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Expression And Function Of Alpha3 And Beta2 Neuronal Nicotinic Acetylcholine Receptor Subunits In Hek-293 Cells, Nathan W. Steinhafel
Expression And Function Of Alpha3 And Beta2 Neuronal Nicotinic Acetylcholine Receptor Subunits In Hek-293 Cells, Nathan W. Steinhafel
Theses and Dissertations
Single-cell real-time quantitative RT-PCR was used to characterize the mRNA expression of rat neuronal nicotinic acetylcholine receptor (nAChR) subunits α3 and β2 in CA1 hippocampus stratum radiatum and stratum oriens interneurons. α3β2 co-expression was detected in 43% of interneurons analyzed. The nAChR subtype α3β2 was transiently expressed in cells derived from the human embryonic kidney cell line 293 at mRNA levels found in the CA1. The functional properties of α3β2 in HEK-293 cells were characterized by whole-cell patch clamping using acetylcholine (ACh) as an agonist. The kinetics of α3β2 channels were further analyzed by altering the level of α3 DNA …
The Role Of Wnt Signaling In Development Of The Ophthalmic Trigeminal Placode., Rhonda Nicole Lassiter
The Role Of Wnt Signaling In Development Of The Ophthalmic Trigeminal Placode., Rhonda Nicole Lassiter
Theses and Dissertations
Cranial placodes are ectodermal regions that contribute extensively to the vertebrate peripheral nervous system. The development of the ophthalmic trigeminal (opV) placode, which gives rise only to sensory neurons of the ophthalmic lobe of the trigeminal ganglion, is a well-studied model of sensory neuron development. While key differentiation processes have been characterized at the tissue and cellular levels, the molecules governing opV placode development have not been well described. This study identifies the canonical Wnt signaling pathway as a regulator of opV trigeminal placode development. Introducing dominant-negative TCF and dominant-active β-catenin expression constructs by in ovo electroporation, we have manipulated …
The Influence Of Membrane Lipid Order On Cell Shape And Microvesiculation In Human Erythrocytes, Laurie Jackson Gonzalez
The Influence Of Membrane Lipid Order On Cell Shape And Microvesiculation In Human Erythrocytes, Laurie Jackson Gonzalez
Theses and Dissertations
Exposure of human erythrocytes to elevated intracellular calcium causes alterations in cell shape and stimulates shedding of the cell membrane in the form of microvesicles. We hypothesized that both the shape transition and microvesiculation are influenced by microscopic membrane physical properties such as lipid order. To test this hypothesis, membrane properties were manipulated by varying the experimental temperature, membrane cholesterol content, and the internal ionic environment. Changes in membrane order were assessed using steady-state fluorescence spectroscopy with an environment-sensitive probe, laurdan. Our observations led us to the following conclusions: 1) the modest temperature dependence of membrane structure observed with laurdan …
Fgf4 And Wnt5a/Pcp Signaling Promote Limb Outgrowth By Polarizing Limb Mesenchyme, Keri Lynn Low
Fgf4 And Wnt5a/Pcp Signaling Promote Limb Outgrowth By Polarizing Limb Mesenchyme, Keri Lynn Low
Theses and Dissertations
The focus of this study was to elucidate the molecular and cellular mechanisms whereby fibroblast growth factors (FGFs) mediate outgrowth of the limb. Specifically, we examined the epistatic relationship between FGF and Wnt/Planar cell polarity (PCP) signaling in establishing cell polarity as a mechanism for outgrowth. By implanting beads into embryonic limbs and lateral plate mesoderm, we established that FGF activates Wnt5a in a gradient fashion. Once it was established that Wnt5a was expressed at the right time and place to turn on PCP signaling, we investigated the ability of Wnt5a to influence cell migration and/or cell polarity. Our analysis …
Construction Of A Col11a1 Transgene Vector, Cameron Mckell Beck
Construction Of A Col11a1 Transgene Vector, Cameron Mckell Beck
Theses and Dissertations
Background: Cartilage disorders affect millions of people in the United States alone, with effects ranging from poor skeletal development and joint pain to shortened lifespan and perinatal lethality. Many of these disorders have their root in defects of collagen, type XI collagen being among the most important. A mouse model of such a type XI collagen defect is the chondrodysplasia (cho) mutant. Mice homozygous for this null mutation in the Col11a1 gene do not express the α1 chain of type XI collagen. This results in a functional knockout of type XI collagen, leading to insufficient skeletal development and perinatal lethality. …
Pka As An Upstream Kinase For Lkb1/Strad/Mo25, Seth Taylor Herway
Pka As An Upstream Kinase For Lkb1/Strad/Mo25, Seth Taylor Herway
Theses and Dissertations
The LKB1/STRAD/MO25 complex (LSMK) has been identified as the major upstream kinase for AMP-activated protein kinase (AMPK). PKA phosphorylates LKB1 at the Ser428 residue in humans and Ser431 residue in mice. We investigated PKA as an upstream kinase for LSMK. LKB1 that had been incubated with PKA prior to incubation with AMPK experienced up to a 51% increase in AMPK Kinase activity compared to LKB1 alone (p < 0.05). When blocked with a PKA Inhibitor, the kinase effect of PKA on LKB1 was eliminated. Rat epitrochlearis muscle tissue incubated with epinephrine experienced no increase in AMPK activity compared with controls indicating that epinephrine does not cause AMPK activity in this type of tissue. In conclusion, phosphorylation by PKA can increase the AMPKK activity of LKB1-STRAD-MO25 in vitro. Because LKB1 has been found to be constitutively active, it is postulated that phosphorylation by PKA may act to enhance LKB1-AMPK interaction and thus achieve its effect.
