Molecular and genetic analysis of synaptic signaling in Drosophila has yielded many insights into nervous system development, properties of synaptic transmission, and how long-lasting changes in neurons occur. Synaptic signaling components required for synaptic transmission and pathways leading to nervous system plasticity are typically conserved from insects to humans. The role of proteins and genes in synaptic function in flies can be analyzed from the level of a single synapse to complex behaviors in the whole organism. Because of a fully sequenced genome and the ease of mutagenesis in flies, genetic screens have been useful in identifying novel regulators of synaptic transmission and long-term memory.In flies, conditional mutations affecting synaptic transmission at nerve terminals often lead to temperature sensitive paralysis. In a screen for mutations that interact with Drosophila shibirets mutants, the stoned gene was identified as a regulator of synaptic vesicle cycling. Stoned encodes two neuronally expressed proteins, stonedA and B, which are required for synaptic vesicle recycling and normal synaptic transmission. However, the exact functions of the two stoned proteins are not fully understood. We investigate distinct roles of the stoned proteins here and show that stoned has a novel role in synaptic growth.Memory in flies can be divided into genetically distinct phases based on the requirement for protein synthesis and activation of the transcription factor CREB. Novel regulators of long-term olfactory avoidance memory were isolated in a mutant screen in flies. Mutants in the Drosophila gene lk6, homologous to the translational regulator MNK, have defects in long-term olfactory avoidance memory. We find that lk6 is highly expressed in the fly nervous system, and is activated by and functions downstream of Ras/ERK signaling in fly neurons. Insights provided here from Drosophila add to the evidence that MNK may be the link between ERK signaling and the regulation of translation in long-term plasticity.Ultimately, understanding synaptic function has therapeutic potential to aid in alleviation of nervous system dysfunction. Insight into the molecular pathways underlying plasticity and long-term memory gained from studies in flies, mollusks, and rodents has been pivotal in the development of potential drugs to aid in memory deficits in humans.

Using an operant-olfactometer, the long term learning and memory, executive function, olfactory sensitivity, and working memory of the 5XFAD mouse model of Alzheimer’s disease was assessed. Six month old male and female 5XFAD and wildtype mice were tested. No deficits were found on an olfactory discrimination task or a reversal learning task. Female and transgenic mice performed better than male and wildtype mice on the higher odour concentrations, but not the lower concentrations, of the sensitivity task, suggesting differences in learning rate or maximum performance on the task, but not olfactory detection. This study demonstrated for the first time that mice are able to learn an olfactory delayed matching to sample task with delays up to 30 seconds long. Female mice showed higher levels of performance on the matching to sample task than male mice, indicative of better working memory.

The LIM-Kinase family of proteins (LIMK) plays an important role in actin dynamics through its regulation of ADF/cofilin. A subtype of LIMK, LIMK1, is mostly expressed in neuronal tissues with high levels in the mature synapse. Previous studies from the Zhen Ping Jia laboratory have shown that LIMK1-/- mice exhibit abnormal spine morphology as well as altered hippocampal synaptic plasticity. LIMK1 has been shown to interact with CREB during neuronal development (Yang et al., 2004). We propose that LIMK1 is able to phosphorylate CREB in response to a synaptic activity. We hypothesize that if LIMK1 activates CREB in mature neurons, then LIMK1 knockout mice will have decreased L-LTP and deficits in long-term memory. My results show that LIMK1 and CREB exist in a complex and are bound to each other in mature neurons. LIMK1-/- mice exhibit deficits in the late phase of long-term potentiation and specific deficits in long-term memory while short-term memory remains unaltered. Pharmacological activation of CREB attenuates the observed deficits in synaptic plasticity and long-term memory. These results show a potentially novel mechanism of CREB activation in response to synaptic activity. Moreover, using peptides to manipulate actin dynamics in LIMK1 lacking animals only has effects on early LTP and is not able to rescue the late phase LTP deficits found in LIMK1 -/- mice. These results indicate a specific role of LIMK1 long-term memory and synaptic plasticity through regulation of CREB and not through regulation of the actin cytoskeleton.

