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Title: Localization of chemical and electrical synapses in the retina.
Creator: Liu, Yufei., Charles E. Schmidt College of Medicine, Department of Biomedical Science
Abstract/Description: The amphibian retina is commonly used as a model system for studying function and mechanism of the visual system in electrophysiology, since the neural structure and synaptic mechanism of the amphibian retina are similar to higher vertebrate retinas. I determined the specific subtypes of receptors and channels that are involved in chemical and electrical synapses in the amphibian retina. My study indicates that glycine receptor subunits of GlyRº1, 3 and 4 and glutamate receptor subunit of...
Show moreThe amphibian retina is commonly used as a model system for studying function and mechanism of the visual system in electrophysiology, since the neural structure and synaptic mechanism of the amphibian retina are similar to higher vertebrate retinas. I determined the specific subtypes of receptors and channels that are involved in chemical and electrical synapses in the amphibian retina. My study indicates that glycine receptor subunits of GlyRº1, 3 and 4 and glutamate receptor subunit of GluR4 are present in bipolar and amacrine dendrites and axons to conduct chemical synapses in the retinal circuit. I also found that the gap junction channel, pannexin 1a (panx1a), is present in cone-dominated On-bipolar cells and rod-dominated amacrine processes possibly to connect rod-and cone-pathway in the inner retina. In addition, panx1a may form hemi-channels that pass ATP and Ca2+ signals. The findings of my study fill the gap of our knowledge about the subtypes of neurotransmitter receptors and gap junction channels conducting visual information in particular cell types and synaptic areas.
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Date Issued: 2011
PURL: http://purl.flvc.org/FAU/3332725
Subject Headings: Synapses, Neurotransmitters, Neural receptors, Cellular signal transduction
Format: Document (PDF)

Title: Molecular Mechanisms Of CaV2.1 Expression and Functional Organization at the Presynaptic Terminal.
Creator: Das, Brati, Young, Samuel M., Florida Atlantic University, Charles E. Schmidt College of Science, Department of Biological Sciences
Abstract/Description: Neuronal circuit output is dependent on the embedded synapses’ precise regulation of Ca2+ mediated release of neurotransmitter filled synaptic vesicles (SVs) in response to action potential (AP) depolarizations. A key determinant of SV release is the specific expression, organization, and abundance of voltage gated calcium channel (VGCC) subtypes at presynaptic active zones (AZs). In particular, the relative distance that SVs are coupled to VGCCs at AZs results in two different modes of SV...
Show moreNeuronal circuit output is dependent on the embedded synapses’ precise regulation of Ca2+ mediated release of neurotransmitter filled synaptic vesicles (SVs) in response to action potential (AP) depolarizations. A key determinant of SV release is the specific expression, organization, and abundance of voltage gated calcium channel (VGCC) subtypes at presynaptic active zones (AZs). In particular, the relative distance that SVs are coupled to VGCCs at AZs results in two different modes of SV release that dramatically impacts synapse release probability and ultimately the neuronal circuit output. They are: “Ca2+ microdomain,” SV release due to cooperative action of many loosely coupled VGCCs to SVs, or “Ca2+ nanodomain,” SV release due to fewer more tightly coupled VGCCs to SVs. VGCCs are multi-subunit complexes with the pore forming a1 subunit (Cav2.1), the critical determinant of the VGCC subtype kinetics, abundance, and organization at the AZ. Although in central synapses Cav2.2 and Cav2.1 mediate synchronous transmitter release, neurons express multiple VGCC subtypes with differential expression patterns between the cell body and the pre-synapse. The calyx of Held, a giant axosomatic glutamatergic presynaptic terminal that arises from the globular bushy cells (GBC) in the cochlear nucleus, exclusively uses Cav2.1 VGCCs to support the early stages of auditory processing. Due to its experimental accessibility the calyx provides unparalleled opportunities to gain mechanistic insights into Cav2.1 expression, organization, and SV release modes at the presynaptic terminal. Although many molecules are implicated in mediating Cav2.1 expression and SV to VGCC coupling through multiple binding domains on the C-terminus of the Cav2.1 a1 subunit, the underlying fundamental molecular mechanisms remain poorly defined. Here, using viral vector mediated approaches in combination with Cav2.1 conditional knock out transgenic mice, we demonstrate that that there a two independent pathways that control Cav2.1 expression and SV to VGCC coupling at the calyx of Held. These implications for the regulation of synaptic transmission in CNS synapses are discussed.
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Date Issued: 2016
PURL: http://purl.flvc.org/fau/fd/FA00004584
Subject Headings: Synapses., Neural transmission., Cellular signal transduction.
Format: Document (PDF)

