

2007-06-01

For sea slugs, chemosensory information represents an important sensory modality, because optical and acoustical information are limited. In the present study, we focussed on the neuroanatomy of the rhinophores and processing of olfactory stimuli in the rhinophore ganglion of Archidoris pseudoargus, belonging to the order of Nudibranchia in the subclass of Opisthobranchia. Histological techniques, fluorescent markers, and immunohistochemistry were used to analyse neuroanatomical features of the rhinophore. A large ganglion and a prominent central lymphatic channel are surrounded by longitudinal muscles. Many serotonin-immunoreactive (IR) processes were found around the centre and between the ganglion and the highly folded lobes of the rhinophore, but serotonin-IR cell bodies were absent inside the rhinophore. In contrast to the conditions recently found in Aplysia punctata, we found no evidence for the presence of olfactory glomeruli within the rhinophore. Using calcium-imaging techniques with Fura II as a calcium indicator, we found differential calcium responses in various regions within the ganglion to stimulation of the rhinophore with different amino acids. The lack of glomeruli in the rhinophores induces functional questions about processing of chemical information in the rhinophore.

Dual-Energy Computed Tomography in Cardiothoracic Vascular Imaging.

PubMed

De Santis, Domenico; Eid, Marwen; De Cecco, Carlo N; Jacobs, Brian E; Albrecht, Moritz H; Varga-Szemes, Akos; Tesche, Christian; Caruso, Damiano; Laghi, Andrea; Schoepf, Uwe Joseph

2018-07-01

Dual energy computed tomography is becoming increasingly widespread in clinical practice. It can expand on the traditional density-based data achievable with single energy computed tomography by adding novel applications to help reach a more accurate diagnosis. The implementation of this technology in cardiothoracic vascular imaging allows for improved image contrast, metal artifact reduction, generation of virtual unenhanced images, virtual calcium subtraction techniques, cardiac and pulmonary perfusion evaluation, and plaque characterization. The improved diagnostic performance afforded by dual energy computed tomography is not associated with an increased radiation dose. This review provides an overview of dual energy computed tomography cardiothoracic vascular applications. Copyright © 2018 Elsevier Inc. All rights reserved.

Single-cell resolution fluorescence imaging of circadian rhythms detected with a Nipkow spinning disk confocal system.

PubMed

Enoki, Ryosuke; Ono, Daisuke; Hasan, Mazahir T; Honma, Sato; Honma, Ken-Ichi

2012-05-30

Single-point laser scanning confocal imaging produces signals with high spatial resolution in living organisms. However, photo-induced toxicity, bleaching, and focus drift remain challenges, especially when recording over several days for monitoring circadian rhythms. Bioluminescence imaging is a tool widely used for this purpose, and does not cause photo-induced difficulties. However, bioluminescence signals are dimmer than fluorescence signals, and are potentially affected by levels of cofactors, including ATP, O(2), and the substrate, luciferin. Here we describe a novel time-lapse confocal imaging technique to monitor circadian rhythms in living tissues. The imaging system comprises a multipoint scanning Nipkow spinning disk confocal unit and a high-sensitivity EM-CCD camera mounted on an inverted microscope with auto-focusing function. Brain slices of the suprachiasmatic nucleus (SCN), the central circadian clock, were prepared from transgenic mice expressing a clock gene, Period 1 (Per1), and fluorescence reporter protein (Per1::d2EGFP). The SCN slices were cut out together with membrane, flipped over, and transferred to the collagen-coated glass dishes to obtain signals with a high signal-to-noise ratio and to minimize focus drift. The imaging technique and improved culture method enabled us to monitor the circadian rhythm of Per1::d2EGFP from optically confirmed single SCN neurons without noticeable photo-induced effects or focus drift. Using recombinant adeno-associated virus carrying a genetically encoded calcium indicator, we also monitored calcium circadian rhythms at a single-cell level in a large population of SCN neurons. Thus, the Nipkow spinning disk confocal imaging system developed here facilitates long-term visualization of circadian rhythms in living cells. Copyright © 2012 Elsevier B.V. All rights reserved.

The role of synaptotagmin I C2A calcium-binding domain in synaptic vesicle clustering during synapse formation

PubMed Central

Gardzinski, Peter; Lee, David W K; Fei, Guang-He; Hui, Kwokyin; Huang, Guan J; Sun, Hong-Shuo; Feng, Zhong-Ping

2007-01-01

Synaptic vesicles aggregate at the presynaptic terminal during synapse formation via mechanisms that are poorly understood. Here we have investigated the role of the putative calcium sensor synaptotagmin I in vesicle aggregation during the formation of soma–soma synapses between identified partner cells using a simple in vitro synapse model in the mollusc Lymnaea stagnalis. Immunocytochemistry, optical imaging and electrophysiological recording techniques were used to monitor synapse formation and vesicle localization. Within 6 h, contact between appropriate synaptic partner cells up-regulated global synaptotagmin I expression, and induced a localized aggregation of synaptotagmin I at the contact site. Cell contacts between non-synaptic partner cells did not affect synaptotagmin I expression. Application of an human immunodeficiency virus type-1 transactivator (HIV-1 TAT)-tagged peptide corresponding to loop 3 of the synaptotagmin I C2A domain prevented synaptic vesicle aggregation and synapse formation. By contrast, a TAT-tagged peptide containing the calcium-binding motif of the C2B domain did not affect synaptic vesicle aggregation or synapse formation. Calcium imaging with Fura-2 demonstrated that TAT–C2 peptides did not alter either basal or evoked intracellular calcium levels. These results demonstrate that contact with an appropriate target cell is necessary to initiate synaptic vesicle aggregation during nascent synapse formation and that the initial aggregation of synaptic vesicles is dependent on loop 3 of the C2A domain of synaptotagmin I. PMID:17317745

An integrative approach for analyzing hundreds of neurons in task performing mice using wide-field calcium imaging.

PubMed

Mohammed, Ali I; Gritton, Howard J; Tseng, Hua-an; Bucklin, Mark E; Yao, Zhaojie; Han, Xue

2016-02-08

Advances in neurotechnology have been integral to the investigation of neural circuit function in systems neuroscience. Recent improvements in high performance fluorescent sensors and scientific CMOS cameras enables optical imaging of neural networks at a much larger scale. While exciting technical advances demonstrate the potential of this technique, further improvement in data acquisition and analysis, especially those that allow effective processing of increasingly larger datasets, would greatly promote the application of optical imaging in systems neuroscience. Here we demonstrate the ability of wide-field imaging to capture the concurrent dynamic activity from hundreds to thousands of neurons over millimeters of brain tissue in behaving mice. This system allows the visualization of morphological details at a higher spatial resolution than has been previously achieved using similar functional imaging modalities. To analyze the expansive data sets, we developed software to facilitate rapid downstream data processing. Using this system, we show that a large fraction of anatomically distinct hippocampal neurons respond to discrete environmental stimuli associated with classical conditioning, and that the observed temporal dynamics of transient calcium signals are sufficient for exploring certain spatiotemporal features of large neural networks.

Fluorescence lifetime imaging of calcium flux in neurons in response to pulsed infrared light

NASA Astrophysics Data System (ADS)

Walsh, Alex J.; Sedelnikova, Anna; Tolstykh, Gleb P.; Ibey, Bennett L.; Beier, Hope T.

2017-02-01

Pulsed infrared light can excite action potentials in neurons; yet, the fundamental mechanism underlying this phenomenon is unknown. Previous work has observed a rise in intracellular calcium concentration following infrared exposure, but the source of the calcium and mechanism of release is unknown. Here, we used fluorescence lifetime imaging of Oregon Green BAPTA-1 to study intracellular calcium dynamics in primary rat hippocampal neurons in response to infrared light exposure. The fluorescence lifetime of Oregon Green BAPTA-1 is longer when bound to calcium, and allows robust measurement of intracellular free calcium concentrations. First, a fluorescence lifetime calcium calibration curve for Oregon Green BAPTA-1 was determined in solutions. The normalized amplitude of the short and long lifetimes was calibrated to calcium concentration. Then, neurons were incubated in Oregon Green BAPTA-1 and exposed to pulses of infrared light (0-1 J/cm2; 0-5 ms; 1869 nm). Fluorescence lifetime images were acquired prior to, during, and after the infrared exposure. Fluorescence lifetime images, 64x64 pixels, were acquired at 12 or 24 ms for frame rates of 83 and 42 Hz, respectively. Accurate α1 approximations were achieved in images with low photon counts by computing an α1 index value from the relative probability of the observed decay events. Results show infrared light exposure increases intracellular calcium in neurons. Altogether, this study demonstrates accurate fluorescence lifetime component analysis from low-photon count data for improved imaging speed.

Studying the mechanism of neurostimulation by infrared laser light using GCaMP6s and Rhodamine B imaging

NASA Astrophysics Data System (ADS)

Moreau, David; Lefort, Claire; Bardet, Sylvia M.; O'Connor, Rodney P.

2016-03-01

Infrared laser light radiation can be used to depolarize neurons and to stimulate neural activity. The absorption of infrared radiation and heating of biological tissue is thought to be the underlying mechanism of this phenomenon whereby local temperature increases in the plasma membrane of cells either directly influence membrane properties or act via temperature sensitive ion channels. Action potentials are typically measured electrically in neurons with microelectrodes, but they can also be observed using fluorescence microscopy techniques that use synthetic or genetically encoded calcium indicators. In this work, we studied the impact of infrared laser light on neuronal calcium signals to address the mechanism of these thermal effects. Cultured primary mouse hippocampal neurons expressing the genetically encoded calcium indicator GCaMP6s were used in combination with the temperature sensitive fluorophore Rhodamine B to measure calcium signals and temperature changes at the cellular level. Here we present our all-optical strategy for studying the influence of infrared laser light on neuronal activity.

Ballistic delivery of dyes for structural and functional studies of the nervous system

PubMed Central

Gan, Wen-Biao; Grutzendler, Jaime; Wong, Rachel O.; Lichtman, Jeff W.

2010-01-01

This chapter describes a detail protocol for rapid labeling of cells in a variety of preparations by means of particle-mediated ballistic (gene gun) delivery of fluorescent dyes. This method has been used for rapid labeling of cells with either lipid or water-soluble dyes in a variety of preparations. In particular, carbocyanine lipophilic dyes such as DiI have been used to obtain Golgi-like labeling of neurons and glia in fixed and live cell cultures, brain slices, as well as fixed post-mortem human brain. Water-soluble calcium indicators such as calcium green-1 dextran have been used to image calcium dynamics in living brain slices and retinal explants. This ballistic labeling technique is thus useful for studying the structure and function of neurons and glia in both living and fixed specimens. PMID:20147144

Detection and quantification of coronary calcium from dual energy chest x-rays: Phantom feasibility study.

PubMed

Zhou, Bo; Wen, Di; Nye, Katelyn; Gilkeson, Robert C; Eck, Brendan; Jordan, David; Wilson, David L

2017-10-01

We have demonstrated the ability to identify coronary calcium, a reliable biomarker of coronary artery disease, using nongated, 2-shot, dual energy (DE) chest x-ray imaging. Here we will use digital simulations, backed up by measurements, to characterize DE calcium signals and the role of potential confounds such as beam hardening, x-ray scatter, cardiac motion, and pulmonary artery pulsation. For the DE calcium signal, we will consider quantification, as compared to CT calcium score, and visualization. We created stylized and anatomical digital 3D phantoms including heart, lung, coronary calcium, spine, ribs, pulmonary artery, and adipose. We simulated high and low kVp x-ray acquisitions with x-ray spectra, energy dependent attenuation, scatter, ideal detector, and automatic exposure control (AEC). Phantoms allowed us to vary adipose thickness, cardiac motion, etc. We used specialized dual energy coronary calcium (DECC) processing that includes corrections for scatter and beam hardening. Beam hardening over a wide range of adipose thickness (0-30 cm) reduced the change in intensity of a coronary artery calcification (ΔI CAC ) by < 3% in DECC images. Scatter correction errors of ±50% affected the calcium signal (ΔI CAC ) in DECC images ±9%. If a simulated pulmonary artery fills with blood between exposures, it can give rise to a residual signal in DECC images, explaining pulmonary artery visibility in some clinical images. Residual misregistration can be mostly compensated by integrating signals in an enlarged region encompassing registration artifacts. DECC calcium score compared favorably to CT mass and volume scores over a number of phantom perturbations. Simulations indicate that proper DECC processing can faithfully recover coronary calcium signals. Beam hardening, errors in scatter estimation, cardiac motion, calcium residual misregistration etc., are all manageable. Simulations are valuable as we continue to optimize DE coronary calcium image processing and quantitative analysis. © 2017 American Association of Physicists in Medicine.

Fast Calcium Imaging with Optical Sectioning via HiLo Microscopy.

PubMed

Lauterbach, Marcel A; Ronzitti, Emiliano; Sternberg, Jenna R; Wyart, Claire; Emiliani, Valentina

2015-01-01

Imaging intracellular calcium concentration via reporters that change their fluorescence properties upon binding of calcium, referred to as calcium imaging, has revolutionized our way to probe neuronal activity non-invasively. To reach neurons densely located deep in the tissue, optical sectioning at high rate of acquisition is necessary but difficult to achieve in a cost effective manner. Here we implement an accessible solution relying on HiLo microscopy to provide robust optical sectioning with a high frame rate in vivo. We show that large calcium signals can be recorded from dense neuronal populations at high acquisition rates. We quantify the optical sectioning capabilities and demonstrate the benefits of HiLo microscopy compared to wide-field microscopy for calcium imaging and 3D reconstruction. We apply HiLo microscopy to functional calcium imaging at 100 frames per second deep in biological tissues. This approach enables us to discriminate neuronal activity of motor neurons from different depths in the spinal cord of zebrafish embryos. We observe distinct time courses of calcium signals in somata and axons. We show that our method enables to remove large fluctuations of the background fluorescence. All together our setup can be implemented to provide efficient optical sectioning in vivo at low cost on a wide range of existing microscopes.

Fast Calcium Imaging with Optical Sectioning via HiLo Microscopy

PubMed Central

Sternberg, Jenna R.; Wyart, Claire; Emiliani, Valentina

2015-01-01

Imaging intracellular calcium concentration via reporters that change their fluorescence properties upon binding of calcium, referred to as calcium imaging, has revolutionized our way to probe neuronal activity non-invasively. To reach neurons densely located deep in the tissue, optical sectioning at high rate of acquisition is necessary but difficult to achieve in a cost effective manner. Here we implement an accessible solution relying on HiLo microscopy to provide robust optical sectioning with a high frame rate in vivo. We show that large calcium signals can be recorded from dense neuronal populations at high acquisition rates. We quantify the optical sectioning capabilities and demonstrate the benefits of HiLo microscopy compared to wide-field microscopy for calcium imaging and 3D reconstruction. We apply HiLo microscopy to functional calcium imaging at 100 frames per second deep in biological tissues. This approach enables us to discriminate neuronal activity of motor neurons from different depths in the spinal cord of zebrafish embryos. We observe distinct time courses of calcium signals in somata and axons. We show that our method enables to remove large fluctuations of the background fluorescence. All together our setup can be implemented to provide efficient optical sectioning in vivo at low cost on a wide range of existing microscopes. PMID:26625116

Mineralogy and chemistry of urinary stones: patients from North Jordan.

PubMed

Abboud, Iyad Ahmed

2008-10-01

Urinary stone diseases are increasing in the Middle East. The majority of urinary stone cases are found in the northern part of the country. Stone samples taken from patients living in the Irbid area were collected from Princess Basma Hospital. The present study concentrates on the mineralogical and chemical composition of the urinary stones and on the effective environmental factors that assist in developing the different types of urinary stones. Using X-ray diffraction techniques, the mineralogical composition of the urinary stones was found to be as follows: oxalate, cholesten, and uric acid, with cystine stones occuring more frequently than the others. Cholesten and calcium oxalate stones are the most dominant types of stones. Calcium oxalate is the most common type of oxalate stone. Calcium oxalate is represented in: whewellite, wheddellite, and calcium carbonate oxalate hydrate minerals, in addition to other minerals such as brushite, ammonium phosphate, vaterite, valleriite, and bobierrite from other types of stones. Bobierrite (phosphate group) is a new mineral reported in urinary stones, and this has not been determined in any previous study worldwide. Apatite (calcium phosphate) is deduced using scanning electron microscope (SEM) images. The SEM technique determined crystal forms and systems, shapes, morphological features, and the names of the minerals forming urine stones, while optical properties are studied by polarizing microscope. X-ray fluorescence technique determined the concentrations of major and some trace elements. It revealed that Ca is the main constituent of the urinary stones, especially those composed of calcium oxalate and calcium phosphate. The concentration of trace elements was Ba = 1.57, P = 3.61, Fe = 1.78, S = 2.08, Zr = 4.63, Mo = 3.92, Cu = 1.89, Co = 1.56, and F = 4.2% and was higher in the urinary stones of Jordanian patients than in foreigners in the country. Questionnaires completed by patients suggest that the most significant factors directly effecting the formation of stones are water, climate conditions, food rich in protein and rich in different chemicals. Moreover, some drugs and diseases might also help in developing other stones.

Beta-Estradiol Regulates Voltage-Gated Calcium Channels and Estrogen Receptors in Telocytes from Human Myometrium.

PubMed

Banciu, Adela; Banciu, Daniel Dumitru; Mustaciosu, Cosmin Catalin; Radu, Mihai; Cretoiu, Dragos; Xiao, Junjie; Cretoiu, Sanda Maria; Suciu, Nicolae; Radu, Beatrice Mihaela

2018-05-09

Voltage-gated calcium channels and estrogen receptors are essential players in uterine physiology, and their association with different calcium signaling pathways contributes to healthy and pathological conditions of the uterine myometrium. Among the properties of the various cell subtypes present in human uterine myometrium, there is increasing evidence that calcium oscillations in telocytes (TCs) contribute to contractile activity and pregnancy. Our study aimed to evaluate the effects of beta-estradiol on voltage-gated calcium channels and estrogen receptors in TCs from human uterine myometrium and to understand their role in pregnancy. For this purpose, we employed patch-clamp recordings, ratiometric Fura-2-based calcium imaging analysis, and qRT-PCR techniques for the analysis of cultured human myometrial TCs derived from pregnant and non-pregnant uterine samples. In human myometrial TCs from both non-pregnant and pregnant uterus, we evidenced by qRT-PCR the presence of genes encoding for voltage-gated calcium channels (Cav3.1, Ca3.2, Cav3.3, Cav2.1), estrogen receptors (ESR1, ESR2, GPR30), and nuclear receptor coactivator 3 (NCOA3). Pregnancy significantly upregulated Cav3.1 and downregulated Cav3.2, Cav3.3, ESR1, ESR2, and NCOA3, compared to the non-pregnant condition. Beta-estradiol treatment (24 h, 10, 100, 1000 nM) downregulated Cav3.2, Cav3.3, Cav1.2, ESR1, ESR2, GRP30, and NCOA3 in TCs from human pregnant uterine myometrium. We also confirmed the functional expression of voltage-gated calcium channels by patch-clamp recordings and calcium imaging analysis of TCs from pregnant human myometrium by perfusing with BAY K8644, which induced calcium influx through these channels. Additionally, we demonstrated that beta-estradiol (1000 nM) antagonized the effect of BAY K8644 (2.5 or 5 µM) in the same preparations. In conclusion, we evidenced the presence of voltage-gated calcium channels and estrogen receptors in TCs from non-pregnant and pregnant human uterine myometrium and their gene expression regulation by beta-estradiol in pregnant conditions. Further exploration of the calcium signaling in TCs and its modulation by estrogen hormones will contribute to the understanding of labor and pregnancy mechanisms and to the development of effective strategies to reduce the risk of premature birth.

Multiparameter imaging of calcium and abscisic acid and high-resolution quantitative calcium measurements using R-GECO1-mTurquoise in Arabidopsis.

PubMed

Waadt, Rainer; Krebs, Melanie; Kudla, Jörg; Schumacher, Karin

2017-10-01

Calcium signals occur in specific spatio-temporal patterns in response to various stimuli and are coordinated with, for example, hormonal signals, for physiological and developmental adaptations. Quantification of calcium together with other signalling molecules is required for correlative analyses and to decipher downstream calcium-decoding mechanisms. Simultaneous in vivo imaging of calcium and abscisic acid has been performed here to investigate the interdependence of the respective signalling processes in Arabidopsis thaliana roots. Advanced ratiometric genetically encoded calcium indicators have been generated and in vivo calcium calibration protocols were established to determine absolute calcium concentration changes in response to auxin and ATP. In roots, abscisic acid induced long-term basal calcium concentration increases, while auxin triggered rapid signals in the elongation zone. The advanced ratiometric calcium indicator R-GECO1-mTurquoise exhibited an increased calcium signal resolution compared to commonly used Förster resonance energy transfer-based indicators. Quantitative calcium measurements in Arabidopsis root tips using R-GECO1-mTurquoise revealed detailed maps of absolute calcium concentration changes in response to auxin and ATP. Calcium calibration protocols using R-GECO1-mTurquoise enabled high-resolution quantitative imaging of resting cytosolic calcium concentrations and their dynamic changes that revealed distinct hormonal and ATP responses in roots. © 2017 The Authors. New Phytologist © 2017 New Phytologist Trust.

Rapid communication between neurons and astrocytes in primary cortical cultures.

PubMed

Murphy, T H; Blatter, L A; Wier, W G; Baraban, J M

1993-06-01

The identification of neurotransmitter receptors and voltage-sensitive ion channels on astrocytes (reviewed by Barres, 1991) has renewed interest in how these cells respond to neuronal activity. To investigate the physiology of neuron astrocyte signaling, we have employed primary cortical cultures that contain both neuronal and glial cells. As the neurons in these cultures exhibit synchronous spontaneous synaptic activity, we have used both calcium imaging and whole-cell recording techniques to identify physiological activity in astrocytes related to neuronal activity. Whole-cell voltage-clamp records from astrocytes revealed rapid inward currents that coincide with bursts of electrical activity in neighboring neurons. Calcium imaging studies demonstrate that these currents in astrocytes are not always associated with slowly propagating calcium waves. Inclusion of the dye Lucifer yellow within patch pipettes confirmed that astrocytes are extensively coupled to each other but not to adjacent neurons, indicating that the currents observed are not due to gap junction connections between these cell types. These currents do not reflect widespread diffusion of glutamate or potassium released during neuronal activity since a population of small, round, multipolar presumed glial cells that are not dye coupled to adjacent cells did not display electrical currents coincident with neuronal firing, even though they respond to locally applied glutamate and potassium. These findings indicate that, in addition to the relatively slow signaling conveyed by calcium waves, astrocytes also display rapid electrical responses to neuronal activity.

Dual-Energy Computed Tomography Gemstone Spectral Imaging: A Novel Technique to Determine Human Cardiac Calculus Composition.

PubMed

Cheng, Ching-Li; Chang, Hsiao-Huang; Ko, Shih-Chi; Huang, Pei-Jung; Lin, Shan-Yang

2016-01-01

Understanding the chemical composition of any calculus in different human organs is essential for choosing the best treatment strategy for patients. The purpose of this study was to assess the capability of determining the chemical composition of a human cardiac calculus using gemstone spectral imaging (GSI) mode on a single-source dual-energy computed tomography (DECT) in vitro. The cardiac calculus was directly scanned on the Discovery CT750 HD FREEdom Edition using GSI mode, in vitro. A portable fiber-optic Raman spectroscopy was also applied to verify the quantitative accuracy of the DECT measurements. The results of spectral DECT measurements indicate that effective Z values in 3 designated positions located in this calculus were 15.02 to 15.47, which are close to values of 15.74 to 15.86, corresponding to the effective Z values of calcium apatite and hydroxyapatite. The Raman spectral data were also reflected by the predominant Raman peak at 960 cm for hydroxyapatite and the minor peak at 875 cm for calcium apatite. A potential single-source DECT with GSI mode was first used to examine the morphological characteristics and chemical compositions of a giant human cardiac calculus, in vitro. The CT results were consistent with the Raman spectral data, suggesting that spectral CT imaging techniques could be accurately used to diagnose and characterize the compositional materials in the cardiac calculus.

A novel approach to breast cancer diagnosis via PET imaging of microcalcifications using 18F-NaF

PubMed Central

Wilson, George H.; Gore, John C.; Yankeelov, Thomas E.; Barnes, Stephanie; Peterson, Todd E.; True, Jarrod M.; Shokouhi, Sepideh; McIntyre, J. Oliver.; Sanders, Melinda; Abramson, Vandana; Ngyuen, The-Quyen; Mahadevan-Jansen, Anita; Tantawy, Mohammed N.

2015-01-01

Rationale Current radiological methods for diagnosing breast cancer detect specific morphological features of solid tumors and/or any associated calcium deposits. These deposits originate from an early molecular microcalcification process which consists of two types: type 1 is calcium oxylate (CO) and type II is carbonated calcium hydroxyapetite (HAP). Type I microcalcifications are mainly associated with benign tumors while type II have been shown to be produced, internally, by malignant cells. No current non-invasive in vivo techniques are available for detecting intratumoral microcalcifications. Such a technique would have a significant impact on breast cancer diagnosis and prognosis in preclinical and clinical settings. 18F-NaF PET has been solely used for bone imaging by targeting the bone HAP. In this work, we provide preliminary evidence that 18F-NaF PET imaging can be used to detect breast cancer by targeting the HAP lattice within the tumor microenvironment with high specificity and soft-tissue contrast-to-background ratio, while delineating tumors from inflammation. METHODS Mice were injected with approximately 106 MDA-MB-231 cells subcutaneously and imaged with 18F-NaF PET/CT in a 120 min dynamic sequence when the tumors reached a size of ~250 mm3. Regions-of-interest (ROIs) were drawn around the tumor, muscle, and bone. The concentration of the radiotracer within those ROIs were compared to one another. For comparison to inflammation, rats with inflammatory paws were subjected to 18F-NaF PET imaging. RESULTS Tumor uptake of 18F− was significantly higher (p<0.05) than muscle uptake where the tumor-to-muscle ratio was ~3.5. The presence of type II microcalcification in the MDA-MB-231 cell line was confirmed histologically using alizarin red S and von Kossa staining as well as Raman microspectroscopy. No uptake of 18F− was observed in the rat inflamed tissue. Lack of HAP in the inflamed tissue was verified histologically. CONCLUSIONS This study provides preliminary evidence suggesting that specific targeting of the HAP within the tumor microenvironment with 18F may be able to distinguishing between inflammation and cancer. PMID:24833491

Bisphenol A stimulates human prostate cancer cell migration via remodelling of calcium signalling.

PubMed

Derouiche, Sandra; Warnier, Marine; Mariot, Pascal; Gosset, Pierre; Mauroy, Brigitte; Bonnal, Jean-Louis; Slomianny, Christian; Delcourt, Philippe; Prevarskaya, Natalia; Roudbaraki, Morad

2013-12-01

Bisphenol A (BPA), the principal constituent of reusable water bottles, metal cans, and plastic food containers, has been shown to be involved in human prostate cancer (PCa) cell proliferation. The aim of the present study was to explore the effect of BPA on PCa cell migration and the pathways involved in these processes. Using the transwell technique, we clearly show for the first time that the pre-treatment of the cells with BPA (1-10 nM) induces human PCa cell migration. Using a calcium imaging technique, we show that BPA pre-treatment induces an amplification of Store-Operated Calcium Entry (SOCE) in LNCaP cells. RT-PCR and Western blot experiments allowed the identification of the ion channel proteins which are up-regulated by BPA pre-treatments. These include the Orai1 protein, which is known as an important SOCE actor in various cell systems, including human PCa cells. Using a siRNA strategy, we observed that BPA-induced amplification of SOCE was Orai1-dependent. Interestingly, the BPA-induced PCa cell migration was suppressed when the calcium entry was impaired by the use of SOCE inhibitors (SKF96365, BTP2), or when the extracellular calcium was chelated. Taken together, the results presented here show that BPA induces PCa cells migration via a modulation of the ion channel protein expression involved in calcium entry and in cancer cell migration. The present data provide novel insights into the molecular mechanisms involved in the effects of an environmental factor on cancer cells and suggest both the necessity of preventive measures and the possibility of targeting ion channels in the treatment of PCa cell metastasis.

Nanoscale analysis of the morphology and surface stability of calcium carbonate polymorphs

PubMed Central

Sekkal, W.; Zaoui, A.

2013-01-01

Under earth surface conditions, in ocean and natural water, calcium carbonate is ubiquitous, forming anhydrous and hydrous minerals. These hydrous phases are of considerable interest for their role as precursors to stable carbonate minerals. Atomistic simulation techniques have been employed here to perform a comprehensive and quantitative study of the structural and energetic stability of dry and hydrous surfaces of calcium carbonate polymorphs using two recently developed forcefields. Results show that the dry forms are prone to ductility; while hydrous phases are found to be brittle. The (001) surface of monohydrocalcite appears to be the most stable (0.99 J/m2) whereas for the ikaite phase, the (001) surface is the most stable. The corresponding value is 0.2 J/m2, i.e. even lower than the surface energy of the Beautiful computed morphology pictures are obtained with Xiao's model and are very similar to the observed SEM images. PMID:23545842

Nanoscale analysis of the morphology and surface stability of calcium carbonate polymorphs.

PubMed

Sekkal, W; Zaoui, A

2013-01-01

Under earth surface conditions, in ocean and natural water, calcium carbonate is ubiquitous, forming anhydrous and hydrous minerals. These hydrous phases are of considerable interest for their role as precursors to stable carbonate minerals. Atomistic simulation techniques have been employed here to perform a comprehensive and quantitative study of the structural and energetic stability of dry and hydrous surfaces of calcium carbonate polymorphs using two recently developed forcefields. Results show that the dry forms are prone to ductility; while hydrous phases are found to be brittle. The (001) surface of monohydrocalcite appears to be the most stable (0.99 J/m(2)) whereas for the ikaite phase, the (001) surface is the most stable. The corresponding value is 0.2 J/m(2), i.e. even lower than the surface energy of the Beautiful computed morphology pictures are obtained with Xiao's model and are very similar to the observed SEM images.

Nanoscale analysis of the morphology and surface stability of calcium carbonate polymorphs

NASA Astrophysics Data System (ADS)

Sekkal, W.; Zaoui, A.

2013-04-01

Under earth surface conditions, in ocean and natural water, calcium carbonate is ubiquitous, forming anhydrous and hydrous minerals. These hydrous phases are of considerable interest for their role as precursors to stable carbonate minerals. Atomistic simulation techniques have been employed here to perform a comprehensive and quantitative study of the structural and energetic stability of dry and hydrous surfaces of calcium carbonate polymorphs using two recently developed forcefields. Results show that the dry forms are prone to ductility; while hydrous phases are found to be brittle. The (001) surface of monohydrocalcite appears to be the most stable (0.99 J/m2) whereas for the ikaite phase, the (001) surface is the most stable. The corresponding value is 0.2 J/m2, i.e. even lower than the surface energy of the Beautiful computed morphology pictures are obtained with Xiao's model and are very similar to the observed SEM images.

Heavy metal cations permeate the TRPV6 epithelial cation channel.

PubMed

Kovacs, Gergely; Danko, Tamas; Bergeron, Marc J; Balazs, Bernadett; Suzuki, Yoshiro; Zsembery, Akos; Hediger, Matthias A

2011-01-01

TRPV6 belongs to the vanilloid family of the transient receptor potential channel (TRP) superfamily. This calcium-selective channel is highly expressed in the duodenum and the placenta, being responsible for calcium absorption in the body and fetus. Previous observations have suggested that TRPV6 is not only permeable to calcium but also to other divalent cations in epithelial tissues. In this study, we tested whether TRPV6 is indeed also permeable to cations such as zinc and cadmium. We found that the basal intracellular calcium concentration was higher in HEK293 cells transfected with hTRPV6 than in non-transfected cells, and that this difference almost disappeared in nominally calcium-free solution. Live cell imaging experiments with Fura-2 and NewPort Green DCF showed that overexpression of human TRPV6 increased the permeability for Ca(2+), Ba(2+), Sr(2+), Mn(2+), Zn(2+), Cd(2+), and interestingly also for La(3+) and Gd(3+). These results were confirmed using the patch clamp technique. (45)Ca uptake experiments showed that cadmium, lanthanum and gadolinium were also highly efficient inhibitors of TRPV6-mediated calcium influx at higher micromolar concentrations. Our results suggest that TRPV6 is not only involved in calcium transport but also in the transport of other divalent cations, including heavy metal ions, which may have toxicological implications. Copyright © 2010 Elsevier Ltd. All rights reserved.

Light Sheet Fluorescence Microscopy Quantifies Calcium Oscillations in Root Hairs of Arabidopsis thaliana.

PubMed

Candeo, Alessia; Doccula, Fabrizio G; Valentini, Gianluca; Bassi, Andrea; Costa, Alex

2017-07-01

Calcium oscillations play a role in the regulation of the development of tip-growing plant cells. Using optical microscopy, calcium oscillations have been observed in a few systems (e.g. pollen tubes, fungal hyphae and algal rhizoids). High-resolution, non-phototoxic and rapid imaging methods are required to study the calcium oscillation in root hairs. We show that light sheet fluorescence microscopy is optimal to image growing root hairs of Arabidopsis thaliana and to follow their oscillatory tip-focused calcium gradient. We describe a protocol for performing live imaging of root hairs in seedlings expressing the cytosol-localized ratiometric calcium indicator Yellow Cameleon 3.6. Using this protocol, we measured the calcium gradient in a large number of root hairs. We characterized their calcium oscillations and correlated them with the rate of hair growth. The method was then used to screen the effect of auxin on the properties of the growing root hairs. © The Author 2017. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists.

Metallic nanoparticles and their medicinal potential. Part II: aluminosilicates, nanobiomagnets, quantum dots and cochleates.

PubMed

Loomba, Leena; Scarabelli, Tiziano

2013-09-01

Metallic miniaturization techniques have taken metals to nanoscale size where they can display fascinating properties and their potential applications in medicine. In recent years, metal nanoparticles such as aluminium, silicon, iron, cadmium, selenium, indium and calcium, which find their presence in aluminosilicates, nanobiomagnets, quantum dots (Q-dots) and cochleates, have caught attention of medical industries. The increasing impact of metallic nanoparticles in life sciences has significantly advanced the production techniques for these nanoparticles. In this Review, the various methods for the synthesis of nanoparticles are outlined, followed by their physicochemical properties, some recent applications in wound healing, diagnostic imaging, biosensing, assay labeling, antimicrobial activity, cancer therapy and drug delivery are listed, and finally their toxicological impacts are revised. The first half of this article describes the medicinal uses of two noble nanoparticles - gold and silver. This Review provides further information on the ability of aluminum, silicon, iron, selenium, indium, calcium and zinc to be used as nanoparticles in biomedical sciences. Aluminosilicates find their utility in wound healing and antibacterial growth. Iron-oxide nanoparticles enhance the properties of MRI contrast agents and are also used as biomagnets. Cadmium, selenium, tellurium and indium form the core nanostructures of tiny Q-dots used in cellular assay labeling, high-resolution cell imaging and biosensing. Cochleates have the bivalent nano ions calcium, magnesium or zinc imbedded in their structures and are considered to be highly effective agents for drug and gene delivery. The aluminosilicates, nanobiomagnets, Q-dots and cochleates are discussed in the light of their properties, synthesis and utility.

Live-cell calcium imaging of adherent and non-adherent GL261 cells reveals phenotype-dependent differences in drug responses.

PubMed

Strong, Averey D; Daniels, Richard L

2017-08-02

The tumor-derived GL261 cell line is used as a model for studying glioblastoma and other high-grade gliomas, and can be cultured adherently or as free-floating aggregates known as neurospheres. These different culture conditions give rise to distinct phenotypes, with increased tumorigenicity displayed by neurosphere-cultured cells. An important technique for understanding GL261 pathobiology is live cell fluorescent imaging of intracellular calcium. However, live cell imaging of GL261 neurospheres presents a technical challenge, as experimental manipulations where drugs are added to the extracellular media cause the cells to move during analysis. Here we present a method to immobilize GL261 neurospheres with low melting point agarose for calcium imaging using the fluorescent calcium sensor fura-2. GL261 cells were obtained from the NCI-Frederick Cancer Research Tumor Repository and cultured as adherent cells or induced to form neurospheres by placing freshly trypsinized cells into serum-free media containing fibroblast growth factor 2, epidermal growth factor, and B-27 supplement. Prior to experiments, adherent cells were loaded with fura-2 and cultured on 8-well chamber slides. Non-adherent neurospheres were first loaded with fura-2, placed in droplets onto an 8-well chamber slide, and finally covered with a thin layer of low melting point agarose to immobilize the cells. Ratiometric pseudocolored images were obtained during treatment with ATP, capsaicin, or vehicle control. Cells were marked as responsive if fluorescence levels increased more than 30% above baseline. Differences between treatment groups were tested using Student's t-tests and one-way ANOVA. We found that cellular responses to pharmacological treatments differ based on cellular phenotype. Adherent cells and neurospheres both responded to ATP with a rise in intracellular calcium. Notably, capsaicin treatment led to robust responses in GL261 neurospheres but not adherent cells. We demonstrate the use of low melting point agarose for immobilizing GL261 cells, a method that is broadly applicable to any cell type cultured in suspension, including acutely trypsinized cells and primary tumor cells. Our results indicate that it is important to consider GL261 phenotype (adherent or neurosphere) when interpreting data regarding physiological responses to experimental compounds.

Preparation and preclinical evaluation of 68Ga-DOTA-amlodipine for L-type calcium channel imaging.

PubMed

Firuzyar, Tahereh; Jalilian, Amir Reza; Aboudzadeh, Mohammad Reza; Sadeghpour, Hossein; Shafiee-Ardestani, Mahdi; Khalaj, Ali

2016-01-01

In order to develop a possible tracer for L-type calcium channel imaging, we here report the development of a Ga-68 amlodipine derivative for possible PET imaging. Amlodipine DOTA conjugate was synthesized, characterized and went through calcium channel blockade, toxicity, apoptosis/necrosis tests. [ 68 Ga] DOTA AMLO was prepared at optimized conditions followed by stability tests, partition coefficient determination and biodistribution studies using tissue counting and co incidence imaging up to 2 h. [ 68 Ga] DOTA AMLO was prepared at pH 4-5 in 7-10 min at 95°C in high radiochemical purity (>99%, radio thin layer chromatography; specific activity: 1.9-2.1 GBq/mmol) and was stable up to 4 h with a log P of -0.94. Calcium channel rich tissues including myocardium, and tissues with smooth muscle cells such as colon, intestine, and lungs demonstrated significant uptake. Co incidence images supported the biodistribution data up to 2 h. The complex can be a candidate for further positron emission tomography imaging for L type calcium channels.

Seizure activity results in calcium- and mitochondria-independent ROS production via NADPH and xanthine oxidase activation

PubMed Central

Kovac, S; Domijan, A-M; Walker, M C; Abramov, A Y

2014-01-01

Seizure activity has been proposed to result in the generation of reactive oxygen species (ROS), which then contribute to seizure-induced neuronal damage and eventually cell death. Although the mechanisms of seizure-induced ROS generation are unclear, mitochondria and cellular calcium overload have been proposed to have a crucial role. We aim to determine the sources of seizure-induced ROS and their contribution to seizure-induced cell death. Using live cell imaging techniques in glioneuronal cultures, we show that prolonged seizure-like activity increases ROS production in an NMDA receptor-dependent manner. Unexpectedly, however, mitochondria did not contribute to ROS production during seizure-like activity. ROS were generated primarily by NADPH oxidase and later by xanthine oxidase (XO) activity in a calcium-independent manner. This calcium-independent neuronal ROS production was accompanied by an increase in intracellular [Na+] through NMDA receptor activation. Inhibition of NADPH or XO markedly reduced seizure-like activity-induced neuronal apoptosis. These findings demonstrate a critical role for ROS in seizure-induced neuronal cell death and identify novel therapeutic targets. PMID:25275601

Hexamethyldisilazane Removal with Mesoporous Materials Prepared from Calcium Fluoride Sludge.

PubMed

Kao, Ching-Yang; Lin, Min-Fa; Nguyen, Nhat-Thien; Tsai, Hsiao-Hsin; Chang, Luh-Maan; Chen, Po-Han; Chang, Chang-Tang

2018-05-01

A large amount of calcium fluoride sludge is generated by the semiconductor industry every year. It also requires a high amount of fuel consumption using rotor concentrators and thermal oxidizers to treat VOCs. The mesoporous adsorbent prepared by calcium fluoride sludge was used for VOCs treatment. The semiconductor industry employs HMDS to promote the adhesion of photo-resistant material to oxide(s) due to the formation of silicon dioxide, which blocks porous adsorbents. The adsorption of HMDS (Hexamethyldisiloxane) was tested with mesoporous silica materials synthesized from calcium fluoride (CF-MCM). The resulting samples were characterized by XRD, XRF, FTIR, N2-adsorption-desorption techniques. The prepared samples possessed high specific surface area, large pore volume and large pore diameter. The crystal patterns of CF-MCM were similar with Mobil composite matter (MCM-41) from TEM image. The adsorption capacity of HMDS with CF-MCM was 40 and 80 mg g-1, respectively, under 100 and 500 ppm HMDS. The effects of operation parameters, such as contact time and mixture concentration, on the performance of CF-MCM were also discussed in this study.

Label-free Imaging of Thyroid and Parathyroid Glands Using Coherent Anti-Stokes Raman Scattering (CARS) Microscopy

NASA Astrophysics Data System (ADS)

Weng, Sheng

Thyroid and parathyroid glands play a vital role in regulating the body's metabolism and calcium levels. Surgical removal of the glands is the main treatment for both thyroid cancer and parathyroid adenoma. In thyroidectomy and parathyroidectomy, it's very important to differentiate thyroid, parathyroid, and the other tissues around the neck. Traditionally, physicians use ultrasound guided fine needle aspiration (FNA) to evaluate thyroid nodules, but up to 30% of FNA results are "inconclusive". The sestamibi scan can localize parathyroid adenoma, but currently it only has 50% accuracy. Here we applied the emerging CARS technique to image both thyroid and parathyroid tissues, which has potential to be used in real-time in vivo examination of different structures. We also developed algorithms to differentiate different cellular structures based on CARS images. When incorporated with a fiber optic endoscope in the future, CARS imaging technique can help surgeons identify cancerous thyroid tissue intraoperatively, preserve good parathyroid glands during thyroidectomy and find parathyroid adenoma during parathyroidectomy.

Pan-neuronal calcium imaging with cellular resolution in freely swimming zebrafish.

PubMed

Kim, Dal Hyung; Kim, Jungsoo; Marques, João C; Grama, Abhinav; Hildebrand, David G C; Gu, Wenchao; Li, Jennifer M; Robson, Drew N

2017-11-01

Calcium imaging with cellular resolution typically requires an animal to be tethered under a microscope, which substantially restricts the range of behaviors that can be studied. To expand the behavioral repertoire amenable to imaging, we have developed a tracking microscope that enables whole-brain calcium imaging with cellular resolution in freely swimming larval zebrafish. This microscope uses infrared imaging to track a target animal in a behavior arena. On the basis of the predicted trajectory of the animal, we applied optimal control theory to a motorized stage system to cancel brain motion in three dimensions. We combined this motion-cancellation system with differential illumination focal filtering, a variant of HiLo microscopy, which enabled us to image the brain of a freely swimming larval zebrafish for more than an hour. This work expands the repertoire of natural behaviors that can be studied with cellular-resolution calcium imaging to potentially include spatial navigation, social behavior, feeding and reward.

Resveratrol Interferes with Fura-2 Intracellular Calcium Measurements.

PubMed

Kopp, Richard F; Leech, Colin A; Roe, Michael W

2014-03-01

Resveratrol, a naturally occurring polyphenol found in some fruits and especially in grapes, has been reported to provide diverse health benefits. Resveratrol's mechanism of action is the subject of many investigations, and some studies using the ratiometric calcium indicator Fura-2 suggest that it modulates cellular calcium responses. In the current study, contradictory cellular calcium responses to resveratrol applied at concentrations exceeding 10 μM were observed during in vitro imaging studies depending on the calcium indicator used, with Fura-2 indicating an increase in intracellular calcium while Fluo-4 and the calcium biosensor YC3.60 indicated no response. When cells loaded with Fura-2 were treated with 100 μM resveratrol, excitation at 340 nm resulted in a large intensity increase at 510 nm, but the expected concurrent decline with 380 nm excitation was not observed. Pre-treatment of cells with the calcium chelator BAPTA-AM did not prevent a rise in the 340/380 ratio when resveratrol was present, but it did prevent an increase in 340/380 when ATP was applied, suggesting that the resveratrol response was an artifact. Cautious data interpretation is recommended from imaging experiments using Fura-2 concurrently with resveratrol in calcium imaging experiments.

GCaMP expression in retinal ganglion cells characterized using a low-cost fundus imaging system

NASA Astrophysics Data System (ADS)

Chang, Yao-Chuan; Walston, Steven T.; Chow, Robert H.; Weiland, James D.

2017-10-01

Objective. Virus-transduced, intracellular-calcium indicators are effective reporters of neural activity, offering the advantage of cell-specific labeling. Due to the existence of an optimal time window for the expression of calcium indicators, a suitable tool for tracking GECI expression in vivo following transduction is highly desirable. Approach. We developed a noninvasive imaging approach based on a custom-modified, low-cost fundus viewing system that allowed us to monitor and characterize in vivo bright-field and fluorescence images of the mouse retina. AAV2-CAG-GCaMP6f was injected into a mouse eye. The fundus imaging system was used to measure fluorescence at several time points post injection. At defined time points, we prepared wholemount retina mounted on a transparent multielectrode array and used calcium imaging to evaluate the responsiveness of retinal ganglion cells (RGCs) to external electrical stimulation. Main results. The noninvasive fundus imaging system clearly resolves individual (RGCs and axons. RGC fluorescence intensity and the number of observable fluorescent cells show a similar rising trend from week 1 to week 3 after viral injection, indicating a consistent increase of GCaMP6f expression. Analysis of the in vivo fluorescence intensity trend and in vitro neurophysiological responsiveness shows that the slope of intensity versus days post injection can be used to estimate the optimal time for calcium imaging of RGCs in response to external electrical stimulation. Significance. The proposed fundus imaging system enables high-resolution digital fundus imaging in the mouse eye, based on off-the-shelf components. The long-term tracking experiment with in vitro calcium imaging validation demonstrates the system can serve as a powerful tool monitoring the level of genetically-encoded calcium indicator expression, further determining the optimal time window for following experiment.

Simultaneous measurement and modulation of multiple physiological parameters in the isolated heart using optical techniques

PubMed Central

Lee, Peter; Yan, Ping; Ewart, Paul; Kohl, Peter

2012-01-01

Whole-heart multi-parametric optical mapping has provided valuable insight into the interplay of electro-physiological parameters, and this technology will continue to thrive as dyes are improved and technical solutions for imaging become simpler and cheaper. Here, we show the advantage of using improved 2nd-generation voltage dyes, provide a simple solution to panoramic multi-parametric mapping, and illustrate the application of flash photolysis of caged compounds for studies in the whole heart. For proof of principle, we used the isolated rat whole-heart model. After characterising the blue and green isosbestic points of di-4-ANBDQBS and di-4-ANBDQPQ, respectively, two voltage and calcium mapping systems are described. With two newly custom-made multi-band optical filters, (1) di-4-ANBDQBS and fluo-4 and (2) di-4-ANBDQPQ and rhod-2 mapping are demonstrated. Furthermore, we demonstrate three-parameter mapping using di-4-ANBDQPQ, rhod-2 and NADH. Using off-the-shelf optics and the di-4-ANBDQPQ and rhod-2 combination, we demonstrate panoramic multi-parametric mapping, affording a 360° spatiotemporal record of activity. Finally, local optical perturbation of calcium dynamics in the whole heart is demonstrated using the caged compound, o-nitrophenyl ethylene glycol tetraacetic acid (NP-EGTA), with an ultraviolet light-emitting diode (LED). Calcium maps (heart loaded with di-4-ANBDQPQ and rhod-2) demonstrate successful NP-EGTA loading and local flash photolysis. All imaging systems were built using only a single camera. In conclusion, using novel 2nd-generation voltage dyes, we developed scalable techniques for multi-parametric optical mapping of the whole heart from one point of view and panoramically. In addition to these parameter imaging approaches, we show that it is possible to use caged compounds and ultraviolet LEDs to locally perturb electrophysiological parameters in the whole heart. PMID:22886365

Video-rate volumetric neuronal imaging using 3D targeted illumination.

PubMed

Xiao, Sheng; Tseng, Hua-An; Gritton, Howard; Han, Xue; Mertz, Jerome

2018-05-21

Fast volumetric microscopy is required to monitor large-scale neural ensembles with high spatio-temporal resolution. Widefield fluorescence microscopy can image large 2D fields of view at high resolution and speed while remaining simple and costeffective. A focal sweep add-on can further extend the capacity of widefield microscopy by enabling extended-depth-of-field (EDOF) imaging, but suffers from an inability to reject out-of-focus fluorescence background. Here, by using a digital micromirror device to target only in-focus sample features, we perform EDOF imaging with greatly enhanced contrast and signal-to-noise ratio, while reducing the light dosage delivered to the sample. Image quality is further improved by the application of a robust deconvolution algorithm. We demonstrate the advantages of our technique for in vivo calcium imaging in the mouse brain.

Silver nanoparticle-induced degranulation observed with quantitative phase microscopy

NASA Astrophysics Data System (ADS)

Yang, Wenzhong; Lee, Seungrag; Lee, Jiyong; Bae, Yoonsung; Kim, Dugyoung

2010-07-01

Monitoring a degranulation process in a live mast cell is a quite important issue in immunology and pharmacology. Because the size of a granule is normally much smaller than the resolution limit of an optical microscope system, there is no direct real-time live cell imaging technique for observing degranulation processes except for fluorescence imaging techniques. In this research, we propose optical quantitative phase microscopy (QPM) as a new observation tool to study degranulation processes in a live mast cell without any fluorescence labeling. We measure the cell volumes and the cross sectional profiles (x-z plane) of an RBL-2H3 cell and a HeLa cell, before and after they are exposed to calcium ionophore A23187 and silver nanoparticles (AgNPs). We verify that the volume and the cross sectional line profile of the RBL-2H3 cell were changed significantly when it was exposed to A23187. When 50 μg/mL of AgNP is used instead of A23187, the measurements of cell volume and cross sectional profiles indicate that RBL-2H3 cells also follow degranulation processes. Degranulation processes for these cells are verified by monitoring the increase of intracellular calcium ([Ca2+]i) and histamine with fluorescent methods.

Cardiovascular Imaging Using Two-Photon Microscopy

PubMed Central

Scherschel, John A.; Rubart, Michael

2008-01-01

Two-photon excitation microscopy has become the standard technique for high resolution deep tissue and intravital imaging. It provides intrinsic three-dimensional resolution in combination with increased penetration depth compared to single-photon confocal microscopy. This article will describe the basic physical principles of two-photon excitation and will review its multiple applications to cardiovascular imaging, including second harmonic generation and fluorescence laser scanning microscopy. In particular, the capability and limitations of multiphoton microscopy to assess functional heterogeneity on a cellular scale deep within intact, Langendorff-perfused hearts are demonstrated. It will also discuss the use of two-photon excitation-induced release of caged compounds for the study of intracellular calcium signaling and intercellular dye transfer. PMID:18986603

Imaging powders with the atomic force microscope: from biominerals to commercial materials.

PubMed

Friedbacher, G; Hansma, P K; Ramli, E; Stucky, G D

1991-09-13

Atomically resolved images of pressed powder samples have been obtained with the atomic force microscope (AFM). The technique was successful in resolving the particle, domain, and atomic structure of pismo clam (Tivela stultorum) and sea urchin (Strongylocentrotus purpuratus) shells and of commercially available calcium carbonate (CaCO(3)) and strontium carbonate (SrCO(3)) powders. Grinding and subsequent pressing of the shells did not destroy the microstructure of these materials. The atomic-resolution imaging capabilities of AFM can be applied to polycrystalline samples by means of pressing powders with a grain size as small as 50 micrometers. These results illustrate that the AFM is a promising tool for material science and the study of biomineralization.

Calcium and bones (image)

MedlinePlus

Calcium is one of the most important minerals for the growth, maintenance, and reproduction of the human ... body, are continually being re-formed and incorporate calcium into their structure. Calcium is essential for the ...

Preparation and preclinical evaluation of 68Ga-DOTA-amlodipine for L-type calcium channel imaging

PubMed Central

Firuzyar, Tahereh; Jalilian, Amir Reza; Aboudzadeh, Mohammad Reza; Sadeghpour, Hossein; Shafiee-Ardestani, Mahdi; Khalaj, Ali

2016-01-01

Aim: In order to develop a possible tracer for L-type calcium channel imaging, we here report the development of a Ga-68 amlodipine derivative for possible PET imaging. Materials and Methods: Amlodipine DOTA conjugate was synthesized, characterized and went through calcium channel blockade, toxicity, apoptosis/necrosis tests. [68Ga] DOTA AMLO was prepared at optimized conditions followed by stability tests, partition coefficient determination and biodistribution studies using tissue counting and co incidence imaging up to 2 h. Results: [68Ga] DOTA AMLO was prepared at pH 4–5 in 7–10 min at 95°C in high radiochemical purity (>99%, radio thin layer chromatography; specific activity: 1.9–2.1 GBq/mmol) and was stable up to 4 h with a log P of −0.94. Calcium channel rich tissues including myocardium, and tissues with smooth muscle cells such as colon, intestine, and lungs demonstrated significant uptake. Co incidence images supported the biodistribution data up to 2 h. Conclusions: The complex can be a candidate for further positron emission tomography imaging for L type calcium channels. PMID:27833311

Coronary calcium visualization using dual energy chest radiography with sliding organ registration

NASA Astrophysics Data System (ADS)

Wen, Di; Nye, Katelyn; Zhou, Bo; Gilkeson, Robert C.; Wilson, David L.

2016-03-01

Coronary artery calcification (CAC) is the lead biomarker for atherosclerotic heart disease. We are developing a new technique to image CAC using ubiquitously ordered, low cost, low radiation dual energy (DE) chest radiography (using the two-shot GE Revolution XRd system). In this paper, we proposed a novel image processing method (CorCalDx) based on sliding organ registration to create a bone-image-like, coronary calcium image (CCI) that significantly reduces motion artifacts and improves CAC conspicuity. Experiments on images of a physical dynamic cardiac phantom showed that CorCalDx reduced 73% of the motion artifact area as compared to standard DE over a range of heart rates up to 90 bpm and varying x-ray radiation exposures. Residual motion artifact in the phantom CCI is greatly suppressed in gray level and area (0.88% of the heart area). In a Functional Measurement Test (FMT) with 20 clinical exams, image quality improvement of CorCalDx against standard DE (measured from -10 to +10) was significantly suggested (p<0.0001) by three radiologists for cardiac motion artifacts (7.2+/-2.1) and cardiac anatomy visibility (6.1+/-3.5). CorCalDx was always chosen best in every image tested. In preliminary assessments of 12 patients with 18 calcifications, 90% of motion artifact regions in standard DE results were removed in CorCalDx results, with 100% sensitivity of calcification detection, showing great potential of CorCalDx to improve CAC detection and grading in DE chest radiography.

Simultaneous Denoising, Deconvolution, and Demixing of Calcium Imaging Data

PubMed Central

Pnevmatikakis, Eftychios A.; Soudry, Daniel; Gao, Yuanjun; Machado, Timothy A.; Merel, Josh; Pfau, David; Reardon, Thomas; Mu, Yu; Lacefield, Clay; Yang, Weijian; Ahrens, Misha; Bruno, Randy; Jessell, Thomas M.; Peterka, Darcy S.; Yuste, Rafael; Paninski, Liam

2016-01-01

SUMMARY We present a modular approach for analyzing calcium imaging recordings of large neuronal ensembles. Our goal is to simultaneously identify the locations of the neurons, demix spatially overlapping components, and denoise and deconvolve the spiking activity from the slow dynamics of the calcium indicator. Our approach relies on a constrained nonnegative matrix factorization that expresses the spatiotemporal fluorescence activity as the product of a spatial matrix that encodes the spatial footprint of each neuron in the optical field and a temporal matrix that characterizes the calcium concentration of each neuron over time. This framework is combined with a novel constrained deconvolution approach that extracts estimates of neural activity from fluorescence traces, to create a spatiotemporal processing algorithm that requires minimal parameter tuning. We demonstrate the general applicability of our method by applying it to in vitro and in vivo multineuronal imaging data, whole-brain light-sheet imaging data, and dendritic imaging data. PMID:26774160

Calcium source (image)

MedlinePlus

Getting enough calcium to keep bones from thinning throughout a person's life may be made more difficult if that person has ... as a tendency toward kidney stones, for avoiding calcium-rich food sources. Calcium deficiency also effects the ...

Discrimination of clinically significant calcium salts using MARS spectral CT

NASA Astrophysics Data System (ADS)

Kirkbride, T. E.; Raja, A.; Mueller, K.; Bateman, C. J.; Becce, F.; Anderson, N.

2017-03-01

Calcium compounds within tissues are usually a sign of pathology, and calcium crystal type is often a pointer to the diagnosis. There are clinical advantages in being able to determine the quantity and type of calcifications non-invasively in cardiovascular, genitourinary and musculoskeletal disorders, and treatment differs depending on the crystal type and quantity. The problem arises when trying to distinguish between different calcium compounds within the same image due to their similar attenuation properties. There are spectroscopic differences between calcium salts at very low energies. As calcium oxalate and calcium hydroxyapatite can co-exist in breast and musculoskeletal pathologies of the breast, we wished to determine whether Spectral CT could distinguish between them in the same image at clinical X-ray energy ranges. Energy thresholds of 15, 22, 29 and 36keV and tube voltages of 50, 80 and 110kVp were chosen, and images were analysed to determine the percentage difference in the attenuation coefficients of calcium hydroxyapatite samples at concentrations of 54.3, 211.7, 808.5 and 1169.3mg/ml, and calcium oxalate at a concentration of 2000 mg/ml. The two lower concentrations of calcium hydroxyapatite were distinguishable from calcium oxalate at all energies and all tube voltages, whereas the ability to discriminate oxalate from hydroxyapatite at higher concentrations was dependent on the threshold energy but only mildly dependent on the tube voltage used. Spectral CT shows promise for distinguishing clinically important calcium salts.

Development of practical red fluorescent probe for cytoplasmic calcium ions with greatly improved cell-membrane permeability.

PubMed

Hirabayashi, Kazuhisa; Hanaoka, Kenjiro; Egawa, Takahiro; Kobayashi, Chiaki; Takahashi, Shodai; Komatsu, Toru; Ueno, Tasuku; Terai, Takuya; Ikegaya, Yuji; Nagano, Tetsuo; Urano, Yasuteru

2016-10-01

Fluorescence imaging of calcium ions (Ca(2+)) has become an essential technique for investigation of signaling pathways involving Ca(2+) as a second messenger. But, Ca(2+) signaling is involved in many biological phenomena, and therefore simultaneous visualization of Ca(2+) and other biomolecules (multicolor imaging) would be particularly informative. For this purpose, we set out to develop a fluorescent probe for Ca(2+) that would operate in a different color region (red) from that of probes for other molecules, many of which show green fluorescence, as exemplified by green fluorescent protein (GFP). We previously developed a red fluorescent probe for monitoring cytoplasmic Ca(2+) concentration, based on our established red fluorophore, TokyoMagenta (TM), but there remained room for improvement, especially as regards efficiency of introduction into cells. We considered that this issue was probably mainly due to limited water solubility of the probe. So, we designed and synthesized a red-fluorescent probe with improved water solubility. We confirmed that this Ca(2+) red-fluorescent probe showed high cell-membrane permeability with bright fluorescence. It was successfully applied to fluorescence imaging of not only live cells, but also brain slices, and should be practically useful for multicolor imaging studies of biological mechanisms. Copyright © 2016 Elsevier Ltd. All rights reserved.

In vivo optoacoustic monitoring of calcium activity in the brain (Conference Presentation)

NASA Astrophysics Data System (ADS)

Deán-Ben, Xose Luís.; Gottschalk, Sven; Sela, Gali; Lauri, Antonella; Kneipp, Moritz; Ntziachristos, Vasilis; Westmeyer, Gil G.; Shoham, Shy; Razansky, Daniel

2017-03-01

Non-invasive observation of spatio-temporal neural activity of large neural populations distributed over the entire brain of complex organisms is a longstanding goal of neuroscience [1,2]. Recently, genetically encoded calcium indicators (GECIs) have revolutionized neuroimaging by enabling mapping the activity of entire neuronal populations in vivo [3]. Visualization of these powerful sensors with fluorescence microscopy has however been limited to superficial regions while deep brain areas have so far remained unreachable [4]. We have developed a volumetric multispectral optoacoustic tomography platform for imaging neural activation deep in scattering brains [5]. The developed methodology can render 100 volumetric frames per second across scalable fields of view ranging between 50-1000 mm3 with respective spatial resolution of 35-150µm. Experiments performed in immobilized and freely swimming larvae and in adult zebrafish brains expressing the genetically-encoded calcium indicator GCaMP5G demonstrated, for the first time, the fundamental ability to directly track neural dynamics using optoacoustics while overcoming the depth barrier of optical imaging in scattering brains [6]. It was further possible to monitor calcium transients in a scattering brain of a living adult transgenic zebrafish expressing GCaMP5G calcium indicator [7]. Fast changes in optoacoustic traces associated to GCaMP5G activity were detectable in the presence of other strongly absorbing endogenous chromophores, such as hemoglobin. The results indicate that the optoacoustic signal traces generally follow the GCaMP5G fluorescence dynamics and further enable overcoming the longstanding optical-diffusion penetration barrier associated to scattering in biological tissues [6]. The new functional optoacoustic neuroimaging method can visualize neural activity at penetration depths and spatio-temporal resolution scales not covered with the existing neuroimaging techniques. Thus, in addition to the well-established capacity of optoacoustics to resolve vascular anatomy and multiple hemodynamic parameters deep in scattering tissues, the newly developed methodology offers unprecedented capabilities for functional whole brain observations of fast calcium dynamics.

Synaptic activation of putative sensory neurons by hexamethonium-sensitive nerve pathways in mouse colon.

PubMed

Hibberd, Timothy J; Travis, Lee; Wiklendt, Lukasz; Costa, Marcello; Brookes, Simon J H; Hu, Hongzhen; Keating, Damien J; Spencer, Nick J

2018-01-01

The gastrointestinal tract contains its own independent population of sensory neurons within the gut wall. These sensory neurons have been referred to as intrinsic primary afferent neurons (IPANs) and can be identified by immunoreactivity to calcitonin gene-related peptide (CGRP) in mice. A common feature of IPANs is a paucity of fast synaptic inputs observed during sharp microelectrode recordings. Whether this is observed using different recording techniques is of particular interest for understanding the physiology of these neurons and neural circuit modeling. Here, we imaged spontaneous and evoked activation of myenteric neurons in isolated whole preparations of mouse colon and correlated recordings with CGRP and nitric oxide synthase (NOS) immunoreactivity, post hoc. Calcium indicator fluo 4 was used for this purpose. Calcium responses were recorded in nerve cell bodies located 5-10 mm oral to transmural electrical nerve stimuli. A total of 618 recorded neurons were classified for CGRP or NOS immunoreactivity. Aboral electrical stimulation evoked short-latency calcium transients in the majority of myenteric neurons, including ~90% of CGRP-immunoreactive Dogiel type II neurons. Activation of Dogiel type II neurons had a time course consistent with fast synaptic transmission and was always abolished by hexamethonium (300 μM) and by low-calcium Krebs solution. The nicotinic receptor agonist 1,1-dimethyl-4-phenylpiperazinium iodide (during synaptic blockade) directly activated Dogiel type II neurons. The present study suggests that murine colonic Dogiel type II neurons receive prominent fast excitatory synaptic inputs from hexamethonium-sensitive neural pathways. NEW & NOTEWORTHY Myenteric neurons in isolated mouse colon were recorded using calcium imaging and then neurochemically defined. Short-latency calcium transients were detected in >90% of calcitonin gene-related peptide-immunoreactive neurons to electrical stimulation of hexamethonium-sensitive pathways. Putative sensory Dogiel type II calcitonin gene-related peptide-immunoreactive myenteric neurons may receive widespread fast synaptic inputs in mouse colon.

Calcium-activated potassium channels in insect pacemaker neurons as unexpected target site for the novel fumigant dimethyl disulfide.

PubMed

Gautier, Hélène; Auger, Jacques; Legros, Christian; Lapied, Bruno

2008-01-01

Dimethyl disulfide (DMDS), a plant-derived insecticide, is a promising fumigant as a substitute for methyl bromide. To further understand the mode of action of DMDS, we examined its effect on cockroach octopaminergic neurosecretory cells, called dorsal unpaired median (DUM) neurons, using whole-cell patch-clamp technique, calcium imaging and antisense oligonucleotide strategy. At low concentration (1 microM), DMDS modified spontaneous regular spike discharge into clear bursting activity associated with a decrease of the amplitude of the afterhyperpolarization. This effect led us to suspect alterations of calcium-activated potassium currents (IKCa) and [Ca(2+)](i) changes. We showed that DMDS reduced amplitudes of both peak transient and sustained components of the total potassium current. IKCa was confirmed as a target of DMDS by using iberiotoxin, cadmium chloride, and pSlo antisense oligonucleotide. In addition, we showed that DMDS induced [Ca(2+)](i) rise in Fura-2-loaded DUM neurons. Using calcium-free solution, and (R,S)-(3,4-dihydro-6,7-dimethoxy-isoquinoline-1-yl)-2-phenyl-N,N-di-[2-(2,3,4-trimethoxy-phenyl)ethyl]-acetamide (LOE 908) [an inhibitor of transient receptor potential (TRP)gamma], we demonstrated that TRPgamma initiated calcium influx. By contrast, omega-conotoxin GVIA (an inhibitor of N-type high-voltage-activated calcium channels), did not affect the DMDS-induced [Ca(2+)](i) rise. Finally, the participation of the calcium-induced calcium release mechanism was investigated using thapsigargin, caffeine, and ryanodine. Our study revealed that DMDS-induced elevation in [Ca(2+)](i) modulated IKCa in an unexpected bell-shaped manner via intracellular calcium. In conclusion, DMDS affects multiple targets, which could be an effective way to improve pest control efficacy of fumigation.

Implementing Patch Clamp and Live Fluorescence Microscopy to Monitor Functional Properties of Freshly Isolated PKD Epithelium

PubMed Central

Pavlov, Tengis S.; Ilatovskaya, Daria V.; Palygin, Oleg; Levchenko, Vladislav; Pochynyuk, Oleh; Staruschenko, Alexander

2015-01-01

Cyst initiation and expansion during polycystic kidney disease is a complex process characterized by abnormalities in tubular cell proliferation, luminal fluid accumulation and extracellular matrix formation. Activity of ion channels and intracellular calcium signaling are key physiologic parameters which determine functions of tubular epithelium. We developed a method suitable for real-time observation of ion channels activity with patch-clamp technique and registration of intracellular Ca2+ level in epithelial monolayers freshly isolated from renal cysts. PCK rats, a genetic model of autosomal recessive polycystic kidney disease (ARPKD), were used here for ex vivo analysis of ion channels and calcium flux. Described here is a detailed step-by-step procedure designed to isolate cystic monolayers and non-dilated tubules from PCK or normal Sprague Dawley (SD) rats, and monitor single channel activity and intracellular Ca2+ dynamics. This method does not require enzymatic processing and allows analysis in a native setting of freshly isolated epithelial monolayer. Moreover, this technique is very sensitive to intracellular calcium changes and generates high resolution images for precise measurements. Finally, isolated cystic epithelium can be further used for staining with antibodies or dyes, preparation of primary cultures and purification for various biochemical assays. PMID:26381526

Subgrain boundary analyses in deformed orthopyroxene by TEM/STEM with EBSD-FIB sample preparation technique

NASA Astrophysics Data System (ADS)

Kogure, Toshihiro; Raimbourg, Hugues; Kumamoto, Akihito; Fujii, Eiko; Ikuhara, Yuichi

2014-12-01

High-resolution structure analyses using electron beam techniques have been performed for the investigation of subgrain boundaries (SGBs) in deformed orthopyroxene (Opx) in mylonite from Hidaka Metamorphic Belt, Hokkaido, Japan, to understand ductile deformation mechanism of silicate minerals in shear zones. Scanning electron microscopy (SEM) and electron backscatter diffraction (EBSD) analysis of Opx porphyroclasts in the mylonitic rock indicated that the crystal orientation inside the Opx crystals gradually changes by rotation about the b-axis by SGBs and crystal folding. In order to observe the SGBs along the b-axis by transmission electron microscopy (TEM) or scanning TEM (STEM), the following sample preparation protocol was adopted. First, petrographic thin sections were slightly etched with hydrofluoric acid to identify SGBs in SEM. The Opx crystals whose b-axes were oriented close to the normal of the surface were identified by EBSD, and the areas containing SGBs were picked and thinned for (S) TEM analysis with a focused ion beam instrument with micro-sampling system. High-resolution TEM imaging of the SGBs in Opx revealed various boundary structures from a periodic array of dissociated (100) [001] edge dislocations to partially or completely incoherent crystals, depending on the misorientation angle. Atomic-resolution STEM imaging clearly confirmed the formation of clinopyroxene (Cpx) structure between the dissociated partial dislocations. Moreover, X-ray microanalysis in STEM revealed that the Cpx contains a considerable amount of calcium replacing iron. Such chemical inhomogeneity may limit glide motion of the dislocation and eventually the plastic deformation of the Opx porphyroclasts at a low temperature. Chemical profiles across the high-angle incoherent SGB also showed an enrichment of the latter in calcium at the boundary, suggesting that SGBs are an efficient diffusion pathway of calcium out of host Opx grain during cooling.

Two-Photon Imaging with Diffractive Optical Elements

PubMed Central

Watson, Brendon O.; Nikolenko, Volodymyr; Yuste, Rafael

2009-01-01

Two-photon imaging has become a useful tool for optical monitoring of neural circuits, but it requires high laser power and serial scanning of each pixel in a sample. This results in slow imaging rates, limiting the measurements of fast signals such as neuronal activity. To improve the speed and signal-to-noise ratio of two-photon imaging, we introduce a simple modification of a two-photon microscope, using a diffractive optical element (DOE) which splits the laser beam into several beamlets that can simultaneously scan the sample. We demonstrate the advantages of DOE scanning by enhancing the speed and sensitivity of two-photon calcium imaging of action potentials in neurons from neocortical brain slices. DOE scanning can easily improve the detection of time-varying signals in two-photon and other non-linear microscopic techniques. PMID:19636390

Sensitivity to detect small coronary artery calcium lesions with varying slice thickness using electron beam tomography.

PubMed

Mao, Songshou; Child, Janis; Carson, Sivi; Liu, Steve C K; Oudiz, Ronald J; Budoff, Matthew J

2003-03-01

To estimate the sensitivity to find small coronary artery calcium lesions with use of different slice widths with electron beam tomography. Two studies were performed. Study 1 utilized double scanning of a stationary cork phantom with three different slice thickness (1.5, 3, and 6 mm). Fifty different calcific lesions (all <20 mm2 in area) fitted in 10 cork coronary arteries were utilized. The calcium foci area, peak value and score were measured and compared. In group 2, 30 patients underwent coronary artery calcium (CAC) screen studies. Each patient was scanned with both 3-mm and 6-mm scan widths in a same study time. Lesions with < 20 mm2 of area of CAC were measured on both 3-mm and 6-mm images. The mean and peak Hounsfield unit measure, and Agatston score were compared between both images. In the cork study, the sensitivity to detect small calcium foci were 96% (48/50), 82% (41/50), and 34% (17/50) in images with 1.5-, 3-, and 6-mm slice thickness, respectively. There is a smaller value in mass, and calcium volume in 6-mm images than 1.5-mm and 3-mm images ( P< 0.001). There was no significant difference between the true value and measured value from 1.5-mm and 3-mm images. In the human study, 18 (30%) of 60 CAC lesions with an area < 20 mm2 defined on 3 mm images were not visible on 6-mm images. Sensitivity of small lesions (P< 5 mm2) was 48% using 6-mm slices. There was a smaller value in CAC area, mean and peak Hounsfield units and score measured from 6-mm images, as compared with 3 mm slices ( P< 0.05). Thinner slice imaging has a higher sensitivity to detect small calcium focus. There was no significant change in score between 3 mm and 1.5 mm on the cork phantom study. However, the use of 6-mm slices should be discouraged, as this protocol both underestimates calcific mass and misses a significant number of calcific lesions in both a phantom and human study.

Combined system for high-time-resolution dual-excitation fluorescence photometry and fluorescence imaging of calcium transients in single normal and diseased skeletal muscle fibers

NASA Astrophysics Data System (ADS)

Uttenweiler, Dietmar; Wojciechowski, Reinhold; Makabe, Makoto; Veigel, Claudia; Fink, Rainer H.

1994-12-01

Fast photometric measurements and video-imaging of fluorescent indicators both are powerful tools in measuring the intracellular free calcium concentration of muscle and many other cells. as photometric systems yield a high temporal resolution, calcium imaging systems have high spatial but significantly reduced temporal resolution. Therefore we have developed an integrated system combining both methods and based mostly on standard components. As a common, sensitive Ca2+- indicator we used the fluorescent probe Fura-2, which is alternatingly excited for ratio measurements at 340/380 nm. We used a commercially available dual excitation photometric system (OSP-3; Olympus) for attaching a CCD-camera and a frame grabber board. To achieve the synchronization we had to design circuitries for external triggering, synchronization and accurate control of the filter changer, which we added to the system. Additionally, the software for a triggered image acquisition was developed. With this integrated setup one can easily switch between the fast photometric mode (ratio frequency 100 Hz) and the imaging mode (ratio frequency 4.17 Hz). The calcium images are correlated with the 25 times faster spot measurements and are analyzed by means of image processing. With this combined system we study release and uptake of calcium ions of normal and diseased skeletal muscle from mdx mice. Such a system will also be important for other cellular studies in which fluorescence indicators are used to monitor similar time dependent alterations as well as changes in cellular distributions of calcium.

a Single-Exposure Dual-Energy Computed Radiography Technique for Improved Nodule Detection and Classification in Chest Imaging

NASA Astrophysics Data System (ADS)

Zink, Frank Edward

The detection and classification of pulmonary nodules is of great interest in chest radiography. Nodules are often indicative of primary cancer, and their detection is particularly important in asymptomatic patients. The ability to classify nodules as calcified or non-calcified is important because calcification is a positive indicator that the nodule is benign. Dual-energy methods offer the potential to improve both the detection and classification of nodules by allowing the formation of material-selective images. Tissue-selective images can improve detection by virtue of the elimination of obscuring rib structure. Bone -selective images are essentially calcium images, allowing classification of the nodule. A dual-energy technique is introduced which uses a computed radiography system to acquire dual-energy chest radiographs in a single-exposure. All aspects of the dual-energy technique are described, with particular emphasis on scatter-correction, beam-hardening correction, and noise-reduction algorithms. The adaptive noise-reduction algorithm employed improves material-selective signal-to-noise ratio by up to a factor of seven with minimal sacrifice in selectivity. A clinical comparison study is described, undertaken to compare the dual-energy technique to conventional chest radiography for the tasks of nodule detection and classification. Observer performance data were collected using the Free Response Observer Characteristic (FROC) method and the bi-normal Alternative FROC (AFROC) performance model. Results of the comparison study, analyzed using two common multiple observer statistical models, showed that the dual-energy technique was superior to conventional chest radiography for detection of nodules at a statistically significant level (p < .05). Discussion of the comparison study emphasizes the unique combination of data collection and analysis techniques employed, as well as the limitations of comparison techniques in the larger context of technology assessment.

Using MRI to detect and differentiate calcium oxalate and calcium hydroxyapatite crystals in air-bubble-free phantom

PubMed Central

Mustafi, Devkumar; Fan, Xiaobing; Peng, Bo; Foxley, Sean; Palgen, Jeremy; Newstead, Gillian M.