The LIM-Kinase family of proteins (LIMK) plays an important role in actin dynamics through its regulation of ADF/cofilin. A subtype of LIMK, LIMK1, is mostly expressed in neuronal tissues with high levels in the mature synapse. Previous studies from the Zhen Ping Jia laboratory have shown that LIMK1-/- mice exhibit abnormal spine morphology as well as altered hippocampal synaptic plasticity. LIMK1 has been shown to interact with CREB during neuronal development (Yang et al., 2004). We propose that LIMK1 is able to phosphorylate CREB in response to a synaptic activity. We hypothesize that if LIMK1 activates CREB in mature neurons, then LIMK1 knockout mice will have decreased L-LTP and deficits in long-term memory. My results show that LIMK1 and CREB exist in a complex and are bound to each other in mature neurons. LIMK1-/- mice exhibit deficits in the late phase of long-term potentiation and specific deficits in long-term memory while short-term memory remains unaltered. Pharmacological activation of CREB attenuates the observed deficits in synaptic plasticity and long-term memory. These results show a potentially novel mechanism of CREB activation in response to synaptic activity. Moreover, using peptides to manipulate actin dynamics in LIMK1 lacking animals only has effects on early LTP and is not able to rescue the late phase LTP deficits found in LIMK1 -/- mice. These results indicate a specific role of LIMK1 long-term memory and synaptic plasticity through regulation of CREB and not through regulation of the actin cytoskeleton.

The behavioral effects of drugs that act on the brain’s dopamine (DA) system change with repeated exposure to the drug. Antipsychotic drugs, that block DA receptors, produce progressively greater effects on behavior with repeated testing. For example, rats repeatedly treated with a low dose of the D2 receptor-preferring antagonist haloperidol do not initially exhibit catalepsy, a response quantified by time spent on a horizontal bar without active movement. However, with repeated drug-environment pairings animals show a reduction in exploration and increases in catalepsy. The current thesis examined the drug-environment relationship in catalepsy sensitization, and how different DA receptor subtypes control this phenomenon. Treatment with a D2, but not D3 or D1-like receptor-preferring antagonist produced catalepsy sensitization. Catalepsy sensitization developed in one test environment did not transfer to another environment. Similarly, rats with a history of haloperidol treatments outside of the test environment (unpaired group) did not exhibit significant catalepsy when given haloperidol for the first time prior to catalepsy testing. Previous exposure to the catalepsy test environment led to a more rapid development of catalepsy sensitization. Thus, drug-environment interaction is critical for the development and expression of catalepsy sensitization. Rats previously given haloperidol and tested with saline in the drug paired environment exhibited conditioned catalepsy. The acquisition of conditioned catalepsy is dependent on D1-like receptors, while its expression is dependent on D3 receptors. Conditioned catalepsy showed gradual day-to-day extinction with repeated saline treatment in the previously haloperidol-paired environment. Following extinction, the response to haloperidol in previously sensitized rats shifted from environment-specific to environment-independent suggesting that a putative haloperidol drug cue alone can elicit conditioned catalepsy. In summary, treatment with a D2, but not D1-like or D3 receptor-preferring antagonist in a particular test environment produces catalepsy sensitization, while acquisition of conditioned catalepsy is dependent on D1-like receptors, and its expression is dependent on D3 receptors. Importantly, the acquisition and expression of sensitization to haloperidol is conditional on the presence of drug-associated environmental stimuli. Our findings provide further insight into the current understanding of learning processes involved in the action of antipsychotic drugs and the dissociable effects of D1-like, D2 and D3 receptors controlling this phenomenon. / Thesis (Ph.D, Neuroscience Studies) -- Queen's University, 2012-08-12 15:51:00.467