Title: Devising a noncancerous model system to study multipolar spindle formation.
Creator: Nagarsheth, Nisha., Harriet L. Wilkes Honors College
Abstract/Description: Aneuploid tumor cells have characteristically unstable genomes which can be caused by mitotic defects such as multipolar spindles. Multipolarity relies upon the presence of extra centrosomes to form. However, some cells, both cancerous and noncancerous are able to avoid the formation of multipolar spindles through centrosomal clustering. Previous research has shown that there are a large number of genes whose activity contributes to the clustering activity, making analysis of individual...
Show moreAneuploid tumor cells have characteristically unstable genomes which can be caused by mitotic defects such as multipolar spindles. Multipolarity relies upon the presence of extra centrosomes to form. However, some cells, both cancerous and noncancerous are able to avoid the formation of multipolar spindles through centrosomal clustering. Previous research has shown that there are a large number of genes whose activity contributes to the clustering activity, making analysis of individual components of the process difficult. In order to better understand centrosomal clustering in cancer cells, we induced supernumerary centrosomes in a genomically normal cell line, RPE, to observe how the normal cells cope with extra centrosomes. Using colcemid to induce extra centrosomes in the RPE cell line, we observed an intact clustering mechanism in fixed cells. Further manipulation of the cells has allowed us to induce multipolarity in this cell line using various disrupters of cell-cycle checkpoint and dynein function.
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Date Issued: 2010
PURL: http://purl.flvc.org/FAU/3335107
Subject Headings: Centrosomes, Research, Cancer, Genetic aspects, Cellular signal transduction, Cell division
Format: Document (PDF)

Title: Deamplification of supernumerary centrosomes by centrosomal clustering.
Creator: Thomas, Ezekiel., Harriet L. Wilkes Honors College
Abstract/Description: Supernumerary centrosomes can arise in a cell through a variety of methods. The presence of supernumerary centrosomes has been observed in nearly all types of cancer and promotes chromosomal instability, with rates of incident increasing as the cancer progresses. An oral squamous cell carcinoma line was treated with hydroxyurea to induce supernumerary centrosomes in the cells. NuMA was then knocked down using shRNA to promote centrosomal clustering and bipolar mitotic division in cells with...
Show moreSupernumerary centrosomes can arise in a cell through a variety of methods. The presence of supernumerary centrosomes has been observed in nearly all types of cancer and promotes chromosomal instability, with rates of incident increasing as the cancer progresses. An oral squamous cell carcinoma line was treated with hydroxyurea to induce supernumerary centrosomes in the cells. NuMA was then knocked down using shRNA to promote centrosomal clustering and bipolar mitotic division in cells with supernumerary centrosomes. Immunofluorescence with an antibody against SAS 6 accuately stained the centrioles for observation. The cells exhibiting supernumerary centrosomes undergoing bipolar mitotic division were studied to look for a possible pattern in centrosomal clustering where the majority of centrosomes are at one pole with a single centrosome at the other pole. Initial results suggest the presence of such a mechanism, which would describe a previously unknown mechanism for cells to deamplify supernumerary centrosomes by centrosomal clustering.
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Date Issued: 2012
PURL: http://purl.flvc.org/FAU/3359328
Subject Headings: Centrosomes, Cell division, Cellular signal transduction, Cancer, Genetic aspects
Format: Document (PDF)

Title: Determination of the acid dissociation constant of cytochrome B5 reductase.
Creator: Chong, Samantha., Harriet L. Wilkes Honors College
Abstract/Description: Most living organisms transduce electron transport chains in order to obtain energy. Flavin adenine dinucleotide (FAD) is a common electron transfer cofactor found in electron transport proteins referred to as flavoproteins. In this study, the different ionization and oxidation states of this cofactor found in cytochrome b5 reductase were identified spectroscopically and quantified as a function of solution potential and pH. The large data sets obtained from these experiments were analyzed...
Show moreMost living organisms transduce electron transport chains in order to obtain energy. Flavin adenine dinucleotide (FAD) is a common electron transfer cofactor found in electron transport proteins referred to as flavoproteins. In this study, the different ionization and oxidation states of this cofactor found in cytochrome b5 reductase were identified spectroscopically and quantified as a function of solution potential and pH. The large data sets obtained from these experiments were analyzed and the acid dissociation constant for reduced cytochrome b5 reductase was determined.
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Date Issued: 2008
PURL: http://purl.flvc.org/FAU/77662
Subject Headings: Cell metabolism, Cellular signal transduction, Chemistry, Statistical methods, Electron spectroscopy
Format: Document (PDF)

Title: Knockdown of KIF9 leads to defects in mitotic entry and progression in mammalian cells.
Creator: Alsina, Laura., Harriet L. Wilkes Honors College
Abstract/Description: Kinesin motors bind to microtubules and function in mitosis and intracellular transport depending on the position of the motor domain within the primary sequence (Hirokawa and Noda 2008). KIF9 has recently been shown to be involved in MTOC positioning and mitotic entry in Dictyostelium (Tikhonenko et al. 2009). To determine if a similar role for KIF9 exists in mammalian cells, we are using siRNA-mediated knockdown of KIF9 in COS-7 cells. Analysis of unsynchronized and cell-cycle synchronized...
Show moreKinesin motors bind to microtubules and function in mitosis and intracellular transport depending on the position of the motor domain within the primary sequence (Hirokawa and Noda 2008). KIF9 has recently been shown to be involved in MTOC positioning and mitotic entry in Dictyostelium (Tikhonenko et al. 2009). To determine if a similar role for KIF9 exists in mammalian cells, we are using siRNA-mediated knockdown of KIF9 in COS-7 cells. Analysis of unsynchronized and cell-cycle synchronized cells treated with siRNA to KIF9 reveal that the transition from G2 to M phase is delayed and that mitotic progression is also affected. Additionally, our data indicates that spindle pole function during anaphase may be abnormal in cells treated with siRNA, suggesting a role for KIF9 during that stage.
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Date Issued: 2010
PURL: http://purl.flvc.org/FAU/3334255
Subject Headings: Cells, Motility, Protoplasmic streaming, Cell organelles, Formation, Cellular signal transduction
Format: Document (PDF)

