scholarly journals Brief Freezing Steps Lead to Robust Immunofluorescence in Drosophila Larval Brains

2018 ◽  
Author(s):  
Dominic Buckley ◽  
Ada Thapa ◽  
Minh Q. Nguyen ◽  
Essence Blankinship ◽  
Diana Williamson ◽  
...  
Keyword(s):  
Drosophila Melanogaster ◽  
Neurodegenerative Disease ◽  
Molecular Mechanisms ◽  
Structural Integrity ◽  
Proteolytic Degradation ◽  
Antibody Specificity ◽  
Neuronal Function ◽  
Larval Brain ◽  
Apparent Loss ◽  
The Brain

AbstractDrosophila melanogaster possess complex neuronal networks regulating sophisticated behavioral outputs that aid in studying the molecular mechanisms of neuronal function and neurodegenerative disease. Immunofluorescence (IF) techniques provide a way to visualize the spatiotemporal organization of these networks, permitting observation of their development, functional location, remodeling, and eventually - degradation. However, general immunostaining techniques do not always result in sufficient antibody penetration through the brain, and techniques used to enhance permeability can compromise structural integrity. We have found that freezing larval brains facilitates permeability with no apparent loss of antibody specificity or structural integrity. To demonstrate the advantage of this freezing technique, we compared results to two commonly used permeation methods: Detergent alone (Basic) and proteolytic degradation (Collagenase) techniques.SummaryHere we compare four different immunofluorescence techniques demonstrating that freezing Drosophila brains results in robust staining of small neurons in the larval brain without compromising structural integrity.

scholarly journals Brief freezing steps lead to robust immunofluorescence in the Drosophila nervous system

BioTechniques ◽  
2019 ◽  
Vol 67 (6) ◽  
pp. 299-305
Author(s):  
Ada Thapa ◽  
Shea M Sullivan ◽  
Minh Q Nguyen ◽  
Dominic Buckley ◽  
Vy T Ngo ◽  
...  
Keyword(s):  
Nervous System ◽  
Drosophila Melanogaster ◽  
Neurodegenerative Disease ◽  
Spatial Organization ◽  
Molecular Mechanisms ◽  
Structural Integrity ◽  
Antibody Specificity ◽  
Proteolytic Digestion ◽  
Neuronal Function ◽  
The Brain

Drosophila melanogaster possesses a complex nervous system, regulating sophisticated behavioral outputs, that serves as a powerful model for dissecting molecular mechanisms underlying neuronal function and neurodegenerative disease. Immunofluorescence techniques provide a way to visualize the spatiotemporal organization of these networks, permitting observation of their development, functional location, remodeling and, eventually, degradation. However, basic immunostaining techniques do not always result in efficient antibody penetration through the brain, and supplemental techniques to enhance permeability can compromise structural integrity, altering spatial organization. Here, slow freezing of brains is shown to facilitate antibody permeability without loss of antibody specificity or brain integrity. To demonstrate the advantages of this freezing technique, the results of two commonly used permeation methods – detergent-based and partial proteolytic digestion – are compared.

scholarly journals Ubiquitin C-terminal hydrolase L1 (UCH-L1): structure, distribution and roles in brain function and dysfunction

2016 ◽  
Vol 473 (16) ◽  
pp. 2453-2462 ◽  
Author(s):  
Paul Bishop ◽  
Dan Rocca ◽  
Jeremy M. Henley
Keyword(s):  
Amino Acids ◽  
Neurodegenerative Disease ◽  
Brain Function ◽  
Neuronal Development ◽  
Complex Structure ◽  
Deubiquitinating Enzyme ◽  
Neuronal Function ◽  
Abundant Protein ◽  
Neuronal Protein ◽  
The Brain

Ubiquitin C-terminal hydrolase L1 (UCH-L1) is an extremely abundant protein in the brain where, remarkably, it is estimated to make up 1–5% of total neuronal protein. Although it comprises only 223 amino acids it has one of the most complicated 3D knotted structures yet discovered. Beyond its expression in neurons UCH-L1 has only very limited expression in other healthy tissues but it is highly expressed in several forms of cancer. Although UCH-L1 is classed as a deubiquitinating enzyme (DUB) the direct functions of UCH-L1 remain enigmatic and a wide array of alternative functions has been proposed. UCH-L1 is not essential for neuronal development but it is absolutely required for the maintenance of axonal integrity and UCH-L1 dysfunction is implicated in neurodegenerative disease. Here we review the properties of UCH-L1, and how understanding its complex structure can provide new insights into its roles in neuronal function and pathology.

scholarly journals Dynamics of Choline-Containing Phospholipids in Traumatic Brain Injury and Associated Comorbidities

2021 ◽  
Vol 22 (21) ◽  
pp. 11313
Author(s):  
Sana Javaid ◽  
Talha Farooq ◽  
Zohabia Rehman ◽  
Ammara Afzal ◽  
Waseem Ashraf ◽  
...  
Keyword(s):  
Molecular Mechanisms ◽  
Brain Injuries ◽  
Structural Integrity ◽  
Preclinical Studies ◽  
Neuronal Transmission ◽  
Expanding Population ◽  
Motorized Vehicles ◽  
The Brain ◽  
And Oxidative Stress

