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Analysis of Optokinetic Response in Zebrafish by Computer-Based Eye Tracking

Large-field movements in the visual surround trigger spontaneous, compensatory eye movements known as optokinetic response (OKR) in all vertebrates. In zebrafish (Danio rerio) the OKR is well developed at 5 days post fertilization and can be used in the laboratory for screening of visual p ...

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N -Methyl-d-Aspartate (NMDA)-Mediated Excitotoxic Damage: A Mouse Model of Acute Retinal Ganglion Cell Damage

The animal model of N-methyl-d-aspartate (NMDA)-induced excitotoxic damage of retinal ganglion cells (RGC) is widely used to study the molecular mechanisms of RGC apoptosis and/or its prevention by neuroprotective agents. This chapter provides protocols for applying NMDA-indu ...

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Light Damage as a Model of Retinal Degeneration

The induction of retinal degeneration by light exposure is widely used to study mechanisms of cell death. The advantage of such light-induced lesions over genetically determined degenerations is that light exposures can be manipulated according to the needs of the experimenter. Brig ...

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Phenotyping of Mouse Models with OCT

Optical coherence tomography (OCT) is an invaluable technique to perform noninvasive retinal imaging in small animal models such as mice. It provides virtual cross sections that correlate well with histomorphometric data with the advantage that multiple iterative measurements ...

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Functional Phenotyping of Mouse Models with ERG

In many situations it is important to be able to assess the degree of retinal function, e.g., for the characterization of mouse models with unknown retinal involvement, when studying degenerative processes, for the analysis of visual signal processing, and during the follow-up of therapeut ...

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Retinal Fundus Imaging in Mouse Models of Retinal Diseases

The development of in vivo retinal fundus imaging in mice has opened a new research horizon, not only in ophthalmic research. The ability to monitor the dynamics of vascular and cellular changes in pathological conditions, such as neovascularization or degeneration, longitudinally wi ...

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Mouse Models for Studies of Retinal Degeneration and Diseases

Mouse models, with their well-developed genetics and similarity to human physiology and anatomy, serve as powerful tools with which to investigate the etiology of human retinal degeneration. Mutant mice also provide reproducible, experimental systems for elucidating pathways ...

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Identification and Analysis of Inherited Retinal Disease Genes

Inherited retinal diseases display a very high degree of clinical and genetic heterogeneity, which poses challenges in identifying the underlying defects in known genes and in identifying novel retinal disease genes. Here, we outline the state-of-the-art techniques to find the causa ...

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Analysis of Photoreceptor Rod Outer Segment Phagocytosis by RPE Cells In Situ

Counting rhodopsin-positive phagosomes residing in the retinal pigment epithelium (RPE) in the eye at different times of day allows a quantitative assessment of engulfment and digestion phases of diurnal RPE phagocytosis, which efficiently clears shed photoreceptor outer seg ...

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Analysis of Photoreceptor Outer Segment Phagocytosis by RPE Cells in Culture

Retinal pigment epithelial (RPE) cells are among the most actively phagocytic cells in nature. Primary RPE and stable RPE cell lines provide experimental model systems that possess the same phagocytic machinery as RPE in situ. Upon experimental challenge with isolated photoreceptor ...

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Functional Analysis of Retinal Microglia and Their Effects on Progenitors

The identification of stem/progenitor cells within the retinal neural environment has opened up the possibility of therapy via cellular replacement and/or reprogramming of resident cell populations (1–4). Within the neuro-retinal niche, following injury or in disease states (i ...

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Ca2+ Microfluorimetry in Retinal Mller Glial Cells

Calcium acts as a prominent second messenger in virtually every cell type and modulates a plethora of cell functions. Thus, Ca2+ microfluorimetry became a valuable tool to assess information about mechanisms involved in the regulation of the intracellular calcium level in research on li ...

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Ca2+-Imaging Techniques to Analyze Ca2+ Signaling in Cells and to Monitor Neuronal Activity in the Retina

Ca2+ is an important regulator of many cell functions including proliferation, apoptosis, movements, secretion, contraction, excitation, and differentiation. The regulation of these different cell functions is encoded by the specific temporal and spatial distribution of Ca2+ ...

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Double Chromatin Immunoprecipitation: Analysis of Target Co-occupancy of Retinal Transcription Factors

Combinatorial binding of transcription factors (TFs) and cofactors to specific regulatory regions of target genes in vivo is an important mechanism of transcriptional regulation. Chromatin immunoprecipitation (ChIP) is a powerful technique to detect protein binding to spec ...

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Quantifying the Activity of cis-Regulatory Elements in the Mouse Retina by Explant Electroporation

Transcription factors control gene expression by binding to noncoding regions of DNA known as �cis-regulatory elements (CREs; i.e., enhancer/promoters). Traditionally, cis-regulatory analysis has been carried out via mouse transgenesis which is time-consuming and nonquan ...

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Optimized Technique for Subretinal Injections in Mice

Subretinal injections in mice become increasingly important. Currently, the most prominent application is in gene therapy of inherited eye diseases by means of viral vector delivery to photoreceptors or the retinal pigment epithelium (RPE). Since there are no large animal models for mo ...

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Adeno-Associated Viral Vectors for Gene Therapy of Inherited Retinal Degenerations

Adeno-associated virus (AAV) vectors are in wide use for in vivo gene transfer for the treatment of inherited retinal disease. AAV vectors have been tested in many animal models and have demonstrated efficacy with low toxicity. In this chapter we describe some of the recent methods for small-sca ...

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Barrier Modulation in Drug Delivery to the Retina

The inner blood–retina barrier (iBRB) is essential in restricting the movement of systemic components such as enzymes, anaphylatoxins, or pathogens that could otherwise enter the neural retina and cause extensive damage. The barrier has evolved to confer protection to the delicate mi ...

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Genetic Lineage Tracing of Beta Cell Neogenesis

Genetic lineage tracing is an invaluable tool to demonstrate and measure neogenesis of beta cells from putative precursor cells. Cre-Lox recombination technology can be used for indelible labeling of a cohort of cells and following the fate of these cells and their progeny in animal models. H ...

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Lineage Tracing of Pancreatic Stem Cells and Beta Cell Regeneration

Restoring a functional β cell mass in diabetes patients by β cell transplantation or stimulation of β cell regeneration are promising approaches. It requires knowledge on the mechanisms of β cell neogenesis, an issue that is still quite controversial. Postnatal islet regeneration may or m ...

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