2015-01-01

Calcium oxalate (CaOX) crystals and calcium hydroxyapatite (CaHA) crystals were commonly associated with breast benign and malignant lesions, respectively. In this research, CaOX (n = 6) and CaHA (n = 6) crystals in air-bubble-free agarose phantom were studied and characterized by using MRI at 9.4 Tesla scanner. Calcium micro-crystals sizes ranged from 200 – 500 microns were made with either 99% pure CaOX or CaHA powder and embedded in agar to mimic the dimensions and calcium content of breast microcalcifications in vivo. MRI data were acquired with high spatial resolution T2-weighted (T2W) images and gradient echo images with five different echo times (TEs). The crystals areas were determined by setting the threshold relative to agarose signal. The ratio of crystals areas were calculated by the measurements from gradient echo images divided by T2W images. Then the ratios as a function of TE were fitted with the radical function. The results showed that the blooming artifacts due to magnetic susceptibility between agar and CaHA crystals were more than twice as large as the susceptibility in CaOX crystals (p < 0.05). In addition, larger bright rings were observed on gradient echo images around CaHA crystals compared to CaOX crystals. Our results suggest that MRI may provide useful information regarding breast microcalcifications by evaluating the apparent area of crystals ratios obtained between gradient echo and T2W images. PMID:26392170

In vivo Post-Cardiac Arrest Myocardial Dysfunction is Supported by CaMKII-Mediated Calcium Long-Term Potentiation and Mitigated by Alda-1, an Agonist of Aldehyde Dehydrogenase Type 2

PubMed Central

Downey, Peter; Zalewski, Adrian; Rubio, Gabriel R.; Liu, Jing; Homburger, Julian R.; Grunwald, Zachary; Qi, Wei; Bollensdorff, Christian; Thanaporn, Porama; Ali, Ayyaz; Riemer, Kirk; Kohl, Peter; Mochly-Rosen, Daria; Gerstenfeld, Edward; Large, Stephen; Ali, Ziad; Ashley, Euan

2016-01-01

Background Survival after sudden cardiac arrest is limited by post-arrest myocardial dysfunction but understanding of this phenomenon is constrained by lack of data from a physiological model of disease. In this study, we established an in vivo model of cardiac arrest and resuscitation, characterized the biology of the associated myocardial dysfunction, and tested novel therapeutic strategies. Methods We developed rodent models of in vivo post-arrest myocardial dysfunction using extra-corporeal membrane oxygenation (ECMO) resuscitation followed by invasive hemodynamics measurement. In post-arrest isolated cardiomyocytes, we assessed mechanical load and Ca2+ induced Ca2+ release (CICR) simultaneously using the micro-carbon-fiber technique and observed reduced function and myofilament calcium sensitivity. We used a novel-designed fiber optic catheter imaging system, and a genetically encoded calcium sensor GCaMP6f, to image CICR in vivo. Results We found potentiation of CICR in isolated cells from this ECMO model and also in cells isolated from an ischemia-reperfusion Langendorff model perfused with oxygenated blood from an arrested animal, but not when reperfused in saline. We established that CICR potentiation begins in vivo. The augmented CICR observed post-arrest was mediated by the activation of Ca2+/calmodulin kinase II (CaMKII). Increased phosphorylation of CaMKII, phospholamban and ryanodine receptor 2 (RyR2) was detected in the post-arrest period. Exogenous adrenergic activation in vivo recapitulated Ca2+ potentiation but was associated with lesser CaMKII activation. Since oxidative stress and aldehydic adduct formation were high post arrest, we tested a small molecule activator of aldehyde dehydrogenase type 2, Alda-1, which reduced oxidative stress, restored calcium and CaMKII homeostasis, and improved cardiac function and post-arrest outcome in vivo. Conclusions Cardiac arrest and reperfusion lead to CaMKII activation and calcium long-term potentiation which support cardiomyocyte contractility in the face of impaired post-ischemic myofilament calcium sensitivity. Alda-1 mitigates these effects, normalizes calcium cycling and improves outcome. PMID:27582424

Simultaneous optical and electrical recording of a single ion-channel.

PubMed

Ide, Toru; Takeuchi, Yuko; Aoki, Takaaki; Yanagida, Toshio

2002-10-01

In recent years, the single-molecule imaging technique has proven to be a valuable tool in solving many basic problems in biophysics. The technique used to measure single-molecule functions was initially developed to study electrophysiological properties of channel proteins. However, the technology to visualize single channels at work has not received as much attention. In this study, we have for the first time, simultaneously measured the optical and electrical properties of single-channel proteins. The large conductance calcium-activated potassium channel (BK-channel) labeled with fluorescent dye molecules was incorporated into a planar bilayer membrane and the fluorescent image captured with a total internal reflection fluorescence microscope simultaneously with single-channel current recording. This innovative technology will greatly advance the study of channel proteins as well as signal transduction processes that involve ion permeation processes.

The necessity of microscopy to characterize the optical properties of size-selected, nonspherical aerosol particles.

PubMed

Veghte, Daniel P; Freedman, Miriam A

2012-11-06

It is currently unknown whether mineral dust causes a net warming or cooling effect on the climate system. This uncertainty stems from the varied and evolving shape and composition of mineral dust, which leads to diverse interactions of dust with solar and terrestrial radiation. To investigate these interactions, we have used a cavity ring-down spectrometer to study the optical properties of size-selected calcium carbonate particles, a reactive component of mineral dust. The size selection of nonspherical particles like mineral dust can differ from spherical particles in the polydispersity of the population selected. To calculate the expected extinction cross sections, we use Mie scattering theory for monodisperse spherical particles and for spherical particles with the polydispersity observed in transmission electron microscopy images. Our results for calcium carbonate are compared to the well-studied system of ammonium sulfate. While ammonium sulfate extinction cross sections agree with Mie scattering theory for monodisperse spherical particles, the results for calcium carbonate deviate at large and small particle sizes. We find good agreement for both systems, however, between the calculations performed using the particle images and the cavity ring-down data, indicating that both ammonium sulfate and calcium carbonate can be treated as polydisperse spherical particles. Our results indicate that having an independent measure of polydispersity is essential for understanding the optical properties of nonspherical particles measured with cavity ring-down spectroscopy. Our combined spectroscopy and microscopy techniques demonstrate a novel method by which cavity ring-down spectroscopy can be extended for the study of more complex aerosol particles.

Molecular imaging of in vivo calcium ion expression in area postrema of total sleep deprived rats: Implications for cardiovascular regulation by TOF-SIMS analysis

NASA Astrophysics Data System (ADS)

Mai, Fu-Der; Chen, Li-You; Ling, Yong-Chien; Chen, Bo-Jung; Wu, Un-In; Chang, Hung-Ming

2010-05-01

Excessive calcium influx in chemosensitive neurons of area postrema (AP) is detrimental for sympathetic activation and participates in the disruption of cardiovascular activities. Since total sleep deprivation (TSD) is a stressful condition known to harm the cardiovascular function, the present study is aimed to determine whether the in vivo calcium expression in AP would significantly alter following TSD by the use of time-of-flight secondary ion mass spectrometry (TOF-SIMS) and calretinin (a specific calcium sensor protein in AP neurons) immunohistochemistry. The results indicated that in normal rats, the calcium intensity was estimated to be 0.5 × 10 5 at m/ z 40.08. However, following TSD, the intensity for calcium ions was greatly increased to 1.2 × 10 5. Molecular imaging revealed that after TSD, various strongly expressed calcium signals were distributed throughout AP with clear identified profiles instead of randomly scattered within this region in normal rats. Immunohistochemical staining corresponded well with ionic image in which a majority of calcium-enriched gathering co-localized with calretinin positive neurons. The functional significance of TSD-induced calcium augmentation was demonstrated by increased heart rate and mean arterial pressure, clinical markers for cardiovascular dysfunction. Considering AP-mediated sympathetic activation is important for cardiovascular regulation, exaggerated calcium influx in AP would render this neurocircuitry more vulnerable to over-excitation, which might serve as the underlying mechanism for the development of TSD-relevant cardiovascular deficiency.

Calcium imaging of neuronal activity in the most rostral parafacial respiratory group of the newborn rat.

PubMed

Onimaru, Hiroshi; Dutschmann, Mathias

2012-01-01

The parafacial respiratory group (pFRG) is thought to be involved in respiratory rhythm generation in neonates. This subgroup expresses the transcription factor, Phox2b, and contains intrinsically CO(2) sensitive neurons. Calcium imaging has been widely used for analysis of neuronal activity at the cellular and network level. In the present study, we applied calcium imaging to analyze neuronal activity of the most-rostral pFRG in an in vitro brainstem-spinal cord preparation from neonatal rats. We detected strong pre-inspiratory neuron activity in the most rostral pFRG, suggesting that significant numbers of pre-inspiratory neurons are localized in the ventrolateral medulla near the rostral end of the medulla. We show that usage of calcium imaging would be very useful for analysis of neuronal activity over different time scales, and discuss the advantages and disadvantages of this method.

Comparative detection of calcium fluctuations in single female sex cells of tobacco to distinguish calcium signals triggered by in vitro fertilization.

PubMed

Peng, Xiong-Bo; Sun, Meng-Xiang; Yang, Hong-Yuan

2009-08-01

Double fertilization is a key process of sexual reproduction in higher plants. The role of calcium in the activation of female sex cells through fertilization has recently received a great deal of attention. The establishment of a Ca(2+)-imaging technique for living, single, female sex cells is a difficult but necessary prerequisite for evaluating the role of Ca(2+) in the transduction of external stimuli, including the fusion with the sperm cell, to internal cellular processes. The present study describes the use of Fluo-3 for reporting the Ca(2+) signal in isolated, single, female sex cells, egg cells and central cells, of tobacco plants. A suitable loading protocol was optimized by loading the cells at pH 5.6 with 2 microM Fluo-3 for 30 min at 30 degrees C. Under these conditions, several key factors related to in vitro fertilization were also investigated in order to test their possible effects on the [Ca(2+)](cyt) of the female sex cells. The results indicated that the bovine serum albumin-fusion system was superior to the polyethlene glycol-fusion system for detecting calcium fluctuations in female sex cells during fertilization. The central cell was fertilized with the sperm cell in bovine serum albumin; however, no evident calcium dynamic was detected, implying that a transient calcium rise might be a specific signal for egg cell fertilization.

Volumetric Two-photon Imaging of Neurons Using Stereoscopy (vTwINS)

PubMed Central

Song, Alexander; Charles, Adam S.; Koay, Sue Ann; Gauthier, Jeff L.; Thiberge, Stephan Y.; Pillow, Jonathan W.; Tank, David W.

2017-01-01

Two-photon laser scanning microscopy of calcium dynamics using fluorescent indicators is a widely used imaging method for large scale recording of neural activity in vivo. Here we introduce volumetric Two-photon Imaging of Neurons using Stereoscopy (vTwINS), a volumetric calcium imaging method that employs an elongated, V-shaped point spread function to image a 3D brain volume. Single neurons project to spatially displaced “image pairs” in the resulting 2D image, and the separation distance between images is proportional to depth in the volume. To demix the fluorescence time series of individual neurons, we introduce a novel orthogonal matching pursuit algorithm that also infers source locations within the 3D volume. We illustrate vTwINS by imaging neural population activity in mouse primary visual cortex and hippocampus. Our results demonstrate that vTwINS provides an effective method for volumetric two-photon calcium imaging that increases the number of neurons recorded while maintaining a high frame-rate. PMID:28319111

Classification of calcium in intravascular OCT images for the purpose of intervention planning

NASA Astrophysics Data System (ADS)

Shalev, Ronny; Bezerra, Hiram G.; Ray, Soumya; Prabhu, David; Wilson, David L.

2016-03-01

The presence of extensive calcification is a primary concern when planning and implementing a vascular percutaneous intervention such as stenting. If the balloon does not expand, the interventionalist must blindly apply high balloon pressure, use an atherectomy device, or abort the procedure. As part of a project to determine the ability of Intravascular Optical Coherence Tomography (IVOCT) to aid intervention planning, we developed a method for automatic classification of calcium in coronary IVOCT images. We developed an approach where plaque texture is modeled by the joint probability distribution of a bank of filter responses where the filter bank was chosen to reflect the qualitative characteristics of the calcium. This distribution is represented by the frequency histogram of filter response cluster centers. The trained algorithm was evaluated on independent ex-vivo image data accurately labeled using registered 3D microscopic cryo-image data which was used as ground truth. In this study, regions for extraction of sub-images (SI's) were selected by experts to include calcium, fibrous, or lipid tissues. We manually optimized algorithm parameters such as choice of filter bank, size of the dictionary, etc. Splitting samples into training and testing data, we achieved 5-fold cross validation calcium classification with F1 score of 93.7+/-2.7% with recall of >=89% and a precision of >=97% in this scenario with admittedly selective data. The automated algorithm performed in close-to-real-time (2.6 seconds per frame) suggesting possible on-line use. This promising preliminary study indicates that computational IVOCT might automatically identify calcium in IVOCT coronary artery images.

Development of a Radiolabeled Amlodipine Analog for L-type Calcium Channel Imaging.

PubMed

Firouzyar, Tahereh; Jalilian, Amir Reza; Aboudzadeh, Mohammad Reza; Sadeghpour, Hossein; Pooladi, Mehrban; Shafiee-Ardestani, Mahdi; Khalaj, Ali

2017-01-01

The non-invasive imaging and quantification of L-type calcium channels (also known as dihydropyridine channels) in living tissues is of great interest in diagnosis of congestive heart failure, myocardial hypertrophy, irritable bowel syndrome etc. Technetium-99m labeled amlodipine conjugate ([99mTc]-DTPA-AMLO) was prepared starting freshly eluted (<1 h) 99mTechnetium pertechnetate (86.5 MBq) and conjugated DTPAAMLO at pH 5 in 30 min at room temperature in high radiochemical purity (>99%, RTLC; specific activity: 55-60 GBq/mmol). The calcium channel blockade activity (CCBA) and apoptosis/necrosis assay of DTPA-amlodipine conjugate evaluations were performed for the conjugate. Log P, stability, bio-distribution and imaging studies were performed for the tracer followed by biodistribution studies as well as imaging. The conjugate demonstrated low toxicity on MCF-7 cells and CCBA (at µm level) compared to the amlodipine. The tracer was stable up to 4 h in final production and presence of human serum and log P (-0.49) was consistent with a water soluble complex. The tracer was excreted through kidneys and liver as expected for dihydropyridines; excluding excretory organs, calcium channel rich smooth muscle cells; including colon, intestine and lungs which demonstrated significant uptake. SPECT images supported the bio-distribution data up to 4 h. significant uptake of [99mTc]-DTPA-AMLO was obtained in calcium channel rich organs. The complex can be a candidate for further SPECT imaging for L-type calcium channels. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

First evidence on phloem transport of nanoscale calcium oxide in groundnut using solution culture technique

NASA Astrophysics Data System (ADS)

Deepa, Manchala; Sudhakar, Palagiri; Nagamadhuri, Kandula Venkata; Balakrishna Reddy, Kota; Giridhara Krishna, Thimmavajjula; Prasad, Tollamadugu Naga Venkata Krishna Vara

2015-06-01

Nanoscale materials, whose size typically falls below 100 nm, exhibit novel chemical, physical and biological properties which are different from their bulk counterparts. In the present investigation, we demonstrated that nanoscale calcium oxide particles (n-CaO) could transport through phloem tissue of groundnut unlike the corresponding bulk materials. n-CaO particles are prepared using sol-gel method. The size of the as prepared n-CaO measured (69.9 nm) using transmission electron microscopic technique (TEM). Results of the hydroponics experiment using solution culture technique revealed that foliar application of n-CaO at different concentrations (10, 50, 100, 500, 1,000 ppm) on groundnut plants confirmed the entry of calcium into leaves and stems through phloem compared to bulk source of calcium sprayed (CaO and CaNO3). After spraying of n-CaO, calcium content in roots, shoots and leaves significantly increased. Based on visual scoring of calcium deficiency correction and calcium content in plant parts, we may establish the fact that nanoscale calcium oxide particles (size 69.9 nm) could move through phloem tissue in groundnut. This is the first report on phloem transport of nanoscale calcium oxide particles in plants and this result points to the use of nanoscale calcium oxide particles as calcium source to the plants through foliar application, agricultural crops in particular, as bulk calcium application through foliar nutrition is restricted due to its non-mobility in phloem.

Imaging of human differentiated 3D neural aggregates using light sheet fluorescence microscopy.

PubMed

Gualda, Emilio J; Simão, Daniel; Pinto, Catarina; Alves, Paula M; Brito, Catarina

2014-01-01

The development of three dimensional (3D) cell cultures represents a big step for the better understanding of cell behavior and disease in a more natural like environment, providing not only single but multiple cell type interactions in a complex 3D matrix, highly resembling physiological conditions. Light sheet fluorescence microscopy (LSFM) is becoming an excellent tool for fast imaging of such 3D biological structures. We demonstrate the potential of this technique for the imaging of human differentiated 3D neural aggregates in fixed and live samples, namely calcium imaging and cell death processes, showing the power of imaging modality compared with traditional microscopy. The combination of light sheet microscopy and 3D neural cultures will open the door to more challenging experiments involving drug testing at large scale as well as a better understanding of relevant biological processes in a more realistic environment.

Imaging of human differentiated 3D neural aggregates using light sheet fluorescence microscopy

PubMed Central

Gualda, Emilio J.; Simão, Daniel; Pinto, Catarina; Alves, Paula M.; Brito, Catarina

2014-01-01

The development of three dimensional (3D) cell cultures represents a big step for the better understanding of cell behavior and disease in a more natural like environment, providing not only single but multiple cell type interactions in a complex 3D matrix, highly resembling physiological conditions. Light sheet fluorescence microscopy (LSFM) is becoming an excellent tool for fast imaging of such 3D biological structures. We demonstrate the potential of this technique for the imaging of human differentiated 3D neural aggregates in fixed and live samples, namely calcium imaging and cell death processes, showing the power of imaging modality compared with traditional microscopy. The combination of light sheet microscopy and 3D neural cultures will open the door to more challenging experiments involving drug testing at large scale as well as a better understanding of relevant biological processes in a more realistic environment. PMID:25161607

Tomographic imaging of bone composition using coherently scattered x rays

NASA Astrophysics Data System (ADS)

Batchelar, Deidre L.; Dabrowski, W.; Cunningham, Ian A.

2000-04-01

Bone tissue consists primarily of calcium hydroxyapatite crystals (bone mineral) and collagen fibrils. Bone mineral density (BMD) is commonly used as an indicator of bone health. Techniques available at present for assessing bone health provide a measure of BMD, but do not provide information about the degree of mineralization of the bone tissue. This may be adequate for assessing diseases in which the collagen-mineral ratio remains constant, as assumed in osteoporosis, but is insufficient when the mineralization state is known to change, as in osteomalacia. No tool exists for the in situ examination of collagen and hydroxyapatite density distributions independently. Coherent-scatter computed tomography (CSCT) is a technique we are developing that produces images of the low- angle scatter properties of tissue. These depend on the molecular structure of the scatterer making it possible to produce material-specific maps of each component in a conglomerate. After corrections to compensate for exposure fluctuations, self-attenuation of scatter and the temporal response of the image intensifier, material-specific images of mineral, collagen, fat and water distributions are obtained. The gray-level in these images provides the volumetric density of each component independently.

A Low Affinity GCaMP3 Variant (GCaMPer) for Imaging the Endoplasmic Reticulum Calcium Store.

PubMed

Henderson, Mark J; Baldwin, Heather A; Werley, Christopher A; Boccardo, Stefano; Whitaker, Leslie R; Yan, Xiaokang; Holt, Graham T; Schreiter, Eric R; Looger, Loren L; Cohen, Adam E; Kim, Douglas S; Harvey, Brandon K

2015-01-01

Endoplasmic reticulum calcium homeostasis is critical for cellular functions and is disrupted in diverse pathologies including neurodegeneration and cardiovascular disease. Owing to the high concentration of calcium within the ER, studying this subcellular compartment requires tools that are optimized for these conditions. To develop a single-fluorophore genetically encoded calcium indicator for this organelle, we targeted a low affinity variant of GCaMP3 to the ER lumen (GCaMPer (10.19)). A set of viral vectors was constructed to express GCaMPer in human neuroblastoma cells, rat primary cortical neurons, and human induced pluripotent stem cell-derived cardiomyocytes. We observed dynamic changes in GCaMPer (10.19) fluorescence in response to pharmacologic manipulations of the ER calcium store. Additionally, periodic calcium efflux from the ER was observed during spontaneous beating of cardiomyocytes. GCaMPer (10.19) has utility in imaging ER calcium in living cells and providing insight into luminal calcium dynamics under physiologic and pathologic states.

Using an Adapted Microfluidic Olfactory Chip for the Imaging of Neuronal Activity in Response to Pheromones in Male C. Elegans Head Neurons

PubMed Central

Reilly, Douglas K.; Lawler, Daniel E.; Albrecht, Dirk R.; Srinivasan, Jagan

2017-01-01

The use of calcium indicators has greatly enhanced our understanding of neural dynamics and regulation. The nematode Caenorhabditis elegans, with its completely mapped nervous system and transparent anatomy, presents an ideal model for understanding real-time neural dynamics using calcium indicators. In combination with microfluidic technologies and experimental designs, calcium-imaging studies using these indicators are performed in both free-moving and trapped animals. However, most previous studies utilizing trapping devices, such as the olfactory chip described in Chronis et al., have devices designed for use in the more common hermaphrodite, as the less common male is both morphologically and structurally dissimilar. An adapted olfactory chip was designed and fabricated for increased efficiency in male neuronal imaging with using young adult animals. A turn was incorporated into the worm loading port to rotate the animals and to allow for the separation of the individual neurons within a bilateral pair in 2D imaging. Worms are exposed to a controlled flow of odorant within the microfluidic device, as described in previous hermaphrodite studies. Calcium transients are then analyzed using the open-source software ImageJ. The procedure described herein should allow for an increased amount of male-based C. elegans calcium imaging studies, deepening our understanding of the mechanisms of sex-specific neuronal signaling. PMID:28930991

Development of large field-of-view two photon microscopy for imaging mouse cortex (Conference Presentation)

NASA Astrophysics Data System (ADS)

Bumstead, Jonathan; Côté, Daniel C.; Culver, Joseph P.

2017-02-01

Spontaneous neuronal activity has been measured at cellular resolution in mice, zebrafish, and C. elegans using optical sectioning microscopy techniques, such as light sheet microscopy (LSM) and two photon microscopy (TPM). Recent improvements in these modalities and genetically encoded calcium indicators (GECI's) have enabled whole brain imaging of calcium dynamics in zebrafish and C. elegans. However, these whole brain microscopy studies have not been extended to mice due to the limited field of view (FOV) of TPM and the cumbersome geometry of LSM. Conventional TPM is restricted to diffraction limited imaging over this small FOV (around 500 x 500 microns) due to the use of high magnification objectives (e.g. 1.0 NA; 20X) and the aberrations introduced by relay optics used in scanning the beam across the sample. To overcome these limitations, we have redesigned the entire optical path of the two photon microscope (scanning optics and objective lens) to support a field of view of Ø7 mm with relatively high spatial resolution (<10 microns). Using optical engineering software Zemax, we designed our system with commercially available optics that minimize astigmatism, field curvature, chromatic focal shift, and vignetting. Performance of the system was also tested experimentally with fluorescent beads in agarose, fixed samples, and in vivo structural imaging. Our large-FOV TPM provides a modality capable of studying distributed brain networks in mice at cellular resolution.

Improvement of cardiac CT reconstruction using local motion vector fields.

PubMed

Schirra, Carsten Oliver; Bontus, Claas; van Stevendaal, Udo; Dössel, Olaf; Grass, Michael

2009-03-01

The motion of the heart is a major challenge for cardiac imaging using CT. A novel approach to decrease motion blur and to improve the signal to noise ratio is motion compensated reconstruction which takes motion vector fields into account in order to correct motion. The presented work deals with the determination of local motion vector fields from high contrast objects and their utilization within motion compensated filtered back projection reconstruction. Image registration is applied during the quiescent cardiac phases. Temporal interpolation in parameter space is used in order to estimate motion during strong motion phases. The resulting motion vector fields are during image reconstruction. The method is assessed using a software phantom and several clinical cases for calcium scoring. As a criterion for reconstruction quality, calcium volume scores were derived from both, gated cardiac reconstruction and motion compensated reconstruction throughout the cardiac phases using low pitch helical cone beam CT acquisitions. The presented technique is a robust method to determine and utilize local motion vector fields. Motion compensated reconstruction using the derived motion vector fields leads to superior image quality compared to gated reconstruction. As a result, the gating window can be enlarged significantly, resulting in increased SNR, while reliable Hounsfield units are achieved due to the reduced level of motion artefacts. The enlargement of the gating window can be translated into reduced dose requirements.

Multi-modal in vivo imaging of brain blood oxygenation, blood flow and neural calcium dynamics during acute seizures

NASA Astrophysics Data System (ADS)

Ringuette, Dene; Jeffrey, Melanie A.; Carlen, Peter L.; Levi, Ofer

2016-03-01

Dysfunction of the vascular endothelium has been implicated in the development of epilepsy. To better understand the relation between vascular function and seizure and provide a foundation for interpreting results from functional imaging in chronic disease models, we investigate the relationship between intracellular calcium dynamics and local cerebral blood flow and blood oxygen saturation during acute seizure-like events and pharmacological seizure rescue. To probe the relation between the aforementioned physiological markers in an acute model of epilepsy in rats, we integrated three different optical modalities together with electrophysiological recordings: Laser speckle contrast imaging (LSCI) was used to study changes in flow speeds, Intrinsic optical signal imaging (IOSI) was used to monitor changes in oxygenated, de-oxygenated, and total hemoglobin concentration, and Calcium-sensitive dye imaging was used to monitor intracellular calcium dynamics. We designed a dedicated cortical flow chamber to remove superficial blood and dye resulting from the injection procedure, which reduced spurious artifacts. The near infrared light used for IOSI and LSCI was delivered via a light pipe integrated with the flow chamber to minimize the effect of fluid surface movement on illumination stability. Calcium-sensitive dye was injected via a glass electrode used for recording the local field potential. Our system allowed us to observe and correlate increases in intracellular calcium, blood flow and blood volume during seizure-like events and provide a quantitative analysis of neurovascular coupling changes associated with seizure rescue via injection of an anti-convulsive agent.

Fast calcium and voltage-sensitive dye imaging in enteric neurones reveal calcium peaks associated with single action potential discharge.

PubMed

Michel, K; Michaelis, M; Mazzuoli, G; Mueller, K; Vanden Berghe, P; Schemann, M

2011-12-15

Slow changes in [Ca(2+)](i) reflect increased neuronal activity. Our study demonstrates that single-trial fast [Ca(2+)](i) imaging (≥200 Hz sampling rate) revealed peaks each of which are associated with single spike discharge recorded by consecutive voltage-sensitive dye (VSD) imaging in enteric neurones and nerve fibres. Fast [Ca(2+)](i) imaging also revealed subthreshold fast excitatory postsynaptic potentials. Nicotine-evoked [Ca(2+)](i) peaks were reduced by -conotoxin and blocked by ruthenium red or tetrodotoxin. Fast [Ca(2+)](i) imaging can be used to directly record single action potentials in enteric neurones. [Ca(2+)](i) peaks required opening of voltage-gated sodium and calcium channels as well as Ca(2+) release from intracellular stores.

Development of a thresholding algorithm for calcium classification at multiple CT energies

NASA Astrophysics Data System (ADS)

Ng, LY.; Alssabbagh, M.; Tajuddin, A. A.; Shuaib, I. L.; Zainon, R.

2017-05-01

The objective of this study was to develop a thresholding method for calcium classification with different concentration using single-energy computed tomography. Five different concentrations of calcium chloride were filled in PMMA tubes and placed inside a water-filled PMMA phantom (diameter 10 cm). The phantom was scanned at 70, 80, 100, 120 and 140 kV using a SECT. CARE DOSE 4D was used and the slice thickness was set to 1 mm for all energies. ImageJ software inspired by the National Institute of Health (NIH) was used to measure the CT numbers for each calcium concentration from the CT images. The results were compared with a developed algorithm for verification. The percentage differences between the measured CT numbers obtained from the developed algorithm and the ImageJ show similar results. The multi-thresholding algorithm was found to be able to distinguish different concentrations of calcium chloride. However, it was unable to detect low concentrations of calcium chloride and iron (III) nitrate with CT numbers between 25 HU and 65 HU. The developed thresholding method used in this study may help to differentiate between calcium plaques and other types of plaques in blood vessels as it is proven to have a good ability to detect the high concentration of the calcium chloride. However, the algorithm needs to be improved to solve the limitations of detecting calcium chloride solution which has a similar CT number with iron (III) nitrate solution.

Direct Imaging of ER Calcium with Targeted-Esterase Induced Dye Loading (TED)

PubMed Central

Samtleben, Samira; Jaepel, Juliane; Fecher, Caroline; Andreska, Thomas; Rehberg, Markus; Blum, Robert

2013-01-01

Visualization of calcium dynamics is important to understand the role of calcium in cell physiology. To examine calcium dynamics, synthetic fluorescent Ca2+ indictors have become popular. Here we demonstrate TED (= targeted-esterase induced dye loading), a method to improve the release of Ca2+ indicator dyes in the ER lumen of different cell types. To date, TED was used in cell lines, glial cells, and neurons in vitro. TED bases on efficient, recombinant targeting of a high carboxylesterase activity to the ER lumen using vector-constructs that express Carboxylesterases (CES). The latest TED vectors contain a core element of CES2 fused to a red fluorescent protein, thus enabling simultaneous two-color imaging. The dynamics of free calcium in the ER are imaged in one color, while the corresponding ER structure appears in red. At the beginning of the procedure, cells are transduced with a lentivirus. Subsequently, the infected cells are seeded on coverslips to finally enable live cell imaging. Then, living cells are incubated with the acetoxymethyl ester (AM-ester) form of low-affinity Ca2+ indicators, for instance Fluo5N-AM, Mag-Fluo4-AM, or Mag-Fura2-AM. The esterase activity in the ER cleaves off hydrophobic side chains from the AM form of the Ca2+ indicator and a hydrophilic fluorescent dye/Ca2+ complex is formed and trapped in the ER lumen. After dye loading, the cells are analyzed at an inverted confocal laser scanning microscope. Cells are continuously perfused with Ringer-like solutions and the ER calcium dynamics are directly visualized by time-lapse imaging. Calcium release from the ER is identified by a decrease in fluorescence intensity in regions of interest, whereas the refilling of the ER calcium store produces an increase in fluorescence intensity. Finally, the change in fluorescent intensity over time is determined by calculation of ΔF/F0. PMID:23685703

Direct imaging of ER calcium with targeted-esterase induced dye loading (TED).

PubMed

Samtleben, Samira; Jaepel, Juliane; Fecher, Caroline; Andreska, Thomas; Rehberg, Markus; Blum, Robert

2013-05-07

Visualization of calcium dynamics is important to understand the role of calcium in cell physiology. To examine calcium dynamics, synthetic fluorescent Ca(2+) indictors have become popular. Here we demonstrate TED (= targeted-esterase induced dye loading), a method to improve the release of Ca(2+) indicator dyes in the ER lumen of different cell types. To date, TED was used in cell lines, glial cells, and neurons in vitro. TED bases on efficient, recombinant targeting of a high carboxylesterase activity to the ER lumen using vector-constructs that express Carboxylesterases (CES). The latest TED vectors contain a core element of CES2 fused to a red fluorescent protein, thus enabling simultaneous two-color imaging. The dynamics of free calcium in the ER are imaged in one color, while the corresponding ER structure appears in red. At the beginning of the procedure, cells are transduced with a lentivirus. Subsequently, the infected cells are seeded on coverslips to finally enable live cell imaging. Then, living cells are incubated with the acetoxymethyl ester (AM-ester) form of low-affinity Ca(2+) indicators, for instance Fluo5N-AM, Mag-Fluo4-AM, or Mag-Fura2-AM. The esterase activity in the ER cleaves off hydrophobic side chains from the AM form of the Ca(2+) indicator and a hydrophilic fluorescent dye/Ca(2+) complex is formed and trapped in the ER lumen. After dye loading, the cells are analyzed at an inverted confocal laser scanning microscope. Cells are continuously perfused with Ringer-like solutions and the ER calcium dynamics are directly visualized by time-lapse imaging. Calcium release from the ER is identified by a decrease in fluorescence intensity in regions of interest, whereas the refilling of the ER calcium store produces an increase in fluorescence intensity. Finally, the change in fluorescent intensity over time is determined by calculation of ΔF/F0.

Two-photon imaging in living brain slices.

PubMed

Mainen, Z F; Maletic-Savatic, M; Shi, S H; Hayashi, Y; Malinow, R; Svoboda, K

1999-06-01

Two-photon excitation laser scanning microscopy (TPLSM) has become the tool of choice for high-resolution fluorescence imaging in intact neural tissues. Compared with other optical techniques, TPLSM allows high-resolution imaging and efficient detection of fluorescence signal with minimal photobleaching and phototoxicity. The advantages of TPLSM are especially pronounced in highly scattering environments such as the brain slice. Here we describe our approaches to imaging various aspects of synaptic function in living brain slices. To combine several imaging modes together with patch-clamp electrophysiological recordings we found it advantageous to custom-build an upright microscope. Our design goals were primarily experimental convenience and efficient collection of fluorescence. We describe our TPLSM imaging system and its performance in detail. We present dynamic measurements of neuronal morphology of neurons expressing green fluorescent protein (GFP) and GFP fusion proteins as well as functional imaging of calcium dynamics in individual dendritic spines. Although our microscope is a custom instrument, its key advantages can be easily implemented as a modification of commercial laser scanning microscopes. Copyright 1999 Academic Press.

Calcium pump kinetics determined in single erythrocyte ghosts by microphotolysis and confocal imaging.

PubMed

Kubitscheck, U; Pratsch, L; Passow, H; Peters, R

1995-07-01

The activity of the plasma membrane calcium pump was measured in single cells. Human red blood cell ghosts were loaded with a fluorescent calcium indicator and either caged calcium and ATP (protocol A) or caged ATP and calcium (protocol B). In a suitably modified laser scanning microscope either calcium or ATP were released by a short UV light pulse. The time-dependent fluorescence intensity of the calcium indicator was then followed in single ghosts by repetitive confocal imaging. The fluorescence intensity was converted into calcium concentration, which in turn was used to derive the kinetic parameters of the calcium pump, the Michaelis-Menten constant Km, and the maximal transport rate vmax. Km and vmax values derived in this manner were 24 +/- 14 microM and 1.0 +/- 0.6 microM/(ghost s) for protocol A, and 4 +/- 3 microM and 1.0 +/- 0.6 microM/(ghost s) for protocol B, respectively. The difference between A and B is presumably caused by calmodulin, which is inactive in the experiments with protocol A. The possibilities to extend the new method to living nucleus-containing cells transiently transfected with mutants of the plasma membrane calcium pump are discussed.

Inference of neuronal network spike dynamics and topology from calcium imaging data

PubMed Central

Lütcke, Henry; Gerhard, Felipe; Zenke, Friedemann; Gerstner, Wulfram; Helmchen, Fritjof

2013-01-01

Two-photon calcium imaging enables functional analysis of neuronal circuits by inferring action potential (AP) occurrence (“spike trains”) from cellular fluorescence signals. It remains unclear how experimental parameters such as signal-to-noise ratio (SNR) and acquisition rate affect spike inference and whether additional information about network structure can be extracted. Here we present a simulation framework for quantitatively assessing how well spike dynamics and network topology can be inferred from noisy calcium imaging data. For simulated AP-evoked calcium transients in neocortical pyramidal cells, we analyzed the quality of spike inference as a function of SNR and data acquisition rate using a recently introduced peeling algorithm. Given experimentally attainable values of SNR and acquisition rate, neural spike trains could be reconstructed accurately and with up to millisecond precision. We then applied statistical neuronal network models to explore how remaining uncertainties in spike inference affect estimates of network connectivity and topological features of network organization. We define the experimental conditions suitable for inferring whether the network has a scale-free structure and determine how well hub neurons can be identified. Our findings provide a benchmark for future calcium imaging studies that aim to reliably infer neuronal network properties. PMID:24399936

Dual-Energy Computed Tomography for the Characterization of Intracranial Hemorrhage and Calcification: A Systematic Approach in a Phantom System.

PubMed

Nute, Jessica L; Jacobsen, Megan C; Chandler, Adam; Cody, Dianna D; Schellingerhout, Dawid

2017-01-01

The aim of this study was to develop a diagnostic framework for distinguishing calcific from hemorrhagic cerebral lesions using dual-energy computed tomography (DECT) in an anthropomorphic phantom system. An anthropomorphic phantom was designed to mimic the CT imaging characteristics of the human head. Cylindrical lesion models containing either calcium or iron, mimicking calcification or hemorrhage, respectively, were developed to exhibit matching, and therefore indistinguishable, single-energy CT (SECT) attenuation values from 40 to 100 HU. These lesion models were fabricated at 0.5, 1, and 1.5 cm in diameter and positioned in simulated cerebrum and skull base locations within the anthropomorphic phantom. All lesion sizes were modeled in the cerebrum, while only 1.5-cm lesions were modeled in the skull base. Images were acquired using a GE 750HD CT scanner and an expansive dual-energy protocol that covered variations in dose (36.7-132.6 mGy CTDIvol, n = 12), image thickness (0.625-5 mm, n = 4), and reconstruction filter (soft, standard, detail, n = 3) for a total of 144 unique technique combinations. Images representing each technique combination were reconstructed into water and calcium material density images, as well as a monoenergetic image chosen to mimic the attenuation of a 120-kVp SECT scan. A true single-energy routine brain protocol was also included for verification of lesion SECT attenuation. Points representing the 3 dual-energy reconstructions were plotted into a 3-dimensional space (water [milligram/milliliter], calcium [milligram/milliliter], monoenergetic Hounsfield unit as x, y, and z axes, respectively), and the distribution of points analyzed using 2 approaches: support vector machines and a simple geometric bisector (GB). Each analysis yielded a plane of optimal differentiation between the calcification and hemorrhage lesion model distributions. By comparing the predicted lesion composition to the known lesion composition, we identified the optimal combination of CTDIvol, image thickness, and reconstruction filter to maximize differentiation between the lesion model types. To validate these results, a new set of hemorrhage and calcification lesion models were created, scanned in a blinded fashion, and prospectively classified using the planes of differentiation derived from support vector machine and GB methods. Accuracy of differentiation improved with increasing dose (CTDIvol) and image thickness. Reconstruction filter had no effect on the accuracy of differentiation. Using an optimized protocol consisting of the maximum CTDIvol of 132.6 mGy, 5-mm-thick images, and a standard filter, hemorrhagic and calcific lesion models with equal SECT attenuation (Hounsfield unit) were differentiated with over 90% accuracy down to 70 HU for skull base lesions of 1.5 cm, and down to 100 HU, 60 HU, and 60 HU for cerebrum lesions of 0.5, 1.0, and 1.5 cm, respectively. The analytic method that yielded the best results was a simple GB plane through the 3-dimensional DECT space. In the validation study, 96% of unknown lesions were correctly classified across all lesion sizes and locations investigated. We define the optimal scan parameters and expected limitations for the accurate classification of hemorrhagic versus calcific cerebral lesions in an anthropomorphic phantom with DECT. Although our proposed DECT protocol represents an increase in dose compared with routine brain CT, this method is intended as a specialized evaluation of potential brain hemorrhage and is thus counterbalanced by increased diagnostic benefit. This work provides justification for the application of this technique in human clinical trials.

Variations in Urine Calcium Isotope: Composition Reflect Changes in Bone Mineral Balance in Humans

NASA Technical Reports Server (NTRS)

Skulan, Joseph; Anbar, Ariel; Bullen, Thomas; Puzas, J. Edward; Shackelford, Linda; Smith, Scott M.

2004-01-01

Changes in bone mineral balance cause rapid and systematic changes in the calcium isotope composition of human urine. Urine from subjects in a 17 week bed rest study was analyzed for calcium isotopic composition. Comparison of isotopic data with measurements of bone mineral density and metabolic markers of bone metabolism indicates the calcium isotope composition of urine reflects changes in bone mineral balance. Urine calcium isotope composition probably is affected by both bone metabolism and renal processes. Calcium isotope. analysis of urine and other tissues may provide information on bone mineral balance that is in important respects better than that available from other techniques, and illustrates the usefulness of applying geochemical techniques to biomedical problems.

Detection of Osteopontin in the pericyst of human hepatic Echinococcus granulosus.

PubMed

Peng, Xinyu; Li, Jianhui; Wu, Xiangwei; Zhang, Shijie; Niu, Jianhua; Chen, Xiaoping; Yao, Jin; Sun, Hong

2006-12-01

It aims at investigating the expression and distribution of the Osteopontin (OPN) in the pericyst of human hepatic Echinococcus granulosus and their related significances. Sixty pericysts excised by "sub-adventitial cystectomy" were studied. OPN was detected in 80% (48/60) of cysts by Western blotting and distributed in the side of "exocyst" layer directing to the parasite, also macrophages were identified in the vicinity of OPN by immunohistochemistry staining. The coexpression of OPN and CD68 was observed by immunofluorescence double labeling and analyzed by Image-Pro Plus 5.1; with special stain techniques, variable degrees of calcium deposits were observed in 80% (48/60) cysts, and the calcium deposits concurrencely found with the OPN expression. The selective distribution of OPN, calcium in the "exocyst" provides a new pathological evidence for the "sub-adventitial cystectomy" we developed. The pericyst of hepatic E. granulosus consists of two detachable layers with different formative mechanisms: the "exocyst" layer directing towards the cyst of parasite was the result of granulomatous reaction; also the results suggest OPN is one regulator in the granulomatous reaction and calcification of "exocyst".

Combining microfluidics, optogenetics and calcium imaging to study neuronal communication in vitro.

PubMed

Renault, Renaud; Sukenik, Nirit; Descroix, Stéphanie; Malaquin, Laurent; Viovy, Jean-Louis; Peyrin, Jean-Michel; Bottani, Samuel; Monceau, Pascal; Moses, Elisha; Vignes, Maéva

2015-01-01

In this paper we report the combination of microfluidics, optogenetics and calcium imaging as a cheap and convenient platform to study synaptic communication between neuronal populations in vitro. We first show that Calcium Orange indicator is compatible in vitro with a commonly used Channelrhodopsine-2 (ChR2) variant, as standard calcium imaging conditions did not alter significantly the activity of transduced cultures of rodent primary neurons. A fast, robust and scalable process for micro-chip fabrication was developed in parallel to build micro-compartmented cultures. Coupling optical fibers to each micro-compartment allowed for the independent control of ChR2 activation in the different populations without crosstalk. By analyzing the post-stimuli activity across the different populations, we finally show how this platform can be used to evaluate quantitatively the effective connectivity between connected neuronal populations.

Dual energy x-ray imaging and scoring of coronary calcium: physics-based digital phantom and clinical studies

NASA Astrophysics Data System (ADS)

Zhou, Bo; Wen, Di; Nye, Katelyn; Gilkeson, Robert C.; Wilson, David L.

2016-03-01

Coronary artery calcification (CAC) as assessed with CT calcium score is the best biomarker of coronary artery disease. Dual energy x-ray provides an inexpensive, low radiation-dose alternative. A two shot system (GE Revolution-XRd) is used, raw images are processed with a custom algorithm, and a coronary calcium image (DECCI) is created, similar to the bone image, but optimized for CAC visualization, not lung visualization. In this report, we developed a physicsbased, digital-phantom containing heart, lung, CAC, spine, ribs, pulmonary artery, and adipose elements, examined effects on DECCI, suggested physics-inspired algorithms to improve CAC contrast, and evaluated the correlation between CT calcium scores and a proposed DE calcium score. In simulation experiment, Beam hardening from increasing adipose thickness (2cm to 8cm) reduced Cg by 19% and 27% in 120kVp and 60kVp images, but only reduced Cg by <7% in DECCI. If a pulmonary artery moves or pulsates with blood filling between exposures, it can give rise to a significantly confounding PA signal in DECCI similar in amplitude to CAC. Observations suggest modifications to DECCI processing, which can further improve CAC contrast by a factor of 2 in clinical exams. The DE score had the best correlation with "CT mass score" among three commonly used CT scores. Results suggest that DE x-ray is a promising tool for imaging and scoring CAC, and there still remains opportunity for further DECCI processing improvements.

Selective laser sintering of calcium phosphate materials for orthopedic implants

NASA Astrophysics Data System (ADS)

Lee, Goonhee

Two technologies, Solid Freeform Fabrication (SFF) and bioceramics are combined in this work to prepare bone replacement implants with complex geometry. SFF has emerged as a crucial technique for rapid prototyping in the last decade. Selective Laser Sintering (SLS) is one of the established SFF manufacturing processes that can build three-dimensional objects directly from computer models without part-specific tooling or human intervention. Meanwhile, there have been great efforts to develop implantable materials that can assist in regeneration of bone defects and injuries. However, little attention has been focused in shaping bones from these materials. The main thrust of this research was to develop a process that can combine those two separate efforts. The specific objective of this research is to develop a process that can construct bone replacement material of complex geometry from synthetic calcium phosphate materials by using the SLS process. The achievement of this goal can have a significant impact on the quality of health care in the sense that complete custom-fit bone and tooth structures suitable for implantation can be prepared within 24--48 hours of receipt of geometric information obtained either from patient Computed Tomographic (CT) data, from Computer Aided Design (CAD) software or from other imaging systems such as Magnetic Resonance Imaging (MRI) and Holographic Laser Range Imaging (HLRI). In this research, two different processes have been developed. First is the SLS fabrication of porous bone implants. In this effort, systematic procedures have been established and calcium phosphate implants were successfully fabricated from various sources of geometric information. These efforts include material selection and preparation, SLS process parameter optimization, and development of post-processing techniques within the 48-hour time frame. Post-processing allows accurate control of geometry and of the chemistry of calcium phosphate, as well as control of micro and macro pore structure, to maximize bone healing and provide sufficient mechanical strength. It also permits the complete removal of the polymeric binders that are resided in the SLS process. In collaboration with the University of Texas Health Science Center at San Antonio and BioMedical Enterprises, Inc., porous implants based on anatomical geometry have been successfully implanted in rabbits and dogs. These histologic animal studies reveal excellent biocompatibility and show its great potential for commercial custom-fit implant manufacture. The second research effort involves fabrication of fully dense bone for application in dental restoration and load-bearing orthopedic functions. Calcium phosphate glass melts, proven to be biocompatible in the first effort, were cast into carbon molds. Processes were developed for preparing the molds. These carbon molds of anatomic shape can be prepared from either Computer Numerical Control (CNC) milling of slab stock or SLS processing of thermoset-coated graphite powder. The CNC milling method provides accurate dimension of the molds in a short period of time, however, the capable geometries are limited; generally two pieces of molds are required for complex shapes. The SLS method provides very complex shape green molds. However, they need to go through pyrolysis of thermoset binder to provide the high temperature capability reached at calcium phosphate melt temperatures (1100°C) and noticeable shrinkage was observed during pyrolysis. The cast glass was annealed to develop polycrystalline calcium phosphate. This process also exhibits great potential.

Functional connectivity structure of cortical calcium dynamics in anesthetized and awake mice.

PubMed

Wright, Patrick W; Brier, Lindsey M; Bauer, Adam Q; Baxter, Grant A; Kraft, Andrew W; Reisman, Matthew D; Bice, Annie R; Snyder, Abraham Z; Lee, Jin-Moo; Culver, Joseph P

2017-01-01

The interplay between hemodynamic-based markers of cortical activity (e.g. fMRI and optical intrinsic signal imaging), which are an indirect and relatively slow report of neural activity, and underlying synaptic electrical and metabolic activity through neurovascular coupling is a topic of ongoing research and debate. As application of resting state functional connectivity measures is extended further into topics such as brain development, aging and disease, the importance of understanding the fundamental physiological basis for functional connectivity will grow. Here we extend functional connectivity analysis from hemodynamic- to calcium-based imaging. Transgenic mice (n = 7) expressing a fluorescent calcium indicator (GCaMP6) driven by the Thy1 promoter in glutamatergic neurons were imaged transcranially in both anesthetized (using ketamine/xylazine) and awake states. Sequential LED illumination (λ = 454, 523, 595, 640nm) enabled concurrent imaging of both GCaMP6 fluorescence emission (corrected for hemoglobin absorption) and hemodynamics. Functional connectivity network maps were constructed for infraslow (0.009-0.08Hz), intermediate (0.08-0.4Hz), and high (0.4-4.0Hz) frequency bands. At infraslow and intermediate frequencies, commonly used in BOLD fMRI and fcOIS studies of functional connectivity and implicated in neurovascular coupling mechanisms, GCaMP6 and HbO2 functional connectivity structures were in high agreement, both qualitatively and also quantitatively through a measure of spatial similarity. The spontaneous dynamics of both contrasts had the highest correlation when the GCaMP6 signal was delayed with a ~0.6-1.5s temporal offset. Within the higher-frequency delta band, sensitive to slow wave sleep oscillations in non-REM sleep and anesthesia, we evaluate the speed with which the connectivity analysis stabilized and found that the functional connectivity maps captured putative network structure within time window lengths as short as 30 seconds. Homotopic GCaMP6 functional connectivity maps at 0.4-4.0Hz in the anesthetized states show a striking correlated and anti-correlated structure along the anterior to posterior axis. This structure is potentially explained in part by observed propagation of delta-band activity from frontal somatomotor regions to visuoparietal areas. During awake imaging, this spatio-temporal quality is altered, and a more complex and detailed functional connectivity structure is observed. The combined calcium/hemoglobin imaging technique described here will enable the dissociation of changes in ionic and hemodynamic functional structure and neurovascular coupling and provide a framework for subsequent studies of neurological disease such as stroke.

Functional connectivity structure of cortical calcium dynamics in anesthetized and awake mice

PubMed Central

Wright, Patrick W.; Brier, Lindsey M.; Bauer, Adam Q.; Baxter, Grant A.; Kraft, Andrew W.; Reisman, Matthew D.; Bice, Annie R.; Snyder, Abraham Z.; Lee, Jin-Moo; Culver, Joseph P.

2017-01-01

The interplay between hemodynamic-based markers of cortical activity (e.g. fMRI and optical intrinsic signal imaging), which are an indirect and relatively slow report of neural activity, and underlying synaptic electrical and metabolic activity through neurovascular coupling is a topic of ongoing research and debate. As application of resting state functional connectivity measures is extended further into topics such as brain development, aging and disease, the importance of understanding the fundamental physiological basis for functional connectivity will grow. Here we extend functional connectivity analysis from hemodynamic- to calcium-based imaging. Transgenic mice (n = 7) expressing a fluorescent calcium indicator (GCaMP6) driven by the Thy1 promoter in glutamatergic neurons were imaged transcranially in both anesthetized (using ketamine/xylazine) and awake states. Sequential LED illumination (λ = 454, 523, 595, 640nm) enabled concurrent imaging of both GCaMP6 fluorescence emission (corrected for hemoglobin absorption) and hemodynamics. Functional connectivity network maps were constructed for infraslow (0.009–0.08Hz), intermediate (0.08–0.4Hz), and high (0.4–4.0Hz) frequency bands. At infraslow and intermediate frequencies, commonly used in BOLD fMRI and fcOIS studies of functional connectivity and implicated in neurovascular coupling mechanisms, GCaMP6 and HbO2 functional connectivity structures were in high agreement, both qualitatively and also quantitatively through a measure of spatial similarity. The spontaneous dynamics of both contrasts had the highest correlation when the GCaMP6 signal was delayed with a ~0.6–1.5s temporal offset. Within the higher-frequency delta band, sensitive to slow wave sleep oscillations in non-REM sleep and anesthesia, we evaluate the speed with which the connectivity analysis stabilized and found that the functional connectivity maps captured putative network structure within time window lengths as short as 30 seconds. Homotopic GCaMP6 functional connectivity maps at 0.4–4.0Hz in the anesthetized states show a striking correlated and anti-correlated structure along the anterior to posterior axis. This structure is potentially explained in part by observed propagation of delta-band activity from frontal somatomotor regions to visuoparietal areas. During awake imaging, this spatio-temporal quality is altered, and a more complex and detailed functional connectivity structure is observed. The combined calcium/hemoglobin imaging technique described here will enable the dissociation of changes in ionic and hemodynamic functional structure and neurovascular coupling and provide a framework for subsequent studies of neurological disease such as stroke. PMID:29049297

A highly sensitive fluorescent indicator dye for calcium imaging of neural activity in vitro and in vivo

PubMed Central

Tada, Mayumi; Takeuchi, Atsuya; Hashizume, Miki; Kitamura, Kazuo; Kano, Masanobu

2014-01-01

Calcium imaging of individual neurons is widely used for monitoring their activity in vitro and in vivo. Synthetic fluorescent calcium indicator dyes are commonly used, but the resulting calcium signals sometimes suffer from a low signal-to-noise ratio (SNR). Therefore, it is difficult to detect signals caused by single action potentials (APs) particularly from neurons in vivo. Here we showed that a recently developed calcium indicator dye, Cal-520, is sufficiently sensitive to reliably detect single APs both in vitro and in vivo. In neocortical neurons, calcium signals were linearly correlated with the number of APs, and the SNR was > 6 for in vitro slice preparations and > 1.6 for in vivo anesthetised mice. In cerebellar Purkinje cells, dendritic calcium transients evoked by climbing fiber inputs were clearly observed in anesthetised mice with a high SNR and fast decay time. These characteristics of Cal-520 are a great advantage over those of Oregon Green BAPTA-1, the most commonly used calcium indicator dye, for monitoring the activity of individual neurons both in vitro and in vivo. PMID:24405482

Effect of extracellular ATP on contraction, cytosolic calcium activity, membrane voltage and ion currents of rat mesangial cells in primary culture.

PubMed Central

Pavenstädt, H.; Gloy, J.; Leipziger, J.; Klär, B.; Pfeilschifter, J.; Schollmeyer, P.; Greger, R.

1993-01-01

1. The effects of extracellular ATP on contraction, membrane voltage (Vm), ion currents and intracellular calcium activity [Ca2+]i were studied in rat mesangial cells (MC) in primary culture. 2. Addition of extracellular ATP (10(-5) and 10(-4) M) to MC led to a cell contraction which was independent of extracellular calcium. 3. Membrane voltage (Vm) and ion currents were measured with the nystatin patch clamp technique. ATP induced a concentration-dependent transient depolarization of Vm (ED50: 2 x 10(-6) M). During the transient depolarization ion currents were monitored simultaneously and showed an increase of the inward- and outward current. 4. In a buffer with a reduced extracellular chloride concentration (from 145 to 30 mM) ATP induced a depolarization augmented to -4 +/- 4 mV. 5. ATP-gamma-S and 2-methylthio-ATP depolarized Vm to the same extent as ATP, whereas alpha,beta-methylene-ATP (all 10(-5) M) had no effect on Vm. 6. The Ca2+ ionophore, A23187, depolarized Vm transiently from -51 +/- 2 to -28 +/- 4 mV and caused an increase of the inward current. 7. The intracellular calcium activity [Ca2+]i was measured with the fura-2 technique. ATP stimulated a concentration-dependent increase of [Ca2+]i (ED50: 5 x 10(-6) M). The increase of [Ca2+]i was biphasic with an initial peak followed by a sustained plateau. 8. The [Ca2+]i peak was still present in an extracellular Ca(2+)-free buffer, whereas the plateau was abolished. Verapamil (10(-4) M) did not inhibit the [Ca2+]i increase induced by ATP.(ABSTRACT TRUNCATED AT 250 WORDS) Images Figure 1 PMID:7691366

Cameleon calcium indicator reports cytoplasmic calcium dynamics in Arabidopsis guard cells

NASA Technical Reports Server (NTRS)

Allen, G. J.; Kwak, J. M.; Chu, S. P.; Llopis, J.; Tsien, R. Y.; Harper, J. F.; Schroeder, J. I.; Evans, M. L. (Principal Investigator)

1999-01-01

Cytoplasmic free calcium ([Ca2+]cyt) acts as a stimulus-induced second messenger in plant cells and multiple signal transduction pathways regulate [Ca2+]cyt in stomatal guard cells. Measuring [Ca2+]cyt in guard cells has previously required loading of calcium-sensitive dyes using invasive and technically difficult micro-injection techniques. To circumvent these problems, we have constitutively expressed the pH-independent, green fluorescent protein-based calcium indicator yellow cameleon 2.1 in Arabidopsis thaliana (Miyawaki et al. 1999; Proc. Natl. Acad. Sci. USA 96, 2135-2140). This yellow cameleon calcium indicator was expressed in guard cells and accumulated predominantly in the cytoplasm. Fluorescence ratio imaging of yellow cameleon 2.1 allowed time-dependent measurements of [Ca2+]cyt in Arabidopsis guard cells. Application of extracellular calcium or the hormone abscisic acid (ABA) induced repetitive [Ca2+]cyt transients in guard cells. [Ca2+]cyt changes could be semi-quantitatively determined following correction of the calibration procedure for chloroplast autofluorescence. Extracellular calcium induced repetitive [Ca2+]cyt transients with peak values of up to approximately 1.5 microM, whereas ABA-induced [Ca2+]cyt transients had peak values up to approximately 0.6 microM. These values are similar to stimulus-induced [Ca2+]cyt changes previously reported in plant cells using ratiometric dyes or aequorin. In some guard cells perfused with low extracellular KCl concentrations, spontaneous calcium transients were observed. As yellow cameleon 2.1 was expressed in all guard cells, [Ca2+]cyt was measured independently in the two guard cells of single stomates for the first time. ABA-induced, calcium-induced or spontaneous [Ca2+]cyt increases were not necessarily synchronized in the two guard cells. Overall, these data demonstrate that that GFP-based cameleon calcium indicators are suitable to measure [Ca2+]cyt changes in guard cells and enable the pattern of [Ca2+]cyt dynamics to be measured with a high level of reproducibility in Arabidopsis cells. This technical advance in combination with cell biological and molecular genetic approaches will become an invaluable tool in the dissection of plant cell signal transduction pathways.

Large-scale imaging of cortical network activity with calcium indicators.

PubMed

Ikegaya, Yuji; Le Bon-Jego, Morgane; Yuste, Rafael

2005-06-01

Bulk loading of calcium indicators has provided a unique opportunity to reconstruct the activity of cortical networks with single-cell resolution. Here we describe the detailed methods of bulk loading of AM dyes we developed and have been improving for imaging with a spinning disk confocal microscope.

Lipofection: a highly efficient, lipid-mediated DNA-transfection procedure.

PubMed Central

Felgner, P L; Gadek, T R; Holm, M; Roman, R; Chan, H W; Wenz, M; Northrop, J P; Ringold, G M; Danielsen, M

1987-01-01

A DNA-transfection protocol has been developed that makes use of a synthetic cationic lipid, N-[1-(2,3-dioleyloxy)propyl]-N,N,N-trimethylammonium chloride (DOTMA). Small unilamellar liposomes containing DOTMA interact spontaneously with DNA to form lipid-DNA complexes with 100% entrapment of the DNA, DOTMA facilitates fusion of the complex with the plasma membrane of tissue culture cells, resulting in both uptake and expression of the DNA. The technique is simple, highly reproducible, and effective for both transient and stable expression of transfected DNA. Depending upon the cell line, lipofection is from 5- to greater than 100-fold more effective than either the calcium phosphate or the DEAE-dextran transfection technique. Images PMID:2823261

Minerals and aligned collagen fibrils in tilapia fish scales: structural analysis using dark-field and energy-filtered transmission electron microscopy and electron tomography.

PubMed

Okuda, Mitsuhiro; Ogawa, Nobuhiro; Takeguchi, Masaki; Hashimoto, Ayako; Tagaya, Motohiro; Chen, Song; Hanagata, Nobutaka; Ikoma, Toshiyuki

2011-10-01

The mineralized structure of aligned collagen fibrils in a tilapia fish scale was investigated using transmission electron microscopy (TEM) techniques after a thin sample was prepared using aqueous techniques. Electron diffraction and electron energy loss spectroscopy data indicated that a mineralized internal layer consisting of aligned collagen fibrils contains hydroxyapatite crystals. Bright-field imaging, dark-field imaging, and energy-filtered TEM showed that the hydroxyapatite was mainly distributed in the hole zones of the aligned collagen fibrils structure, while needle-like materials composed of calcium compounds including hydroxyapatite existed in the mineralized internal layer. Dark-field imaging and three-dimensional observation using electron tomography revealed that hydroxyapatite and needle-like materials were mainly found in the matrix between the collagen fibrils. It was observed that hydroxyapatite and needle-like materials were preferentially distributed on the surface of the hole zones in the aligned collagen fibrils structure and in the matrix between the collagen fibrils in the mineralized internal layer of the scale.

The need for calcium imaging in nonhuman primates: New motor neuroscience and brain-machine interfaces.

PubMed

O'Shea, Daniel J; Trautmann, Eric; Chandrasekaran, Chandramouli; Stavisky, Sergey; Kao, Jonathan C; Sahani, Maneesh; Ryu, Stephen; Deisseroth, Karl; Shenoy, Krishna V

2017-01-01

A central goal of neuroscience is to understand how populations of neurons coordinate and cooperate in order to give rise to perception, cognition, and action. Nonhuman primates (NHPs) are an attractive model with which to understand these mechanisms in humans, primarily due to the strong homology of their brains and the cognitively sophisticated behaviors they can be trained to perform. Using electrode recordings, the activity of one to a few hundred individual neurons may be measured electrically, which has enabled many scientific findings and the development of brain-machine interfaces. Despite these successes, electrophysiology samples sparsely from neural populations and provides little information about the genetic identity and spatial micro-organization of recorded neurons. These limitations have spurred the development of all-optical methods for neural circuit interrogation. Fluorescent calcium signals serve as a reporter of neuronal responses, and when combined with post-mortem optical clearing techniques such as CLARITY, provide dense recordings of neuronal populations, spatially organized and annotated with genetic and anatomical information. Here, we advocate that this methodology, which has been of tremendous utility in smaller animal models, can and should be developed for use with NHPs. We review here several of the key opportunities and challenges for calcium-based optical imaging in NHPs. We focus on motor neuroscience and brain-machine interface design as representative domains of opportunity within the larger field of NHP neuroscience. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

Comparison of the x-ray attenuation properties of breast calcifications, aluminium, hydroxyapatite and calcium oxalate.

PubMed

Warren, L M; Mackenzie, A; Dance, D R; Young, K C

2013-04-07

Aluminium is often used as a substitute material for calcifications in phantom measurements in mammography. Additionally, calcium oxalate, hydroxyapatite and aluminium are used in simulation studies. This assumes that these materials have similar attenuation properties to calcification, and this assumption is examined in this work. Sliced mastectomy samples containing calcification were imaged at ×5 magnification using a digital specimen cabinet. Images of the individual calcifications were extracted, and the diameter and contrast of each calculated. The thicknesses of aluminium required to achieve the same contrast as each calcification when imaged under the same conditions were calculated using measurements of the contrast of aluminium foils. As hydroxyapatite and calcium oxalate are also used to simulate calcifications, the equivalent aluminium thicknesses of these materials were also calculated using tabulated attenuation coefficients. On average the equivalent aluminium thickness was 0.85 times the calcification diameter. For calcium oxalate and hydroxyapatite, the equivalent aluminium thicknesses were 1.01 and 2.19 times the thickness of these materials respectively. Aluminium and calcium oxalate are suitable substitute materials for calcifications. Hydroxyapatite is much more attenuating than the calcifications and aluminium. Using solid hydroxyapatite as a substitute for calcification of the same size would lead to excessive contrast in the mammographic image.

Space and Time Resolved Detection of Platelet Activation and von Willebrand Factor Conformational Changes in Deep Suspensions.

PubMed

Biasetti, Jacopo; Sampath, Kaushik; Cortez, Angel; Azhir, Alaleh; Gilad, Assaf A; Kickler, Thomas S; Obser, Tobias; Ruggeri, Zaverio M; Katz, Joseph

2017-01-01

Tracking cells and proteins' phenotypic changes in deep suspensions is critical for the direct imaging of blood-related phenomena in in vitro replica of cardiovascular systems and blood-handling devices. This paper introduces fluorescence imaging techniques for space and time resolved detection of platelet activation, von Willebrand factor (VWF) conformational changes, and VWF-platelet interaction in deep suspensions. Labeled VWF, platelets, and VWF-platelet strands are suspended in deep cuvettes, illuminated, and imaged with a high-sensitivity EM-CCD camera, allowing detection using an exposure time of 1 ms. In-house postprocessing algorithms identify and track the moving signals. Recombinant VWF-eGFP (rVWF-eGFP) and VWF labeled with an FITC-conjugated polyclonal antibody are employed. Anti-P-Selectin FITC-conjugated antibodies and the calcium-sensitive probe Indo-1 are used to detect activated platelets. A positive correlation between the mean number of platelets detected per image and the percentage of activated platelets determined through flow cytometry is obtained, validating the technique. An increase in the number of rVWF-eGFP signals upon exposure to shear stress demonstrates the technique's ability to detect breakup of self-aggregates. VWF globular and unfolded conformations and self-aggregation are also observed. The ability to track the size and shape of VWF-platelet strands in space and time provides means to detect pro- and antithrombotic processes.

Confocal microscopy for astrocyte in vivo imaging: Recycle and reuse in microscopy

PubMed Central

Pérez-Alvarez, Alberto; Araque, Alfonso; Martín, Eduardo D.

2013-01-01

In vivo imaging is one of the ultimate and fundamental approaches for the study of the brain. Two-photon laser scanning microscopy (2PLSM) constitutes the state-of-the-art technique in current neuroscience to address questions regarding brain cell structure, development and function, blood flow regulation and metabolism. This technique evolved from laser scanning confocal microscopy (LSCM), which impacted the field with a major improvement in image resolution of live tissues in the 1980s compared to widefield microscopy. While nowadays some of the unparalleled features of 2PLSM make it the tool of choice for brain studies in vivo, such as the possibility to image deep within a tissue, LSCM can still be useful in this matter. Here we discuss the validity and limitations of LSCM and provide a guide to perform high-resolution in vivo imaging of the brain of live rodents with minimal mechanical disruption employing LSCM. We describe the surgical procedure and experimental setup that allowed us to record intracellular calcium variations in astrocytes evoked by sensory stimulation, and to monitor intact neuronal dendritic spines and astrocytic processes as well as blood vessel dynamics. Therefore, in spite of certain limitations that need to be carefully considered, LSCM constitutes a useful, convenient, and affordable tool for brain studies in vivo. PMID:23658537

Dual-slit confocal light sheet microscopy for in vivo whole-brain imaging of zebrafish

PubMed Central

Yang, Zhe; Mei, Li; Xia, Fei; Luo, Qingming; Fu, Ling; Gong, Hui

2015-01-01

In vivo functional imaging at single-neuron resolution is an important approach to visualize biological processes in neuroscience. Light sheet microscopy (LSM) is a cutting edge in vivo imaging technique that provides micron-scale spatial resolution at high frame rate. Due to the scattering and absorption of tissue, however, conventional LSM is inadequate to resolve cells because of the attenuated signal to noise ratio (SNR). Using dual-beam illumination and confocal dual-slit detection, here a dual-slit confocal LSM is demonstrated to obtain the SNR enhanced images with frame rate twice as high as line confocal LSM method. Through theoretical calculations and experiments, the correlation between the slit’s width and SNR was determined to optimize the image quality. In vivo whole brain structural imaging stacks and the functional imaging sequences of single slice were obtained for analysis of calcium activities at single-cell resolution. A two-fold increase in imaging speed of conventional confocal LSM makes it possible to capture the sequence of the neurons’ activities and help reveal the potential functional connections in the whole zebrafish’s brain. PMID:26137381

An open source tool for automatic spatiotemporal assessment of calcium transients and local ‘signal-close-to-noise’ activity in calcium imaging data

PubMed Central

Martin, Corinna; Jablonka, Sibylle

2018-01-01

Local and spontaneous calcium signals play important roles in neurons and neuronal networks. Spontaneous or cell-autonomous calcium signals may be difficult to assess because they appear in an unpredictable spatiotemporal pattern and in very small neuronal loci of axons or dendrites. We developed an open source bioinformatics tool for an unbiased assessment of calcium signals in x,y-t imaging series. The tool bases its algorithm on a continuous wavelet transform-guided peak detection to identify calcium signal candidates. The highly sensitive calcium event definition is based on identification of peaks in 1D data through analysis of a 2D wavelet transform surface. For spatial analysis, the tool uses a grid to separate the x,y-image field in independently analyzed grid windows. A document containing a graphical summary of the data is automatically created and displays the loci of activity for a wide range of signal intensities. Furthermore, the number of activity events is summed up to create an estimated total activity value, which can be used to compare different experimental situations, such as calcium activity before or after an experimental treatment. All traces and data of active loci become documented. The tool can also compute the signal variance in a sliding window to visualize activity-dependent signal fluctuations. We applied the calcium signal detector to monitor activity states of cultured mouse neurons. Our data show that both the total activity value and the variance area created by a sliding window can distinguish experimental manipulations of neuronal activity states. Notably, the tool is powerful enough to compute local calcium events and ‘signal-close-to-noise’ activity in small loci of distal neurites of neurons, which remain during pharmacological blockade of neuronal activity with inhibitors such as tetrodotoxin, to block action potential firing, or inhibitors of ionotropic glutamate receptors. The tool can also offer information about local homeostatic calcium activity events in neurites. PMID:29601577

In vivo alterations in calcium buffering capacity in transgenic mouse model of synucleinopathy.

PubMed

Reznichenko, Lidia; Cheng, Qun; Nizar, Krystal; Gratiy, Sergey L; Saisan, Payam A; Rockenstein, Edward M; González, Tanya; Patrick, Christina; Spencer, Brian; Desplats, Paula; Dale, Anders M; Devor, Anna; Masliah, Eliezer

2012-07-18

Abnormal accumulation of α-synuclein is centrally involved in the pathogenesis of many disorders with Parkinsonism and dementia. Previous in vitro studies suggest that α-synuclein dysregulates intracellular calcium. However, it is unclear whether these alterations occur in vivo. For this reason, we investigated calcium dynamics in transgenic mice expressing human WT α-synuclein using two-photon microscopy. We imaged spontaneous and stimulus-induced neuronal activity in the barrel cortex. Transgenic mice exhibited augmented, long-lasting calcium transients characterized by considerable deviation from the exponential decay. The most evident pathology was observed in response to a repetitive stimulation in which subsequent stimuli were presented before relaxation of calcium signal to the baseline. These alterations were detected in the absence of significant increase in neuronal spiking response compared with age-matched controls, supporting the possibility that α-synuclein promoted alterations in calcium dynamics via interference with intracellular buffering mechanisms. The characteristic shape of calcium decay and augmented response during repetitive stimulation can serve as in vivo imaging biomarkers in this model of neurodegeneration, to monitor progression of the disease and screen candidate treatment strategies.

High-throughput automated home-cage mesoscopic functional imaging of mouse cortex

PubMed Central

Murphy, Timothy H.; Boyd, Jamie D.; Bolaños, Federico; Vanni, Matthieu P.; Silasi, Gergely; Haupt, Dirk; LeDue, Jeff M.

2016-01-01

Mouse head-fixed behaviour coupled with functional imaging has become a powerful technique in rodent systems neuroscience. However, training mice can be time consuming and is potentially stressful for animals. Here we report a fully automated, open source, self-initiated head-fixation system for mesoscopic functional imaging in mice. The system supports five mice at a time and requires minimal investigator intervention. Using genetically encoded calcium indicator transgenic mice, we longitudinally monitor cortical functional connectivity up to 24 h per day in >7,000 self-initiated and unsupervised imaging sessions up to 90 days. The procedure provides robust assessment of functional cortical maps on the basis of both spontaneous activity and brief sensory stimuli such as light flashes. The approach is scalable to a number of remotely controlled cages that can be assessed within the controlled conditions of dedicated animal facilities. We anticipate that home-cage brain imaging will permit flexible and chronic assessment of mesoscale cortical function. PMID:27291514

Calcium-sensitive MRI contrast agents based on superparamagnetic iron oxide nanoparticles and calmodulin

PubMed Central

Atanasijevic, Tatjana; Shusteff, Maxim; Fam, Peter; Jasanoff, Alan

2006-01-01

We describe a family of calcium indicators for magnetic resonance imaging (MRI), formed by combining a powerful iron oxide nanoparticle-based contrast mechanism with the versatile calcium-sensing protein calmodulin and its targets. Calcium-dependent protein–protein interactions drive particle clustering and produce up to 5-fold changes in T2 relaxivity, an indication of the sensors' potency. A variant based on conjugates of wild-type calmodulin and the peptide M13 reports concentration changes near 1 μM Ca2+, suitable for detection of elevated intracellular calcium levels. The midpoint and cooperativity of the response can be tuned by mutating the protein domains that actuate the sensor. Robust MRI signal changes are achieved even at nanomolar particle concentrations (<1 μM in calmodulin) that are unlikely to buffer calcium levels. When combined with technologies for cellular delivery of nanoparticulate agents, these sensors and their derivatives may be useful for functional molecular imaging of biological signaling networks in live, opaque specimens. PMID:17003117

Calcium-sensitive MRI contrast agents based on superparamagnetic iron oxide nanoparticles and calmodulin.

PubMed

Atanasijevic, Tatjana; Shusteff, Maxim; Fam, Peter; Jasanoff, Alan

2006-10-03

We describe a family of calcium indicators for magnetic resonance imaging (MRI), formed by combining a powerful iron oxide nanoparticle-based contrast mechanism with the versatile calcium-sensing protein calmodulin and its targets. Calcium-dependent protein-protein interactions drive particle clustering and produce up to 5-fold changes in T2 relaxivity, an indication of the sensors' potency. A variant based on conjugates of wild-type calmodulin and the peptide M13 reports concentration changes near 1 microM Ca(2+), suitable for detection of elevated intracellular calcium levels. The midpoint and cooperativity of the response can be tuned by mutating the protein domains that actuate the sensor. Robust MRI signal changes are achieved even at nanomolar particle concentrations (<1 microM in calmodulin) that are unlikely to buffer calcium levels. When combined with technologies for cellular delivery of nanoparticulate agents, these sensors and their derivatives may be useful for functional molecular imaging of biological signaling networks in live, opaque specimens.

Efficient multi-site two-photon functional imaging of neuronal circuits.

PubMed

Castanares, Michael Lawrence; Gautam, Vini; Drury, Jack; Bachor, Hans; Daria, Vincent R

2016-12-01

Two-photon imaging using high-speed multi-channel detectors is a promising approach for optical recording of cellular membrane dynamics at multiple sites. A main bottleneck of this technique is the limited number of photons captured within a short exposure time (~1ms). Here, we implement temporal gating to improve the two-photon fluorescence yield from holographically projected multiple foci whilst maintaining a biologically safe incident average power. We observed up to 6x improvement in the signal-to-noise ratio (SNR) in Fluorescein and cultured hippocampal neurons showing evoked calcium transients. With improved SNR, we could pave the way to achieving multi-site optical recording of fluorogenic probes with response times in the order of ~1ms.

Efficient multi-site two-photon functional imaging of neuronal circuits

PubMed Central

Castanares, Michael Lawrence; Gautam, Vini; Drury, Jack; Bachor, Hans; Daria, Vincent R.

2016-01-01

Two-photon imaging using high-speed multi-channel detectors is a promising approach for optical recording of cellular membrane dynamics at multiple sites. A main bottleneck of this technique is the limited number of photons captured within a short exposure time (~1ms). Here, we implement temporal gating to improve the two-photon fluorescence yield from holographically projected multiple foci whilst maintaining a biologically safe incident average power. We observed up to 6x improvement in the signal-to-noise ratio (SNR) in Fluorescein and cultured hippocampal neurons showing evoked calcium transients. With improved SNR, we could pave the way to achieving multi-site optical recording of fluorogenic probes with response times in the order of ~1ms. PMID:28018745

Fast Kinetics of Calcium Signaling and Sensor Design

PubMed Central

Tang, Shen; Reddish, Florence; Zhuo, You; Yang, Jenny J.

2015-01-01

Fast calcium signaling is regulated by numerous calcium channels exhibiting high spatiotemporal profiles which are currently measured by fluorescent calcium sensors. There is still a strong need to improve the kinetics of genetically encoded calcium indicators (sensors) to capture calcium dynamics in the millisecond time frame. In this review, we summarize several major fast calcium signaling pathways and discuss the recent developments and application of genetically encoded calcium indicators to detect these pathways. A new class of genetically encoded calcium indicators designed with site-directed mutagenesis on the surface of beta-barrel fluorescent proteins to form a pentagonal bipyramidal-like calcium binding domain dramatically accelerates calcium binding kinetics. Furthermore, novel genetically encoded calcium indicators with significantly increased fluorescent lifetime change are advantageous in deep-field imaging with high light-scattering and notable morphology change. PMID:26151819

[Myofibroblasts and afferent signalling in the urinary bladder. A concept].

PubMed

Neuhaus, J; Scholler, U; Freick, K; Schwalenberg, T; Heinrich, M; Horn, L C; Stolzenburg, J U

2008-09-01

Afferent signal transduction in the urinary bladder is still not clearly understood. An increasing body of evidence supports the view of complex interactions between urothelium, suburothelial myofibroblasts, and sensory nerves. Bladder tissue from tumour patients was used in this study. Methods included confocal immunofluorescence, polymerase chain reaction, calcium imaging, and fluorescence recovery after photobleaching (FRAP).Myofibroblasts express muscarinic and purinergic receptors. They show constitutive spontaneous activity in calcium imaging, which completely depends on extracellular calcium. Stimulation with carbachol and ATP-evoked intracellular calcium transients also depend on extracellular calcium. The intensive coupling between the cells is significantly diminished by incubation with TGF-beta 1. Myofibroblasts form an important cellular element within the afferent signalling of the urinary bladder. They possess all features required to take part in the complex interactions with urothelial cells and sensory nerves. Modulation of their function by cytokines may provide a pathomechanism for bladder dysfunction.

Ground-to-air flow visualization using Solar Calcium-K line Background-Oriented Schlieren

NASA Astrophysics Data System (ADS)

Hill, Michael A.; Haering, Edward A.

