Browsing by Subject "Homeostasis"
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Item Activity-Dependent Regulation of Inhibition from Different Inhibitory Subtypes(2007-08-08) Bartley, Aundrea Frances; Gibson, Jay R.Neuronal activity, in the form of action potential firing, is critical in the maturation and maintenance of neocortical circuitry. A negative feedback mechanism by which neuronal circuits adapt to changing levels of average activity on a time scale of hours to days is known as homeostatic plasticity. At the simplest level, homeostatic adaptations occur to maintain firing rate of neurons at a particular set-point. To better understand homeostatic plasticity at the network level, one must understand the activity-dependent adaptations that occur in the different neocortical cells types. To this end, I examined the regulation of inhibitory neurons and their synapses. I used chronic pharmacological block of activity in a neocortical slice cultures to examine the role activity plays in regulating feedback inhibition defined by two biochemical inhibitory neuron subtypes - parvalbumin-positive (Parv+) and somatostatin-positive (Som+). The cellular and synaptic components of local feedback inhibition were examined. I found that chronic activity blockade caused the following: 1) an increase in the intrinsic excitability of Som+ neurons through the downregulation of 2 substhreshold currents. While not thoroughly examined in Parv+ neurons, a similar, but weaker, increase in excitability may occur in these neurons as well. These< changes are consistent with a homeostatic maintenance of firing rate in these neurons. 2) a differential regulation of monosynaptic inhibition based on subtype that was frequency dependent. At low frequency action potential firing, Parv+ mediated inhibitory drive was downregulated while Som+ was unchanged. Both subtypes were likely downregulated at high frequency firing. 3) an upregulation of excitatory drive onto both Parv+ and Som+ neurons. This was most dramatic at low frequency firing where both subtypes displayed an almost 3-fold increase. This is also consistent with homeostatic maintenance of firing rate in inhibitory neurons. 4) based on the above, a clear change in recurrent inhibition occurred at low frequency firing. First, net recurrent inhibition was increased for both subtypes, but the relative influence of the two changed, such that Som+ recurrent inhibition contributed more relative to that of Parv+ circuitry. At high frequency firing, a slight, but less resolvable, increase in net recurrent inhibition may have occurred in both subtypes without any change in relative contribution. 5) all of the synaptic changes were likely due to increases in presynaptic release probability and/or decreases in synapse number.Item Activity-dependent regulation of ion channel gene expression: a homeostatic hypothesis for drug tolerance(2006) Ghezzi, Alfredo; Atkinson, Nigel (Nigel S.)Conservation of the balance between excitation and inhibition of neural activity is critically important for the proper function of the nervous system. Upon alterations in excitability, the nervous system may thus trigger mechanisms that attempt to restore homeostasis. Many alcohols, anesthetics, and other abused volatile solvents such as ethanol, benzyl alcohol, toluene, trichloroethylene, and chloroform alter neural excitability and trigger homeostatic adaptations that through the modulation of gene expression and signaling between nerve cells act to counteract these alterations. Many of these adaptations may account for the development of tolerance, dependence and addiction to these drugs. Here, I demonstrate that in Drosophila, tolerance to the sedative effects of alcohols and anesthetics, is mediated by an increase in expression of the Ca2+ activated K+ channel gene, slowpoke. A mutation that eliminates slowpoke expression prevents tolerance, while expression from an inducible slowpoke transgene mimics tolerance in naive animals. Furthermore, the behavioral and molecular response to volatile solvents can be separated into an initial phase of hyperkinesis that causes a drop in slowpoke gene expression and makes animals more sensitive to subsequent sedation by these drugs, and a sedative phase that stimulates slowpoke gene expression and induces tolerance. This demonstrates that the changes in expression level of slowpoke act as a modulator of drug sensitivity. Because of it's central role as a regulator of electrical activity in nerve terminals this channel gene is a likely contributor to the homeostatic mechanism that resists untoward changes in net cellular excitability and mediates tolerance to sedation. If hyperexcitability is induced, the proposed mechanism alters channel expression to reduce this excitability, whereas if cellular excitability is suppressed, channel gene expression changes to enhance excitability. An electrophysiological test of this hypothesis shows that increased slowpoke expression enhances the excitability of a neural pathway in a way that opposes the effects of sedative drugs. This data supports the notion that slowpoke mediated tolerance to sedation is part of a homeostatic adaptation that compensates for changes in neural activity caused by drugs and represents a step forward in the understanding of the molecular basis of drug addiction.Item Calcium-mediated change in neuronal intrinsic excitability in weakly electric fish: biasing mechanisms of homeostatis for those of plasticity(2009-12) George, Andrew Anthony; Zakon, H. H.; Aldrich, Richard W.; Atkinson, Nigel S.; Mihic, S. John; Golding, Nace L.; Dalby, Kevin N.Although