Title: Approaches for raising the level of FOXO3a in animal cells.
Creator: Navarro, Diana., Charles E. Schmidt College of Medicine, Department of Biomedical Science
Abstract/Description: The turtle is a unique model of anoxic survival. The turtle's brain can tolerate total oxygen deprivation for hours to days as well as prevent high levels of mitochondrial-derived free radicals upon re-oxygenation. Because of its ability to prevent elevated free radical generation, the turtle has also become recognized as a model of exceptional longevity. We are employing the turtle model for an investigation into the regulation of a key antioxidant enzyme system - methionine sulfoxide...
Show moreThe turtle is a unique model of anoxic survival. The turtle's brain can tolerate total oxygen deprivation for hours to days as well as prevent high levels of mitochondrial-derived free radicals upon re-oxygenation. Because of its ability to prevent elevated free radical generation, the turtle has also become recognized as a model of exceptional longevity. We are employing the turtle model for an investigation into the regulation of a key antioxidant enzyme system - methionine sulfoxide reductases (Msrs), primarily MsrA and MsrB. The Msr system is capable of reversing oxidation of methionines in proteins and Msr subtypes have been implicated in protecting tissues against oxidative stress, as well as, enhancing the longevity of organisms from yeast to mammals. Preliminary data, unpublished results, indicate that MsrA protein and transcripts are elevated by anoxia. A recent study on Caenorhabditis elegans demonstrated that FOXO is involved in activation of the MsrA promoter. Using the turtle MsrA promoter sequence we worked to determine which regions in the promoter are necessary for activation by anoxia. The results of the present study were 1) to prepare a TAT-FOXO3a fusion protein which could penetrate animal cells and 2) to construct a FOXO3a expression vector for transcription studies on MsrA expression.
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Date Issued: 2012
PURL: http://purl.flvc.org/FAU/3342243
Subject Headings: Cellular signal transduction, Cell proliferation, Oxidative stress, Prevention, Adaptation (Physiology)
Format: Document (PDF)

Title: Characterization of the MHC II B of the bald eagle.
Creator: Smith, Andrew., Charles E. Schmidt College of Science, Department of Biological Sciences
Abstract/Description: The Major Histocompatibility Complex class II B (MHC II B) gene encodes a protein that is part of the adaptive immune system and critical for the non-self recognition ability of immune cells. This gene has been characterized in the Bald Eagle, ten unique alleles were found in two subpopulations at the geographic extremes of the range margins. Geographic genetic variation is suggested by the presence of population specific alleles. The results showed considerable divergence of groups of Bald...
Show moreThe Major Histocompatibility Complex class II B (MHC II B) gene encodes a protein that is part of the adaptive immune system and critical for the non-self recognition ability of immune cells. This gene has been characterized in the Bald Eagle, ten unique alleles were found in two subpopulations at the geographic extremes of the range margins. Geographic genetic variation is suggested by the presence of population specific alleles. The results showed considerable divergence of groups of Bald Eagle alleles when compared to alleles from other birds of prey. Particular codons within the exon II show signs of balancing selection driving the evolution of the MHC II B. Transcription data showed statistically significant differential expression of alleles. This can be interpreted as meaning a particular locus is being preferentially expressed in blood. The analysis of the polymorphism of this adaptive marker may aid managers of wildlife during this age of global climate change and the biodiversity crisis.
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Date Issued: 2010
PURL: http://purl.flvc.org/FAU/2979375
Subject Headings: Major histocompatibility complex, Cellular signal transduction, Immunogenetics, Genetic polymorphisms
Format: Document (PDF)

Title: Function of glycinergic interplexiform cells in rod synaptic transmission.
Creator: Jiang, Zheng., Charles E. Schmidt College of Science, Department of Biological Sciences
Abstract/Description: The interplexiform cells(IP cells) are the most recently discovered neurons in the retina and their function is to provide centrifugal feedback in retina. The anatomical structure of the IP cells has been well studied, but the function of these neurons is largely unknown. I systematically studied the excitatory and inhibitory inputs from IP cells in salamander retina. I found that L-EPSCs in IP cells are mediated by AMPA and NMDA receptors; in addition, L-IPSCs are mediated by glycine...
Show moreThe interplexiform cells(IP cells) are the most recently discovered neurons in the retina and their function is to provide centrifugal feedback in retina. The anatomical structure of the IP cells has been well studied, but the function of these neurons is largely unknown. I systematically studied the excitatory and inhibitory inputs from IP cells in salamander retina. I found that L-EPSCs in IP cells are mediated by AMPA and NMDA receptors; in addition, L-IPSCs are mediated by glycine receptors and GABAC receptors. In response to light, IP cells reaction potentials transiently at the onset and onset of light stimulation. The major neural transmitter of IP cells in salamander retina is glycine. We also studied the distribution and function of glycine transporters. Our result indicates that GlyT1- and GlyT2-like transporters were present in Muller cells and neurons. The glycine feedback at outer plexiform layer (OPL) has effects on both the bipolar cell dendrites and rod photoreceptor terminals. At bipolar cell dendrites, glycine selectively depolarizes rod-dominant On-bipolar cells, and hyperpolarizes Off- bipolar cells. At rod photoreceptor terminals, 10 M glycine activates voltage-gated Ca2+ channels. These effects facilitated glutamate vesicle release in photoreceptors. It increases the sEPSC in OFF bipolar cells. The combined effect of glycine at rod terminals and bipolar cell dendrites leads to enhanced dim light signal transduction in the rod photoreceptor to ganglion cell pathway. This study provides a model that displays the function of centrifugal feedback through IP cells in the retina.
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Date Issued: 2009
PURL: http://purl.flvc.org/FAU/2708369
Subject Headings: Cellular signal transduction, Neurochemistry, Neuroplasticity, Synapses, Retina, Cytology, Visual pathways
Format: Document (PDF)