2017-01-01

The Calcium-K Eclipse Background-Oriented Schlieren experiment was performed as a proof of concept test to evaluate the effectiveness of using the solar disk as a background to perform the Background-Oriented Schlieren (BOS) method of flow visualization. A ground-based imaging system was equipped with a Calcium-K line optical etalon filter to enable the use of the chromosphere of the sun as the irregular background to be used for BOS. A US Air Force T-38 aircraft performed three supersonic runs which eclipsed the sun as viewed from the imaging system. The images were successfully post-processed using optical flow methods to qualitatively reveal the density gradients in the flow around the aircraft.

Electron microprobe analysis and histochemical examination of the calcium distribution in human bone trabeculae: a methodological study using biopsy specimens from post-traumatic osteopenia

DOE Office of Scientific and Technical Information (OSTI.GOV)

Obrant, K.J.; Odselius, R.

1984-01-01

Energy dispersive X-ray microanalysis (EDX) (or electron microprobe analysis) of the relative intensity for calcium in different bone trabeculae from the tibia epiphysis, and in different parts of one and the same trabecula, was performed on 3 patients who had earlier had a fracture of the ipsilateral tibia-diaphysis. The variation in intensity was compared with the histochemical patterns obtained with both the Goldner and the von Kossa staining techniques for detecting calcium in tissues. Previously reported calcium distribution features, found to be typical for posttraumatic osteopenia, such as striated mineralization patterns in individual trabeculae and large differences in mineralization levelmore » between different trabeculae, could be verified both by means of the two histochemical procedures and from the electron microprobe analysis. A pronounced difference was observed, however, between the two histochemical staining techniques as regards their sensitivity to detect calcium. To judge from the values obtained from the EDX measurements, the sensitivity of the Goldner technique should be more than ten times higher than that of von Kossa. The EDX measurements gave more detailed information than either of the two histochemical techniques: great variations in the intensity of the calcium peak were found in trabeculae stained as unmineralized as well as mineralized.« less

Dual-energy digital mammography for calcification imaging: scatter and nonuniformity corrections.

PubMed

Kappadath, S Cheenu; Shaw, Chris C

2005-11-01

Mammographic images of small calcifications, which are often the earliest signs of breast cancer, can be obscured by overlapping fibroglandular tissue. We have developed and implemented a dual-energy digital mammography (DEDM) technique for calcification imaging under full-field imaging conditions using a commercially available aSi:H/CsI:Tl flat-panel based digital mammography system. The low- and high-energy images were combined using a nonlinear mapping function to cancel the tissue structures and generate the dual-energy (DE) calcification images. The total entrance-skin exposure and mean-glandular dose from the low- and high-energy images were constrained so that they were similar to screening-examination levels. To evaluate the DE calcification image, we designed a phantom using calcium carbonate crystals to simulate calcifications of various sizes (212-425 microm) overlaid with breast-tissue-equivalent material 5 cm thick with a continuously varying glandular-tissue ratio from 0% to 100%. We report on the effects of scatter radiation and nonuniformity in x-ray intensity and detector response on the DE calcification images. The nonuniformity was corrected by normalizing the low- and high-energy images with full-field reference images. Correction of scatter in the low- and high-energy images significantly reduced the background signal in the DE calcification image. Under the current implementation of DEDM, utilizing the mammography system and dose level tested, calcifications in the 300-355 microm size range were clearly visible in DE calcification images. Calcification threshold sizes decreased to the 250-280 microm size range when the visibility criteria were lowered to barely visible. Calcifications smaller than approximately 250 microm were usually not visible in most cases. The visibility of calcifications with our DEDM imaging technique was limited by quantum noise, not system noise.

Dual-energy digital mammography for calcification imaging: Scatter and nonuniformity corrections

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kappadath, S. Cheenu; Shaw, Chris C.

Mammographic images of small calcifications, which are often the earliest signs of breast cancer, can be obscured by overlapping fibroglandular tissue. We have developed and implemented a dual-energy digital mammography (DEDM) technique for calcification imaging under full-field imaging conditions using a commercially available aSi:H/CsI:Tl flat-panel based digital mammography system. The low- and high-energy images were combined using a nonlinear mapping function to cancel the tissue structures and generate the dual-energy (DE) calcification images. The total entrance-skin exposure and mean-glandular dose from the low- and high-energy images were constrained so that they were similar to screening-examination levels. To evaluate the DEmore » calcification image, we designed a phantom using calcium carbonate crystals to simulate calcifications of various sizes (212-425 {mu}m) overlaid with breast-tissue-equivalent material 5 cm thick with a continuously varying glandular-tissue ratio from 0% to 100%. We report on the effects of scatter radiation and nonuniformity in x-ray intensity and detector response on the DE calcification images. The nonuniformity was corrected by normalizing the low- and high-energy images with full-field reference images. Correction of scatter in the low- and high-energy images significantly reduced the background signal in the DE calcification image. Under the current implementation of DEDM, utilizing the mammography system and dose level tested, calcifications in the 300-355 {mu}m size range were clearly visible in DE calcification images. Calcification threshold sizes decreased to the 250-280 {mu}m size range when the visibility criteria were lowered to barely visible. Calcifications smaller than {approx}250 {mu}m were usually not visible in most cases. The visibility of calcifications with our DEDM imaging technique was limited by quantum noise, not system noise.« less

CALCIUM ABSORPTION IN MAN: BASED ON LARGE VOLUME LIQUID SCINTILLATION COUNTER STUDIES.

PubMed

LUTWAK, L; SHAPIRO, J R

1964-05-29

A technique has been developed for the in vivo measurement of absorption of calcium in man after oral administration of 1 to 5 microcuries of calcium-47 and continuous counting of the radiation in the subject's arm with a large volume liquid scintillation counter. The maximum value for the arm counting technique is proportional to the absorption of tracer as measured by direct stool analysis. The rate of uptake by the arm is lower in subjects with either the malabsorption syndrome or hypoparathyroidism. The administration of vitamin D increases both the absorption rate and the maximum amount of calcium absorbed.

In vivo optical activation of astrocytes as a potential therapeutic strategy for neurodegenerative diseases

NASA Astrophysics Data System (ADS)

Chen, Yuanxin; Mancuso, James; Zhao, Zhen; Li, Xuping; Xue, Zhong; Wong, Stephen T. C.

2013-03-01

Neurovascular dysfunction in many neurodegenerative diseases, such as Alzheimer's disease (AD), reduces blood flow to affected brain areas and causes neuronal dysfunction and loss. A new optical imaging technique is developed to activate astrocytes in live animal models in order to investigate the increase of local cerebral blood flow as a potential therapeutic strategy for AD. The technique uses fluorescent labeling of vasculature and astrocytes coupled with intravital 2-photon microscopy to visualize the astrocyte-vasculature interactions in live animals. Using femtosecond laser stimulation, calcium uncaging is applied to specifically target and activate astrocytes in vivo with high spatial and temporal resolutions. Intravital 2-photon microscopy imaging was employed to demonstrate that single endfoot optical activation around an arteriole induced a 25% increase in arteriole diameter, which in turn increased cerebral local blood flow in down-stream capillaries. This quantitative result indicates the potential of using optical activation of astrocytes in afflicted brain areas of neurodegeneration to restore normal neurovascular functions.

Bone metabolism in oxalosis: a single-center study using new imaging techniques and biomarkers.

PubMed

Bacchetta, Justine; Fargue, Sonia; Boutroy, Stéphanie; Basmaison, Odile; Vilayphiou, Nicolas; Plotton, Ingrid; Guebre-Egziabher, Fitsum; Dohin, Bruno; Kohler, Rémi; Cochat, Pierre

2010-06-01

The deposition of calcium oxalate crystals in the kidney and bone is a hallmark of primary hyperoxaluria type 1 (PH1). We report here an evaluation of the bone status of 12 PH1 children based on bone biomarkers [parathyroid hormone, vitamin D, fibroblast growth factor 23 (FGF23)] and radiological assessments (skeletal age, three-dimensional high-resolution peripheral quantitative computed tomography, HR-pQCT) carried out within the framework of a cross-sectional single-center study. The controls consisted of healthy and children with chronic kidney disease already enrolled in local bone and mineral metabolism studies. The mean age (+ or - standard deviation) age of the patients was 99 (+ or - 63) months. Six children suffered from fracture. Bone maturation was accelerated in five patients, four of whom were <5 years. The combination of new imaging techniques and biomarkers highlighted new and unexplained features of PH1: advanced skeletal age in young PH1 patients, increased FGF23 levels and decreased total volumetric bone mineral density with bone microarchitecture alteration.

Space Flight Calcium: Implications for Astronaut Health, Spacecraft Operations, and Earth

PubMed Central

Smith, Scott M.; McCoy, Torin; Gazda, Daniel; Morgan, Jennifer L. L.; Heer, Martina; Zwart, Sara R.

2012-01-01

The space flight environment is known to induce bone loss and, subsequently, calcium loss. The longer the mission, generally the more bone and calcium are lost. This review provides a history of bone and calcium studies related to space flight and highlights issues related to calcium excretion that the space program must consider so that urine can be recycled. It also discusses a novel technique using natural stable isotopes of calcium that will be helpful in the future to determine calcium and bone balance during space flight. PMID:23250146

Space flight calcium: implications for astronaut health, spacecraft operations, and Earth.

PubMed

Smith, Scott M; McCoy, Torin; Gazda, Daniel; Morgan, Jennifer L L; Heer, Martina; Zwart, Sara R

2012-12-18

The space flight environment is known to induce bone loss and, subsequently, calcium loss. The longer the mission, generally the more bone and calcium are lost. This review provides a history of bone and calcium studies related to space flight and highlights issues related to calcium excretion that the space program must consider so that urine can be recycled. It also discusses a novel technique using natural stable isotopes of calcium that will be helpful in the future to determine calcium and bone balance during space flight.

Surface analysis and depth profiling of corrosion products formed in lead pipes used to supply low alkalinity drinking water.

PubMed

Davidson, C M; Peters, N J; Britton, A; Brady, L; Gardiner, P H E; Lewis, B D

2004-01-01

Modern analytical techniques have been applied to investigate the nature of lead pipe corrosion products formed in pH adjusted, orthophosphate-treated, low alkalinity water, under supply conditions. Depth profiling and surface analysis have been carried out on pipe samples obtained from the water distribution system in Glasgow, Scotland, UK. X-ray diffraction spectrometry identified basic lead carbonate, lead oxide and lead phosphate as the principal components. Scanning electron microscopy/energy-dispersive x-ray spectrometry revealed the crystalline structure within the corrosion product and also showed spatial correlations existed between calcium, iron, lead, oxygen and phosphorus. Elemental profiling, conducted by means of secondary ion mass spectrometry (SIMS) and secondary neutrals mass spectrometry (SNMS) indicated that the corrosion product was not uniform with depth. However, no clear stratification was apparent. Indeed, counts obtained for carbonate, phosphate and oxide were well correlated within the depth range probed by SIMS. SNMS showed relationships existed between carbon, calcium, iron, and phosphorus within the bulk of the scale, as well as at the surface. SIMS imaging confirmed the relationship between calcium and lead and suggested there might also be an association between chloride and phosphorus.

Automatic coronary calcium scoring using noncontrast and contrast CT images

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yang, Guanyu, E-mail: yang.list@seu.edu.cn; Chen, Yang; Shu, Huazhong

Purpose: Calcium scoring is widely used to assess the risk of coronary heart disease (CHD). Accurate coronary artery calcification detection in noncontrast CT image is a prerequisite step for coronary calcium scoring. Currently, calcified lesions in the coronary arteries are manually identified by radiologists in clinical practice. Thus, in this paper, a fully automatic calcium scoring method was developed to alleviate the work load of the radiologists or cardiologists. Methods: The challenge of automatic coronary calcification detection is to discriminate the calcification in the coronary arteries from the calcification in the other tissues. Since the anatomy of coronary arteries ismore » difficult to be observed in the noncontrast CT images, the contrast CT image of the same patient is used to extract the regions of the aorta, heart, and coronary arteries. Then, a patient-specific region-of-interest (ROI) is generated in the noncontrast CT image according to the segmentation results in the contrast CT image. This patient-specific ROI focuses on the regions in the neighborhood of coronary arteries for calcification detection, which can eliminate the calcifications in the surrounding tissues. A support vector machine classifier is applied finally to refine the results by removing possible image noise. Furthermore, the calcified lesions in the noncontrast images belonging to the different main coronary arteries are identified automatically using the labeling results of the extracted coronary arteries. Results: Forty datasets from four different CT machine vendors were used to evaluate their algorithm, which were provided by the MICCAI 2014 Coronary Calcium Scoring (orCaScore) Challenge. The sensitivity and positive predictive value for the volume of detected calcifications are 0.989 and 0.948. Only one patient out of 40 patients had been assigned to the wrong risk category defined according to Agatston scores (0, 1–100, 101–300, >300) by comparing with the ground truth. Conclusions: The calcified lesions in the noncontrast CT images can be detected automatically by using the segmentation results of the aorta, heart, and coronary arteries obtained in the contrast CT images with a very high accuracy.« less

Video-rate volumetric functional imaging of the brain at synaptic resolution.

PubMed

Lu, Rongwen; Sun, Wenzhi; Liang, Yajie; Kerlin, Aaron; Bierfeld, Jens; Seelig, Johannes D; Wilson, Daniel E; Scholl, Benjamin; Mohar, Boaz; Tanimoto, Masashi; Koyama, Minoru; Fitzpatrick, David; Orger, Michael B; Ji, Na

2017-04-01

Neurons and neural networks often extend hundreds of micrometers in three dimensions. Capturing the calcium transients associated with their activity requires volume imaging methods with subsecond temporal resolution. Such speed is a challenge for conventional two-photon laser-scanning microscopy, because it depends on serial focal scanning in 3D and indicators with limited brightness. Here we present an optical module that is easily integrated into standard two-photon laser-scanning microscopes to generate an axially elongated Bessel focus, which when scanned in 2D turns frame rate into volume rate. We demonstrated the power of this approach in enabling discoveries for neurobiology by imaging the calcium dynamics of volumes of neurons and synapses in fruit flies, zebrafish larvae, mice and ferrets in vivo. Calcium signals in objects as small as dendritic spines could be resolved at video rates, provided that the samples were sparsely labeled to limit overlap in their axially projected images.

A brief review of space flight calcium metabolism: Results and methodologies

NASA Technical Reports Server (NTRS)

1978-01-01

Space flight induced osteoporosis was described. The techniques that were used to measure and detect the osteoporosis were also described. Areas of calcium metabolism were shown to be very important in the investigation into more sensitive detection and measurement techniques of bone demineralization.

Technical feasibility and validation of a coronary artery calcium scoring system using CT coronary angiography images.

PubMed

Pavitt, Christopher W; Harron, Katie; Lindsay, Alistair C; Zielke, Sayeh; Ray, Robin; Gordon, Daniel; Rubens, Michael B; Padley, Simon P; Nicol, Edward D

2016-05-01

We validate a novel CT coronary angiography (CCTA) coronary calcium scoring system. Calcium was quantified on CCTA images using a new patient-specific attenuation threshold: mean + 2SD of intra-coronary contrast density (HU). Using 335 patient data sets a conversion factor (CF) for predicting CACS from CCTA scores (CCTAS) was derived and validated in a separate cohort (n = 168). Bland-Altman analysis and weighted kappa for MESA centiles and Agatston risk groupings were calculated. Multivariable linear regression yielded a CF: CACS = (1.185 × CCTAS) + (0.002 × CCTAS × attenuation threshold). When applied to CCTA data sets there was excellent correlation (r = 0.95; p < 0.0001) and agreement (mean difference -10.4 [95% limits of agreement -258.9 to 238.1]) with traditional calcium scores. Agreement was better for calcium scores below 500; however, MESA percentile agreement was better for high risk patients. Risk stratification was excellent (Agatston groups k = 0.88 and MESA centiles k = 0.91). Eliminating the dedicated CACS scan decreased patient radiation exposure by approximately one-third. CCTA calcium scores can accurately predict CACS using a simple, individualized, semiautomated approach reducing acquisition time and radiation exposure when evaluating patients for CAD. This method is not affected by the ROI location, imaging protocol, or tube voltage strengthening its clinical applicability. • Coronary calcium scores can be reliably determined on contrast-enhanced cardiac CT • This score can accurately risk stratify patients • Elimination of a dedicated calcium scan reduces patient radiation by a third.

A mutation in the extracellular domain of the α7 nAChR reduces calcium permeability.

PubMed

Colón-Sáez, José O; Yakel, Jerrel L

2014-08-01

The α7 neuronal nicotinic acetylcholine receptor (nAChR) displays the highest calcium permeability among the different subtypes of nAChRs expressed in the mammalian brain and can impact cellular events including neurotransmitter release, second messenger cascades, cell survival, and apoptosis. The selectivity for cations in nAChRs is thought to be achieved in part by anionic residues which are located on either side of the channel mouth and increase relative cationic concentration. Mutagenesis studies have improved our understanding of the role of the second transmembrane domain and the intracellular loop of the channel in ion selectivity. However, little is known about the influence that the extracellular domain (ECD) plays in ion permeation. In the α7 nAChR, it has been found that the ECD contains a ring of ten aspartates (two per subunit) that is believed to face the lumen of the pore and could attract cations for permeation. Using mutagenesis and a combination of electrophysiology and imaging techniques, we tested the possible involvement of these aspartate residues in the calcium permeability of the rat α7 nAChR. We found that one of these residues (the aspartate at position 44) appears to be essential since mutating it to alanine resulted in a decrease in amplitude for both whole cell and single-channel responses and in the complete disappearance of detectable calcium changes in most cells, which indicates that the ECD of the α7 nAChR plays a key role in calcium permeation.

A mutation in the extracellular domain of the α7 nAChR reduces calcium permeability

PubMed Central

Colón-Sáez, José O.

2013-01-01

The α7 neuronal nicotinic acetylcholine receptor (nAChR) displays the highest calcium permeability among the different subtypes of nAChRs expressed in the mammalian brain and can impact cellular events including neurotransmitter release, second messenger cascades, cell survival, and apoptosis. The selectivity for cations in nAChRs is thought to be achieved in part by anionic residues which are located on either side of the channel mouth and increase relative cationic concentration. Mutagenesis studies have improved our understanding of the role of the second transmembrane domain and the intracellular loop of the channel in ion selectivity. However, little is known about the influence that the extracellular domain (ECD) plays in ion permeation. In the α7 nAChR, it has been found that the ECD contains a ring of ten aspartates (two per subunit) that is believed to face the lumen of the pore and could attract cations for permeation. Using mutagenesis and a combination of electrophysiology and imaging techniques, we tested the possible involvement of these aspartate residues in the calcium permeability of the rat α7 nAChR. We found that one of these residues (the aspartate at position 44) appears to be essential since mutating it to alanine resulted in a decrease in amplitude for both whole cell and single-channel responses and in the complete disappearance of detectable calcium changes in most cells, which indicates that the ECD of the α7 nAChR plays a key role in calcium permeation. PMID:24177919

Present and future of glass-ionomers and calcium-silicate cements as bioactive materials in dentistry: Biophotonics-based interfacial analyses in health and disease

PubMed Central

Watson, Timothy F.; Atmeh, Amre R.; Sajini, Shara; Cook, Richard J.; Festy, Frederic

2014-01-01

Objective Since their introduction, calcium silicate cements have primarily found use as endodontic sealers, due to long setting times. While similar in chemistry, recent variations such as constituent proportions, purities and manufacturing processes mandate a critical understanding of service behavior differences of the new coronal restorative material variants. Of particular relevance to minimally invasive philosophies is the potential for ion supply, from initial hydration to mature set in dental cements. They may be capable of supporting repair and remineralization of dentin left after decay and cavity preparation, following the concepts of ion exchange from glass ionomers. Methods This paper reviews the underlying chemistry and interactions of glass ionomer and calcium silicate cements, with dental tissues, concentrating on dentin–restoration interface reactions. We additionally demonstrate a new optical technique, based around high resolution deep tissue, two-photon fluorescence and lifetime imaging, which allows monitoring of undisturbed cement–dentin interface samples behavior over time. Results The local bioactivity of the calcium-silicate based materials has been shown to produce mineralization within the subjacent dentin substrate, extending deep within the tissues. This suggests that the local ion-rich alkaline environment may be more favorable to mineral repair and re-construction, compared with the acidic environs of comparable glass ionomer based materials. Significance The advantages of this potential re-mineralization phenomenon for minimally invasive management of carious dentin are self-evident. There is a clear need to improve the bioactivity of restorative dental materials and these calcium silicate cement systems offer exciting possibilities in realizing this goal. PMID:24113131

Voltage imaging to understand connections and functions of neuronal circuits.

PubMed

Antic, Srdjan D; Empson, Ruth M; Knöpfel, Thomas

2016-07-01

Understanding of the cellular mechanisms underlying brain functions such as cognition and emotions requires monitoring of membrane voltage at the cellular, circuit, and system levels. Seminal voltage-sensitive dye and calcium-sensitive dye imaging studies have demonstrated parallel detection of electrical activity across populations of interconnected neurons in a variety of preparations. A game-changing advance made in recent years has been the conceptualization and development of optogenetic tools, including genetically encoded indicators of voltage (GEVIs) or calcium (GECIs) and genetically encoded light-gated ion channels (actuators, e.g., channelrhodopsin2). Compared with low-molecular-weight calcium and voltage indicators (dyes), the optogenetic imaging approaches are 1) cell type specific, 2) less invasive, 3) able to relate activity and anatomy, and 4) facilitate long-term recordings of individual cells' activities over weeks, thereby allowing direct monitoring of the emergence of learned behaviors and underlying circuit mechanisms. We highlight the potential of novel approaches based on GEVIs and compare those to calcium imaging approaches. We also discuss how novel approaches based on GEVIs (and GECIs) coupled with genetically encoded actuators will promote progress in our knowledge of brain circuits and systems. Copyright © 2016 the American Physiological Society.

Voltage imaging to understand connections and functions of neuronal circuits

PubMed Central

Antic, Srdjan D.; Empson, Ruth M.

2016-01-01

Understanding of the cellular mechanisms underlying brain functions such as cognition and emotions requires monitoring of membrane voltage at the cellular, circuit, and system levels. Seminal voltage-sensitive dye and calcium-sensitive dye imaging studies have demonstrated parallel detection of electrical activity across populations of interconnected neurons in a variety of preparations. A game-changing advance made in recent years has been the conceptualization and development of optogenetic tools, including genetically encoded indicators of voltage (GEVIs) or calcium (GECIs) and genetically encoded light-gated ion channels (actuators, e.g., channelrhodopsin2). Compared with low-molecular-weight calcium and voltage indicators (dyes), the optogenetic imaging approaches are 1) cell type specific, 2) less invasive, 3) able to relate activity and anatomy, and 4) facilitate long-term recordings of individual cells' activities over weeks, thereby allowing direct monitoring of the emergence of learned behaviors and underlying circuit mechanisms. We highlight the potential of novel approaches based on GEVIs and compare those to calcium imaging approaches. We also discuss how novel approaches based on GEVIs (and GECIs) coupled with genetically encoded actuators will promote progress in our knowledge of brain circuits and systems. PMID:27075539

Plastic fiber optics for micro-imaging of fluorescence signals in living cells

NASA Astrophysics Data System (ADS)

Sakurai, Takashi; Natsume, Mitsuo; Koida, Kowa

2015-03-01

The fiber-coupled microscope (FCM) enables in vivo imaging at deep sites in the tissues or organs that other optical techniques are unable to reach. To develop FCM-based intravital imaging, we employed a plastic optical fiber (POF) bundle that included more than 10,000-units of polystyrene core and polymethyl methacrylate cladding. Each POF had a diameter of less than 5 μm the tip of the bundle was less than 0.5 mm wide, and the flexible wire had a length of 1,000 mm. The optical performance of the plastic FCM was sufficient for detection of significant signal changes in an acinus of rat pancreas labeled with a calcium ion-sensitive fluorescent dye. In the future, the potential power of plastic FCM is expected to increase, enabling analysis of structure and organization of specific functions in live cells within vulnerable organs.

Multi-layer Cortical Ca2+ Imaging in Freely Moving Mice with Prism Probes and Miniaturized Fluorescence Microscopy

PubMed Central

Gulati, Srishti; Cao, Vania Y.; Otte, Stephani

2017-01-01

In vivo circuit and cellular level functional imaging is a critical tool for understanding the brain in action. High resolution imaging of mouse cortical neurons with two-photon microscopy has provided unique insights into cortical structure, function and plasticity. However, these studies are limited to head fixed animals, greatly reducing the behavioral complexity available for study. In this paper, we describe a procedure for performing chronic fluorescence microscopy with cellular-resolution across multiple cortical layers in freely behaving mice. We used an integrated miniaturized fluorescence microscope paired with an implanted prism probe to simultaneously visualize and record the calcium dynamics of hundreds of neurons across multiple layers of the somatosensory cortex as the mouse engaged in a novel object exploration task, over several days. This technique can be adapted to other brain regions in different animal species for other behavioral paradigms. PMID:28654056

Microstructure of bentonite in iron ore green pellets.

PubMed

Bhuiyan, Iftekhar U; Mouzon, Johanne; Schröppel, Birgit; Kaech, Andres; Dobryden, Illia; Forsmo, Seija P E; Hedlund, Jonas

2014-02-01

Sodium-activated calcium bentonite is used as a binder in iron ore pellets and is known to increase strength of both wet and dry iron ore green pellets. In this article, the microstructure of bentonite in magnetite pellets is revealed for the first time using scanning electron microscopy. The microstructure of bentonite in wet and dry iron ore pellets, as well as in distilled water, was imaged by various imaging techniques (e.g., imaging at low voltage with monochromatic and decelerated beam or low loss backscattered electrons) and cryogenic methods (i.e., high pressure freezing and plunge freezing in liquid ethane). In wet iron ore green pellets, clay tactoids (stacks of parallel primary clay platelets) were very well dispersed and formed a voluminous network occupying the space available between mineral particles. When the pellet was dried, bentonite was drawn to the contact points between the particles and formed solid bridges, which impart strength to the solid compact.

Large-Scale Fluorescence Calcium-Imaging Methods for Studies of Long-Term Memory in Behaving Mammals

PubMed Central

Jercog, Pablo; Rogerson, Thomas; Schnitzer, Mark J.

2016-01-01

During long-term memory formation, cellular and molecular processes reshape how individual neurons respond to specific patterns of synaptic input. It remains poorly understood how such changes impact information processing across networks of mammalian neurons. To observe how networks encode, store, and retrieve information, neuroscientists must track the dynamics of large ensembles of individual cells in behaving animals, over timescales commensurate with long-term memory. Fluorescence Ca2+-imaging techniques can monitor hundreds of neurons in behaving mice, opening exciting avenues for studies of learning and memory at the network level. Genetically encoded Ca2+ indicators allow neurons to be targeted by genetic type or connectivity. Chronic animal preparations permit repeated imaging of neural Ca2+ dynamics over multiple weeks. Together, these capabilities should enable unprecedented analyses of how ensemble neural codes evolve throughout memory processing and provide new insights into how memories are organized in the brain. PMID:27048190

Calcium transport into the cells of the sea urchin larva in relation to spicule formation

PubMed Central

Vidavsky, Netta; Addadi, Sefi; Schertel, Andreas; Ben-Ezra, David; Shpigel, Muki; Addadi, Lia; Weiner, Steve

2016-01-01

We investigated the manner in which the sea urchin larva takes up calcium from its body cavity into the primary mesenchymal cells (PMCs) that are responsible for spicule formation. We used the membrane-impermeable fluorescent dye calcein and alexa-dextran, with or without a calcium channel inhibitor, and imaged the larvae in vivo with selective-plane illumination microscopy. Both fluorescent molecules are taken up from the body cavity into the PMCs and ectoderm cells, where the two labels are predominantly colocalized in particles, whereas the calcium-binding calcein label is mainly excluded from the endoderm and is concentrated in the spicules. The presence of vesicles and vacuoles inside the PMCs that have openings through the plasma membrane directly to the body cavity was documented using high-resolution cryo-focused ion beam-SEM serial imaging. Some of the vesicles and vacuoles are interconnected to form large networks. We suggest that these vacuolar networks are involved in direct sea water uptake. We conclude that the calcium pathway from the body cavity into cells involves nonspecific endocytosis of sea water with its calcium. PMID:27791140

Calcium transport into the cells of the sea urchin larva in relation to spicule formation.

PubMed

Vidavsky, Netta; Addadi, Sefi; Schertel, Andreas; Ben-Ezra, David; Shpigel, Muki; Addadi, Lia; Weiner, Steve

2016-10-24

We investigated the manner in which the sea urchin larva takes up calcium from its body cavity into the primary mesenchymal cells (PMCs) that are responsible for spicule formation. We used the membrane-impermeable fluorescent dye calcein and alexa-dextran, with or without a calcium channel inhibitor, and imaged the larvae in vivo with selective-plane illumination microscopy. Both fluorescent molecules are taken up from the body cavity into the PMCs and ectoderm cells, where the two labels are predominantly colocalized in particles, whereas the calcium-binding calcein label is mainly excluded from the endoderm and is concentrated in the spicules. The presence of vesicles and vacuoles inside the PMCs that have openings through the plasma membrane directly to the body cavity was documented using high-resolution cryo-focused ion beam-SEM serial imaging. Some of the vesicles and vacuoles are interconnected to form large networks. We suggest that these vacuolar networks are involved in direct sea water uptake. We conclude that the calcium pathway from the body cavity into cells involves nonspecific endocytosis of sea water with its calcium.

Detection of microcalcifications by characteristic magnetic susceptibility effects using MR phase image cross-correlation analysis

PubMed Central

Baheza, Richard A.; Welch, E. Brian; Gochberg, Daniel F.; Sanders, Melinda; Harvey, Sara; Gore, John C.; Yankeelov, Thomas E.

2015-01-01

Purpose: To develop and evaluate a new method for detecting calcium deposits using their characteristic magnetic susceptibility effects on magnetic resonance (MR) images at high fields and demonstrate its potential in practice for detecting breast microcalcifications. Methods: Characteristic dipole signatures of calcium deposits were detected in magnetic resonance phase images by computing the cross-correlation between the acquired data and a library of templates containing simulated phase patterns of spherical deposits. The influence of signal-to-noise ratio and various other MR parameters on the results were assessed using simulations and validated experimentally. The method was tested experimentally for detection of calcium fragments within gel phantoms and calcium-like inhomogeneities within chicken tissue at 7 T with optimized MR acquisition parameters. The method was also evaluated for detection of simulated microcalcifications, modeled from biopsy samples of malignant breast cancer, inserted in silico into breast magnetic resonance imaging (MRIs) of healthy subjects at 7 T. For both assessments of calcium fragments in phantoms and biopsy-based simulated microcalcifications in breast MRIs, receiver operator characteristic curve analyses were performed to determine the cross-correlation index cutoff, for achieving optimal sensitivity and specificity, and the area under the curve (AUC), for measuring the method’s performance. Results: The method detected calcium fragments with sizes of 0.14–0.79 mm, 1 mm calcium-like deposits, and simulated microcalcifications with sizes of 0.4–1.0 mm in images with voxel sizes between (0.2 mm)3 and (0.6 mm)3. In images acquired at 7 T with voxel sizes of (0.2 mm)3–(0.4 mm)3, calcium fragments (size 0.3–0.4 mm) were detected with a sensitivity, specificity, and AUC of 78%–90%, 51%–68%, and 0.77%–0.88%, respectively. In images acquired with a human 7 T scanner, acquisition times below 12 min, and voxel sizes of (0.4 mm)3–(0.6 mm)3, simulated microcalcifications with sizes of 0.6–1.0 mm were detected with a sensitivity, specificity, and AUC of 75%–87%, 54%–87%, and 0.76%–0.90%, respectively. However, different microcalcification shapes were indistinguishable. Conclusions: The new method is promising for detecting relatively large microcalcifications (i.e., 0.6–0.9 mm) within the breast at 7 T in reasonable times. Detection of smaller deposits at high field may be possible with higher spatial resolution, but such images require relatively long scan times. Although mammography can detect and distinguish the shape of smaller microcalcifications with superior sensitivity and specificity, this alternative method does not expose tissue to ionizing radiation, is not affected by breast density, and can be combined with other MRI methods (e.g., dynamic contrast-enhanced MRI and diffusion weighted MRI), to potentially improve diagnostic performance. PMID:25735297

Detection of microcalcifications by characteristic magnetic susceptibility effects using MR phase image cross-correlation analysis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Baheza, Richard A.; Welch, E. Brian; Gochberg, Daniel F.

Purpose: To develop and evaluate a new method for detecting calcium deposits using their characteristic magnetic susceptibility effects on magnetic resonance (MR) images at high fields and demonstrate its potential in practice for detecting breast microcalcifications. Methods: Characteristic dipole signatures of calcium deposits were detected in magnetic resonance phase images by computing the cross-correlation between the acquired data and a library of templates containing simulated phase patterns of spherical deposits. The influence of signal-to-noise ratio and various other MR parameters on the results were assessed using simulations and validated experimentally. The method was tested experimentally for detection of calcium fragmentsmore » within gel phantoms and calcium-like inhomogeneities within chicken tissue at 7 T with optimized MR acquisition parameters. The method was also evaluated for detection of simulated microcalcifications, modeled from biopsy samples of malignant breast cancer, inserted in silico into breast magnetic resonance imaging (MRIs) of healthy subjects at 7 T. For both assessments of calcium fragments in phantoms and biopsy-based simulated microcalcifications in breast MRIs, receiver operator characteristic curve analyses were performed to determine the cross-correlation index cutoff, for achieving optimal sensitivity and specificity, and the area under the curve (AUC), for measuring the method’s performance. Results: The method detected calcium fragments with sizes of 0.14–0.79 mm, 1 mm calcium-like deposits, and simulated microcalcifications with sizes of 0.4–1.0 mm in images with voxel sizes between (0.2 mm){sup 3} and (0.6 mm){sup 3}. In images acquired at 7 T with voxel sizes of (0.2 mm){sup 3}–(0.4 mm){sup 3}, calcium fragments (size 0.3–0.4 mm) were detected with a sensitivity, specificity, and AUC of 78%–90%, 51%–68%, and 0.77%–0.88%, respectively. In images acquired with a human 7 T scanner, acquisition times below 12 min, and voxel sizes of (0.4 mm){sup 3}–(0.6 mm){sup 3}, simulated microcalcifications with sizes of 0.6–1.0 mm were detected with a sensitivity, specificity, and AUC of 75%–87%, 54%–87%, and 0.76%–0.90%, respectively. However, different microcalcification shapes were indistinguishable. Conclusions: The new method is promising for detecting relatively large microcalcifications (i.e., 0.6–0.9 mm) within the breast at 7 T in reasonable times. Detection of smaller deposits at high field may be possible with higher spatial resolution, but such images require relatively long scan times. Although mammography can detect and distinguish the shape of smaller microcalcifications with superior sensitivity and specificity, this alternative method does not expose tissue to ionizing radiation, is not affected by breast density, and can be combined with other MRI methods (e.g., dynamic contrast-enhanced MRI and diffusion weighted MRI), to potentially improve diagnostic performance.« less

Calcium homeostasis in intraerythrocytic malaria parasites.

PubMed

Garcia, C R; Dluzewski, A R; Catalani, L H; Burting, R; Hoyland, J; Mason, W T

1996-12-01

The fluorescent indicator, fura-2, AM, was used to measure free calcium concentrations in the intraerythrocytic malaria parasites of Plasmodium chabaudi and Plasmodium falciparum. In both species the free cytosolic calcium concentration was maintained at low levels (between 40 and 100 nM throughout the maturation process. Digital image analysis of the indicator fluorescence was performed on parasites and evaluated with the aid of a calibration of the calcium response, based on permeabilized parasites, exposed to calcium buffers. This again revealed that free calcium concentrations in the intact parasite are maintained at a predetermined level, regardless of the free calcium in the surrounding milieu. Both species of parasites are thus capable of regulating their internal free calcium levels with high precision, presumably by means of calcium pump ATPases. A small but significant elevation of the cytosolic free calcium concentration by the tumor promoter, thapsigargin, may be taken to reflect the presence of calcium stores in the endoplasmic reticulum in P. falciparum.

High-Speed Fluorescence Microscopy: Lifetime Imaging in the Biomedical Sciences

NASA Astrophysics Data System (ADS)

Periasamy, Ammasi; Wang, Xue F.; Wodnick, Pawel; Gordon, Gerald W.; Kwon, Seongwook; Diliberto, Pamela A.; Herman, Brian

1995-02-01

The ability to observe the behavior of living cells and tissues provides unparalleled access to information regarding the organization and dynamics of complex cellular structures. While great strides have been made over the past 30 to 40 years in the design and application of a variety of novel optical microscopic techniques, until recently, it has not been possible to image biological phenomena that occur over very short time periods (nanosecond to millisecond) or over short distances (10 to 1000 [Angstrom capital A, ring]). However, the recent combination of (1) very rapidly gated and sensitive image intensifiers and (2) the ability to deliver fluorescence excitation energy to intact living biological specimens in a pulsed or sinusoidally modulated fashion has allowed such measurements to become a reality through the imaging of the lifetimes of fluorescent molecules. This capability has resulted in the ability to observe the dynamic organization and interaction of cellular components on a spatial and temporal scale previously not possible using other microscopic techniques. This paper discusses the implementation of a fluorescence lifetime imaging microscope (FLIM) and provides a review of some of the applications of such an instrument. These include measurements of receptor topography and subunit interactions using fluorescence resonance energy transfer (FRET), fluorescence anisotropy of phospholipids in cell membranes, cytosolic free calcium (Ca2+)i and the detection of human papillomavirus (HPV) infection in clinical cervicovaginal smears.

Contrasting calcium localization and speciation in leaves of Medicago trunculata mutant COD5 analyzed via synchrotron X-ray techniques

USDA-ARS?s Scientific Manuscript database

Oxalate-producing plants accumulate calcium oxalate crystals (CaOx(C)) in the range of 3-80%(w/w) of their dry weight, reducing calcium (Ca) bioavailability. The calcium oxalate deficient 5 (cod5) mutant of Medicago truncatula has been previously shown to contain similar Ca, but lower oxalate and Ca...

Primary hyperparathyroidism: recent advances.

PubMed

Walker, Marcella D; Bilezikian, John P

2018-07-01

The purpose of this review is to describe recent advances and changes in the evaluation and management of primary hyperparathyroidism (PHPT). Although it has long been recognized that asymptomatic PHPT is associated with bone loss, particularly at cortical skeletal sites when evaluated with dual-energy X-ray absorptiometry, new imaging techniques suggest that trabecular skeletal deterioration as well as clinically silent vertebral fractures and nephrolithiasis are common. Nonclassical targets of asymptomatic PHPT as well as the effect of vitamin D deficiency and treatment upon PHPT presentation have been the subject of recent intense investigation. Randomized clinical trials are now available regarding the effect of parathyroidectomy (PTX) upon both classical and nonclassical target organs. They have confirmed results from observational studies with regard to the skeletal benefits of PTX but have not consistently shown improvements in nonclassical symptoms. These findings have led to recommendations for more extensive renal and skeletal evaluation and broader criteria for PTX in PHPT. In addition to dual-energy X-ray absorptiometry, vertebral and renal imaging is recommended. When available, trabecular imaging techniques may be helpful. PTX criteria now include subclinical kidney stones, vertebral fractures and hypercalciuria, in addition to those based on age, serum calcium, bone densitometry and renal function.

Assessment of calcium scoring performance in cardiac computed tomography.

PubMed

Ulzheimer, Stefan; Kalender, Willi A

2003-03-01

Electron beam tomography (EBT) has been used for cardiac diagnosis and the quantitative assessment of coronary calcium since the late 1980s. The introduction of mechanical multi-slice spiral CT (MSCT) scanners with shorter rotation times opened new possibilities of cardiac imaging with conventional CT scanners. The purpose of this work was to qualitatively and quantitatively evaluate the performance for EBT and MSCT for the task of coronary artery calcium imaging as a function of acquisition protocol, heart rate, spiral reconstruction algorithm (where applicable) and calcium scoring method. A cardiac CT semi-anthropomorphic phantom was designed and manufactured for the investigation of all relevant image quality parameters in cardiac CT. This phantom includes various test objects, some of which can be moved within the anthropomorphic phantom in a manner that mimics realistic heart motion. These tools were used to qualitatively and quantitatively demonstrate the accuracy of coronary calcium imaging using typical protocols for an electron beam (Evolution C-150XP, Imatron, South San Francisco, Calif.) and a 0.5-s four-slice spiral CT scanner (Sensation 4, Siemens, Erlangen, Germany). A special focus was put on the method of quantifying coronary calcium, and three scoring systems were evaluated (Agatston, volume, and mass scoring). Good reproducibility in coronary calcium scoring is always the result of a combination of high temporal and spatial resolution; consequently, thin-slice protocols in combination with retrospective gating on MSCT scanners yielded the best results. The Agatston score was found to be the least reproducible scoring method. The hydroxyapatite mass, being better reproducible and comparable on different scanners and being a physical quantitative measure, appears to be the method of choice for future clinical studies. The hydroxyapatite mass is highly correlated to the Agatston score. The introduced phantoms can be used to quantitatively assess the performance characteristics of, for example, different scanners, reconstruction algorithms, and quantification methods in cardiac CT. This is especially important for quantitative tasks, such as the determination of the amount of calcium in the coronary arteries, to achieve high and constant quality in this field.

Dual-Energy CT: New Horizon in Medical Imaging

PubMed Central

Goo, Jin Mo

2017-01-01

Dual-energy CT has remained underutilized over the past decade probably due to a cumbersome workflow issue and current technical limitations. Clinical radiologists should be made aware of the potential clinical benefits of dual-energy CT over single-energy CT. To accomplish this aim, the basic principle, current acquisition methods with advantages and disadvantages, and various material-specific imaging methods as clinical applications of dual-energy CT should be addressed in detail. Current dual-energy CT acquisition methods include dual tubes with or without beam filtration, rapid voltage switching, dual-layer detector, split filter technique, and sequential scanning. Dual-energy material-specific imaging methods include virtual monoenergetic or monochromatic imaging, effective atomic number map, virtual non-contrast or unenhanced imaging, virtual non-calcium imaging, iodine map, inhaled xenon map, uric acid imaging, automatic bone removal, and lung vessels analysis. In this review, we focus on dual-energy CT imaging including related issues of radiation exposure to patients, scanning and post-processing options, and potential clinical benefits mainly to improve the understanding of clinical radiologists and thus, expand the clinical use of dual-energy CT; in addition, we briefly describe the current technical limitations of dual-energy CT and the current developments of photon-counting detector. PMID:28670151

Live Imaging of Calcium Dynamics during Axon Degeneration Reveals Two Functionally Distinct Phases of Calcium Influx

PubMed Central

Yamagishi, Yuya; Tessier-Lavigne, Marc

2015-01-01

Calcium is a key regulator of axon degeneration caused by trauma and disease, but its specific spatial and temporal dynamics in injured axons remain unclear. To clarify the function of calcium in axon degeneration, we observed calcium dynamics in single injured neurons in live zebrafish larvae and tested the temporal requirement for calcium in zebrafish neurons and cultured mouse DRG neurons. Using laser axotomy to induce Wallerian degeneration (WD) in zebrafish peripheral sensory axons, we monitored calcium dynamics from injury to fragmentation, revealing two stereotyped phases of axonal calcium influx. First, axotomy triggered a transient local calcium wave originating at the injury site. This initial calcium wave only disrupted mitochondria near the injury site and was not altered by expression of the protective WD slow (WldS) protein. Inducing multiple waves with additional axotomies did not change the kinetics of degeneration. In contrast, a second phase of calcium influx occurring minutes before fragmentation spread as a wave throughout the axon, entered mitochondria, and was abolished by WldS expression. In live zebrafish, chelating calcium after the first wave, but before the second wave, delayed the progress of fragmentation. In cultured DRG neurons, chelating calcium early in the process of WD did not alter degeneration, but chelating calcium late in WD delayed fragmentation. We propose that a terminal calcium wave is a key instructive component of the axon degeneration program. SIGNIFICANCE STATEMENT Axon degeneration resulting from trauma or neurodegenerative disease can cause devastating deficits in neural function. Understanding the molecular and cellular events that execute axon degeneration is essential for developing treatments to address these conditions. Calcium is known to contribute to axon degeneration, but its temporal requirements in this process have been unclear. Live calcium imaging in severed zebrafish neurons and temporally controlled pharmacological treatments in both zebrafish and cultured mouse sensory neurons revealed that axonal calcium influx late in the degeneration process regulates axon fragmentation. These findings suggest that temporal considerations will be crucial for developing treatments for diseases associated with axon degeneration. PMID:26558774

Comparative Behaviour of Nitrite and Nitrate for the Protection of Rebar Corrosion

NASA Astrophysics Data System (ADS)

Ahmad, Altaf; Kumar, Anil

2017-10-01

Corrosion of rebar steel due to environmental causes has been studied through various approaches, and among the protection techniques use of inhibitors has gained encouragement. Nitrites and nitrates of sodium have gained sufficient scientific coverage. Recently, nitrites and nitrates of calcium have been verified in some studies, which, however, needs further experimentation through different angles. Simple polarization technique has been utilized in the present study to compare inhibitive efficiency of these salts of sodium and calcium, which indicate that calcium salts are more efficient.

Measurement of Total Calcium in Neurons by Electron Probe X-ray Microanalysis

PubMed Central

Pivovarova, Natalia B.; Andrews, S. Brian

2013-01-01

In this article the tools, techniques, and instruments appropriate for quantitative measurements of intracellular elemental content using the technique known as electron probe microanalysis (EPMA) are described. Intramitochondrial calcium is a particular focus because of the critical role that mitochondrial calcium overload plays in neurodegenerative diseases. The method is based on the analysis of X-rays generated in an electron microscope (EM) by interaction of an electron beam with the specimen. In order to maintain the native distribution of diffusible elements in electron microscopy specimens, EPMA requires "cryofixation" of tissue followed by the preparation of ultrathin cryosections. Rapid freezing of cultured cells or organotypic slice cultures is carried out by plunge freezing in liquid ethane or by slam freezing against a cold metal block, respectively. Cryosections nominally 80 nm thick are cut dry with a diamond knife at ca. -160 °C, mounted on carbon/pioloform-coated copper grids, and cryotransferred into a cryo-EM using a specialized cryospecimen holder. After visual survey and location mapping at ≤-160 °C and low electron dose, frozen-hydrated cryosections are freeze-dried at -100 °C for ~30 min. Organelle-level images of dried cryosections are recorded, also at low dose, by means of a slow-scan CCD camera and subcellular regions of interest selected for analysis. X-rays emitted from ROIs by a stationary, focused, high-intensity electron probe are collected by an energy-dispersive X-ray (EDX) spectrometer, processed by associated electronics, and presented as an X-ray spectrum, that is, a plot of X-ray intensity vs. energy. Additional software facilitates: 1) identification of elemental components by their "characteristic" peak energies and fingerprint; and 2) quantitative analysis by extraction of peak areas/background. This paper concludes with two examples that illustrate typical EPMA applications, one in which mitochondrial calcium analysis provided critical insight into mechanisms of excitotoxic injury and another that revealed the basis of ischemia resistance. PMID:24300079

Optical Imaging of Flow Pattern and Phantom

NASA Technical Reports Server (NTRS)

Galland, Pierre A.; Liang, X.; Wang, L.; Ho, P. P.; Alfano, R. R.; Breisacher, K.

1999-01-01

Time-resolved optical imaging technique has been used to image the spatial distribution of small droplets and jet sprays in a highly scattering environment. The snake and ballistic components of the transmitted pulse are less scattered, and contain direct information about the sample to facilitate image formation as opposed to the diffusive components which are due to multiple collisions as a light pulse propagates through a scattering medium. In a time-gated imaging scheme, these early-arriving, image-bearing components of the incident pulse are selected by opening a gate for an ultrashort period of time and a shadowgram image is detected. Using a single shot cooled CCD camera system, the formation of water droplets is monitored as a function of time. Picosecond time-gated image of drop in scattering cells, spray droplets as a function of let speed and gas pressure, and model calcification samples consisted of calcium carbonate particles of irregular shapes ranging in size from 0. 1 to 1.5 mm affixed to a microscope slide have been measured. Formation produced by an impinging jet will be further monitored using a CCD with 1 kHz framing illuminated with pulsed light. The desired image resolution of the fuel droplets is on the 20 pm scale using early light through a highly scattering medium. A 10(exp -6)m displacement from a jet spray with a flow speed of 100 m/sec introduced by the ns grating pulse used in the imaging is negligible. Early ballistic/snake light imaging offers nondestructive and noninvasive method to observe the spatial distribution of hidden objects inside a highly scattering environment for space, biomedical, and materials applications. In this paper, the techniques we will present are time-resolved K-F transillumination imaging and time-gated scattered light imaging. With a large dynamic range and high resolution, time-gated early light imaging has the potential for improving rocket/aircraft design by determining jets shape and particle sizes. Refinements to these techniques may enable drop size measurements in the highly scattering, optically dense region of multi-element rocket injectors. These types of measurements should greatly enhance the design of stable, and higher performing rocket engines.

Visualizing Calcium Flux in Freely Moving Nematode Embryos.

PubMed

Ardiel, Evan L; Kumar, Abhishek; Marbach, Joseph; Christensen, Ryan; Gupta, Rishi; Duncan, William; Daniels, Jonathan S; Stuurman, Nico; Colón-Ramos, Daniel; Shroff, Hari

2017-05-09

The lack of physiological recordings from Caenorhabditis elegans embryos stands in stark contrast to the comprehensive anatomical and gene expression datasets already available. Using light-sheet fluorescence microscopy to address the challenges associated with functional imaging at this developmental stage, we recorded calcium dynamics in muscles and neurons and developed analysis strategies to relate activity and movement. In muscles, we found that the initiation of twitching was associated with a spreading calcium wave in a dorsal muscle bundle. Correlated activity in muscle bundles was linked with early twitching and eventual coordinated movement. To identify neuronal correlates of behavior, we monitored brainwide activity with subcellular resolution and identified a particularly active cell associated with muscle contractions. Finally, imaging neurons of a well-defined adult motor circuit, we found that reversals in the eggshell correlated with calcium transients in AVA interneurons. Published by Elsevier Inc.

A Neuron-Based Screening Platform for Optimizing Genetically-Encoded Calcium Indicators

PubMed Central

Schreiter, Eric R.; Hasseman, Jeremy P.; Tsegaye, Getahun; Fosque, Benjamin F.; Behnam, Reza; Shields, Brenda C.; Ramirez, Melissa; Kimmel, Bruce E.; Kerr, Rex A.; Jayaraman, Vivek; Looger, Loren L.; Svoboda, Karel; Kim, Douglas S.

2013-01-01

Fluorescent protein-based sensors for detecting neuronal activity have been developed largely based on non-neuronal screening systems. However, the dynamics of neuronal state variables (e.g., voltage, calcium, etc.) are typically very rapid compared to those of non-excitable cells. We developed an electrical stimulation and fluorescence imaging platform based on dissociated rat primary neuronal cultures. We describe its use in testing genetically-encoded calcium indicators (GECIs). Efficient neuronal GECI expression was achieved using lentiviruses containing a neuronal-selective gene promoter. Action potentials (APs) and thus neuronal calcium levels were quantitatively controlled by electrical field stimulation, and fluorescence images were recorded. Images were segmented to extract fluorescence signals corresponding to individual GECI-expressing neurons, which improved sensitivity over full-field measurements. We demonstrate the superiority of screening GECIs in neurons compared with solution measurements. Neuronal screening was useful for efficient identification of variants with both improved response kinetics and high signal amplitudes. This platform can be used to screen many types of sensors with cellular resolution under realistic conditions where neuronal state variables are in relevant ranges with respect to timing and amplitude. PMID:24155972

Sensitive red protein calcium indicators for imaging neural activity

PubMed Central

Dana, Hod; Mohar, Boaz; Sun, Yi; Narayan, Sujatha; Gordus, Andrew; Hasseman, Jeremy P; Tsegaye, Getahun; Holt, Graham T; Hu, Amy; Walpita, Deepika; Patel, Ronak; Macklin, John J; Bargmann, Cornelia I; Ahrens, Misha B; Schreiter, Eric R; Jayaraman, Vivek; Looger, Loren L; Svoboda, Karel; Kim, Douglas S

2016-01-01

Genetically encoded calcium indicators (GECIs) allow measurement of activity in large populations of neurons and in small neuronal compartments, over times of milliseconds to months. Although GFP-based GECIs are widely used for in vivo neurophysiology, GECIs with red-shifted excitation and emission spectra have advantages for in vivo imaging because of reduced scattering and absorption in tissue, and a consequent reduction in phototoxicity. However, current red GECIs are inferior to the state-of-the-art GFP-based GCaMP6 indicators for detecting and quantifying neural activity. Here we present improved red GECIs based on mRuby (jRCaMP1a, b) and mApple (jRGECO1a), with sensitivity comparable to GCaMP6. We characterized the performance of the new red GECIs in cultured neurons and in mouse, Drosophila, zebrafish and C. elegans in vivo. Red GECIs facilitate deep-tissue imaging, dual-color imaging together with GFP-based reporters, and the use of optogenetics in combination with calcium imaging. DOI: http://dx.doi.org/10.7554/eLife.12727.001 PMID:27011354

Specific Adsorption of Osteopontin and Synthetic Polypeptides to Calcium Oxalate Monohydrate Crystals

PubMed Central

Taller, Adam; Grohe, Bernd; Rogers, Kem A.; Goldberg, Harvey A.; Hunter, Graeme K.

2007-01-01

Protein-crystal interactions are known to be important in biomineralization. To study the physicochemical basis of such interactions, we have developed a technique that combines confocal microscopy of crystals with fluorescence imaging of proteins. In this study, osteopontin (OPN), a protein abundant in urine, was labeled with the fluorescent dye AlexaFluor-488 and added to crystals of calcium oxalate monohydrate (COM), the major constituent of kidney stones. In five to seven optical sections along the z axis, scanning confocal microscopy was used to visualize COM crystals and fluorescence imaging to map OPN adsorbed to the crystals. To quantify the relative adsorption to different crystal faces, fluorescence intensity was measured around the perimeter of the crystal in several sections. Using this method, it was shown that OPN adsorbs with high specificity to the edges between {100} and {121} faces of COM and much less so to {100}, {121}, or {010} faces. By contrast, poly-L-aspartic acid adsorbs preferentially to {121} faces, whereas poly-L-glutamic acid adsorbs to all faces approximately equally. Growth of COM in the presence of rat bone OPN results in dumbbell-shaped crystals. We hypothesize that the edge-specific adsorption of OPN may be responsible for the dumbbell morphology of COM crystals found in human urine. PMID:17496021

Fine structures and ion images on fresh frozen dried ultrathin sections by transmission electron and scanning ion microscopy

NASA Astrophysics Data System (ADS)

Takaya, K.; Okabe, M.; Sawataishi, M.; Takashima, H.; Yoshida, T.

2003-01-01

Ion microscopy (IM) of air-dried or freeze-dried cryostat and semi-thin cryosections has provided ion images of elements and organic substances in wide areas of the tissue. For reproducible ion images by a shorter time of exposure to the primary ion beam, fresh frozen dried ultrathin sections were prepared by freezing the tissue in propane chilled with liquid nitrogen, cryocut at 60 nm, mounted on grids and silicon wafer pieces, and freeze-dried. Rat Cowper gland and sciatic nerve, bone marrow of the rat administered of lithium carbonate, tree frog and African toad spleen and buffy coat of atopic dermatitis patients were examined. Fine structures and ion images of the corresponding areas in the same or neighboring sections were observed by transmission electron microscopy (TEM) followed by sector type and time-of-flight type IM. Cells in the buffy coat contained larger amounts of potassium and magnesium while plasma had larger amounts of sodium and calcium. However, in the tissues, lithium, sodium, magnesium, calcium and potassium were distributed in the cell and calcium showed a granular appearance. A granular cell of the tree frog spleen contained sodium and potassium over the cell and magnesium and calcium were confined to granules.

Rapid hydrothermal flow synthesis and characterisation of carbonate- and silicate-substituted calcium phosphates

PubMed Central

Knowles, Jonathan C; Rehman, Ihtesham; Darr, Jawwad A

2013-01-01

A range of crystalline and nano-sized carbonate- and silicate-substituted hydroxyapatite has been successfully produced by using continuous hydrothermal flow synthesis technology. Ion-substituted calcium phosphates are better candidates for bone replacement applications (due to improved bioactivity) as compared to phase-pure hydroxyapatite. Urea was used as a carbonate source for synthesising phase pure carbonated hydroxyapatite (CO3-HA) with ≈5 wt% substituted carbonate content (sample 7.5CO3-HA) and it was found that a further increase in urea concentration in solution resulted in biphasic mixtures of carbonate-substituted hydroxyapatite and calcium carbonate. Transmission electron microscopy images revealed that the particle size of hydroxyapatite decreased with increasing urea concentration. Energy-dispersive X-ray spectroscopy result revealed a calcium deficient apatite with Ca:P molar ratio of 1.45 (±0.04) in sample 7.5CO3-HA. For silicate-substituted hydroxyapatite (SiO4-HA) silicon acetate was used as a silicate ion source. It was observed that a substitution threshold of ∼1.1 wt% exists for synthesis of SiO4-HA in the continuous hydrothermal flow synthesis system, which could be due to the decreasing yields with progressive increase in silicon acetate concentration. All the as-precipitated powders (without any additional heat treatments) were analysed using techniques including Transmission electron microscopy, X-ray powder diffraction, Differential scanning calorimetry, Thermogravimetric analysis, Raman spectroscopy and Fourier transform infrared spectroscopy. PMID:22983020

Application of low-frequency alternating current electric fields via interdigitated electrodes: effects on cellular viability, cytoplasmic calcium, and osteogenic differentiation of human adipose-derived stem cells.

PubMed

McCullen, Seth D; McQuilling, John P; Grossfeld, Robert M; Lubischer, Jane L; Clarke, Laura I; Loboa, Elizabeth G

2010-12-01

Electric stimulation is known to initiate signaling pathways and provides a technique to enhance osteogenic differentiation of stem and/or progenitor cells. There are a variety of in vitro stimulation devices to apply electric fields to such cells. Herein, we describe and highlight the use of interdigitated electrodes to characterize signaling pathways and the effect of electric fields on the proliferation and osteogenic differentiation of human adipose-derived stem cells (hASCs). The advantage of the interdigitated electrode configuration is that cells can be easily imaged during short-term (acute) stimulation, and this identical configuration can be utilized for long-term (chronic) studies. Acute exposure of hASCs to alternating current (AC) sinusoidal electric fields of 1 Hz induced a dose-dependent increase in cytoplasmic calcium in response to electric field magnitude, as observed by fluorescence microscopy. hASCs that were chronically exposed to AC electric field treatment of 1 V/cm (4 h/day for 14 days, cultured in the osteogenic differentiation medium containing dexamethasone, ascorbic acid, and β-glycerol phosphate) displayed a significant increase in mineral deposition relative to unstimulated controls. This is the first study to evaluate the effects of sinusoidal AC electric fields on hASCs and to demonstrate that acute and chronic electric field exposure can significantly increase intracellular calcium signaling and the deposition of accreted calcium under osteogenic stimulation, respectively.

Measuring calcium dynamics in living cells with Genetically Encodable Calcium Indicators

PubMed Central

McCombs, Janet E.

2008-01-01

Genetically encoded calcium indicators (GECIs) allow researchers to measure calcium dynamics in specific targeted locations within living cells. Such indicators enable dissection of the spatial and temporal control of calcium signaling processes. Here we review recent progress in the development of GECIs, highlighting which indicators are most appropriate for measuring calcium in specific organelles and localized domains in mammalian tissue culture cells. An overview of recent approaches that have been undertaken to ensure that the GECIs are minimally perturbed by the cellular environment is provided. Additionally, the procedures for introducing GECIs into mammalian cells, conducting calcium imaging experiments, and analyzing data are discussed. Because organelle-targeted indicators often pose an additional challenge, we underscore strategies for calibrating GECIs in these locations. PMID:18848629

Femtosecond laser induced fixation of calcium alkali phosphate ceramics on titanium alloy bone implant material.

PubMed

Symietz, Christian; Lehmann, Erhard; Gildenhaar, Renate; Krüger, Jörg; Berger, Georg

2010-08-01

Femtosecond lasers provide a novel method of attaching bioceramic material to a titanium alloy, thereby improving the quality of bone implants. The ultrashort 30 fs laser pulses (790 nm wavelength) penetrate a thin dip-coated layer of fine ceramic powder, while simultaneously melting a surface layer of the underlying metal. The specific adjustment of the laser parameters (pulse energy and number of pulses per spot) avoids unnecessary melting of the bioactive calcium phosphate, and permits a defined thin surface melting of the metal, which in turn is not heated throughout, and therefore maintains its mechanical stability. It is essential to choose laser energy densities that correspond to the interval between the ablation fluences of both materials involved: about 0.1-0.4 Jcm(-2). In this work, we present the first results of this unusual technique, including laser ablation studies, scanning electron microscopy and optical microscope images, combined with EDX data. Copyright 2010 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

[Experimental research on the effect of nanophase ceramics on osteoblasts functions].

PubMed

Wen, Bo; Chen, Zhiqing; Jiang, Yinshan; Yang, Zhengwen; Xu, Yongzhong

2005-06-01

In order to study the cytocompatibility of nanophase hydroxyapatite ceramic in vitro, we prepared hydroxyapatite by use of the wet chemistry techniques. The grain size of hydroxyapatite of interest to the present study was determined by scanning electron microscopy and atomic force microscopy with image analysis software. Primary culture of osteoblast from rat calvaria was established. Protein content, synthesis of alkaline phosphatase and deposition of calcium-containing mineral by osteoblasts cultured on nanophase hydroxyapatite ceramics and on conventional hydroxyapatite ceramics for 7, 14, 21 and 28 days were examined. The results showed that the average surface grain size of the nanophase and that of the conventional HA compact formulations was 55 (nanophase) and 780 (conventional) nm, respectively. More importantly, compared to the synthesis of alkaline phosphatase and deposition of calcium-containing mineral by osteoblasts cultured on nanophase was significantly greater than that on conventional ceramics after 21 and 28 days. The cytocompatibility was significantly greater on nanophase HA than on conventional formulations of the same ceramic.

THE COMPARISON OF SEVERAL STANDARD MATERIALS AND TECHNIQUES FOR THE WARREN-AVERBACH DETERMINATION OF MICRO-STRUCTURE CHARACTERISTICS OF CALCIUM HYDROXIDE SORBENT MATERIALS

EPA Science Inventory

The paper gives results of a comparison of several standard materials and techniques for the Warren-Averbach determination of microstructure characteristics of calcium hydroxide--Ca(OH)2--sorbent materials. The comparison is part of an investigation of the injection of dry Ca(OH)...

Present and future of glass-ionomers and calcium-silicate cements as bioactive materials in dentistry: biophotonics-based interfacial analyses in health and disease.

PubMed

Watson, Timothy F; Atmeh, Amre R; Sajini, Shara; Cook, Richard J; Festy, Frederic

2014-01-01

Since their introduction, calcium silicate cements have primarily found use as endodontic sealers, due to long setting times. While similar in chemistry, recent variations such as constituent proportions, purities and manufacturing processes mandate a critical understanding of service behavior differences of the new coronal restorative material variants. Of particular relevance to minimally invasive philosophies is the potential for ion supply, from initial hydration to mature set in dental cements. They may be capable of supporting repair and remineralization of dentin left after decay and cavity preparation, following the concepts of ion exchange from glass ionomers. This paper reviews the underlying chemistry and interactions of glass ionomer and calcium silicate cements, with dental tissues, concentrating on dentin-restoration interface reactions. We additionally demonstrate a new optical technique, based around high resolution deep tissue, two-photon fluorescence and lifetime imaging, which allows monitoring of undisturbed cement-dentin interface samples behavior over time. The local bioactivity of the calcium-silicate based materials has been shown to produce mineralization within the subjacent dentin substrate, extending deep within the tissues. This suggests that the local ion-rich alkaline environment may be more favorable to mineral repair and re-construction, compared with the acidic environs of comparable glass ionomer based materials. The advantages of this potential re-mineralization phenomenon for minimally invasive management of carious dentin are self-evident. There is a clear need to improve the bioactivity of restorative dental materials and these calcium silicate cement systems offer exciting possibilities in realizing this goal. Copyright © 2013 Academy of Dental Materials. Published by Elsevier Ltd. All rights reserved.

Calcium transport into the cells of the sea urchin larva in relation to spicule formation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Vidavsky, Netta; Addadi, Sefi; Schertel, Andreas

We investigated the manner in which the sea urchin larva takes up calcium from its body cavity into the primary mesenchymal cells (PMCs) that are responsible for spicule formation. We used the membrane-impermeable fluorescent dye calcein and alexa-dextran, with or without a calcium channel inhibitor, and imaged the larvae in vivo with selective-plane illumination microscopy. Both fluorescent molecules are taken up from the body cavity into the PMCs and ectoderm cells, where the two labels are predominantly colocalized in particles, whereas the calcium-binding calcein label is mainly excluded from the endoderm and is concentrated in the spicules. The presence ofmore » vesicles and vacuoles inside the PMCs that have openings through the plasma membrane directly to the body cavity was documented using high-resolution cryo-focused ion beam-SEM serial imaging. Some of the vesicles and vacuoles are interconnected to form large networks. We suggest that these vacuolar networks are involved in direct sea water uptake. We conclude that the calcium pathway from the body cavity into cells involves nonspecific endocytosis of sea water with its calcium.« less

Calcium transport into the cells of the sea urchin larva in relation to spicule formation

DOE PAGES

Vidavsky, Netta; Addadi, Sefi; Schertel, Andreas; ...

2016-10-24

We investigated the manner in which the sea urchin larva takes up calcium from its body cavity into the primary mesenchymal cells (PMCs) that are responsible for spicule formation. We used the membrane-impermeable fluorescent dye calcein and alexa-dextran, with or without a calcium channel inhibitor, and imaged the larvae in vivo with selective-plane illumination microscopy. Both fluorescent molecules are taken up from the body cavity into the PMCs and ectoderm cells, where the two labels are predominantly colocalized in particles, whereas the calcium-binding calcein label is mainly excluded from the endoderm and is concentrated in the spicules. The presence ofmore » vesicles and vacuoles inside the PMCs that have openings through the plasma membrane directly to the body cavity was documented using high-resolution cryo-focused ion beam-SEM serial imaging. Some of the vesicles and vacuoles are interconnected to form large networks. We suggest that these vacuolar networks are involved in direct sea water uptake. We conclude that the calcium pathway from the body cavity into cells involves nonspecific endocytosis of sea water with its calcium.« less

Thick, Dark Veins at Garden City, Mars

NASA Image and Video Library

2015-11-11

These images from the Chemistry and Camera (ChemCam) instrument on NASA's Curiosity Mars rover indicate similarly dark material, but with very different chemistries, in mineral veins at "Garden City." Each of the side-by-side circular images covers an area about 2 inches (5 centimeters) in diameter. The images were taken by ChemCam's Remote Micro-Imager. Researchers used ChemCam's laser, telescope and spectrometers to examine the chemistry of material in these veins. While both of these veins are dark, their chemistries are very different, indicating that they were formed by different fluids. One common aspect of the chemistry in the dark material is an iron content higher than nearby bedrock. Thus the dark appearance may be result of similar iron content. The dark maerial in the vein on the left is enriched in calcium and contains calcium fluorine. The dark material in the vein on the right is enriched in magnesium, but not in calcium or calcium fluorine. Thus, the veins were formed by different fluids that deposited minerals in rock fractures. The Remote Micro-Imager took the image on the left on March 27, 2015, during the 938th Martian day, or sol, of Curiosity's work on Mars. The next day, it took the image on the right. A broader view of the prominent mineral veins at Garden City is at PIA19161. ChemCam is one of 10 instruments in Curiosity's science payload. The U.S. Department of Energy's Los Alamos National Laboratory, in Los Alamos, New Mexico, developed ChemCam in partnership with scientists and engineers funded by the French national space agency (CNES), the University of Toulouse and the French national research agency (CNRS). More information about ChemCam is available at http://www.msl-chemcam.com. http://photojournal.jpl.nasa.gov/catalog/PIA19924

Development of optical neuroimaging to detect drug-induced brain functional changes in vivo

NASA Astrophysics Data System (ADS)

Du, Congwu; Pan, Yingtian

2014-03-01

Deficits in prefrontal function play a crucial role in compulsive cocaine use, which is a hallmark of addiction. Dysfunction of the prefrontal cortex might result from effects of cocaine on neurons as well as from disruption of cerebral blood vessels. However, the mechanisms underlying cocaine's neurotoxic effects are not fully understood, partially due to technical limitations of current imaging techniques (e.g., PET, fMRI) to differentiate vascular from neuronal effects at sufficiently high temporal and spatial resolutions. We have recently developed a multimodal imaging platform which can simultaneously characterize the changes in cerebrovascular hemodynamics, hemoglobin oxygenation and intracellular calcium fluorescence for monitoring the effects of cocaine on the brain. Such a multimodality imaging technique (OFI) provides several uniquely important merits, including: 1) a large field-of-view, 2) high spatiotemporal resolutions, 3) quantitative 3D imaging of the cerebral blood flow (CBF) networks, 4) label-free imaging of hemodynamic changes, 5) separation of vascular compartments (e.g., arterial and venous vessels) and monitoring of cortical brain metabolic changes, 6) discrimination of cellular (neuronal) from vascular responses. These imaging features have been further advanced in combination with microprobes to form micro-OFI that allows quantification of drug effects on subcortical brain. In addition, our ultrahigh-resolution ODT (μODT) enables 3D microangiography and quantitative imaging of capillary CBF networks. These optical strategies have been used to investigate the effects of cocaine on brain physiology to facilitate the studies of brain functional changes induced by addictive substance to provide new insights into neurobiological effects of the drug on the brain.

A simple microfluidic device for live cell imaging of Arabidopsis cotyledons, leaves, and seedlings.

PubMed

Vang, Shia; Seitz, Kati; Krysan, Patrick J

2018-06-01

One of the challenges of performing live-cell imaging in plants is establishing a system for securing the sample during imaging that allows for the rapid addition of treatments. Here we report how a commercially available device called a HybriWell ™ can be repurposed to create an imaging chamber suitable for Arabidopsis seedlings, cotyledons and leaves. Liquid in the imaging chamber can be rapidly exchanged to introduce chemical treatments via microfluidic passive pumping. When used in conjunction with fluorescent biosensors, this system can facilitate live-cell imaging studies of signal transduction pathways triggered by different treatments. As a demonstration, we show how the HybriWell can be used to monitor flg22-induced calcium transients using the R-GECO1 calcium indicator in detached Arabidopsis leaves.

Rapid flow-induced responses in endothelial cells

NASA Technical Reports Server (NTRS)

Stamatas, G. N.; McIntire, L. V.

2001-01-01

Endothelial cells alter their morphology, growth rate, and metabolism in response to fluid shear stress. To study rapid flow-induced responses in the 3D endothelial cell morphology and calcium distribution, coupled fluorescence microscopy with optical sectioning, digital imaging, and numerical deconvolution techniques have been utilized. Results demonstrate that within the first minutes of flow application nuclear calcium is increasing. In the same time frame whole cell height and nuclear height are reduced by about 1 microm. Whole cell height changes may facilitate reduction of shear stress gradients on the luminal surface, whereas nuclear structural changes may be important for modulating endothelial growth rate and metabolism. To study the role of the cytoskeleton in these responses, endothelial cells have been treated with specific disrupters (acrylamide, cytochalasin D, and colchicine) of each of the cytoskeleton elements (intermediate filaments, microfilaments, and microtubules, respectively). None of these compounds had any effect on the shear-induced calcium response. Cytochalasin D and acrylamide did not affect the shear-induced nuclear morphology changes. Colchicine, however, completely abrogated the response, indicating that microtubules may be implicated in force transmission from the plasma membrane to the nucleus. A pedagogical model based on tensegrity theory principles is presented that is consistent with the results on the 3D endothelial morphology.

[Acute pancreatitis associated with hypercalcaemia].

PubMed

Tun-Abraham, Mauro Enrique; Obregón-Guerrero, Gabriela; Romero-Espinoza, Larry; Valencia-Jiménez, Javier

2015-01-01

Hypercalcaemia due to primary hyperparathyroidism is a rare cause of acute pancreatitis, with a reported prevalence of 1.5 to 8%. There is no clear pathophysiological basis, but elevated parathyroid hormone and high serum calcium levels could be responsible for calcium deposit in the pancreatic ducts and activation of pancreatic enzymes, which may be the main risk factor for developing acute pancreatitis. The aim of this report is to describe four cases. Four cases are reported of severe pancreatitis associated with hypercalcaemia secondary to primary hyperparathyroidism; three of them with complications (two pseudocysts and one pancreatic necrosis). Cervical ultrasound, computed tomography, and scintigraphy using 99mTc-Sestambi, studies showed the parathyroid adenoma. Surgical resection was the definitive treatment in all four cases. None of the patients had recurrent acute pancreatitis events during follow-up. Acute pancreatitis secondary to hypercalcaemia of primary hyperparathyroidism is rare; however, when it occurs it is associated with severe pancreatitis. It is suspected in patients with elevated serum calcium and high parathyroid hormone levels. Imaging techniques such as cervical ultrasound, computed tomography, and scintigraphy using 99mTc-Sestambi, should be performed, to confirm clinical suspicion. Surgical resection is the definitive treatment with excellent results. Copyright © 2015 Academia Mexicana de Cirugía A.C. Published by Masson Doyma México S.A. All rights reserved.

Close-up View of Homestake Vein

NASA Image and Video Library

2011-12-07

This close-up view of a mineral vein called Homestake comes from the microscopic imager on NASA Mars Exploration Rover Opportunity; the vein is found to be rich in calcium and sulfur, possibly the calcium-sulfate mineral gypsum.

Nitric oxide augments voltage-activated calcium currents of crustacea (Idotea baltica) skeletal muscle.

PubMed

Erxleben, C; Hermann, A

2001-03-16

Invertebrate skeletal muscle contraction is regulated by calcium influx through voltage-dependent calcium channels in the sarcolemmal membrane. In present study we investigated the effects of nitric oxide (NO) donors on calcium currents of single skeletal muscle fibres from the marine isopod, Idotea baltica, using two-electrode voltage clamp recording techniques. The NO donors, S-nitrosocysteine, S-nitroso-N-acetyl-penicillamine or hydroxylamine reversibly increased calcium inward currents in a time dependent manner. The increase of the current was prevented by methylene blue. Our experiments suggest that NO increases calcium inward currents. NO, by acting on calcium ion channels in the sarcolemmal membrane, therefore, may directly be involved in the modulation of muscle contraction.

Advanced Characterization of Rare Earth Elements in Coal Utilization Byproducts

NASA Astrophysics Data System (ADS)

Verba, C.; Scott, M.; Dieterich, M.; Poston, J.; Collins, K.

2016-12-01

Rare earth elements (REE) in various forms (e.g., crystalline mineral phases; adsorbed/absorbed state on and into organic macerals, neoformed glass from flyash or bottom ash) from domestic feedstocks such as coal deposits to coal utilization byproducts (CUB) have the potential to reduce foreign REE dependence and increase domestic resource security. Characterization is critical for understanding environmental risks related to their fate and transport as well as determining the most practical and economical techniques for concentrating the REE and converting them into chemical stocks for manufacturing. Several complementary electron microscopy (SEM-EDS, EPMA-WDS, FIB-SEM, cathodoluminescence, and XRD) and post image processing techniques were used to understand REE transition from coal to CUB. Sites of interest were identified and imaged and respective elemental x-ray maps acquired and montaged. Pixel classification of SEM imagers was completed using image analysis techniques to quantify the distribution of REE associated features. Quantitative elemental analysis of phases were completed using EMPA-WDS followed by FIB-SEM. The FIB-SEM results were reconstructed into 3D volumes and features of interest (e.g. monazite) were analyzed to determine the structure and volumetric estimation of REEs and thus predict detrital REE phases to ICP-MS results. Trace minerals were identified as pyrite, zircon, REE-phosphates' (monazite, xenotime), and barite within the coal tailings. In CUB, amorphous aluminosilicates, iron oxide cenospheres, and calcium oxides were present; monazite appear to be unaltered and unaffected by the combustion process in these samples. Thermal decomposition may have occurred due to presence of detrital zircon and xenotime and subsequent thin Ca-oxide coating enriched in trace REEs.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Solla, E.L., E-mail: esolla@uvigo.es

Herein, we report on the micro- and nanostructure of the calcium phosphate coating produced by pulsed laser deposition (PLD), using focused ion beam (FIB) lamella sample preparation and transmission electron microscopy (TEM) as the characterization technique. The initial selected area electron diffraction (SAED) data demonstrated the presence of hydroxyapatite (HA) over any other possible calcium phosphate crystalline structure and the polycrystalline nature of the coating. Moreover, the SAED analyses showed clear textured ring patterns coherent with the presence of a preferred orientation in the HA nano-crystal growth. The SAED data also indicated that the coating appears to be textured inmore » the 〈002〉 crystalline direction. Dark-field images obtained using 002 as the working reflection showed a clear oriented crystal growth in columns, from bottom to top. These columns have a peculiar arrangement of nano-crystals since, in some cases, the preferred orientation appears to start at a certain distance from the substrate. Direct d-spacing measurements on high-resolution TEM images provided further proof of the presence of an HA nano-crystal structure. The reported data may be of interest in the future to adjust the microstructure of the HA coatings. - Highlights: •The FIB lift-out technique allows a very site-specific sample preparation method for HRTEM analysis. •It also permits a fast assessment of the HA coating thickness and elemental composition (EDS). •The coatings exhibit a nano-crystalline nature, with a texturing effect along the 002 planes. •PLD is suitable for the production of crystalline c-axis oriented hydroxyapatite coatings. •The crystalline HA phase in the PLD coating is very similar to the present in bone.« less

[In vitro study with techniques of imaging of the composition of urinary calculi].