the processes used for temporarily storing and manipulating neural information have been extensively studied at the synaptic level far less attention has been given to the underlying cellular and molecular mechanisms that contribute to change in the intrinsic excitability of neurons. More importantly, how do these mechanisms of plasticity integrate with ongoing mechanisms of regulation of neural intrinsic excitability and, in turn, homeostasis of entire neural circuits? In this dissertation I describe the underlying mechanisms that contribute to persistent neural activity and, more globally, sensorimotor adaptation using weakly electric fish as my model system. Weakly electric fish have evolved a behavior adaptation known as the jamming avoidance response (JAR), and it is this adaptation that allows the organism to elevate its own electrical discharge in response to intraspecific interactions and subsequent distortions of the animal’s electric field. The elevation operates over a wide range and in vivo can last tens of hours upon cessation of a jamming stimulus. I demonstrate that the underlying mechanisms of the adaptation are mediated by calcium-dependent signaling in the pacemaker nucleus and that calcium-mediated phosphorylation plays an important role in the regulation of the long-term frequency elevation (LTFE). I demonstrate using an in vitro brain slice preparation from the weakly electric fish, Apteronotus leptorhynchus that the engram of memory formation depends on the cooperativity of calcium-dependent protein kinases and protein phosphatases. In addition, I show that the memory formation (in the form of LTFE) does not depend on the continued flux of calcium, but rather the phosphorylation events downstream of NMDA receptor activation. Moreover, I describe the differences in the expression of protein phosphatases and protein kinases as they relate to species-specific differences in sensorimotor adaptation. It is important to note that this is the first time that the cooperativity between different isoforms of protein kinase C (PKC) have been shown to play a role in graded long-term change in neuronal activity and, in turn, providing the neural basis of species-specific behavior. The neural adaptation of the electromotor system in weakly electric fish provides an excellent model system to study the underlying cellular and molecular events of vertebrate memory formation.Item Homeostasis and synaptic scaling : a theoretical perspective(2012-12) Corey, Joseph Harrod; Priebe, Nicholas; Cormack, Lawrence; Huk, Alexander; Pillow, JonathanAbstract The synaptic input received by neurons in cortical circuits is in constant flux. From both environmental sensory changes and learning mechanisms that modify synaptic strengths, the excitatory and inhibitory signals received by a post-synaptic cell vary on a continuum of time scales. These variable inputs inherent in different sensory environments, as well as inputs changed by Hebbian learning mechanisms (which have been shown to destabilize the activity of neural circuits) serve to limit the input ranges over which a neural network can effectively operate. To avoid circuit behavior which is either quiescent or epileptic, there are a variety of homeostatic mechanisms in place to maintain proper levels of circuit activity. This article provides a basic overview of the biological mechanisms, and consider the advantages and disadvantages of homeostasis on a theoretical level.Item Mechanisms of benzyl alcohol tolerance in Drosophila melanogaster(2009-12) Alhasan, Yazan Mahmoud; Atkinson, Nigel (Nigel S.); Zakon, Harold H.; Gonzales, Rueben A.; Singer, Michael C.; Bergeson, Susan E.Proper neuronal function requires the preservation of appropriate neural excitability. An adaptive increase in neural excitability after exposure to agents that depress neuronal signaling blunts the sedative drug effects upon subsequent drug exposure. This adaptive response to drug exposure leads to changes in drug induced behaviors such as tolerance, withdrawal and addiction. Here I use Drosophila melanogaster to study the cellular and neuronal components which mediate behavioral tolerance to the anesthetic benzyl alcohol. I demonstrate that rapid tolerance to benzyl alcohol is a pharmacodynamic mechanism independent of drug metabolism. Furthermore, tolerance is a cell autonomous response which occurs in the absence of neural signaling. Using genetic and pharmacological manipulations I find the synapse to play an important role in the development of tolerance. In addition, the neural circuits that regulate arousal and sleep also alter benzyl alcohol sensitivity. Beyond previously described transcriptional mechanisms I find a post-translational role of the Ca2+-activated K+-channel, slowpoke in the development of tolerance. Finally, I explore a form of juvenile onset tolerance, which may have origins that differ from rapid tolerance. The implications of this study go beyond tolerance in Drosophila melanogaster to benzyl alcohol and can shed light on human drug tolerance, withdrawal and addiction.Item Regulation of calcium stores in normal and diabetic endothelial cells(Texas Tech University, 2000-12) Sanka, Shankar ChittaranjanCytosolic Ca^^ ([Ca^^]*'^^) mediates many cellular ftinctions, e.g.. cell growth, motility, secretion, etc. In many cell types, ion transport processes appear to be dependent on metabolism of glucose for maximal activity. In certain cell types, a strict coupling between glycolysis and the acfivity of Endoplasmic Reticulum Ca^"-ATPases (SERCA). involved in regulating Ca^^ homeostasis, has been suggested. In