Title: GAD 65 and its role in pancreatic tissue survival.
Creator: Kumari, Neeta., Charles E. Schmidt College of Medicine, Department of Biomedical Science
Abstract/Description: We employed three genotypes of GAD 65, wildtype (GAD 65 +/+), heterozygous (GAD 65 +/-) and knockout (GAD 65 -/-) to investigate the role of GAD 65 in survival of pancreatic islets. We analyzed the mRNA expression of pro-survival proteins including Bcl2 and Bax in pancreas of wildtype, heterozygous and knockout using Reverse Transcriptase Polymerase Chain Reaction (RTPCR). The level of expression of Bcl2 mRNA was down regulated in knockout mice pancreas and Bax to Bcl2 ratio was found higher...
Show moreWe employed three genotypes of GAD 65, wildtype (GAD 65 +/+), heterozygous (GAD 65 +/-) and knockout (GAD 65 -/-) to investigate the role of GAD 65 in survival of pancreatic islets. We analyzed the mRNA expression of pro-survival proteins including Bcl2 and Bax in pancreas of wildtype, heterozygous and knockout using Reverse Transcriptase Polymerase Chain Reaction (RTPCR). The level of expression of Bcl2 mRNA was down regulated in knockout mice pancreas and Bax to Bcl2 ratio was found higher in knockout mice pancreas suggesting higher cell death rate. However, further studies are required to recognize and understand the specific connections between apoptotic pathways and GAD 65 in pancreatic islets.
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Date Issued: 2012
PURL: http://purl.flvc.org/FAU/3342206
Subject Headings: Glutamic acids, Antagonists, Cellular signal transduction, Glutamic acid, Metabolism
Format: Document (PDF)

Title: Discovery and visualization of co-regulated genes relevant to target diseases.
Creator: Lad, Vaibhan., College of Engineering and Computer Science, Department of Computer and Electrical Engineering and Computer Science
Abstract/Description: In this thesis, we propose to discover co-regulated genes using microarray expression data, as well as providing visualization functionalities for domain experts to study relationships among discovered co-regulated genes. To discover co-regulated genes, we first use existing gene selection methods to select a small portion of genes which are relevant to the target diseases, on which we build an ordered similarity matrix by using nearest neighbor based similarity assessment criteria. We then...
Show moreIn this thesis, we propose to discover co-regulated genes using microarray expression data, as well as providing visualization functionalities for domain experts to study relationships among discovered co-regulated genes. To discover co-regulated genes, we first use existing gene selection methods to select a small portion of genes which are relevant to the target diseases, on which we build an ordered similarity matrix by using nearest neighbor based similarity assessment criteria. We then apply a threshold based clustering algorithm named Spectral Clustering to the matrix to obtain a number of clusters. The genes which are clustered together in one cluster represent a group of co-regulated genes and to visualize them, we use Java Swings as the tool and develop a visualization platform which provides functionalities for domain experts to study relationships between different groups of co-regulated genes; study internal structures within each group of genes, and investigate details of each individual gene and of course for gene function prediction. Results are analyzed based on microarray expression datasets collected from brain tumor, lung cancers and leukemia samples.
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Date Issued: 2010
PURL: http://purl.flvc.org/FAU/2976447
Subject Headings: Genomics, Gene mapping, Cell transformation, Cellular signal transduction
Format: Document (PDF)