PubMed

Tellez Martínez-Fornés, M; Burgos Revilla, F J; Sáez Garrido, J C; Soria Descalzo, J; Barbero González, J; Sánchez Corral, J; Minaya Minaya, A; Vallejo Herrador, J

1997-02-01

Pre-treatment knowledge of the lithiasic composition can be useful to design the most appropriate therapeutic scheme for each kind of stone. The relationship between the stone's densitometry information provided by the different imaging techniques, conventional radiology (RX), computerized axial tomography (CAT) and dual energy radiographic densitometry (DO) is analyzed, as well as the elemental composition determined by the microanalysis of fragments obtained post-lithotrity using a scanning electronic microscope (SEM) associated to X-ray dispersion energy (XDE). 60 stones, 12 for each pure composition selected (calcium oxalate mono and dihydro, phosphocarbonate, magnesium ammonium phosphate and uric acid), were studied with XR, CAT and DO and were later subjected to lithofragmentation in vitro. Fragments analysis was carried out post-lithotrity with SEM associated to XDE. The X-ray does not allow to establish the composition of some calculi. CAT quantifies the mineral contents of the oxalocalcic and infective calculi and differentiates the uric acid from the other compositions because the mean density values are under 500 Hounsfield Units. DO evaluates the lithiasic content in phosphocarbonate salts which are structurally similar to bone hydroxyapatite.

Visualisation of an nsPEF induced calcium wave using the genetically encoded calcium indicator GCaMP in U87 human glioblastoma cells.

PubMed

Carr, Lynn; Bardet, Sylvia M; Arnaud-Cormos, Delia; Leveque, Philippe; O'Connor, Rodney P

2018-02-01

Cytosolic, synthetic chemical calcium indicators are typically used to visualise the rapid increase in intracellular calcium ion concentration that follows nanosecond pulsed electric field (nsPEF) application. This study looks at the application of genetically encoded calcium indicators (GECIs) to investigate the spatiotemporal nature of nsPEF-induced calcium signals using fluorescent live cell imaging. Calcium responses to 44kV/cm, 10ns pulses were observed in U87-MG cells expressing either a plasma membrane targeted GECI (GCaMP5-G), or one cytosolically expressed (GCaMP6-S), and compared to the response of cells loaded with cytosolic or plasma membrane targeted chemical calcium indicators. Application of 100 pulses, to cells containing plasma membrane targeted indicators, revealed a wave of calcium across the cell initiating at the cathode side. A similar spatial wave was not observed with cytosolic indicators with mobile calcium buffering properties. The speed of the wave was related to pulse application frequency and it was not propagated by calcium induced calcium release. Copyright © 2017 Elsevier B.V. All rights reserved.

An Overview of the Percutaneous Antibiotic Delivery Technique for Osteomyelitis Treatment and a Case Study of Calcaneal Osteomyelitis.

PubMed

Karr, Jeffrey C

2017-11-01

A percutaneous antibiotic delivery technique (PAD-T) used for the adjunctive management of osteomyelitis is presented. This surgical technique incorporates a calcium sulfate and hydroxyapatite (calcium phosphate) bone void filler acting as a carrier vehicle with either an antibiotic or an antifungal medicine, delivering this combination directly into the area of osteomyelitis. The benefit of the PAD-T is reviewed with a case presentation of a successfully treated calcaneal osteomyelitis. No previously reported PAD-T using a simple bone cortex incision in the adjunctive treatment of osteomyelitis has been reported. The PAD-T safely and effectively uses a calcium sulfate and hydroxyapatite bone void filler carrier vehicle to deliver either an antibiotic or an antifungal medicine directly into the area of osteomyelitis.

The role of radiology in the evolution of the understanding of articular disease.

PubMed

Huang, Mingqian; Schweitzer, Mark E

2014-11-01

Both the clinical practice of radiology and the journal Radiology have had an enormous effect on our understanding of articular disease. Early descriptions of osteoarthritis (OA) appeared in Radiology. More recently, advanced physiologic magnetic resonance (MR) techniques have furthered our understanding of the early prestructural changes in patients with OA. Sodium imaging, delayed gadolinium-enhanced MR imaging of cartilage, and spin-lattice relaxation in the rotating frame (or T1ρ) sequences have advanced understanding of the pathophysiology and pathoanatomy of OA. Many pioneering articles on rheumatoid arthritis (RA) also have been published in Radiology. In the intervening decades, our understanding of the natural history of RA has been altered by these articles. Many of the first descriptions of crystalline arthropathies, including gout, calcium pyrophosphate deposition, and hydroxyapatite deposition disease, appeared in Radiology.

Expression of TRPV1 channels by Cajal-Retzius cells and layer-specific modulation of synaptic transmission by capsaicin in the mouse hippocampus.

PubMed

Anstötz, Max; Lee, Sun Kyong; Maccaferri, Gianmaria

2018-05-28

By taking advantage of calcium imaging and electrophysiology, we provide direct pharmacological evidence for the functional expression of TRPV1 channels in hippocampal Cajal-Retzius cells. Application of the TRPV1 activator capsaicin powerfully enhances spontaneous synaptic transmission in the hippocampal layers that are innervated by the axons of Cajal-Retzius cells. Capsaicin-triggered calcium responses and membrane currents in Cajal-Retzius cells, as well as layer-specific modulation of spontaneous synaptic transmission, are absent when the drug is applied to slices prepared from TRPV1 - / - animals. We discuss the implications of the functional expression of TRPV1 channels in Cajal-Retzius cells and of the observed TRPV1-dependent layer-specific modulation of synaptic transmission for physiological and pathological network processing. The vanilloid receptor TRPV1 forms complex polymodal channels that are expressed by sensory neurons and play a critical role in nociception. Their distribution pattern and functions in cortical circuits are, however, much less understood. Although TRPV1 reporter mice have suggested that, in the hippocampus, TRPV1 is predominantly expressed by Cajal-Retzius cells (CRs), direct functional evidence is missing. As CRs powerfully excite GABAergic interneurons of the molecular layers, TRPV1 could play important roles in the regulation of layer-specific processing. Here, we have taken advantage of calcium imaging with the genetically encoded indicator GCaMP6s and patch-clamp techniques to study the responses of hippocampal CRs to the activation of TRPV1 by capsaicin, and have compared the effect of TRPV1 stimulation on synaptic transmission in layers innervated or non-innervated by CRs. Capsaicin induced both calcium responses and membrane currents in ∼50% of the cell tested. Neither increases of intracellular calcium nor whole-cell currents were observed in the presence of the TRPV1 antagonists capsazepine/Ruthenium Red or in slices prepared from TRPV1 knockout mice. We also report a powerful TRPV1-dependent enhancement of spontaneous synaptic transmission onto interneurons with dendritic trees confined to the layers innervated by CRs. In conclusion, our work establishes that functional TRPV1 is expressed by a significant fraction of CRs and we propose that TRPV1 activity may regulate layer-specific synaptic transmission in the hippocampus. Lastly, as CR density decreases during postnatal development, we also propose that functional TRPV1 receptors may be related to mechanisms involved in CR progressive reduction by calcium-dependent toxicity/apoptosis. © 2018 The Authors. The Journal of Physiology © 2018 The Physiological Society.

Analysis of calcium-induced conformational changes in calcium-binding allergens and quantitative determination of their IgE binding properties.

PubMed

Parody, Nuria; Fuertes, Miguel Angel; Alonso, Carlos; Pico de Coaña, Yago

2013-01-01

The polcalcin family is one of the most epidemiologically relevant families of calcium-binding allergens. Polcalcins are potent plant allergens that contain one or several EF-hand motifs and their allergenicity is primarily associated with the Ca(2+)-bound form of the protein. Conformation, stability, as well as IgE recognition of calcium-binding allergens greatly depend on the presence of protein-bound calcium ions. We describe a protocol that uses three techniques (SDS-PAGE, circular dichroism spectroscopy, and ELISA) to describe the effects that calcium has on the structural changes in an allergen and its IgE binding properties.

A correlative and quantitative imaging approach enabling characterization of primary cell-cell communication: Case of human CD4+ T cell-macrophage immunological synapses.

PubMed

Kasprowicz, Richard; Rand, Emma; O'Toole, Peter J; Signoret, Nathalie

2018-05-22

Cell-to-cell communication engages signaling and spatiotemporal reorganization events driven by highly context-dependent and dynamic intercellular interactions, which are difficult to capture within heterogeneous primary cell cultures. Here, we present a straightforward correlative imaging approach utilizing commonly available instrumentation to sample large numbers of cell-cell interaction events, allowing qualitative and quantitative characterization of rare functioning cell-conjugates based on calcium signals. We applied this approach to examine a previously uncharacterized immunological synapse, investigating autologous human blood CD4 + T cells and monocyte-derived macrophages (MDMs) forming functional conjugates in vitro. Populations of signaling conjugates were visualized, tracked and analyzed by combining live imaging, calcium recording and multivariate statistical analysis. Correlative immunofluorescence was added to quantify endogenous molecular recruitments at the cell-cell junction. By analyzing a large number of rare conjugates, we were able to define calcium signatures associated with different states of CD4 + T cell-MDM interactions. Quantitative image analysis of immunostained conjugates detected the propensity of endogenous T cell surface markers and intracellular organelles to polarize towards cell-cell junctions with high and sustained calcium signaling profiles, hence defining immunological synapses. Overall, we developed a broadly applicable approach enabling detailed single cell- and population-based investigations of rare cell-cell communication events with primary cells.

Fast online deconvolution of calcium imaging data

PubMed Central

Zhou, Pengcheng; Paninski, Liam

2017-01-01

Fluorescent calcium indicators are a popular means for observing the spiking activity of large neuronal populations, but extracting the activity of each neuron from raw fluorescence calcium imaging data is a nontrivial problem. We present a fast online active set method to solve this sparse non-negative deconvolution problem. Importantly, the algorithm 3progresses through each time series sequentially from beginning to end, thus enabling real-time online estimation of neural activity during the imaging session. Our algorithm is a generalization of the pool adjacent violators algorithm (PAVA) for isotonic regression and inherits its linear-time computational complexity. We gain remarkable increases in processing speed: more than one order of magnitude compared to currently employed state of the art convex solvers relying on interior point methods. Unlike these approaches, our method can exploit warm starts; therefore optimizing model hyperparameters only requires a handful of passes through the data. A minor modification can further improve the quality of activity inference by imposing a constraint on the minimum spike size. The algorithm enables real-time simultaneous deconvolution of O(105) traces of whole-brain larval zebrafish imaging data on a laptop. PMID:28291787

Programmable Illumination and High-Speed, Multi-Wavelength, Confocal Microscopy Using a Digital Micromirror

PubMed Central

Martial, Franck P.; Hartell, Nicholas A.

2012-01-01

Confocal microscopy is routinely used for high-resolution fluorescence imaging of biological specimens. Most standard confocal systems scan a laser across a specimen and collect emitted light passing through a single pinhole to produce an optical section of the sample. Sequential scanning on a point-by-point basis limits the speed of image acquisition and even the fastest commercial instruments struggle to resolve the temporal dynamics of rapid cellular events such as calcium signals. Various approaches have been introduced that increase the speed of confocal imaging. Nipkov disk microscopes, for example, use arrays of pinholes or slits on a spinning disk to achieve parallel scanning which significantly increases the speed of acquisition. Here we report the development of a microscope module that utilises a digital micromirror device as a spatial light modulator to provide programmable confocal optical sectioning with a single camera, at high spatial and axial resolution at speeds limited by the frame rate of the camera. The digital micromirror acts as a solid state Nipkov disk but with the added ability to change the pinholes size and separation and to control the light intensity on a mirror-by-mirror basis. The use of an arrangement of concave and convex mirrors in the emission pathway instead of lenses overcomes the astigmatism inherent with DMD devices, increases light collection efficiency and ensures image collection is achromatic so that images are perfectly aligned at different wavelengths. Combined with non-laser light sources, this allows low cost, high-speed, multi-wavelength image acquisition without the need for complex wavelength-dependent image alignment. The micromirror can also be used for programmable illumination allowing spatially defined photoactivation of fluorescent proteins. We demonstrate the use of this system for high-speed calcium imaging using both a single wavelength calcium indicator and a genetically encoded, ratiometric, calcium sensor. PMID:22937130

Programmable illumination and high-speed, multi-wavelength, confocal microscopy using a digital micromirror.

PubMed

Martial, Franck P; Hartell, Nicholas A

2012-01-01

Confocal microscopy is routinely used for high-resolution fluorescence imaging of biological specimens. Most standard confocal systems scan a laser across a specimen and collect emitted light passing through a single pinhole to produce an optical section of the sample. Sequential scanning on a point-by-point basis limits the speed of image acquisition and even the fastest commercial instruments struggle to resolve the temporal dynamics of rapid cellular events such as calcium signals. Various approaches have been introduced that increase the speed of confocal imaging. Nipkov disk microscopes, for example, use arrays of pinholes or slits on a spinning disk to achieve parallel scanning which significantly increases the speed of acquisition. Here we report the development of a microscope module that utilises a digital micromirror device as a spatial light modulator to provide programmable confocal optical sectioning with a single camera, at high spatial and axial resolution at speeds limited by the frame rate of the camera. The digital micromirror acts as a solid state Nipkov disk but with the added ability to change the pinholes size and separation and to control the light intensity on a mirror-by-mirror basis. The use of an arrangement of concave and convex mirrors in the emission pathway instead of lenses overcomes the astigmatism inherent with DMD devices, increases light collection efficiency and ensures image collection is achromatic so that images are perfectly aligned at different wavelengths. Combined with non-laser light sources, this allows low cost, high-speed, multi-wavelength image acquisition without the need for complex wavelength-dependent image alignment. The micromirror can also be used for programmable illumination allowing spatially defined photoactivation of fluorescent proteins. We demonstrate the use of this system for high-speed calcium imaging using both a single wavelength calcium indicator and a genetically encoded, ratiometric, calcium sensor.

Non-Invasive In Vivo Imaging of Calcium Signaling in Mice

PubMed Central

Rogers, Kelly L.; Picaud, Sandrine; Roncali, Emilie; Boisgard, Raphaël; Colasante, Cesare; Stinnakre, Jacques; Tavitian, Bertrand; Brûlet, Philippe

2007-01-01

Rapid and transient elevations of Ca2+ within cellular microdomains play a critical role in the regulation of many signal transduction pathways. Described here is a genetic approach for non-invasive detection of localized Ca2+ concentration ([Ca2+]) rises in live animals using bioluminescence imaging (BLI). Transgenic mice conditionally expressing the Ca2+-sensitive bioluminescent reporter GFP-aequorin targeted to the mitochondrial matrix were studied in several experimental paradigms. Rapid [Ca2+] rises inside the mitochondrial matrix could be readily detected during single-twitch muscle contractions. Whole body patterns of [Ca2+] were monitored in freely moving mice and during epileptic seizures. Furthermore, variations in mitochondrial [Ca2+] correlated to behavioral components of the sleep/wake cycle were observed during prolonged whole body recordings of newborn mice. This non-invasive imaging technique opens new avenues for the analysis of Ca2+ signaling whenever whole body information in freely moving animals is desired, in particular during behavioral and developmental studies. PMID:17912353

Inhibition of Prostate Cancer Skeletal Metastases by Targeting Cathepsin K

DTIC Science & Technology

2009-05-01

micro synthetic calcium phosphate thin films coated onto the culture vessels. As a parallel study, a 96-well plate which contained dentin slice...bone resorption in vitro. (A) Representative images of resorption pits on dentin slices or synthetic calcium phosphate thin films are shown. Left...Osteologic Bone cell culture system (BD Bioscience) that consist of sub-micro synthetic calcium phosphate thin films coated on to the culture vessels and

Coronary artery calcium: a multi-institutional, multimanufacturer international standard for quantification at cardiac CT.

PubMed

McCollough, Cynthia H; Ulzheimer, Stefan; Halliburton, Sandra S; Shanneik, Kaiss; White, Richard D; Kalender, Willi A

2007-05-01

To develop a consensus standard for quantification of coronary artery calcium (CAC). A standard for CAC quantification was developed by a multi-institutional, multimanufacturer international consortium of cardiac radiologists, medical physicists, and industry representatives. This report specifically describes the standardization of scan acquisition and reconstruction parameters, the use of patient size-specific tube current values to achieve a prescribed image noise, and the use of the calcium mass score to eliminate scanner- and patient size-based variations. An anthropomorphic phantom containing calibration inserts and additional phantom rings were used to simulate small, medium-size, and large patients. The three phantoms were scanned by using the recommended protocols for various computed tomography (CT) systems to determine the calibration factors that relate measured CT numbers to calcium hydroxyapatite density and to determine the tube current values that yield comparable noise values. Calculation of the calcium mass score was standardized, and the variance in Agatston, volume, and mass scores was compared among CT systems. Use of the recommended scanning parameters resulted in similar noise for small, medium-size, and large phantoms with all multi-detector row CT scanners. Volume scores had greater interscanner variance than did Agatston and calcium mass scores. Use of a fixed calcium hydroxyapatite density threshold (100 mg/cm(3)), as compared with use of a fixed CT number threshold (130 HU), reduced interscanner variability in Agatston and calcium mass scores. With use of a density segmentation threshold, the calcium mass score had the smallest variance as a function of patient size. Standardized quantification of CAC yielded comparable image noise, spatial resolution, and mass scores among different patient sizes and different CT systems and facilitated reduced radiation dose for small and medium-size patients.

Supramolecular structure of the casein micelle.

PubMed

McMahon, D J; Oommen, B S

2008-05-01

The supramolecular structure of colloidal casein micelles in milk was investigated by using a sample preparation protocol based on adsorption of proteins onto a poly-l-lysine and parlodion-coated copper grid, staining of proteins and calcium phosphate by uranyl oxalate, instantaneous freezing, and drying under a high vacuum. High-resolution transmission electron microscopy stereo-images were obtained showing the interior structure of casein micelles. On the basis of our interpretation of these images, an interlocked lattice model was developed in which both casein-calcium phosphate aggregates and casein polymer chains act together to maintain casein micelle integrity. The caseins form linear and branched chains (2 to 5 proteins long) interlocked by the casein-stabilized calcium phosphate nanoclusters. This model suggests that stabilization of calcium phosphate nanoclusters by phosphoserine domains of alpha(s1)-, alpha(s2)-, or beta-casein, or their combination, would orient their hydrophobic domains outward, allowing interaction and binding to other casein molecules. Other interactions between the caseins, such as calcium bridging, could also occur and further stabilize the supramolecule. The combination of having an interlocked lattice structure and multiple interactions results in an open, sponge-like colloidal supramolecule that is resistant to spatial changes and disintegration. Hydrophobic interactions between caseins surrounding a calcium phosphate nanocluster would prevent complete dissociation of casein micelles when the calcium phosphate nanoclusters are solubilized. Likewise, calcium bridging and other electrostatic interactions between caseins would prevent dissociation of the casein micelles into casein-calcium phosphate nanocluster aggregates when milk is cooled or urea is added to milk, and hydrophobic interactions are reduced. The appearance of both polymer chains and small aggregate particles during milk synthesis would also be expected based on this interlocked lattice model of casein micelles, and its supramolecule structure thus exhibits the principles of self-aggregation, interdependence, and diversity observed in nature.

A Low-Cost System Based on Image Analysis for Monitoring the Crystal Growth Process.

PubMed

Venâncio, Fabrício; Rosário, Francisca F do; Cajaiba, João

2017-05-31

Many techniques are used to monitor one or more of the phenomena involved in the crystallization process. One of the challenges in crystal growth monitoring is finding techniques that allow direct interpretation of the data. The present study used a low-cost system, composed of a commercial webcam and a simple white LED (Light Emitting Diode) illuminator, to follow the calcium carbonate crystal growth process. The experiments were followed with focused beam reflectance measurement (FBRM), a common technique for obtaining information about the formation and growth of crystals. The images obtained in real time were treated with the red, blue, and green (RGB) system. The results showed a qualitative response of the system to crystal formation and growth processes, as there was an observed decrease in the signal as the growth process occurred. Control of the crystal growth was managed by increasing the viscosity of the test solution with the addition of monoethylene glycol (MEG) at 30% and 70% in a mass to mass relationship, providing different profiles of the RGB average curves. The decrease in the average RGB value became slower as the concentration of MEG was increased; this reflected a lag in the growth process that was proven by the FBRM.

In vivo Labeling of Constellations of Functionally Identified Neurons for Targeted in vitro Recordings

PubMed Central

Lien, Anthony D.; Scanziani, Massimo

2011-01-01

Relating the functional properties of neurons in an intact organism with their cellular and synaptic characteristics is necessary for a mechanistic understanding of brain function. However, while the functional properties of cortical neurons (e.g., tuning to sensory stimuli) are necessarily determined in vivo, detailed cellular and synaptic analysis relies on in vitro techniques. Here we describe an approach that combines in vivo calcium imaging (for functional characterization) with photo-activation of fluorescent proteins (for neuron labeling), thereby allowing targeted in vitro recording of multiple neurons with known functional properties. We expressed photo-activatable GFP rendered non-diffusible through fusion with a histone protein (H2B–PAGFP) in the mouse visual cortex to rapidly photo-label constellations of neurons in vivo at cellular and sub-cellular resolution using two-photon excitation. This photo-labeling method was compatible with two-photon calcium imaging of neuronal responses to visual stimuli, allowing us to label constellations of neurons with specific functional properties. Photo-labeled neurons were easily identified in vitro in acute brain slices and could be targeted for whole-cell recording. We also demonstrate that in vitro and in vivo image stacks of the same photo-labeled neurons could be registered to one another, allowing the exact in vivo response properties of individual neurons recorded in vitro to be known. The ability to perform in vitro recordings from neurons with known functional properties opens up exciting new possibilities for dissecting the cellular, synaptic, and circuit mechanisms that underlie neuronal function in vivo. PMID:22144948

The Relationship between Membrane Potential and Calcium Dynamics in Glucose-Stimulated Beta Cell Syncytium in Acute Mouse Pancreas Tissue Slices

PubMed Central

Miller, Evan W.; Slak Rupnik, Marjan

2013-01-01

Oscillatory electrical activity is regarded as a hallmark of the pancreatic beta cell glucose-dependent excitability pattern. Electrophysiologically recorded membrane potential oscillations in beta cells are associated with in-phase oscillatory cytosolic calcium activity ([Ca2+]i) measured with fluorescent probes. Recent high spatial and temporal resolution confocal imaging revealed that glucose stimulation of beta cells in intact islets within acute tissue slices produces a [Ca2+]i change with initial transient phase followed by a plateau phase with highly synchronized [Ca2+]i oscillations. Here, we aimed to correlate the plateau [Ca2+]i oscillations with the oscillations of membrane potential using patch-clamp and for the first time high resolution voltage-sensitive dye based confocal imaging. Our results demonstrated that the glucose-evoked membrane potential oscillations spread over the islet in a wave-like manner, their durations and wave velocities being comparable to the ones for [Ca2+]i oscillations and waves. High temporal resolution simultaneous records of membrane potential and [Ca2+]i confirmed tight but nevertheless limited coupling of the two processes, with membrane depolarization preceding the [Ca2+]i increase. The potassium channel blocker tetraethylammonium increased the velocity at which oscillations advanced over the islet by several-fold while, at the same time, emphasized differences in kinetics of the membrane potential and the [Ca2+]i. The combination of both imaging techniques provides a powerful tool that will help us attain deeper knowledge of the beta cell network. PMID:24324777

Effect of Mitral Annular Calcium on Left Ventricular Diastolic Parameters.

PubMed

Codolosa, Jose N; Koshkelashvili, Nikoloz; Alnabelsi, Talal; Goykhman, Igor; Romero-Corral, Abel; Pressman, Gregg S

2016-03-01

Assessment of left ventricular (LV) diastolic function by Doppler flow imaging and tissue Doppler is an integral part of the echocardiographic examination. Mitral annular calcium (MAC) is frequently encountered on echocardiography. The aim of this study was to assess the impact of MAC, quantitatively measured by computed tomography scan, on echocardiographic LV diastolic parameters. We included 155 patients aged ≥65 years. Computed tomography reconstructions of the mitral annulus were created, and calcium identified and quantified by Agatston technique. Calcium locations were assigned using an overlaid template depicting the annular segments in relation to surrounding anatomic structures. Echocardiographic assessment of diastolic function was performed in standard fashion. Mean age was 77 years; 49% were men; and 43% were black. Patients with MAC had lower septal e' (p = 0.003), lateral e' (p = 0.04), and average e' (p = 0.01) compared with those without MAC. They also had a higher E-wave velocity (p = 0.01) and E/e' ratio (p <0.001). When evaluated by severity of MAC, and after adjustment for multiple clinical factors, there was a graded (inverse) relation between MAC severity and septal e' (p = 0.01), lateral e' (p = 0.01), and average e' (p = 0.01). In conclusion, LV diastolic parameters, as measured by Doppler echocardiography, are altered in the presence of MAC. This could be due to direct effects of MAC on annular function or might reflect truly reduced diastolic function. Interpretation of diastolic parameters in patients with MAC should be performed with caution. Copyright © 2016 Elsevier Inc. All rights reserved.

Calcium Sulfate Characterized by Chemcam/Curiousity at Gale Crater, Mars

NASA Technical Reports Server (NTRS)

Nachon, M.; Clegg, S. M.; Mangold, N.; Schroeder, S.; Kah, L. C.; Dromart, G.; Ollila, A.; Johnson, J. R; Oehler, D. Z.; Bridges, J. C.;

Imaging calcium carbonate distribution in human sweat pore in vivo using nonlinear microscopy

NASA Astrophysics Data System (ADS)

Chen, Xueqin; Gasecka, Alicja; Formanek, Florian; Galey, Jean-Baptiste; Rigneault, Hervé

2015-03-01

Nonlinear microscopies, including two-photon excited autofluorescence (TPEF) and coherent anti-Stokes Raman scattering (CARS), were used to study individual human sweat pore morphology and topically applied antiperspirant salt penetration inside sweat pore, in vivo on human palms. Sweat pore inner morphology in vivo was imaged up to the depth of 100 μm by TPEF microscopy. The 3D penetration and distribution of "in situ calcium carbonate" (isCC), an antiperspirant salt model, was investigated using CARS microscopy.

In vivo wide-field calcium imaging of mouse thalamocortical synapses with an 8 K ultra-high-definition camera.

PubMed

Yoshida, Eriko; Terada, Shin-Ichiro; Tanaka, Yasuyo H; Kobayashi, Kenta; Ohkura, Masamichi; Nakai, Junichi; Matsuzaki, Masanori

2018-05-29

In vivo wide-field imaging of neural activity with a high spatio-temporal resolution is a challenge in modern neuroscience. Although two-photon imaging is very powerful, high-speed imaging of the activity of individual synapses is mostly limited to a field of approximately 200 µm on a side. Wide-field one-photon epifluorescence imaging can reveal neuronal activity over a field of ≥1 mm 2 at a high speed, but is not able to resolve a single synapse. Here, to achieve a high spatio-temporal resolution, we combine an 8 K ultra-high-definition camera with spinning-disk one-photon confocal microscopy. This combination allowed us to image a 1 mm 2 field with a pixel resolution of 0.21 µm at 60 fps. When we imaged motor cortical layer 1 in a behaving head-restrained mouse, calcium transients were detected in presynaptic boutons of thalamocortical axons sparsely labeled with GCaMP6s, although their density was lower than when two-photon imaging was used. The effects of out-of-focus fluorescence changes on calcium transients in individual boutons appeared minimal. Axonal boutons with highly correlated activity were detected over the 1 mm 2 field, and were probably distributed on multiple axonal arbors originating from the same thalamic neuron. This new microscopy with an 8 K ultra-high-definition camera should serve to clarify the activity and plasticity of widely distributed cortical synapses.

Calcium phosphate coatings on magnesium alloys for biomedical applications: a review.

PubMed

Shadanbaz, Shaylin; Dias, George J

2012-01-01

Magnesium has been suggested as a revolutionary biodegradable metal for use as an orthopaedic material. As a biocompatible and degradable metal, it has several advantages over the permanent metallic materials currently in use, including eliminating the effects of stress shielding, improving biocompatibility concerns in vivo and improving degradation properties, removing the requirement of a second surgery for implant removal. The rapid degradation of magnesium, however, is a double-edged sword as it is necessary to control the corrosion rates of the materials to match the rates of bone healing. In response, calcium phosphate coatings have been suggested as a means to control these corrosion rates. The potential calcium phosphate phases and their coating techniques on substrates are numerous and can provide several different properties for different applications. The reactivity and low melting point of magnesium, however, require specific parameters for calcium phosphate coatings to be successful. Within this review, an overview of the different calcium phosphate phases, their properties and their behaviour in vitro and in vivo has been provided, followed by the current coating techniques used for calcium phosphates that may be or may have been adapted for magnesium substrates. Copyright © 2011 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

A prospective evaluation of laparoscopic exploration with intraoperative ultrasound as a technique for localizing sporadic insulinomas.

PubMed

Grover, Amelia C; Skarulis, Monica; Alexander, H Richard; Pingpank, James F; Javor, Edward D; Chang, Richard; Shawker, Thomas; Gorden, Phil; Cochran, Craig; Libutti, Steven K

2005-12-01

Preoperative imaging studies localize insulinomas in less than 50% of patients. Arteriography with calcium stimulation and venous sampling (ASVS) regionalizes greater than 90% of insulinomas but requires specialized expertise and an invasive procedure. This prospective study evaluated laparoscopic exploration with IOUS compared with the other localization procedures in patients with a sporadic insulinoma. Between March 2001 and October 2004, 14 patients (7 women and 7 men; mean age, 53) with an insulinoma were enrolled in an IRB-approved protocol. Computed tomography, magnetic resonance imaging, ultrasound scan, and arteriography with calcium stimulation and venous sampling were performed preoperatively. A surgeon, blinded to the results of the localizing studies, performed a laparoscopic exploration with intraoperative ultrasound (IOUS). At the completion of the exploration, the success of laparoscopy for localization was scored, and the tumor was resected. Twelve of 14 tumors were localized successfully before laparoscopy (noninvasive, 7 of 14; invasive, 11 of 14). Laparoscopic IOUS localized successfully 12 of 14 tumors. All lesions were resected, and all patients were cured (median follow-up, 36 months). Laparoscopic IOUS identified 86% of tumors. The authors consider laparoscopic IOUS to be equivalent to ASVS in localizing insulinomas. Further study is therefore warranted to determine the role of laparoscopy with IOUS in the localization and treatment algorithm for patients with sporadic insulinoma.

Community-based benchmarking improves spike rate inference from two-photon calcium imaging data.

PubMed

Berens, Philipp; Freeman, Jeremy; Deneux, Thomas; Chenkov, Nikolay; McColgan, Thomas; Speiser, Artur; Macke, Jakob H; Turaga, Srinivas C; Mineault, Patrick; Rupprecht, Peter; Gerhard, Stephan; Friedrich, Rainer W; Friedrich, Johannes; Paninski, Liam; Pachitariu, Marius; Harris, Kenneth D; Bolte, Ben; Machado, Timothy A; Ringach, Dario; Stone, Jasmine; Rogerson, Luke E; Sofroniew, Nicolas J; Reimer, Jacob; Froudarakis, Emmanouil; Euler, Thomas; Román Rosón, Miroslav; Theis, Lucas; Tolias, Andreas S; Bethge, Matthias

2018-05-01

In recent years, two-photon calcium imaging has become a standard tool to probe the function of neural circuits and to study computations in neuronal populations. However, the acquired signal is only an indirect measurement of neural activity due to the comparatively slow dynamics of fluorescent calcium indicators. Different algorithms for estimating spike rates from noisy calcium measurements have been proposed in the past, but it is an open question how far performance can be improved. Here, we report the results of the spikefinder challenge, launched to catalyze the development of new spike rate inference algorithms through crowd-sourcing. We present ten of the submitted algorithms which show improved performance compared to previously evaluated methods. Interestingly, the top-performing algorithms are based on a wide range of principles from deep neural networks to generative models, yet provide highly correlated estimates of the neural activity. The competition shows that benchmark challenges can drive algorithmic developments in neuroscience.

Measuring and Modeling Sonoporation Dynamics in Mammalian Cells via Calcium Imaging

NASA Astrophysics Data System (ADS)

Kumon, R. E.; Parikh, P.; Sabens, D.; Aehle, M.; Kourennyi, D.; Deng, C. X.

2007-05-01

In this study, calcium imaging via the fluorescent indicator Fura-2 is used to characterize the sonoporation of Chinese Hamster Ovarian (CHO) cells in the presence of Optison™ microbubbles. Evolution of the calcium concentration within cells is determined from real-time fluorescence intensity measurements before, during, and after exposure to a 1 MHz ultrasound tone burst (0.2 s, 0.45 MPa). To relate microscopic sonoporation parameters to the measurements, an analytical model that includes sonoporation and plasma membrane transport is developed, assuming rapid mixing (uniform spatial distribution) in the cell. Fitting the measured data to the model provides estimated values for the poration area as a function of poration relaxation rate as well as plasma membrane pump and leakage rates. A modified compartment model that includes the effects of sonoporation, buffering proteins, and transport across the plasma membrane, endoplasmic reticulum, and mitochondria is also investigated. Numerical 3solutions of this model show a variety of behaviors for the calcium dynamics of the cell.

Calcium imaging of neural circuits with extended depth-of-field light-sheet microscopy

PubMed Central

Quirin, Sean; Vladimirov, Nikita; Yang, Chao-Tsung; Peterka, Darcy S.; Yuste, Rafael; Ahrens, Misha B.

2016-01-01

Increasing the volumetric imaging speed of light-sheet microscopy will improve its ability to detect fast changes in neural activity. Here, a system is introduced for brain-wide imaging of neural activity in the larval zebrafish by coupling structured illumination with cubic phase extended depth-of-field (EDoF) pupil encoding. This microscope enables faster light-sheet imaging and facilitates arbitrary plane scanning—removing constraints on acquisition speed, alignment tolerances, and physical motion near the sample. The usefulness of this method is demonstrated by performing multi-plane calcium imaging in the fish brain with a 416 × 832 × 160 µm field of view at 33 Hz. The optomotor response behavior of the zebrafish is monitored at high speeds, and time-locked correlations of neuronal activity are resolved across its brain. PMID:26974063

Effects of process variables on the encapsulation of oil in ca-alginate capsules using an inverse gelation technique.

PubMed

Abang, Sariah; Chan, Eng-Seng; Poncelet, Denis

2012-01-01

The objective of this study was to investigate the effects of process variables on the encapsulation of oil in a calcium alginate membrane using an inverse gelation technique. A dispersion of calcium chloride solution in sunflower oil (water-in-oil emulsion) was added dropwise to the alginate solution. The migration of calcium ions to the alginate solution initiates the formation of a ca-alginate membrane around the emulsion droplets. The membrane thickness of wet capsules and the elastic modulus of dry capsules increased following first-order kinetics with an increasing curing time. An increase in the calcium chloride concentration increased the membrane thickness of wet capsules and the elastic modulus of dry capsules. An increase in the alginate concentration decreased the mean diameter of wet capsules but increased the elastic modulus of dry capsules.

Two-photon imaging of spatially extended neuronal network dynamics with high temporal resolution.

PubMed

Lillis, Kyle P; Eng, Alfred; White, John A; Mertz, Jerome

2008-07-30

We describe a simple two-photon fluorescence imaging strategy, called targeted path scanning (TPS), to monitor the dynamics of spatially extended neuronal networks with high spatiotemporal resolution. Our strategy combines the advantages of mirror-based scanning, minimized dead time, ease of implementation, and compatibility with high-resolution low-magnification objectives. To demonstrate the performance of TPS, we monitor the calcium dynamics distributed across an entire juvenile rat hippocampus (>1.5mm), at scan rates of 100 Hz, with single cell resolution and single action potential sensitivity. Our strategy for fast, efficient two-photon microscopy over spatially extended regions provides a particularly attractive solution for monitoring neuronal population activity in thick tissue, without sacrificing the signal-to-noise ratio or high spatial resolution associated with standard two-photon microscopy. Finally, we provide the code to make our technique generally available.

Influence of calcium oxide level and time of exposure to sugarcane on in vitro and in situ digestion kinetics

USDA-ARS?s Scientific Manuscript database

Experiments were carried out to evaluate, using in vitro and in situ techniques, the effects of three inclusion levels of calcium oxide (0, 5, and 10 g/kg of sugarcane fresh matter) and four exposure times (0, 24, 48, and 72 h) of sugarcane to calcium oxide on the chemical composition and digestive ...

Calcium influx through the TRPV1 channel of endothelial cells (ECs) correlates with a stronger adhesion between monocytes and ECs.

PubMed

Himi, N; Hamaguchi, A; Hashimoto, K; Koga, T; Narita, K; Miyamoto, O

2012-01-01

Atherosclerosis is thought to be initiated by the transendothelial migration of monocytes. In the early stage of this process, the adhesion of monocytes to endothelial cells is supported by an increase in the intracellular concentration of calcium ion ([Ca(2+)]i) in endothelial cells. However, the main source of Ca(2+) has been unclear. In this study, the changes in ionic transmittance and [Ca(2+)]i due to the adhesion of monocytes were continuously measured by an electrophysiological technique and fluorescent imaging. Especially, we focused on transient receptor potential vanilloid channel 1 (TRPV1) as a Ca(2+) channel that could influence the adhesion of monocytes. Whole-cell current was continuously recorded in human umbilical vein endothelial cells (HUVECs) by a patch electrode. The adhesion of monocytes (THP-1) induced a transient inward current in HUVECs, as well as an elevation of [Ca(2+)]i. This inward element was abolished by the application of 100 nM SB366,791, a selective antagonist of TRPV1 channel. Furthermore, SB366,791 significantly decreased the number of THP-1 cells that adhered to HUVECs (control: 231 ± 38, SB366,791: 96 ± 16 cells/mm2). These results suggest that an inward calcium current via the TRPV1 channels of endothelial cells correlates with a stronger adhesion between monocytes and endothelial cells.

Synaptically evoked Ca2+ release from intracellular stores is not influenced by vesicular zinc in CA3 hippocampal pyramidal neurones.

PubMed

Evstratova, Alesya; Tóth, Katalin

2011-12-01

The co-release of neuromodulatory substances in combination with classic neurotransmitters such as glutamate and GABA from individual presynaptic nerve terminals has the capacity to dramatically influence synaptic efficacy and plasticity. At hippocampal mossy fibre synapses vesicular zinc is suggested to serve as a cotransmitter capable of regulating calcium release from internal stores in postsynaptic CA3 pyramidal cells. Here we investigated this possibility using combined intracellular ratiometric calcium imaging and patch-clamp recording techniques. In acute hippocampal slices a brief train of mossy fibre stimulation produced a large, delayed postsynaptic Ca(2+) wave that was spatially restricted to the proximal apical dendrites of CA3 pyramidal cells within stratum lucidum. This calcium increase was sensitive to intracellularly applied heparin indicating reliance upon release from internal stores and was triggered by activation of both group I metabotropic glutamate and NMDA receptors. Importantly, treatment of slices with the membrane-impermeant zinc chelator CaEDTA did not influence the synaptically evoked postsynaptic Ca(2+) waves. Moreover, mossy fibre stimulus evoked postsynaptic Ca(2+) signals were not significantly different between wild-type and zinc transporter 3 (ZnT3) knock-out animals. Considered together our data do not support a role for vesicular zinc in regulating mossy fibre evoked Ca(2+) release from CA3 pyramidal cell internal stores.

Synaptically evoked Ca2+ release from intracellular stores is not influenced by vesicular zinc in CA3 hippocampal pyramidal neurones

PubMed Central

Evstratova, Alesya; Tóth, Katalin

2011-01-01

Abstract The co-release of neuromodulatory substances in combination with classic neurotransmitters such as glutamate and GABA from individual presynaptic nerve terminals has the capacity to dramatically influence synaptic efficacy and plasticity. At hippocampal mossy fibre synapses vesicular zinc is suggested to serve as a cotransmitter capable of regulating calcium release from internal stores in postsynaptic CA3 pyramidal cells. Here we investigated this possibility using combined intracellular ratiometric calcium imaging and patch-clamp recording techniques. In acute hippocampal slices a brief train of mossy fibre stimulation produced a large, delayed postsynaptic Ca2+ wave that was spatially restricted to the proximal apical dendrites of CA3 pyramidal cells within stratum lucidum. This calcium increase was sensitive to intracellularly applied heparin indicating reliance upon release from internal stores and was triggered by activation of both group I metabotropic glutamate and NMDA receptors. Importantly, treatment of slices with the membrane-impermeant zinc chelator CaEDTA did not influence the synaptically evoked postsynaptic Ca2+ waves. Moreover, mossy fibre stimulus evoked postsynaptic Ca2+ signals were not significantly different between wild-type and zinc transporter 3 (ZnT3) knock-out animals. Considered together our data do not support a role for vesicular zinc in regulating mossy fibre evoked Ca2+ release from CA3 pyramidal cell internal stores. PMID:21986206

An overview of techniques for the measurement of calcium distribution, calcium fluxes, and cytosolic free calcium in mammalian cells.

PubMed Central

Borle, A B

1990-01-01

An array of techniques can be used to study cell calcium metabolism that comprises several calcium compartments and many types of transport systems such as ion channels, ATP-dependent pumps, and antiporters. The measurement of total cell calcium brings little information of value since 60 to 80% of total cell calcium is actually bound to the extracellular glycocalyx. Cell fractionation and differential centrifugation have been used to study intracellular Ca2+ compartmentalization, but the methods suffer from the possibility of Ca2+ loss or redistribution among cell fractions. Steady-state kinetic analyses of 45Ca uptake or desaturation curves have been used to study the distribution of Ca2+ among various kinetic pools in living cells and their rate of Ca2+ exchange, but the analyses are constrained by many limitations. Nonsteady-state tracer studies can provide information about rapid changes in calcium influx or efflux in and out of the cell. Zero-time kinetics of 45Ca uptake can detect instantaneous changes in calcium influx, while 45Ca fractional efflux ratio, can detect rapid stimulations or inhibitions of calcium efflux out of cells. Permeabilized cells have been successfully used to gauge the relative role of intracellular organelles in controlling [Ca2+]i. The measurement of the cytosolic ionized calcium ([Ca2+]i) is undoubtedly the most important and, physiologically, the most relevant method available. The choice of the appropriate calcium indicator, fluorescent, bioluminescent, metallochromic, or Ca2(+)-sensitive microelectrodes depends on the cell type and the magnitude and time constant of the event under study. Each probe has specific assets and drawbacks. The study of plasma membrane vesicles derived from baso-lateral or apical plasmalemma can also bring important information on the (Ca2(+)-Mg2+) ATPase-dependent calcium pump and on the kinetics and stoichiometry of the Na(+)-Ca2+ antiporter. The best strategy to study cell calcium metabolism is to use several different methods that focus on a specific problem from widely different angles. PMID:2190818

CalQuo: automated, simultaneous single-cell and population-level quantification of global intracellular Ca2+ responses.

PubMed

Fritzsche, Marco; Fernandes, Ricardo A; Colin-York, Huw; Santos, Ana M; Lee, Steven F; Lagerholm, B Christoffer; Davis, Simon J; Eggeling, Christian

2015-11-13

Detecting intracellular calcium signaling with fluorescent calcium indicator dyes is often coupled with microscopy techniques to follow the activation state of non-excitable cells, including lymphocytes. However, the analysis of global intracellular calcium responses both at the single-cell level and in large ensembles simultaneously has yet to be automated. Here, we present a new software package, CalQuo (Calcium Quantification), which allows the automated analysis and simultaneous monitoring of global fluorescent calcium reporter-based signaling responses in up to 1000 single cells per experiment, at temporal resolutions of sub-seconds to seconds. CalQuo quantifies the number and fraction of responding cells, the temporal dependence of calcium signaling and provides global and individual calcium-reporter fluorescence intensity profiles. We demonstrate the utility of the new method by comparing the calcium-based signaling responses of genetically manipulated human lymphocytic cell lines.

Calcium-pH crosstalks in rat mast cells: cytosolic alkalinization, but not intracellular calcium release, is a sufficient signal for degranulation

PubMed Central

Alfonso, A; Cabado, A G; Vieytes, M R; Botana, L M

2000-01-01

The aim of this work was to study the relationship between intracellular alkalinization, calcium fluxes and histamine release in rat mast cells. Intracellular alkalinization was induced by nigericin, a monovalent cation ionophore, and by NH4Cl (ammonium chloride). Calcium cytosolic and intracellular pH were measured by fluorescence digital imaging using Fura-2-AM and BCECF-AM.In rat mast cells, nigericin and NH4Cl induce a dose-dependent intracellular alkalinization, a dose-dependent increase in intracellular calcium levels by releasing calcium from intracellular pools, and an activation of capacitative calcium influx.The increase in both intracellular calcium and pH activates exocytosis (histamine release) in the absence of external calcium. Under the same conditions, thapsigargin does not activate exocytosis, the main difference being that thapsigargin does not alkalinize the cytosol.After alkalinization, histamine release is intracellular-calcium dependent. With 2.5 mM EGTA and thapsigargin the cell response decreases by 62%.The cytosolic alkalinization, in addition to the calcium increase it is enough signal to elicit the exocytotic process in rat mast cells. PMID:10952669

Synthesis of Calcite Nano Particles from Natural Limestone assisted with Ultrasonic Technique

NASA Astrophysics Data System (ADS)

Handayani, M.; Sulistiyono, E.; Firdiyono, F.; Fajariani, E. N.

2018-03-01

This article represents a precipitation method assisted with ultrasonic process to synthesize precipitated calcium carbonate nano particles from natural limestone. The synthesis of nanoparticles material of precipitated calcium carbonate from commercial calcium carbonate was done for comparison. The process was performed using ultrasonic waves at optimum condition, that is, at temperature of 80oC for 10 minutes with various amplitudes. Synthesized precipitated calcium carbonate nanoparticles were characterized using X-Ray Diffraction (XRD), Scanning Electron Microscope (SEM) and Particle Size Analyzer (PSA). The result of PSA measurements showed that precipitated calcium carbonate nano particles was obtained with the average size of 109 nm.

Influence of water vapour and carbon dioxide on free lime during storage at 80 °C, studied by Raman spectroscopy.

PubMed

Dubina, E; Korat, L; Black, L; Strupi-Šuput, J; Plank, J

2013-07-01

Micro-Raman spectroscopy has been used to follow the reaction of free lime (CaO) exposed for 24h to moist air at 80 °C under conditions of different relative humidities (10-80% RH). X-ray diffraction and SEM imaging were applied as complementary techniques. The conversion of lime to calcium hydroxide and its subsequent carbonation to various calcium carbonate polymorphs was found to strongly depend on the relative humidity. At low RH (10-20%), only Raman spectroscopy revealed the formation of early amorphous CaCO3 which in the XRD patterns was detected only at ≥40% RH. However, XRD analysis could identify the crystalline polymorphs formed at higher relative humidities. Thus, between 20 and 60% RH, all three CaCO3 polymorphs (calcite, aragonite and vaterite) were observed via XRD whereas at high relative humidity (80%), calcite was the predominant reaction product. The results demonstrate the usefulness of Raman spectroscopy in the study of minor cement constituents and their reaction products on air, especially of amorphous character. Copyright © 2013 Elsevier B.V. All rights reserved.

Photolysis of caged calcium in femtoliter volumes using two-photon excitation.

PubMed Central

Brown, E B; Shear, J B; Adams, S R; Tsien, R Y; Webb, W W

1999-01-01

A new technique for the determination of the two-photon uncaging action cross section (deltau) of photolyzable calcium cages is described. This technique is potentially applicable to other caged species that can be chelated by a fluorescent indicator dye, as well as caged fluorescent compounds. The two-photon action cross sections of three calcium cages, DM-nitrophen, NP-EGTA, and azid-1, are studied in the range of excitation wavelengths between 700 and 800 nm. Azid-1 has a maximum deltau of approximately 1.4 GM at 700 nm, DM-nitrophen has a maximum deltau of approximately 0.013 GM at 730 nm, and NP-EGTA has no measurable uncaging yield. The equations necessary to predict the amount of cage photolyzed and the temporal behavior of the liberated calcium distribution under a variety of conditions are derived. These equations predict that by using 700-nm light from a Ti:sapphire laser focused with a 1.3-NA objective, essentially all of the azid-1 within the two-photon focal volume would be photolyzed with a 10-micros pulse train of approximately 7 mW average power. The initially localized distributions of free calcium will dissipate rapidly because of diffusion of free calcium and uptake by buffers. In buffer-free cytoplasm, the elevation of the calcium concentration at the center of the focal volume is expected to last for approximately 165 micros. PMID:9876162

Ultracytochemical visualization of calcium distribution in heart cells and erythrocytes of zebrafish Danio rerio.

PubMed

Niksirat, Hamid; Steinbach, Christoph

2018-05-24

Detection of patterns of subcellular calcium distribution in the cardiovascular system can contribute to understanding its role in cardiac and blood function. The present study localized calcium in heart atrium, ventricle, and bulbus arteriosus as well as in erythrocytes of zebrafish Danio rerio using an oxalate-pyroantimonate technique combined with transmission electron microscopy. Intracellular calcium stores were detected in caveolae, mitochondria, and the nuclei of several zebrafish cardiac cell types. Melanin pigmentation containing calcium stores was detected in the pericardial cavity. Melanin might be an extracellular source of calcium for heart beating and/or a lubricant to prevent friction during beating process. Calcium deposits were also detected in the plasma membrane, cytoplasm and nucleus of erythrocytes as well as in blood plasma. Possible exchange of calcium between erythrocytes and blood plasma was observed. Interactions of such calcium stores and possible contribution of extracellular calcium stores such as melanin pigmentation to supply calcium for vital functions of heart cells should be addressed in future studies. Copyright © 2018 Elsevier Ltd. All rights reserved.

Homo-FRET Based Biosensors and Their Application to Multiplexed Imaging of Signalling Events in Live Cells

PubMed Central

Warren, Sean C.; Margineanu, Anca; Katan, Matilda; Dunsby, Chris; French, Paul M. W.

2015-01-01

Multiplexed imaging of Förster Resonance Energy Transfer (FRET)-based biosensors potentially presents a powerful approach to monitoring the spatio-temporal correlation of signalling pathways within a single live cell. Here, we discuss the potential of homo-FRET based biosensors to facilitate multiplexed imaging. We demonstrate that the homo-FRET between pleckstrin homology domains of Akt (Akt-PH) labelled with mCherry may be used to monitor 3′-phosphoinositide accumulation in live cells and show how global analysis of time resolved fluorescence anisotropy measurements can be used to quantify this accumulation. We further present multiplexed imaging readouts of calcium concentration, using fluorescence lifetime measurements of TN-L15-a CFP/YFP based hetero-FRET calcium biosensor-with 3′-phosphoinositide accumulation. PMID:26133241

NoRMCorre: An online algorithm for piecewise rigid motion correction of calcium imaging data.

PubMed

Pnevmatikakis, Eftychios A; Giovannucci, Andrea

2017-11-01

Motion correction is a challenging pre-processing problem that arises early in the analysis pipeline of calcium imaging data sequences. The motion artifacts in two-photon microscopy recordings can be non-rigid, arising from the finite time of raster scanning and non-uniform deformations of the brain medium. We introduce an algorithm for fast Non-Rigid Motion Correction (NoRMCorre) based on template matching. NoRMCorre operates by splitting the field of view (FOV) into overlapping spatial patches along all directions. The patches are registered at a sub-pixel resolution for rigid translation against a regularly updated template. The estimated alignments are subsequently up-sampled to create a smooth motion field for each frame that can efficiently approximate non-rigid artifacts in a piecewise-rigid manner. Existing approaches either do not scale well in terms of computational performance or are targeted to non-rigid artifacts arising just from the finite speed of raster scanning, and thus cannot correct for non-rigid motion observable in datasets from a large FOV. NoRMCorre can be run in an online mode resulting in comparable to or even faster than real time motion registration of streaming data. We evaluate its performance with simple yet intuitive metrics and compare against other non-rigid registration methods on simulated data and in vivo two-photon calcium imaging datasets. Open source Matlab and Python code is also made available. The proposed method and accompanying code can be useful for solving large scale image registration problems in calcium imaging, especially in the presence of non-rigid deformations. Copyright © 2017 The Author(s). Published by Elsevier B.V. All rights reserved.

A single cell observation of staurosporine effect on the Ca2+ signals in rat basophilic leukemia cells.

PubMed

Teshima, R; Ikebuchi, H; Terao, T; Miyagawa, T; Arata, Y; Nakanishi, M

1990-09-17

A digital imaging fluorescence microscope was used to study the effect of a protein kinase inhibitor staurosporine on the antigen-dependent calcium signals in an individual rat basophilic leukemia cell (RBL-2H3). Although dose dependency of staurosporine was different from one cell to another, staurosporine inhibited, at low concentration, the calcium influx from the external medium into RBL-2H3 cells. At high concentration, however, it inhibited both the removal of calcium ion from internal stores and the calcium influx from the external medium. These results indicated that staurosporine is necessary for the inhibition of the calcium influx from the external medium and that a protein kinase (possibly protein kinase C) is involved in the calcium influx from the external medium into the cytoplasm.

Metabotropic glutamate receptors activate dendritic calcium waves and TRPM channels which drive rhythmic respiratory patterns in mice

PubMed Central

Mironov, S L

2008-01-01

Respiration in vertebrates is generated by a compact network which is located in the lower brainstem but cellular mechanisms which underlie persistent oscillatory activity of the respiratory network are yet unknown. Using two-photon imaging and patch-clamp recordings in functional brainstem preparations of mice containing pre-Bötzinger complex (preBötC), we examined the actions of metabotropic glutamate receptors (mGluR1/5) on the respiratory patterns. The agonist DHPG potentiated and antagonist LY367385 depressed respiration-related activities. In the inspiratory neurons, we observed rhythmic activation of non-selective channels which had a conductance of 24 pS. Their activity was enhanced with membrane depolarization and after elevation of calcium from the cytoplasmic side of the membrane. They were activated by a non-hydrolysable PIP2 analogue and blocked by flufenamate, ATP4− and Gd3+. All these properties correspond well to those of TRPM4 channels. Calcium imaging of functional slices revealed rhythmic transients in small clusters of neurons present in a network. Calcium transients in the soma were preceded by the waves in dendrites which were dependent on mGluR activation. Initiation and propagation of waves required calcium influx and calcium release from internal stores. Calcium waves activated TPRM4-like channels in the soma and promoted generation of inspiratory bursts. Simulations of activity of neurons communicated via dendritic calcium waves showed emerging activity within neuronal clusters and its synchronization between the clusters. The experimental and theoretical data provide a subcellular basis for a recently proposed group-pacemaker hypothesis and describe a novel mechanism of rhythm generation in neuronal networks. PMID:18308826

High-yield in vitro recordings from neurons functionally characterized in vivo.

PubMed

Weiler, Simon; Bauer, Joel; Hübener, Mark; Bonhoeffer, Tobias; Rose, Tobias; Scheuss, Volker

2018-06-01

In vivo two-photon calcium imaging provides detailed information about the activity and response properties of individual neurons. However, in vitro methods are often required to study the underlying neuronal connectivity and physiology at the cellular and synaptic levels at high resolution. This protocol provides a fast and reliable workflow for combining the two approaches by characterizing the response properties of individual neurons in mice in vivo using genetically encoded calcium indicators (GECIs), followed by retrieval of the same neurons in brain slices for further analysis in vitro (e.g., circuit mapping). In this approach, a reference frame is provided by fluorescent-bead tracks and sparsely transduced neurons expressing a structural marker in order to re-identify the same neurons. The use of GECIs provides a substantial advancement over previous approaches by allowing for repeated in vivo imaging. This opens the possibility of directly correlating experience-dependent changes in neuronal activity and feature selectivity with changes in neuronal connectivity and physiology. This protocol requires expertise both in in vivo two-photon calcium imaging and in vitro electrophysiology. It takes 3 weeks or more to complete, depending on the time allotted for repeated in vivo imaging of neuronal activity.

Estimation of calcium and magnesium in serum and urine by atomic absorption spectrophotometry

PubMed Central

Thin, Christian G.; Thomson, Patricia A.

1967-01-01

A method has been described for the estimation of calcium and magnesium in serum and urine using atomic absorption spectrophotometry. The precision and accuracy of the techniques have been determined and were found to be acceptable. The range of values for calcium and magnesium in the sera of normal adults was found to be: serum calcium (corrected to a specific gravity of 1·026) 8·38-10·08 mg. per 100 ml.; serum magnesium 1·83-2·43 mg. per 100 ml. PMID:5602562

Selective labeling of retinal ganglion cells with calcium indicators by retrograde loading in vitro

PubMed Central

Behrend, Matthew R.; Ahuja, Ashish K.; Humayun, Mark S.; Weiland, James D.; Chow, Robert H.

2012-01-01

Here we present a retrograde loading technique that makes it possible for the first time to rapidly load a calcium indicator in the majority of retinal ganglion cells (RGCs) in salamander retina, and then to observe physiological activity of these dye-loaded cells. Dextran-conjugated calcium indicator, dissolved in water, was applied to the optic nerve stump. Following dye loading, the isolated retina was mounted on a microelectrode array to demonstrate that electrical activity and calcium activity were preserved, as the retina responded to electrical stimuli. PMID:19428523

Mechanically induced intercellular calcium communication in confined endothelial structures.

PubMed

Junkin, Michael; Lu, Yi; Long, Juexuan; Deymier, Pierre A; Hoying, James B; Wong, Pak Kin

2013-03-01

Calcium signaling in the diverse vascular structures is regulated by a wide range of mechanical and biochemical factors to maintain essential physiological functions of the vasculature. To properly transmit information, the intercellular calcium communication mechanism must be robust against various conditions in the cellular microenvironment. Using plasma lithography geometric confinement, we investigate mechanically induced calcium wave propagation in networks of human umbilical vein endothelial cells organized. Endothelial cell networks with confined architectures were stimulated at the single cell level, including using capacitive force probes. Calcium wave propagation in the network was observed using fluorescence calcium imaging. We show that mechanically induced calcium signaling in the endothelial networks is dynamically regulated against a wide range of probing forces and repeated stimulations. The calcium wave is able to propagate consistently in various dimensions from monolayers to individual cell chains, and in different topologies from linear patterns to cell junctions. Our results reveal that calcium signaling provides a robust mechanism for cell-cell communication in networks of endothelial cells despite the diversity of the microenvironmental inputs and complexity of vascular structures. Copyright © 2012 Elsevier Ltd. All rights reserved.

Activity-Dependent Gating of Calcium Spikes by A-type K+ Channels Controls Climbing Fiber Signaling in Purkinje Cell Dendrites

PubMed Central

Otsu, Yo; Marcaggi, Païkan; Feltz, Anne; Isope, Philippe; Kollo, Mihaly; Nusser, Zoltan; Mathieu, Benjamin; Kano, Masanobu; Tsujita, Mika; Sakimura, Kenji; Dieudonné, Stéphane

2014-01-01

Summary In cerebellar Purkinje cell dendrites, heterosynaptic calcium signaling induced by the proximal climbing fiber (CF) input controls plasticity at distal parallel fiber (PF) synapses. The substrate and regulation of this long-range dendritic calcium signaling are poorly understood. Using high-speed calcium imaging, we examine the role of active dendritic conductances. Under basal conditions, CF stimulation evokes T-type calcium signaling displaying sharp proximodistal decrement. Combined mGluR1 receptor activation and depolarization, two activity-dependent signals, unlock P/Q calcium spikes initiation and propagation, mediating efficient CF signaling at distal sites. These spikes are initiated in proximal smooth dendrites, independently from somatic sodium action potentials, and evoke high-frequency bursts of all-or-none fast-rising calcium transients in PF spines. Gradual calcium spike burst unlocking arises from increasing inactivation of mGluR1-modulated low-threshold A-type potassium channels located in distal dendrites. Evidence for graded activity-dependent CF calcium signaling at PF synapses refines current views on cerebellar supervised learning rules. PMID:25220810

Activity-dependent gating of calcium spikes by A-type K+ channels controls climbing fiber signaling in Purkinje cell dendrites.

PubMed

Otsu, Yo; Marcaggi, Païkan; Feltz, Anne; Isope, Philippe; Kollo, Mihaly; Nusser, Zoltan; Mathieu, Benjamin; Kano, Masanobu; Tsujita, Mika; Sakimura, Kenji; Dieudonné, Stéphane

2014-10-01

In cerebellar Purkinje cell dendrites, heterosynaptic calcium signaling induced by the proximal climbing fiber (CF) input controls plasticity at distal parallel fiber (PF) synapses. The substrate and regulation of this long-range dendritic calcium signaling are poorly understood. Using high-speed calcium imaging, we examine the role of active dendritic conductances. Under basal conditions, CF stimulation evokes T-type calcium signaling displaying sharp proximodistal decrement. Combined mGluR1 receptor activation and depolarization, two activity-dependent signals, unlock P/Q calcium spikes initiation and propagation, mediating efficient CF signaling at distal sites. These spikes are initiated in proximal smooth dendrites, independently from somatic sodium action potentials, and evoke high-frequency bursts of all-or-none fast-rising calcium transients in PF spines. Gradual calcium spike burst unlocking arises from increasing inactivation of mGluR1-modulated low-threshold A-type potassium channels located in distal dendrites. Evidence for graded activity-dependent CF calcium signaling at PF synapses refines current views on cerebellar supervised learning rules. Copyright © 2014 Elsevier Inc. All rights reserved.

Neural data science: accelerating the experiment-analysis-theory cycle in large-scale neuroscience.

PubMed

Paninski, L; Cunningham, J P

2018-06-01

Modern large-scale multineuronal recording methodologies, including multielectrode arrays, calcium imaging, and optogenetic techniques, produce single-neuron resolution data of a magnitude and precision that were the realm of science fiction twenty years ago. The major bottlenecks in systems and circuit neuroscience no longer lie in simply collecting data from large neural populations, but also in understanding this data: developing novel scientific questions, with corresponding analysis techniques and experimental designs to fully harness these new capabilities and meaningfully interrogate these questions. Advances in methods for signal processing, network analysis, dimensionality reduction, and optimal control-developed in lockstep with advances in experimental neurotechnology-promise major breakthroughs in multiple fundamental neuroscience problems. These trends are clear in a broad array of subfields of modern neuroscience; this review focuses on recent advances in methods for analyzing neural time-series data with single-neuronal precision. Copyright © 2018 Elsevier Ltd. All rights reserved.

Macroindentation hardness measurement-Modernization and applications.

PubMed

Patel, Sarsvat; Sun, Changquan Calvin

2016-06-15

In this study, we first developed a modernized indentation technique for measuring tablet hardness. This technique is featured by rapid digital image capture, using a calibrated light microscope, and precise area-determination. We then systematically studied effects of key experimental parameters, including indentation force, speed, and holding time, on measured hardness of a very soft material, hydroxypropyl cellulose, and a very hard material, dibasic calcium phosphate, to cover a wide range of material properties. Based on the results, a holding period of 3min at the peak indentation load is recommended to minimize the effect of testing speed on H. Using this method, we show that an exponential decay function well describes the relationship between tablet hardness and porosity for seven commonly used pharmaceutical powders investigated in this work. We propose that H and H at zero porosity may be used to quantify the tablet deformability and powder plasticity, respectively. Copyright © 2016 Elsevier B.V. All rights reserved.

Velocity and attenuation of sound in arterial tissues

NASA Technical Reports Server (NTRS)

Rooney, J. A.; Gammell, P. M.; Hestenes, J. D.; Chin, H. P.; Blankenhorn, D. H.

1982-01-01

The velocity of sound in excised human and canine arterial tissues is measured in order to serve as a basis for the development and application of ultrasonic techniques for the diagnosis of atherosclerotic lesions. Measurements of sound velocity at different regions of 11 human and six canine aortas were made by a time delay spectrometer technique at frequencies from 2 to 10 MHz, and compared with ultrasonic attenuation parameters and the results of biochemical assays. Sound velocity is found to increase with increasing attenuation at all frequencies, and with increasing collagen content. A strong dependence of sound velocity on cholesterol content or low calcium contents is not observed, although velocities of up to 2000 m/sec are observed in highly organized calcified lesions. A decrease in velocity with decreasing temperature is also noted. It is thus concluded that it is principally the differences in tissue collagen levels that contribute to image formation according to sound velocity.

Calcium Sulfate Characterized by ChemCam/Curiosity at Gale Crater, Mars

NASA Technical Reports Server (NTRS)

Nachon, M.; Clegg, S. N.; Mangold, N.; Schroeder, S.; Kah, L. C.; Dromart, G.; Ollila, A.; Johnson, J. R.; Oehler, D. Z.; Bridges, J. C.;

Imaging of drug loading distributions in individual microspheres of calcium silicate hydrate - an X-ray spectromicroscopy study

NASA Astrophysics Data System (ADS)

Guo, Xiaoxuan; Wang, Zhiqiang; Wu, Jin; Wang, Jian; Zhu, Ying-Jie; Sham, Tsun-Kong

2015-04-01

Imaging is one of the most direct and ideal ways to track drug loading distributions in drug carriers on the molecular level, which will facilitate the optimization of drug carriers and drug loading capacities. Herein, we report the mapping of an individual mesoporous calcium silicate hydrate (CSH) microsphere before and after the loading of ibuprofen (IBU) and the interactions between drug carriers and drug molecules simultaneously by scanning transmission X-ray microscopy (STXM). Nanoscaled X-ray absorption near edge structure (XANES) spectroscopy clearly indicates that IBU is bonded to calcium and silicate sites via carboxylic acid groups. More importantly, STXM has been successfully used to determine the absolute thickness of IBU, revealing its distribution in the CSH microsphere.Imaging is one of the most direct and ideal ways to track drug loading distributions in drug carriers on the molecular level, which will facilitate the optimization of drug carriers and drug loading capacities. Herein, we report the mapping of an individual mesoporous calcium silicate hydrate (CSH) microsphere before and after the loading of ibuprofen (IBU) and the interactions between drug carriers and drug molecules simultaneously by scanning transmission X-ray microscopy (STXM). Nanoscaled X-ray absorption near edge structure (XANES) spectroscopy clearly indicates that IBU is bonded to calcium and silicate sites via carboxylic acid groups. More importantly, STXM has been successfully used to determine the absolute thickness of IBU, revealing its distribution in the CSH microsphere. Electronic supplementary information (ESI) available. See DOI: 10.1039/c4nr07471h

Imaging light responses of foveal ganglion cells in the living macaque eye.

PubMed

Yin, Lu; Masella, Benjamin; Dalkara, Deniz; Zhang, Jie; Flannery, John G; Schaffer, David V; Williams, David R; Merigan, William H

2014-05-07

The fovea dominates primate vision, and its anatomy and perceptual abilities are well studied, but its physiology has been little explored because of limitations of current physiological methods. In this study, we adapted a novel in vivo imaging method, originally developed in mouse retina, to explore foveal physiology in the macaque, which permits the repeated imaging of the functional response of many retinal ganglion cells (RGCs) simultaneously. A genetically encoded calcium indicator, G-CaMP5, was inserted into foveal RGCs, followed by calcium imaging of the displacement of foveal RGCs from their receptive fields, and their intensity-response functions. The spatial offset of foveal RGCs from their cone inputs makes this method especially appropriate for fovea by permitting imaging of RGC responses without excessive light adaptation of cones. This new method will permit the tracking of visual development, progression of retinal disease, or therapeutic interventions, such as insertion of visual prostheses.

Direct chemical reduction of neptunium oxide to neptunium metal using calcium and calcium chloride

DOE PAGES

Squires, Leah N.; Lessing, Paul

2016-01-13

A process of direct reduction of neptunium oxide to neptunium metal using calcium metal as the reducing agent is discussed. After reduction of the oxide to metal, the metal is separated by density from the other components of the reaction mixture and can easily removed upon cooling. Furthermore, the direct reduction technique consistently produces high purity (98%–99% pure) neptunium metal.

Effects of calcium and magnesium on strontium distribution coefficients

USGS Publications Warehouse

Bunde, R.L.; Rosentreter, J.J.; Liszewski, M.J.; Hemming, C.H.; Welhan, J.

1997-01-01

The effects of calcium and magnesium on the distribution of strontium between a surficial sediment and simulated wastewater solutions were measured as part of an investigation to determine strontium transport properties of surficial sediment at the Idaho National Engineering Laboratory (INEL), Idaho. The investigation was conducted by the U.S. Geological Survey and Idaho State University, in cooperation with the U.S. Department of Energy. Batch experimental techniques were used to determine strontium linear sorption isotherms and distribution coefficients (K(d)'s) using simulated wastewater solutions prepared at pH 8.0??0.1 with variable concentrations of calcium and magnesium. Strontium linear sorption isotherm K(d)'s ranged from 12??1 to 85??3 ml/g, increasing as the concentration of calcium and magnesium decreased. The concentration of sorbed strontium and the percentage of strontium retained by the sediment were correlated to aqueous concentrations of strontium, calcium, and magnesium. The effect of these cation concentrations on strontium sorption was quantified using multivariate least-squares regression techniques. Analysis of data from these experiments indicates that increased concentrations of calcium and magnesium in wastewater discharged to waste disposal ponds at the INEL increases the availability of strontium for transport beneath the ponds by decreasing strontium sorption to the surficial sediment.

Imaging ion and molecular transport at subcellular resolution by secondary ion mass spectrometry

NASA Astrophysics Data System (ADS)

Chandra, Subhash; Morrison, George H.

1995-05-01

The transport of K+, Na+, and Ca2+ were imaged in individual cells with a Cameca IMS-3f ion microscope. Strict cryogenic frozen freeze-dry sample preparations were employed. Ion redistribution artifacts in conventional chemical preparations are discussed. Cryogenically prepared freeze-fractured freeze-dried cultured cells allowed the three-dimensional ion microscopic imaging of elements. As smaller structures in calcium images can be resolved with the 0.5 [mu]m spatial resolution, correlative techniques are needed to confirm their identity. The potentials of reflected light microscopy, scanning electron microscopy and laser scanning confocal microscopy are discussed for microfeature recognition in freeze-fractured freeze-dried cells. The feasibility of using frozen freeze-dried cells for imaging molecular transport at subcellular resolution was tested. Ion microscopy successfully imaged the transport of the isotopically tagged (13C, 15N) amino acid, -arginine. The labeled amino acid was imaged at mass 28 with a Cs+ primary ion beam as the 28(13C15N)- species. After a 4 h exposure of LLC-PK1 kidney cells to 4 mM labeled arginine, the amino acid was localized throughout the cell with a preferential incorporation into the nucleus and nucleolus. An example is also shown of the ion microscopic imaging of sodium borocaptate, an experimental therapeutic drug for brain tumors, in cryogenically prepared frozen freeze-dried Swiss 3T3 cells.

Effect of simvastatin on vascular tone in porcine coronary artery: Potential role of the mitochondria

DOE Office of Scientific and Technical Information (OSTI.GOV)

Almukhtar, H.; Garle, M.J.; Smith, P.A.

2016-08-15

Statins induce acute vasorelaxation which may contribute to the overall benefits of statins in the treatment of cardiovascular disease. The mechanism underlying this relaxation is unknown. As statins have been shown to alter mitochondrial function, in this study we investigated the role of mitochondria in the relaxation to simvastatin. Relaxation of porcine coronary artery segments by statins was measured using isolated tissue baths. Mitochondrial activity was determined by measuring changes in rhodamine 123 fluorescence. Changes in intracellular calcium levels were determined in freshly isolated smooth muscle cells with Fluo-4 using standard epifluorescent imaging techniques. Simvastatin, but not pravastatin, produced amore » slow relaxation of the coronary artery, which was independent of the endothelium. The relaxation was attenuated by the mitochondrial complex I inhibitor rotenone (10 μM) and the complex III inhibitor myxothiazol (10 μM), or a combination of the two. The complex III inhibitor antimycin A (10 μM) produced a similar time-dependent relaxation of the porcine coronary artery, which was attenuated by rotenone. Changes in rhodamine 123 fluorescence showed that simvastatin (10 μM) depolarized the membrane potential of mitochondria in both isolated mitochondria and intact blood vessels. Simvastatin and antimycin A both inhibited calcium-induced contractions in isolated blood vessels and calcium influx in smooth muscle cells and this inhibition was prevented by rotenone. In conclusion, simvastatin produces an endothelium-independent relaxation of the porcine coronary artery which is dependent, in part, upon effects on the mitochondria. The effects on the mitochondria may lead to a reduction in calcium influx and hence relaxation of the blood vessel. - Highlights: • Simvastatin produces a relaxation of the porcine coronary artery. • This relaxation is inhibited by mitochondrial complex inhibitors. • Simvastatin alters mitochondrial membrane potential in intact blood vessels. • Simvastatin inhibits calcium influx in smooth muscle cells, prevented by a mitochondrial inhibitor. • Simvastatin-induced relaxations are dependent upon mitochondrial activity.« less

Roentgen Examination of Soft Tissues of the Pelvis

PubMed Central

Noonan, Charles D.

1964-01-01

With meticulous preparation of the patient and with careful technique, the soft tissues of the pelvis are identifiable in most cases. Search should be made for the traces of abnormal pelvic structures on plain-film studies. Once the normal is recognized, any variations are easily identified. The fundamental differences between various radiologic densities—air, fat, fluid, muscle, calcium, bone and metal—should be observed. Special procedures can be used to enhance the contrasts after adequate evaluation of the simplest and, on many occasions, the invaluable, plain-film study of the soft tissues of the pelvis. ImagesFigure 2.Figure 3.Figure 3.Figure 4.Figure 5.Figure 6.Figure 7.Figure 8. PMID:14232160

Arterial Structure and Function in Mild Primary Hyperparathyroidism Is Not Directly Related to Parathyroid Hormone, Calcium, or Vitamin D

PubMed Central

Ring, Margareta; Farahnak, Parastou; Gustavsson, Tomas; Nilsson, Inga-Lena; Eriksson, Maria J.; Caidahl, Kenneth

2012-01-01

Objective Elevated levels of calcium and parathyroid hormone (PTH), characteristics of primary hyperparathyroidism (PHPT), may be associated with cardiovascular morbidity and mortality in the general population. We evaluated the possible vascular effects of these risk factors in patients with mild PHPT by using standard methods and new imaging techniques. Design A prospective case-control study. Subjects and Methods Forty-eight patients with mild PHPT without any known cardiovascular risk factors were studied at baseline and at one year after parathyroidectomy (PTX) in comparison with 48 healthy age- and gender-matched controls. We measured biochemical variables, augmentation index (AIx), aortic pulse wave velocity (PWVao), radial (IMTrad) and common carotid artery (IMTcca) intima media thicknesses, and the grayscale median (IM-GSM) of the latter. Results No significant differences were observed between PHPT patients and controls at baseline for AIx (28.6±12.2 vs. 27.7±12.8%), IMTrad (0.271±0.060 vs. 0.255±0.053 mm), IMTcca (0.688±0.113 vs. 0.680±0.135 mm), or IM-GSM (82.3±17.2 vs. 86.5±15.3), while PWVao was slightly higher in patients (8.68±1.50 vs. 8.13±1.55, p<0.05). Systolic blood pressure (SBP), calcium, and PTH were higher in patients compared with controls, and decreased after PTX, while vitamin D was lower in patients and increased after PTX. While AIx, PWVao, IMTrad, and IMTcca were related to SBP, neither correlated to vitamin D levels. Only PWVao correlated weakly to plasma PTH (r = 0.29, p<0.01) and ionized calcium (r = 0.22, p<0.05) but showed no relation when age and SBP were adjusted for. Conclusion We found normal arterial function despite high calcium, PTH, and low vitamin D levels, in patients with mild PHPT without cardiovascular risk factors. The cardiovascular risk associated with low vitamin D and/or high PTH and calcium levels may be explained by their coupling to blood pressure and other risk factors rather than direct effects on arterial structure. PMID:22815708

Electrophysiological Features of Single Store-Operated Calcium Channels in HEK S4 Cell Line with Stable STIM1 Protein Knockdown.

PubMed

Shalygin, A V; Vigont, V A; Glushankova, L N; Zimina, O A; Kolesnikov, D O; Skopin, A Yu; Kaznacheeva, E V

2017-07-01

An important role in intracellular calcium signaling is played by store-operated channels activated by STIM proteins, calcium sensors of the endoplasmic reticulum. In stable STIM1 knockdown HEK S4 cells, single channels activated by depletion of intracellular calcium stores were detected by cell-attached patch-clamp technique and their electrophysiological parameters were described. Comparison of the properties of single channels in HEK293 and HEK S4 cells revealed no significant differences in their current-voltage curves, while regulation of store-operated calcium channels in these cell lines depended on the level of STIM1 expression. We can conclude that electrophysiological peculiarities of store-regulated calcium entry observed in different cells can be explained by differences in STIM1 expression.

