Ex Vivo Brain Preparation to Analyze Vocal Pathways of Xenopus Frogs

2021 ◽  
Vol 2021 (9) ◽  
pp. pdb.prot106872
Author(s):  
Ayako Yamaguchi
Keyword(s):  
Neural Activity ◽  
Ex Vivo ◽  
Neural Circuitry ◽  
Neural Basis ◽  
Vocal Production ◽  
Basic Principles ◽  
Electrophysiological Recordings ◽  
The Brain

Understanding the neural basis of behavior is a challenging task for technical reasons. Most methods of recording neural activity require animals to be immobilized, but neural activity associated with most behavior cannot be recorded from an anesthetized, immobilized animal. Using amphibians, however, there has been some success in developing in vitro brain preparations that can be used for electrophysiological and anatomical studies. Here, we describe an ex vivo frog brain preparation from which fictive vocalizations (the neural activity that would have produced vocalizations had the brain been attached to the muscle) can be elicited repeatedly. When serotonin is applied to the isolated brains of male and female African clawed frogs, Xenopus laevis, laryngeal nerve activity that is a facsimile of those that underlie sex-specific vocalizations in vivo can be readily recorded. Recently, this preparation was successfully used in other species within the genus including Xenopus tropicalis and Xenopus victorianus. This preparation allows a variety of techniques to be applied including extracellular and intracellular electrophysiological recordings and calcium imaging during vocal production, surgical and pharmacological manipulation of neurons to evaluate their impact on motor output, and tract tracing of the neural circuitry. Thus, the preparation is a powerful tool with which to understand the basic principles that govern the production of coherent and robust motor programs in vertebrates.

scholarly journals A method for single-neuron chronic recording from the retina in awake mice

Science ◽  
2018 ◽  
Vol 360 (6396) ◽  
pp. 1447-1451 ◽  
Author(s):  
Guosong Hong ◽  
Tian-Ming Fu ◽  
Mu Qiao ◽  
Robert D. Viveros ◽  
Xiao Yang ◽  
...  
Keyword(s):  
Retinal Ganglion Cells ◽  
Visual Information ◽  
Single Neuron ◽  
Ganglion Cells ◽  
Ex Vivo ◽  
Neural Circuitry ◽  
Retinal Ganglion ◽  
Chronic Recording ◽  
The Brain

The retina, which processes visual information and sends it to the brain, is an excellent model for studying neural circuitry. It has been probed extensively ex vivo but has been refractory to chronic in vivo electrophysiology. We report a nonsurgical method to achieve chronically stable in vivo recordings from single retinal ganglion cells (RGCs) in awake mice. We developed a noncoaxial intravitreal injection scheme in which injected mesh electronics unrolls inside the eye and conformally coats the highly curved retina without compromising normal eye functions. The method allows 16-channel recordings from multiple types of RGCs with stable responses to visual stimuli for at least 2 weeks, and reveals circadian rhythms in RGC responses over multiple day/night cycles.

scholarly journals Latest Trends in Electrochemical Sensors for Neurotransmitters: A Review

Sensors ◽  
2019 ◽  
Vol 19 (9) ◽  
pp. 2037 ◽  
Author(s):  
Tavakolian-Ardakani ◽  
Hosu ◽  
Cristea ◽  
Mazloum-Ardakani ◽  
Marrazza
Keyword(s):  
Electrochemical Sensors ◽  
Ex Vivo ◽  
Analytical Techniques ◽  
Substantial Improvement ◽  
Robust Detection ◽  
Brain Functions ◽  
The Brain ◽  
Made In

Neurotransmitters are endogenous chemical messengers which play an important role in many of the brain functions, abnormal levels being correlated with physical, psychotic and neurodegenerative diseases such as Alzheimer's, Parkinson's, and Huntington's disease. Therefore, their sensitive and robust detection is of great clinical significance. Electrochemical methods have been intensively used in the last decades for neurotransmitter detection, outclassing more complicated analytical techniques such as conventional spectrophotometry, chromatography, fluorescence, flow injection, and capillary electrophoresis. In this manuscript, the most successful and promising electrochemical enzyme-free and enzymatic sensors for neurotransmitter detection are reviewed. Focusing on the activity of worldwide researchers mainly during the last ten years (2010–2019), without pretending to be exhaustive, we present an overview of the progress made in sensing strategies during this time. Particular emphasis is placed on nanostructured-based sensors, which show a substantial improvement of the analytical performances. This review also examines the progress made in biosensors for neurotransmitter measurements in vitro, in vivo and ex vivo.