Title: Study of cell penetrating peptides with Raman spectroscopy and microscopy.
Creator: Ye, Jing., Charles E. Schmidt College of Science, Department of Chemistry and Biochemistry
Abstract/Description: Cell penetrating peptides (CPPs) have drawn the attention of researchers due to their ability to internalize large cargos into cells including cancer cells. The mechanism(s) with which the peptides enter the cell, however, is/are not clear and full of controversy. The peptide conformations and their microenvironment in live cells had been unknown until the development of a technique developed in our lab. As a first demonstration of principle, penetratin, a 16-residue CPP derived from the...
Show moreCell penetrating peptides (CPPs) have drawn the attention of researchers due to their ability to internalize large cargos into cells including cancer cells. The mechanism(s) with which the peptides enter the cell, however, is/are not clear and full of controversy. The peptide conformations and their microenvironment in live cells had been unknown until the development of a technique developed in our lab. As a first demonstration of principle, penetratin, a 16-residue CPP derived from the Antennapedia homeodomain protein of Drosophila, was measured in single, living melanoma cells. Carbon-13 labeling of the Phe residue of penetratin was used to shift the intense aromatic ring-breathing vibrational mode from 1003 to 967 cm-1, thereby enabling the peptide to be traced in cells. Difference spectroscopy and principal components analysis (PCA) were used independently to resolve the Raman spectrum of the peptide from the background cellular Raman signals., On the basis of the position of the amide I vibrational band in the Raman spectra, the secondary structure of the peptide was found to be mainly random coil and b-strand in the cytoplasm, and possibly assembling as b-sheets in the nucleus. Next, label-free transportan was studied with the same methodology. The peptide, besides predominantly a-helix, adopted a significant portion of b-sheet conformation in the cytoplasm and nucleolus, which is different from the peptide in aqueous solution. The peptide microenvironment was also probed through H-bonding reported by the tyrosine Fermi doublet. Transportan displayed a tendency to accumulate in the cytoplasm over time which was unlike penetratin, which concentrated in the nucleus. The relative concentration of CPPs in various locations of live melanoma cells was directly estimated from the Raman spectra using average Phe concentration in the cell as an internal standard., The rapid entry and almost uniform cellular distribution of both peptides, as well as the lack of correlation between peptide and lipid Raman signatures, indicated that the mechanism of CPP internalization under the conditions of study was probably non-endocytotic. Last, transportan and penetratin were studied using polarized Raman spectroscopy for more detailed vibrational spectroscopic information of the two peptides in water and TFE solutions. The majority of the bands in the Raman spectra of the peptides were highly polarized, consistent with the high symmetry of aromatic ring side chain vibrational bands dispersed throughout the spectra. This work has provided new insights into the structure of CPPs in live cells and in solutions.
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Date Issued: 2011
PURL: http://purl.flvc.org/FAU/3342344
Subject Headings: Peptides, Analysis, Infrared spectroscopy, Raman spectroscopy, Cellular signal transduction
Format: Document (PDF)

Title: RNA oxidative damage and ribosomal RNA surveillance under oxidative stress.
Creator: Liu, Min, Charles E. Schmidt College of Science, Department of Biological Sciences
Abstract/Description: We have studies oxidative damage of RNA, a major type of cellular macromolecules. RNA is a primary target of reactive oxygen species (ROS). Under oxidative stress, most nucleic acid damages in Escherichia coli (E.coli) are present in RNA as shown by high levels of 8-oxo-G, an oxidized form of guanine. Increased RNA oxidation is closely correlated to cell death under oxidative stress. Surprisingly, neither RNA structure nor association with proteins protects RNA from oxidation... Our results...
Show moreWe have studies oxidative damage of RNA, a major type of cellular macromolecules. RNA is a primary target of reactive oxygen species (ROS). Under oxidative stress, most nucleic acid damages in Escherichia coli (E.coli) are present in RNA as shown by high levels of 8-oxo-G, an oxidized form of guanine. Increased RNA oxidation is closely correlated to cell death under oxidative stress. Surprisingly, neither RNA structure nor association with proteins protects RNA from oxidation... Our results demonstrate a major role for RNA degradation in controlling oxidized RNA. We have identified activities that may work in specific pathways for selectively degrading damaged RNA. These activities may play pivotal rold in controlling oxidized RNA and protecting cells under oxidative stress.
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Date Issued: 2012
PURL: http://purl.flvc.org/FAU/3355620
Subject Headings: RNA, Metabolism, Cellular signal transduction, Genetic translation, Molecular biology
Format: Document (PDF)