DDPH ameliorated oxygen and glucose deprivation-induced injury in rat hippocampal neurons via interrupting Ca2+ overload and glutamate release.

PubMed

He, Zhi; Lu, Qing; Xu, Xulin; Huang, Lin; Chen, Jianguo; Guo, Lianjun

2009-01-28

Our previous work has demonstrated that DDPH (1-(2, 6-dimethylphenoxy)-2-(3, 4-dimethoxyphenylethylamino) propane hydrochloride), a competitive alpha(1)-adrenoceptor antagonist, could improve cognitive deficits, reduce histopathological damage and facilitate synaptic plasticity in vivo possibly via increasing NR2B (NMDA receptor 2B) expression and antioxidation of DDPH itself. The present study further evaluated effects of DDPH on OGD (Oxygen and glucose deprivation)-induced neuronal damage in rat primary hippocampal cells. The addition of DDPH to the cultured cells 12 h before OGD for 4 h significantly reduced neuronal damage as determined by MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay and LDH (lactate dehydrogenase) release experiments. The effects of DDPH on intracellular calcium concentration were explored by Fura-2 based calcium imaging techniques and results showed that DDPH at the dosages of 5 microM and 10 microM suppressed the increase of intracellular calcium ([Ca(2+)](i)) stimulated by 50 mM KCl in Ca(2+)-containing extracellular solutions. However, DDPH couldn't suppress the increase of [Ca(2+)](i) induced by both 50 microM glutamate in Ca(2+)-containing extracellular solutions and 20 microM ATP (Adenosine Triphosphate) in Ca(2+)-free solution. These results indicated that DDPH prevented [Ca(2+)](i) overload in hippocampal neurons by blocking Ca(2+) influx (voltage-dependent calcium channel) but not Ca(2+) mobilization from the intracellular Ca(2+) store in endoplasm reticulum (ER). We also demonstrated that DDPH could decrease glutamate release when hippocampal cells were subjected to OGD. These observations demonstrated that DDPH protected hippocampal neurons against OGD-induced damage by preventing the Ca(2+) influx and decreasing glutamate release.

Complementary information on in vitro conversion of amorphous (precursor) calcium phosphate to hydroxyapatite from Raman microspectroscopy and wide-angle X-ray scattering.

PubMed

Kazanci, M; Fratzl, P; Klaushofer, K; Paschalis, E P

2006-11-01

In addition to mechanical functions, bones have an essential role in metabolic activity as mineral reservoirs that are able to absorb and release ions. Bioapatite, considered the major component in the mineralized part of mammalian bones, is a calcium phosphate mineral with a structure that closely resembles hydroxyapatite (HA, Ca10[PO4]6[OH]2) with variable chemical substitutions. It is important to note that it continues to be chemically active long after it has been initially deposited. Detailed understanding of changes in the mineral phase as HA matures is essential for understanding how normal bone achieves its remarkable mechanical performance, how it is altered in disease, as well as the effects of therapeutic interventions. A model system for investigation of the in vivo maturation of HA is available, namely, the in vitro conversion of amorphous calcium phosphate (ACP) to HA in a supersaturated solution of calcium and phosphate ions. In the present study, this system was employed to correlate with the changes in chemistry and poorly crystalline HAP crystal size, shape, and habit. The results of the X-ray diffraction as well as Raman analyses showed that as the crystallites mature in the 002 and 310 directions both the full width at half-height and wavelength at maximum of the Raman peaks change as a function of reaction extent and crystallite maturation, size, and shape. Moreover, such analyses can be performed in intact bone specimens through Raman microspectroscopic and imaging analyses with a spatial resolution of 0.6-1 mu, by far superior to the one offered by other microspectroscopic techniques, thus potentially yielding important new information on the organization and mineral quality of normal and fragile bone.

Plant Calcium Content: Ready to Remodel

PubMed Central

Yang, Jian; Punshon, Tracy; Guerinot, Mary Lou; Hirschi, Kendal D.

2012-01-01

By identifying the relationship between calcium location in the plant cell and nutrient bioavailability, the plant characteristics leading to maximal calcium absorption by humans can be identified. Knowledge of plant cellular and molecular targets controlling calcium location in plants is emerging. These insights should allow for better strategies for increasing the nutritional content of foods. In particular, the use of preparation-free elemental imaging technologies such as synchrotron X-ray fluorescence (SXRF) microscopy in plant biology may allow researchers to understand the relationship between subcellular location and nutrient bioavailability. These approaches may lead to better strategies for altering the location of calcium within the plant to maximize its absorption from fruits and vegetables. These modified foods could be part of a diet for children and adults identified as at-risk for low calcium intake or absorption with the ultimate goal of decreasing the incidence and severity of inadequate bone mineralization. PMID:23016135

Evidence for calcium soaps in human hair shaft revealed by sub-micrometer X-ray fluorescence

NASA Astrophysics Data System (ADS)

Briki, F.; Mérigoux, C.; Sarrot-Reynauld, F.; Salomé, M.; Fayard, B.; Susini, J.; Doucet, J.

2003-03-01

New information about calcium status in human scalp hair shaft, deduced from X-ray microfluorescence imaging, including its distribution over the hair section, the existence of one or several binding-types and its variation between people, is presented. The existence of two different calcium types is inferred. The first one corresponds to atoms (or ions) easily removable by hydrochloric acid, located in the cortex (granules), in the cuticle zone and also in the core of the medulla, which are identified as calcium soaps cy comparison with X-ray diffraction and IR spectromicroscopy data. The second type consists of non-easily removable calcium atoms (or ions) that are located in the medulla wall, probably also the cuticle, and rather uniformly in the cortex; these calcium atoms may be involved in Ca^{2+}-binding proteins, their concentration is fairly constant from one subject to another.

A comparison of the effects of commercially available hawthorn preparations on calcium transients of isolated cardiomyocytes.

PubMed

Rodriguez, Michelle E; Poindexter, Brian J; Bick, Roger J; Dasgupta, Amitava

2008-12-01

We studied the potential cardiac effects of two alcohol extracts of commercially available hawthorn using rat cardiomyocytes and measuring calcium transients by real-time fluorescence spectrophotometry. One preparation was a blend of hawthorn flowers, leaves, and berries (extract #1), and the other (extract #2) was from a "berries-only" preparation. Fluorescent images and calcium transients were acquired concurrently. Addition of extract #1 resulted in the initiation of robust calcium transients and eventual calcium overload, while addition of extract #2 caused increased calcium sparking, initiation of calcium transients, and an increased beating rate but no calcium overload. To identify the mechanisms of increased calcium influx, adult rat cardiomyocytes were challenged with 10 microM ouabain, a Na(+),K(+)-ATPase inhibitor, and the calcium channel blocker nifedipine. The findings revealed that equal volumes of the two readily available hawthorn preparations demonstrated markedly different effects on isolated adult rat cardiomyocytes, suggesting important implications for patients who are using these preparations to supplement or even replace their prescribed cardiac medications as to which preparation(s) to use, and potential dire consequences, particularly in cardiac patients. Our study indicates that the mechanism of cardiac activity of hawthorn is via the Na(+),K(+)-ATPase and intracellular calcium concentrations are influenced.

Crystal structures of the GCaMP calcium sensor reveal the mechanism of fluorescence signal change and aid rational design.

PubMed

Akerboom, Jasper; Rivera, Jonathan D Vélez; Guilbe, María M Rodríguez; Malavé, Elisa C Alfaro; Hernandez, Hector H; Tian, Lin; Hires, S Andrew; Marvin, Jonathan S; Looger, Loren L; Schreiter, Eric R

2009-03-06

The genetically encoded calcium indicator GCaMP2 shows promise for neural network activity imaging, but is currently limited by low signal-to-noise ratio. We describe x-ray crystal structures as well as solution biophysical and spectroscopic characterization of GCaMP2 in the calcium-free dark state, and in two calcium-bound bright states: a monomeric form that dominates at intracellular concentrations observed during imaging experiments and an unexpected domain-swapped dimer with decreased fluorescence. This series of structures provides insight into the mechanism of Ca2+-induced fluorescence change. Upon calcium binding, the calmodulin (CaM) domain wraps around the M13 peptide, creating a new domain interface between CaM and the circularly permuted enhanced green fluorescent protein domain. Residues from CaM alter the chemical environment of the circularly permuted enhanced green fluorescent protein chromophore and, together with flexible inter-domain linkers, block solvent access to the chromophore. Guided by the crystal structures, we engineered a series of GCaMP2 point mutants to probe the mechanism of GCaMP2 function and characterized one mutant with significantly improved signal-to-noise. The mutation is located at a domain interface and its effect on sensor function could not have been predicted in the absence of structural data.

Visualization of Plasticity in Fear-Evoked Calcium Signals in Midbrain Dopamine Neurons

ERIC Educational Resources Information Center

Gore, Bryan B.; Soden, Marta E.; Zweifel, Larry S.

2014-01-01

Dopamine is broadly implicated in fear-related processes, yet we know very little about signaling dynamics in these neurons during active fear conditioning. We describe the direct imaging of calcium signals of dopamine neurons during Pavlovian fear conditioning using fiber-optic confocal microscopy coupled with the genetically encoded calcium…

Calcium-Responsive Liposomes via a Synthetic Lipid Switch.

PubMed

Lou, Jinchao; Carr, Adam J; Watson, Alexa J; Mattern-Schain, Samuel I; Best, Michael D

2018-03-07

Liposomal drug delivery would benefit from enhanced control over content release. Here, we report a novel avenue for triggering release driven by chemical composition using liposomes sensitized to calcium-a target chosen due to its key roles in biology and disease. To demonstrate this principle, we synthesized calcium-responsive lipid switch 1, designed to undergo conformational changes upon calcium binding. The conformational change perturbs membrane integrity, thereby promoting cargo release. This was shown through fluorescence-based release assays via dose-dependent response depending on the percentage of 1 in liposomes, with minimal background leakage in controls. DLS experiments indicated dramatic changes in particle size upon treatment of liposomes containing 1 with calcium. In a comparison of ten naturally occurring metal cations, calcium provided the greatest release. Finally, STEM images showed significant changes in liposome morphology upon treatment of liposomes containing 1 with calcium. These results showcase lipid switches driven by molecular recognition principles as an exciting avenue for controlling membrane properties. © 2018 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

Drosophila wing imaginal discs respond to mechanical injury via slow InsP3R-mediated intercellular calcium waves

NASA Astrophysics Data System (ADS)

Restrepo, Simon; Basler, Konrad

2016-08-01

Calcium signalling is a highly versatile cellular communication system that modulates basic functions such as cell contractility, essential steps of animal development such as fertilization and higher-order processes such as memory. We probed the function of calcium signalling in Drosophila wing imaginal discs through a combination of ex vivo and in vivo imaging and genetic analysis. Here we discover that wing discs display slow, long-range intercellular calcium waves (ICWs) when mechanically stressed in vivo or cultured ex vivo. These slow imaginal disc intercellular calcium waves (SIDICs) are mediated by the inositol-3-phosphate receptor, the endoplasmic reticulum (ER) calcium pump SERCA and the key gap junction component Inx2. The knockdown of genes required for SIDIC formation and propagation negatively affects wing disc recovery after mechanical injury. Our results reveal a role for ICWs in wing disc homoeostasis and highlight the utility of the wing disc as a model for calcium signalling studies.

Calcium as a signal integrator in developing epithelial tissues.

PubMed

Brodskiy, Pavel A; Zartman, Jeremiah J

2018-05-16

Decoding how tissue properties emerge across multiple spatial and temporal scales from the integration of local signals is a grand challenge in quantitative biology. For example, the collective behavior of epithelial cells is critical for shaping developing embryos. Understanding how epithelial cells interpret a diverse range of local signals to coordinate tissue-level processes requires a systems-level understanding of development. Integration of multiple signaling pathways that specify cell signaling information requires second messengers such as calcium ions. Increasingly, specific roles have been uncovered for calcium signaling throughout development. Calcium signaling regulates many processes including division, migration, death, and differentiation. However, the pleiotropic and ubiquitous nature of calcium signaling implies that many additional functions remain to be discovered. Here we review a selection of recent studies to highlight important insights into how multiple signals are transduced by calcium transients in developing epithelial tissues. Quantitative imaging and computational modeling have provided important insights into how calcium signaling integration occurs. Reverse-engineering the conserved features of signal integration mediated by calcium signaling will enable novel approaches in regenerative medicine and synthetic control of morphogenesis.

Advances in two photon scanning and scanless microscopy technologies for functional neural circuit imaging.

PubMed

Schultz, Simon R; Copeland, Caroline S; Foust, Amanda J; Quicke, Peter; Schuck, Renaud

2017-01-01

Recent years have seen substantial developments in technology for imaging neural circuits, raising the prospect of large scale imaging studies of neural populations involved in information processing, with the potential to lead to step changes in our understanding of brain function and dysfunction. In this article we will review some key recent advances: improved fluorophores for single cell resolution functional neuroimaging using a two photon microscope; improved approaches to the problem of scanning active circuits; and the prospect of scanless microscopes which overcome some of the bandwidth limitations of current imaging techniques. These advances in technology for experimental neuroscience have in themselves led to technical challenges, such as the need for the development of novel signal processing and data analysis tools in order to make the most of the new experimental tools. We review recent work in some active topics, such as region of interest segmentation algorithms capable of demixing overlapping signals, and new highly accurate algorithms for calcium transient detection. These advances motivate the development of new data analysis tools capable of dealing with spatial or spatiotemporal patterns of neural activity, that scale well with pattern size.

Physiological imaging of electrical trauma and therapeutic responses

NASA Astrophysics Data System (ADS)

Chen, Chin-Tu; Matthews, K.; Aarsvold, John N.; Mintzer, Robert A.; Yasillo, Nicholas J.; Hannig, Jurgen; Capelli-Schellpfefer, M.; Cooper, Malcolm; Lee, Raphael C.

2000-04-01

In victims of electrical trauma, electroporation of cell membrane, in which lipid bilayer is permeabilized by thermal and electrical forces, is thought to be a substantial cause of tissue damage. It has been suggested that certain mild surfactant in low concentration could induce sealing of permeabilized lipid bilayers, thus repairing cell membranes that had not been extensively damaged. With an animal model of electrically injured hind limb of rats, we have demonstrated and validated the use of radiotracer imaging technique to assess the physiology of the damaged tissues after electrical shock and of their repairs after applying surfactant as a therapeutic strategy. For example, using Tc-99m labeled pyrophosphate (PYP), which follows calcium in cellular function and is known to accumulate in damaged tissues, we have established a physiological imaging approach for assessment of the extent of tissue injury for diagnosis and surgical planning, as well as for evaluation of responses to therapy. With the use of a small, hand-held, miniature gamma camera, this physiological imaging method can be employed at patient's bedside and even in the field, for example, at accident site or during transfer for emergency care, rapid diagnosis, and prompt treatment in order to maximize the chance for tissue survival.

Advances in two photon scanning and scanless microscopy technologies for functional neural circuit imaging

PubMed Central

Schultz, Simon R.; Copeland, Caroline S.; Foust, Amanda J.; Quicke, Peter; Schuck, Renaud

2017-01-01

Recent years have seen substantial developments in technology for imaging neural circuits, raising the prospect of large scale imaging studies of neural populations involved in information processing, with the potential to lead to step changes in our understanding of brain function and dysfunction. In this article we will review some key recent advances: improved fluorophores for single cell resolution functional neuroimaging using a two photon microscope; improved approaches to the problem of scanning active circuits; and the prospect of scanless microscopes which overcome some of the bandwidth limitations of current imaging techniques. These advances in technology for experimental neuroscience have in themselves led to technical challenges, such as the need for the development of novel signal processing and data analysis tools in order to make the most of the new experimental tools. We review recent work in some active topics, such as region of interest segmentation algorithms capable of demixing overlapping signals, and new highly accurate algorithms for calcium transient detection. These advances motivate the development of new data analysis tools capable of dealing with spatial or spatiotemporal patterns of neural activity, that scale well with pattern size. PMID:28757657

Synthesis of Calcium Phosphate Composite Organogels by Using Emulsion Method for Dentine Occlusion Materials

NASA Astrophysics Data System (ADS)

Nopteeranupharp, C.; Akkarachaneeyakorn, K.; Songsasaen, A.

2018-03-01

Dentinal hypersensitivity (DH) is one of the most human’s problems caused by the erosion of enamel. There are many methods and materials to solve this problem. Calcium phosphate is an excellent alternative for curing this symptom because of its osteoconductivity, and biocompatibility properties. The low-cost and low-toxicity calcium phosphate nanogel was fabricated by using emulsion method and characterized by using TEM, EDX, and DLS techniques. The results showed that P123 (poly (ethylene oxide)19-block-Poly (propylene oxide)69-block-poly (ethylene oxide)19) has played a major role as template and gel formation, SDS was used as a surfactant to form water-in-oil emulsion nanodroplets with circle-like shape. Moreover, the ability of synthesised organogel to occlude the exposed dentine tubules was tested on the model of human’s dentine slices. The results showed that calcium phosphate composite organogel can be efficiently occluded on dentine slice, characterized by SEM technique, after 1 day.

Development of a sensitive setup for laser spectroscopy studies of very exotic calcium isotopes

NASA Astrophysics Data System (ADS)

Garcia Ruiz, R. F.; Gorges, C.; Bissell, M.; Blaum, K.; Gins, W.; Heylen, H.; Koenig, K.; Kaufmann, S.; Kowalska, M.; Krämer, J.; Lievens, P.; Malbrunot-Ettenauer, S.; Neugart, R.; Neyens, G.; Nörtershäuser, W.; Yordanov, D. T.; Yang, X. F.

2017-04-01

An experimental setup for sensitive high-resolution measurements of hyperfine structure spectra of exotic calcium isotopes has been developed and commissioned at the COLLAPS beam line at ISOLDE, CERN. The technique is based on the radioactive detection of decaying isotopes after optical pumping and state selective neutralization (ROC) (Vermeeren et al 1992 Phys. Rev. Lett. 68 1679). The improvements and developments necessary to extend the applicability of the experimental technique to calcium isotopes produced at rates as low as few ions s-1 are discussed. Numerical calculations of laser-ion interaction and ion-beam simulations were explored to obtain the optimum performance of the experimental setup. Among the implemented features are a multi-step optical pumping region for sensitive measurements of isotopes with hyperfine splitting, a high-voltage platform for adequate control of low-energy ion beams and simultaneous β-detection of neutralized and remaining ions. The commissioning of the experimental setup, and the first online results on neutron-rich calcium isotopes are presented.

Dual-energy CT revisited with multidetector CT: review of principles and clinical applications.

PubMed

Karçaaltıncaba, Muşturay; Aktaş, Aykut

2011-09-01

Although dual-energy CT (DECT) was first conceived in the 1970s, it was not widely used for CT indications. Recently, the simultaneous acquisition of volumetric dual-energy data has been introduced using multidetector CT (MDCT) with two X-ray tubes and rapid kVp switching (gemstone spectral imaging). Two major advantages of DECT are material decomposition by acquiring two image series with different kVp and the elimination of misregistration artifacts. Hounsfield unit measurements by DECT are not absolute and can change depending on the kVp used for an acquisition. Typically, a combination of 80/140 kVp is used for DECT, but for some applications, 100/140 kVp is preferred. In this study, we summarized the clinical applications of DECT and included images that were acquired using the dual-source CT and rapid kVp switching. In general, unenhanced images can be avoided by using DECT for body and neurological applications; iodine can be removed from the image, and a virtual, non-contrast (water) image can be obtained. Neuroradiological applications allow for the removal of bone and calcium from the carotid and brain CT angiography. Thorax applications include perfusion imaging in patients with pulmonary thromboemboli and other chest diseases, xenon ventilation-perfusion imaging and solitary nodule characterization. Cardiac applications include dual-energy cardiac perfusion, viability and cardiac iron detection. The removal of calcific plaques from arteries, bone removal and aortic stent graft evaluation may be achieved in the vascular system. Abdominal applications include the detection and characterization of liver and pancreas masses, the diagnosis of steatosis and iron overload, DECT colonoscopy and CT cholangiography. Urinary system applications are urinary calculi characterization (uric acid vs. non-uric acid), renal cyst characterization and mass characterization. Musculoskeletal applications permit the differentiation of gout from pseudogout and a reduction of metal artifacts. Recent introduction of iterative reconstruction techniques can increase the use of DECT techniques; the use of dual energy in patients with a high BMI is limited due to noise and the radiation dose. DECT may be a good alternative to PET-CT. Iodine map images can quantify iodine uptake, and this approach may be more effective than obtaining non-contrast and post-contrast images for the diagnosis of a solid mass. Thus, computer-aided detection may be used more effectively in CT applications. DECT is a promising technique with potential clinical applications.

Intravascular photoacoustic imaging: a new tool for vulnerable plaque identification.

PubMed

Jansen, Krista; van Soest, Gijs; van der Steen, Antonius F W

2014-06-01

The vulnerable atherosclerotic plaque is believed to be at the root of the majority of acute coronary events. Even though the exact origins of plaque vulnerability remain elusive, the thin-cap fibroatheroma, characterized by a lipid-rich necrotic core covered by a thin fibrous cap, is considered to be the most prominent type of vulnerable plaque. No clinically available imaging technique can characterize atherosclerotic lesions to the extent needed to determine plaque vulnerability prognostically. Intravascular photoacoustic imaging (IVPA) has the potential to take a significant step in that direction by imaging both plaque structure and composition. IVPA is a natural extension of intravascular ultrasound that adds tissue type specificity to the images. IVPA utilizes the optical contrast provided by the differences in the absorption spectra of plaque components to image composition. Its capability to image lipids in human coronary atherosclerosis has been shown extensively ex vivo and has recently been translated to an in vivo animal model. Other disease markers that have been successfully targeted are calcium and inflammatory markers, such as macrophages and matrix metalloproteinase; the latter two through application of exogenous contrast agents. By simultaneously displaying plaque morphology and composition, IVPA can provide a powerful prognostic marker for disease progression, and as such has the potential to transform the current practice in percutaneous coronary intervention. Copyright © 2014 World Federation for Ultrasound in Medicine & Biology. Published by Elsevier Inc. All rights reserved.

Ancient Chinese medicine and mechanistic evidence of acupuncture physiology.

PubMed

Yang, Edward S; Li, Pei-Wen; Nilius, Bernd; Li, Geng

2011-11-01

Acupuncture has been widely used in China for three millennia as an art of healing. Yet, its physiology is not yet understood. The current interest in acupuncture started in 1971. Soon afterward, extensive research led to the concept of neural signaling with possible involvement of opioid peptides, glutamate, adenosine and identifying responsive parts in the central nervous system. In the last decade scientists began investigating the subject with anatomical and molecular imaging. It was found that mechanical movements of the needle, ignored in the past, appear to be central to the method and intracellular calcium ions may play a pivotal role. In this review, we trace the technique of clinical treatment from the first written record about 2,200 years ago to the modern time. The ancient texts have been used to introduce the concepts of yin, yang, qi, de qi, and meridians, the traditional foundation of acupuncture. We explore the sequence of the physiological process, from the turning of the needle, the mechanical wave activation of calcium ion channel to beta-endorphin secretion. By using modern terminology to re-interpret the ancient texts, we have found that the 2nd century B.C.: physiologists were meticulous investigators and their explanation fits well with the mechanistic model derived from magnetic resonance imaging (MRI) and confocal microscopy. In conclusion, the ancient model appears to have withstood the test of time surprisingly well confirming the popular axiom that the old wine is better than the new.

Analysis of Spontaneous and Nerve-Evoked Calcium Transients in Intact Extraocular Muscles in Vitro

PubMed Central

Feng, Cheng-Yuan; Hennig, Grant W.; Corrigan, Robert D.; Smith, Terence K.; von Bartheld, Christopher S.

2012-01-01

Extraocular muscles (EOMs) have unique calcium handling properties, yet little is known about the dynamics of calcium events underlying ultrafast and tonic contractions in myofibers of intact EOMs. Superior oblique EOMs of juvenile chickens were dissected with their nerve attached, maintained in oxygenated Krebs buffer, and loaded with fluo-4. Spontaneous and nerve stimulation-evoked calcium transients were recorded and, following calcium imaging, some EOMs were double-labeled with rhodamine-conjugated alpha-bungarotoxin (rhBTX) to identify EOM myofiber types. EOMs showed two main types of spontaneous calcium transients, one slow type (calcium waves with 1/2max duration of 2–12 s, velocity of 25–50 μm/s) and two fast “flash-like” types (Type 1, 30–90 ms; Type 2, 90–150 ms 1/2max duration). Single pulse nerve stimulation evoked fast calcium transients identical to the fast (Type 1) calcium transients. Calcium waves were accompanied by a local myofiber contraction that followed the calcium transient wavefront. The magnitude of calcium-wave induced myofiber contraction far exceeded those of movement induced by nerve stimulation and associated fast calcium transients. Tetrodotoxin eliminated nerve-evoked transients, but not spontaneous transients. Alpha-bungarotoxin eliminated both spontaneous and nerve-evoked fast calcium transients, but not calcium waves, and caffeine increased wave activity. Calcium waves were observed in myofibers lacking spontaneous or evoked fast transients, suggestive of multiply-innervated myofibers, and this was confirmed by double-labeling with rhBTX. We propose that the abundant spontaneous calcium transients and calcium waves with localized contractions that do not depend on innervation may contribute to intrinsic generation of tonic functions of EOMs. PMID:22579493

Development of calcium bodies in Hylonsicus riparius (Crustacea: Isopoda).

PubMed

Vittori, Miloš; Khurshed, Mohammed; Picavet, Daisy I; van Noorden, Cornelis J F; Štrus, Jasna

2018-03-01

Calcium bodies are internal epithelial sacs found in terrestrial isopods of the family Trichoniscidae that contain a mineralized extracellular matrix that is deposited and resorbed in relation to the molt cycle. Calcium bodies in several trichoniscids are filled with bacteria, the function of which is currently unknown. The woodlouse Hyloniscus riparius differs from other trichoniscids in that it possesses two different pairs of calcium bodies, the posterior pair being filled with bacteria and the anterior pair being devoid of bacteria. We explored the development of these organs and bacterial colonization of their lumen during the postmarsupial development with the use of optical clearing and whole-body confocal imaging of larval and juvenile stages. Our results show that calcium bodies are formed as invaginations of the epidermis in the region of intersegmental membranes during the postmarsupial development. The anterior pair of calcium bodies is generated during the first postmarsupial manca stage, whereas the posterior calcium bodies first appear in juveniles and are immediately colonized by bacteria, likely through a connection between the calcium body lumen and the body surface. Mineral is deposited in calcium bodies as soon as they are present. Copyright © 2018 Elsevier Ltd. All rights reserved.

Combined use of UV-labile calcium chelators and calcium-sensitive dyes in a microscope with two light sources influencing different regions in a group of coordinated contracting cardiac myocytes

NASA Astrophysics Data System (ADS)

Pilarczyk, Goetz; Greulich, Karl-Otto

1997-12-01

The coordination of excitation in a biological system of cells such as cardiac myocytes in heart tissue has crucial influence on the function of the entire organ. This coordinated behavior can be visualized in a small group of embryonic cardiac myocytes derived from the hearts of unborn chicken. Loaded with a calcium sensitive dye the excitation can be imaged via the occurring transient rise in cytosolic calcium concentration. It can be shown that in regions with physiological or morphological restrictions the transient rise in cytosolic calcium occurs with a temporal delay compared to the ordinary array of coupled myocytes. The height of the transient rise of cytosolic calcium is related to the ability of the individual cell to participate in the coordinated contraction. The free cytosolic calcium concentration is decreased with the UV-labile calcium, chelator diazo-2. Our setup allows to decrease the free cytosolic calcium in a single cell of the contracting array of cells. This allows us to introduce mismatches in selected regions of the coordinated contraction and to visualize the effects simultaneously.

Calcium waves.

PubMed

Jaffe, Lionel F

2008-04-12

Waves through living systems are best characterized by their speeds at 20 degrees C. These speeds vary from those of calcium action potentials to those of ultraslow ones which move at 1-10 and/or 10-20 nm s(-1). All such waves are known or inferred to be calcium waves. The two classes of calcium waves which include ones with important morphogenetic effects are slow waves that move at 0.2-2 microm s(-1) and ultraslow ones. Both may be propagated by cycles in which the entry of calcium through the plasma membrane induces subsurface contraction. This contraction opens nearby stretch-sensitive calcium channels. Calcium entry through these channels propagates the calcium wave. Many slow waves are seen as waves of indentation. Some are considered to act via cellular peristalsis; for example, those which seem to drive the germ plasm to the vegetal pole of the Xenopus egg. Other good examples of morphogenetic slow waves are ones through fertilizing maize eggs, through developing barnacle eggs and through axolotl embryos during neural induction. Good examples of ultraslow morphogenetic waves are ones during inversion in developing Volvox embryos and across developing Drosophila eye discs. Morphogenetic waves may be best pursued by imaging their calcium with aequorins.

A technique of experimental and numerical analysis of influence of defects in the intraocular lens on the retinal image quality

NASA Astrophysics Data System (ADS)

Geniusz, Malwina; ZajÄ c, Marek

2016-09-01

Intraocular lens (IOL) is an artificial lens implanted into the eye in order to restore correct vision after the removal of natural lens cloudy due to cataract. The IOL prolonged stay in the eyeball causes the creation of different changes on the surface and inside the implant mainly in form of small-size local defects such as vacuoles and calcium deposites. Their presence worsens the imaging properties of the eye mainly due to occurence of scattered light thus deteriorating the vision quality of patients after cataract surgery. It is very difficult to study influence the effects of these changes on image quality in real patients. To avoid these difficulties two other possibilities were chosen: the analysis of the image obtained in an optomechanical eye model with artificially aged IOL as well as numerical calculation of the image characteristics while the eye lens is burdened with adequately modeled defects. In experiments the optomechanical model of an eye consisting of a glass "cornea", chamber filled with liquid where the IOL under investigation was inserted and a high resulution CCC detector serving as a "retina" was used. The Modulation Transfer Function (MTF) of such "eye" was evaluated on the basis of image of an edge. Experiments show that there is significant connection between ageing defects and decrease in MTF parameters. Numerical part was performed with a computer programme for optical imaging analysis (OpticStudio Professional, Zemax Professional from Radiant Zemax, LLC). On the basis of Atchison eye model with lens burdened with defects Modulation Transfer Functio was calculated. Particular parameters of defects used in a numerical model were based on own measurements. Numerical simulation also show significant connection between ageing defects and decrease of MTF parameters. With this technique the influence of types, density and distribution of local defect in the IOL on the retinal image quality can be evaluated quickly without the need of performing very difficult and even dangereous experiments on real human patients.

Effects of calcium leaching on diffusion properties of hardened and altered cement pastes

NASA Astrophysics Data System (ADS)

Kurumisawa, Kiyofumi; Haga, Kazuko; Hayashi, Daisuke; Owada, Hitoshi

2017-06-01

It is very important to predict alterations in the concrete used for fabricating disposal containers for radioactive waste. Therefore, it is necessary to understand the alteration of cementitious materials caused by calcium leaching when they are in contact with ground water in the long term. To evaluate the long-term transport characteristics of cementitious materials, the microstructural behavior of these materials should be considered. However, many predictive models of transport characteristics focus on the pore structure, while only few such models consider both, the spatial distribution of calcium silicate hydrate (C-S-H), portlandite, and the pore spaces. This study focused on the spatial distribution of these cement phases. The auto-correlation function of each phase of cementitious materials was calculated from two-dimensional backscattered electron imaging, and the three-dimensional spatial image of the cementitious material was produced using these auto-correlation functions. An attempt was made to estimate the diffusion coefficient of chloride from the three-dimensional spatial image. The estimated diffusion coefficient of the altered sample from the three-dimensional spatial image was found to be comparable to the measured value. This demonstrated that it is possible to predict the diffusion coefficient of the altered cement paste by using the proposed model.

FocusStack and StimServer: a new open source MATLAB toolchain for visual stimulation and analysis of two-photon calcium neuronal imaging data.

PubMed

Muir, Dylan R; Kampa, Björn M

2014-01-01

Two-photon calcium imaging of neuronal responses is an increasingly accessible technology for probing population responses in cortex at single cell resolution, and with reasonable and improving temporal resolution. However, analysis of two-photon data is usually performed using ad-hoc solutions. To date, no publicly available software exists for straightforward analysis of stimulus-triggered two-photon imaging experiments. In addition, the increasing data rates of two-photon acquisition systems imply increasing cost of computing hardware required for in-memory analysis. Here we present a Matlab toolbox, FocusStack, for simple and efficient analysis of two-photon calcium imaging stacks on consumer-level hardware, with minimal memory footprint. We also present a Matlab toolbox, StimServer, for generation and sequencing of visual stimuli, designed to be triggered over a network link from a two-photon acquisition system. FocusStack is compatible out of the box with several existing two-photon acquisition systems, and is simple to adapt to arbitrary binary file formats. Analysis tools such as stack alignment for movement correction, automated cell detection and peri-stimulus time histograms are already provided, and further tools can be easily incorporated. Both packages are available as publicly-accessible source-code repositories.

FocusStack and StimServer: a new open source MATLAB toolchain for visual stimulation and analysis of two-photon calcium neuronal imaging data

PubMed Central

Muir, Dylan R.; Kampa, Björn M.

2015-01-01

Two-photon calcium imaging of neuronal responses is an increasingly accessible technology for probing population responses in cortex at single cell resolution, and with reasonable and improving temporal resolution. However, analysis of two-photon data is usually performed using ad-hoc solutions. To date, no publicly available software exists for straightforward analysis of stimulus-triggered two-photon imaging experiments. In addition, the increasing data rates of two-photon acquisition systems imply increasing cost of computing hardware required for in-memory analysis. Here we present a Matlab toolbox, FocusStack, for simple and efficient analysis of two-photon calcium imaging stacks on consumer-level hardware, with minimal memory footprint. We also present a Matlab toolbox, StimServer, for generation and sequencing of visual stimuli, designed to be triggered over a network link from a two-photon acquisition system. FocusStack is compatible out of the box with several existing two-photon acquisition systems, and is simple to adapt to arbitrary binary file formats. Analysis tools such as stack alignment for movement correction, automated cell detection and peri-stimulus time histograms are already provided, and further tools can be easily incorporated. Both packages are available as publicly-accessible source-code repositories1. PMID:25653614

Total body calcium analysis using the Ca-12(n, alpha) Ar-37 reaction

NASA Technical Reports Server (NTRS)

Lewellen, T. K.; Nelp, W. B.

1977-01-01

A low dose neutron activation technique was developed to measure total body calcium in vivo. The effort had included development of irradiation and processing facilities and conduction of human studies to determine the accuracy and precision of measurement attainable with the systems.

St. John’s Wort enhances the synaptic activity of the nucleus of the solitary tract

PubMed Central

Vance, Katie M.; Ribnicky, David M.; Hermann, Gerlinda E.; Rogers, Richard C.

2014-01-01

Objective St. John’s Wort extract, which is commonly used to treat depression, inhibits the reuptake of several neurotransmitters, including glutamate, serotonin, norepinephrine, and dopamine. Glutamatergic visceral vagal afferents synapse upon neurons of the solitary tract (NST); thus, we evaluated whether St. John’s Wort extract modulates glutamatergic neurotransmission within the NST. Materials and Methods We used live cell calcium imaging to evaluate whether St. John’s Wort and its isolated components hypericin and hyperforin increase the excitability of pre-labeled vagal afferent terminals synapsing upon the NST. We used voltage-clamp recordings of spontaneous miniature excitatory postsynaptic currents (mEPSCs) to evaluate whether St. John’s Wort alters glutamate release from vagal afferents onto NST neurons. Results Our imaging data show that St. John’s Wort (50 μg/mL) increased the intracellular calcium levels of stimulated vagal afferent terminals compared to the bath control. This increase in presynaptic vagal afferent calcium by the extract coincides with an increase in neurotransmitter release within the nucleus of the solitary tract, as the frequency of mEPSCs is significantly higher in the presence of the extract compared to the control. Finally, our imaging data show that hyperforin, a known component of St. John’s Wort extract, also significantly increases terminal calcium levels. Conclusion These data suggest that St. John’s Wort extract can significantly increase the probability of glutamate release from vagal afferents onto the NST by increasing presynaptic calcium. The in vitro vagal afferent synapse with NST neurons is an ideal model system to examine the mechanism of action of botanical agents on glutamatergic neurotransmission. PMID:24985104

Accurately Diagnosing Uric Acid Stones from Conventional Computerized Tomography Imaging: Development and Preliminary Assessment of a Pixel Mapping Software.

PubMed

Ganesan, Vishnu; De, Shubha; Shkumat, Nicholas; Marchini, Giovanni; Monga, Manoj

2018-02-01

Preoperative determination of uric acid stones from computerized tomography imaging would be of tremendous clinical use. We sought to design a software algorithm that could apply data from noncontrast computerized tomography to predict the presence of uric acid stones. Patients with pure uric acid and calcium oxalate stones were identified from our stone registry. Only stones greater than 4 mm which were clearly traceable from initial computerized tomography to final composition were included in analysis. A semiautomated computer algorithm was used to process image data. Average and maximum HU, eccentricity (deviation from a circle) and kurtosis (peakedness vs flatness) were automatically generated. These parameters were examined in several mathematical models to predict the presence of uric acid stones. A total of 100 patients, of whom 52 had calcium oxalate and 48 had uric acid stones, were included in the final analysis. Uric acid stones were significantly larger (12.2 vs 9.0 mm, p = 0.03) but calcium oxalate stones had higher mean attenuation (457 vs 315 HU, p = 0.001) and maximum attenuation (918 vs 553 HU, p <0.001). Kurtosis was significantly higher in each axis for calcium oxalate stones (each p <0.001). A composite algorithm using attenuation distribution pattern, average attenuation and stone size had overall 89% sensitivity, 91% specificity, 91% positive predictive value and 89% negative predictive value to predict uric acid stones. A combination of stone size, attenuation intensity and attenuation pattern from conventional computerized tomography can distinguish uric acid stones from calcium oxalate stones with high sensitivity and specificity. Copyright © 2018 American Urological Association Education and Research, Inc. Published by Elsevier Inc. All rights reserved.

Imaging Wellbore Cement Degradation by Carbon Dioxide under Geologic Sequestration Conditions Using X-ray Computed Microtomography

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jung, Hun Bok; Jansik, Danielle; Um, Wooyong

2013-01-02

ABSTRACT: X-ray microtomography (XMT), a nondestructive three-dimensional imaging technique, was applied to demonstrate its capability to visualize the mineralogical alteration and microstructure changes in hydrated Portland cement exposed to carbon dioxide under geologic sequestration conditions. Steel coupons and basalt fragments were added to the cement paste in order to simulate cement-steel and cement-rock interfaces. XMT image analysis showed the changes of material density and porosity in the degradation front (density: 1.98 g/cm3, porosity: 40%) and the carbonated zone (density: 2.27 g/cm3, porosity: 23%) after reaction with CO2- saturated water for 5 months compared to unaltered cement (density: 2.15 g/cm3, porosity:more » 30%). Three-dimensional XMT imaging was capable of displaying spatially heterogeneous alteration in cement pores, calcium carbonate precipitation in cement cracks, and preferential cement alteration along the cement-steel and cement-rock interfaces. This result also indicates that the interface between cement and host rock or steel casing is likely more vulnerable to a CO2 attack than the cement matrix in a wellbore environment. It is shown here that XMT imaging can potentially provide a new insight into the physical and chemical degradation of wellbore cement by CO2 leakage.« less

Calcium Apatite Deposition Disease: Diagnosis and Treatment

PubMed Central

2016-01-01

Calcium apatite deposition disease (CADD) is a common entity characterized by deposition of calcium apatite crystals within and around connective tissues, usually in a periarticular location. CADD most frequently involves the rotator cuff. However, it can theoretically occur in almost any location in the musculoskeletal system, and many different locations of CADD have been described. When CADD presents in an unexpected location it can pose a diagnostic challenge, particularly when associated with pain or swelling, and can be confused with other pathologic processes, such as infection or malignancy. However, CADD has typical imaging characteristics that usually allows for a correct diagnosis to be made without additional imaging or laboratory workup, even when presenting in unusual locations. This is a review of the common and uncommon presentations of CADD in the appendicular and axial skeleton as well as an updated review of pathophysiology of CADD and current treatments. PMID:28042481

Occult urolithiasis in asymptomatic primary hyperparathyroidism.

PubMed

Tay, Yu-Kwang Donovan; Liu, Minghao; Bandeira, Leonardo; Bucovsky, Mariana; Lee, James A; Silverberg, Shonni J; Walker, Marcella D

2018-05-01

Recent international guidelines suggest renal imaging to detect occult urolithiasis in all patients with asymptomatic primary hyperparathyroidism (PHPT), but data regarding their prevalence and associated risk factors are limited. We evaluated the prevalence and risk factors for occult urolithiasis. Cross-sectional analysis of 96 asymptomatic PHPT patients from a university hospital in the United States with and without occult nephrolithiasis. Occult urolithiasis was identified in 21% of patients. Stone formers had 47% higher 24-hour urinary calcium excretion (p = 0.002). Although available in only a subset of patients (n = 28), activated vitamin D [1,25(OH) 2 D] was 29% higher (p = 0.02) in stone formers. There was no difference in demographics, BMI, calcium or vitamin D intake, other biochemistries, renal function, BMD, or fractures. Receiver operating characteristic curves indicated that urinary calcium excretion and 1,25(OH) 2 D had an area under the curve of 0.724 (p = 0.003) and 0.750 (p = 0.04), respectively. A urinary calcium threshold of >211mg/day provided a sensitivity of 84.2% and a specificity of 55.3% while a 1,25(OH) 2 D threshold of >91pg/mL provided a sensitivity and specificity of 62.5% and 90.0% respectively for the presence of stones. Occult urolithiasis is present in about one-fifth of patients with asymptomatic PHPT and is associated with higher urinary calcium and 1,25(OH) 2 D. Given that most patients will not have occult urolithiasis, targeted imaging in those most likely to have occult stones rather than screening all asymptomatic PHPT patients may be useful. The higher sensitivity of urinary calcium versus 1,25(OH) 2 D suggests screening those with higher urinary calcium may be an appropriate approach.

Population calcium imaging of spontaneous respiratory and novel motor activity in the facial nucleus and ventral brainstem in newborn mice

PubMed Central

Persson, Karin; Rekling, Jens C

2011-01-01

Abstract The brainstem contains rhythm and pattern forming circuits, which drive cranial and spinal motor pools to produce respiratory and other motor patterns. Here we used calcium imaging combined with nerve recordings in newborn mice to reveal spontaneous population activity in the ventral brainstem and in the facial nucleus. In Fluo-8 AM loaded brainstem–spinal cord preparations, respiratory activity on cervical nerves was synchronized with calcium signals at the ventrolateral brainstem surface. Individual ventrolateral neurons at the level of the parafacial respiratory group showed perfect or partial synchrony with respiratory nerve bursts. In brainstem–spinal cord preparations, cut at the level of the mid-facial nucleus, calcium signals were recorded in the dorsal, lateral and medial facial subnuclei during respiratory activity. Strong activity initiated in the dorsal subnucleus, followed by activity in lateral and medial subnuclei. Whole-cell recordings from facial motoneurons showed weak respiratory drives, and electrical field potential recordings confirmed respiratory drive to particularly the dorsal and lateral subnuclei. Putative facial premotoneurons showed respiratory-related calcium signals, and were predominantly located dorsomedial to the facial nucleus. A novel motor activity on facial, cervical and thoracic nerves was synchronized with calcium signals at the ventromedial brainstem extending from the level of the facial nucleus to the medulla–spinal cord border. Cervical dorsal root stimulation induced similar ventromedial activity. The medial facial subnucleus showed calcium signals synchronized with this novel motor activity on cervical nerves, and cervical dorsal root stimulation induced similar medial facial subnucleus activity. In conclusion, the dorsal and lateral facial subnuclei are strongly respiratory-modulated, and the brainstem contains a novel pattern forming circuit that drives the medial facial subnucleus and cervical motor pools. PMID:21486812

Stone size limits the use of Hounsfield units for prediction of calcium oxalate stone composition.

PubMed

Stewart, Gregory; Johnson, Lewis; Ganesh, Halemane; Davenport, Daniel; Smelser, Woodson; Crispen, Paul; Venkatesh, Ramakrishna

2015-02-01

To evaluate the role of stone size in predicting urinary calculus composition using Hounsfield units on noncontrasted computed tomography (CT) scan. A retrospective review was performed for all patients who underwent ureteroscopy or percutaneous nephrolithotomy during a 1-year period, had a stone analysis performed, and had CT imaging available for review. All CT scans were reviewed by a board-certified radiologist. Variables evaluated included age, sex, body mass index, stone size, stone location, Hounsfield units (HUs), and stone composition. We identified a total of 91 patients (41 men and 50 women) with CT imaging and stone analysis available for review. Stone analysis showed 41 calcium oxalate monohydrate (CaOxMH), 13 calcium oxalate dihydrate, 29 calcium phosphate, 5 uric acid, 2 struvite, and 1 cystine stone. Average age was 46 years, and average body mass index was 32 kg/m2. Measured HUs varied significantly with size for CaOxMH and calcium oxalate dihydrate stones (P values <.05), but not for calcium phosphate stones (P = .126). Using a CaOxMH identification value of 700-1000 HUs, 28 of 41 stone compositions (68%) would not have been correctly identified, including all 10 (100%) small (<5 mm) stones, 13 of 22 (59%) medium (5-10 mm) stones, and 5 of 9 large (>10 mm) stones (55%). For calcium stones, the ability of CT HUs to predict stone composition was limited, likely due to the mixed stone composition. Within a cohort of CaOxMH stone formers, measured HUs varied linearly with stone size. All stones <5 mm were below thresholds for CaOxMH composition. Copyright © 2015 Elsevier Inc. All rights reserved.

Ionotropic and Metabotropic Mechanisms of Allosteric Modulation of α7 Nicotinic Receptor Intracellular Calcium.

PubMed

King, Justin R; Ullah, Aman; Bak, Ellen; Jafri, M Saleet; Kabbani, Nadine

2018-06-01

The pharmacological targeting of the α 7 nicotinic acetylcholine receptor ( α 7) is a promising strategy in the development of new drugs for neurologic diseases. Because α 7 receptors regulate cellular calcium, we investigated how the prototypical type II-positive allosteric modulator PNU120596 affects α 7-mediated calcium signaling. Live imaging experiments show that PNU120596 augments ryanodine receptor-driven calcium-induced calcium release (CICR), inositol-induced calcium release (IICR), and phospholipase C activation by the α 7 receptor. Both influx of calcium through the α 7 nicotinic acetylcholine receptor (nAChR) channel as well as the binding of intracellular G proteins were involved in the effect of PNU120596 on intracellular calcium. This is evidenced by the findings that chelation of extracellular calcium, expression of α 7 D44A or α 7 345-348A mutant subunits, or blockade of calcium store release compromised the ability of PNU120596 to increase intracellular calcium transients generated by α 7 ligand activation. Spatiotemporal stochastic modeling of calcium transient responses corroborates these results and indicates that α 7 receptor activation enables calcium microdomains locally and to lesser extent in the distant cytosol. From the model, allosteric modulation of the receptor activates CICR locally via ryanodine receptors and augments IICR through enhanced calcium influx due to prolonged α 7 nAChR opening. These findings provide a new mechanistic framework for understanding the effect of α 7 receptor allosteric modulation on both local and global calcium dynamics. Copyright © 2018 by The American Society for Pharmacology and Experimental Therapeutics.

Scaffolds with a standardized macro-architecture fabricated from several calcium phosphate ceramics using an indirect rapid prototyping technique

PubMed Central

Wilson, C. E.; van Blitterswijk, C. A.; Verbout, A. J.; de Bruijn, J. D.

2010-01-01

Calcium phosphate ceramics, commonly applied as bone graft substitutes, are a natural choice of scaffolding material for bone tissue engineering. Evidence shows that the chemical composition, macroporosity and microporosity of these ceramics influences their behavior as bone graft substitutes and bone tissue engineering scaffolds but little has been done to optimize these parameters. One method of optimization is to place focus on a particular parameter by normalizing the influence, as much as possible, of confounding parameters. This is difficult to accomplish with traditional fabrication techniques. In this study we describe a design based rapid prototyping method of manufacturing scaffolds with virtually identical macroporous architectures from different calcium phosphate ceramic compositions. Beta-tricalcium phosphate, hydroxyapatite (at two sintering temperatures) and biphasic calcium phosphate scaffolds were manufactured. The macro- and micro-architectures of the scaffolds were characterized as well as the influence of the manufacturing method on the chemistries of the calcium phosphate compositions. The structural characteristics of the resulting scaffolds were remarkably similar. The manufacturing process had little influence on the composition of the materials except for the consistent but small addition of, or increase in, a beta-tricalcium phosphate phase. Among other applications, scaffolds produced by the method described provide a means of examining the influence of different calcium phosphate compositions while confidently excluding the influence of the macroporous structure of the scaffolds. PMID:21069558

Action of cholecystokinin and cholinergic agents on calcium transport in isolated pancreatic acinar cells.

PubMed Central

Gardner, J D; Conlon, T P; Kleveman, H L; Adams, T D; Ondetti, M A

1975-01-01

COOH-terminal octapeptide of cholecystokinin (CCK-octapeptide) and the cholinergic agent carbamylcholine each produced a fourfold stimulation of calcium outflux in guinea pig isolated pancreatic acinar cells. Neither agent altered calcium influx. Stimulation of calcium outflux was rapid and specific, was abolished by reducing the incubation temperature to 4 degrees C, and was a saturable function of the secretagogue concentration. The concentrations of CCK-octapeptide and carbamylcholine that produced half-maximal stimulation of calcium outflux were 3.1 x 10(-10) M and 4.9 x 10(-5) M, respectively. The cholinergic antagonist antropine competitively inhibited carbamylcholine stimulation of calcium outflux but did not alter stimulation produced by CCK-octapeptide. Stimulation of calcium outflux by maximal concentrations of carbamycholine plus CCK-octapeptide was the same as that produced by a maximal concentration of either agent alone.Calcium outflux became refractory to stimulation by secretagogues, and incubation with either CCK-ostapeptide or carbamylcholine produced a refractoriness to both agents. The relative potencies with CCK and its related fragments stimulated calcium outflux were CCK-octapeptide greater than heptapeptide greater than CCK greater than hexapeptide = gastrin. Secretin, glucagon, and vasoactive intestinal peptide, at concentrations as high as 10(-5) M, failed to alter calcium outflux and did not affect stimulation by CCK-octapeptide or by carbamycholine. Images PMID:1150877

Efficacy of Mixtures of Magnesium, Citrate and Phytate as Calcium Oxalate Crystallization Inhibitors in Urine.

PubMed

Grases, Felix; Rodriguez, Adrian; Costa-Bauza, Antonia

2015-09-01

The main aim of the current study was to evaluate the effectiveness of mixtures of magnesium, citrate and phytate as calcium oxalate crystallization inhibitors. A turbidimetric assay in synthetic urine was performed to obtain induction times for calcium oxalate crystallization in the absence and presence of different mixtures of inhibitors. The morphology of calcium oxalate crystals in the absence or presence of inhibitors and mixtures of the inhibitors was evaluated in 2 crystallization experiments at low and high calcium oxalate supersaturation. The crystals formed were examined using scanning electron microscopy. Examination of crystallization induction times revealed clear inhibitory effects of magnesium, citrate and phytate on calcium oxalate crystallization, supporting usefulness in the treatment and prevention of calcium oxalate nephrolithiasis. Significant synergistic effects between magnesium and phytate were observed. Scanning electron microscopy images revealed that phytate is a powerful crystal growth inhibitor of calcium oxalate, totally preventing the formation of trihydrate and monohydrate. In addition to crystallization inhibition capacity, citrate and magnesium avoided calcium oxalate crystallization by decreasing its supersaturation. The synergistic effect between magnesium and phytate on calcium oxalate crystallization suggests that a combination of these 2 compounds may be highly useful as antilithiasis therapy. Copyright © 2015 American Urological Association Education and Research, Inc. Published by Elsevier Inc. All rights reserved.

Calcium release-dependent inactivation precedes formation of the tubular system in developing rat cardiac myocytes.

PubMed

Macková, Katarina; Zahradníková, Alexandra; Hoťka, Matej; Hoffmannová, Barbora; Zahradník, Ivan; Zahradníková, Alexandra

2017-12-01

Developing cardiac myocytes undergo substantial structural and functional changes transforming the mechanism of excitation-contraction coupling from the embryonic form, based on calcium influx through sarcolemmal DHPR calcium channels, to the adult form, relying on local calcium release through RYR calcium channels of sarcoplasmic reticulum stimulated by calcium influx. We characterized day-by-day the postnatal development of the structure of sarcolemma, using techniques of confocal fluorescence microscopy, and the development of the calcium current, measured by the whole-cell patch-clamp in isolated rat ventricular myocytes. We characterized the appearance and expansion of the t-tubule system and compared it with the appearance and progress of the calcium current inactivation induced by the release of calcium ions from sarcoplasmic reticulum as structural and functional measures of direct DHPR-RYR interaction. The release-dependent inactivation of calcium current preceded the development of the t-tubular system by several days, indicating formation of the first DHPR-RYR couplons at the surface sarcolemma and their later spreading close to contractile myofibrils with the growing t-tubules. Large variability of both of the measured parameters among individual myocytes indicates uneven maturation of myocytes within the growing myocardium.

Sub-micron opto-chemical probes for studying living neurons

NASA Astrophysics Data System (ADS)

Hossein-Zadeh, M.; Delgado, J.; Schweizer, F.; Lieberman, R.

2017-02-01

We have fabricated sub-micron opto-chemical probes for pH, oxygen and calcium monitoring and demonstrated their application in intracellular and extracellular monitoring of neurons (cortical neuronal cultures and acute hippocampal slices). Using these probes, we have measured extracellular pH in the stratum radiatum of the CA1 region of mouse hippocampus upon stimulation of presynaptic Schaffer collateral axons. Synaptic transmission was monitored using standard electrophysiological techniques. We find that the local pH transiently changes in response to synaptic stimulation. In addition, the geometry of the functionalized region on the probe combined with high sensitivity imaging enables simultaneous monitoring of spatially adjacent but distinct compartments. As proof of concept we impaled cultured neurons with the probe measured calcium and pH inside as well as directly outside of neurons as we changed the pH and calcium concentration in the physiological solution in the perfusion chamber. As such these probes can be used to study the impact of the environment on both cellular and extra-cellular space. Additionally as the chemical properties of the surrounding medium can be controlled and monitored with high precision, these probes enable differential measurement of the target parameter referenced to a stable bath. This approach eliminates the uncertainties associated with non-chemical fluctuations in the fluorescent emission and result in a self-calibrated opto-chemical probe. We have also demonstrated multifunctional probes that are capable of measuring up to three parameters in the extracellular space in brain slices.

Basolateral K+ channel involvement in forskolin-activated chloride secretion in human colon

PubMed Central

McNamara, Brian; Winter, Desmond C; Cuffe, John E; O'Sullivan, Gerald C; Harvey, Brian J

1999-01-01

In this study we investigated the role of basolateral potassium transport in maintaining cAMP-activated chloride secretion in human colonic epithelium. Ion transport was quantified in isolated human colonic epithelium using the short-circuit current technique. Basolateral potassium transport was studied using nystatin permeabilization. Intracellular calcium measurements were obtained from isolated human colonic crypts using fura-2 spectrofluorescence imaging. In intact isolated colonic strips, forskolin and prostaglandin E2 (PGE2) activated an inward transmembrane current (ISC) consistent with anion secretion (for forskolin ΔISC = 63.8 ± 6.2 μA cm−2, n = 6; for PGE2 ΔISC = 34.3 ± 5.2 μA cm−2, n = 6). This current was inhibited in chloride-free Krebs solution or by inhibiting basolateral chloride uptake with bumetanide and 4,4′-diisothiocyanatostilbene-2,2′-disulfonic acid (DIDS). The forskolin- and PGE2-induced chloride secretion was inhibited by basolateral exposure to barium (5 mM), tetrapentylammonium (10 μM) and tetraethylammonium (10 mM). The transepithelial current produced under an apical to serosal K+ gradient in nystatin-perforated colon is generated at the basolateral membrane by K+ transport. Forskolin failed to activate this current under conditions of high or low calcium and failed to increase the levels of intracellular calcium in isolated crypts In conclusion, we propose that potassium recycling through basolateral K+ channels is essential for cAMP-activated chloride secretion. PMID:10432355

Functional Assay of Cancer Cell Invasion Potential Based on Mechanotransduction of Focused Ultrasound.

PubMed

Weitz, Andrew C; Lee, Nan Sook; Yoon, Chi Woo; Bonyad, Adrineh; Goo, Kyo Suk; Kim, Seaok; Moon, Sunho; Jung, Hayong; Zhou, Qifa; Chow, Robert H; Shung, K Kirk

2017-01-01

Cancer cells undergo a number of biophysical changes as they transform from an indolent to an aggressive state. These changes, which include altered mechanical and electrical properties, can reveal important diagnostic information about disease status. Here, we introduce a high-throughput, functional technique for assessing cancer cell invasion potential, which works by probing for the mechanically excitable phenotype exhibited by invasive cancer cells. Cells are labeled with fluorescent calcium dye and imaged during stimulation with low-intensity focused ultrasound, a non-contact mechanical stimulus. We show that cells located at the focus of the stimulus exhibit calcium elevation for invasive prostate (PC-3 and DU-145) and bladder (T24/83) cancer cell lines, but not for non-invasive cell lines (BPH-1, PNT1A, and RT112/84). In invasive cells, ultrasound stimulation initiates a calcium wave that propagates from the cells at the transducer focus to other cells, over distances greater than 1 mm. We demonstrate that this wave is mediated by extracellular signaling molecules and can be abolished through inhibition of transient receptor potential channels and inositol trisphosphate receptors, implicating these proteins in the mechanotransduction process. If validated clinically, our technology could provide a means to assess tumor invasion potential in cytology specimens, which is not currently possible. It may therefore have applications in diseases such as bladder cancer, where cytologic diagnosis of tumor invasion could improve clinical decision-making.

Functional Assay of Cancer Cell Invasion Potential Based on Mechanotransduction of Focused Ultrasound

PubMed Central

Weitz, Andrew C.; Lee, Nan Sook; Yoon, Chi Woo; Bonyad, Adrineh; Goo, Kyo Suk; Kim, Seaok; Moon, Sunho; Jung, Hayong; Zhou, Qifa; Chow, Robert H.; Shung, K. Kirk

2017-01-01

Cancer cells undergo a number of biophysical changes as they transform from an indolent to an aggressive state. These changes, which include altered mechanical and electrical properties, can reveal important diagnostic information about disease status. Here, we introduce a high-throughput, functional technique for assessing cancer cell invasion potential, which works by probing for the mechanically excitable phenotype exhibited by invasive cancer cells. Cells are labeled with fluorescent calcium dye and imaged during stimulation with low-intensity focused ultrasound, a non-contact mechanical stimulus. We show that cells located at the focus of the stimulus exhibit calcium elevation for invasive prostate (PC-3 and DU-145) and bladder (T24/83) cancer cell lines, but not for non-invasive cell lines (BPH-1, PNT1A, and RT112/84). In invasive cells, ultrasound stimulation initiates a calcium wave that propagates from the cells at the transducer focus to other cells, over distances greater than 1 mm. We demonstrate that this wave is mediated by extracellular signaling molecules and can be abolished through inhibition of transient receptor potential channels and inositol trisphosphate receptors, implicating these proteins in the mechanotransduction process. If validated clinically, our technology could provide a means to assess tumor invasion potential in cytology specimens, which is not currently possible. It may therefore have applications in diseases such as bladder cancer, where cytologic diagnosis of tumor invasion could improve clinical decision-making. PMID:28824873

Registering 2D and 3D imaging data of bone during healing.

PubMed

Hoerth, Rebecca M; Baum, Daniel; Knötel, David; Prohaska, Steffen; Willie, Bettina M; Duda, Georg N; Hege, Hans-Christian; Fratzl, Peter; Wagermaier, Wolfgang

2015-04-01

PURPOSE/AIMS OF THE STUDY: Bone's hierarchical structure can be visualized using a variety of methods. Many techniques, such as light and electron microscopy generate two-dimensional (2D) images, while micro-computed tomography (µCT) allows a direct representation of the three-dimensional (3D) structure. In addition, different methods provide complementary structural information, such as the arrangement of organic or inorganic compounds. The overall aim of the present study is to answer bone research questions by linking information of different 2D and 3D imaging techniques. A great challenge in combining different methods arises from the fact that they usually reflect different characteristics of the real structure. We investigated bone during healing by means of µCT and a couple of 2D methods. Backscattered electron images were used to qualitatively evaluate the tissue's calcium content and served as a position map for other experimental data. Nanoindentation and X-ray scattering experiments were performed to visualize mechanical and structural properties. We present an approach for the registration of 2D data in a 3D µCT reference frame, where scanning electron microscopies serve as a methodic link. Backscattered electron images are perfectly suited for registration into µCT reference frames, since both show structures based on the same physical principles. We introduce specific registration tools that have been developed to perform the registration process in a semi-automatic way. By applying this routine, we were able to exactly locate structural information (e.g. mineral particle properties) in the 3D bone volume. In bone healing studies this will help to better understand basic formation, remodeling and mineralization processes.

Calcium activation of frog slow muscle fibres

PubMed Central

Costantin, L. L.; Podolsky, R. J.; Tice, Lois W.

1967-01-01

1. Skinned muscle fibres were prepared from the tonus bundle of the frog iliofibularis muscle and the contractile response elicited by applied calcium ions was studied. The fibre type was determined by electron microscopy. 2. Fast fibres shortened many times more rapidly than slow fibres, indicating that the slow contraction of slow fibres is an inherent property of the contractile mechanism. 3. The extent of spread of contraction following local calcium application was much greater in slow than in fast fibres, a difference which is consistent with the relative sparsity of the sarcoplasmic reticulum in slow fibres. 4. The ability of the sarcoplasmic reticulum of slow fibres to accumulate calcium was demonstrated by the in situ immobilization of calcium when oxalate solutions were added to the skinned fibre. ImagesPlate 1Plate 2Plate 3Plate 4Plate 5AB PMID:6030519

Mystery #16 Answer

Atmospheric Science Data Center

2013-04-22

... were acquired by the Multi-angle Imaging SpectroRadiometer (MISR) during October and November 2003. The two images represent about 310 ... obtain calcium from the seawater and carbon dioxide from cell respiration, and bring these products into the internal tissues of the ...

Calcium neuroimaging in behaving zebrafish larvae using a turn-key light field camera

NASA Astrophysics Data System (ADS)

Cruz Perez, Carlos; Lauri, Antonella; Symvoulidis, Panagiotis; Cappetta, Michele; Erdmann, Arne; Westmeyer, Gil Gregor

2015-09-01

Reconstructing a three-dimensional scene from multiple simultaneously acquired perspectives (the light field) is an elegant scanless imaging concept that can exceed the temporal resolution of currently available scanning-based imaging methods for capturing fast cellular processes. We tested the performance of commercially available light field cameras on a fluorescent microscopy setup for monitoring calcium activity in the brain of awake and behaving reporter zebrafish larvae. The plenoptic imaging system could volumetrically resolve diverse neuronal response profiles throughout the zebrafish brain upon stimulation with an aversive odorant. Behavioral responses of the reporter fish could be captured simultaneously together with depth-resolved neuronal activity. Overall, our assessment showed that with some optimizations for fluorescence microscopy applications, commercial light field cameras have the potential of becoming an attractive alternative to custom-built systems to accelerate molecular imaging research on cellular dynamics.

Calcium neuroimaging in behaving zebrafish larvae using a turn-key light field camera.

PubMed

Perez, Carlos Cruz; Lauri, Antonella; Symvoulidis, Panagiotis; Cappetta, Michele; Erdmann, Arne; Westmeyer, Gil Gregor

2015-09-01

Reconstructing a three-dimensional scene from multiple simultaneously acquired perspectives (the light field) is an elegant scanless imaging concept that can exceed the temporal resolution of currently available scanning-based imaging methods for capturing fast cellular processes. We tested the performance of commercially available light field cameras on a fluorescent microscopy setup for monitoring calcium activity in the brain of awake and behaving reporter zebrafish larvae. The plenoptic imaging system could volumetrically resolve diverse neuronal response profiles throughout the zebrafish brain upon stimulation with an aversive odorant. Behavioral responses of the reporter fish could be captured simultaneously together with depth-resolved neuronal activity. Overall, our assessment showed that with some optimizations for fluorescence microscopy applications, commercial light field cameras have the potential of becoming an attractive alternative to custom-built systems to accelerate molecular imaging research on cellular dynamics.

The Determination of Vitamin D-Dependent Calcium Binding Protein in Chick Intesting: An Undergraduate Biochemistry Laboratory Experiment.

ERIC Educational Resources Information Center

Lessard, George M.

1980-01-01

Described is an experiment used in an undergraduate biochemistry laboratory involving inducing rickets in chicks and correlating the disease to a reduction in vitamin D-dependent calcium binding protein. Techniques involved are hormone induction, protein isolation, and radioisotope methodology. (Author/DS)

Imaging extracellular calcium in endolymph

NASA Astrophysics Data System (ADS)

Strimbu, C. Elliott; Fridberger, Anders

2018-05-01

Hair cell mechanoelectrical transduction and adaptation are believed to be regulated by extracellular calcium. However, the majority of experiments addressing calcium's role have been performed on reduced preparations in conditions that do not mimic those present in vivo. We used confocal microscopy and a low affinity (kd ˜11 µM) ratiometric fluorescent indicator to measure the extracellular calcium concentration in scala media in an in vitro preparation of the guinea pig cochlea. Microelectrodes were used to measure the cochlear microphonic potential during acoustic stimulation. The mean calcium concentration is significantly higher in the tectorial membrane (TM) than the surrounding endolymph, suggesting that the membrane acts as a calcium sink. We also observe calcium hot spots along the underside of the TM, near the outer hair cell bundles and near Hensens stripe close to the inner hair cell bundle. This suggests that the local calcium concentration near the hair bundles exceeds 100 µM, significantly higher than the bulk endolymph. These results were corroborated with fluorescence correlation spectroscopy using a second calcium sensitive dye, Oregon Green 488-BAPTA. Following a brief exposure to loud sound, TM calcium drops dramatically and shows recovery on a similar timescale as the microphonic potential. Our results suggest that the extracellular calcium concentration near the hair bundles is much higher than previously believed and may also serve as a partial control parameter for temporary threshold shifts.

Musculoskeletal ultrasound: how to treat calcific tendinitis of the rotator cuff by ultrasound-guided single-needle lavage technique.

PubMed

Lee, Kenneth S; Rosas, Humberto G

2010-09-01

The purpose of this video article is to illustrate the ultrasound appearance of calcium deposition in the rotator cuff and provide a detailed step-by-step protocol for performing the ultrasound-guided single-needle lavage technique for the treatment of calcific tendinitis with emphasis on patient positioning, necessary supplies, real-time lavage technique, and steroid injection into the subacromial subdeltoid bursa. Musculoskeletal ultrasound is well established as a safe, cost-effective imaging tool in diagnosing and treating common musculoskeletal disorders. Calcific tendinitis of the rotator cuff is a common disabling cause of shoulder pain. Although most cases are self-limiting, a subset of patients is refractory to conservative therapy and requires treatment intervention. Ultrasound-guided lavage is an effective and safe minimally-invasive treatment not readily offered in the United States as an alternative to surgery, perhaps because of the limited prevalence of musculoskeletal ultrasound programs and limited training. On completion of this video article, the participant should be able to develop an appropriate diagnostic and therapeutic algorithm for the treatment of calcific tendinitis of the rotator cuff using ultrasound.

Observation of the molecular organization of calcium release sites in fast- and slow-twitch skeletal muscle with nanoscale imaging.

PubMed

Jayasinghe, Isuru D; Munro, Michelle; Baddeley, David; Launikonis, Bradley S; Soeller, Christian

2014-10-06

Localization microscopy is a fairly recently introduced super-resolution fluorescence imaging modality capable of achieving nanometre-scale resolution. We have applied the dSTORM variation of this method to image intracellular molecular assemblies in skeletal muscle fibres which are large cells that critically rely on nanoscale signalling domains, the triads. Immunofluorescence staining in fixed adult rat skeletal muscle sections revealed clear differences between fast- and slow-twitch fibres in the molecular organization of ryanodine receptors (RyRs; the primary calcium release channels) within triads. With the improved resolution offered by dSTORM, abutting arrays of RyRs in transverse view of fast fibres were observed in contrast to the fragmented distribution on slow-twitch muscle that were approximately 1.8 times shorter and consisted of approximately 1.6 times fewer receptors. To the best of our knowledge, for the first time, we have quantified the nanometre-scale spatial association between triadic proteins using multi-colour super-resolution, an analysis difficult to conduct with electron microscopy. Our findings confirm that junctophilin-1 (JPH1), which tethers the sarcoplasmic reticulum ((SR) intracellular calcium store) to the tubular (t-) system at triads, was present throughout the RyR array, whereas JPH2 was contained within much smaller nanodomains. Similar imaging of the primary SR calcium buffer, calsequestrin (CSQ), detected less overlap of the triad with CSQ in slow-twitch muscle supporting greater spatial heterogeneity in the luminal Ca2+ buffering when compared with fast twitch muscle. Taken together, these nanoscale differences can explain the fundamentally different physiologies of fast- and slow-twitch muscle. © 2014 The Author(s) Published by the Royal Society. All rights reserved.

Observation of the molecular organization of calcium release sites in fast- and slow-twitch skeletal muscle with nanoscale imaging

PubMed Central

Jayasinghe, Isuru D.; Munro, Michelle; Baddeley, David; Launikonis, Bradley S.; Soeller, Christian

2014-01-01

Localization microscopy is a fairly recently introduced super-resolution fluorescence imaging modality capable of achieving nanometre-scale resolution. We have applied the dSTORM variation of this method to image intracellular molecular assemblies in skeletal muscle fibres which are large cells that critically rely on nanoscale signalling domains, the triads. Immunofluorescence staining in fixed adult rat skeletal muscle sections revealed clear differences between fast- and slow-twitch fibres in the molecular organization of ryanodine receptors (RyRs; the primary calcium release channels) within triads. With the improved resolution offered by dSTORM, abutting arrays of RyRs in transverse view of fast fibres were observed in contrast to the fragmented distribution on slow-twitch muscle that were approximately 1.8 times shorter and consisted of approximately 1.6 times fewer receptors. To the best of our knowledge, for the first time, we have quantified the nanometre-scale spatial association between triadic proteins using multi-colour super-resolution, an analysis difficult to conduct with electron microscopy. Our findings confirm that junctophilin-1 (JPH1), which tethers the sarcoplasmic reticulum ((SR) intracellular calcium store) to the tubular (t-) system at triads, was present throughout the RyR array, whereas JPH2 was contained within much smaller nanodomains. Similar imaging of the primary SR calcium buffer, calsequestrin (CSQ), detected less overlap of the triad with CSQ in slow-twitch muscle supporting greater spatial heterogeneity in the luminal Ca2+ buffering when compared with fast twitch muscle. Taken together, these nanoscale differences can explain the fundamentally different physiologies of fast- and slow-twitch muscle. PMID:25100314

Genetically encoded calcium indicators for multi-color neural activity imaging and combination with optogenetics

PubMed Central

Akerboom, Jasper; Carreras Calderón, Nicole; Tian, Lin; Wabnig, Sebastian; Prigge, Matthias; Tolö, Johan; Gordus, Andrew; Orger, Michael B.; Severi, Kristen E.; Macklin, John J.; Patel, Ronak; Pulver, Stefan R.; Wardill, Trevor J.; Fischer, Elisabeth; Schüler, Christina; Chen, Tsai-Wen; Sarkisyan, Karen S.; Marvin, Jonathan S.; Bargmann, Cornelia I.; Kim, Douglas S.; Kügler, Sebastian; Lagnado, Leon; Hegemann, Peter; Gottschalk, Alexander; Schreiter, Eric R.; Looger, Loren L.

2013-01-01

Genetically encoded calcium indicators (GECIs) are powerful tools for systems neuroscience. Here we describe red, single-wavelength GECIs, “RCaMPs,” engineered from circular permutation of the thermostable red fluorescent protein mRuby. High-resolution crystal structures of mRuby, the red sensor RCaMP, and the recently published red GECI R-GECO1 give insight into the chromophore environments of the Ca2+-bound state of the sensors and the engineered protein domain interfaces of the different indicators. We characterized the biophysical properties and performance of RCaMP sensors in vitro and in vivo in Caenorhabditis elegans, Drosophila larvae, and larval zebrafish. Further, we demonstrate 2-color calcium imaging both within the same cell (registering mitochondrial and somatic [Ca2+]) and between two populations of cells: neurons and astrocytes. Finally, we perform integrated optogenetics experiments, wherein neural activation via channelrhodopsin-2 (ChR2) or a red-shifted variant, and activity imaging via RCaMP or GCaMP, are conducted simultaneously, with the ChR2/RCaMP pair providing independently addressable spectral channels. Using this paradigm, we measure calcium responses of naturalistic and ChR2-evoked muscle contractions in vivo in crawling C. elegans. We systematically compare the RCaMP sensors to R-GECO1, in terms of action potential-evoked fluorescence increases in neurons, photobleaching, and photoswitching. R-GECO1 displays higher Ca2+ affinity and larger dynamic range than RCaMP, but exhibits significant photoactivation with blue and green light, suggesting that integrated channelrhodopsin-based optogenetics using R-GECO1 may be subject to artifact. Finally, we create and test blue, cyan, and yellow variants engineered from GCaMP by rational design. This engineered set of chromatic variants facilitates new experiments in functional imaging and optogenetics. PMID:23459413

Increased calcium absorption from synthetic stable amorphous calcium carbonate: double-blind randomized crossover clinical trial in postmenopausal women.

PubMed

Vaisman, Nachum; Shaltiel, Galit; Daniely, Michal; Meiron, Oren E; Shechter, Assaf; Abrams, Steven A; Niv, Eva; Shapira, Yami; Sagi, Amir

2014-10-01

Calcium supplementation is a widely recognized strategy for achieving adequate calcium intake. We designed this blinded, randomized, crossover interventional trial to compare the bioavailability of a new stable synthetic amorphous calcium carbonate (ACC) with that of crystalline calcium carbonate (CCC) using the dual stable isotope technique. The study was conducted in the Unit of Clinical Nutrition, Tel Aviv Sourasky Medical Center, Israel. The study population included 15 early postmenopausal women aged 54.9 ± 2.8 (mean ± SD) years with no history of major medical illness or metabolic bone disorder, excess calcium intake, or vitamin D deficiency. Standardized breakfast was followed by randomly provided CCC or ACC capsules containing 192 mg elemental calcium labeled with 44Ca at intervals of at least 3 weeks. After swallowing the capsules, intravenous CaCl2 labeled with 42Ca on was administered on each occasion. Fractional calcium absorption (FCA) of ACC and CCC was calculated from the 24-hour urine collection following calcium administration. The results indicated that FCA of ACC was doubled (± 0.96 SD) on average compared to that of CCC (p < 0.02). The higher absorption of the synthetic stable ACC may serve as a more efficacious way of calcium supplementation. © 2014 American Society for Bone and Mineral Research.

Whiting in Lake Michigan

NASA Technical Reports Server (NTRS)

2002-01-01

Satellites provide a view from space of changes on the Earth's surface. This series of images from the Sea-viewing Wide Field-of-view Sensor (SeaWiFS) aboard the Orbview-2 satellite shows the dramatic change in the color of Lake Michigan during the summer. The bright color that appears in late summer is probably caused by calcium carbonate-chalk-in the water. Lake Michigan always has a lot of calcium carbonate in it because the floor of the lake is limestone. During most of the year the calcium carbonate remains dissolved in the cold water, but at the end of summer the lake warms up, lowering the solubility of calcium carbonate. As a result, the calcium carbonate precipitates out of the water, forming clouds of very small solid particles that appear as bright swirls from above. The phenomenon is appropriately called a whiting event. A similar event occured in 1999, but appears to have started later and subsided earlier. It is also possible that a bloom of the algae Microcystis is responsible for the color change, but unlikely because of Lake Michigan's depth and size. Microcystis blooms have occured in other lakes in the region, however. On the shore of the lake it is possible to see the cities of Chicago, Illinois, and Milwaukee, Wisconsin. Both appear as clusters of gray-brown pixels. Image courtesy the SeaWiFS Project, NASA/Goddard Space Flight Center, and ORBIMAGE

Crystal Structures of the GCaMP Calcium Sensor Reveal the Mechanism of Fluorescence Signal Change and Aid Rational Design

DOE Office of Scientific and Technical Information (OSTI.GOV)

Akerboom, Jasper; Velez Rivera, Jonathan D.; Rodriguez Guilbe, María M.