scholarly journals Inhibition of Tryptophan Catabolism Is Associated With Neuroprotection During Zika Virus Infection

2021 ◽  
Vol 12 ◽  
Author(s):  
Fernanda Martins Marim ◽  
Danielle Cunha Teixeira ◽  
Celso Martins Queiroz-Junior ◽  
Bruno Vinicius Santos Valiate ◽  
Jose Carlos Alves-Filho ◽  
...  
Keyword(s):  
Zika Virus ◽  
Ex Vivo ◽  
Neuroprotective Effect ◽  
Neuronal Cell Death ◽  
Neuronal Cell ◽  
Kynurenine Pathway ◽  
Pcr Analysis ◽  
The Brain

Zika virus (ZIKV) is an arbovirus belonging to Flaviviridae family that emerged as a global health threat due to its association with microcephaly and other severe neurological complications, including Guillain-Barré Syndrome (GBS) and Congenital Zika Syndrome (CZS). ZIKV disease has been linked to neuroinflammation and neuronal cell death. Neurodegenerative processes may be exacerbated by metabolites produced by the kynurenine pathway, an important pathway for the degradation of tryptophan, which induces neuronal dysfunction due to enhanced excitotoxicity. Here, we exploited the hypothesis that ZIKV-induced neurodegeneration can be rescued by blocking a target enzyme of the kynurenine pathway, the Indoleamine 2,3-dioxygenase (IDO-1). RT-PCR analysis showed increased levels of IDO-1 RNA expression in undifferentiated primary neurons isolated from wild type (WT) mice infected by ZIKV ex vivo, as well as in the brain of ZIKV-infected A129 mice. Pharmacological inhibition of IDO-1 enzyme with 1-methyl-D-tryptophan (1-MT), in both in vitro and in vivo systems, led to significant reduction of ZIKV-induced neuronal death without interfering with the ability of ZIKV to replicate in those cells. Furthermore, in vivo analyses using both genetically modified mice (IDO-/- mice) and A129 mice treated with 1-MT resulted in reduced microgliosis, astrogliosis and Caspase-3 positive cells in the brain of ZIKV-infected A129 mice. Interestingly, increased levels of CCL5 and CXCL-1 chemokines were found in the brain of 1-MT treated-mice. Together, our data indicate that IDO-1 blockade provides a neuroprotective effect against ZIKV-induced neurodegeneration, and this is amenable to inhibition by pharmacological treatment.

scholarly journals Preclinical and clinical study on [18F]DRKXH1: a novel β-amyloid PET tracer for Alzheimer’s disease

2021 ◽  
Author(s):  
MiaoMiao Xu ◽  
Jun Guo ◽  
JiaCheng Gu ◽  
LinLin Zhang ◽  
ZiHao Liu ◽  
...  
Keyword(s):  
Transgenic Mice ◽  
Ex Vivo ◽  
Small Animal ◽  
Small Animal Pet ◽  
Animal Pet ◽  
In Vitro Autoradiography ◽  
Healthy Control ◽  
The Brain