Dynorphins, endogenous peptide neurotransmitters expressed in the central nervous system, have been implicated in diverse pathophysiological processes, including excitotoxicity, chronic inflammation, traumatic injury, cognitive impairment, and motor dysfunction, with significant changes with aging or age-related disease processes. This has led to the hypothesis that the suppression of dynorphin expression would produce beneficial effects on learning and memory and motor function. To assess the phenotypic manifestations of chronic suppression of endogenous dynorphin, knockout (KO) mice lacking the coding exons of the gene encoding the prodynorphin (Pdyn) precursor protein, were tested in a series of behavioral, biochemical, and molecular biological studies. Moderately aged Pdyn KO perform comparatively better than similarly aged wild-type (WT) mice in the water maze task, although no Pdyn effect was seen among young adult mice. In addition, young adult Pdyn KO mice show mildly improved performance on a passive avoidance task. Minimal baseline differences were noted in spontaneous locomotor activity in an open-field assay, but Pdyn deletion produced a relative sparing of motor dysfunction induced by the neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). To investigate the relationship between aging and brain dynorphin expression in mice, we examined dynorphin peptide levels at varying ages in hippocampus, striatum, and frontal cortex of WT mice by quantitative radioimmunoassay. While aging produces progressive decline in Dyn B in striatum and frontal cortex, Dyn A shows an upward trend in frontal cortex without significant change in striatum. Systemic MPTP produces significant short-term elevations in dynorphin peptides that regress to below baseline by 7 days. HPLC analysis of striatal dopamine shows an age-dependent increase in basal dopamine levels in Pdyn KO mice, an effect that is abolished after MPTP. Western blotting experiments demonstrate that Pdyn deletion is associated with greater phosphorylation at the serine-40 site of tyrosine hydroxylase (TH) despite relatively less total TH immunoreactivity, suggesting a suppressive effect of dynorphins on dopamine synthesis. Microarray analysis of hippocampal tissue from young and aged WT and Pdyn KO mice reveals a number of functional groups of genes demonstrating altered expression. The results of this dissertation support a role of endogenous dynorphins in age-associated cognitive and motor dysfunction.

Research on the neural and cognitive basis of spatial navigation over the last 30 years has been largely guided by cognitive map theory and many of the studies have used a standardized procedure in a single task, the Morris Water Maze (MWM). Although this theory proposes that acquisition of place knowledge should be very rapid, little evidence has been provided to support this point. The present study investigates the possibility that a new procedure for measuring place knowledge in the MWM will show that place learning is faster than previously shown. In a virtual MWM with a fixed goal location, participants were given pairs of standard learning trials plus new explicit probe trials in which they were directed to go to where they found the goal on the immediately preceding trial. The distance between their estimate and the actual location was measured as “Place Error”. Results indicated that Place Errors were surprisingly small after just one learning trial and were equivalent for females and males. These findings provide new evidence for the fast learning proposed by cognitive map theory and demonstrate the value of this new method for measuring place learning. / Graduate

Thesis (Ph. D.)--University of Chicago, Dept. of Psychology and Dept. of Linguistics, June 1999. / Includes bibliographical references. Also available on the Internet.

Thesis (Ph. D.)--University of Wisconsin--Madison, 1969. / Typescript. Vita. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references.

The epigenetics of the aging brain is a growing field of study that holds great promise for the discovery of mechanisms and potential treatments for neurodegenerative diseases. In this current study, a novel, accelerated aging murine model, the I-PpoI/Cre, or ICE (Inducible Changes in the Epigenome) mouse, is studied to test its potential for demonstrating the theory of the rearrangement of chromatin (RCM) as the main cause of aging, and in particular, the mechanism through which the brain ages. Immunohistochemistry and behavioral assays are utilized to determine whether there are morphological changes, inflammatory response, and changes in learning and memory. Results showed a significant increase in microglia and astrocytes, markers of inflammation, in I-PpoI/Cre mice compared to their Cre controls. Long term memory performance was also significantly decreased in the I-PpoI/Cre mice, demonstrated through contextual fear conditioning (CFC) testing, and Morris Water Maze (MWM) testing. Results from this study are in support of the I-PpoI/Cre mouse as a model of accelerated aging of the brain, with deficits in learning and memory. Further studies are needed to further characterize this murine model of accelerated aging.