The genetically encoded calcium indicator GCaMP2 shows promise for neural network activity imaging, but is currently limited by low signal-to-noise ratio. We describe x-ray crystal structures as well as solution biophysical and spectroscopic characterization of GCaMP2 in the calcium-free dark state, and in two calcium-bound bright states: a monomeric form that dominates at intracellular concentrations observed during imaging experiments and an unexpected domain-swapped dimer with decreased fluorescence. This series of structures provides insight into the mechanism of Ca{sup 2+}-induced fluorescence change. Upon calcium binding, the calmodulin (CaM) domain wraps around the M13 peptide, creating a new domain interface between CaMmore » and the circularly permuted enhanced green fluorescent protein domain. Residues from CaM alter the chemical environment of the circularly permuted enhanced green fluorescent protein chromophore and, together with flexible inter-domain linkers, block solvent access to the chromophore. Guided by the crystal structures, we engineered a series of GCaMP2 point mutants to probe the mechanism of GCaMP2 function and characterized one mutant with significantly improved signal-to-noise. The mutation is located at a domain interface and its effect on sensor function could not have been predicted in the absence of structural data.« less

Modification of the N-Terminus of a Calcium Carbonate Precipitating Peptide Affects Calcium Carbonate Mineralization.

PubMed

Usui, Kenji; Yokota, Shin-Ichiro; Ozaki, Makoto; Sakashita, Shungo; Imai, Takahito; Tomizaki, Kin-Ya

2018-01-01

A core sequence (the 9 C-terminal residues) of calcification-associated peptide (CAP- 1) isolated from the exoskeleton of the red swamp crayfish was previously shown to control calcium carbonate precipitation with chitin. In addition, a modified core sequence in which the phosphorylated serine at the N terminus is replaced with serine exhibits was also previously shown to alter precipitation characteristics with chitin. We focused on calcium carbonate precipitation and attempted to elucidate aspects of the mechanism underlying mineralization. We attempted to evaluate in detail the effects of modifying the N-terminus in the core sequence on calcium carbonate mineralization without chitin. The peptide modifications included phosphorylation, dephosphorylation, and a free or acetylated Nterminus. The peptides were synthesized manually on Wang resin using the DIPCI-DMAP method for the first residue, and Fmoc solid phase peptide synthesis with HBTU-HOBt for the subsequent residues. Prior to calcium carbonate precipitation, calcium carbonate was suspended in MilliQ water. Carbon dioxide gas was bubbled into the stirred suspension, then the remaining solid CaCO3 was removed by filtration. The concentration of calcium ions in the solution was determined by standard titration with ethylenediaminetetraacetate. Calcium carbonate precipitation was conducted in a micro tube for 3 h at 37°C. We used the micro-scale techniques AFM (atomic force microscopy) and TEM (transmission electron microscopy), and the macro-scale techniques chelate titration, HPLC, gel filtration, CD (circular dichroism) and DLS (dynamic light scattering). We determined the morphologies of the calcium carbonate deposits using AFM and TEM. The pS peptide provided the best control of the shape and size of the calcium carbonate round particles. The acetylated peptides (Ac-S and Ac-pS) provided bigger particles with various shapes. S peptide provided a mixture of bigger particles and amorphous particles. We verified these findings using DLS. All the peptide samples produced nanostructures of the expected size in agreement with the AFM and TEM results. We estimated the abilities of these peptides to precipitate calcium carbonate by determining the residual calcium hydrogen carbonate concentration by standard titration with ethylenediaminetetraacetate after calcium carbonate precipitation. The Ac-pS peptide showed the lowest residual calcium hydrogen carbonate concentration whereas the S peptide showed the highest, suggesting that the precipitating activities of these peptides towards calcium carbonate correlated with peptide net charge. Then the gel filtration results showed a large oligomer peak and a small oligomer/monomer peak for all peptide samples in agreement with the AFM, TEM and DLS results. CD measurements showed that all the peptides formed random-coil-like structures. Thus, we used both macro- and micro-observation techniques such as chelate titration, DLS, AFM and TEM to show that the calcium carbonate precipitating activities of four derivatives of the core sequence of CAP-1 may correlate with the peptide net charge. These peptides mainly act as a catalyst rather than as a binder or component of the calcium carbonate deposits (as a template). On the other hand, the morphologies of the calcium carbonate deposits appeared to be dependent on the ability of the peptide to assemble and act as a template. Consequently, elucidating the relationship between peptide sequence and the ability of the peptide to assemble would be indispensable for controlling precipitate morphologies in the near future. This knowledge would provide important clues for elucidating the relationship between peptide sequence and mineralization ability, including deposit morphology and precipitating activity, for use in nanobiochemistry and materials chemistry research. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Fourier transform infrared spectroscopy imaging of live epithelial cancer cells under non-aqueous media.

PubMed

Soh, JunYi; Chueng, Adeline; Adio, Aminat; Cooper, Alan J; Birch, Brian R; Lwaleed, Bashir A

2013-04-01

Fourier transform infrared (FT-IR) imaging is increasingly being applied to biomedical specimens, but strong IR absorption by water complicates live cell imaging. This study investigates the viability of adherent epithelial cells maintained for short periods under mineral oils in order to facilitate live cell spectroscopy using FT-IR with subsequent imaging. The MGH-U1 urothelial or CaCo2 colorectal cancer cell lines were grown on plastic surfaces or mid-range infrared transparent windows. Medium in established cultures was replaced with paraffin mineral oil, or Fluorolube, for up to 2 h, and viability assessed by supravital staining. Drug handling characteristics were also assessed. Imaging of preparations was attempted by reflectance and transmission using a Varian FT-IR microscope. Cells covered by mineral oil remained viable for 2 h, with recovery into normal medium possible. MTT ((3-(4,5-dimethylthlazol-2-yl)-2,5-diphenyl tetrazolium) conversion to crystalline formazan and differential patterns of drug uptake were maintained. The combination of a calcium fluoride substrate, Fluorolube oil, and transmission optics proved best for spectroscopy. Spectral features were used to obtain images of live cells. The viability of cells overlaid with IR transparent oils was assessed as part of a technique to optimise conditions for FT-IR imaging. Images of untreated cells were obtained using both reflectance and transmission. This represents an effective means of imaging live cells by IR spectroscopy, and also means that imaging is not necessarily a terminal event. It also increases options for producing images based on real-time biochemistry in a range of in vitro experimental and 'optical biopsy' contexts.

Interfacial Ca2+ environments in nanocrystalline apatites revealed by dynamic nuclear polarization enhanced 43Ca NMR spectroscopy

NASA Astrophysics Data System (ADS)

Lee, Daniel; Leroy, César; Crevant, Charlène; Bonhomme-Coury, Laure; Babonneau, Florence; Laurencin, Danielle; Bonhomme, Christian; de Paëpe, Gaël

2017-01-01

The interfaces within bones, teeth and other hybrid biomaterials are of paramount importance but remain particularly difficult to characterize at the molecular level because both sensitive and selective techniques are mandatory. Here, it is demonstrated that unprecedented insights into calcium environments, for example the differentiation of surface and core species of hydroxyapatite nanoparticles, can be obtained using solid-state NMR, when combined with dynamic nuclear polarization. Although calcium represents an ideal NMR target here (and de facto for a large variety of calcium-derived materials), its stable NMR-active isotope, calcium-43, is a highly unreceptive probe. Using the sensitivity gains from dynamic nuclear polarization, not only could calcium-43 NMR spectra be obtained easily, but natural isotopic abundance 2D correlation experiments could be recorded for calcium-43 in short experimental time. This opens perspectives for the detailed study of interfaces in nanostructured materials of the highest biological interest as well as calcium-based nanosystems in general.

Preparation and evaluation of solid dispersion of atorvastatin calcium with Soluplus® by spray drying technique.

PubMed

Ha, Eun-Sol; Baek, In-hwan; Cho, Wonkyung; Hwang, Sung-Joo; Kim, Min-Soo

2014-01-01

The aim of the present study was to investigate the effect of Soluplus® on the solubility of atorvastatin calcium and to develop a solid dispersion formulation that can improve the oral bioavailability of atorvastatin calcium. We demonstrated that Soluplus® increases the aqueous solubility of atorvastatin calcium. Several solid dispersion formulations of atorvastatin calcium with Soluplus® were prepared at various drug : carrier ratios by spray drying. Physicochemical analysis demonstrated that atorvastatin calcium is amorphous in each solid dispersion, and the 2 : 8 drug : carrier ratio provided the highest degree of sustained atorvastatin supersaturation. Pharmacokinetic analysis in rats revealed that the 2 : 8 dispersion significantly improved the oral bioavailability of atorvastatin. This study demonstrates that spray-dried Soluplus® solid dispersions can be an effective method for achieving higher atorvastatin plasma levels.

Biophysically realistic minimal model of dopamine neuron

NASA Astrophysics Data System (ADS)

Oprisan, Sorinel

2008-03-01

We proposed and studied a new biophysically relevant computational model of dopaminergic neurons. Midbrain dopamine neurons are involved in motivation and the control of movement, and have been implicated in various pathologies such as Parkinson's disease, schizophrenia, and drug abuse. The model we developed is a single-compartment Hodgkin-Huxley (HH)-type parallel conductance membrane model. The model captures the essential mechanisms underlying the slow oscillatory potentials and plateau potential oscillations. The main currents involved are: 1) a voltage-dependent fast calcium current, 2) a small conductance potassium current that is modulated by the cytosolic concentration of calcium, and 3) a slow voltage-activated potassium current. We developed multidimensional bifurcation diagrams and extracted the effective domains of sustained oscillations. The model includes a calcium balance due to the fundamental importance of calcium influx as proved by simultaneous electrophysiological and calcium imaging procedure. Although there are significant evidences to suggest a partially electrogenic calcium pump, all previous models considered only elecrtogenic pumps. We investigated the effect of the electrogenic calcium pump on the bifurcation diagram of the model and compared our findings against the experimental results.

Calcium absorption is not increased by caseinophosphopeptides.

PubMed

Teucher, Birgit; Majsak-Newman, Gosia; Dainty, Jack R; McDonagh, David; FitzGerald, Richard J; Fairweather-Tait, Susan J

2006-07-01

One of the suggested health benefits of caseinophosphopeptides (CPPs) is their ability to enhance calcium absorption. This possibility is based on the assumption that they resist proteolysis in the upper gastrointestinal tract and maintain calcium in a soluble form at alkaline pH in the distal ileum. The effects of CPP-enriched preparations (containing candidate functional food ingredients) on calcium absorption from a calcium lactate drink were tested. A randomized crossover trial was undertaken in 15 adults in whom we measured the absorption of calcium from a calcium lactate drink (drink A: 400 mg Ca as lactate) and 2 preparations enriched with forms of CPP (1.7 g each; drinks B and C). Both drinks B and C contained 400 mg Ca as calcium lactate plus approximately 100 mg CPP-derived calcium). Each volunteer received the 3 drinks in random order. Absorption was measured by the dual-label calcium stable-isotope technique. The quantity of calcium absorbed was significantly lower from drink A (103 mg) than from drink B (117 mg; P = 0.012) or drink C (121 mg; P = 0.002), which indicated a positive effect of the CPPs. However, because the CPP preparations contributed additional calcium besides that found in the calcium lactate (drink A), fractional absorption of calcium from drink B (23%) was slightly but significantly (P = 0.015) lower than that from drink A (26%). The differences in calcium absorption are unlikely to have any biological significance. CPPs are unsuitable as candidate ingredients for functional foods that are designed to deliver improved calcium nutrition.

Non-invasive techniques for determining musculoskeleton body composition

DOE Office of Scientific and Technical Information (OSTI.GOV)

Cohn, S.H.

1984-01-01

In vivo neutron activation analysis, combined with gamma spectrometry, has ushered in a new era of clinical diagnosis and evaluation of therapies, as well as investigation into and modelling of body composition in both normal individuals and patients suffering from various diseases and dysfunctions. Body composition studies have provided baseline data on such vital constituents as nitrogen, potassium and calcium. The non-invasive measurement techniques are particularly suitable for study of the musculo-skeletal changes in body composition. Of particular relevance here is the measurement of calcium loss in astronauts during prolonged space flights.

Characterization of crystalline structures in Opuntia ficus-indica.

PubMed

Contreras-Padilla, Margarita; Rivera-Muñoz, Eric M; Gutiérrez-Cortez, Elsa; del López, Alicia Real; Rodríguez-García, Mario Enrique

2015-01-01

This research studies the crystalline compounds present in nopal (Opuntia ficus-indica) cladodes. The identification of the crystalline structures was performed using X-ray diffraction, scanning electron microscopy, mass spectrometry, and Fourier transform infrared spectroscopy. The crystalline structures identified were calcium carbonate (calcite) [CaCO3], calcium-magnesium bicarbonate [CaMg(CO3)2], magnesium oxide [MgO], calcium oxalate monohydrate [Ca(C2O4)•(H2O)], potassium peroxydiphosphate [K4P2O8] and potassium chloride [KCl]. The SEM images indicate that calcite crystals grow to dipyramidal, octahedral-like, prismatic, and flower-like structures; meanwhile, calcium-magnesium bicarbonate structures show rhombohedral exfoliation and calcium oxalate monohydrate is present in a drusenoid morphology. These calcium carbonate compounds have a great importance for humans because their bioavailability. This is the first report about the identification and structural analysis of calcium carbonate and calcium-magnesium bicarbonate in nopal cladodes, as well as the presence of magnesium oxide, potassium peroxydiphosphate and potassium chloride in these plants. The significance of the study of the inorganic components of these cactus plants is related with the increasing interest in the potential use of Opuntia as a raw material of products for the food, pharmaceutical, and cosmetic industries.

The effect of calcium on the composition and physical properties of whey protein particles prepared using emulsification.

PubMed

Westerik, Nieke; Scholten, Elke; Corredig, Milena

2015-06-15

Protein microparticles were formed through emulsification of 25% (w/w) whey protein isolate (WPI) solutions containing various concentrations of calcium (0.0-400.0mM) in an oil phase stabilized by polyglycerol polyricinoleate (PGPR). The emulsions were heated (at 80°C) and the microparticles subsequently re-dispersed in an aqueous phase. Light microscopy and scanning electron microscopy (SEM) images revealed that control particles and those prepared with 7.4mM calcium were spherical and smooth. Particles prepared with 15.0mM calcium gained an irregular, cauliflower-like structure, and at concentrations larger than 30.0mM, shells formed and the particles were no longer spherical. These results describe, for the first time, the potential of modulating the properties of dense whey protein particles by using calcium, and may be used as structuring agents for the design of functional food matrices with increased protein and calcium content. Copyright © 2015. Published by Elsevier Ltd.

Imaging Intracellular Ca2+ Signals in Striatal Astrocytes from Adult Mice Using Genetically-encoded Calcium Indicators

PubMed Central

Jiang, Ruotian; Haustein, Martin D.; Sofroniew, Michael V.; Khakh, Baljit S.

2014-01-01

Astrocytes display spontaneous intracellular Ca2+ concentration fluctuations ([Ca2+]i) and in several settings respond to neuronal excitation with enhanced [Ca2+]i signals. It has been proposed that astrocytes in turn regulate neurons and blood vessels through calcium-dependent mechanisms, such as the release of signaling molecules. However, [Ca2+]i imaging in entire astrocytes has only recently become feasible with genetically encoded calcium indicators (GECIs) such as the GCaMP series. The use of GECIs in astrocytes now provides opportunities to study astrocyte [Ca2+]i signals in detail within model microcircuits such as the striatum, which is the largest nucleus of the basal ganglia. In the present report, detailed surgical methods to express GECIs in astrocytes in vivo, and confocal imaging approaches to record [Ca2+]i signals in striatal astrocytes in situ, are described. We highlight precautions, necessary controls and tests to determine if GECI expression is selective for astrocytes and to evaluate signs of overt astrocyte reactivity. We also describe brain slice and imaging conditions in detail that permit reliable [Ca2+]i imaging in striatal astrocytes in situ. The use of these approaches revealed the entire territories of single striatal astrocytes and spontaneous [Ca2+]i signals within their somata, branches and branchlets. The further use and expansion of these approaches in the striatum will allow for the detailed study of astrocyte [Ca2+]i signals in the striatal microcircuitry. PMID:25490346

Imaging intracellular Ca²⁺ signals in striatal astrocytes from adult mice using genetically-encoded calcium indicators.

PubMed

Jiang, Ruotian; Haustein, Martin D; Sofroniew, Michael V; Khakh, Baljit S

2014-11-19

Astrocytes display spontaneous intracellular Ca(2+) concentration fluctuations ([Ca(2+)]i) and in several settings respond to neuronal excitation with enhanced [Ca(2+)]i signals. It has been proposed that astrocytes in turn regulate neurons and blood vessels through calcium-dependent mechanisms, such as the release of signaling molecules. However, [Ca(2+)]i imaging in entire astrocytes has only recently become feasible with genetically encoded calcium indicators (GECIs) such as the GCaMP series. The use of GECIs in astrocytes now provides opportunities to study astrocyte [Ca(2+)]i signals in detail within model microcircuits such as the striatum, which is the largest nucleus of the basal ganglia. In the present report, detailed surgical methods to express GECIs in astrocytes in vivo, and confocal imaging approaches to record [Ca(2+)]i signals in striatal astrocytes in situ, are described. We highlight precautions, necessary controls and tests to determine if GECI expression is selective for astrocytes and to evaluate signs of overt astrocyte reactivity. We also describe brain slice and imaging conditions in detail that permit reliable [Ca(2+)]i imaging in striatal astrocytes in situ. The use of these approaches revealed the entire territories of single striatal astrocytes and spontaneous [Ca(2+)]i signals within their somata, branches and branchlets. The further use and expansion of these approaches in the striatum will allow for the detailed study of astrocyte [Ca(2+)]i signals in the striatal microcircuitry.

Stationary digital chest tomosynthesis for coronary artery calcium scoring

NASA Astrophysics Data System (ADS)

Wu, Gongting; Wang, Jiong; Potuzko, Marci; Harman, Allison; Pearce, Caleb; Shan, Jing; Lee, Yueh Z.; Zhou, Otto; Lu, Jianping

2016-03-01

The coronary artery calcium score (CACS) measures the buildup of calcium on the coronary artery wall and has been shown to be an important predictor of the risk of coronary artery diseases (CAD). Currently CACS is measured using CT, though the relatively high cost and high radiation dose has limited its adoption as a routine screening procedure. Digital Chest Tomosynthesis (DCT), a low dose and low cost alternative to CT, and has been shown to achieve 90% of sensitivity of CT in lung disease screening. However commercial DCT requires long scanning time and cannot be adapted for high resolution gated cardiac imaging, necessary for CACS. The stationary DCT system (s- DCT), developed in our lab, has the potential to significantly shorten the scanning time and enables high resolution cardiac gated imaging. Here we report the preliminary results of using s-DCT to estimate the CACS. A phantom heart model was developed and scanned by the s-DCT system and a clinical CT in a phantom model with realistic coronary calcifications. The adapted fan-beam volume reconstruction (AFVR) method, developed specifically for stationary tomosynthesis systems, is used to obtain high resolution tomosynthesis images. A trained cardiologist segmented out the calcifications and the CACS was obtained. We observed a strong correlation between the tomosynthesis derived CACS and CT CACS (r2 = 0.88). Our results shows s-DCT imaging has the potential to estimate CACS, thus providing a possible low cost and low dose imaging protocol for screening and monitoring CAD.

The effect of sol-gel-formed calcium phosphate coatings on bone ingrowth and osteoconductivity of porous-surfaced Ti alloy implants.

PubMed

Nguyen, H Q; Deporter, D A; Pilliar, R M; Valiquette, N; Yakubovich, R

2004-02-01

Ti-6Al-4V implants formed with a sintered porous surface for implant fixation by bone ingrowth were prepared with or without the addition of a thin surface layer of calcium phosphate (Ca-P) formed using a sol-gel coating technique over the porous surface. The implants were placed transversely across the tibiae of 17 rabbits. Implanted sites were allowed to heal for 2 weeks, after which specimens were retrieved for morphometric assessment using backscattered scanning electron microscopy and quantitative image analysis. Bone formation along the porous-structured implant surface, was measured in relation to the medial and lateral cortices as an indication of implant surface osteoconductivity. The Absolute Contact Length measurements of endosteal bone growth along the porous-surfaced zone were greater with the Ca-P-coated implants compared to the non-Ca-P-coated implants. The Ca-P-coated implants also displayed a trend towards a significant increase in the area of bone ingrowth (Bone Ingrowth Fraction). Finally, there was significantly greater bone-to-implant contact within the sinter neck regions of the Ca-P-coated implants.

Central Nervous Activity upon Systemic Salicylate Application in Animals with Kanamycin-Induced Hearing Loss - A Manganese-Enhanced MRI (MEMRI) Study

PubMed Central

Gröschel, Moritz; Götze, Romy; Müller, Susanne; Ernst, Arne; Basta, Dietmar

2016-01-01

This study investigated the effect of systemic salicylate on central auditory and non-auditory structures in mice. Since cochlear hair cells are known to be one major target of salicylate, cochlear effects were reduced by using kanamycin to remove or impair hair cells. Neuronal brain activity was measured using the non-invasive manganese-enhanced magnetic resonance imaging technique. For all brain structures investigated, calcium-related neuronal activity was increased following systemic application of a sodium salicylate solution: probably due to neuronal hyperactivity. In addition, it was shown that the central effect of salicylate was not limited to the auditory system. A general alteration of calcium-related activity was indicated by an increase in manganese accumulation in the preoptic area of the anterior hypothalamus, as well as in the amygdala. The present data suggest that salicylate-induced activity changes in the auditory system differ from those shown in studies of noise trauma. Since salicylate action is reversible, central pharmacological effects of salicylate compared to those of (permanent) noise-induced hearing impairment and tinnitus might induce different pathophysiologies. These should therefore, be treated as different causes with the same symptoms. PMID:27078034

TEM preparation methods and influence of radiation damage on the beam sensitive CaCO3 shell of Emiliania huxleyi.

PubMed

Hoffmann, Ramona; Wochnik, Angela S; Betzler, Sophia B; Matich, Sonja; Griesshaber, Erika; Schmahl, Wolfgang W; Scheu, Christina

2014-07-01

The ultrastructure of biologically formed calcium carbonate crystals like the shell of Emiliania huxleyi depends on the environmental conditions such as pH value, temperature and salinity. Therefore, they can be used as indicator for climate changes. However, for this a detailed understanding of their crystal structure and chemical composition is required. High resolution methods like transmission electron microscopy can provide those information on the nanoscale, given that sufficiently thin samples can be prepared. In our study, we developed sample preparation techniques for cross-section and plan-view investigations and studied the sample stability under electron bombardment. In addition to the biological material (Emiliania huxleyi) we also prepared mineralogical samples (Iceland spar) for comparison. High resolution transmission electron microscopy imaging, electron diffraction and electron energy-loss spectroscopy studies revealed that all prepared samples are relatively stable under electron bombardment at an acceleration voltage of 300 kV when using a parallel illumination. Above an accumulated dose of ∼10(5) e/nm2 the material--independent whether its origin is biological or geological--transformed to poly-crystalline calcium oxide. Copyright © 2014 Elsevier Ltd. All rights reserved.

Structural changes in calcium-binding allergens: use of circular dichroism to study binding characteristics.

PubMed

Hebenstreit, D; Ferreira, F

2005-09-01

Several studies showed that calcium-binding proteins have a fixed place in the spectrum of allergenic substances. Often the binding of a calcium ion induces conformational changes and affects immunoglobulin E-binding to the allergen. Hence, the quantitative characterization of the binding to calcium is of importance to understand both the biologic and allergenic activity of these proteins. In the present study we describe a procedure for determining the stoichiometry and dissociation constant (K(D)) of calcium-binding allergens using circular dichroism (CD) techniques. For the experiments, we used recombinant Bet v 4, a two EF-hand allergen from birch pollen. Solutions of Bet v 4 were titrated with calcium and the change in molar ellipticity at 222 nm was monitored with a CD spectropolarimeter. The determination of the binding stoichiometry as well as of the K(D) for one EF-hand (4 microM) demonstrated the applicability of the method. CD-monitored calcium-titration of protein solutions represents a fast and easy method for determining the binding characteristics of calcium-binding allergens.

A novel biphasic calcium phosphate derived from fish otoliths

NASA Astrophysics Data System (ADS)

Montañez-Supelano, N. D.; Sandoval-Amador, A.; Estupiñan-Durán, H. A.; Y Peña-Ballesteros, D.

2017-12-01

Calcium phosphates are bioceramics that have been widely used as bone substitutes because they encourage the formation of bone on their surface and can improve the healing of the bone. Hydroxyapatite HA (calcium/phosphorus ratio of 1.67) and tricalcium phosphate TCP (calcium/phosphorus ratio of 1.50) are the most common calcium phosphates. Natural materials have begun to be tested to make HA or TCP such as shells of cardiidae (family of mollusks) and eggshells. The calcium phosphate obtained has a high ability to precipitate apatite. In this work, the mixed phase ceramic of beta-Tri-calcium phosphate / hydroxyapatite (β-TCP/HA) was synthesized by aqueous precipitation from fish otoliths, which are monomineralic species composed of aragonite. Otoliths of the specie Plagioscion squamosissimus, commonly called the river croaker, were used. Techniques such as DRX, Raman spectroscopy and SEM-EDS were used to characterize the raw material and the obtained material. X-ray diffraction analysis revealed the presence of two crystalline phases of calcium phosphates with 86.2% crystallinity. SEM micrographs showed agglomeration of particles with porous structure and submicron particle sizes.

Variation in band gap energy and electrical analysis of double doped cobalt ferrite

NASA Astrophysics Data System (ADS)

Parveen, Azra; Agrawal, Shraddha; Azam, Ameer

2018-05-01

The Ca and Cr doped cobalt ferrite nanoparticles (Co0.9Ca0.1) (Fe0.8 Cr0.2)2O4 were synthesized by microwave gel combustion method. Microstructural studies were carried out by XRD and SEM. Structural studies suggest that the crystal system remains spinal even with the doping of calcium and chromium. The SEM image shows the spherical morphology of surface of the sample. Optical properties of Ca and Cr doped cobalt ferrite were studied by UV-visible technique in the range of 400-600 nm. The electrical conductivity of pure and doped cobalt ferrite were studied as a function of frequency and were explained on the basis of electron hopping.

Fast functional imaging of multiple brain regions in intact zebrafish larvae using selective plane illumination microscopy.

PubMed

Panier, Thomas; Romano, Sebastián A; Olive, Raphaël; Pietri, Thomas; Sumbre, Germán; Candelier, Raphaël; Debrégeas, Georges

2013-01-01

The optical transparency and the small dimensions of zebrafish at the larval stage make it a vertebrate model of choice for brain-wide in-vivo functional imaging. However, current point-scanning imaging techniques, such as two-photon or confocal microscopy, impose a strong limit on acquisition speed which in turn sets the number of neurons that can be simultaneously recorded. At 5 Hz, this number is of the order of one thousand, i.e., approximately 1-2% of the brain. Here we demonstrate that this limitation can be greatly overcome by using Selective-plane Illumination Microscopy (SPIM). Zebrafish larvae expressing the genetically encoded calcium indicator GCaMP3 were illuminated with a scanned laser sheet and imaged with a camera whose optical axis was oriented orthogonally to the illumination plane. This optical sectioning approach was shown to permit functional imaging of a very large fraction of the brain volume of 5-9-day-old larvae with single- or near single-cell resolution. The spontaneous activity of up to 5,000 neurons was recorded at 20 Hz for 20-60 min. By rapidly scanning the specimen in the axial direction, the activity of 25,000 individual neurons from 5 different z-planes (approximately 30% of the entire brain) could be simultaneously monitored at 4 Hz. Compared to point-scanning techniques, this imaging strategy thus yields a ≃20-fold increase in data throughput (number of recorded neurons times acquisition rate) without compromising the signal-to-noise ratio (SNR). The extended field of view offered by the SPIM method allowed us to directly identify large scale ensembles of neurons, spanning several brain regions, that displayed correlated activity and were thus likely to participate in common neural processes. The benefits and limitations of SPIM for functional imaging in zebrafish as well as future developments are briefly discussed.

Optimization of the K-edge imaging for vulnerable plaques using gold nanoparticles and energy-resolved photon counting detectors: a simulation study

PubMed Central

Alivov, Yahya; Baturin, Pavlo; Le, Huy Q.; Ducote, Justin; Molloi, Sabee

2014-01-01

We investigated the effect of different imaging parameters such as dose, beam energy, energy resolution, and number of energy bins on image quality of K-edge spectral computed tomography (CT) of gold nanoparticles (GNP) accumulated in an atherosclerotic plaque. Maximum likelihood technique was employed to estimate the concentration of GNP, which served as a targeted intravenous contrast material intended to detect the degree of plaque's inflammation. The simulations studies used a single slice parallel beam CT geometry with an X-ray beam energy ranging between 50 and 140 kVp. The synthetic phantoms included small (3 cm in diameter) cylinder and chest (33x24 cm2) phantom, where both phantoms contained tissue, calcium, and gold. In the simulation studies GNP quantification and background (calcium and tissue) suppression task were pursued. The X-ray detection sensor was represented by an energy resolved photon counting detector (e.g., CdZnTe) with adjustable energy bins. Both ideal and more realistic (12% FWHM energy resolution) implementations of photon counting detector were simulated. The simulations were performed for the CdZnTe detector with pixel pitch of 0.5-1 mm, which corresponds to the performance without significant charge sharing and cross-talk effects. The Rose model was employed to estimate the minimum detectable concentration of GNPs. A figure of merit (FOM) was used to optimize the X-ray beam energy (kVp) to achieve the highest signal-to-noise ratio (SNR) with respect to patient dose. As a result, the successful identification of gold and background suppression was demonstrated. The highest FOM was observed at 125 kVp X-ray beam energy. The minimum detectable GNP concentration was determined to be approximately 1.06 μmol/mL (0.21 mg/mL) for an ideal detector and about 2.5 μmol/mL (0.49 mg/mL) for more realistic (12% FWHM) detector. The studies show the optimal imaging parameters at lowest patient dose using an energy resolved photon counting detector to image GNP in an atherosclerotic plaque. PMID:24334301

Preparation and properties of calcium oxide from eggshells via calcination

NASA Astrophysics Data System (ADS)

Tangboriboon, N.; Kunanuruksapong, R.; Sirivat, A.

2012-12-01

Duck eggs are one of the most versatile cooking ingredients in which residue eggshells are discarded. Raw duck eggshells were calcined at temperatures between 300 to 900 °C, for 1, 3, and 5 h. Both the raw and calcined duck eggshells were characterized by FTIR, STA, XRD, XRF, TEM, BET, a particle size analyzer, and an impedance analyzer. The proper calcination conditions are: 900 °C and 1 h, yielding calcium oxide with a purity of 99.06 % w/w. The calcium carbonate of the rhombohedral form (CaCO3) transforms completely into the calcium oxide or lime of the face centered cubic form (CaO) at 900 °C, as shown by XRD diffraction patterns. The transmission electron microscopy (TEM) images of the calcium oxide reveal a moderately good dispersion of nearly uniform particles. The calcium oxide has a white color, a spherical shape, high porosity, and narrow particles size distribution. The percentage of ceramic yield of the calcium oxide is 53.53, as measured by STA (TG-DTA-DTG). The calcium oxide has a N2 adsorption-desorption isotherm indicating the meso-porosity range. The dielectric constant and the electrical conductivity of the calcined calcium oxide are 35 and 1:0×10-6(Ω·m)-1, respectively, at the frequency of 500 Hz.

FISSA: A neuropil decontamination toolbox for calcium imaging signals.

PubMed

Keemink, Sander W; Lowe, Scott C; Pakan, Janelle M P; Dylda, Evelyn; van Rossum, Mark C W; Rochefort, Nathalie L

2018-02-22

In vivo calcium imaging has become a method of choice to image neuronal population activity throughout the nervous system. These experiments generate large sequences of images. Their analysis is computationally intensive and typically involves motion correction, image segmentation into regions of interest (ROIs), and extraction of fluorescence traces from each ROI. Out of focus fluorescence from surrounding neuropil and other cells can strongly contaminate the signal assigned to a given ROI. In this study, we introduce the FISSA toolbox (Fast Image Signal Separation Analysis) for neuropil decontamination. Given pre-defined ROIs, the FISSA toolbox automatically extracts the surrounding local neuropil and performs blind-source separation with non-negative matrix factorization. Using both simulated and in vivo data, we show that this toolbox performs similarly or better than existing published methods. FISSA requires only little RAM, and allows for fast processing of large datasets even on a standard laptop. The FISSA toolbox is available in Python, with an option for MATLAB format outputs, and can easily be integrated into existing workflows. It is available from Github and the standard Python repositories.

Crucibleless crystal growth and Radioluminescence study of calcium tungstate single crystal fiber

NASA Astrophysics Data System (ADS)

Silva, M. S.; Jesus, L. M.; Barbosa, L. B.; Ardila, D. R.; Andreeta, J. P.; Silva, R. S.

2014-11-01

In this article, single phase and high optical quality scheelite calcium tungstate single crystal fibers were grown by using the crucibleless laser heated pedestal growth technique. The as-synthesized calcium tungstate powders used for shaping seed and feed rods were investigated by X-ray diffraction technique. As-grown crystals were studied by Raman spectroscopy and Radioluminescence measurements. The results indicate that in both two cases, calcined powder and single crystal fiber, only the expected scheelite CaWO4 phase was observed. It was verified large homogeneity in the crystal composition, without the presence of secondary phases. The Radioluminescence spectra of the as-grown single crystal fibers are in agreement with that present in Literature for bulk single crystals, presented a single emission band centered at 420 nm when irradiated with β-rays.

Neuropathic Pain Causes Pyramidal Neuronal Hyperactivity in the Anterior Cingulate Cortex.

PubMed

Zhao, Ruohe; Zhou, Hang; Huang, Lianyan; Xie, Zhongcong; Wang, Jing; Gan, Wen-Biao; Yang, Guang

2018-01-01

The anterior cingulate cortex (ACC) is thought to be important for acute pain perception as well as the development of chronic pain after peripheral nerve injury. Nevertheless, how ACC neurons respond to sensory stimulation under chronic pain states is not well understood. Here, we used an in vivo two-photon imaging technique to monitor the activity of individual neurons in the ACC of awake, head restrained mice. Calcium imaging in the dorsal ACC revealed robust somatic activity in layer 5 (L5) pyramidal neurons in response to peripheral noxious stimuli, and the degree of evoked activity was correlated with the intensity of noxious stimulation. Furthermore, the activation of ACC neurons occurred bilaterally upon noxious stimulation to either contralateral or ipsilateral hind paws. Notably, with nerve injury-induced neuropathic pain in one limb, L5 pyramidal neurons in both sides of the ACC showed enhanced activity in the absence or presence of pain stimuli. These results reveal hyperactivity of L5 pyramidal neurons in the bilateral ACC during the development of neuropathic pain.

Optical manipulation for optogenetics: otoliths manipulation in zebrafish (Conference Presentation)

NASA Astrophysics Data System (ADS)

Favre-Bulle, Itia A.; Scott, Ethan; Rubinsztein-Dunlop, Halina

2016-03-01

Otoliths play an important role in Zebrafish in terms of hearing and sense of balance. Many studies have been conducted to understand its structure and function, however the encoding of its movement in the brain remains unknown. Here we developed a noninvasive system capable of manipulating the otolith using optical trapping while we image its behavioral response and brain activity. We'll also present our tools for behavioral response detection and brain activity mapping. Acceleration is sensed through movements of the otoliths in the inner ear. Because experimental manipulations involve movements, electrophysiology and fluorescence microscopy are difficult. As a result, the neural codes underlying acceleration sensation are poorly understood. We have developed a technique for optically trapping otoliths, allowing us to simulate acceleration in stationary larval zebrafish. By applying forces to the otoliths, we can elicit behavioral responses consistent with compensation for perceived acceleration. Since the animal is stationary, we can use calcium imaging in these animals' brains to identify the functional circuits responsible for mediating responses to acceleration in natural settings.

Mapping piezoelectric response in nanomaterials using a dedicated non-destructive scanning probe technique.

PubMed

Calahorra, Yonatan; Smith, Michael; Datta, Anuja; Benisty, Hadas; Kar-Narayan, Sohini

2017-12-14

There has been tremendous interest in piezoelectricity at the nanoscale, for example in nanowires and nanofibers where piezoelectric properties may be enhanced or controllably tuned, thus necessitating robust characterization techniques of piezoelectric response in nanomaterials. Piezo-response force microscopy (PFM) is a well-established scanning probe technique routinely used to image piezoelectric/ferroelectric domains in thin films, however, its applicability to nanoscale objects is limited due to the requirement for physical contact with an atomic force microscope (AFM) tip that may cause dislocation or damage, particularly to soft materials, during scanning. Here we report a non-destructive PFM (ND-PFM) technique wherein the tip is oscillated into "discontinuous" contact during scanning, while applying an AC bias between tip and sample and extracting the piezoelectric response for each contact point by monitoring the resulting localized deformation at the AC frequency. ND-PFM is successfully applied to soft polymeric (poly-l-lactic acid) nanowires, as well as hard ceramic (barium zirconate titanate-barium calcium titanate) nanowires, both previously inaccessible by conventional PFM. Our ND-PFM technique is versatile and compatible with commercial AFMs, and can be used to correlate piezoelectric properties of nanomaterials with their microstructural features thus overcoming key characterisation challenges in the field.

Analysis of the Chemical Modification of Dental Enamel Submitted to 35% Hydrogen Peroxide “In-Office” Whitening, with or without Calcium

PubMed Central

Santos, Fábio Pinheiro; Santos, Estevão Antero; dos Santos, Ramon Silva; dos Anjos, Marcelino José; de Miranda, Mauro Sayão

2017-01-01

Purpose The purpose of this study was to evaluate changes in calcium and phosphorus content in dental enamel when subjected to “in-office” whitening for an extended time by using a 35% hydrogen peroxide solution, with and without calcium. Materials and Methods 10 human teeth, from which the roots had been removed, were embedded in epoxy resin, and their surfaces were smoothed. The specimens were divided into two groups; in group 1, a whitening solution without calcium was used, while in group 2, the solution included calcium. Each specimen was evaluated at 6 different points before the bleaching treatment, and these points were reassessed after each session. A total of five sessions were carried out. Concentrations of calcium and phosphorus were measured by using the technique of X-ray fluorescence. Results After performing a statistical analysis, it was found that there was no statistically significant loss of calcium and phosphorus during the whitening treatment, and the groups showed no statistical differences. Conclusion Excessive use of hydrogen peroxide, with or without calcium, causes no loss of calcium and phosphorus. PMID:28932242

Electrophoretic mobility shift in native gels indicates calcium-dependent structural changes of neuronal calcium sensor proteins.

PubMed

Viviano, Jeffrey; Krishnan, Anuradha; Wu, Hao; Venkataraman, Venkat

2016-02-01

In proteins of the neuronal calcium sensor (NCS) family, changes in structure as well as function are brought about by the binding of calcium. In this article, we demonstrate that these structural changes, solely due to calcium binding, can be assessed through electrophoresis in native gels. The results demonstrate that the NCS proteins undergo ligand-dependent conformational changes that are detectable in native gels as a gradual decrease in mobility with increasing calcium but not other tested divalent cations such as magnesium, strontium, and barium. Surprisingly, such a gradual change over the entire tested range is exhibited only by the NCS proteins but not by other tested calcium-binding proteins such as calmodulin and S100B, indicating that the change in mobility may be linked to a unique NCS family feature--the calcium-myristoyl switch. Even within the NCS family, the changes in mobility are characteristic of the protein, indicating that the technique is sensitive to the individual features of the protein. Thus, electrophoretic mobility on native gels provides a simple and elegant method to investigate calcium (small ligand)-induced structural changes at least in the superfamily of NCS proteins. Copyright © 2015 Elsevier Inc. All rights reserved.

Mineral balance and bone turnover in adolescents with anorexia nervosa.

PubMed

Abrams, S A; Silber, T J; Esteban, N V; Vieira, N E; Stuff, J E; Meyers, R; Majd, M; Yergey, A L

1993-08-01

We evaluated seven female adolescents with anorexia nervosa to determine whether calcium metabolism was affected by their disorder. We measured calcium absorption, urinary calcium excretion, and calcium kinetics, using a dual-tracer, stable-isotope technique during the first weeks of an inpatient nutritional rehabilitation program. Results were compared with those from a control group of seven healthy adolescent girls of similar ages. The percentage of absorption of calcium was lower in subjects with anorexia nervosa than in control subjects (16.2% +/- 6.3% vs 24.6% +/- 7.2%; p < 0.05). Urinary calcium excretion was greater in subjects with anorexia nervosa than in control subjects (6.4 +/- 2.5 vs 1.6 +/- 0.7 mg.kg-1 x day-1; p < 0.01) and was associated with bone resorption rather than calcium hyper-absorption. Calcium kinetic studies demonstrated a decreased rate of bone formation and an increased rate of bone resorption. These results suggest marked abnormalities in mineral metabolism in patients with anorexia nervosa. From these results, we hypothesize that improvement in bone mineralization during recovery from anorexia nervosa will require resolution of hormonal abnormalities, including hypercortisolism, in addition to increased calcium intake.

Characterization of calcium and zinc spatial distributions at the fibrocartilage zone of bone-tendon junction by synchrotron radiation-based micro X-ray fluorescence analysis combined with backscattered electron imaging

NASA Astrophysics Data System (ADS)

Lu, Hongbin; Chen, Can; Wang, Zhanwen; Qu, Jin; Xu, Daqi; Wu, Tianding; Cao, Yong; Zhou, Jingyong; Zheng, Cheng; Hu, Jianzhong

2015-09-01

Tendon attaches to bone through a functionally graded fibrocartilage zone, including uncalcified fibrocartilage (UF), tidemark (TM) and calcified fibrocartilage (CF). This transition zone plays a pivotal role in relaxing load transfer between tendon and bone, and serves as a boundary between otherwise structurally and functionally distinct tissue types. Calcium and zinc are believed to play important roles in the normal growth, mineralization, and repair of the fibrocartilage zone of bone-tendon junction (BTJ). However, spatial distributions of calcium and zinc at the fibrocartilage zone of BTJ and their distribution-function relationship are not totally understood. Thus, synchrotron radiation-based micro X-ray fluorescence analysis (SR-μXRF) in combination with backscattered electron imaging (BEI) was employed to characterize the distributions of calcium and zinc at the fibrocartilage zone of rabbit patella-patellar tendon complex (PPTC). For the first time, the unique distributions of calcium and zinc at the fibrocartilage zone of the PPTC were clearly mapped by this method. The distributions of calcium and zinc at the fibrocartilage zone of the PPTC were inhomogeneous. A significant accumulation of zinc was exhibited in the transition region between UF and CF. The highest zinc content (3.17 times of that of patellar tendon) was found in the TM of fibrocartilage zone. The calcium content began to increase near the TM and increased exponentially across the calcified fibrocartilage region towards the patella. The highest calcium content (43.14 times of that of patellar tendon) was in the transitional zone of calcified fibrocartilage region and the patella, approximately 69 μm from the location with the highest zinc content. This study indicated, for the first time, that there is a differential distribution of calcium and zinc at the fibrocartilage zone of PPTC. These observations reveal new insights into region-dependent changes across the fibrocartilage zone of BTJ and will serve as critical benchmark parameters for current efforts in BTJ repair.

Does Increased Expression of the Plasma Membrane Calcium-ATPase Isoform 2 Confer Resistance to Apoptosis on Breast Cancer Cells?

DTIC Science & Technology

2008-09-01

T47D control cells with the highest sustained levels of intracellular calcium in the  live   cell   imaging  experiments (Figure 9). Membrane blebbing is a...classic hallmark, and early indicator of apoptosis. No  membrane blebbing was observed in T47D/PMCA2 cells during the  live   cell   imaging  studies.     10

Spatiotemporal dynamics of rhythmic spinal interneurons measured with two-photon calcium imaging and coherence analysis.

PubMed

Kwan, Alex C; Dietz, Shelby B; Zhong, Guisheng; Harris-Warrick, Ronald M; Webb, Watt W

2010-12-01

In rhythmic neural circuits, a neuron often fires action potentials with a constant phase to the rhythm, a timing relationship that can be functionally significant. To characterize these phase preferences in a large-scale, cell type-specific manner, we adapted multitaper coherence analysis for two-photon calcium imaging. Analysis of simulated data showed that coherence is a simple and robust measure of rhythmicity for calcium imaging data. When applied to the neonatal mouse hindlimb spinal locomotor network, the phase relationships between peak activity of >1,000 ventral spinal interneurons and motor output were characterized. Most interneurons showed rhythmic activity that was coherent and in phase with the ipsilateral motor output during fictive locomotion. The phase distributions of two genetically identified classes of interneurons were distinct from the ensemble population and from each other. There was no obvious spatial clustering of interneurons with similar phase preferences. Together, these results suggest that cell type, not neighboring neuron activity, is a better indicator of an interneuron's response during fictive locomotion. The ability to measure the phase preferences of many neurons with cell type and spatial information should be widely applicable for studying other rhythmic neural circuits.

Evidence of Multi-Component Ion Exchange in Dolomite Formation during Low Salinity Waterflooding

NASA Astrophysics Data System (ADS)

Srisuriyachai, Falan; Meekangwal, Suthida

2017-12-01

Low salinity waterflooding is a technique performed in many oil reservoirs around the globe. The technique is simply implemented by injecting water with very low ionic activity compared to formation water into an injection well. The injected water will increase reservoir pressure that is compulsory to drive oil moving toward production well. More than just maintaining reservoir pressure as obtained from conventional waterflooding, low salinity water creates shifting of surface condition, resulting in additional amount of liberated oil. Nevertheless, exact oil recovery mechanisms are still discussed. Among these proposed mechanisms, Multi-component Ion Exchange (MIE) together with wettability alteration is believed to be a major mechanism leading to higher oil recovery compared to conventional waterflooding. In this study, detection of calcium and magnesium ions which are Potential Determining Ions (PDI) for carbonate reservoirs are detected during the coreflood experiment. Dolomite rock sample is used to represent carbonate formation and detection of previously mentioned ions is performed by complexometric titration of the effluents. From the study, it is observed that during conventional waterflooding and low salinity waterflooding at low temperature of 30 degrees Celsius, calcium and magnesium ions in the produced water is increased compared to the amount of these ions in the injected water. This incremental of ions can be explained by the dissolution of calcium and magnesium from dolomite which is chemically composed of calcium magnesium carbonate. At this temperature, the portion of calcium ion is always less than magnesium ion even though the amount of calcium ion is higher than magnesium ion in injected water. However, at higher temperatures which are 50 and 70 degrees Celsius, ratio of calcium and magnesium ions in injected and produced water is reversed. Disappearance of magnesium ion in the effluent is more obvious especially at 70 degrees Celsius and by low salinity waterflooding. This can be explained that at lower temperature, calcium ion disappears to form of calcium carboxylate complex with oil and at higher temperature, magnesium ion disappears as magnesium can start to form magnesium carboxylate complex with oil and hence, the amount of both calcium and magnesium ions is decreased compared to lower temperature. In dolomite reservoir, since both calcium ions and magnesium ions are provided from dissolution mechanism, the benefit from multi-component ion exchange will occur at high temperature as both calcium and magnesium ions will be consumed for oil recovery mechanism.

Calcium-induced calcium release in rod photoreceptor terminals boosts synaptic transmission during maintained depolarization

PubMed Central

Cadetti, Lucia; Bryson, Eric J.; Ciccone, Cory A.; Rabl, Katalin; Thoreson, Wallace B.

2008-01-01

We examined the contribution of calcium-induced calcium release (CICR) to synaptic transmission from rod photoreceptor terminals. Whole-cell recording and confocal calcium imaging experiments were conducted on rods with intact synaptic terminals in a retinal slice preparation from salamander. Low concentrations of ryanodine stimulated calcium increases in rod terminals, consistent with the presence of ryanodine receptors. Application of strong depolarizing steps (−70 to −10 mV) exceeding 200 ms or longer in duration evoked a wave of calcium that spread across the synaptic terminals of voltage-clamped rods. This secondary calcium increase was blocked by high concentrations of ryanodine, indicating it was due to CICR. Ryanodine (50 μM) had no significant effect on rod calcium current (Ica) although it slightly diminished rod light-evoked voltage responses. Bath application of 50 μM ryanodine strongly inhibited light-evoked currents in horizontal cells. Whether applied extracellularly or delivered into the rod cell through the patch pipette, ryanodine (50 μM) also inhibited excitatory post-synaptic currents (EPSCs) evoked in horizontal cells by depolarizing steps applied to rods. Ryanodine caused a preferential reduction in the later portions of EPSCs evoked by depolarizing steps of 200 ms or longer. These results indicate that CICR enhances calcium increases in rod terminals evoked by sustained depolarization, which in turn acts to boost synaptic exocytosis from rods. PMID:16819987

Vacuum impregnation: a promising way for mineral fortification in potato porous matrix (potato chips).

PubMed

Joshi, Alka; Kar, A; Rudra, S G; Sagar, V R; Varghese, E; Lad, M; Khan, I; Singh, B

2016-12-01

Potato chips can be considered as an ideal carrier for targeted nutrient/s delivery as mostly consumed by the vulnerable group (children and teen agers). The present study was planned to fortifiy potato chips with calcium (Calcium lactate) and zinc (Zinc sulphate) using vacuum impregnation technique. At about 70-80 mm Hg vacuum pressure, maximum level of impregnation of both the minerals was achieved. Results showed that after optimization, calcium lactate at 4.81%, zinc sulphate at 0.72%, and vacuum of 33.53 mm Hg with restoration period of 19.52 min can fortify potato chips that can fulfil 10 and 21% need of calcium and zinc, respectively of targeted group (age 4-17 years). The present research work has shown that through this technique, fortification can be done in potato chips which are generally considered as a poor source of minerals. Further to make potato chips more fit to health conscious consumers, rather frying microwaving was done to develop mineral fortified low fat potato chips.

Deficits in memory and hippocampal long-term potentiation in mice with reduced calbindin D28K expression.

PubMed Central

Molinari, S; Battini, R; Ferrari, S; Pozzi, L; Killcross, A S; Robbins, T W; Jouvenceau, A; Billard, J M; Dutar, P; Lamour, Y; Baker, W A; Cox, H; Emson, P C

1996-01-01

The influx of calcium into the postsynaptic neuron is likely to be an important event in memory formation. Among the mechanisms that nerve cells may use to alter the time course or size of a spike of intracellular calcium are cytosolic calcium binding or "buffering" proteins. To consider the role in memory formation of one of these proteins, calbindin D28K, which is abundant in many neurons, including the CA1 pyramidal cells of the hippocampus, transgenic mice deficient in calbindin D28K have been created. These mice show selective impairments in spatial learning paradigms and fail to maintain long-term potentiation. These results suggest a role for calbindin D28K protein in temporally extending a neuronal calcium signal, allowing the activation of calcium-dependent intracellular signaling pathways underlying memory function. Images Fig. 1 PMID:8755597

Subcellular localization of calcium deposits in the noble crayfish Astacus astacus spermatophore: Implications for post-mating spermatophore hardening and spermatozoon maturation.

PubMed

Niksirat, Hamid; Kouba, Antonín

2016-04-01

The freshly ejaculated spermatophore of crayfish undergoes a hardening process during post-mating storage on the body surface of female. The ultrastructural distribution of calcium deposits were studied and compared in freshly ejaculated and post-mating noble crayfish spermatophores, using the oxalate-pyroantimonate technique, to determine possible roles of calcium in post-mating spermatophore hardening and spermatozoon maturation. Small particles of sparsely distributed calcium deposits were visible in the wall of freshly ejaculated spermatophore. Also, large amount of calcium deposits were visible in the membranes of the freshly ejaculated spermatozoon. Five minutes post-ejaculation, granules in the spermatophore wall appeared as porous formations with numerous electron lucent spaces. Calcium deposits were visible within the spaces and scattered in the spermatophore wall matrix, where smaller calcium deposits combined to form globular calcium deposits. Large numbers of the globular calcium deposits were visible in the wall of the post-mating spermatophore. Smaller calcium deposits were detected in the central area of post-mating spermatophore, which contains the sperm mass, and in the extracellular matrix and capsule. While the density of calcium deposits decreased in the post-mating spermatozoon membranes, numerous small calcium deposits appeared in the subacrosomal zone and nucleus. Substantial changes in calcium deposit distribution in the crayfish spermatophore during post-mating storage on the body of female may be involved in the processes of the spermatophore hardening and spermatozoon maturation. © 2016 Wiley Periodicals, Inc.

Calcium overloading in traumatic axonal injury by lateral head rotation: a morphological evidence in rat model.

PubMed

He, Xiao-Sheng; Xiang, Zhang; Zhou, Fei; Fu, Luo-An; Shuang, Wang

2004-05-01

The study investigated morphologically axonal calcium overloading and its relationship with axonal structural changes. Twelve SD rats were divided into an injury and a sham group. The rat model of traumatic axonal injury (TAI) by lateral head rotation was produced. The oxalate-pyroantimonate technique for calcium localization was used to process the rat's medulla oblongata tissues with thin sections observed electron-microscopically for axonal structure and calcium precipitates on it. The axonal damage in medulla oblongata appeared at 2 h post-injury, gradually became diffuse and severe, and continued to exist at 24 hours. At 2 hours, calcium precipitates were deposited on separated lamellae and axolemma, but were rarely distributed in the axoplasm. At 6 hours, calcium precipitates occurred on separated lamellae and axolemma in much higher density, but on axoplasm in extremely small amounts. Some axons, though lacking structural changes of the myelin sheath, sequestered plenty of calcium deposits on their swollen mitochondria. At 24 hours, damaged axons presented with much more severe lamellae separation and calcium deposits. Axonal calcium overloading developed in rat TAI model using lateral head rotation. This was significantly related to structural damage in the axons. These findings suggest the feasibility of using calcium antagonists in cope the management of human DAI in its very early stage.

Novel in vivo techniques to visualize kidney anatomy and function.

PubMed

Peti-Peterdi, János; Kidokoro, Kengo; Riquier-Brison, Anne

2015-07-01

Intravital imaging using multiphoton microscopy (MPM) has become an increasingly popular and widely used experimental technique in kidney research over the past few years. MPM allows deep optical sectioning of the intact, living kidney tissue with submicron resolution, which is unparalleled among intravital imaging approaches. MPM has solved a long-standing critical technical barrier in renal research to study several complex and inaccessible cell types and anatomical structures in vivo in their native environment. Comprehensive and quantitative kidney structure and function MPM studies helped our better understanding of the cellular and molecular mechanisms of the healthy and diseased kidney. This review summarizes recent in vivo MPM studies with a focus on the glomerulus and the filtration barrier, although select, glomerulus-related renal vascular and tubular functions are also mentioned. The latest applications of serial MPM of the same glomerulus in vivo, in the intact kidney over several days, during the progression of glomerular disease are discussed. This visual approach, in combination with genetically encoded fluorescent markers of cell lineage, has helped track the fate and function (e.g., cell calcium changes) of single podocytes during the development of glomerular pathologies, and provided visual proof for the highly dynamic, rather than static, nature of the glomerular environment. Future intravital imaging applications have the promise to further push the limits of optical microscopy, and to advance our understanding of the mechanisms of kidney injury. Also, MPM will help to study new mechanisms of tissue repair and regeneration, a cutting-edge area of kidney research.

Impact of calcium signaling during infection of Neisseria meningitidis to human brain microvascular endothelial cells.

PubMed

Asmat, Tauseef M; Tenenbaum, Tobias; Jonsson, Ann-Beth; Schwerk, Christian; Schroten, Horst

2014-01-01

The pili and outer membrane proteins of Neisseria meningitidis (meningococci) facilitate bacterial adhesion and invasion into host cells. In this context expression of meningococcal PilC1 protein has been reported to play a crucial role. Intracellular calcium mobilization has been implicated as an important signaling event during internalization of several bacterial pathogens. Here we employed time lapse calcium-imaging and demonstrated that PilC1 of meningococci triggered a significant increase in cytoplasmic calcium in human brain microvascular endothelial cells, whereas PilC1-deficient meningococci could not initiate this signaling process. The increase in cytosolic calcium in response to PilC1-expressing meningococci was due to efflux of calcium from host intracellular stores as demonstrated by using 2-APB, which inhibits the release of calcium from the endoplasmic reticulum. Moreover, pre-treatment of host cells with U73122 (phospholipase C inhibitor) abolished the cytosolic calcium increase caused by PilC1-expressing meningococci demonstrating that active phospholipase C (PLC) is required to induce calcium transients in host cells. Furthermore, the role of cytosolic calcium on meningococcal adherence and internalization was documented by gentamicin protection assay and double immunofluorescence (DIF) staining. Results indicated that chelation of intracellular calcium by using BAPTA-AM significantly impaired PilC1-mediated meningococcal adherence to and invasion into host endothelial cells. However, buffering of extracellular calcium by BAPTA or EGTA demonstrated no significant effect on meningococcal adherence to and invasion into host cells. Taken together, these results indicate that meningococci induce calcium release from intracellular stores of host endothelial cells via PilC1 and cytoplasmic calcium concentrations play a critical role during PilC1 mediated meningococcal adherence to and subsequent invasion into host endothelial cells.

The search for a hippocampal engram.

PubMed

Mayford, Mark

2014-01-05

Understanding the molecular and cellular changes that underlie memory, the engram, requires the identification, isolation and manipulation of the neurons involved. This presents a major difficulty for complex forms of memory, for example hippocampus-dependent declarative memory, where the participating neurons are likely to be sparse, anatomically distributed and unique to each individual brain and learning event. In this paper, I discuss several new approaches to this problem. In vivo calcium imaging techniques provide a means of assessing the activity patterns of large numbers of neurons over long periods of time with precise anatomical identification. This provides important insight into how the brain represents complex information and how this is altered with learning. The development of techniques for the genetic modification of neural ensembles based on their natural, sensory-evoked, activity along with optogenetics allows direct tests of the coding function of these ensembles. These approaches provide a new methodological framework in which to examine the mechanisms of complex forms of learning at the level of the neurons involved in a specific memory.

The search for a hippocampal engram

PubMed Central

Mayford, Mark

2014-01-01

Understanding the molecular and cellular changes that underlie memory, the engram, requires the identification, isolation and manipulation of the neurons involved. This presents a major difficulty for complex forms of memory, for example hippocampus-dependent declarative memory, where the participating neurons are likely to be sparse, anatomically distributed and unique to each individual brain and learning event. In this paper, I discuss several new approaches to this problem. In vivo calcium imaging techniques provide a means of assessing the activity patterns of large numbers of neurons over long periods of time with precise anatomical identification. This provides important insight into how the brain represents complex information and how this is altered with learning. The development of techniques for the genetic modification of neural ensembles based on their natural, sensory-evoked, activity along with optogenetics allows direct tests of the coding function of these ensembles. These approaches provide a new methodological framework in which to examine the mechanisms of complex forms of learning at the level of the neurons involved in a specific memory. PMID:24298162

Synthesis of brushite particles in reverse microemulsions of the biosurfactant surfactin.

PubMed

Maity, Jyoti Prakash; Lin, Tz-Jiun; Cheng, Henry Pai-Heng; Chen, Chien-Yen; Reddy, A Satyanarayana; Atla, Shashi B; Chang, Young-Fo; Chen, Hau-Ren; Chen, Chien-Cheng

2011-01-01

In this study the "green chemistry" use of the biosurfactant surfactin for the synthesis of calcium phosphate using the reverse microemulsion technique was demonstrated. Calcium phosphates are bioactive materials that are a major constituent of human teeth and bone tissue. A reverse microemulsion technique with surfactin was used to produce nanocrystalline brushite particles. Structural diversity (analyzed by SEM and TEM) resulted from different water to surfactin ratios (W/S; 250, 500, 1000 and 40,000). The particle sizes were found to be in the 16-200 nm range. Morphological variety was observed in the as-synthesized microemulsions, which consisted of nanospheres (~16 nm in diameter) and needle-like (8-14 nm in diameter and 80-100 nm in length) noncalcinated particles. However, the calcinated products included nanospheres (50-200 nm in diameter), oval (~300 nm in diameter) and nanorod (200-400 nm in length) particles. FTIR and XRD analysis confirmed the formation of brushite nanoparticles in the as-synthesized products, while calcium pyrophosphate was produced after calcination. These results indicate that the reverse microemulsion technique using surfactin is a green process suitable for the synthesis of nanoparticles.

Multi-scale approaches for high-speed imaging and analysis of large neural populations

PubMed Central

Ahrens, Misha B.; Yuste, Rafael; Peterka, Darcy S.; Paninski, Liam

2017-01-01

Progress in modern neuroscience critically depends on our ability to observe the activity of large neuronal populations with cellular spatial and high temporal resolution. However, two bottlenecks constrain efforts towards fast imaging of large populations. First, the resulting large video data is challenging to analyze. Second, there is an explicit tradeoff between imaging speed, signal-to-noise, and field of view: with current recording technology we cannot image very large neuronal populations with simultaneously high spatial and temporal resolution. Here we describe multi-scale approaches for alleviating both of these bottlenecks. First, we show that spatial and temporal decimation techniques based on simple local averaging provide order-of-magnitude speedups in spatiotemporally demixing calcium video data into estimates of single-cell neural activity. Second, once the shapes of individual neurons have been identified at fine scale (e.g., after an initial phase of conventional imaging with standard temporal and spatial resolution), we find that the spatial/temporal resolution tradeoff shifts dramatically: after demixing we can accurately recover denoised fluorescence traces and deconvolved neural activity of each individual neuron from coarse scale data that has been spatially decimated by an order of magnitude. This offers a cheap method for compressing this large video data, and also implies that it is possible to either speed up imaging significantly, or to “zoom out” by a corresponding factor to image order-of-magnitude larger neuronal populations with minimal loss in accuracy or temporal resolution. PMID:28771570

Na+/H+ exchanger 3 inhibitor diminishes hepcidin-enhanced duodenal calcium transport in hemizygous β-globin knockout thalassemic mice.

PubMed

Charoenphandhu, Narattaphol; Kraidith, Kamonshanok; Lertsuwan, Kornkamon; Sripong, Chanakarn; Suntornsaratoon, Panan; Svasti, Saovaros; Krishnamra, Nateetip; Wongdee, Kannikar

2017-03-01

Recent investigation has shown that the liver-derived iron-regulating hormone, hepcidin, can potentiate intestinal calcium absorption in hemizygous β-globin knockout thalassemic (BKO) mice. Since the upregulation of Fe 2+ and H + cotransporter, divalent metal transporter (DMT)-1, has been shown to correlate with thalassemia-induced intestinal calcium absorption impairment, the inhibition of the apical Na + /H + exchanger (NHE)-3 that is essential for cytoplasmic pH regulation and transepithelial sodium absorption was hypothesized to negatively affect hepcidin action. Herein, the positive effect of hepcidin on the duodenal calcium transport was evaluated using Ussing chamber technique. The results showed that BKO mice had lower absorptive surface area and duodenal calcium transport than wild-type mice. Besides, paracellular transport of zinc in BKO mice was compromised. Hepcidin administration completely restored calcium transport. Since this hepcidin action was totally abolished by inhibitors of the basolateral calcium transporters, Na + /Ca 2+ exchanger (NCX1) and plasma membrane Ca 2+ -ATPase (PMCA 1b ), the enhanced calcium flux potentially occurred through the transcellular pathway rather than paracellular pathway. Interestingly, the selective NHE3 inhibitor, 100 nM tenapanor, markedly inhibited hepcidin-enhanced calcium transport. Accordingly, hepcidin is one of the promising therapeutic agents for calcium malabsorption in β-thalassemia. It mainly stimulates the transcellular calcium transport across the duodenal epithelium in an NHE3-dependent manner.

Parallel temperature dependence of contracture-associated enzyme release due to anoxia, 2,4-dinitrophenol (DNP), or caffeine and the calcium paradox.

PubMed Central

Ganote, C. E.; Sims, M. A.

1984-01-01

Hypothermia during calcium-free perfusion of hearts protects them from injury caused by subsequent calcium repletion at 37 C (calcium paradox). Injury to calcium-free hearts is also associated with contracture caused by anoxia, 2,4-dinitrophenol (DNP), or caffeine. This study was done for the purpose of determining whether hypothermia during calcium-free perfusions protects hearts from contracture-associated injury. Langendorff-perfused rat hearts were studied in four experimental groups: I) Anoxia: Thirty minutes of anoxic perfusion at 37 C was followed by thirty minutes of anoxic calcium-free perfusion at 37-18 C. II) Calcium paradox: Five minutes of calcium-free perfusion at 37-18 C was followed by calcium repletion at 37 C. III, IVa) Caffeine or DNP: Five minutes of calcium-free perfusion at 37-18 C was followed by addition of 10 mM caffeine or 1 mM DNP in calcium-free medium at 37 C or, IVb) 1 mM DNP in calcium-free medium at 22 C. Injury was assessed by measurement of serial releases of creatine kinase (CK) in effluents and by cellular morphology. The results show that progressive hypothermia to 22 C during calcium-free perfusion periods produced a progressive reduction of CK release and morphologic evidence of injury due to anoxia, caffeine, or DNP, which closely paralleled protection of hearts from the calcium paradox. Protection from injury in all experimental groups was associated with preservation of sarcolemmal membrane integrity and prevention of cell separations at intercalated disk junctions. It is proposed that weakening of intercalated disks occurs during calcium-free perfusions and may be a cause of mechanical fragility of the sarcolemma. Hypothermia may protect hearts from contracture-associated injury by preserving the integrity of intercalated disk junctions during periods of extracellular calcium depletion. Images Figure 4 Figure 5 Figure 6 Figure 7 Figure 8 Figure 9 Figure 10 Figure 11 PMID:6742111

Enteroaggregative Escherichia coli strains secrete a heat-labile toxin antigenically related to E. coli hemolysin.

PubMed Central

Baldwin, T J; Knutton, S; Sellers, L; Hernandez, H A; Aitken, A; Williams, P H

1992-01-01

A protein toxin of approximately 120,000 Da secreted by nonhemolytic enteroaggregative Escherichia coli strains cross-reacted in Western blots (immunoblots) with antibodies raised against the C-terminal region of E. coli hemolysin. Treatment of HEp-2 cells with enteroaggregative E. coli or culture supernatants caused elevation of intracellular calcium and stimulated calcium-dependent protein phosphorylation. Images PMID:1563799

Real-time In Vivo Recording of Arabidopsis Calcium Signals During Insect Feeding Using a Fluorescent Biosensor

PubMed Central

Vincent, Thomas R.; Canham, James; Toyota, Masatsugu; Avramova, Marieta; Mugford, Sam T.; Gilroy, Simon; Miller, Anthony J.; Hogenhout, Saskia; Sanders, Dale

2017-01-01

Calcium ions are predicted to be key signaling entities during biotic interactions, with calcium signaling forming an established part of the plant defense response to microbial elicitors and to wounding caused by chewing insects, eliciting systemic calcium signals in plants. However, the role of calcium in vivo during biotic stress is still unclear. This protocol describes the use of a genetically-encoded calcium sensor to detect calcium signals in plants during feeding by a hemipteran pest. Hemipterans such as aphids pierce a small number of cells with specialized, elongated sucking mouthparts, making them the ideal tool to study calcium dynamics when a plant is faced with a biotic stress, which is distinct from a wounding response. In addition, fluorescent biosensors are revolutionizing the measurement of signaling molecules in vivo in both animals and plants. Expressing a GFP-based calcium biosensor, GCaMP3, in the model plant Arabidopsis thaliana allows for the real-time imaging of plant calcium dynamics during insect feeding, with a high spatial and temporal resolution. A repeatable and robust assay has been developed using the fluorescence microscopy of detached GCaMP3 leaves, allowing for the continuous measurement of cytosolic calcium dynamics before, during, and after insect feeding. This reveals a highly-localized rapid calcium elevation around the aphid feeding site that occurs within a few minutes. The protocol can be adapted to other biotic stresses, such as additional insect species, while the use of Arabidopsis thaliana allows for the rapid generation of mutants to facilitate the molecular analysis of the phenomenon. PMID:28829425

An automated multi-modal object analysis approach to coronary calcium scoring of adaptive heart isolated MSCT images

NASA Astrophysics Data System (ADS)

Wu, Jing; Ferns, Gordon; Giles, John; Lewis, Emma

2012-02-01

Inter- and intra- observer variability is a problem often faced when an expert or observer is tasked with assessing the severity of a disease. This issue is keenly felt in coronary calcium scoring of patients suffering from atherosclerosis where in clinical practice, the observer must identify firstly the presence, followed by the location of candidate calcified plaques found within the coronary arteries that may prevent oxygenated blood flow to the heart muscle. This can be challenging for a human observer as it is difficult to differentiate calcified plaques that are located in the coronary arteries from those found in surrounding anatomy such as the mitral valve or pericardium. The inclusion or exclusion of false positive or true positive calcified plaques respectively will alter the patient calcium score incorrectly, thus leading to the possibility of incorrect treatment prescription. In addition to the benefits to scoring accuracy, the use of fast, low dose multi-slice CT imaging to perform the cardiac scan is capable of acquiring the entire heart within a single breath hold. Thus exposing the patient to lower radiation dose, which for a progressive disease such as atherosclerosis where multiple scans may be required, is beneficial to their health. Presented here is a fully automated method for calcium scoring using both the traditional Agatston method, as well as the Volume scoring method. Elimination of the unwanted regions of the cardiac image slices such as lungs, ribs, and vertebrae is carried out using adaptive heart isolation. Such regions cannot contain calcified plaques but can be of a similar intensity and their removal will aid detection. Removal of both the ascending and descending aortas, as they contain clinical insignificant plaques, is necessary before the final calcium scores are calculated and examined against ground truth scores of three averaged expert observer results. The results presented here are intended to show the requirement and feasibility for an automated scoring method that reduces the subjectivity and reproducibility error inherent with manual clinical calcium scoring.

Italian Chapter of the International Society of Cardiovascular Ultrasound expert consensus document on coronary computed tomography angiography: overview and new insights.

PubMed

Sozzi, Fabiola B; Maiello, Maria; Pelliccia, Francesco; Parato, Vito Maurizio; Canetta, Ciro; Savino, Ketty; Lombardi, Federico; Palmiero, Pasquale

2016-09-01

Coronary computed tomography angiography is a noninvasive heart imaging test currently undergoing rapid development and advancement. The high resolution of the three-dimensional pictures of the moving heart and great vessels is performed during a coronary computed tomography to identify coronary artery disease and classify patient risk for atherosclerotic cardiovascular disease. The technique provides useful information about the coronary tree and atherosclerotic plaques beyond simple luminal narrowing and plaque type defined by calcium content. This application will improve image-guided prevention, medical therapy, and coronary interventions. The ability to interpret coronary computed tomography images is of utmost importance as we develop personalized medical care to enable therapeutic interventions stratified on the bases of plaque characteristics. This overview provides available data and expert's recommendations in the utilization of coronary computed tomography findings. We focus on the use of coronary computed tomography to detect coronary artery disease and stratify patients at risk, illustrating the implications of this test on patient management. We describe its diagnostic power in identifying patients at higher risk to develop acute coronary syndrome and its prognostic significance. Finally, we highlight the features of the vulnerable plaques imaged by coronary computed tomography angiography. © 2016, Wiley Periodicals, Inc.

Numerical Simulation of Permeability Change in Wellbore Cement Fractures after Geomechanical Stress and Geochemical Reactions Using X-ray Computed Tomography Imaging.

PubMed

Kabilan, Senthil; Jung, Hun Bok; Kuprat, Andrew P; Beck, Anthon N; Varga, Tamas; Fernandez, Carlos A; Um, Wooyong

2016-06-21

X-ray microtomography (XMT) imaging combined with three-dimensional (3D) computational fluid dynamics (CFD) modeling technique was used to study the effect of geochemical and geomechanical processes on fracture permeability in composite Portland cement-basalt caprock core samples. The effect of fluid density and viscosity and two different pressure gradient conditions on fracture permeability was numerically studied by using fluids with varying density and viscosity and simulating two different pressure gradient conditions. After the application of geomechanical stress but before CO2-reaction, CFD revealed fluid flow increase, which resulted in increased fracture permeability. After CO2-reaction, XMT images displayed preferential precipitation of calcium carbonate within the fractures in the cement matrix and less precipitation in fractures located at the cement-basalt interface. CFD estimated changes in flow profile and differences in absolute values of flow velocity due to different pressure gradients. CFD was able to highlight the profound effect of fluid viscosity on velocity profile and fracture permeability. This study demonstrates the applicability of XMT imaging and CFD as powerful tools for characterizing the hydraulic properties of fractures in a number of applications like geologic carbon sequestration and storage, hydraulic fracturing for shale gas production, and enhanced geothermal systems.

Numerical Simulation of Permeability Change in Wellbore Cement Fractures after Geomechanical Stress and Geochemical Reactions Using X-ray Computed Tomography Imaging

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kabilan, Senthil; Jung, Hun Bok; Kuprat, Andrew P.

X-ray microtomography (XMT) imaging combined with a three-dimensional (3D) computational fluid dynamics (CFD) modeling technique was used to study the effect of geochemical and geomechanical processes on fracture properties in composite Portland cement–basalt caprock core samples. The effect of fluid properties and flow conditions on fracture permeability was numerically studied by using fluids with varying physical properties and simulating different pressure conditions. CFD revealed that the application of geomechanical stress led to increased fluid flow, which resulted in increased fracture permeability. After CO2-reaction, XMT images displayed preferential precipitation of calcium carbonate within the fractures in the cement matrix and lessmore » precipitation in fractures located at the cement–basalt interface. CFD predicted changes in flow characteristics and differences in absolute values of flow properties due to different pressure gradients. CFD was able to highlight the profound effect of fluid properties on flow characteristics and hydraulic properties of fractures. This study demonstrates the applicability of XMT imaging and CFD as powerful tools for characterizing the hydraulic properties of fractures in a number of applications like geologic carbon sequestration and storage, hydraulic fracturing for shale gas production, and enhanced geothermal systems.« less

Comparison of the Pullout Strength of Different Pedicle Screw Designs and Augmentation Techniques in an Osteoporotic Bone Model.

PubMed

Kiyak, Gorkem; Balikci, Tevfik; Heydar, Ahmed Majid; Bezer, Murat

2018-02-01

Mechanical study. To compare the pullout strength of different screw designs and augmentation techniques in an osteoporotic bone model. Adequate bone screw pullout strength is a common problem among osteoporotic patients. Various screw designs and augmentation techniques have been developed to improve the biomechanical characteristics of the bone-screw interface. Polyurethane blocks were used to mimic human osteoporotic cancellous bone, and six different screw designs were tested. Five standard and expandable screws without augmentation, eight expandable screws with polymethylmethacrylate (PMMA) or calcium phosphate augmentation, and distal cannulated screws with PMMA and calcium phosphate augmentation were tested. Mechanical tests were performed on 10 unused new screws of each group. Screws with or without augmentation were inserted in a block that was held in a fixture frame, and a longitudinal extraction force was applied to the screw head at a loading rate of 5 mm/min. Maximum load was recorded in a load displacement curve. The peak pullout force of all tested screws with or without augmentation was significantly greater than that of the standard pedicle screw. The greatest pullout force was observed with 40-mm expandable pedicle screws with four fins and PMMA augmentation. Augmented distal cannulated screws did not have a greater peak pullout force than nonaugmented expandable screws. PMMA augmentation provided a greater peak pullout force than calcium phosphate augmentation. Expandable pedicle screws had greater peak pullout forces than standard pedicle screws and had the advantage of augmentation with either PMMA or calcium phosphate cement. Although calcium phosphate cement is biodegradable, osteoconductive, and nonexothermic, PMMA provided a significantly greater peak pullout force. PMMA-augmented expandable 40-mm four-fin pedicle screws had the greatest peak pullout force.

STUDYING THE EFFECTS OF CALCIUM AND MAGNESIUM ON SIZE-DISTRIBUTED NITRATE AND AMMONIUM WITH EQUISOLV II. (R823186)

EPA Science Inventory

Abstract

A chemical equilibrium code was improved and used to show that calcium and magnesium have a large yet different effect on the aerosol size distribution in different regions of Los Angeles. In the code, a new technique of solving individual equilibrium equation...

Kinetic aspects of bone mineral metabolism

NASA Technical Reports Server (NTRS)

Palmer, H. E.

1973-01-01

Two techniques were studied for measuring changes in bone mass in rats. One technique measures the Ar-37 produced from calcium during neutron irradiation and the other measures the changes in the Na-22 content which has been incorporated within the rat bone. Both methods are performed in VIVO and cause no significant physiological damage. The Ar-37 leaves the body of a rat within an hour after being produced, and it can be quantitatively collected and measured with a precision of - or + 2% on the same rat. With appropriate irradiation conditions it appears that the absolute quantity of calcuim in any rat can be determined within - or + 3% regardless of animal size. The Na-22 when uniformly distributed in bone, can be used to monitor bone mineral turnover and this has been demonstrated in conditions of calcium deficiency during growth and also pregnancy coupled with calcium deficiency.

Hierarchically porous structure, mechanical strength and cell biological behaviors of calcium phosphate composite scaffolds prepared by combination of extrusion and porogen burnout technique and enhanced by gelatin.