Abstract Background The deposition of β-amyloid (Aβ) in the brain is a biomarker of Alzheimer’s disease (AD). Highly sensitive Aβ positron emission tomography (PET) imaging plays an essential role in diagnosing and evaluating the therapeutic effects of AD. Aim To synthesize a new Aβ tracer [18F]DRKXH1 (5-(4-(6-(2-[18]fluoroethoxy)ethoxy)imidazo[1,2-alpha]pyridin-2-yl)phenyl) and evaluate the tracer performance by biodistribution analysis, in vivo small-animal PET-CT dynamic scan, ex vivo and in vitro autoradiography, and PET in human subjects. Methods [18F]DRKXH1 was synthesized automatically by the GE FN module. Log D (pH 7.4) and biodistribution of [18F]DRKXH1 were investigated. Small-animal-PET was used for [18F]DRKXH1 and [18F]AV45 imaging study in AD transgenic mice (APPswe/PSEN1dE9) and age-matched normal mice. The distribution volume ratios (DVR) and standardized uptake value ratios (SUVRs) were calculated with the cerebellum as the reference region. The deposition of Aβ plaques in the brain of AD transgenic mice was determined by ex vivo autoradiography and immunohistochemistry. In vitro autoradiography was performed in the postmortem brain sections of AD patients and healthy controls. Two healthy control subjects and one AD patient was subjected to in vivo PET study using [18F]DRKXH1. Results The yield of [18F]DRKXH1 was 40%, and the specific activity was 156.64 ± 11.55 GBq/μmol. [18F]DRKXH1 was mainly excreted through the liver and kidney. The small-animal PET study showed high initial brain uptake and rapid washout of [18F]DRKXH1. The concentration of [18F]DRKXH1 was detected in the cortex and hippocampus of AD transgenic mice brain. The cortex DVR of AD transgenic mice was higher than that of WT mice (P < 0.0001). Moreover, the SUVRs of AD transgenic mice were higher than those of WT mice based on the 0–60-min dynamic scanning. In vitro autoradiography showed a significant concentration of tracer in the Aβ plaque-rich areas in the brain of AD transgenic mice. The DVR value of [18F]-DRKXH1 is higher than that of [18F]-AV45 (1.29 ± 0.05 vs. 1.05 ± 0.08; t = 5.33, P = 0.0003). Autoradiography of postmortem human brain sections showed [18F]DRKXH1-labeled Aβ plaques in the AD brain. The AD patients had high retention in cortical regions, while healthy control subjects had uniformly low radioactivity uptake. Conclusions [18F]DRKXH1 is an Aβ tracer with high sensitivity in preclinical study and has the potential for in vivo detection of the human brain.

scholarly journals Brain pharmacokinetics of two BBB penetrating bispecific antibodies of different size

2021 ◽  
Vol 18 (1) ◽  
Author(s):  
Rebecca Faresjö ◽  
Gillian Bonvicini ◽  
Xiaotian T. Fang ◽  
Ximena Aguilar ◽  
Dag Sehlin ◽  
...  
Keyword(s):  
Ex Vivo ◽  
Elimination Rate ◽  
Brain Parenchyma ◽  
Bispecific Antibodies ◽  
Igg Antibody ◽  
Nuclear Track Emulsion ◽  
Ex Vivo Autoradiography ◽  
The Brain

Abstract Background Transferrin receptor (TfR1) mediated enhanced brain delivery of antibodies have been studied extensively in preclinical settings. However, the brain pharmacokinetics, i.e. brain entry, distribution and elimination are still not fully understood for this class of antibodies. The overall aim of the study was to compare the brain pharmacokinetics of two BBB-penetrating bispecific antibodies of different size (210 vs 58 kDa). Specifically, we wanted to investigate if the faster systemic clearance of the smaller non-IgG antibody di-scFv3D6-8D3, in comparison with the IgG-based bispecific antibody mAb3D6-scFv8D3, was also reflected in the brain. Methods Wild-type (C57/Bl6) mice were injected with 125I-iodinated ([125I]) mAb3D6-scFv8D3 (n = 46) or [125I]di-scFv3D6-8D3 (n = 32) and euthanized 2, 4, 6, 8, 10, 12, 16, or 24 h post injection. Ex vivo radioactivity in whole blood, peripheral organs and brain was measured by γ-counting. Ex vivo autoradiography and nuclear track emulsion were performed on brain sections to investigate brain and parenchymal distribution. Capillary depletion was carried out at 2, 6, and 24 h after injection of [125I]mAb3D6-scFv8D3 (n = 12) or [125I]di-scFv3D6-8D3 (n = 12), to estimate the relative levels of radiolabelled antibody in brain capillaries versus brain parenchyma. In vitro binding kinetics for [125I]mAb3D6-scFv8D3 or [125I]di-scFv3D6-8D3 to murine TfR were determined by LigandTracer. Results [125I]di-scFv3D6-8D3 showed faster elimination from blood, lower brain Cmax, and Tmax, a larger parenchymal-to-capillary concentration ratio, and a net elimination from brain at an earlier time point after injection compared with the larger [125I]mAb3D6-scFv8D3. However, the elimination rate from brain did not differ between the antibodies. The study also indicated that [125I]di-scFv3D6-8D3 displayed lower avidity than [125I]mAb3D6-scFv8D3 towards TfR1 in vitro and potentially in vivo, at least at the BBB. Conclusion A smaller size and lower TfR1 avidity are likely important for fast parenchymal delivery, while elimination of brain-associated bispecific antibodies may not be dependent on these characteristics.