The incidences of traumatic brain injuries (TBIs) are increasing globally because of expanding population and increased dependencies on motorized vehicles and machines. This has resulted in increased socio-economic burden on the healthcare system, as TBIs are often associated with mental and physical morbidities with lifelong dependencies, and have severely limited therapeutic options. There is an emerging need to identify the molecular mechanisms orchestrating these injuries to life-long neurodegenerative disease and a therapeutic strategy to counter them. This review highlights the dynamics and role of choline-containing phospholipids during TBIs and how they can be used to evaluate the severity of injuries and later targeted to mitigate neuro-degradation, based on clinical and preclinical studies. Choline-based phospholipids are involved in maintaining the structural integrity of the neuronal/glial cell membranes and are simultaneously the essential component of various biochemical pathways, such as cholinergic neuronal transmission in the brain. Choline or its metabolite levels increase during acute and chronic phases of TBI because of excitotoxicity, ischemia and oxidative stress; this can serve as useful biomarker to predict the severity and prognosis of TBIs. Moreover, the effect of choline-replenishing agents as a post-TBI management strategy has been reviewed in clinical and preclinical studies. Overall, this review determines the theranostic potential of choline phospholipids and provides new insights in the management of TBI.

Organization of endogenous clocks in insects

2005 ◽  
Vol 33 (5) ◽  
pp. 957-961 ◽  
Author(s):  
C. Helfrich-Förster
Keyword(s):  
Drosophila Melanogaster ◽  
Molecular Mechanisms ◽  
Clock Genes ◽  
Fruit Fly ◽  
Circadian Clocks ◽  
Rhythm Generation ◽  
Similarities And Differences ◽  
The Brain ◽  
Master Clock

Insect and mammalian circadian clocks show striking similarities. They utilize homologous clock genes, generating self-sustained circadian oscillations in distinct master clocks of the brain, which then control rhythmic behaviour. The molecular mechanisms of rhythm generation were first uncovered in the fruit fly Drosophila melanogaster, whereas cockroaches were among the first animals where the brain master clock was localized. Despite many similarities, there exist obvious differences in the organization and functioning of insect master clocks. These similarities and differences are reviewed on a molecular and anatomical level.

scholarly journals Pituitary Adenylate Cyclase-Activating Polypeptide: A Potent Therapeutic Agent in Oxidative Stress

Antioxidants ◽  
2021 ◽  
Vol 10 (3) ◽  
pp. 354
Author(s):  
Nadia Sadanandan ◽  
Blaise Cozene ◽  
You Jeong Park ◽  
Jeffrey Farooq ◽  
Chase Kingsbury ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Cell Death ◽  
Adenylate Cyclase ◽  
Molecular Mechanisms ◽  
Structural Integrity ◽  
Treatment Options ◽  
Preclinical Research ◽  
Adequate Treatment ◽  
Pituitary Adenylate Cyclase ◽  
The Brain

Stroke is a life-threatening condition that is characterized by secondary cell death processes that occur after the initial disruption of blood flow to the brain. The inability of endogenous repair mechanisms to sufficiently support functional recovery in stroke patients and the inadequate treatment options available are cause for concern. The pathology behind oxidative stress in stroke is of particular interest due to its detrimental effects on the brain. The oxidative stress caused by ischemic stroke overwhelms the neutralization capacity of the body’s endogenous antioxidant system, which leads to an overproduction of reactive oxygen species (ROS) and reactive nitrogen species (RNS) and eventually results in cell death. The overproduction of ROS compromises the functional and structural integrity of brain tissue. Therefore, it is essential to investigate the mechanisms involved in oxidative stress to help obtain adequate treatment options for stroke. Here, we focus on the latest preclinical research that details the mechanisms behind secondary cell death processes that cause many central nervous system (CNS) disorders, as well as research that relates to how the neuroprotective molecular mechanisms of pituitary adenylate cyclase-activating polypeptides (PACAPs) could make these molecules an ideal candidate for the treatment of stroke.