PubMed

Feng, Shenglei; He, Fupo; Ye, Jiandong

2018-01-01

In this study, hierarchically porous calcium phosphate scaffolds (HTCP) with unidirectional pores, transversely interconnected pores, and micropores were fabricated by the combination of extrusion and porogen burnout technique. Gelatin was incorporated into the HTCP scaffolds by vacuum-impregnation of gelatin solution and subsequent freeze-drying. The phase composition, microstructure, physical and cytobiological properties were analyzed. The results showed that the HTCP scaffolds were composed of β-tricalcium phosphate with minor hydroxyapatite. The HTCP scaffolds had unidirectional pores (~400μm), transversely interconnected pores (~130μm) and micropores (~1μm). The incorporation of gelatin significantly increased the compressive strength, toughness, and cell seeding of the HTCP scaffolds. The composite scaffolds showed excellent cytocompatibility. The hierarchically porous calcium phosphate composite scaffolds may have potential application prospects in bone tissue engineering. Copyright © 2017 Elsevier B.V. All rights reserved.

Calcium/Vitamin D Supplementation and Coronary Artery Calcification

PubMed Central

Manson, JoAnn E.; Allison, Matthew A.; Carr, J. Jeffrey; Langer, Robert D.; Cochrane, Barbara B.; Hendrix, Susan L.; Hsia, Judith; Hunt, Julie R.; Lewis, Cora E.; Margolis, Karen L.; Robinson, Jennifer G.; Rodabough, Rebecca J.; Thomas, Asha M.

2010-01-01

Objectives Coronary artery calcified plaque is a marker for atheromatous plaque burden and predicts future risk of cardiovascular events. The relationship between calcium plus vitamin D supplementation and coronary artery calcium (CAC) has not been previously assessed in a randomized trial setting. We compared coronary artery calcium scores among women randomized to calcium/vitamin D supplementation versus placebo following trial completion. Methods In an ancillary substudy of women randomized to calcium carbonate (1000 mg of elemental calcium daily) plus vitamin D3 (400 IU daily) versus placebo, nested within the Women’s Health Initiative trial of estrogen among women with hysterectomy, we measured CAC with cardiac computed tomography in 754 women aged 50–59 years at randomization. Imaging for CAC was performed at 28 of 40 centers following a mean of 7 years of treatment and scans were read centrally. Coronary artery calcium scores were measured by a central reading center with masking to randomization assignments. Results Post-trial CAC measurements were similar in women randomized to calcium/vitamin D supplementation (calcium/D) and those receiving placebo. The mean CAC score was 91.6 for calcium/D and 100.5 for placebo (rank test p-value=0.74). After adjustment for coronary risk factors, multivariate odds ratios for increasing CAC score cutpoints (CAC >0, ≥10, and ≥100) for calcium/D vs placebo were 0.92 (95% confidence interval, 0.64–1.34), 1.29 (0.88–1.87), and 0.90 (0.56–1.44), respectively. Corresponding odds ratios among women with >50% adherence to study pills and for higher levels of CAC (>300), were similar. Conclusions Treatment with moderate doses of calcium plus vitamin D3 did not appear to alter coronary artery calcified plaque burden among postmenopausal women. PMID:20551849

Hydrostatic Pressure–Induced Release of Stored Calcium in Cultured Rat Optic Nerve Head Astrocytes

PubMed Central

Mandal, Amritlal; Delamere, Nicholas A.

2010-01-01

Purpose. Elevated intraocular pressure is associated with glaucomatous optic nerve damage. Other investigators have shown functional changes in optic nerve head astrocytes subjected to elevated hydrostatic pressure (HP) for 1 to 5 days. Recently, the authors reported ERK1/2, p90RSK and NHE1 phosphorylation after 2 hours. Here they examine calcium responses at the onset of HP to determine what precedes ERK1/2 phosphorylation. Methods. Cytoplasmic calcium concentration ([Ca2+]i) was measured in cultured rat optic nerve astrocytes loaded with fura-2. The cells were placed in a closed imaging chamber and subjected to an HP increase of 15 mm Hg. Protein phosphorylation was detected by Western blot analysis. Results. The increase of HP caused an immediate slow increase in [Ca2+]i. The response persisted in calcium-free solution and when nickel chloride (4 mM) was added to suppress channel-mediated calcium entry. Previous depletion of the ER calcium stores by cyclopiazonic acid abolished the HP-induced calcium level increase. The HP-induced increase persisted in cells exposed to xestospongin C, an inhibitor of IP3R-mediated calcium release. In contrast, ryanodine receptor (RyR) antagonist ruthenium red (10 μM) or dantrolene (25 μM) inhibited the HP-induced calcium increase. The HP-induced calcium increase was abolished when ryanodine-sensitive calcium stores were pre-depleted with caffeine (3 mM). HP caused ERK1/2 phosphorylation. The magnitude of the ERK1/2 phosphorylation response was reduced by ruthenium red and dantrolene. Conclusions. Increasing HP causes calcium release from a ryanodine-sensitive cytoplasmic store and subsequent ERK1/2 activation. Calcium store release appears to be a required early step in the initial astrocyte response to an HP increase. PMID:20071675

Electrodeposition on nanofibrous polymer scaffolds: Rapid mineralization, tunable calcium phosphate composition and topography

PubMed Central

He, Chuanglong; Xiao, Guiyong; Jin, Xiaobing; Sun, Chenghui; Ma, Peter X.

2011-01-01

We developed a straightforward, fast, and versatile technique to fabricate mineralized nanofibrous polymer scaffolds for bone regeneration in this work. Nanofibrous poly(l-lactic acid) scaffolds were fabricated using both electrospinning and phase separation techniques. An electrodeposition process was designed to deposit calcium phosphate on the nanofibrous scaffolds. Such scaffolds contain a high quality mineral coating on the fiber surface with tunable surface topography and chemical composition by varying the processing parameters, which can mimic the composition and structure of natural bone extracellular matrix and provide a more biocompatible interface for bone regeneration. PMID:21673827

Evaluation of the influence of acquisition and reconstruction parameters for 16-row multidetector CT on coronary calcium scoring using a stationary and dynamic cardiac phantom.

PubMed

Begemann, Philipp G C; van Stevendaal, Udo; Koester, Ralph; Mahnken, Andreas H; Koops, Andreas; Adam, Gerhard; Grass, Michael; Nolte-Ernsting, Claus

2007-08-01

A calcium-scoring phantom with hydroxyapatite-filled cylindrical holes (0.5 to 4 mm) was used. High-resolution scans were performed for an accuracy baseline. The phantom was mounted to a moving heart phantom. Non-moving data with the implementation of an ECG-signal were acquired for different pitches (0.2/0.3), heart rates (60/80/95 bpm) and collimations (16 x 0.75/16 x 1.5 mm). Images were reconstructed with a cone-beam multi-cycle algorithm at a standard thickness/increment of 3 mm/1.5 mm and the thinnest possible thickness (0.8/0.4 and 2/1). Subsequently, ECG-gated moving calcium-scoring phantom data were acquired. The calcium volume and Agatston score were measured. The temporal resolution and reconstruction cycles were calculated. High-resolution scans determine the calcium volume with a high accuracy (mean overestimation, 0.8%). In the non-moving measurements, the volume underestimation ranged from about 6% (16 x 0.75 mm; 0.8/0.4 mm) to nearly 25% (16 x 1.5 mm; 3/1.5 mm). Moving scans showed increased measurement errors depending on the reconstructed RR interval, collimation, pitch, heart rate and gantry rotation time. Also, a correlation with the temporal resolution could be found. The reliability of calcium-scoring results can be improved with the use of a narrower collimation, a lower pitch and the reconstruction of thinner images, resulting in higher patient doses. The choice of the correct cardiac phase within the RR interval is essential to minimize measurement errors.

Cortical Circuit Activity Evokes Rapid Astrocyte Calcium Signals on a Similar Timescale to Neurons.

PubMed

Stobart, Jillian L; Ferrari, Kim David; Barrett, Matthew J P; Glück, Chaim; Stobart, Michael J; Zuend, Marc; Weber, Bruno

2018-05-16

Sensory stimulation evokes intracellular calcium signals in astrocytes; however, the timing of these signals is disputed. Here, we used novel combinations of genetically encoded calcium indicators for concurrent two-photon imaging of cortical astrocytes and neurons in awake mice during whisker deflection. We identified calcium responses in both astrocyte processes and endfeet that rapidly followed neuronal events (∼120 ms after). These fast astrocyte responses were largely independent of IP 3 R2-mediated signaling and known neuromodulator activity (acetylcholine, serotonin, and norepinephrine), suggesting that they are evoked by local synaptic activity. The existence of such rapid signals implies that astrocytes are fast enough to play a role in synaptic modulation and neurovascular coupling. VIDEO ABSTRACT. Copyright © 2018 Elsevier Inc. All rights reserved.

Synthesis and characterization of bioresorbable calcium phosphosilicate nanocomposite particles for fluorescence imaging and biomedical applications

NASA Astrophysics Data System (ADS)

Morgan, Thomas T.

Organically doped calcium phosphosilicate nanoparticles (CPSNPs) were developed and characterized, driven by the need for non-toxic vectors for drug delivery and fluorescence biological imaging applications. In particular, advancement in drug delivery for the chemotherapeutic treatment of cancers is required to increase drug efficacy and improve patient quality of life. Additionally, brighter and more photostable fluorophores are needed to meet demands for improved sensitivity and experimental diversity, which may lead to improvements in early detection of solid tumors and advancement in understanding of biological processes. A literature survey on the state of the field for nanoparticle based biological fluorescence imaging and drug delivery is presented in Chapter 1. Chapter 2 focuses on the characterization techniques used in this work. The development and optical characterization of 20-40 nm diameter, citrate functionalized Cy3 amidite doped calcium phosphosilicate nanoparticles (Cy3 CPSNPs) for in vitro fluorescence imaging is outlined in Chapters 3 and 4, respectively. In particular, sodium citrate was used to functionalize the surface and provide electrosteric dispersion of these particles. CPSNPs stabilized with sodium citrate routinely exhibited highly negative zeta potentials greater than -25 mV in magnitude. Furthermore, the fluorescence quantum yield of the encapsulated fluorophore was improved by more than 4.5-fold when compared to the unencapsulated dye. The bioimaging and drug delivery capability of CPSNPs was explored. Cy3 CPSNPs dissolved quickly in the acidic environment experienced during endocytosis, releasing the encapsulated fluorophore. This is consistent with solution phase experiments that show the particles are dissolved at pH 5. CPSNPs loaded with fluorescein and a hydrophobic growth inhibitor, ceramide C6, proved the ability to simultaneously image and delivery of the hydrophobic drug to cells in vitro. Chapter 5 examined the colloidal stability of citrate and polyethylene glycol (PEG) functionalized CPSNPs in 70 volume % ethanol/30% water, both experimentally using TEM and theoretically using DLVO and polymeric steric dispersion theories. There are three basic mechanisms for colloidal stability for macroscopic suspensions (i.e., for particulate diameters down to ˜100nm), metastable electrostatic in which some finite degree of agglomeration continuously takes place because a finite energy barrier against agglomeration; and electrosteric and steric mechanisms in which infinitely high potential energy barriers toward agglomeration are present leading to thermodynamically stable suspensions. One of the fundamental issues addressed in this chapter was whether the mechanisms of electrosteric or steric dispersion, based on relatively large adsorbed polyelectrolytes for macroscopic size particulates, scales with particles in the range of ˜40 nm diameter such that a small, charged organic molecule such as citrate provides the thermodynamic colloidal stability of electrosteric mechanisms as suggested by preliminary theoretical calculations. The particle diameter-number distributions for as-prepared and after drying (at 25°C) and redispersion were used as metrics for thermodynamic colloidal stability. How efficiently particles redispersed after drying and reintroduction into the 70:30 ethanol:water solvent was used as the primary metric for whether the metastable electrostatic mechanism or thermodynamically stable electrosteric or steric approaches were responsible for the robust dispersion experimentally observed in the colloids. These experiments found that, even with the thin electrosteric layer provided by the adsorbed citrate, particles were electrosterically dispersed, and were unagglomerated when dried under argon and redispersed. Preliminary work outlining the synthesis and characterization of silver core, calcium phosphosilicate shell nanoparticles for surface plasmon coupled emission and metal enhanced fluorescence applications is discussed in Chapter 6. Thin (2-5 nm) calcium phosphosilicate shells were formed around agglomerated silver cores in the presence of 8-Methoxypyrene-1,3,6-trisulfonic acid trisodium salt (MPTS). Calcium phosphosilicate shells were consistently formed after 72 hours in the presence of 5x10-5 M CaCl2, 3x10 -5 M Na2HPO4, 3x10-6 M Na 2SiO3, and silver core nanoparticles prepared by citrate reduction in aqueous solution. However, the particles synthesized were agglomerated, resulting in a loss of the plasmon resonance peak, and the shells prepared were not thick enough to provide sufficient separation of the fluorophore from the surface to prevent quenching and allow plasmon resonance enhanced fluorescence. (Abstract shortened by UMI.)

Quantitative Mineralogical Composition of Calculi and Urine Abnormalities for Calcium Oxalate Stone Formers: A Single-Center Results.

PubMed

Kustov, Andrey V; Strelnikov, Alexander I

2018-05-03

The paper focuses on the relationship of risk factors and metabolic disorders with mineralogical composition of calculi, age and gender of calcium oxalate stone formers. Stone mineralogical composition, 24 hour biochemistry and pH-profile of urine were examined for sixty four stone formers using powder X-ray diffraction, spectrophotometric and potentiometric techniques. The analysis indicated that 44 % of calculi were composed of pure calcium oxalate monohydrate, whereas other 56 % contained both monohydrate and dihydrate or usually their mixtures with hydroxyl apatite. Hypocitraturia, hypercalciuria and hyperuricosuria were identified as the most frequent disorders. Patients with pure calcium oxalate stones and calcium oxalate mixed with apatite revealed different patterns including age, acid-base balance of urine, calcium, citrate excretion etc.Conclusions: Our results demonstrate that most patients simultaneously reveal several risk factors. The special attention should be paid to normalize the daily citrate, calcium and urate excretion. High risk patients, such as postmenopausal females or stone formers with a high apatite content require a specific metabolic evaluation towards in highlighting abnormalities associated with stone formation.

Intracellular Calcium Mobilization in Response to Ion Channel Regulators via a Calcium-Induced Calcium Release Mechanism

PubMed Central

Petrou, Terry; Olsen, Hervør L.; Thrasivoulou, Christopher; Masters, John R.; Ashmore, Jonathan F.

2017-01-01

Free intracellular calcium ([Ca2+]i), in addition to being an important second messenger, is a key regulator of many cellular processes including cell membrane potential, proliferation, and apoptosis. In many cases, the mobilization of [Ca2+]i is controlled by intracellular store activation and calcium influx. We have investigated the effect of several ion channel modulators, which have been used to treat a range of human diseases, on [Ca2+]i release, by ratiometric calcium imaging. We show that six such modulators [amiodarone (Ami), dofetilide, furosemide (Fur), minoxidil (Min), loxapine (Lox), and Nicorandil] initiate release of [Ca2+]i in prostate and breast cancer cell lines, PC3 and MCF7, respectively. Whole-cell currents in PC3 cells were inhibited by the compounds tested in patch-clamp experiments in a concentration-dependent manner. In all cases [Ca2+]i was increased by modulator concentrations comparable to those used clinically. The increase in [Ca2+]i in response to Ami, Fur, Lox, and Min was reduced significantly (P < 0.01) when the external calcium was reduced to nM concentration by chelation with EGTA. The data suggest that many ion channel regulators mobilize [Ca2+]i. We suggest a mechanism whereby calcium-induced calcium release is implicated; such a mechanism may be important for understanding the action of these compounds. PMID:27980039

Crystallization and preliminary X-ray characterization of the genetically encoded fluorescent calcium indicator protein GCaMP2

DOE Office of Scientific and Technical Information (OSTI.GOV)

Rodríguez Guilbe, María M.; Protein Research and Development Center, University of Puerto Rico; Alfaro Malavé, Elisa C.

The genetically encoded fluorescent calcium-indicator protein GCaMP2 was crystallized in the calcium-saturated form. X-ray diffraction data were collected to 2.0 Å resolution and the structure was solved by molecular replacement. Fluorescent proteins and their engineered variants have played an important role in the study of biology. The genetically encoded calcium-indicator protein GCaMP2 comprises a circularly permuted fluorescent protein coupled to the calcium-binding protein calmodulin and a calmodulin target peptide, M13, derived from the intracellular calmodulin target myosin light-chain kinase and has been used to image calcium transients in vivo. To aid rational efforts to engineer improved variants of GCaMP2, thismore » protein was crystallized in the calcium-saturated form. X-ray diffraction data were collected to 2.0 Å resolution. The crystals belong to space group C2, with unit-cell parameters a = 126.1, b = 47.1, c = 68.8 Å, β = 100.5° and one GCaMP2 molecule in the asymmetric unit. The structure was phased by molecular replacement and refinement is currently under way.« less

MARS spectral molecular imaging of lamb tissue: data collection and image analysis

NASA Astrophysics Data System (ADS)

Aamir, R.; Chernoglazov, A.; Bateman, C. J.; Butler, A. P. H.; Butler, P. H.; Anderson, N. G.; Bell, S. T.; Panta, R. K.; Healy, J. L.; Mohr, J. L.; Rajendran, K.; Walsh, M. F.; de Ruiter, N.; Gieseg, S. P.; Woodfield, T.; Renaud, P. F.; Brooke, L.; Abdul-Majid, S.; Clyne, M.; Glendenning, R.; Bones, P. J.; Billinghurst, M.; Bartneck, C.; Mandalika, H.; Grasset, R.; Schleich, N.; Scott, N.; Nik, S. J.; Opie, A.; Janmale, T.; Tang, D. N.; Kim, D.; Doesburg, R. M.; Zainon, R.; Ronaldson, J. P.; Cook, N. J.; Smithies, D. J.; Hodge, K.

2014-02-01

Spectral molecular imaging is a new imaging technique able to discriminate and quantify different components of tissue simultaneously at high spatial and high energy resolution. Our MARS scanner is an x-ray based small animal CT system designed to be used in the diagnostic energy range (20-140 keV). In this paper, we demonstrate the use of the MARS scanner, equipped with the Medipix3RX spectroscopic photon-processing detector, to discriminate fat, calcium, and water in tissue. We present data collected from a sample of lamb meat including bone as an illustrative example of human tissue imaging. The data is analyzed using our 3D Algebraic Reconstruction Algorithm (MARS-ART) and by material decomposition based on a constrained linear least squares algorithm. The results presented here clearly show the quantification of lipid-like, water-like and bone-like components of tissue. However, it is also clear to us that better algorithms could extract more information of clinical interest from our data. Because we are one of the first to present data from multi-energy photon-processing small animal CT systems, we make the raw, partial and fully processed data available with the intention that others can analyze it using their familiar routines. The raw, partially processed and fully processed data of lamb tissue along with the phantom calibration data can be found at http://hdl.handle.net/10092/8531.

Application of microscopy technique and high performance liquid chromatography for quality assessment of Polygonum multiflorum Thunb. (Heshouwu)

PubMed Central

Liang, Li; Zhao, Zhongzhen; Kang, Tingguo

2014-01-01

Background: The technique of microscopy has been applied for identification of Chinese materia medica (CMM) since decades. However, very few scientific publications report the combination of conventional microscopy and high performance liquid chromatography (HPLC) techniques for further application to quality assessment of CMM. Objective: The objective of this study is to analyze the quality of the dried root tuber of Polygonum multiflorum Thunb. (Heshouwu) and to establish the relationships between 2,3,5,4’-tetrahydroxystilbene-2-O-β-glucoside, combined anthraquinone (CAQ) and quantity of clusters of calcium oxalate. Materials and Methods: In this study, microscopy and HPLC techniques were applied to assess the quality of P. multiflorum Thunb., and SPSS software was used to establish the relationship between microscopic characteristics and chemical components. Results: The results showed close and direct correlations between the quantity of clusters of calcium oxalate in P. multiflorum Thunb. and the contents of 2,3,5,4’-tetrahydroxystilbene-2-O-β-glucoside and CAQ. From these results, it can be deduced that Polygoni Multiflori Radix with a higher quantity of clusters of calcium oxalate should be of better quality. Conclusion: The established method can be helpful for evaluating the quality of CMM based upon the identification and quantitation of chemical and ergastic substance of cells. PMID:25422540

Impact of mineral precipitation on flow and mixing in porous media determined by microcomputed tomography and MRI

DOE PAGES

Bray, Joshua M.; Lauchnor, Ellen G.; Redden, George D.; ...

2016-12-21

Here, precipitation reactions in porous media influence transport properties of the environment and can control advective and dispersive transport. In subsurface environments, mixing of saline groundwater or injected solutions for remediation with fresh groundwater can induce supersaturation of constituents and drive precipitation reactions. Magnetic resonance imaging (MRI) and micro-computed tomography (µ-CT) were employed as complimentary techniques to evaluate advection, dispersion and formation of precipitate in a 3D porous media flow cell. Two parallel fluids were flowed concentrically through the porous media under two flow rate conditions with Na 2CO 3 and CaCl 2 in the inner and outer fluids, respectively.more » Upon mixing, calcium carbonate became supersaturated and formed a precipitate at the interface of the two fluids. Spatial maps of changing local velocity fields and dispersion in the flow cell were generated from MRI, while high resolution imaging of the precipitate formed in the porous media was achieved via µ-CT imaging. Formation of a precipitate layer minimized dispersive and advective transport between the two fluids and the shape of the precipitation was influenced by the flow rate condition.« less

Impact of mineral precipitation on flow and mixing in porous media determined by microcomputed tomography and MRI

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bray, Joshua M.; Lauchnor, Ellen G.; Redden, George D.

Here, precipitation reactions in porous media influence transport properties of the environment and can control advective and dispersive transport. In subsurface environments, mixing of saline groundwater or injected solutions for remediation with fresh groundwater can induce supersaturation of constituents and drive precipitation reactions. Magnetic resonance imaging (MRI) and micro-computed tomography (µ-CT) were employed as complimentary techniques to evaluate advection, dispersion and formation of precipitate in a 3D porous media flow cell. Two parallel fluids were flowed concentrically through the porous media under two flow rate conditions with Na 2CO 3 and CaCl 2 in the inner and outer fluids, respectively.more » Upon mixing, calcium carbonate became supersaturated and formed a precipitate at the interface of the two fluids. Spatial maps of changing local velocity fields and dispersion in the flow cell were generated from MRI, while high resolution imaging of the precipitate formed in the porous media was achieved via µ-CT imaging. Formation of a precipitate layer minimized dispersive and advective transport between the two fluids and the shape of the precipitation was influenced by the flow rate condition.« less

A novel approach for detection and classification of mammographic microcalcifications using wavelet analysis and extreme learning machine.

PubMed

Malar, E; Kandaswamy, A; Chakravarthy, D; Giri Dharan, A

2012-09-01

The objective of this paper is to reveal the effectiveness of wavelet based tissue texture analysis for microcalcification detection in digitized mammograms using Extreme Learning Machine (ELM). Microcalcifications are tiny deposits of calcium in the breast tissue which are potential indicators for early detection of breast cancer. The dense nature of the breast tissue and the poor contrast of the mammogram image prohibit the effectiveness in identifying microcalcifications. Hence, a new approach to discriminate the microcalcifications from the normal tissue is done using wavelet features and is compared with different feature vectors extracted using Gray Level Spatial Dependence Matrix (GLSDM) and Gabor filter based techniques. A total of 120 Region of Interests (ROIs) extracted from 55 mammogram images of mini-Mias database, including normal and microcalcification images are used in the current research. The network is trained with the above mentioned features and the results denote that ELM produces relatively better classification accuracy (94%) with a significant reduction in training time than the other artificial neural networks like Bayesnet classifier, Naivebayes classifier, and Support Vector Machine. ELM also avoids problems like local minima, improper learning rate, and over fitting. Copyright © 2012 Elsevier Ltd. All rights reserved.

A Microfluidic System with Surface Patterning for Investigating Cavitation Bubble(s)-Cell Interaction and the Resultant Bioeffects at the Single-cell Level.

PubMed

Li, Fenfang; Yuan, Fang; Sankin, Georgy; Yang, Chen; Zhong, Pei

2017-01-10

In this manuscript, we first describe the fabrication protocol of a microfluidic chip, with gold dots and fibronectin-coated regions on the same glass substrate, that precisely controls the generation of tandem bubbles and individual cells patterned nearby with well-defined locations and shapes. We then demonstrate the generation of tandem bubbles by using two pulsed lasers illuminating a pair of gold dots with a few-microsecond time delay. We visualize the bubble-bubble interaction and jet formation by high-speed imaging and characterize the resultant flow field using particle image velocimetry (PIV). Finally, we present some applications of this technique for single cell analysis, including cell membrane poration with macromolecule uptake, localized membrane deformation determined by the displacements of attached integrin-binding beads, and intracellular calcium response from ratiometric imaging. Our results show that a fast and directional jetting flow is produced by the tandem bubble interaction, which can impose a highly localized shear stress on the surface of a cell grown in close proximity. Furthermore, different bioeffects can be induced by altering the strength of the jetting flow by adjusting the standoff distance from the cell to the tandem bubbles.

A spectral X-ray CT simulation study for quantitative determination of iron

NASA Astrophysics Data System (ADS)

Su, Ting; Kaftandjian, Valérie; Duvauchelle, Philippe; Zhu, Yuemin

2018-06-01

Iron is an essential element in the human body and disorders in iron such as iron deficiency or overload can cause serious diseases. This paper aims to explore the ability of spectral X-ray CT to quantitatively separate iron from calcium and potassium and to investigate the influence of different acquisition parameters on material decomposition performance. We simulated spectral X-ray CT imaging of a PMMA phantom filled with iron, calcium, and potassium solutions at various concentrations (15-200 mg/cc). Different acquisition parameters were considered, such as the number of energy bins (6, 10, 15, 20, 30, 60) and exposure factor per projection (0.025, 0.1, 1, 10, 100 mA s). Based on the simulation data, we investigated the performance of two regularized material decomposition approaches: projection domain method and image domain method. It was found that the former method discriminated iron from calcium, potassium and water in all cases and tended to benefit from lower number of energy bins for lower exposure factor acquisition. The latter method succeeded in iron determination only when the number of energy bins equals 60, and in this case, the contrast-to-noise ratios of the decomposed iron images are higher than those obtained using the projection domain method. The results demonstrate that both methods are able to discriminate and quantify iron from calcium, potassium and water under certain conditions. Their performances vary with the acquisition parameters of spectral CT. One can use one method or the other to benefit better performance according to the data available.

Boron in Calcium Sulfate Vein at Catabola, Mars

NASA Image and Video Library

2016-12-13

The highest concentration of boron measured on Mars, as of late 2016, is in this mineral vein, called "Catabola," examined with the Chemistry and Camera (ChemCam) instrument on NASA's Curiosity rover on Aug, 25, 2016, during Sol 1441 of the mission. This two-part illustration shows the context of the erosion-resistant, raised vein, in an image from Curiosity's Mast Camera (Mastcam), and a detailed inset image from ChemCam's remote micro-imager. The inset includes indicators of the boron content measured at 10 points along the vein that were analyzed with ChemCam's laser-firing spectrometer. The vein's main component is calcium sulfate. The highest boron content identified is less than one-tenth of one percent. The heights of the orange bars at each point indicate relative abundance of boron, compared with boron content at other points. The scale bar for the inset is 9.2 millimeters, or about 0.36 inch. The ChemCam image is enhanced with color information from Mastcam. http://photojournal.jpl.nasa.gov/catalog/PIA21251

In situ intracellular calcium oscillations in osteocytes in intact mouse long bones under dynamic mechanical loading

PubMed Central

Jing, Da; Baik, Andrew D.; Lu, X. Lucas; Zhou, Bin; Lai, Xiaohan; Wang, Liyun; Luo, Erping; Guo, X. Edward

2014-01-01

Osteocytes have been hypothesized to be the major mechanosensors in bone. How in situ osteocytes respond to mechanical stimuli is still unclear because of technical difficulties. In vitro studies have shown that osteocytes exhibited unique calcium (Ca2+) oscillations to fluid shear. However, whether this mechanotransduction phenomenon holds for in situ osteocytes embedded within a mineralized bone matrix under dynamic loading remains unknown. Using a novel synchronized loading/imaging technique, we successfully visualized in real time and quantified Ca2+ responses in osteocytes and bone surface cells in situ under controlled dynamic loading on intact mouse tibia. The resultant fluid-induced shear stress on the osteocyte in the lacunocanalicular system (LCS) was also quantified. Osteocytes, but not surface cells, displayed repetitive Ca2+ spikes in response to dynamic loading, with spike frequency and magnitude dependent on load magnitude, tissue strain, and shear stress in the LCS. The Ca2+ oscillations were significantly reduced by endoplasmic reticulum (ER) depletion and P2 purinergic receptor (P2R)/phospholipase C (PLC) inhibition. This study provides direct evidence that osteocytes respond to in situ mechanical loading by Ca2+ oscillations, which are dependent on the P2R/PLC/inositol trisphosphate/ER pathway. This study develops a novel approach in skeletal mechanobiology and also advances our fundamental knowledge of bone mechanotransduction.—Jing, D., Baik, A. D., Lu, X. L., Zhou, B., Lai, X., Wang, L., Luo, E., Guo, X. E. In situ intracellular calcium oscillations in osteocytes in intact mouse long bones under dynamic mechanical loading. PMID:24347610

Densitometric and biochemical values of broiler tibias at different ages.

PubMed

Barreiro, F R; Sagula, A L; Junqueira, O M; Pereira, G T; Baraldi-Artoni, S M

2009-12-01

The objective of this experiment was to determine the normal values of bone radiographic density (BRD) by using the optical densitometry in radiographic images and the biochemical values represented by serum calcium, ash percentage, and minerals (calcium, phosphorus, and magnesium) from tibia ash of Cobb broilers at 8, 22, and 43 d of age. A total of 14 broilers were used for densitometric analysis, and 15 were used for biochemical dosages. The BRD values increased (P < 0.05) with age and in all tibia regions (proximal epiphysis, diaphysis, and distal epiphysis), concluding that growth was a determinative factor for bone performance, demanding a higher BRD during broiler development. Tibia proximal epiphysis presented higher BRD values in relation to the other bone regions (P < 0.05), as a result of a possible biomechanical adaptation to ligaments and tension of the muscle tendons at this region, allowing the support of the muscle mass increase. The serum calcium values were kept constant, as a result of the appropriate nutritional levels of the diet that supported the animal homeostasis. The bone ash and mineral percentage increased (P < 0.05) at 22 d of age, due to the higher mineral requirement in this age. The correlation between bone densitometry and the invasive techniques showed that the bone densitometry can substitute the determination of mineral percentage in the ash. This experiment presented normal values of the noninvasive and invasive methods more used in aviculture, allowing us to compare, subsequently, pathological and physiological values or results of broilers fed with different diets.

Calcium impregnation of coal enriched in CO.sub.2 using high-pressure techniques

NASA Technical Reports Server (NTRS)

Gavalas, George R. (Inventor); Sharma, Pramod K. (Inventor); Voecks, Gerald E. (Inventor)

1990-01-01

Methods are described for impregnating coal with calcium carbonate by utilizing an aqueous phase ionic reaction between calcium acetate, calcium hydroxide, and water with CO.sub.2 contained within the coal. The coal is enriched in CO.sub.2 by contacting it with CO.sub.2 at high pressure, in either a continuous or pulsed mode. The inclusion of CO.sub.2 in the coal during the process does not involve evacuating the coal and subsequently absorbing CO.sub.2 onto the coal as in prior methods. Rather, the coal is treated with carbon dioxide at high pressure in a practical and viable approach. The impregnation of coal by calcium compounds not only reduces sulfur emissions by effectively tying up the sulfur as calcium sulfide or sulfate, but also increases the gasification or combustion rate. The invention also encompasses the use of other Group IIA elements, as well as the coal products resulting from the methods of treatment described.

Low temperature pyrolysis of coal or oil shale in the presence of calcium compounds

DOEpatents

Khan, M. Rashid

1988-01-01

A coal pyrolysis technique or process is described in which particulate coal is pyrolyzed in the presence of about 5 to 21 wt. % of a calcium compound selected from calcium oxide, calcined (hydrate) dolomite, or calcined calcium hydrate to produce a high quality hydrocarbon liquid and a combustible product gas which are characterized by low sulfur content. The pyrolysis is achieved by heating the coal-calcium compound mixture at a relatively slow rate at a temperature of about 450.degree. to 700.degree. C. over a duration of about 10 to 60 minutes in a fixed or moving bed reactor. The gas exhibits an increased yield in hydrogen and C.sub.1 -C.sub.8 hydrocarbons and a reduction in H.sub.2 S over gas obtainable by pyrolyzing cola without the calcium compound. The liquid product obtained is of a sufficient quality to permit its use directly as a fuel and has a reduced sulfur and oxygen content which inhibits polymerization during storage.

Regression-Based Identification of Behavior-Encoding Neurons During Large-Scale Optical Imaging of Neural Activity at Cellular Resolution

PubMed Central

Miri, Andrew; Daie, Kayvon; Burdine, Rebecca D.; Aksay, Emre

2011-01-01

The advent of methods for optical imaging of large-scale neural activity at cellular resolution in behaving animals presents the problem of identifying behavior-encoding cells within the resulting image time series. Rapid and precise identification of cells with particular neural encoding would facilitate targeted activity measurements and perturbations useful in characterizing the operating principles of neural circuits. Here we report a regression-based approach to semiautomatically identify neurons that is based on the correlation of fluorescence time series with quantitative measurements of behavior. The approach is illustrated with a novel preparation allowing synchronous eye tracking and two-photon laser scanning fluorescence imaging of calcium changes in populations of hindbrain neurons during spontaneous eye movement in the larval zebrafish. Putative velocity-to-position oculomotor integrator neurons were identified that showed a broad spatial distribution and diversity of encoding. Optical identification of integrator neurons was confirmed with targeted loose-patch electrical recording and laser ablation. The general regression-based approach we demonstrate should be widely applicable to calcium imaging time series in behaving animals. PMID:21084686

Simultaneous imaging of neural activity in three dimensions

PubMed Central

Quirin, Sean; Jackson, Jesse; Peterka, Darcy S.; Yuste, Rafael

2014-01-01

We introduce a scanless optical method to image neuronal activity in three dimensions simultaneously. Using a spatial light modulator and a custom-designed phase mask, we illuminate and collect light simultaneously from different focal planes and perform calcium imaging of neuronal activity in vitro and in vivo. This method, combining structured illumination with volume projection imaging, could be used as a technological platform for brain activity mapping. PMID:24772066

Experiment K-6-01. Distribution and biochemistry of mineral and matrix in the femurs of rats

NASA Technical Reports Server (NTRS)

Arnaud, S.; Mechanic, G.; Buckendahl, P.; Bromage, T.; Boyde, A.; Elliott, J.; Katz, E.; Durnova, G

1990-01-01

Previous analyses of the composition of mineral and matrix in the bone of young rats following space flight has revealed deficits in calcium, phosphorus, and osteocalcin, a non-collagenous protein, without an associated decrease in collagen. To characterize the location and nature of this mineralization defect in a weight bearing long bone, the femur, researchers attempted to relate the spatial distribution of mineral in situ in the proximal, central and distal thirds of the femoral diaphysis to the biochemical composition of bone from the same area. Biochemical analyses revealed lower concentrations of calcium, phosphorus and osteocalcin but not collagen only in the central third of the diaphysis of the flight animals (F) compared to synchronous controls (S). Collagen concentration was reduced only in the proximal third of the diaphysis, where all 3 crosslinks, expressed as nM/mol collagen were higher in F than S. A new technique, x ray microtomography, with a resolution of 26 microns, was used to obtain semi-quantitative data on mineral distribution in reconstructed sections of wet whole bone. To improve the resolution of the mineral density distribution, images of the surfaces of cut sections were analyzed by backscattered electrons in a scanning electron microscope (BSE). There was good agreement between the results of the two stereochemical techniques which revealed distinct patterns of mineralization in transverse and longitudinal directions of the diaphysis. The novel methodology developed for this flight experiment shows considerable promise in elucidating the biochemical nature of what appear to be regional alterations in the mineralization of long bones of animals exposed to spaceflight.

A fully automated multi-modal computer aided diagnosis approach to coronary calcium scoring of MSCT images

NASA Astrophysics Data System (ADS)

Wu, Jing; Ferns, Gordon; Giles, John; Lewis, Emma

2012-03-01

Inter- and intra- observer variability is a problem often faced when an expert or observer is tasked with assessing the severity of a disease. This issue is keenly felt in coronary calcium scoring of patients suffering from atherosclerosis where in clinical practice, the observer must identify firstly the presence, followed by the location of candidate calcified plaques found within the coronary arteries that may prevent oxygenated blood flow to the heart muscle. However, it can be difficult for a human observer to differentiate calcified plaques that are located in the coronary arteries from those found in surrounding anatomy such as the mitral valve or pericardium. In addition to the benefits to scoring accuracy, the use of fast, low dose multi-slice CT imaging to perform the cardiac scan is capable of acquiring the entire heart within a single breath hold. Thus exposing the patient to lower radiation dose, which for a progressive disease such as atherosclerosis where multiple scans may be required, is beneficial to their health. Presented here is a fully automated method for calcium scoring using both the traditional Agatston method, as well as the volume scoring method. Elimination of the unwanted regions of the cardiac image slices such as lungs, ribs, and vertebrae is carried out using adaptive heart isolation. Such regions cannot contain calcified plaques but can be of a similar intensity and their removal will aid detection. Removal of both the ascending and descending aortas, as they contain clinical insignificant plaques, is necessary before the final calcium scores are calculated and examined against ground truth scores of three averaged expert observer results. The results presented here are intended to show the feasibility and requirement for an automated scoring method to reduce the subjectivity and reproducibility error inherent with manual clinical calcium scoring.

High-dose calcium stimulation test in a case of insulinoma masquerading as hysteria.

PubMed

Nakamura, Yoshio; Doi, Ryuichiro; Kohno, Yasuhiro; Shimono, Dai; Kuwamura, Naomitsu; Inoue, Koichi; Koshiyama, Hiroyuki; Imamura, Masayuki

2002-11-01

It is reported that some cases with insulinoma present with neuropsychiatric symptoms and are often misdiagnosed as psychosis. Here we report a case of insulinoma masquerading as hysteria, whose final diagnosis could be made using high-dose calcium stimulation test. A 28-yr-old woman was referred presenting with substupor, mutism, mannerism, restlessness, and incoherence. Laboratory examinations revealed hypoglycemia (33 mg/dL) and detectable insulin levels (9.7 microU/mL), suggesting the diagnosis of insulinoma. However, neither imaging studies nor selective arterial calcium injection (SACI) test with a conventional dose of calcium (0.025 mEq/kg) indicated the tumor. High-dose calcium injection (0.05 mEq/kg) evoked insulin secretion when injected into superior mesenteric artery. A solitary tumor in the head of the pancreas was resected, and her plasma glucose returned to normal. Postoperatively, iv injection of secretin resulted in a normal response of insulin, which was not found preoperatively. This case suggests the usefulness of the SACI test with high-dose of calcium in the case of insulinoma when the standard dose fails to detect such a tumor.

In vivo imaging of neuronal calcium during electrode implantation: Spatial and temporal mapping of damage and recovery.

PubMed

Eles, James R; Vazquez, Alberto L; Kozai, Takashi D Y; Cui, X Tracy

2018-08-01

Implantable electrode devices enable long-term electrophysiological recordings for brain-machine interfaces and basic neuroscience research. Implantation of these devices, however, leads to neuronal damage and progressive neural degeneration that can lead to device failure. The present study uses in vivo two-photon microscopy to study the calcium activity and morphology of neurons before, during, and one month after electrode implantation to determine how implantation trauma injures neurons. We show that implantation leads to prolonged, elevated calcium levels in neurons within 150 μm of the electrode interface. These neurons show signs of mechanical distortion and mechanoporation after implantation, suggesting that calcium influx is related to mechanical trauma. Further, calcium-laden neurites develop signs of axonal injury at 1-3 h post-insert. Over the first month after implantation, physiological neuronal calcium activity increases, suggesting that neurons may be recovering. By defining the mechanisms of neuron damage after electrode implantation, our results suggest new directions for therapies to improve electrode longevity. Copyright © 2018 Elsevier Ltd. All rights reserved.

General anesthesia selectively disrupts astrocyte calcium signaling in the awake mouse cortex

PubMed Central

Thrane, Alexander Stanley; Zeppenfeld, Douglas; Lou, Nanhong; Xu, Qiwu; Nagelhus, Erlend Arnulf; Nedergaard, Maiken

2012-01-01

Calcium signaling represents the principle pathway by which astrocytes respond to neuronal activity. General anesthetics are routinely used in clinical practice to induce a sleep-like state, allowing otherwise painful procedures to be performed. Anesthetic drugs are thought to mainly target neurons in the brain and act by suppressing synaptic activity. However, the direct effect of general anesthesia on astrocyte signaling in awake animals has not previously been addressed. This is a critical issue, because calcium signaling may represent an essential mechanism through which astrocytes can modulate synaptic activity. In our study, we performed calcium imaging in awake head-restrained mice and found that three commonly used anesthetic combinations (ketamine/xylazine, isoflurane, and urethane) markedly suppressed calcium transients in neocortical astrocytes. Additionally, all three anesthetics masked potentially important features of the astrocyte calcium signals, such as synchronized widespread transients that appeared to be associated with arousal in awake animals. Notably, anesthesia affected calcium transients in both processes and soma and depressed spontaneous signals, as well as calcium responses, evoked by whisker stimulation or agonist application. We show that these calcium transients are inositol 1,4,5-triphosphate type 2 receptor (IP3R2)-dependent but resistant to a local blockade of glutamatergic or purinergic signaling. Finally, we found that doses of anesthesia insufficient to affect neuronal responses to whisker stimulation selectively suppressed astrocyte calcium signals. Taken together, these data suggest that general anesthesia may suppress astrocyte calcium signals independently of neuronal activity. We propose that these glial effects may constitute a nonneuronal mechanism for sedative action of anesthetic drugs. PMID:23112168

Hemispheric asymmetry of macroscopic and elementary calcium signals mediated by InsP3 in Xenopus oocytes.

PubMed

Callamaras, N; Sun, X P; Ivorra, I; Parker, I

1998-09-01

1. The mechanisms underlying hemispheric asymmetry of the inositol 1, 4,5-trisphosphate (InsP3)-calcium signalling pathway in Xenopus oocytes were examined by fluorescence imaging of calcium signals and recording calcium-activated Cl- currents (ICl,Ca) evoked by intracellular calcium injections and photorelease of InsP3. 2. The maximal ICl,Ca evoked by strong photorelease of InsP3 was 8 times greater in the animal than the vegetal hemisphere, but the average threshold amounts of InsP3 required to evoke detectable currents were similar in each hemisphere. 3. Currents evoked by injections of calcium were about 2.5 times greater near the animal pole than near the vegetal pole, whereas fluorescence signals evoked by injections were similar in each hemisphere. 4. Calcium waves were evoked by photolysis flashes of similar strengths in both hemispheres of albino oocytes, but peak calcium levels evoked by supramaximal stimuli were 70 % greater in the animal hemisphere. 5. Elementary calcium release events (puffs) in the animal hemisphere had amplitudes about double that in the vegetal hemisphere, and more often involved coupled release from adjacent sites. Calcium release sites were more closely packed in the animal hemisphere, with a mean spacing of about 1.5 micro m compared with 2.25 micro m in the vegetal hemisphere. 6. The larger amplitude of currents mediated by InsP3 in the animal hemisphere, therefore, involves an increased flux of calcium at individual release units, a more dense packing of release units and a higher density of Cl- channels.

Hemispheric asymmetry of macroscopic and elementary calcium signals mediated by InsP3 in Xenopus oocytes

PubMed Central

Callamaras, Nick; Sun, Xiao-Ping; Ivorra, Isabel; Parker, Ian

1998-01-01

The mechanisms underlying hemispheric asymmetry of the inositol 1,4,5-trisphosphate (InsP3)-calcium signalling pathway in Xenopus oocytes were examined by fluorescence imaging of calcium signals and recording calcium-activated Cl− currents (ICl,Ca) evoked by intracellular calcium injections and photorelease of InsP3. The maximal ICl,Ca evoked by strong photorelease of InsP3 was 8 times greater in the animal than the vegetal hemisphere, but the average threshold amounts of InsP3 required to evoke detectable currents were similar in each hemisphere. Currents evoked by injections of calcium were about 2.5 times greater near the animal pole than near the vegetal pole, whereas fluorescence signals evoked by injections were similar in each hemisphere. Calcium waves were evoked by photolysis flashes of similar strengths in both hemispheres of albino oocytes, but peak calcium levels evoked by supramaximal stimuli were 70% greater in the animal hemisphere. Elementary calcium release events (puffs) in the animal hemisphere had amplitudes about double that in the vegetal hemisphere, and more often involved coupled release from adjacent sites. Calcium release sites were more closely packed in the animal hemisphere, with a mean spacing of about 1.5 μm compared with 2.25 μm in the vegetal hemisphere. The larger amplitude of currents mediated by InsP3 in the animal hemisphere, therefore, involves an increased flux of calcium at individual release units, a more dense packing of release units and a higher density of Cl− channels. PMID:9706018

Negative Inotropic Effects of High Mobility Box Group 1 Protein in Isolated Contracting Cardiac Myocytes

PubMed Central

Tzeng, Huei-Ping; Fan, Jinping; Vallejo, Jesus G.; Dong, Jian Wen; Chen, Xiongwen; Houser, Steven R.; Mann, Douglas L.

2013-01-01

HMGB1 released from necrotic cells or macrophages functions as a late inflammatory mediator, and has been shown to induce cardiovascular collapse during sepsis. Thus far, however, the effect(s) of HMGB1 in the heart are not known. We determined the effects of HMGB1 on isolated feline cardiac myocytes by measuring sarcomere shortening in contracting cardiac myocytes, intracellular Ca2+ transients using fluo-3, and L-type calcium currents using whole cell perforate configuration of the patch clamp technique. Treatment of isolated myocytes with HMGB1 (100 ng/ml) resulted in a 70% decrease in sarcomere shortening and a 50% decrease in the height of the peak Ca++ transient within 5 min (p <0.01). The immediate negative inotropic effects HMGB1 on cell contractility and calcium homeostasis were partially reversible upon washout of HMGB1. A significant inhibition of the inward L-type calcium currents also was documented by the patch clamp technique. HMGB1 induced the PKCε translocation and a PKC inhibitor significantly attenuated the negative inotropic effects of HMGB1. These studies show for the first time that HMGB1 impairs sarcomere shortening by decreasing calcium availability in cardiac myocytes through modulating membrane calcium influx, and suggest that HMGB1 maybe act as a novel myocardial depressant factor during cardiac injury. PMID:18223193

Novel botanical drug DA-9803 prevents deficits in Alzheimer's mouse models.

PubMed

Pagnier, Guillaume J; Kastanenka, Ksenia V; Sohn, Miwon; Choi, Sangzin; Choi, Song-Hyen; Soh, HyeYeon; Bacskai, Brian J

2018-01-29

Alzheimer's disease (AD) is a neurodegenerative disorder characterized by deposition of amyloid plaques and disruption of neural circuitry, leading to cognitive decline. Animal models of AD deposit senile plaques and exhibit structural and functional deficits in neurons and neural networks. An effective treatment would prevent or restore these deficits, including calcium dyshomeostasis observed with in-vivo imaging. We examined the effects of DA-9803, a multimodal botanical drug, in 5XFAD and APP/PS1 transgenic mice which underwent daily oral treatment with 30 or 100 mg/kg DA-9803 or vehicle alone. Behavioral testing and longitudinal imaging of amyloid deposits and intracellular calcium in neurons with multiphoton microscopy was performed. Chronic administration of DA-9803 restored behavioral deficits in 5XFAD mice and reduced amyloid-β levels. DA-9803 also prevented progressive amyloid plaque deposition in APP/PS1 mice. Elevated calcium, detected in a subset of neurons before the treatment, was restored and served as a functional indicator of treatment efficacy in addition to the behavioral readout. In contrast, mice treated with vehicle alone continued to progressively accumulate amyloid plaques and calcium overload. In summary, treatment with DA-9803 prevented structural and functional outcome measures in mouse models of AD. Thus, DA-9803 shows promise as a novel therapeutic approach for Alzheimer's disease.

Calcium absorption from fortified ice cream formulations compared with calcium absorption from milk.

PubMed

van der Hee, Regine M; Miret, Silvia; Slettenaar, Marieke; Duchateau, Guus S M J E; Rietveld, Anton G; Wilkinson, Joy E; Quail, Patricia J; Berry, Mark J; Dainty, Jack R; Teucher, Birgit; Fairweather-Tait, Susan J

2009-05-01

Optimal bone mass in early adulthood is achieved through appropriate diet and lifestyle, thereby protecting against osteoporosis and risk of bone fracture in later life. Calcium and vitamin D are essential to build adequate bones, but calcium intakes of many population groups do not meet dietary reference values. In addition, changes in dietary patterns are exacerbating the problem, thereby emphasizing the important role of calcium-rich food products. We have designed a calcium-fortified ice cream formulation that is lower in fat than regular ice cream and could provide a useful source of additional dietary calcium. Calcium absorption from two different ice cream formulations was determined in young adults and compared with milk. Sixteen healthy volunteers (25 to 45 years of age), recruited from the general public of The Netherlands, participated in a randomized, reference-controlled, double-blind cross-over study in which two test products and milk were consumed with a light standard breakfast on three separate occasions: a standard portion of ice cream (60 g) fortified with milk minerals and containing a low level (3%) of butter fat, ice cream (60 g) fortified with milk minerals and containing a typical level (9%) of coconut oil, and reduced-fat milk (1.7% milk fat) (200 mL). Calcium absorption was measured by the dual-label stable isotope technique. Effects on calcium absorption were evaluated by analysis of variance. Fractional absorption of calcium from the 3% butterfat ice cream, 9% coconut oil ice cream, and milk was 26%+/-8%, 28%+/-5%, and 31%+/-9%, respectively, and did not differ significantly (P=0.159). Results indicate that calcium bioavailability in the two calcium-fortified ice cream formulations used in this study is as high as milk, indicating that ice cream may be a good vehicle for delivery of calcium.

Presentation and conservative management of acute calcific tendinopathy: a case study and literature review.

PubMed

Scibek, Jason S; Carcia, Christopher R

2012-11-01

The efficacy of a variety of noninvasive, conservative management techniques for calcific tendinopathy has been investigated and established for improving pain and function and/or facilitating a decrease in the size or presence of calcium deposits. Surprisingly, few have reported on the use of traditional therapeutic exercise and rehabilitation alone in the management of this condition, given the often spontaneous resorptive nature of calcium deposits. The purpose of this case is to present the results of a conservative approach, including therapeutic exercise, for the management of calcific tendinopathy of the supraspinatus, with an emphasis on patient outcomes. The patient was a self-referred 41-y-old man with complaints of acute right-shoulder pain and difficulty sleeping. Imaging studies revealed liquefied calcium deposits in the right supraspinatus. The patient reported constant pain at rest (9/10) and tenderness in the area of the greater tuberosity. He exhibited a decrease in all shoulder motions and had reduced strength. The simple shoulder test (SST) revealed limited function (0/12). Conservative management included superficial modalities and medication for pain and a regimen of scapulothoracic and glenohumeral range-of-motion (ROM) and strengthening exercises. At discharge, pain levels decreased to 0/10 and SST scores increased to 12/12. ROM was full in all planes, and resisted motion was strong and pain free. The patient was able to engage in endurance activities and continue practicing as a health care provider. The outcomes with respect to pain, function, and patient satisfaction provide evidence to support the use of conservative therapeutic interventions when managing patients with acute cases of calcific tendinopathy. Successful management of calcific tendinopathy requires attention to outcomes and an understanding of the pathophysiology, prognostic factors, and physical interventions based on the current stage of the calcium deposits and the patient's status in the healing continuum.

Analysis of calcium-induced effects on the conformation of fengycin.

PubMed

Nasir, Mehmet Nail; Laurent, Pascal; Flore, Christelle; Lins, Laurence; Ongena, Marc; Deleu, Magali

2013-06-01

Fengycin is a natural lipopeptide with antifungal and eliciting properties and able to inhibit the activity of phospholipase A2. A combination of CD, FT-IR, NMR and fluorescence spectroscopic techniques was applied to elucidate its conformation in a membrane-mimicking environment and to investigate the effect of calcium ions on it. We mainly observed that fengycin adopts a turn conformation. Our results showed that calcium ions are bound by the two charged glutamates. The calcium binding has an influence on the fengycin conformation and more particularly, on the environment of the tyrosine residues. The modulation of the fengycin conformation by the environmental conditions may influence its biological properties. Copyright © 2013 Elsevier B.V. All rights reserved.

Releasing effects in flame photometry: Determination of calcium

USGS Publications Warehouse

Dinnin, J.I.

1960-01-01

Strontium, lanthanum, neodymium, samarium, and yttrium completely release the flame emission of calcium from the depressive effects of sulfate, phosphate, and aluminate. Magnesium, beryllium, barium, and scandium release most of the calcium emission. These cations, when present in high concentration, preferentially form compounds with the depressing anions when the solution is evaporated rapidly in the flame. The mechanism of the interference and releasing effects is explained on the basis of the chemical equilibria in the evaporating droplets of solution and is shown to depend upon the nature of the compounds present in the aqueous phase of the solution. The need for background correction techniques is stressed. The releasing effect is used in the determination of calcium in silicate rocks without the need for separations.

Crystallization from Gels

NASA Astrophysics Data System (ADS)

Narayana Kalkura, S.; Natarajan, Subramanian

Among the various crystallization techniques, crystallization in gels has found wide applications in the fields of biomineralization and macromolecular crystallization in addition to crystallizing materials having nonlinear optical, ferroelectric, ferromagnetic, and other properties. Furthermore, by using this method it is possible to grow single crystals with very high perfection that are difficult to grow by other techniques. The gel method of crystallization provides an ideal technique to study crystal deposition diseases, which could lead to better understanding of their etiology. This chapter focuses on crystallization in gels of compounds that are responsible for crystal deposition diseases. The introduction is followed by a description of the various gels used, the mechanism of gelling, and the fascinating phenomenon of Liesegang ring formation, along with various gel growth techniques. The importance and scope of study on crystal deposition diseases and the need for crystal growth experiments using gel media are stressed. The various crystal deposition diseases, viz. (1) urolithiasis, (2) gout or arthritis, (3) cholelithiasis and atherosclerosis, and (4) pancreatitis and details regarding the constituents of the crystal deposits responsible for the pathological mineralization are discussed. Brief accounts of the theories of the formation of urinary stones and gallstones and the role of trace elements in urinary stone formation are also given. The crystallization in gels of (1) the urinary stone constituents, viz. calcium oxalate, calcium phosphates, uric acid, cystine, etc., (2) the constituents of the gallstones, viz. cholesterol, calcium carbonate, etc., (3) the major constituent of the pancreatic calculi, viz., calcium carbonate, and (4) cholic acid, a steroidal hormone are presented. The effect of various organic and inorganic ions, trace elements, and extracts from cereals, herbs, and fruits on the crystallization of major urinary stone and gallstone constituents are described. In addition, tables of gel-grown organic and inorganic crystals are provided.

Printability of calcium phosphate: calcium sulfate powders for the application of tissue engineered bone scaffolds using the 3D printing technique.

PubMed

Zhou, Zuoxin; Buchanan, Fraser; Mitchell, Christina; Dunne, Nicholas

2014-05-01

In this study, calcium phosphate (CaP) powders were blended with a three-dimensional printing (3DP) calcium sulfate (CaSO4)-based powder and the resulting composite powders were printed with a water-based binder using the 3DP technology. Application of a water-based binder ensured the manufacture of CaP:CaSO4 constructs on a reliable and repeatable basis, without long term damage of the printhead. Printability of CaP:CaSO4 powders was quantitatively assessed by investigating the key 3DP process parameters, i.e. in-process powder bed packing, drop penetration behavior and the quality of printed solid constructs. Effects of particle size, CaP:CaSO4 ratio and CaP powder type on the 3DP process were considered. The drop penetration technique was used to reliably identify powder formulations that could be potentially used for the application of tissue engineered bone scaffolds using the 3DP technique. Significant improvements (p<0.05) in the 3DP process parameters were found for CaP (30-110 μm):CaSO4 powders compared to CaP (<20 μm):CaSO4 powders. Higher compressive strength was obtained for the powders with the higher CaP:CaSO4 ratio. Hydroxyapatite (HA):CaSO4 powders showed better results than beta-tricalcium phosphate (β-TCP):CaSO4 powders. Solid and porous constructs were manufactured using the 3DP technique from the optimized CaP:CaSO4 powder formulations. High-quality printed constructs were manufactured, which exhibited appropriate green compressive strength and a high level of printing accuracy. Copyright © 2014 Elsevier B.V. All rights reserved.

Imaging-based assessment of the mineral composition of urinary stones: an in vitro study of the combination of hounsfield unit measurement in noncontrast helical computerized tomography and the twinkling artifact in color Doppler ultrasound.

PubMed

Hassani, Hakim; Raynal, Gauthier; Spie, Romain; Daudon, Michel; Vallée, Jean-Noël

2012-05-01

We evaluated the value of combining noncontrast helical computerized tomography (NCHCT) and color Doppler ultrasound in the assessment of the composition of urinary stones. In vitro, we studied 120 stones of known composition, that separate into the five main types: 18 calcium oxalate monohydrate (COM) stones, 41 calcium oxalate dihydrate (COD) stones, 24 uric acid stones, 25 calcium phosphate stones and 12 cystine calculi. Stones were characterized in terms of their Hounsfield density (HU) in NCHCT and the presence of a twinkling artifact (TA) in color Doppler ultrasound. There were statistically significant HU differences between calcium and non-calcium stones (p < 0.001), calcium oxalate stones and calcium phosphate stones (p < 0.001) and uric acid stones and cystine calculi (p < 0.001) but not between COM and COD stones (p = 0.786). Hence, the HU was a predictive factor of the composition of all types of stones, other than for COM and COD stones within the calcium oxalate class (p > 0.05). We found that the TA does not enable differentiation between calcium and non-calcium stones (p > 0.999), calcium oxalate stones and calcium phosphate stones (p = 0.15), or uric acid stones and cystine calculi (p = 0.079). However, it did reveal a significant difference between COM and COD stones (p = 0.002). The absence of a TA is a predictive factor for the presence of COM stones (p = 0.008). Hence, the association of NCHCT and Doppler enables the accurate classification of the five types of stones in vitro. Copyright © 2012 World Federation for Ultrasound in Medicine & Biology. Published by Elsevier Inc. All rights reserved.

Calcium scoring with dual-energy CT in men and women: an anthropomorphic phantom study

NASA Astrophysics Data System (ADS)

Li, Qin; Liu, Songtao; Myers, Kyle; Gavrielides, Marios A.; Zeng, Rongping; Sahiner, Berkman; Petrick, Nicholas

2016-03-01

This work aimed to quantify and compare the potential impact of gender differences on coronary artery calcium scoring with dual-energy CT. An anthropomorphic thorax phantom with four synthetic heart vessels (diameter 3-4.5 mm: female/male left main and left circumflex artery) were scanned with and without female breast plates. Ten repeat scans were acquired in both single- and dual-energy modes and reconstructed at six reconstruction settings: two slice thicknesses (3 mm, 0.6 mm) and three reconstruction algorithms (FBP, IR3, IR5). Agatston and calcium volume scores were estimated from the reconstructed data using a segmentation-based approach. Total calcium score (summation of four vessels), and male/female calcium scores (summation of male/female vessels scanned in phantom without/with breast plates) were calculated accordingly. Both Agatston and calcium volume scores were found comparable between single- and dual-energy scans (Pearson r= 0.99, p<0.05). The total calcium scores were larger for the thinner slice thickness. Among the scores obtained from the three reconstruction algorithms, FBP yielded the highest and IR5 yielded the lowest scores. The total calcium scores from the phantom without breast plates were significantly larger than those from the phantom with breast plates, and the difference increased with the stronger denoising in iterative algorithm and with thicker slices. Both gender-based anatomical differences and vessel size impacted the calcium scores. The calcium volume scores tended to be underestimated when the vessels were smaller. These findings are valuable for understanding inconsistencies between women and men in calcium scoring, and for standardizing imaging protocols for improved gender-specific calcium scoring.

Proteins on exocytic vesicles mediate calcium-triggered fusion.

PubMed Central

Vogel, S S; Zimmerberg, J

1992-01-01

In many exocytic systems, micromolar concentrations of intracellular Ca2+ trigger fusion. We find that aggregates of secretory granules isolated from sea urchin eggs fuse together when perfused with greater than or equal to 10 microM free Ca2+. Mixing of membrane components was demonstrated by transfer of fluorescent lipophilic dye, and melding of granule contents was seen with differential interference microscopy. A technique based upon light scattering was developed to conveniently detect fusion. Two protein modifiers, trypsin and N-ethylmaleimide, inhibit granule-granule fusion at concentrations similar to those that inhibit granule-plasma membrane fusion. We suggest that molecular machinery sufficient for Ca(2+)-triggered fusion resides on secretory granules as purified and that at least some of these essential components are proteinaceous. Images PMID:1584814

Image Information Obtained Using a Charge-Coupled Device (CCD) Camera During an Immersion Liquid Evaporation Process for Measuring the Refractive Index of Solid Particles.

PubMed

Niskanen, Ilpo; Sutinen, Veijo; Thungström, Göran; Räty, Jukka

2018-06-01

The refractive index is a fundamental physical property of a medium, which can be used for the identification and purity issues of all media. Here we describe a refractive index measurement technique to determine simultaneously the refractive index of different solid particles by monitoring the transmittance of light from a suspension using a charge-coupled device (CCD) camera. An important feature of the measurement is the liquid evaporation process for the refractive index matching of the solid particle and the immersion liquid; this was realized by using a pair of volatile and non-volatile immersion liquids. In this study, refractive indices of calcium fluoride (CaF 2 ) and barium fluoride (BaF 2 ) were determined using the proposed method.

Towards the Physics of Calcium Signalling in Plants

PubMed Central

Vaz Martins, Teresa; Evans, Matthew J.; Woolfenden, Hugh C.; Morris, Richard J.

2013-01-01

Calcium is an abundant element with a wide variety of important roles within cells. Calcium ions are inter- and intra-cellular messengers that are involved in numerous signalling pathways. Fluctuating compartment-specific calcium ion concentrations can lead to localised and even plant-wide oscillations that can regulate downstream events. Understanding the mechanisms that give rise to these complex patterns that vary both in space and time can be challenging, even in cases for which individual components have been identified. Taking a systems biology approach, mathematical and computational techniques can be employed to produce models that recapitulate experimental observations and capture our current understanding of the system. Useful models make novel predictions that can be investigated and falsified experimentally. This review brings together recent work on the modelling of calcium signalling in plants, from the scale of ion channels through to plant-wide responses to external stimuli. Some in silico results that have informed later experiments are highlighted. PMID:27137393

Resolution Properties of a Calcium Tungstate (CaWO4) Screen Coupled to a CMOS Imaging Detector

NASA Astrophysics Data System (ADS)

Koukou, Vaia; Martini, Niki; Valais, Ioannis; Bakas, Athanasios; Kalyvas, Nektarios; Lavdas, Eleftherios; Fountos, George; Kandarakis, Ioannis; Michail, Christos

2017-11-01

The aim of the current work was to assess the resolution properties of a calcium tungstate (CaWO4) screen (screen coating thickness: 50.09 mg/cm2, actual thickness: 167.2 μm) coupled to a high resolution complementary metal oxide semiconductor (CMOS) digital imaging sensor. A 2.7x3.6 cm2 CaWO4 sample was extracted from an Agfa Curix universal screen and was coupled directly with the active area of the active pixel sensor (APS) CMOS sensor. Experiments were performed following the new IEC 62220-1-1:2015 International Standard, using an RQA-5 beam quality. Resolution was assessed in terms of the Modulation Transfer Function (MTF), using the slanted-edge method. The CaWO4/CMOS detector configuration was found with linear response, in the exposure range under investigation. The final MTF was obtained through averaging the oversampled edge spread function (ESF), using a custom-made software developed by our team, according to the IEC 62220-1-1:2015. Considering the renewed interest in calcium tungstate for various applications, along with the resolution results of this work, CaWO4 could be also considered for use in X-ray imaging devices such as charged-coupled devices (CCD) and CMOS.

Ultrasound neuro-modulation chip: activation of sensory neurons in Caenorhabditis elegans by surface acoustic waves.

PubMed

Zhou, Wei; Wang, Jingjing; Wang, Kaiyue; Huang, Bin; Niu, Lili; Li, Fei; Cai, Feiyan; Chen, Yan; Liu, Xin; Zhang, Xiaoyan; Cheng, Hankui; Kang, Lijun; Meng, Long; Zheng, Hairong

2017-05-16

Ultrasound neuro-modulation has gained increasing attention as a non-invasive method. In this paper, we present an ultrasound neuro-modulation chip, capable of initiating reversal behaviour and activating neurons of C. elegans under the stimulation of a single-shot, short-pulsed ultrasound. About 85.29% ± 6.17% of worms respond to the ultrasound stimulation exhibiting reversal behaviour. Furthermore, the worms can adapt to the ultrasound stimulation with a lower acoustic pulse duration of stimulation. In vivo calcium imaging shows that the activity of ASH, a polymodal sensory neuron in C. elegans, can be directly evoked by the ultrasound stimulation. On the other hand, AFD, a thermal sensitive neuron, cannot be activated by the ultrasound stimulation using the same parameter and the temperature elevation during the stimulation process is relatively small. Consistent with the calcium imaging results, the tax-4 mutants, which are insensitive to temperature increase, do not show a significant difference in avoidance probability compared to the wild type. Therefore, the mechanical effects induced by ultrasound are the main reason for neural and behavioural modulation of C. elegans. With the advantages of confined acoustic energy on the surface, compatible with standard calcium imaging, this neuro-modulation chip could be a powerful tool for revealing the molecular mechanisms of ultrasound neuro-modulation.

Superresolution imaging of viral protein trafficking

PubMed Central

Salka, Kyle; Bhuvanendran, Shivaprasad; Yang, David

2015-01-01

The endoplasmic reticulum (ER) membrane is closely apposed to the outer mitochondrial membrane (OMM), which facilitates communication between these organelles. These contacts, known as mitochondria-associated membranes (MAM), facilitate calcium signaling, lipid transfer, as well as antiviral and stress responses. How cellular proteins traffic to the MAM, are distributed therein, and interact with ER and mitochondrial proteins are subject of great interest. The human cytomegalovirus UL37 exon 1 protein or viral mitochondria-localized inhibitor of apoptosis (vMIA) is crucial for viral growth. Upon synthesis at the ER, vMIA traffics to the MAM and OMM, where it reprograms the organization and function of these compartments. vMIA significantly changes the abundance of cellular proteins at the MAM and OMM, including proteins that regulate calcium homeostasis and cell death. Through the use of superresolution imaging, we have shown that vMIA is distributed at the OMM in nanometer scale clusters. This is similar to the clusters reported for the mitochondrial calcium channel, VDAC, as well as electron transport chain, translocase of the OMM complex, and mitochondrial inner membrane organizing system components. Thus, aside from addressing how vMIA targets the MAM and regulates survival of infected cells, biochemical studies and superresolution imaging of vMIA offer insights into the formation, organization, and functioning of MAM. Here, we discuss these insights into trafficking, function, and organization of vMIA at the MAM and OMM and discuss how the use of superresolution imaging is contributing to the study of the formation and trafficking of viruses. PMID:25724304

Multifunctional calcium phosphate nanoparticles for combining near-infrared fluorescence imaging and photodynamic therapy.

PubMed

Haedicke, Katja; Kozlova, Diana; Gräfe, Susanna; Teichgräber, Ulf; Epple, Matthias; Hilger, Ingrid

2015-03-01

Photodynamic therapy (PDT) of tumors causes skin photosensitivity as a result of unspecific accumulation behavior of the photosensitizers. PDT of tumors was improved by calcium phosphate nanoparticles conjugated with (i) Temoporfin as a photosensitizer, (ii) the RGDfK peptide for favored tumor targeting and (iii) the fluorescent dye molecule DY682-NHS for enabling near-infrared fluorescence (NIRF) optical imaging in vivo. The nanoparticles were characterized with regard to size, spectroscopic properties and uptake into CAL-27 cells. The nanoparticles had a hydrodynamic diameter of approximately 200 nm and a zeta potential of around +22mV. Their biodistribution at 24h after injection was investigated via NIRF optical imaging. After treating tumor-bearing CAL-27 mice with nanoparticle-PDT, the therapeutic efficacy was assessed by a fluorescent DY-734-annexin V probe at 2 days and 2 weeks after treatment to detect apoptosis. Additionally, the contrast agent IRDye® 800CW RGD was used to assess tumor vascularization (up to 4 weeks after PDT). After nanoparticle-PDT in mice, apoptosis in the tumor was detected after 2 days. Decreases in tumor vascularization and tumor volume were detected in the next few days. Calcium phosphate nanoparticles can be used as multifunctional tools for NIRF optical imaging, PDT and tumor targeting as they exhibited a high therapeutic efficacy, being capable of inducing apoptosis and destroying tumor vascularization. Copyright © 2014 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Reduction of orthophosphates loss in agricultural soil by nano calcium sulfate.

PubMed

Chen, Dong; Szostak, Paul; Wei, Zongsu; Xiao, Ruiyang

2016-01-01

Nutrient loss from soil, especially phosphorous (P) from farmlands to natural water bodies via surface runoff or infiltration, have caused significant eutrophication problems. This is because dissolved orthophosphates are usually the limiting nutrient for algal blooms. Currently, available techniques to control eutrophication are surprisingly scarce. Calcium sulfate or gypsum is a common soil amendment and has a strong complexation to orthophosphates. The results showed that calcium sulfate reduced the amount of water extractable P (WEP) through soil incubation tests, suggesting less P loss from farmlands. A greater decrease in WEP occurred with a greater dosage of calcium sulfate. Compared to conventional coarse calcium sulfate, nano calcium sulfate further reduced WEP by providing a much greater specific surface area, higher solubility, better contact with the fertilizer and the soil particles, and superior dispersibility. The enhancement of the nano calcium sulfate for WEP reduction is more apparent for a pellet- than a powdered- fertilizer. At the dosage of Ca/P weight ratio of 2.8, the WEP decreased by 31±5% with the nano calcium sulfate compared to 20±5% decrease with the coarse calcium sulfate when the pellet fertilizer was used. Computation of the chemical equilibrium speciation shows that calcium hydroxyapatite has the lowest solubility. However, other mineral phases such as hydroxydicalcium phosphate, dicalcium phosphate dihydrate, octacalcium phosphate, and tricalcium phosphate might form preceding to calcium hydroxyapatite. Since calcium sulfate is the major product of the flue gas desulfurization (FGD) process, this study demonstrates a potential beneficial reuse and reduction of the solid FGD waste. Copyright © 2015 Elsevier B.V. All rights reserved.

Effect of aluminum on bidirectional calcium flux in rat everted intestinal sacs.

PubMed

Adler, A J; Zara, C; Berlyne, G M

1989-09-01

The effect of aluminum on intestinal calcium absorption was determined in male Sprague-Dawley rats using an everted intestinal sac technique. Bidirectional calcium flux in the duodena and ilea of normal rats was assessed by means of dual calcium isotopes. Two micromolar aluminum significantly inhibited net calcium absorption (J net) in the duodenum through suppression of mucosato-serosa flux (J m----s). Jm----s was reduced from 2.21 +/- 0.50 mumol Ca.h-1.g wet wt-1 in controls to 0.93 +/- 0.35 mumol Ca.h-1.g-1 in aluminum exposed sacs, and Jnet was reduced from 1.88 +/- 0.14 mumol Ca.h-1.g-1 to 0.55 +/- 0.41 mumol Ca.h-1.g-1 (P less than 0.001). Serosa-to-mucosa calcium flux (Js----m) was not similarly influenced by aluminum. Inhibition of Jm----s occurred whether aluminum was initially present on the mucosal or serosal side of the duodenal sac and inhibition of Jnet calcium by 2 muM A1 occurred at all ambient concentrations of calcium studied. In the ileum, aluminum had no effect on any component of calcium flux. Aluminum did not induce any suppression of glucose transport in either the duodenum or ileum, suggesting that the effect on calcium transport is relatively specific. These results suggest that aluminum inhibits calcium absorption in the duodenum through an effect on active mucosa-to-serosa transport, but has no effect on ileal calcium absorption, which in the rat is not mediated by an active process.

Characteristics and nutritional value of whole wheat cracker fortified with tuna bone bio-calcium powder.

PubMed

Benjakul, Soottawat; Karnjanapratum, Supatra

2018-09-01

Whole wheat cracker fortified with tuna bone bio-calcium (Bio-Ca) powder was developed as health-promoting food rich in calcium. Fortification with different levels of Bi-Ca, over the range of 0-50% of whole wheat flour (w/w) on quality and sensory properties of crackers, were determined. Color, thickness, weight and textural properties of crackers varied with the different levels of Bio-Ca powder added, but it was found that up to 30% could be added without detrimental effect on sensory properties. Scanning electron microscopic images showed that the developed crackers were less porous and had a denser structure, compared to the control. Based on scanning electron microscopy-energy dispersive X-ray spectroscopic (SEM-EDX), the cracker containing Bio-Ca powder had calcium and phosphorous distribution with higher intensity, compared to the control. The fortified crackers were rich in calcium and phosphorous with higher protein content but lower fat, carbohydrate, cholesterol and energy value, compared to the control. Copyright © 2018 Elsevier Ltd. All rights reserved.

Optimized ratiometric calcium sensors for functional in vivo imaging of neurons and T lymphocytes.

PubMed

Thestrup, Thomas; Litzlbauer, Julia; Bartholomäus, Ingo; Mues, Marsilius; Russo, Luigi; Dana, Hod; Kovalchuk, Yuri; Liang, Yajie; Kalamakis, Georgios; Laukat, Yvonne; Becker, Stefan; Witte, Gregor; Geiger, Anselm; Allen, Taylor; Rome, Lawrence C; Chen, Tsai-Wen; Kim, Douglas S; Garaschuk, Olga; Griesinger, Christian; Griesbeck, Oliver

2014-02-01

The quality of genetically encoded calcium indicators (GECIs) has improved dramatically in recent years, but high-performing ratiometric indicators are still rare. Here we describe a series of fluorescence resonance energy transfer (FRET)-based calcium biosensors with a reduced number of calcium binding sites per sensor. These 'Twitch' sensors are based on the C-terminal domain of Opsanus troponin C. Their FRET responses were optimized by a large-scale functional screen in bacterial colonies, refined by a secondary screen in rat hippocampal neuron cultures. We tested the in vivo performance of the most sensitive variants in the brain and lymph nodes of mice. The sensitivity of the Twitch sensors matched that of synthetic calcium dyes and allowed visualization of tonic action potential firing in neurons and high resolution functional tracking of T lymphocytes. Given their ratiometric readout, their brightness, large dynamic range and linear response properties, Twitch sensors represent versatile tools for neuroscience and immunology.

Fast-Spiking Interneurons Supply Feedforward Control of Bursting, Calcium, and Plasticity for Efficient Learning.

PubMed

Owen, Scott F; Berke, Joshua D; Kreitzer, Anatol C

2018-02-08

Fast-spiking interneurons (FSIs) are a prominent class of forebrain GABAergic cells implicated in two seemingly independent network functions: gain control and network plasticity. Little is known, however, about how these roles interact. Here, we use a combination of cell-type-specific ablation, optogenetics, electrophysiology, imaging, and behavior to describe a unified mechanism by which striatal FSIs control burst firing, calcium influx, and synaptic plasticity in neighboring medium spiny projection neurons (MSNs). In vivo silencing of FSIs increased bursting, calcium transients, and AMPA/NMDA ratios in MSNs. In a motor sequence task, FSI silencing increased the frequency of calcium transients but reduced the specificity with which transients aligned to individual task events. Consistent with this, ablation of FSIs disrupted the acquisition of striatum-dependent egocentric learning strategies. Together, our data support a model in which feedforward inhibition from FSIs temporally restricts MSN bursting and calcium-dependent synaptic plasticity to facilitate striatum-dependent sequence learning. Copyright © 2018 Elsevier Inc. All rights reserved.

Crystallization and preliminary X-ray characterization of the genetically encoded fluorescent calcium indicator protein GCaMP2

PubMed Central

Rodríguez Guilbe, María M.; Alfaro Malavé, Elisa C.; Akerboom, Jasper; Marvin, Jonathan S.; Looger, Loren L.; Schreiter, Eric R.