The Effects Of 3-Phosphoglycerate And Other Metabolites On The Activation Of Amp-Activated Protein Kinase By Lkb1/Strad/Mo25, William John Ellingson
The Effects Of 3-Phosphoglycerate And Other Metabolites On The Activation Of Amp-Activated Protein Kinase By Lkb1/Strad/Mo25, William John Ellingson
Theses and Dissertations
Skeletal muscle contraction results in the phosphorylation and activation of the AMP-activated protein kinase (AMPK) by an upstream kinase, AMPKK. The LKB1-STRAD-MO25 complex is the major AMPKK in skeletal muscle; however, LKB1-STRAD-MO25 activity is not increased by muscle contraction. This relationship suggests that phosphorylation of AMPK by LKB1-STRAD-MO25 during skeletal muscle contraction may be regulated by allosteric mechanisms. In this study we tested an array of metabolites including glucose-6-phosphate (G6P), fructose-6-phosphate (F6P), fructose 1,6-bisphosphate (F1,6-P2), 3-phosphoglycerate (3PG), glucose-1-phosphate (G1P), glucose-1,6-bisphosphate (G1,6-P2), adenosine diphosphate (ADP), carnitine (Carn), acetyl-carnitine (Acarn), inosine monophosphate (IMP), inosine, and ammonia for allosteric regulation. We found that …
Properties Of Conductance And Inhibition Of Proton Channels: M2 From Influenza A Virus And Fo From Escherichia Coli Atp Synthase, Jeffrey C. Moffat
Properties Of Conductance And Inhibition Of Proton Channels: M2 From Influenza A Virus And Fo From Escherichia Coli Atp Synthase, Jeffrey C. Moffat
Theses and Dissertations
Proton channels are essential for many of the processes of life. The influenza A viral protein M2 is responsible for sensing the conditions necessary for viral RNA release. The proton-translocating FoF1 ATPase (ATP synthase) uses a proton gradient to drive adenosine triphosphate (ATP) synthesis. We have directly measured proton uptake in vesicles containing reconstituted M2 or FO by monitoring external pH after addition of valinomycin to vesicles with 100-fold diluted external [K+]. This proton flux assay was utilized to quantify proton flux through single M2 and Fo channels. Contrary to previous reports, proton uptake by M2 was not significantly altered …
The Role Of Fgfr4 In Trigeminal Placode Cell Development, Stephanie Beth Reynolds
The Role Of Fgfr4 In Trigeminal Placode Cell Development, Stephanie Beth Reynolds
Theses and Dissertations
In vertebrate embryos, the ophthalmic trigeminal (opV) placode contributes sensory neurons to the trigeminal ganglion during development of the peripheral nervous system. FGFR4 is expressed transiently in the trigeminal placode as cells undergo an epithelial-mesenchymal transition and begin to migrate toward the condensing ganglion. Because of the restricted spatiotemporal expression of FGFR4 in the opV placode, it was hypothesized that FGFR4 is necessary for the process of epithelial-mesenchymal transition in the opV placode. Using electroporation, an FGFR4 inhibitory gene construct was introduced into 6—10 somite stage chick embryos. This secreted inhibitory form of the FGFR4 gene blocked endogenous FGFR4 signaling, …
The Role Of Pax3 In Neuronal Differentiation Of The Ophthalmic (Opv) Trigeminal Placode And Neural Tube During Chicken Embryonic Development, James R. Bradshaw
The Role Of Pax3 In Neuronal Differentiation Of The Ophthalmic (Opv) Trigeminal Placode And Neural Tube During Chicken Embryonic Development, James R. Bradshaw
Theses and Dissertations
Pax3 has been used as a valuable marker in research aimed at understanding tissue interactions involved in trigeminal ophthalmic (opV) placode development. While Pax3 expression coincides with opV neuron specification, the function of Pax3 in these cells has not previously been investigated. Splotch mutant mice (which lack Pax3) have a reduced trigeminal ganglion; however it is not clear whether this reduction is due to neural crest or placode cells. We have used electroporation in the chick model system to block or ectopically express Pax3 at key times in opV placode development. Using several markers of placode cell differentiation, we have …
Apparent Digestibility In Lamas (Lama Glama), Fed With Brazilian Grass And Wheat In The C.E.A.C., William Edson Velez Contacayo
Apparent Digestibility In Lamas (Lama Glama), Fed With Brazilian Grass And Wheat In The C.E.A.C., William Edson Velez Contacayo
Theses and Dissertations