Title: Potential therapies and neuroprotective cascades in anoxia tolerant freshwater turtle Trachemys scripta ellegans.
Creator: Nayak, Gauri., Charles E. Schmidt College of Science, Department of Biomedical Science
Abstract/Description: Mammalian neurons exhibit extreme sensitivity to oxygen deprivation and undergo rapid and irreversible degeneration when oxygen supply is curtailed. Though several neuroprotective pathways are activated during oxygen deprivation, their analyses are masked by the complex series of pathological events which are triggered simultaneously. Such events can be analyzed in the anoxia tolerant fresh water turtle, which can inherently survive the conditions of oxygen deprivation and post-anoxic...
Show moreMammalian neurons exhibit extreme sensitivity to oxygen deprivation and undergo rapid and irreversible degeneration when oxygen supply is curtailed. Though several neuroprotective pathways are activated during oxygen deprivation, their analyses are masked by the complex series of pathological events which are triggered simultaneously. Such events can be analyzed in the anoxia tolerant fresh water turtle, which can inherently survive the conditions of oxygen deprivation and post-anoxic reoxygenation without brain damage. It is likely in such a model that modulation of a particular molecular pathway is adaptive rather than pathological. The major objective behind this study was to analyze the intracellular signaling pathways mediating the protective effects of adenosine, a potential neuromodulator, and its effect on cell survival by influencing the key prosurvival proteins that prevent apoptosis. In vivo and in vitro studies have shown that adenosine acts as a neuroprotective metabolite and its action can be duplicated or abrogated using specific agonist and antagonists. Stimulating the adenosine receptors using selective A1 receptor agonist N6-cyclopentyladenosine (CPA) activated the presumed prosurvival ERK and P13-K/AKT cascade promoting cell survival, and suppression of the receptor using the selective antagonist DPCPX (8- cyclopentyl-1,3-dipropylxanthine) activated the prodeath JNK and P38 pathways. The complex regulation of the MAPK's/AKT signaling cascades was also analyzed using their specific inhibitors. The inhibiton of the ERK and AKT pathway increased cell death, indicating a prosurvival role, whereas inhibiton of the JNK and p38 pathway increased cell survival in this model. In vitro studies have also shown a high Bcl-2/BAX ratio during anoxia and reoxygenation, indicating a strong resistance to cell death via apoptosis., Silencing of the anti-apoptotic Bcl-2 gene using specific siRNA upregulated levels of prodeath BAX, thus altering the Bcl-2/BAX ratio and elevating cleaved Caspase-3 levels leading to increased cell death. Another promising neuroprotective target which we analyzed was Neuroglobin, which was induced during oxygen crisis and silencing this gene indicated that its plays a major role in modulation of ROS. This study strongly emphasizes the advantages of an alternate animal model in elucidating neuroprotective mechanisms and revealing novel therapeutic targets which could eventually help clinicians to design new stroke therapies based on naturally tolerant organisms.
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Date Issued: 2009
PURL: http://purl.flvc.org/FAU/186762
Subject Headings: Turtles, Physiology, Adenosine, Receptors, Cellular signal transduction, Molecular neurobiology, Apoptosis, Research, Cellular control mechanisms
Format: Document (PDF)

Title: An investigation of the role of PAK6 tumorigenesis.
Creator: Roberts, JoAnn, Charles E. Schmidt College of Medicine, Department of Biomedical Science
Abstract/Description: The function and role of PAK6, serine/threonone kinase, in cancer progressionhas not yet been clearly identified. Several studies reveal that PAK6 may participate in key changes contributing to cancer progression such as cell survival, cell motility, and invasiveness. Basedon the membrane localization of PAK6 in prostate and breast cancer cells,we speculated that PAK6 plays a rolein cancer progression cells by localizing on the membrane and modifying proteins linked to motility and...
Show moreThe function and role of PAK6, serine/threonone kinase, in cancer progressionhas not yet been clearly identified. Several studies reveal that PAK6 may participate in key changes contributing to cancer progression such as cell survival, cell motility, and invasiveness. Basedon the membrane localization of PAK6 in prostate and breast cancer cells,we speculated that PAK6 plays a rolein cancer progression cells by localizing on the membrane and modifying proteins linked to motility and proliferation. We isolated the raft domain of breast cancer cells expressing either wild type (WT), constitutively active (SN), or kinase dead PAK6 (KM) and found that PAK6 is a membrane associated kinase which translocates from the plasma membrane to the cytosol when activated. The downstream effects of PAK6 are unknown ; however, results from cell proliferation assays suggest a growth regulatory mechanism.
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Date Issued: 2012
PURL: http://purl.flvc.org/FAU/3356888
Subject Headings: Apoptosis, Cancer, Etiology, Cancer cells, Proliferation, Cellular signal transduction, Cellular control mechanisms, Cell cycle, Regulation
Format: Document (PDF)

Title: Synaptic Rearrangements and the Role of Netrin-Frazzled Signaling in Shaping the Drosophila Giant Fiber Circuit.
Creator: Lloyd, Brandon N., Murphey, Rodney K., Florida Atlantic University, Charles E. Schmidt College of Science, Department of Biological Sciences
Abstract/Description: In the developing CNS, presynaptic neurons often have exuberant overgrowth and form excess (and overlapping) postsynaptic connections. Importantly, these excess connections are refined during circuit maturation so that only the appropriate connections remain. This synaptic rearrangement phenomenon has been studied extensively in vertebrates but many of those models involve complex neuronal circuits with multiple presynaptic inputs and postsynaptic outputs. Using a simple escape circuit in...
Show moreIn the developing CNS, presynaptic neurons often have exuberant overgrowth and form excess (and overlapping) postsynaptic connections. Importantly, these excess connections are refined during circuit maturation so that only the appropriate connections remain. This synaptic rearrangement phenomenon has been studied extensively in vertebrates but many of those models involve complex neuronal circuits with multiple presynaptic inputs and postsynaptic outputs. Using a simple escape circuit in Drosophila melanogaster (the giant fiber circuit), we developed tools that enabled us to study the molecular development of this circuit; which consists of a bilaterally symmetrical pair of presynaptic interneurons and postsynaptic motorneurons. In the adult circuit, each presynaptic interneuron (giant fiber) forms a single connection with the ipsilateral, postsynaptic motorneuron (TTMn). Using new tools that we developed we labeled both giant fibers throughout their development and saw that these neurons overgrew their targets and formed overlapping connections. As the circuit matured, giant fibers pruned their terminals and refined their connectivity such that only a single postsynaptic connection remained with the ipsilateral target. Furthermore, if we ablated one of the two giant fibers during development in wildtype animals, the remaining giant fiber often retained excess connections with the contralateral target that persisted into adulthood. After demonstrating that the giant fiber circuit was suitable to study synaptic rearrangement, we investigated two proteins that might mediate this process. First, we were able to prevent giant fibers from refining their connectivity by knocking out highwire, a ubiquitin ligase that prevented pruning. Second, we investigated whether overexpressing Netrin (or Frazzled), part of a canonical axon guidance system, would affect the refinement of giant fiber connectivity. We found that overexpressing Netrin (or Frazzled) pre- & postsynaptically resulted in some giant fibers forming or retaining excess connections, while exclusively presynaptic (or postsynaptic) expression of either protein had no effect. We further showed that by simultaneously reducing (Slit-Robo) midline repulsion and elevating Netrin (or Frazzled) pre- & postsynaptically, we significantly enhanced the proportion of giant fibers that formed excess connections. Our findings suggest that Netrin-Frazzled and Slit-Robo signaling play a significant role in refining synaptic circuits and shaping giant fiber circuit connectivity.
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Date Issued: 2016
PURL: http://purl.flvc.org/fau/fd/FA00004758, http://purl.flvc.org/fau/fd/FA00004758
Subject Headings: Drosophila melanogaster--Cytogenetics., Genetic transcription., Transcription factors., Cellular signal transduction., Cellular control mechanisms., Cell receptors.
Format: Document (PDF)