scholarly journals Preliminary Assessment of the Anti-Inflammatory Activity of New Structural Honokiol Analogs with a 4′-O-(2-fluoroethyl) Moiety and the Potential of Their 18F-Labeled Derivatives for Neuroinflammation Imaging

Molecules ◽  
2021 ◽  
Vol 26 (21) ◽  
pp. 6630
Author(s):  
Daria D. Vaulina ◽  
Kira I. Stosman ◽  
Konstantin V. Sivak ◽  
Andrey G. Aleksandrov ◽  
Nikolai B. Viktorov ◽  
...  
Keyword(s):  
Ex Vivo ◽  
Inflammatory Activity ◽  
Preliminary Evaluation ◽  
Moderate Increase ◽  
Biodistribution Studies ◽  
Cox 2 ◽  
Anti Inflammatory ◽  
The Brain

Neolignans honokiol and 4′-O-methylhonokiol (MH) and their derivatives have pronounced anti-inflammatory activity, as evidenced by numerous pharmacological studies. Literature data suggested that cyclooxygenase type 2 (COX-2) may be a target for these compounds in vitro and in vivo. Recent studies of [11C]MPbP (4′-[11С]methoxy-5-propyl-1,1′-biphenyl-2-ol) biodistribution in LPS (lipopolysaccharide)-treated rats have confirmed the high potential of MH derivatives for imaging neuroinflammation. Here, we report the synthesis of four structural analogs of honokiol, of which 4′-(2-fluoroethoxy)-2-hydroxy-5-propyl-1, 1′-biphenyl (F-IV) was selected for labeling with fluorine-18 (T1/2 = 109.8 min) due to its high anti-inflammatory activity confirmed by enzyme immunoassays (EIA) and neuromorphological studies. The high inhibitory potency of F-IV to COX-2 and its moderate lipophilicity and chemical stability are favorable factors for the preliminary evaluation of the radioligand [18F]F-IV in a rodent model of neuroinflammation. [18F]F-IV was prepared with good radiochemical yield and high molar activity and radiochemical purity by 18F-fluoroethylation of the precursor with Boc-protecting group (15) with [18F]2-fluoro-1-bromoethane ([18F]FEB). Ex vivo biodistribution studies revealed a small to moderate increase in radioligand uptake in the brain and peripheral organs of LPS-induced rats compared to control animals. Pretreatment with celecoxib resulted in significant blocking of radioactivity uptake in the brain (pons and medulla), heart, lungs, and kidneys, indicating that [18F]F-IV is likely to specifically bind to COX-2 in a rat model of neuroinflammation. However, in comparison with [11C]MPbP, the new radioligand showed decreased brain uptake in LPS rats and high retention in the blood pool, which apparently could be explained by its high plasma protein binding. We believe that the structure of [18F]F-IV can be optimized by replacing the substituents in the biphenyl core to eliminate these disadvantages and develop new radioligands for imaging activated microglia.

scholarly journals Assessing Microglial Dynamics by Live Imaging

2021 ◽  
Vol 12 ◽  
Author(s):  
Megumi Andoh ◽  
Ryuta Koyama
Keyword(s):  
Molecular Mechanisms ◽  
Ex Vivo ◽  
Experimental System ◽  
Live Imaging ◽  
Culture Conditions ◽  
Experimental Conditions ◽  
Advantages And Disadvantages ◽  
The Brain

Microglia are highly dynamic in the brain in terms of their ability to migrate, proliferate, and phagocytose over the course of an individual's life. Real-time imaging is a useful tool to examine how microglial behavior is regulated and how it affects the surrounding environment. However, microglia are sensitive to environmental stimuli, so they possibly change their state during live imaging in vivo, mainly due to surgical damage, and in vitro due to various effects associated with culture conditions. Therefore, it is difficult to perform live imaging without compromising the properties of the microglia under physiological conditions. To overcome this barrier, various experimental conditions have been developed; recently, it has become possible to perform live imaging of so-called surveillant microglia in vivo, ex vivo, and in vitro, although there are various limitations. Now, we can choose in vivo, ex vivo, or in vitro live imaging systems according to the research objective. In this review, we discuss the advantages and disadvantages of each experimental system and outline the physiological significance and molecular mechanisms of microglial behavior that have been elucidated by live imaging.

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