Considering the activity of introducing genetic resources with a high potential for yield, and with a wide adaptation range, it is possible to strengthen the cattle production system, through the production of adequate forage in quality, and quantity. Brazilian grass was introduced to Bolivia in 1974 for demonstrating excellent forage characteristics, and great adaptation qualities to the different ecosystems of the Andes. The material was introduced through the Experimental Station of Patacamaya, with their material supplied through the Experimental Station of Obonuco (Colombia). The following study was conducted at the Agropecuary Experimental Center of Condori, located 49 km. north of …
The Effects Of Hemoglobin-Based Oxygen Carriers On Mean Arterial Pressure, Arteriolar Diameter, And Nitric Oxide In The Microcirculation, Veronique C. Hionis
The Effects Of Hemoglobin-Based Oxygen Carriers On Mean Arterial Pressure, Arteriolar Diameter, And Nitric Oxide In The Microcirculation, Veronique C. Hionis
Theses and Dissertations
In the US today, blood transfusion is safer than ever. Nevertheless, the century-old quest for a suitable blood substitute persists. The elimination of unwanted side effects, especially transfusion-transmitted diseases, the problems and high cost factor involved in collecting and storing human blood, the pending worldwide shortages, and the need for compatibility testing are the driving forces contributing towards the development of blood substitutes. The leading research is focusing on hemoglobin-based oxygen carriers (HBOCs), which are limited in clinical application due to the pressor effect they induce. In this study, the mechanisms through which HBOCs affect mean arterial pressure (MAP), arteriolar …
The Effect Of Adding Drag-Reducing Polymers To Resuscitation Fluid During Hemorrhagic Shock On Skeletal Muscle Microcirculation, Geoffrey C. Alexander
The Effect Of Adding Drag-Reducing Polymers To Resuscitation Fluid During Hemorrhagic Shock On Skeletal Muscle Microcirculation, Geoffrey C. Alexander
Theses and Dissertations
Previous studies have shown an increase in survival when a minute amount of drag-reducing polymers were added to a resuscitation fluid. The purpose of this investigation was to examine the effect of adding a minute amount of the drag-reducing polymer polyethylene glycol to a resuscitation fluid, on the microcirculation of skeletal muscle during a volume-controlled hemorrhage model. The spinotrapezius muscle in twelve male Sprague Dawley rats was exteriorized for microvascular measurements of the arterioles. The diameters of the three levels of arterioles, interstitial fluid PO2, and RBC velocity in the feed arteriole were measured. Flow in the feed arteriole was …
Isolation Of Mesenchymal Stem Cells Derived From Adult Bone Marrow And Umbilical Cord Blood And Their Potential To Differentiate Into Osteoblasts, Andrew P. Pacitti
Isolation Of Mesenchymal Stem Cells Derived From Adult Bone Marrow And Umbilical Cord Blood And Their Potential To Differentiate Into Osteoblasts, Andrew P. Pacitti
Theses and Dissertations
The demand for treatment strategies of musculoskeletal tissues is continuously growing, especially considering the increasing number of elderly people with degenerative diseases of the skeletal system. Despite major strides in the field of bone regenerative medicine during the years, current therapies, such as bone grafts, still have several limitations. Multipotent stem cells, such as mesenchymal stem cells (MSCs) are promising candidates for tissue repair because of their differentiation potential and their capacity to undergo extensive replication. However, isolating a homogeneous population of MSCs from multiple sources is an area that needs to be addressed. Also, the knowledge regarding the mechanisms …
Effect Of Nitric Oxide On Oxygen Consumption Of Skeletal Muscle, Christina Lyn Cox
Effect Of Nitric Oxide On Oxygen Consumption Of Skeletal Muscle, Christina Lyn Cox
Theses and Dissertations
Mammalian cells require a continuous and sufficient supply of oxygen to carry out their functions. The oxygen pathway has an overall direction taking O2 from the air to the mitochondria, which is a result of the mitochondrial O2 consumption (VO2) NO various effects on the mitochondria: at low concentrations for short periods NO specifically and irreversibly inhibits cytochrome c and reversibly inhibits cytochrome c oxidase, to decrease VO2. Thus, NO can modulate VO2 of skeletal muscle. The purpose of the present study was to measure VO2 of the rat spinotrapezius muscle under conditions of altered NO. The methods used provide …
Early Mediators Of Cutaneous Ischemia Reperfusion Injury: A Mouse Model, Christopher Rogers Pruitt
Early Mediators Of Cutaneous Ischemia Reperfusion Injury: A Mouse Model, Christopher Rogers Pruitt