Title: Representation of object-in-context within mouse hippocampal neuronal activity.
Creator: Asgeirsdottir, Herborg Nanna, Charles E. Schmidt College of Science, Department of Psychology
Abstract/Description: The rodent hippocampus is critical for processing spatial memory but its contribution to non-spatial, specifically object memory is debated. The cognitive map theory of hippocampal function states that the hippocampus stores relationships of goal locations (places) to discrete items (objects) encountered within environments. Dorsal CA1 place cells were recorded in male C57BL/6J mice performing three variations of the novel object recognition paradigm to define "object-in-context"...
Show moreThe rodent hippocampus is critical for processing spatial memory but its contribution to non-spatial, specifically object memory is debated. The cognitive map theory of hippocampal function states that the hippocampus stores relationships of goal locations (places) to discrete items (objects) encountered within environments. Dorsal CA1 place cells were recorded in male C57BL/6J mice performing three variations of the novel object recognition paradigm to define "object-in-context" representation of hippocampal neuronal activity that may support object memory. Results indicate, (i) that place field stability is higher when polarizing environmental cues are provided during object recognition; (ii) hippocampal place fields remain stable throughout the novel object recognition testing without a polarizing cue; and (iii) time dependent effects on stability when objects were dissociated from the context. These data indirectly support that the rodent hippocampus processes object memory, and challenge the view that "object-in-context" representations are formed when mice perform novel object recognition task.
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Date Issued: 2013
PURL: http://purl.flvc.org/fcla/dt/3362339
Subject Headings: Mice as laboratory animals, Hippocampus (Brain), Neurotransmitter receptors, Cellular control mechanisms, Cellular signal transduction
Format: Document (PDF)

Title: Highwire coordinates synapse formation and maturation by regulating both a map kinase cascade and the ability of the axon to respond to external cues in the giant fiber system of Drosophila Melanogaster.
Creator: Borgen, Melissa A., Murphey, Rodney K., Florida Atlantic University, Charles E. Schmidt College of Science, Department of Biological Sciences
Abstract/Description: The ubiquitin ligase Highwire is responsible for cell-autonomously promoting synapse formation in the Drosophila Giant Fiber system. highwire mutants show defects in synaptic function and extra branching at the axon terminal, corresponding to transient branching that occur in the course of giant synapse formation during metamorphosis. The MAP kinase pathway, including Wallenda and JNK/Basket, plus the transcription factor Jun, act to suppress synaptic function and axon pruning in a dosage...
Show moreThe ubiquitin ligase Highwire is responsible for cell-autonomously promoting synapse formation in the Drosophila Giant Fiber system. highwire mutants show defects in synaptic function and extra branching at the axon terminal, corresponding to transient branching that occur in the course of giant synapse formation during metamorphosis. The MAP kinase pathway, including Wallenda and JNK/Basket, plus the transcription factor Jun, act to suppress synaptic function and axon pruning in a dosage sensitive manner, suggesting different molecular mechanisms downstream of the MAP kinase pathway govern function and pruning. A novel role for Highwire is revealed, regulating the giant fiber axon’s ability to respond to external cues regulated by Fos. When expression of the transcription factor Fos is disrupted in the post-synaptic TTMn or surrounding midline glia of highwire mutants, the giant fiber axons show a marked increase in axon overgrowth and midline crossing. However, synaptic function is rescued by the cell nonautonomous manipulation of Fos, indicating distinct mechanisms downstream of Highwire regulating synaptic function and axon morphology.
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Date Issued: 2014
PURL: http://purl.flvc.org/fau/fd/FA00004081, http://purl.flvc.org/fau/fd/FA00004081
Subject Headings: Cell differentiation, Cellular control mechanisms, Cellular signal transduction, Drosophila melanogaster -- Cytogenetics, Gene expression, Genetic transcription
Format: Document (PDF)