2008-01-01

Fluorescent proteins and their engineered variants have played an important role in the study of biology. The genetically encoded calcium-indicator protein GCaMP2 comprises a circularly permuted fluorescent protein coupled to the calcium-binding protein calmodulin and a calmodulin target peptide, M13, derived from the intracellular calmodulin target myosin light-chain kinase and has been used to image calcium transients in vivo. To aid rational efforts to engineer improved variants of GCaMP2, this protein was crystallized in the calcium-saturated form. X-ray diffraction data were collected to 2.0 Å resolution. The crystals belong to space group C2, with unit-cell parameters a = 126.1, b = 47.1, c = 68.8 Å, β = 100.5° and one GCaMP2 molecule in the asymmetric unit. The structure was phased by molecular replacement and refinement is currently under way. PMID:18607093

The organic matrix of gallstones

PubMed Central

Sutor, D. June; Wooley, Susan E.

1974-01-01

Dissolution of gallstones consisting of cholesterol, calcium carbonate, or calcium phosphate in different solvents left an amorphous organic gel-like substance (the matrix). Matrix from cholesterol stones could be colourless but was usually orange, yellow, or brown while that from calcium carbonate and calcium phosphate stones was almost invariably coloured black or dark brown. These pigments were also shown to be organic and amorphous. The amount of matrix present and its structure varied with the texture of the crystalline material. Irrespective of their composition, laminated pieces of material yielded compact laminated matrix of the same shape as the original piece and areas of loose crystalline material gave small pieces of non-cohesive matrix. Only large cholesterol crystals which usually radiate from the stone nucleus had no associated matrix. ImagesFig 1Fig 2Fig 3Fig 4Fig 5 PMID:4854981

Evaluation of Changes in Morphology and Function of Human Induced Pluripotent Stem Cell Derived Cardiomyocytes (HiPSC-CMs) Cultured on an Aligned-Nanofiber Cardiac Patch

PubMed Central

Khan, Mahmood; Xu, Yanyi; Hua, Serena; Johnson, Jed; Belevych, Andriy; Janssen, Paul M. L.; Gyorke, Sandor; Guan, Jianjun; Angelos, Mark G.

2015-01-01

Introduction Dilated cardiomyopathy is a major cause of progressive heart failure. Utilization of stem cell therapy offers a potential means of regenerating viable cardiac tissue. However, a major obstacle to stem cell therapy is the delivery and survival of implanted stem cells in the ischemic heart. To address this issue, we have developed a biomimetic aligned nanofibrous cardiac patch and characterized the alignment and function of human inducible pluripotent stem cell derived cardiomyocytes (hiPSC-CMs) cultured on this cardiac patch. This hiPSC-CMs seeded patch was compared with hiPSC-CMs cultured on standard flat cell culture plates. Methods hiPSC-CMs were cultured on; 1) a highly aligned polylactide-co-glycolide (PLGA) nanofiber scaffold (~50 microns thick) and 2) on a standard flat culture plate. Scanning electron microscopy (SEM) was used to determine alignment of PLGA nanofibers and orientation of the cells on the respective surfaces. Analysis of gap junctions (Connexin-43) was performed by confocal imaging in both the groups. Calcium cycling and patch-clamp technique were performed to measure calcium transients and electrical coupling properties of cardiomyocytes. Results SEM demonstrated >90% alignment of the nanofibers in the patch which is similar to the extracellular matrix of decellularized rat myocardium. Confocal imaging of the cardiomyocytes demonstrated symmetrical alignment in the same direction on the aligned nanofiber patch in sharp contrast to the random appearance of cardiomyocytes cultured on a tissue culture plate. The hiPSC-CMs cultured on aligned nanofiber cardiac patches showed more efficient calcium cycling compared with cells cultured on standard flat surface culture plates. Quantification of mRNA with qRT-PCR confirmed that these cardiomyocytes expressed α-actinin, troponin-T and connexin-43 in-vitro. Conclusions Overall, our results demonstrated changes in morphology and function of human induced pluripotent derived cardiomyocytes cultured in an anisotropic environment created by an aligned nanofiber patch. In this environment, these cells better approximate normal cardiac tissue compared with cells cultured on flat surface and can serve as the basis for bioengineering of an implantable cardiac patch. PMID:25993466

Evaluation of Changes in Morphology and Function of Human Induced Pluripotent Stem Cell Derived Cardiomyocytes (HiPSC-CMs) Cultured on an Aligned-Nanofiber Cardiac Patch.

PubMed

Khan, Mahmood; Xu, Yanyi; Hua, Serena; Johnson, Jed; Belevych, Andriy; Janssen, Paul M L; Gyorke, Sandor; Guan, Jianjun; Angelos, Mark G

2015-01-01

Dilated cardiomyopathy is a major cause of progressive heart failure. Utilization of stem cell therapy offers a potential means of regenerating viable cardiac tissue. However, a major obstacle to stem cell therapy is the delivery and survival of implanted stem cells in the ischemic heart. To address this issue, we have developed a biomimetic aligned nanofibrous cardiac patch and characterized the alignment and function of human inducible pluripotent stem cell derived cardiomyocytes (hiPSC-CMs) cultured on this cardiac patch. This hiPSC-CMs seeded patch was compared with hiPSC-CMs cultured on standard flat cell culture plates. hiPSC-CMs were cultured on; 1) a highly aligned polylactide-co-glycolide (PLGA) nanofiber scaffold (~50 microns thick) and 2) on a standard flat culture plate. Scanning electron microscopy (SEM) was used to determine alignment of PLGA nanofibers and orientation of the cells on the respective surfaces. Analysis of gap junctions (Connexin-43) was performed by confocal imaging in both the groups. Calcium cycling and patch-clamp technique were performed to measure calcium transients and electrical coupling properties of cardiomyocytes. SEM demonstrated >90% alignment of the nanofibers in the patch which is similar to the extracellular matrix of decellularized rat myocardium. Confocal imaging of the cardiomyocytes demonstrated symmetrical alignment in the same direction on the aligned nanofiber patch in sharp contrast to the random appearance of cardiomyocytes cultured on a tissue culture plate. The hiPSC-CMs cultured on aligned nanofiber cardiac patches showed more efficient calcium cycling compared with cells cultured on standard flat surface culture plates. Quantification of mRNA with qRT-PCR confirmed that these cardiomyocytes expressed α-actinin, troponin-T and connexin-43 in-vitro. Overall, our results demonstrated changes in morphology and function of human induced pluripotent derived cardiomyocytes cultured in an anisotropic environment created by an aligned nanofiber patch. In this environment, these cells better approximate normal cardiac tissue compared with cells cultured on flat surface and can serve as the basis for bioengineering of an implantable cardiac patch.

An integrated platform for simultaneous multi-well field potential recording and Fura-2-based calcium transient ratiometry in human induced pluripotent stem cell (hiPSC)-derived cardiomyocytes.

PubMed

Rast, Georg; Weber, Jürgen; Disch, Christoph; Schuck, Elmar; Ittrich, Carina; Guth, Brian D

2015-01-01

Human induced pluripotent stem cell-derived cardiomyocytes are available from various sources and they are being evaluated for safety testing. Several platforms are available offering different assay principles and read-out parameters: patch-clamp and field potential recording, imaging or photometry, impedance measurement, and recording of contractile force. Routine use will establish which assay principle and which parameters best serve the intended purpose. We introduce a combination of field potential recording and calcium ratiometry from spontaneously beating cardiomyocytes as a novel assay providing a complementary read-out parameter set. Field potential recording is performed using a commercial multi-well multi-electrode array platform. Calcium ratiometry is performed using a fiber optic illumination and silicon avalanche photodetectors. Data condensation and statistical analysis are designed to enable statistical inference of differences and equivalence with regard to a solvent control. Simultaneous recording of field potentials and calcium transients from spontaneously beating monolayers was done in a nine-well format. Calcium channel blockers (e.g. nifedipine) and a blocker of calcium store release (ryanodine) can be recognized and discriminated based on the calcium transient signal. An agonist of L-type calcium channels, FPL 64176, increased and prolonged the calcium transient, whereas BAY K 8644, another L-type calcium channel agonist, had no effect. Both FPL 64176 and various calcium channel antagonists have chronotropic effects, which can be discriminated from typical "chronotropic" compounds, like (±)isoprenaline (positive) and arecaidine propargyl ester (negative), based on their effects on the calcium transient. Despite technical limitations in temporal resolution and exact matching of composite calcium transient with the field potential of a subset of cells, the combined recording platform enables a refined interpretation of the field potential recording and a more reliable identification of drug effects on calcium handling. Copyright © 2015 Elsevier Inc. All rights reserved.

Calcium Kinetics During Space Flight

NASA Technical Reports Server (NTRS)

Smith, Scott M.; Wastney, Meryl E.; OBrien, Kimberly O.; Lane, Helen W.

1999-01-01

Bone loss is one of the most detrimental effects of space flight, threatening to limit the duration of human space missions. The ability to understand and counteract this loss will be critical for crew health and safety during and after extended-duration missions. The hypotheses to be tested in this project are that space flight alters calcium homeostasis and bone mineral metabolism, and that calcium homeostasis and bone mineral metabolism will return to baseline within days to weeks of return to Earth. These hypotheses will be evidenced by elevated rates of bone mineral resorption and decreased bone mineral deposition, decreased absorption of dietary calcium, altered calcitropic endocrine profiles, elevated excretion of calcium in urine and feces, and elevated excretion of markers of bone resorption. The second hypothesis will be evidenced by return of indices of calcium homeostasis and bone metabolism to preflight levels within days to weeks of return to Earth. Studies will be conducted on International Space Station astronauts before, during, and after extended-duration flights. Measurements of calcium kinetics, bone mass, and endocrine/biochemical markers of bone and calcium homeostasis will be conducted. Kinetic studies utilizing dual isotope tracer kinetic studies and mathematical modeling techniques will allow for determination of bone calcium deposition, bone calcium resorption, dietary calcium absorption and calcium excretion (both urinary and endogenous fecal excretion). These studies will build upon preliminary work conducted on the Russian Mir space station. The results from this project will be critical for clarifying how microgravity affects bone and calcium homeostasis, and will provide an important control point for assessment of countermeasure efficacy. These results are expected to aid in developing countermeasures for bone loss, both for space crews and for individuals on Earth who have metabolic bone diseases.

Generative Adversarial Networks for Noise Reduction in Low-Dose CT.

PubMed

Wolterink, Jelmer M; Leiner, Tim; Viergever, Max A; Isgum, Ivana

2017-12-01

Noise is inherent to low-dose CT acquisition. We propose to train a convolutional neural network (CNN) jointly with an adversarial CNN to estimate routine-dose CT images from low-dose CT images and hence reduce noise. A generator CNN was trained to transform low-dose CT images into routine-dose CT images using voxelwise loss minimization. An adversarial discriminator CNN was simultaneously trained to distinguish the output of the generator from routine-dose CT images. The performance of this discriminator was used as an adversarial loss for the generator. Experiments were performed using CT images of an anthropomorphic phantom containing calcium inserts, as well as patient non-contrast-enhanced cardiac CT images. The phantom and patients were scanned at 20% and 100% routine clinical dose. Three training strategies were compared: the first used only voxelwise loss, the second combined voxelwise loss and adversarial loss, and the third used only adversarial loss. The results showed that training with only voxelwise loss resulted in the highest peak signal-to-noise ratio with respect to reference routine-dose images. However, CNNs trained with adversarial loss captured image statistics of routine-dose images better. Noise reduction improved quantification of low-density calcified inserts in phantom CT images and allowed coronary calcium scoring in low-dose patient CT images with high noise levels. Testing took less than 10 s per CT volume. CNN-based low-dose CT noise reduction in the image domain is feasible. Training with an adversarial network improves the CNNs ability to generate images with an appearance similar to that of reference routine-dose CT images.

Spectroscopic studies of gel grown zinc doped calcium hydrogen phosphate dihydrate crystals

NASA Astrophysics Data System (ADS)

Suryawanshi, V. B.; Chaudhari, R. T.

2018-05-01

The influence of zinc doping on the gel grown calcium hydrogen phosphate dihydrate crystals was studied using the spectroscopic techniques, which included SEM, FTIR and EDAX. It was found that, zinc ions transform the morphology of brushite crystals from rectangular plate shaped crystals to branching microcrystal patterns. However in FT-IR spectroscopy, as compared to undoped brushite crystals few vibrations were shifted to higher value. The observed changes in the vibrations were due to the impact of zinc ions. EDAX techniques is use to determine the percentage composition of elements present in the doped crystals. It revealed that the sample was of a mixed composition.

Evaluation of the phase properties of hydrating cement composite using simulated nanoindentation technique

NASA Astrophysics Data System (ADS)

Gautham, S.; Sindu, B. S.; Sasmal, Saptarshi

2017-10-01

Properties and distribution of the products formed during the hydration of cementitious composite at the microlevel are investigated using a nanoindentation technique. First, numerical nanoindentation using nonlinear contact mechanics is carried out on three different phase compositions of cement paste, viz. mono-phase Tri-calcium Silicate (C3S), Di-calcium Silicate (C2S) and Calcium-Silicate-Hydrate (CSH) individually), bi-phase (C3S-CSH, C2S-CSH) and multi-phase (more than 10 individual phases including water pores). To reflect the multi-phase characteristics of hydrating cement composite, a discretized multi-phase microstructural model of cement composite during the progression of hydration is developed. Further, a grid indentation technique for simulated nanoindentation is established, and employed to evaluate the mechanical characteristics of the hydrated multi-phase cement paste. The properties obtained from the numerical studies are compared with those obtained from experimental grid nanoindentation. The influence of composition and distribution of individual phase properties on the properties obtained from indentation are closely investigated. The study paves the way to establishing the procedure for simulated grid nanoindentation to evaluate the mechanical properties of heterogeneous composites, and facilitates the design of experimental nanoindentation.

Synthesis of Brushite Particles in Reverse Microemulsions of the Biosurfactant Surfactin

PubMed Central

Maity, Jyoti Prakash; Lin, Tz-Jiun; Cheng, Henry Pai-Heng; Chen, Chien-Yen; Reddy, A. Satyanarayana; Atla, Shashi B.; Chang, Young-Fo; Chen, Hau-Ren; Chen, Chien-Cheng

2011-01-01

In this study the “green chemistry” use of the biosurfactant surfactin for the synthesis of calcium phosphate using the reverse microemulsion technique was demonstrated. Calcium phosphates are bioactive materials that are a major constituent of human teeth and bone tissue. A reverse microemulsion technique with surfactin was used to produce nanocrystalline brushite particles. Structural diversity (analyzed by SEM and TEM) resulted from different water to surfactin ratios (W/S; 250, 500, 1000 and 40,000). The particle sizes were found to be in the 16–200 nm range. Morphological variety was observed in the as-synthesized microemulsions, which consisted of nanospheres (~16 nm in diameter) and needle-like (8–14 nm in diameter and 80–100 nm in length) noncalcinated particles. However, the calcinated products included nanospheres (50–200 nm in diameter), oval (~300 nm in diameter) and nanorod (200–400 nm in length) particles. FTIR and XRD analysis confirmed the formation of brushite nanoparticles in the as-synthesized products, while calcium pyrophosphate was produced after calcination. These results indicate that the reverse microemulsion technique using surfactin is a green process suitable for the synthesis of nanoparticles. PMID:21747709

Arterial Smooth Muscle Mitochondria Amplify Hydrogen Peroxide Microdomains Functionally Coupled to L-Type Calcium Channels

PubMed Central

Chaplin, Nathan L.; Nieves-Cintrón, Madeline; Fresquez, Adriana M.; Navedo, Manuel F.; Amberg, Gregory C.

2015-01-01

Rationale Mitochondria are key integrators of convergent intracellular signaling pathways. Two important second messengers modulated by mitochondria are calcium and reactive oxygen species. To date, coherent mechanisms describing mitochondrial integration of calcium and oxidative signaling in arterial smooth muscle are incomplete. Objective To address and add clarity to this issue we tested the hypothesis that mitochondria regulate subplasmalemmal calcium and hydrogen peroxide microdomain signaling in cerebral arterial smooth muscle. Methods and Results Using an image-based approach we investigated the impact of mitochondrial regulation of L-type calcium channels on subcellular calcium and ROS signaling microdomains in isolated arterial smooth muscle cells. Our single cell observations were then related experimentally to intact arterial segments and to living animals. We found that subplasmalemmal mitochondrial amplification of hydrogen peroxide microdomain signaling stimulates L-type calcium channels and that this mechanism strongly impacts the functional capacity of the vasoconstrictor angiotensin II. Importantly, we also found that disrupting this mitochondrial amplification mechanism in vivo normalized arterial function and attenuated the hypertensive response to systemic endothelial dysfunction. Conclusions From these observations we conclude that mitochondrial amplification of subplasmalemmal calcium and hydrogen peroxide microdomain signaling is a fundamental mechanism regulating arterial smooth muscle function. As the principle components involved are fairly ubiquitous and positioning of mitochondria near the plasma membrane is not restricted to arterial smooth muscle, this mechanism could occur in many cell types and contribute to pathological elevations of intracellular calcium and increased oxidative stress associated with many diseases. PMID:26390880

Effect of particle size of calcium phosphate based bioceramic drug delivery carrier on the release kinetics of ciprofloxacin hydrochloride: an in vitro study

NASA Astrophysics Data System (ADS)

Sasikumar, Swamiappan

2013-09-01

Hydroxyapatite (HAP) is the constituent of calcium phosphate based bone cement and it is extensively used as a bone substitute and drug delivery vehicle in various biomedical applications. In the present study we investigated the release kinetics of ciprofloxacin loaded HAP and analyzed its ability to function as a targeted and sustained release drug carrier. Synthesis of HAP was carried out by combustion method using tartaric acid as a fuel and nitric acid as an oxidizer. Powder XRD and FTIR techniques were employed to characterize the phase purity of the drug carrier and to verify the chemical interaction between the drug and carrier. The synthesized powders were sieve separated to make two different drug carriers with different particle sizes and the surface topography of the pellets of the drug carrier was imaged by AFM. Surface area and porosity of the drug carrier was carried out using surface area analyzer. The in-vitro drug release kinetics was performed in simulated body fluid, at 37.3°C. The amount of ciprofloxacin released is measured using UV-visible spectroscopy following the characteristic λ max of 278 nm. The release saturates around 450 h which indicates that it can be used as a targeted and sustained release carrier for bone infections.

Osteocyte calcium signals encode strain magnitude and loading frequency in vivo.

PubMed

Lewis, Karl J; Frikha-Benayed, Dorra; Louie, Joyce; Stephen, Samuel; Spray, David C; Thi, Mia M; Seref-Ferlengez, Zeynep; Majeska, Robert J; Weinbaum, Sheldon; Schaffler, Mitchell B

2017-10-31

Osteocytes are considered to be the major mechanosensory cells of bone, but how osteocytes in vivo process, perceive, and respond to mechanical loading remains poorly understood. Intracellular calcium (Ca 2+ ) signaling resulting from mechanical stimulation has been widely studied in osteocytes in vitro and in bone explants, but has yet to be examined in vivo. This is achieved herein by using a three-point bending device which is capable of delivering well-defined mechanical loads to metatarsal bones of living mice while simultaneously monitoring the intracellular Ca 2+ responses of individual osteocytes by using a genetically encoded fluorescent Ca 2+ indicator. Osteocyte responses are imaged by using multiphoton fluorescence microscopy. We investigated the in vivo responses of osteocytes to strains ranging from 250 to 3,000 [Formula: see text] and frequencies from 0.5 to 2 Hz, which are characteristic of physiological conditions reported for bone. At all loading frequencies examined, the number of responding osteocytes increased strongly with applied strain magnitude. However, Ca 2+ intensity within responding osteocytes did not change significantly with physiological loading magnitudes. Our studies offer a glimpse into how these critical bone cells respond to mechanical load in vivo, as well as provide a technique to determine how the cells encode magnitude and frequency of loading. Published under the PNAS license.

Simulation of heart infarction by laser microbeams and induction of arrhythmias by optical tweezers

NASA Astrophysics Data System (ADS)

Perner, Birgit; Monajembashi, Shamci; Rapp, Alexander; Wollweber, Leo; Greulich, Karl Otto

2004-10-01

Laser microbeam and optical tweezers were used for micromanipulation of a heart tissue model consisting of embryonic chicken cardiomyocytes and bibroblasts. Using the laser microbeam a would was created, i.e. a sort of artificial heart infarction was generated. The first steps of wound repair were observed by live cell imaging. A complete filling of teh would primarily by migrating fibroblasts but not by cardiomyocytes was detected 18 hours after wounding. In another set of experiments erythrocyte mediated force application (EMFA) by optical tweezers was applied for optomechanical manipulatoin of cardiomyocytes and fibroblasts. Here we demonstrate induction of dramatic distrubances of calcium waves in a group of synchronously beating cardiomyocytes by an optomechanical input that results in cellular deformation. Surprisingly, it was found that putatively non-excitable fibroblasts respond to this mechanical stress with calcium oscillations. The results reported here indicate that the induction of artificial heart infarction can provide insights into healing processes after mycardial injury. EMFA is capable to examine effects of myocardial overload and to provide important information about processes triggered by mechanical stress on the level of single or very few cells. As a perspective, the preseneted techniques may be used to study the influence of drugs on wound healing and coordination of beating in the heart.

Back-Propagation of Physiological Action Potential Output in Dendrites of Slender-Tufted L5A Pyramidal Neurons

PubMed Central

Grewe, Benjamin F.; Bonnan, Audrey; Frick, Andreas

2009-01-01

Pyramidal neurons of layer 5A are a major neocortical output type and clearly distinguished from layer 5B pyramidal neurons with respect to morphology, in vivo firing patterns, and connectivity; yet knowledge of their dendritic properties is scant. We used a combination of whole-cell recordings and Ca2+ imaging techniques in vitro to explore the specific dendritic signaling role of physiological action potential patterns recorded in vivo in layer 5A pyramidal neurons of the whisker-related ‘barrel cortex’. Our data provide evidence that the temporal structure of physiological action potential patterns is crucial for an effective invasion of the main apical dendrites up to the major branch point. Both the critical frequency enabling action potential trains to invade efficiently and the dendritic calcium profile changed during postnatal development. In contrast to the main apical dendrite, the more passive properties of the short basal and apical tuft dendrites prevented an efficient back-propagation. Various Ca2+ channel types contributed to the enhanced calcium signals during high-frequency firing activity, whereas A-type K+ and BKCa channels strongly suppressed it. Our data support models in which the interaction of synaptic input with action potential output is a function of the timing, rate and pattern of action potentials, and dendritic location. PMID:20508744

Automated quantification of neuronal networks and single-cell calcium dynamics using calcium imaging

PubMed Central

Patel, Tapan P.; Man, Karen; Firestein, Bonnie L.; Meaney, David F.

2017-01-01

Background Recent advances in genetically engineered calcium and membrane potential indicators provide the potential to estimate the activation dynamics of individual neurons within larger, mesoscale networks (100s–1000 +neurons). However, a fully integrated automated workflow for the analysis and visualization of neural microcircuits from high speed fluorescence imaging data is lacking. New method Here we introduce FluoroSNNAP, Fluorescence Single Neuron and Network Analysis Package. FluoroSNNAP is an open-source, interactive software developed in MATLAB for automated quantification of numerous biologically relevant features of both the calcium dynamics of single-cells and network activity patterns. FluoroSNNAP integrates and improves upon existing tools for spike detection, synchronization analysis, and inference of functional connectivity, making it most useful to experimentalists with little or no programming knowledge. Results We apply FluoroSNNAP to characterize the activity patterns of neuronal microcircuits undergoing developmental maturation in vitro. Separately, we highlight the utility of single-cell analysis for phenotyping a mixed population of neurons expressing a human mutant variant of the microtubule associated protein tau and wild-type tau. Comparison with existing method(s) We show the performance of semi-automated cell segmentation using spatiotemporal independent component analysis and significant improvement in detecting calcium transients using a template-based algorithm in comparison to peak-based or wavelet-based detection methods. Our software further enables automated analysis of microcircuits, which is an improvement over existing methods. Conclusions We expect the dissemination of this software will facilitate a comprehensive analysis of neuronal networks, promoting the rapid interrogation of circuits in health and disease. PMID:25629800

Na+/H+ exchanger 3 inhibitor diminishes the amino-acid-enhanced transepithelial calcium transport across the rat duodenum.

PubMed

Thammayon, Nithipak; Wongdee, Kannikar; Lertsuwan, Kornkamon; Suntornsaratoon, Panan; Thongbunchoo, Jirawan; Krishnamra, Nateetip; Charoenphandhu, Narattaphol

2017-04-01

Na + /H + exchanger (NHE)-3 is important for intestinal absorption of nutrients and minerals, including calcium. The previous investigations have shown that the intestinal calcium absorption is also dependent on luminal nutrients, but whether aliphatic amino acids and glucose, which are abundant in the luminal fluid during a meal, similarly enhance calcium transport remains elusive. Herein, we used the in vitro Ussing chamber technique to determine epithelial electrical parameters, i.e., potential difference (PD), short-circuit current (Isc), and transepithelial resistance, as well as 45 Ca flux in the rat duodenum directly exposed on the mucosal side to glucose or various amino acids. We found that mucosal glucose exposure led to the enhanced calcium transport, PD, and Isc, all of which were insensitive to NHE3 inhibitor (100 nM tenapanor). In the absence of mucosal glucose, several amino acids (12 mM in the mucosal side), i.e., alanine, isoleucine, leucine, proline, and hydroxyproline, markedly increased the duodenal calcium transport. An inhibitor for NHE3 exposure on the mucosal side completely abolished proline- and leucine-enhanced calcium transport, but not transepithelial transport of both amino acids themselves. In conclusion, glucose and certain amino acids in the mucosal side were potent stimulators of the duodenal calcium absorption, but only amino-acid-enhanced calcium transport was NHE3-dependent.

Calcium supplementation improves clinical outcome in intensive care unit patients: a propensity score matched analysis of a large clinical database MIMIC-II.

PubMed

Zhang, Zhongheng; Chen, Kun; Ni, Hongying

2015-01-01

Observational studies have linked hypocalcemia with adverse clinical outcome in critically ill patients. However, calcium supplementation has never been formally investigated for its beneficial effect in critically ill patients. To investigate whether calcium supplementation can improve 28-day survival in adult critically ill patients. Secondary analysis of a large clinical database consisting over 30,000 critical ill patients was performed. Multivariable analysis was performed to examine the independent association of calcium supplementation and 28-day morality. Furthermore, propensity score matching technique was employed to investigate the role of calcium supplementation in improving survival. none. Primary outcome was the 28-day mortality. 90-day mortality was used as secondary outcome. A total of 32,551 adult patients, including 28,062 survivors and 4489 non-survivors (28-day mortality rate: 13.8 %) were included. Calcium supplementation was independently associated with improved 28-day mortality after adjusting for confounding variables (hazard ratio: 0.51; 95 % CI 0.47-0.56). Propensity score matching was performed and the after-matching cohort showed well balanced covariates. The results showed that calcium supplementation was associated with improved 28- and 90-day mortality (p < 0.05 for both Log-rank test). In adult critically ill patients, calcium supplementation during their ICU stay improved 28-day survival. This finding supports the use of calcium supplementation in critically ill patients.

Heat stress responses modulate calcium regulations and electrophysiological characteristics in atrial myocytes.

PubMed

Chen, Yao-Chang; Kao, Yu-Hsun; Huang, Chun-Feng; Cheng, Chen-Chuan; Chen, Yi-Jen; Chen, Shih-Ann

2010-04-01

Heat stress-induced responses change the ionic currents and calcium homeostasis. However, the molecular insights into the heat stress responses on calcium homeostasis remain unclear. The purposes of this study were to examine the mechanisms of heat stress responses on calcium handling and electrophysiological characteristics in atrial myocytes. We used indo-1 fluorimetric ratio technique and whole-cell patch clamp to investigate the intracellular calcium, action potentials, and ionic currents in isolated rabbit single atrial cardiomyocytes with or without (control) exposure to heat stress (43 degrees C, 15 min) 5+/-1 h before experiments. The expressions of sarcoplasmic reticulum ATPase (SERCA2a), and Na(+)-Ca(2+) exchanger (NCX) in the control and heat stress-treated atrial myocytes were evaluated by Western blot and real-time PCR. As compared with control myocytes, the heat stress-treated myocytes had larger sarcoplasmic reticulum calcium content and larger intracellular calcium transient with a shorter decay portion. Heat stress-treated myocytes also had larger L-type calcium currents, transient outward potassium currents, but smaller NCX currents. Heat stress responses increased the protein expressions, SERCA2a, NCX, and heat shock protein. However, heat stress responses did not change the RNA expression of SERCA2a and NCX. In conclusion, heat stress responses change calcium handling through protein but not RNA regulation. Copyright (c) 2009 Elsevier Ltd. All rights reserved.

[Application of T grain technique to the diagnosis of lung disease and analysis of its image quality].

PubMed

Liu, Xin-jun; Liu, Pei-cheng; Cai, Pei; Zhang, Dun; Yao, Shi-sheng

2003-06-01

To compare the image quality of T grain green sensitive film (TML-1) and Lanex Gd(2)O(2)S rare earth intensifying screen with that of XK-1 blue sensitive film and calcium tungstate (CaWO(4)) intensifying screen, and to study the application of T grain technic to the diagnosis of lung diseases. 160 coal miners were randomly selected to take both TML-1 and XK-1 chest film of high kV radiographs at the same time. Silver halide granule, fluorescence of intensifying screen, radiographic parameters, the density at different points in the lung and chest radiographs were observed. Silver grains in TML-1 film were more homogeneous in distribution than in XK-1 film. Luminous intensity of Lanex Gd(2)O(2)S rare earth intensifying screen was brighter than CaWO(4) intensifying screen in the same exposure. The exposure doses of TML-1 film was reduced to one third of XK-1 film. The density of chest radiographs was 0.24 to 2.74 in TML-1 film, and 0.30 to 2.60 in XK-1 film. There were greater exposure latitude and more informations in TML-1 film. By apertured-disc observation, the fine structure of lung in TML-1 film was clearer than in XK-1 film, the shape was more concrete and reliable, visualizability was stronger. T grain technique may obviously improve the clearness and resolution of image, and enhance the transmission of information, as well as increase the diagnostic informations.

Endotrophic Calcium, Strontium, and Barium Spores of Bacillus megaterium and Bacillus cereus1

PubMed Central

Foerster, Harold F.; Foster, J. W.

1966-01-01

Foerster, Harold F. (The University of Texas, Austin), and J. W. Foster. Endotrophic calcium, strontium, and barium spores of Bacillus megaterium and Bacillus cereus. J. Bacteriol. 91:1333–1345. 1966.—Spores were produced by washed vegetative cells suspended in deionized water supplemented with CaCl2, SrCl2, or BaCl2. Normal, refractile spores were produced in each case; a portion of the barium spores lost refractility and darkened. Thin-section electron micrographs revealed no apparent anatomical differences among the three types of spores. Analyses revealed that the different spore types were enriched specifically in the metal to which they were exposed during sporogenesis. The calcium content of the strontium and the barium spores was very small. From binary equimolar mixtures of the metal salts, endotrophic spores accumulated both metals to nearly the same extent. Viability of the barium spores was considerably less than that of the other two types. Strontium and barium spores were heat-resistant; however, calcium was essential for maximal heat resistance. Significant differences existed in the rates of germination; calcium spores germinated fastest, strontium spores were slower, and barium spores were slowest. Calcium-barium and calcium-strontium spores germinated readily. Endotrophic calcium and strontium spores germinated without the prior heat activation essential for growth spores. Chemical germination of the different metal-type spores with n-dodecylamine took place at the same relative rates as physiological germination. Heat-induced release of dipicolinic acid occurred much faster with barium and strontium spores than with calcium spores. The washed “coat fraction” from disrupted spores contained little of the spore calcium but most of the spore barium. The metal in this fraction was released by dilute acid. The demineralized coats reabsorbed calcium and barium at neutral pH. Images PMID:4956334

Overexpression of human mutated G93A SOD1 changes dynamics of the ER mitochondria calcium cycle specifically in mouse embryonic motor neurons.

PubMed

Lautenschläger, Janin; Prell, Tino; Ruhmer, Julia; Weidemann, Lisa; Witte, Otto W; Grosskreutz, Julian

2013-09-01

Motor neurons vulnerable to the rapidly progressive deadly neurodegenerative disease amyotrophic lateral sclerosis (ALS) inherently express low amounts of calcium binding proteins (CaBP), likely to allow physiological motor neuron firing frequency modulation. At the same time motor neurons are susceptible to AMPA receptor mediated excitotoxicity and internal calcium deregulation which is not fully understood. We analysed ER mitochondria calcium cycle (ERMCC) dynamics with subsecond resolution in G93A hSOD1 overexpressing motor neurons as a model of ALS using fluorescent calcium imaging. When comparing vulnerable motor neurons and non-motor neurons from G93A hSOD1 mice and their non-transgenic littermates, we found a decelerated cytosolic calcium clearance in the presence of G93A hSOD1. While both non-transgenic as well as G93A hSOD1 motor neurons displayed large mitochondrial calcium uptake by the mitochondrial uniporter (mUP), the mitochondrial calcium extrusion system was altered in the presence of G93A hSOD1. In addition, ER calcium uptake by the sarco-/endoplasmic reticulum ATPase (SERCA) was increased in G93A hSOD1 motor neurons. In survival assays, blocking the mitochondrial sodium calcium exchanger (mNCE) by CGP37157 as well as inhibiting SERCA by cyclopiazonic acid showed protective effects against kainate induced excitotoxicity. Thus, our study shows for the first time that the functional consequence of G93A hSOD1 overexpression in intact motor neurons is indeed a disturbance of the ER mitochondria calcium cycle, and identified two promising targets for therapeutic intervention in the pathology of ALS. Copyright © 2013 Elsevier Inc. All rights reserved.

WE-FG-207B-12: Quantitative Evaluation of a Spectral CT Scanner in a Phantom Study: Results of Spectral Reconstructions

DOE Office of Scientific and Technical Information (OSTI.GOV)

Duan, X; Arbique, G; Guild, J

Purpose: To evaluate the quantitative image quality of spectral reconstructions of phantom data from a spectral CT scanner. Methods: The spectral CT scanner (IQon Spectral CT, Philips Healthcare) is equipped with a dual-layer detector and generates conventional 80-140 kVp images and variety of spectral reconstructions, e.g., virtual monochromatic (VM) images, virtual non-contrast (VNC) images, iodine maps, and effective atomic number (Z) images. A cylindrical solid water phantom (Gammex 472, 33 cm diameter and 5 cm thick) with iodine (2.0-20.0 mg I/ml) and calcium (50-600 mg/ml) rod inserts was scanned at 120 kVp and 27 mGy CTDIvol. Spectral reconstructions were evaluatedmore » by comparing image measurements with theoretical values calculated from nominal rod compositions provided by the phantom manufacturer. The theoretical VNC was calculated using water and iodine basis material decomposition, and the theoretical Z was calculated using two common methods, the chemical formula method (Z1) and the dual-energy ratio method (Z2). Results: Beam-hardening-like artifacts between high-attenuation calcium rods (≥300 mg/ml, >800 HU) influenced quantitative measurements, so the quantitative analysis was only performed on iodine rods using the images from the scan with all the calcium rods removed. The CT numbers of the iodine rods in the VM images (50∼150 keV) were close to theoretical values with average difference of 2.4±6.9 HU. Compared with theoretical values, the average difference for iodine concentration, VNC CT number and effective Z of iodine rods were −0.10±0.38 mg/ml, −0.1±8.2 HU, 0.25±0.06 (Z1) and −0.23±0.07 (Z2). Conclusion: The results indicate that the spectral CT scanner generates quantitatively accurate spectral reconstructions at clinically relevant iodine concentrations. Beam-hardening-like artifacts still exist when high-attenuation objects are present and their impact on patient images needs further investigation. YY is an employee of Philips Healthcare.« less

Alpha Hemolysin Induces an Increase of Erythrocytes Calcium: A FLIM 2-Photon Phasor Analysis Approach

PubMed Central

Sanchez, Susana; Bakás, Laura; Gratton, Enrico; Herlax, Vanesa

2011-01-01

α-hemolysin (HlyA) from Escherichia coli is considered as the prototype of a family of toxins called RTX (repeat in toxin), a group of proteins that share genetic and structural features. HlyA is an important virulence factor in E. coli extraintestinal infections, such as meningitis, septicemia and urinary infections. High concentrations of the toxin cause the lysis of several cells such as erythrocytes, granulocytes, monocytes, endothelial and renal epithelial cells of different species. At low concentrations it induces the production of cytokines and apoptosis. Since many of the subcytolytic effects in other cells have been reported to be triggered by the increase of intracellular calcium, we followed the calcium concentration inside the erythrocytes while incubating with sublytic concentrations of HlyA. Calcium concentration was monitored using the calcium indicator Green 1, 2-photon excitation, and fluorescence lifetime imaging microscopy (FLIM). Data were analyzed using the phasor representation. In this report, we present evidence that, at sublytic concentrations, HlyA induces an increase of calcium concentration in rabbit erythrocytes in the first 10 s. Results are discussed in relation to the difficulties of measuring calcium concentrations in erythrocytes where hemoglobin is present, the contribution of the background and the heterogeneity of the response observed in individual cells. PMID:21698153

Porcine malignant hyperthermia susceptibility: hypersensitive calcium-release mechanism of skeletal muscle sarcoplasmic reticulum.

PubMed Central

O'Brien, P J

1986-01-01

This study tested the hypothesis that calcium-release from sarcoplasmic reticulum isolated from malignant hyperthermia swine had abnormal concentration-dependency on release modulators. Halothane stimulated half-maximal calcium-release at similar concentrations for malignant hyperthermia and control sarcoplasmic reticulum (0.10 +/- 0.04 mM). However, concentrations causing half-maximal calcium-release were lower for malignant hyperthermia sarcoplasmic reticulum (P less than 0.001) by an order of magnitude for Ca2+ (28.1 +/- 8.3 versus 1.23 +/- 0.45 nM), adenosine triphosphate (0.33 +/- 0.09 versus 0.023 +/- 0.014 mM) and caffeine (7.79 +/- 1.56 versus 0.80 +/- 0.44 mM). Half-maximal inhibition by Mg2+ occurred at threefold higher concentrations for malignant hyperthermia sarcoplasmic reticulum (0.23 +/- 0.02 versus 0.78 +/- 0.17 mM). The Ca2+-sensitivity curves for calcium-release by sarcoplasmic reticulum isolated from heterozygotes for the malignant hyperthermia-defect were indistinguishable from the averages of the curves for controls and malignant hyperthermia-homozygotes. Results of this study suggest that malignant hyperthermia is initiated due to a hypersensitive calcium-release mechanism which is inherited in an autosomal, codominant pattern and may be diagnosed using calcium-release sensitivity-tests on isolated sarcoplasmic reticulum. Images Fig. 1. PMID:3742367

Randomised controlled trial of a synthetic triglyceride milk formula for preterm infants

PubMed Central

Lucas, A; Quinlan, P; Abrams, S; Ryan, S; Meah, S; Lucas, P

1997-01-01

AIMS—To test whether use of infant formula containing synthetic structured triglycerides results in: (i) increased palmitate absorption; (ii) increased total fat absorption; (iii) reduction in calcium soap formation in the gut; and hence (iv) increased calcium absorption.
METHODS—A randomised study was made of 24 infants comparing three formulas, one containing the synthetic fat Betapol with 74% of palmitate in the 2-position, which was substantially higher than in the two comparison diets (8.4% and 28%). The hypothesised outcomes were tested using balance studies, detailed chemical analysis of stool specimens and dual calcium isotope tracers (44calcium orally and 46calcium intravenously). 
RESULTS—Three of the four hypotheses were confirmed: use of a formula rich in 2-position palmitate (i) improved palmitate (16:0) and also (18:0) absorption; (ii) reduced the formation of insoluble calcium soaps in the stool; and (iii) improved calcium absorption, determined by the dual tracer technique from 42 (SE 3)% to 57 (7)%.
CONCLUSION—Synthetic triglycerides that mimic the stereoisometric structure of those in breast milk may have a valuable role in the design of formulas used for preterm infants in neonatal intensive care units.

 PMID:9462186

Antimicrobial activity of calcium hydroxide and chlorhexidine on intratubular Candida albicans

PubMed Central

Jacques Rezende Delgado, Ronan; Helena Gasparoto, Thaís; Renata Sipert, Carla; Ramos Pinheiro, Claudia; Gomes de Moraes, Ivaldo; Brandão Garcia, Roberto; Antônio Hungaro Duarte, Marco; Monteiro Bramante, Clóvis; Aparecido Torres, Sérgio; Pompermaier Garlet, Gustavo; Paula Campanelli, Ana; Bernardineli, Norberti

2013-01-01

This study investigated the efficacy of calcium hydroxide and chlorhexidine gel for the elimination of intratubular Candida albicans (C. albicans). Human single-rooted teeth contaminated with C. albicans were treated with calcium hydroxide, 2% chlorhexidine gel, calcium hydroxide plus 2% chlorhexidine gel, or saline (0.9% sodium chloride) as a positive control. The samples obtained at depths of 0–100 and 100–200 µm from the root canal system were analyzed for C. albicans load by counting the number of colony forming units and for the percentage of viable C. albicans using fluorescence microscopy. First, the antimicrobial activity of calcium hydroxide and the 2% chlorhexidine gel was evaluated by counting the number of colony forming units. After 14 days of intracanal medication, there was a significant decrease in the number of C. albicans colony forming units at a depth of 0–100 µm with chlorhexidine treatment either with or without calcium hydroxide compared with the calcium hydroxide only treatment. However, there were no differences in the number of colony forming units at the 100–200 µm depth for any of the medications investigated. C. albicans viability was also evaluated by vital staining techniques and fluorescence microscopy analysis. Antifungal activity against C. albicans significantly increased at both depths in the chlorhexidine groups with and without calcium hydroxide compared with the groups treated with calcium hydroxide only. Treatments with only chlorhexidine or chlorhexidine in combination with calcium hydroxide were effective for elimination of C. albicans. PMID:23538639

INDUCIBLE TRANSIENT CENTRAL RETINAL ARTERY VASOSPASM: A CASE REPORT.

PubMed

Mishulin, Aleksey; Ghandi, Sachin; Apple, Daniel; Lin, Xihui; Hu, Jonathan; Abrams, Gary W

2017-09-27

To report a case of inducible transient central retinal artery vasospasm with associated imaging. Observational case report. A 51-year-old man presented for outpatient follow-up for recurrent inducible transient vision loss in his right eye. He experienced an episode during examination and was found to have central retinal artery vasospasm. Fundus photography and fluorescein angiography obtained during his vasospastic attack confirmed retinal arterial vasospasm. Treatment with a calcium-channel blocker (nifedipine) has been effective in preventing recurrent attacks. Idiopathic primary vasospasm is a rare cause of transient vision loss that is difficult to confirm because of the transient nature. We obtained imaging showing the initiation and resolution of the vasospastic event. The patient was then successfully treated with a calcium-channel blocker.

Releasing-addition method for the flame-photometric determination of calcium in thermal waters

USGS Publications Warehouse

Rowe, J.J.

1963-01-01

Study of the interferences of silica and sulfate in the flame-photometric determination of calcium in thermal waters has led to the development of a method requiring no prior chemical separations. The interference effects of silica, sulfate, potassium, sodium, aluminum, and phosphate are overcome by an addition technique coupled with the use of magnesium as a releasing agent. ?? 1963.

Investigation of free fatty acid associated recombinant membrane receptor protein expression in HEK293 cells using Raman spectroscopy, calcium imaging, and atomic force microscopy.

PubMed

Lin, Juqiang; Xu, Han; Wu, Yangzhe; Tang, Mingjie; McEwen, Gerald D; Liu, Pin; Hansen, Dane R; Gilbertson, Timothy A; Zhou, Anhong

2013-02-05

G-protein-coupled receptor 120 (GPR120) is a previously orphaned G-protein-coupled receptor that apparently functions as a sensor for dietary fat in the gustatory and digestive systems. In this study, a cDNA sequence encoding a doxycycline (Dox)-inducible mature peptide of GPR120 was inserted into an expression vector and transfected in HEK293 cells. We measured Raman spectra of single HEK293 cells as well as GPR120-expressing HEK293-GPR120 cells at a 48 h period following the additions of Dox at several concentrations. We found that the spectral intensity of HEK293-GPR120 cells is dependent upon the dose of Dox, which correlates with the accumulation of GPR120 protein in the cells. However, the amount of the fatty acid activated changes in intracellular calcium (Ca(2+)) as measured by ratiometric calcium imaging was not correlated with Dox concentration. Principal components analysis (PCA) of Raman spectra reveals that the spectra from different treatments of HEK293-GPR120 cells form distinct, completely separated clusters with the receiver operating characteristic (ROC) area of 1, while those spectra for the HEK293 cells form small overlap clusters with the ROC area of 0.836. It was also found that expression of GPR120 altered the physiochemical and biomechanical properties of the parental cell membrane surface, which was quantitated by atomic force microscopy (AFM). These findings demonstrate that the combination of Raman spectroscopy, calcium imaging, and AFM may provide new tools in noninvasive and quantitative monitoring of membrane receptor expression induced alterations in the biophysical and signaling properties of single living cells.

Lidocaine bolus may facilitate computed tomographic coronary angiography in patients with frequent premature ventricular contractions

PubMed Central

Jastrzębski, Jan; Kępka, Cezary; Kruk, Mariusz; Demkow, Marcin; Kalińczuk, Łukasz; Wolny, Rafał; Ciszewski, Michał; Michałowska, Ilona; Witkowski, Adam

2013-01-01

Introduction Heart rate irregularities are the major limitations of computed tomographic coronary angiography (CTCA) due to severe motion artifacts. Aim To evaluate the safety and efficacy of a lidocaine intravenous bolus in preserving good image quality by the transient suppression of premature ventricular contractions (PVC) during the CTCA scan. Material and methods The study group comprised 67 consecutive patients with sinus rhythm and numerous PVC scheduled for CTCA. Intravenous boluses of 25–50 mg lidocaine were given after calcium score assessment and immediately before CTCA. The control group comprised 67 patients with sinus rhythm without PVC matched according to the body mass index (BMI), age, sex, and calcium score. All coronary vessel segments were assessed for image quality and presence of significant stenosis. Results As compared with calcium score assessment, after administration of lidocaine and during the CTCA scan PVC were completely suppressed in 22 (40%), reduced in 10 (18%), unchanged in 18 (32%), and intensified in 5 (10%) patients. Overall, there were 32 (58%) patients with sinus rhythm during CTCA as compared with only 11 (20%) patients free from PVC during calcium score assessment (p < 0.001). Image quality in 871 coronary segments including both the study group and control patients was worse in patients with PVC (p < 0.0001). However, there was no statistically significant difference in the number of patients with at least one segment of non-diagnostic quality (6% vs. 12%, p = 0.36; respectively). Conclusions Single lidocaine bolus given prior to CTCA is safe, may temporarily eliminate or reduce the intensity of arrhythmia, and hence results in improved quality of CTCA in patients with numerous PVC. PMID:24570719

Effect of calcium source on structure and properties of sol-gel derived bioactive glasses.

PubMed

Yu, Bobo; Turdean-Ionescu, Claudia A; Martin, Richard A; Newport, Robert J; Hanna, John V; Smith, Mark E; Jones, Julian R

2012-12-18

The aim was to determine the most effective calcium precursor for synthesis of sol-gel hybrids and for improving homogeneity of sol-gel bioactive glasses. Sol-gel derived bioactive calcium silicate glasses are one of the most promising materials for bone regeneration. Inorganic/organic hybrid materials, which are synthesized by incorporating a polymer into the sol-gel process, have also recently been produced to improve toughness. Calcium nitrate is conventionally used as the calcium source, but it has several disadvantages. Calcium nitrate causes inhomogeneity by forming calcium-rich regions, and it requires high temperature treatment (>400 °C) for calcium to be incorporated into the silicate network. Nitrates are also toxic and need to be burnt off. Calcium nitrate therefore cannot be used in the synthesis of hybrids as the highest temperature used in the process is typically 40-60 °C. Therefore, a different precursor is needed that can incorporate calcium into the silica network and enhance the homogeneity of the glasses at low (room) temperature. In this work, calcium methoxyethoxide (CME) was used to synthesize sol-gel bioactive glasses with a range of final processing temperatures from 60 to 800 °C. Comparison is made between the use of CME and calcium chloride and calcium nitrate. Using advanced probe techniques, the temperature at which Ca is incorporated into the network was identified for 70S30C (70 mol % SiO(2), 30 mol % CaO) for each of the calcium precursors. When CaCl(2) was used, the Ca did not seem to enter the network at any of the temperatures used. In contrast, Ca from CME entered the silica network at room temperature, as confirmed by X-ray diffraction, (29)Si magic angle spinning nuclear magnetic resonance spectroscopy, and dissolution studies. CME should be used in preference to calcium salts for hybrid synthesis and may improve homogeneity of sol-gel glasses.

Real-World Bioresorbable Vascular Scaffold Experience Compared With Second-Generation Metallic Drug-Eluting Stents in Complex Coronary Lesions.

PubMed

Okamoto, Naotaka; Ueda, Hiroshi; Yoshimura, Takahiro; Chamaria, Surbhi; Bhatheja, Samit; Vengrenyuk, Yuliya; Rabiei, Samaneh; Barrientos, Yonandy; Kapur, Vishal; Barman, Nitin; Sweeny, Joseph; Baber, Usman; Mehran, Roxana; Sharma, Samin K; Kini, Annapoorna S

2018-04-15

The aim of the study was to compare the acute outcomes of Absorb bioresorbable vascular scaffolds (BVS) and second-generation drug-eluting stent (DES) implantation in routine clinical practice. There is a paucity of data regarding BVS use in a real-world patient population. The study population comprised 40 consecutive patients who underwent percutaneous coronary intervention (PCI) with BVS implantation at a tertiary-care center in New York, New York between July and December of 2016. An optimal implantation technique including adequate lesion preparation, mandatory postdilation, and optical coherence tomography (OCT) imaging was used in all cases. De novo lesions treated with BVS were compared to lesions treated with DES matched by OCT calcium arc, scaffold/stent size, use of atherectomy device, and lesion postdilation. Acute lumen gain, minimal device area, malapposition, eccentricity, and symmetry index were assessed using OCT. We analyzed OCT images of 40 BVS cases and 40 matching DES cases from 35 and 40 patients, respectively. Compared to the DES group, the BVS group demonstrated similar acute lumen gain, minimal scaffold/stent area, eccentricity index, and symmetry index after PCI. There were fewer malapposed struts detected after BVS implantation; however, malapposition distance and length were not different between the groups. BVS implantation in a real-world patient population with optimal implantation technique resulted in similar stent expansion and better strut apposition compared to DES implantation.

Integrationof Remote Sensing and Geographic information system in Ground Water Quality Assessment and Management

NASA Astrophysics Data System (ADS)

Shakak, N.

2015-04-01

Spatial variations in ground water quality in the Khartoum state, Sudan, have been studied using geographic information system (GIS) and remote sensing technique. Gegraphical informtion system a tool which is used for storing, analyzing and displaying spatial data is also used for investigating ground water quality information. Khartoum landsat mosac image aquired in 2013was used, Arc/Gis software applied to extract the boundary of the study area, the image was classified to create land use/land cover map. The land use map,geological and soil map are used for correlation between land use , geological formations, and soil types to understand the source of natural pollution that can lower the ground water quality. For this study, the global positioning system (GPS), used in the field to identify the borehole location in a three dimentional coordinate (Latitude, longitude, and altitude), water samples were collected from 156 borehole wells, and analyzed for physico-chemical parameters like electrical conductivity, Total dissolved solid,Chloride, Nitrate, Sodium, Magnisium, Calcium,and Flouride, using standard techniques in the laboratory and compared with the standards.The ground water quality maps of the entire study area have been prepared using spatial interpolation technique for all the above parameters.then the created maps used to visualize, analyze, and understand the relationship among the measured points. Mapping was coded for potable zones, non-potable zones in the study area, in terms of water quality sutability for drinking water and sutability for irrigation. In general satellite remote sensing in conjunction with geographical information system (GIS) offers great potential for water resource development and management.

Application of CdZnTe Gamma-Ray Detector for Imaging Corrosion under Insulation

NASA Astrophysics Data System (ADS)

Abdullah, J.; Yahya, R.

2007-05-01

Corrosion under insulation (CUI) on the external wall of steel pipes is a common problem in many types of industrial plants. This is mainly due to the presence of moisture or water in the insulation materials. This type of corrosion can cause failures in areas that are not normally of a primary concern to an inspection program. The failures are often the result of localised corrosion and not general wasting over a large area. These failures can tee catastrophic in nature or at least have an adverse economic effect in terms of downtime and repairs. There are a number of techniques used today for CUI investigations. The main ones are profile radiography, pulse eddy current, ultrasonic spot readings and insulation removal. A new system now available is portable Pipe-CUI-Profiler. The nucleonic system is based on dual-beam gamma-ray absorption technique using Cadmium Zinc Telluride (CdZnTe) semiconductor detectors. The Pipe-CUI-Profiler is designed to inspect pipes of internal diameter 50, 65, 80, 90, 100, 125 and 150 mm. Pipeline of these sizes with aluminium or thin steel sheathing, containing fibreglass or calcium silicate insulation to thickness of 25, 40 and 50 mm can be inspected. The system has proven to be a safe, fast and effective method of inspecting pipe in industrial plant operations. This paper describes the application of gamma-ray techniques and CdZnTe semiconductor detectors in the development of Pipe-CUI-Profiler for non-destructive imaging of corrosion under insulation of steel pipes. Some results of actual pipe testing in large-scale industrial plant will be presented.

Modification of caffeine-induced injury in Ca2+-free perfused rat hearts. Relationship to the calcium paradox.

PubMed Central

Vander Heide, R. S.; Altschuld, R. A.; Lamka, K. G.; Ganote, C. E.

1986-01-01

The pathogenesis of the calcium paradox has not been established. In calcium-free perfused hearts, caffeine, which releases calcium from the sarcoplasmic reticulum, causes severe myocardial injury, with creatine kinase (CK) release and contraction band necrosis similar in many respects to the calcium paradox. It has been postulated that contracture, initiated by a small rise in intracellular calcium, may cause sarcolemmal injury in both the calcium paradox and caffeine-induced myocardial injury. The present study was initiated to determine whether interventions which modulate caffeine-induced contracture will also correspondingly alter cellular injury. The effects of caffeine dose, procaine, extended calcium-free perfusion, elevated potassium, temperature, and increasing intracellular sodium on caffeine-induced contracture were examined in Langendorff-perfused adult rat hearts. Caffeine-induced contracture at 22 C increased over a dose range of 5-40 mM caffeine. Procaine, which inhibits caffeine-induced calcium release at doses between 5 and 20 mM, progressively reduced contracture caused by addition of 20 mM caffeine at 22 C. Hearts perfused with calcium-free solution containing 16 mM K+ showed a reduction in caffeine-induced contracture. Extended calcium-free perfusion (20 minutes) at temperatures from 18 to 37 C resulted in a progressive reduction of caffeine-induced contracture. Each of these interventions was also found to inhibit caffeine-induced injury at 37 C. Low temperature was found to have complex effects. Hypothermia enhanced caffeine contractures but also protected hearts from cell separations and CK release. Increasing intracellular sodium was found to enhance caffeine-induced contracture at 37 C. There was a direct correlation between measured intracellular sodium levels and the magnitude and duration of caffeine-induced contracture. These results demonstrate a direct correlation between the magnitude of contracture and myocardial injury in calcium-free hearts. It is proposed that contracture is the primary mediator of sarcolemmal membrane injury in hearts with intercalated disks weakened by prior calcium-free perfusion. Images Figure 11 PMID:3706496

Real-time scratch assay reveals mechanisms of early calcium signaling in breast cancer cells in response to wounding

PubMed Central

Pratt, Stephen J.P.; Hernández-Ochoa, Erick O.; Lee, Rachel M.; Ory, Eleanor C.; Lyons, James S.; Joca, Humberto C.; Johnson, Ashley; Thompson, Keyata; Bailey, Patrick; Lee, Cornell J.; Mathias, Trevor; Vitolo, Michele I.; Trudeau, Matt; Stains, Joseph P.; Ward, Christopher W.; Schneider, Martin F.; Martin, Stuart S.

2018-01-01

Aggressive cellular phenotypes such as uncontrolled proliferation and increased migration capacity engender cellular transformation, malignancy and metastasis. While genetic mutations are undisputed drivers of cancer initiation and progression, it is increasingly accepted that external factors are also playing a major role. Two recently studied modulators of breast cancer are changes in the cellular mechanical microenvironment and alterations in calcium homeostasis. While many studies investigate these factors separately in breast cancer cells, very few do so in combination. This current work sets a foundation to explore mechano-calcium relationships driving malignant progression in breast cancer. Utilizing real-time imaging of an in vitro scratch assay, we were able to resolve mechanically-sensitive calcium signaling in human breast cancer cells. We observed rapid initiation of intracellular calcium elevations within seconds in cells at the immediate wound edge, followed by a time-dependent increase in calcium in cells at distances up to 500μm from the scratch wound. Calcium signaling to neighboring cells away from the wound edge returned to baseline within seconds. Calcium elevations at the wound edge however, persisted for up to 50 minutes. Rigorous quantification showed that extracellular calcium was necessary for persistent calcium elevation at the wound edge, but intercellular signal propagation was dependent on internal calcium stores. In addition, intercellular signaling required extracellular ATP and activation of P2Y2 receptors. Through comparison of scratch-induced signaling from multiple cell lines, we report drastic reductions in response from aggressively tumorigenic and metastatic cells. The real-time scratch assay established here provides quantitative data on the molecular mechanisms that support rapid scratch-induced calcium signaling in breast cancer cells. These mechanisms now provide a clear framework for investigating which short-term calcium signals promote long-term changes in cancer cell biology. PMID:29861849

Label-free biomolecular characterization of human breast cancer tissue with stimulated Raman scattering (SRS) spectral imaging (Conference Presentation)

NASA Astrophysics Data System (ADS)

Lu, Fa-Ke F.; Calligaris, David; Suo, Yuanzhen; Santagata, Sandro; Golby, Alexandra J.; Xie, X. Sunney; Mallory, Melissa A.; Golshan, Mehra; Dillon, Deborah A.; Agar, Nathalie Y. R.

2017-02-01

Stimulated Raman scattering (SRS) microscopy has been used for rapid label-free imaging of various biomolecules and drugs in living cells and tissues (Science, doi:10.1126/science.aaa8870). Our recent work has demonstrated that lipid and protein mapping of cancer tissue renders pathology-like images, providing essential histopathological information with subcellular resolution of the entire specimen (Cancer Research, doi: 10.1158/0008-5472.CAN-16-027). We have also established the first SRS imaging Atlas of human brain tumors (Harvard Dataverse, doi: (doi:10.7910/DVN/EZW4EK). SRS imaging of tissue could provide invaluable information for cancer diagnosis and surgical guidance in two aspects: rapid surgical pathology and quantitative biomolecular characterization. In this work, we present the use of SRS microscopy for characterization of a few essential biomolecules in breast cancer. Human breast cancer tissue specimens at the tumor core, tumor margin and normal area (5 cm away from the tumor) from surgical cases will be imaged with SRS at multiple Raman shifts, including the peaks for lipid, protein, blood (absorption), collagen, microcalcification (calcium phosphates and calcium oxalate) and carotenoids. Most of these Raman shifts have relatively strong Raman cross sections, which ensures high-quality and fast imaging. This proof-of-principle study is sought to demonstrate the feasibility and potential of SRS imaging for ambient diagnosis and surgical guidance of breast cancer.

Use of aromatic salts for simultaneously removing SO.sub.2 and NO.sub.x pollutants from exhaust of a combustion system

DOEpatents

Levendis, Yiannis A.; Wise, Donald L.

1994-10-04

A method is disclosed for removing pollutants from the exhaust of combustion systems burning fuels containing substantial amounts of sulfur and nitrogen. An exemplary method of the invention involves the formation and reaction of a sorbent comprising calcium benzoate. The calcium benzoate is either dry-sprayed (in the form of a fine powder) or wet-sprayed in an aqueous solution in a high temperature environment such as a combustion chamber. The latter technique is feasible since calcium benzoate is a water-soluble form of calcium. When the dispersed particles of calcium benzoate are heated to a high temperature, the organic benzoate burns off and fine calcium oxide particles are formed. These particles are cenospheric (hollow) and have thin and highly porous walls, thus, affording optimum external and internal accessibility for reacting with toxic gaseous emissions such as SO.sub.2. Further, the combustion of the organic benzoate portion of the sorbent results in the conversion of NO.sub.x to N.sub.2.

Calcium kinetics with microgram stable isotope doses and saliva sampling

NASA Technical Reports Server (NTRS)

Smith, S. M.; Wastney, M. E.; Nyquist, L. E.; Shih, C. Y.; Wiesmann, H.; Nillen, J. L.; Lane, H. W.

1996-01-01

Studies of calcium kinetics require administration of tracer doses of calcium and subsequent repeated sampling of biological fluids. This study was designed to develop techniques that would allow estimation of calcium kinetics by using small (micrograms) doses of isotopes instead of the more common large (mg) doses to minimize tracer perturbation of the system and reduce cost, and to explore the use of saliva sampling as an alternative to blood sampling. Subjects received an oral dose (133 micrograms) of 43Ca and an i.v. dose (7.7 micrograms) of 46Ca. Isotopic enrichment in blood, urine, saliva and feces was well above thermal ionization mass spectrometry measurement precision up to 170 h after dosing. Fractional calcium absorptions determined from isotopic ratios in blood, urine and saliva were similar. Compartmental modeling revealed that kinetic parameters determined from serum or saliva data were similar, decreasing the necessity for blood samples. It is concluded from these results that calcium kinetics can be assessed with micrograms doses of stable isotopes, thereby reducing tracer costs and with saliva samples, thereby reducing the amount of blood needed.

Photothermal gold nanoparticle mediated stimulation of cardiomyocyte beating (Conference Presentation)

NASA Astrophysics Data System (ADS)

Kalies, Stefan; Gentemann, Lara; Coffee, Michelle; Zweigerdt, Robert; Heinemann, Dag; Heisterkamp, Alexander

2017-03-01

Photothermal manipulation of cells via heating of gold nanoparticles has proven to be an efficient tool for molecular delivery into cells via cell perforation with short laser pulses. We investigated a potential extension of this technique for cell stimulation of cardiomyocytes using a 532 nm and 850 ps laser system and a surface concentration of 0.5 μg/cm2 of 200 nm gold nanoparticles. The gold nanoparticles were unspecifically attached to the cardiomyocytes after an incubation period of three hours. The laser irradiation leads to a temperature rise directly at the particles of several hundred degrees K which evokes bubble formation and membrane perforation. We examined the effect of laser based photothermal manipulation at different laser powers, with different calcium concentrations, and for a cardiomyocyte-like cell line (HL1 cells), neonatal rat cardiomyocytes and human embryonic stem cell (hESC)-derived cardiomyocytes. Fast calcium oscillations in HL1 cells were observed in the presence and absence of extracellular calcium and most pronounced in the area next to the laser spot after irradiation. Within the laser spot, in particular high laser powers led to a single rise in calcium over a time period of several seconds. These results were confirmed in stem cell-derived cardiomyocytes. In the presence of normal and high calcium concentrations, the spontaneous contraction frequency increased after laser irradiation in neonatal rat cardiomyocytes. Consequently, gold nanoparticle mediated photothermal cell manipulation via pulsed lasers may serve as a potential pacemaker-technique in regenerative approaches, next to optogenetics.

[Applicability of laser-based geological techniques in bone research: analysis of calcium oxide distribution in thin-cut animal bones].

PubMed

Andrássy, László; Maros, Gyula; Kovács, István János; Horváth, Ágnes; Gulyás, Katalin; Bertalan, Éva; Besnyi, Anikó; Füri, Judit; Fancsik, Tamás; Szekanecz, Zoltán; Bhattoa, Harjit Pal

2014-11-09

The structural similarities between the inorganic component of bone tissue and geological formations make it possible that mathematic models may be used to determine weight percentage composition of different mineral element oxides constituting the inorganic component of bone tissue. The determined weight percentage composition can be verified with the determination of element oxide concentration values by laser induced plasma spectroscopy and inductively coupled plasma optical emission spectrometry. It can be concluded from calculated weight percentage composition of the inorganic component of bone tissue and laboratory analyses that the properties of bone tissue are determined primarily by hydroxylapatite. The inorganic bone structure can be studied well by determining the calcium oxide concentration distribution using the laser induced plasma spectroscopy technique. In the present study, thin polished bone slides prepared from male bovine tibia were examined with laser induced plasma spectroscopy in a regular network and combined sampling system to derive the calculated calcium oxide concentration distribution. The superficial calcium oxide concentration distribution, as supported by "frequency distribution" curves, can be categorized into a number of groups. This, as such, helps in clearly demarcating the cortical and trabecular bone structures. Following analyses of bovine tibial bone, the authors found a positive association between the attenuation value, as determined by quantitative computer tomography and the "ρ" density, as used in geology. Furthermore, the calculated "ρ" density and the measured average calcium oxide concentration values showed inverse correlation.

Fluorescence Fluctuations and Equivalence Classes of Ca 2+ Imaging Experiments

PubMed Central

Piegari, Estefanía; Lopez, Lucía; Perez Ipiña, Emiliano; Ponce Dawson, Silvina

2014-01-01

release into the cytosol through inositol 1,4,5-trisphosphate receptors (IP3Rs) plays a relevant role in numerous physiological processes. IP3R-mediated signals involve -induced -release (CICR) whereby release through one open IP3R induces the opening of other channels. IP3Rs are apparently organized in clusters. The signals can remain localized (i.e., puffs) if CICR is limited to one cluster or become waves that propagate between clusters. puffs are the building blocks of waves. Thus, there is great interest in determining puff properties, especially in view of the current controversy on the spatial distribution of activatable IP3Rs. puffs have been observed in intact cells with optical techniques proving that they are intrinsically stochastic. Obtaining a correct picture of their dynamics then entails being able to detect the whole range of puff sizes. puffs are observed using visible single-wavelength dyes, slow exogenous buffers (e.g., EGTA) to disrupt inter-cluster CICR and UV-photolyzable caged IP3. Single-wavelength dyes increase their fluorescence upon calcium binding producing images that are strongly dependent on their kinetic, transport and photophysical properties. Determining the artifacts that the imaging setting introduces is particularly relevant when trying to analyze the smallest signals. In this paper we introduce a method to estimate the expected signal-to-noise ratio of imaging experiments that use single-wavelength dyes. The method is based on the Number and Brightness technique. It involves the performance of a series of experiments and their subsequent analysis in terms of a fluorescence fluctuation model with which the model parameters are quantified. Using the model, the expected signal-to-noise ratio is then computed. Equivalence classes between different experimental conditions that produce images with similar signal-to-noise ratios can then be established. The method may also be used to estimate the smallest signals that can reliably be observed with each setting. PMID:24776736

Static antibiotic spacers augmented by calcium sulphate impregnated beads in revision TKA: Surgical technique and review of literature.

PubMed

Risitano, Salvatore; Sabatini, Luigi; Atzori, Francesco; Massè, Alessandro; Indelli, Pier Francesco

2018-06-01

Periprosthetic joint infection (PJI) is a serious complication in total knee arthroplasty (TKA) and represents one of the most common causes of revision. The challenge for surgeons treating an infected TKA is to quickly obtain an infection-free joint in order to re-implant, when possible, a new TKA. Recent literature confirms the role of local antibiotic-loaded beads as a strong bactericidal, allowing higher antibiotic elution when compared with antibiotic loaded spacers only. Unfortunately, classical Polymethylmethacrylate (PMMA) beads might allow bacteria adhesion, secondary development of antibiotic resistance and eventually surgical removal once antibiotics have eluted. This article describes a novel surgical technique using static, custom-made antibiotic loaded spacers augmented by calcium sulphate antibiotic-impregnated beads to improve the success rate of revision TKA in a setting of PJI. The use of calcium sulphate beads has several potential benefits, including a longer sustained local antibiotic release when compared with classical PMMA beads and, being resorbable, not requiring accessory surgical interventions.

Space Flight Effects on Intracellular Ions in Sublingual Cells of Non-Human Primates

NASA Technical Reports Server (NTRS)

Arnaud, Sara B.; Dotsenko, R.; Fung, P.; Navidi, M.; Silver, B.; Wade, Charles E. (Technical Monitor)

1994-01-01

We have used a novel technique that quantifies minerals and electrolytes from smears of sublingual cells by x-ray microanalysis to monitor metabolic changes in bed rest subjects. Increases in intracellular calcium (Ca), phosphorus (P), and potassium (K) were characteristic of subjects whose exercise regimen was inadequate to maintain calcium metabolism. To test the effects of space flight on intracellular ions, we analyzed cells from 2-4 kg Rhesus monkeys before and after 2 weeks in space or chair restraint (CR). There were increases in sublingual cell Ca, P and K after space flight which paralleled the clinical estimates of metabolic status of the animals and exceeded the levels found during CR on R+11. Increases after 2 weeks CR were 26% in Ca, 6% in P and 29% in K. Species similarity ill responses of intracellular ions to inactivity imposed by bed rest, restraint or microgravity suggest that this innovative non-invasive technique would be a useful in-flight monitor of exercise countermeasures directed toward maintaining calcium balance.

Correlating Calcium Binding, Förster Resonance Energy Transfer, and Conformational Change in the Biosensor TN-XXL

PubMed Central

Geiger, Anselm; Russo, Luigi; Gensch, Thomas; Thestrup, Thomas; Becker, Stefan; Hopfner, Karl-Peter; Griesinger, Christian; Witte, Gregor; Griesbeck, Oliver

2012-01-01

Genetically encoded calcium indicators have become instrumental in imaging signaling in complex tissues and neuronal circuits in vivo. Despite their importance, structure-function relationships of these sensors often remain largely uncharacterized due to their artificial and multimodular composition. Here, we describe a combination of protein engineering and kinetic, spectroscopic, and biophysical analysis of the Förster resonance energy transfer (FRET)-based calcium biosensor TN-XXL. Using fluorescence spectroscopy of engineered tyrosines, we show that two of the four calcium binding EF-hands dominate the FRET output of TN-XXL and that local conformational changes of these hands match the kinetics of FRET change. Using small-angle x-ray scattering and NMR spectroscopy, we show that TN-XXL changes from a flexible elongated to a rigid globular shape upon binding calcium, thus resulting in FRET signal output. Furthermore, we compare calcium titrations using fluorescence lifetime spectroscopy with the ratiometric approach and investigate potential non-FRET effects that may affect the fluorophores. Thus, our data characterize the biophysics of TN-XXL in detail and may form a basis for further rational engineering of FRET-based biosensors. PMID:22677394

Optimization of K-edge imaging for vulnerable plaques using gold nanoparticles and energy resolved photon counting detectors: a simulation study.

PubMed

Alivov, Yahya; Baturin, Pavlo; Le, Huy Q; Ducote, Justin; Molloi, Sabee

2014-01-06

We investigated the effect of different imaging parameters, such as dose, beam energy, energy resolution and the number of energy bins, on the image quality of K-edge spectral computed tomography (CT) of gold nanoparticles (GNP) accumulated in an atherosclerotic plaque. A maximum likelihood technique was employed to estimate the concentration of GNP, which served as a targeted intravenous contrast material intended to detect the degree of the plaque's inflammation. The simulation studies used a single-slice parallel beam CT geometry with an x-ray beam energy ranging between 50 and 140 kVp. The synthetic phantoms included small (3 cm in diameter) cylinder and chest (33 × 24 cm(2)) phantoms, where both phantoms contained tissue, calcium and gold. In the simulation studies, GNP quantification and background (calcium and tissue) suppression tasks were pursued. The x-ray detection sensor was represented by an energy resolved photon counting detector (e.g., CdZnTe) with adjustable energy bins. Both ideal and more realistic (12% full width at half maximum (FWHM) energy resolution) implementations of the photon counting detector were simulated. The simulations were performed for the CdZnTe detector with a pixel pitch of 0.5-1 mm, which corresponds to a performance without significant charge sharing and cross-talk effects. The Rose model was employed to estimate the minimum detectable concentration of GNPs. A figure of merit (FOM) was used to optimize the x-ray beam energy (kVp) to achieve the highest signal-to-noise ratio with respect to the patient dose. As a result, the successful identification of gold and background suppression was demonstrated. The highest FOM was observed at the 125 kVp x-ray beam energy. The minimum detectable GNP concentration was determined to be approximately 1.06 µmol mL(-1) (0.21 mg mL(-1)) for an ideal detector and about 2.5 µmol mL(-1) (0.49 mg mL(-1)) for a more realistic (12% FWHM) detector. The studies show the optimal imaging parameters at the lowest patient dose using an energy resolved photon counting detector to image GNP in an atherosclerotic plaque.

Chimeric green fluorescent protein-aequorin as bioluminescent Ca2+ reporters at the single-cell level

PubMed Central

Baubet, Valérie; Le Mouellic, Hervé; Campbell, Anthony K.; Lucas-Meunier, Estelle; Fossier, Philippe; Brûlet, Philippe

2000-01-01

Monitoring calcium fluxes in real time could help to understand the development, the plasticity, and the functioning of the central nervous system. In jellyfish, the chemiluminescent calcium binding aequorin protein is associated with the green fluorescent protein and a green bioluminescent signal is emitted upon Ca2+ stimulation. We decided to use this chemiluminescence resonance energy transfer between the two molecules. Calcium-sensitive bioluminescent reporter genes have been constructed by fusing green fluorescent protein and aequorin, resulting in much more light being emitted. Chemiluminescent and fluorescent activities of these fusion proteins have been assessed in mammalian cells. Cytosolic Ca2+ increases were imaged at the single-cell level with a cooled intensified charge-coupled device camera. This bifunctional reporter gene should allow the investigation of calcium activities in neuronal networks and in specific subcellular compartments in transgenic animals. PMID:10860991

Field populations of native Indian honey bees from pesticide intensive agricultural landscape show signs of impaired olfaction

NASA Astrophysics Data System (ADS)

Chakrabarti, Priyadarshini; Rana, Santanu; Bandopadhyay, Sreejata; Naik, Dattatraya G.; Sarkar, Sagartirtha; Basu, Parthiba

2015-07-01

Little information is available regarding the adverse effects of pesticides on natural honey bee populations. This study highlights the detrimental effects of pesticides on honey bee olfaction through behavioural studies, scanning electron microscopic imaging of antennal sensillae and confocal microscopic studies of honey bee brains for calcium ions on Apis cerana, a native Indian honey bee species. There was a significant decrease in proboscis extension response and biologically active free calcium ions and adverse changes in antennal sensillae in pesticide exposed field honey bee populations compared to morphometrically similar honey bees sampled from low/no pesticide sites. Controlled laboratory experiments corroborated these findings. This study reports for the first time the changes in antennal sensillae, expression of Calpain 1(an important calcium binding protein) and resting state free calcium in brains of honey bees exposed to pesticide stress.

Field populations of native Indian honey bees from pesticide intensive agricultural landscape show signs of impaired olfaction

PubMed Central

Chakrabarti, Priyadarshini; Rana, Santanu; Bandopadhyay, Sreejata; Naik, Dattatraya G.; Sarkar, Sagartirtha; Basu, Parthiba

2015-01-01

Little information is available regarding the adverse effects of pesticides on natural honey bee populations. This study highlights the detrimental effects of pesticides on honey bee olfaction through behavioural studies, scanning electron microscopic imaging of antennal sensillae and confocal microscopic studies of honey bee brains for calcium ions on Apis cerana, a native Indian honey bee species. There was a significant decrease in proboscis extension response and biologically active free calcium ions and adverse changes in antennal sensillae in pesticide exposed field honey bee populations compared to morphometrically similar honey bees sampled from low/no pesticide sites. Controlled laboratory experiments corroborated these findings. This study reports for the first time the changes in antennal sensillae, expression of Calpain 1(an important calcium binding protein) and resting state free calcium in brains of honey bees exposed to pesticide stress. PMID:26212690

Visualization of cortical, subcortical, and deep brain neural circuit dynamics during naturalistic mammalian behavior with head-mounted microscopes and chronically implanted lenses

PubMed Central

Resendez, Shanna L.; Jennings, Josh H.; Ung, Randall L.; Namboodiri, Vijay Mohan K.; Zhou, Zhe Charles; Otis, James M.; Nomura, Hiroshi; McHenry, Jenna A.; Kosyk, Oksana; Stuber, Garret D.

2016-01-01

Genetically encoded calcium indicators for visualizing dynamic cellular activity have greatly expanded our understanding of the brain. However, due to light scattering properties of the brain as well as the size and rigidity of traditional imaging technology, in vivo calcium imaging has been limited to superficial brain structures during head fixed behavioral tasks. This limitation can now be circumvented by utilizing miniature, integrated microscopes in conjunction with an implantable microendoscopic lens to guide light into and out of the brain, thus permitting optical access to deep brain (or superficial) neural ensembles during naturalistic behaviors. Here, we describe procedural steps to conduct such imaging studies using mice. However, we anticipate the protocol can be easily adapted for use in other small vertebrates. Successful completion of this protocol will permit cellular imaging of neuronal activity and the generation of data sets with sufficient statistical power to correlate neural activity with stimulus presentation, physiological state, and other aspects of complex behavioral tasks. This protocol takes 6–11 weeks to complete. PMID:26914316