Theses and Dissertations
Chronic tissue injuries present an enormous problem to both patients and healthcare professionals, and yet little is definitively known as to the underlying pathophysiology. While there are numerous comorbidities associated with these wounds, a unifying theme has been proposed to be the repetitive incidence of ischemia reperfusion injury.A mouse model of cyclic ischemia reperfusion was employed to examine the initiation events in the pathology of chronic wounds. Mice were subjected to eight hours of magnetic skin compression via four cycles of two-hours ischemia and thirty minutes of reperfusion. The presence of neutrophil markers of oxidative stress and inflammation such as …
Functional Consequences Of Matrix Metalloproteinase-1 Over-Expression In Human Gliomas, Emily Mullet
Functional Consequences Of Matrix Metalloproteinase-1 Over-Expression In Human Gliomas, Emily Mullet
Theses and Dissertations
Malignant brain tumors are among the deadliest of human cancers. Despit recent advancements in conventional therapies, glioblastomas remain incurable, largel y due to their ability to invade surrounding tissue. Matrix metalloproteinases are thought to contribute to the invaseive phenotype of human gliomas. Absent in normal brain, matrix metalloproteinase-1 (MMP-1) has been shown to be present in gliomas, and in particular in glioblastoma multiforme (GBM). To begin to examine the role of MMP-1 in these tumore, two human glioma cell lines were stably transfected with MMP-1 cDNA. Confirmation of MMP-1 over-expression in these cells was achieved through real-time PCR and Western …
Comparison Of Leukocyte Classes Most Likely To Cause Vascular Dysfunction In Preeclampsia, Kristen Anne Cadden
Comparison Of Leukocyte Classes Most Likely To Cause Vascular Dysfunction In Preeclampsia, Kristen Anne Cadden
Theses and Dissertations
Leukocytes are activated in women with preeclampsia, but the class of leukocyte most likely to cause vascular dysfunction is not known. We hypothesized that neutrophils may be the class of leukocyte most involved in causing this dysfunction because neutrophils are the most abundant of the leukocytes and their numbers increase during pregnancy. In this study we compared vascular infiltration of neutrophils (CD66b) with monocytes/macrophages (CD14) and lymphocytes (CD99) in normal non-pregnant women (NNP), normal pregnant women (NP), and preeclamptic women (PE). There was no significant difference in the infiltration of lymphocytes into the maternal vasculature in PE as compared to …
Anatomical Bases For Auditory Projections To Suprasylvian Visual Areas In The Cat Cerebral Cortex, Giriaj K. Sharma
Anatomical Bases For Auditory Projections To Suprasylvian Visual Areas In The Cat Cerebral Cortex, Giriaj K. Sharma
Theses and Dissertations
How the neural representations of different sensory modalities transition from one to another is an unexplored issue of cortical organization. The present experiments addressed this problem by examining auditory projections to the lateral suprasylvian visual area (LSS) of the cat using neuroanatomical tract tracing methods. Injections of tracer (Biotinylated Dextran Amine, BDA) were made into defined areas of the cat auditory cortex. Following transport and tissue processing, a light microscope with a digitizing stage was used to visualize and plot labeled auditory projections to the LSS. The results showed that all auditory cortices tested demonstrated projections to a restricted portion …
Energy Value In Llamas (Lama Glama) Fed With Alfalfa Hay (Modicago Sativa) And Barley (Hordeum Vulgare) For Apparent Digestibility In Vivo In The National Camelid Germplasm Bank (Ceac), Edwin Gonzalo Ugarte Huanca
Energy Value In Llamas (Lama Glama) Fed With Alfalfa Hay (Modicago Sativa) And Barley (Hordeum Vulgare) For Apparent Digestibility In Vivo In The National Camelid Germplasm Bank (Ceac), Edwin Gonzalo Ugarte Huanca
Theses and Dissertations
In this experiment, we used four male llamas that were three to four years of age from the Khara race. They were divided into two ad libitum feeding groups. The digestible energy (ED) was determined using samples that were sent to the laboratory. These samples were derived from a conventional in vivo assay using the method of total fecal collection in order to determine the difference between energy consumed and energy excreted. Metabolizable energy was calculated by deducting digestible energy, energy loss in the urine and energy loss in the gases (methane). Urinary energy was determined through the measurement of …