diabetes, glucose homeostasis is altered. We hypothesize that Ca^^ homeostasis in microvascular endothelial cells from diabetic animals is altered due to a dysfunction of glycolysis coupling the activity of SERCA. We further hypothesize that endosomal/lysosomal (E/L) compartments exhibiting SERCA are involved in this dysfunction. Our data indicated that agonist stimulation (ATP, vasopressin, angiotensin-II)elicited [Ca^"]^^* increases (independent of extracellular Ca^^) that were larger in endothelial cells from diabetic than from normal animals. Simultaneous measurements of [Ca^^]'^^' and Ca^^ in E/L compartments ([Ca^^]^) using fluorescence spectroscopy, indicated that E/L compartments released Ca^^ following agonist-stimulation. The magnitude of the Ca'* release was significantly larger in microvascular endothelial cells from diabetic rats. SERCA inhibitors elicited Ca^^ releases from E/L compartments in both normal and diabetic models. The magnitude of the [Ca^^]^ release was however similar among normal and diabetic cells. Immunocytochemical experiments demonstrated that 60% of E/L compartments exhibited SERCA. These data indicate that (a) E/L compartments are important for Ca^^ homeostasis in microvascular endothelial cells from both normal and diabetic models; (b) Ca^^ regulation in E/L compartments is different in cells from a diabefic model, (c) the compartment involved in altered Ca'* homeostasis in diabetes is unknown.Item The Role of Foxo Transcription Factors in B Cell Development and Activation(2010-01-12T18:52:39Z) Hinman, Rochelle Marie; Satterhwait, Anne B.A functional immune system depends on a diverse, self tolerant B cell repertoire. Mature B cells distributed throughout secondary lymphoid organs respond to antigenic stimuli by dividing and differentiating into plasma cells and other effector cell types. Signaling from the B cell receptor (BCR) plays a critical role at several points during this developmental process. Cell survival, proliferation, differentiation, death, anergy, and receptor editing may occur in response to BCR stimulation. A variety of factors, including signal strength and duration, cytokine presence, and co-stimulation determine the ultimate B cell fate. In this thesis, the roles Foxo transcription factors play in maintaining B cell homeostasis will be explored. Foxo1, Foxo3, and Foxo4 have both anti-mitogenic and pro-apoptotic properties. The transcription factors are posttranslationally controlled via Akt. When a mature B lymphocyte is stimulated through the BCR, Akt-mediated phosphorylation of Foxos results in their exclusion from the nucleus. In the absence of Foxo nuclear activity, the B cell progresses into the cell cycle. We have discovered a second PI3K-dependent means of control for Foxos, at the level of mRNA expression. Downstream of the BCR, this means of control is unique and functionally relevant. Mature B cells proliferating in response to anti-IgM downregulate Foxo mRNA expression. This is via activation of the PI3K/Btk/BLNK/PLC-gamma2 pathway. Conversely, Foxo mRNA expression is upregulated in immature B cells, both when the tonic/basal signal through the BCR is disrupted and when the BCR is engaged with anti-IgM. Overexpression of Foxo3 mRNA in an immature B cell line promotes anti- IgM induced apoptosis. Primary immature B cells from Foxo3-/- mice have decreased apoptotic response to BCR crosslinking. Thus, at the immature stage of development our work has revealed a potential role for Foxo3 in promoting clonal deletion. Foxo3-/- mice also have reduced frequencies of pre-B and mature recirculating B cells in the blood and bone marrow. The mice demonstrate increased basal levels of IgG2a, IgG3, and IgA. Thus, Foxo3 deficiency affects numerous aspects of B cell development. [Keywords: B Lymphocyte; Foxo transcription factors; B cell receptor (BCR); PI3K; Foxo3; Btk; B cell development; B cell activation]Item Studies of Bile Acid-Like Signaling Pathways in Mammals and Nematodes(2010-01-12T18:59:50Z) Zhi, Xiaoyong; Mangelsdorf, David J.Bile acids are not only detergents for lipid solubilization and absorption, but also important signaling molecules. They regulate biological events in mammals by acting on nuclear receptors and membrane-bound receptors. Bile acid homeostasis is maintained in part through a FXR-SHP signaling circuit, in which SHP functions as a transcriptional corepressor. The mechanism whereby SHP represses was one focus of my thesis research. I used a number of biochemical strategies including tandem affinity purification to identify SHP interacting proteins. I also successfully solubilized SHP recombinant protein, which was used to generate crystals that diffracted to 3.2 Angstroms. Bile acid-like molecules function in nematodes to control a variety of life history traits such as dauer and infective L3 formation through the nuclear receptor DAF-12. Although DAF-12 homologues from different nematode species are functionally and structurally conserved, they show differential pharmacological responses to ligands. To that end, I solved the X-ray crystal structure of the hookworm Ancylostoma ceylanicum DAF-12 ligand binding domain and revealed the molecular basis underlying species specific-ligand binding for DAF-12. Furthermore, DAF-12 was shown to be structurally similar to the bile acid sensor FXR, suggesting bile acid-like signaling pathways have been conserved across evolution. In conclusion, my studies provide new insights into how bile acids are sensed and regulated in mammals and nematodes. [Keywords: bile acids; SHP; FXR; DAF-12; Dafachronic acids; hookworm; protein expression; crystallization]