Title: Netrin-Frazzled signaling instructs synaptogenesis and plasticity at an identified central synapse in Drosophila.
Creator: Orr, Brian, Murphey, Rodney K., Charles E. Schmidt College of Science, Department of Biological Sciences
Abstract/Description: The classic guidance molecules, Netrin and its receptor Frazzled (Fra), dictate the strength of synaptic connections in the giant fiber system (GFS) of Drosophila melanogaster by regulating gap junction localization in the pre-synaptic terminal. In Netrin mutant animals the synaptic coupling between a giant interneuron and the jump motor neuron was weakened. Dye-coupling between these two neurons was severely compromised or absent. These mutants exhibited anatomically and physiologically...
Show moreThe classic guidance molecules, Netrin and its receptor Frazzled (Fra), dictate the strength of synaptic connections in the giant fiber system (GFS) of Drosophila melanogaster by regulating gap junction localization in the pre-synaptic terminal. In Netrin mutant animals the synaptic coupling between a giant interneuron and the jump motor neuron was weakened. Dye-coupling between these two neurons was severely compromised or absent. These mutants exhibited anatomically and physiologically defective synapses between the giant fiber (GF) and tergotrochanteral motor neuron (TTMn). In cases where Netrin mutants displayed apparently normal synaptic anatomy, half of the specimens exhibited physiologically defective synapses. Dye-coupling between the giant fiber and the motor neuron was reduced or eliminated, suggesting that gap junctions were disrupted in the Netrin mutants. When we examined the gap junctions with antibodies to Shaking-B Innexin (ShakB), they were significantly decreased or absent in the pre-synaptic terminal of the mutant GF. This data is the first to show that Netrin and Frazzled regulate placement of gap junctions pre-synaptically at a central synapse. In the Drosophila Giant Fiber System, we demonstrate a mechanism that ensures the monoinnervation of two homologous motor neurons by two homologous interneurons. In a scenario where both interneurons could synapse with both motor neuron targets, each interneuron exclusively synapsed with only one target and the circuit functions at normal physiological levels. This innervation pattern depended on the ratio of netrin-to-frazzled expression. When Netrin was over expressed in the system, shifting the ratio in favor of Netrin, both interneurons synapsed with both target motor neurons and physiological function was reduced. This resulted in the polyinnervationof a single target. In contrast, when Frazzled was over expressed in the system, one interneuron innervated both targets and excluded the remaining interneuron from making any synaptic contact. This resulted in a single interneuron mono-innervating both motor neurons and physiological function was mutant. The orphaned interneuron made no synaptic contact with either motor neuron target. Physiological function was only normal when the Netrin-Frazzled ratio was at endogenous levels and each GF monoinnervated one motor neuron. When we examined the gap junctions at this synapse in experimental animals, there was a significant reduction of gap junction hemichannels in the presynaptic terminal of axons that deviated from normal innervation patterns. While the synapse dyecoupled, the reduction in gap junction hemichannels reduced function in the circuit.
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Date Issued: 2013
PURL: http://purl.flvc.org/fau/fd/FA0004041
Subject Headings: Cellular control mechanisms, Cellular signal transduction, Drosophila melanogaster -- Cytogenetics, Genetic transcription, Transcription factors
Format: Document (PDF)

Title: Highwire's characterization and signaling roles in Drosophila central synapse formation.
Creator: Rowland, Kimberly Diane., Charles E. Schmidt College of Science, Department of Biological Sciences
Abstract/Description: The assembly and maintenance of central synapses is a complex process, requiring myriad genes and their products. Highwire is a large gene containing a RING domain, characteristic of ubiquitin E3 ligases. Highwire has been shown to restrain axon growth and control synaptogenesis at a peripheral synapse. Here I examine the roles of Highwire at a central synapse in the adult Drosophila Giant Fiber System (GFS). Highwire is indeed necessary for proper axonal growth as well as synaptic...
Show moreThe assembly and maintenance of central synapses is a complex process, requiring myriad genes and their products. Highwire is a large gene containing a RING domain, characteristic of ubiquitin E3 ligases. Highwire has been shown to restrain axon growth and control synaptogenesis at a peripheral synapse. Here I examine the roles of Highwire at a central synapse in the adult Drosophila Giant Fiber System (GFS). Highwire is indeed necessary for proper axonal growth as well as synaptic transmission in the GFS. Differences arise between the central synapse and the neuromuscular junction (NMJ), where highwire was initially characterized : expresion from the postsynaptic cell can rescue highwire synaptic defects, which is not seen at the NMJ. In addition, a MAP kinase signaling pathway regulated by highwire at the NMJ has differing roles at a central synapse. Wallenda MAPK can rescue not only the highwire anatomical phenotype but also the defects seen in transmission. Another distinction is seen here : loss of function basket and Dfos enhance the highwire anatomical phenotype while expression of dominant negative basket and Dfos suppress the highwire phenotype. As a result we have compared the signaling pathway in flies and worms and found that the NMJ in the two organisms use a parallel pathway while the central synapse uses a distinct pathway.
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Date Issued: 2012
PURL: http://purl.flvc.org/FAU/3352826
Subject Headings: Cellular control mechanisms, Cellular signal transduction, Cell differentiation, Gene expression, Genetic transcription, Transcription factors, Drosophila melanogaster, Cytogenetics
Format: Document (PDF)

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