Brain Drug Delivery: Overcoming the Blood-brain Barrier to Treat Tauopathies

2020 ◽  
Vol 26 (13) ◽  
pp. 1448-1465 ◽  
Author(s):  
Jozef Hanes ◽  
Eva Dobakova ◽  
Petra Majerova
Keyword(s):  
Drug Delivery ◽  
Blood Brain Barrier ◽  
Neurodegenerative Disorders ◽  
Brain Barrier ◽  
Neuronal Function ◽  
Brain Drug Delivery ◽  
Naturally Occurring ◽  
Blood Brain ◽  
Traditional Approaches ◽  
The Brain

Tauopathies are neurodegenerative disorders characterized by the deposition of abnormal tau protein in the brain. The application of potentially effective therapeutics for their successful treatment is hampered by the presence of a naturally occurring brain protection layer called the blood-brain barrier (BBB). BBB represents one of the biggest challenges in the development of therapeutics for central nervous system (CNS) disorders, where sufficient BBB penetration is inevitable. BBB is a heavily restricting barrier regulating the movement of molecules, ions, and cells between the blood and the CNS to secure proper neuronal function and protect the CNS from dangerous substances and processes. Yet, these natural functions possessed by BBB represent a great hurdle for brain drug delivery. This review is concentrated on summarizing the available methods and approaches for effective therapeutics’ delivery through the BBB to treat neurodegenerative disorders with a focus on tauopathies. It describes the traditional approaches but also new nanotechnology strategies emerging with advanced medical techniques. Their limitations and benefits are discussed.

scholarly journals Correction: The Nutrient-Responsive Hormone CCHamide-2 Controls Growth by Regulating Insulin-like Peptides in the Brain of Drosophila melanogaster

PLoS Genetics ◽  
2015 ◽  
Vol 11 (9) ◽  
pp. e1005481 ◽  
Author(s):  
Hiroko Sano ◽  
Akira Nakamura ◽  
Michael J. Texada ◽  
James W. Truman ◽  
Hiroshi Ishimoto ◽  
...  
Keyword(s):  
Drosophila Melanogaster ◽  
The Brain

scholarly journals Sublethal Exposure Effects of the Neonicotinoid Clothianidin Strongly Modify the Brain Transcriptome and Proteome in the Male Moth Agrotis ipsilon

Insects ◽  
2021 ◽  
Vol 12 (2) ◽  
pp. 152
Author(s):  
Camille Meslin ◽  
Françoise Bozzolan ◽  
Virginie Braman ◽  
Solenne Chardonnet ◽  
Cédric Pionneau ◽  
...  
Keyword(s):  
Sex Pheromone ◽  
Molecular Mechanisms ◽  
Insect Pest ◽  
Agrotis Ipsilon ◽  
Positive Effects ◽  
Male Moth ◽  
Hormetic Effect ◽  
Pest Insects ◽  
Neuronal Processes ◽  
The Brain

Insect pest management relies mainly on neurotoxic insecticides, including neonicotinoids such as clothianidin. The residual accumulation of low concentrations of these insecticides can have positive effects on target pest insects by enhancing various life traits. Because pest insects often rely on sex pheromones for reproduction and olfactory synaptic transmission is cholinergic, neonicotinoid residues could indeed modify chemical communication. We recently showed that treatments with low doses of clothianidin could induce hormetic effects on behavioral and neuronal sex pheromone responses in the male moth, Agrotis ipsilon. In this study, we used high-throughput RNAseq and proteomic analyses from brains of A. ipsilon males that were intoxicated with a low dose of clothianidin to investigate the molecular mechanisms leading to the observed hormetic effect. Our results showed that clothianidin induced significant changes in transcript levels and protein quantity in the brain of treated moths: 1229 genes and 49 proteins were differentially expressed upon clothianidin exposure. In particular, our analyses highlighted a regulation in numerous enzymes as a possible detoxification response to the insecticide and also numerous changes in neuronal processes, which could act as a form of acclimatization to the insecticide-contaminated environment, both leading to enhanced neuronal and behavioral responses to sex pheromone.

scholarly journals DNA Methylation Profiles of Tph1A and BDNF in Gut and Brain of L. Rhamnosus-Treated Zebrafish

Biomolecules ◽  
2021 ◽  
Vol 11 (2) ◽  
pp. 142
Author(s):  
Mariella Cuomo ◽  
Luca Borrelli ◽  
Rosa Della Monica ◽  
Lorena Coretti ◽  
Giulia De Riso ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Molecular Mechanisms ◽  
The Past ◽  
Targeted Analysis ◽  
Lack Of Information ◽  
High Resolution Power ◽  
Resolution Level ◽  
Health And Disease ◽  
High Doses ◽  
The Brain

The bidirectional microbiota–gut–brain axis has raised increasing interest over the past years in the context of health and disease, but there is a lack of information on molecular mechanisms underlying this connection. We hypothesized that change in microbiota composition may affect brain epigenetics leading to long-lasting effects on specific brain gene regulation. To test this hypothesis, we used Zebrafish (Danio Rerio) as a model system. As previously shown, treatment with high doses of probiotics can modulate behavior in Zebrafish, causing significant changes in the expression of some brain-relevant genes, such as BDNF and Tph1A. Using an ultra-deep targeted analysis, we investigated the methylation state of the BDNF and Tph1A promoter region in the brain and gut of probiotic-treated and untreated Zebrafishes. Thanks to the high resolution power of our analysis, we evaluated cell-to-cell methylation differences. At this resolution level, we found slight DNA methylation changes in probiotic-treated samples, likely related to a subgroup of brain and gut cells, and that specific DNA methylation signatures significantly correlated with specific behavioral scores.

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