Browsing by Author "Robert Grossfeld, Committee Member"

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Analysis of Xenopus laevis claudin (Xcla) Tight Junction Genes in Development
(2005-07-27) Xie, Jianzhen; Robert Grossfeld, Committee Member; Brenda Brizuela, Committee Chair; Pat Estes, Committee Member
Eight Xenopus laevis claudin genes, Xcla1, Xcla4B, Xcla5, Xcla6, Xcla12, Xcla16, Xcla18 and Xcla19, were cloned and sequenced. Their normal mRNA expression was determined from cleavage stage to tadpole stage by whole mount in situ hybridization. The protein expression of Xcla5 was detected at the neural stage by whole mount immunostaining. Overexpression of Xcla5 by injection of synthetic mRNA into embryos caused morphological defects similar to those in embryos exposed to Bisphenol A (BPA). Altered patterns of claudin gene expression in the presence of BPA can be correlated with these developmental defects. The results suggest that claudins may play an important role in neural crest cell migration, epithelial-mesenchymal transition and ultimately organogenesis during embryonic development.
Developmental Exposure to Environmental Estrogens Alters Adult Behavior in Female Rodents
(2005-08-16) Ryan, Bryce Clair; John G. Vandenbergh, Committee Chair; L. Earl Gray, Committee Member; Robert Grossfeld, Committee Member; Robert MacPhail, Committee Member; Gerald LeBlanc, Committee Member
Humans and wildlife are exposed to numerous anthropogenic drugs and pollutants. Many of these compounds are hormonally active and recent evidence strongly suggests that the presence of these endocrine disruptors can permanently alter normal development and physiology in a variety of vertebrate species. The experiments in this project investigated the effects of two common estrogenic pollutants. Bisphenol a is a monomer of polycarbonate plastic used to make resins for the food and dental industries. Ethinyl estradiol is used pharmaceutically as the active estrogen in the oral contraceptive pill. The majority of past research on these chemicals has focused on reproductive physiology. The focus of my research in on the behavioral consequences of developmental exposure to these compounds. Estrogens will feminize the reproductive system but will masculinize the rodent nervous system, so I focused on identifying whether females would show masculinization of sexually dimorphic traits. The effects of these compounds were studied on two commonly used laboratory species, the mouse (Mus musculus domesticus) and the rat (Rattus norvegicus). The test animals were exposed to environmentally relevant levels of these compounds (ranging from 2 — 200 μg/kg/day for bisphenol A and 0.05 to 50 μg/kg/day for ethinyl estradiol) throughout prenatal and early postnatal development. After this exposure, the animals were allowed to reach adulthood and then observed in a variety tests measuring sexually dimorphic behaviors. These include short-term spatial memory, anxiety, saccharin preference, motor activity and lordosis. Developmental exposure to ethinyl estradiol was found to masculinize every behavior measured in both species in a dose-dependent fashion. Bisphenol A disrupted selected behaviors, namely anxiety and motor activity, and was active in both rodent species, but did not always follow a clear dose response. These results indicate that sexually dimorphic behavior can be exquisitely sensitive to endocrine disruption. In addition, these experiments suggest that both humans and wildlife are presently being exposed to levels of these endocrine disrupting compounds that are sufficient to disrupt the development of the nervous system and that may have permanent consequences on sexually dimorphic behaviors.
The Genetic Architecture of Locomotor Behavior in Drosophila melanogaster
(2006-12-20) Jordan, Katherine Wells; Trudy Mackay, Committee Chair; Bruce Weir, Committee Member; Robert Grossfeld, Committee Member; Michael Puruganan, Committee Member
Locomotion is an integral component of most animal behaviors: movement is required for localization of food and mates, escape from predators, defense of territory, and response to stress. Many human neurological diseases (e.g., Parkinson's Disease and Huntington's Disease) are associated with locomotor deficits. Locomotion is a complex behavior, with variation in nature attributable to the joint segregation of multiple interacting quantitative trait loci (QTLs), with effects that are sensitive to the environment. Thus, understanding the genetic architecture of locomotor behavior is important from the dual perspectives of evolutionary biology and human health. However, our current knowledge falls short of the level of detail with which we ultimately seek to describe variation in locomotor behavior. We used complementary approaches in the model system Drosophila melanogaster to identify genes affecting locomotion: QTL mapping, followed by linkage disequilbrium mapping and association testing; artificial selection to derive lines for transcriptome analysis using microarrays; and P-element insertional mutagenesis to confirm the microarray results. QTL mapping uncovered four regions that contribute to variation in locomotor reactivity (a component of locomotor behavior) between two lab stocks. Deficiency complementation mapping refined our large QTL into 12 smaller QTL, then complementation tests to mutations identified 13 positional candidate genes affecting locomotor reactivity, including Dopa decarboxylase (Ddc) and Catecholamines Up (Catsup). Linkage disequilibrium mapping in a natural population of 164 second chromosome substitution lines suggested polymorphisms at Ddc and Catsup were associated with naturally occurring genetic variation in locomotion. Another strategy to discover genes affecting complex behaviors is to combine artificial selection for divergent phenotypes with whole genome expression profiling. Artificial selection lines created from a genetically heterogeneous background were selected for 25 generations to derive replicate lines with divergent levels of locomotor reactivity. Transcription profiling identified nearly 1,800 probe sets that were differentially expressed between the selection lines. Functional tests of P-element mutations in ten differentially expressed genes confirmed seven novel candidate genes affecting locomotion. Many of the genes identified in this study have other functions in metabolism, nervous system development, and response to different stimuli, suggesting extensive pleiotropy among the genes affecting locomotor behavior.
The Genomic Architecture of Aggressive Behavior in Drosophila melanogaster
(2008-09-25) Edwards, Alexis C; Robert Grossfeld, Committee Member; Jane Lubischer, Committee Member; Patricia Estes, Committee Member; Robert Anholt, Committee Member; Trudy F.C. Mackay, Committee Chair
Pharmacogenetic Analysis of Nicotine and Caffeine Resistance in Drosophila Melanogaster.
(2004-12-01) Carrillo, Roland Javier; Robert Grossfeld, Committee Member; Michael Purugganan, Committee Member; James Mahaffey, Committee Member; Greg Gibson, Committee Chair
Drug response is a polygenic trait that varies as a result of many factors, including the rate of drug absorption, metabolism and secretion. It is an important trait that can result in physiological and behavioral changes and can affect both health and survival. Associations between drug response and genes have been suggested but no clear picture of the relationship between genetic and pharmacological variation has yet emerged. Dissection of the genetic architecture of drug resistance is further complicated in that it involves the activity of multiple genes, which can interact with each other and the environment. My research uses Drosophila melanogaster to study resistance to the behaviorally active substances nicotine and caffeine. Both of these substances can exhibit adverse health effects at high doses or after chronic use by humans and are lethal when added to the diet of Drosophila. For this study, several approaches were used to study drug response, including an analysis of quantitative genetic variation for drug resistance in natural populations, a P-element mutagenesis screen and association tests with candidate genes. These were used to assess drug resistance by measuring survival time on diets containing either nicotine or caffeine, and revealed that abundant genetic variation exists for drug resistance in Drosophila. This variation involves a complex genetic architecture and the interaction of many genes. Nevertheless, a classical forward genetic mutagenesis screen identified individual genes involved in drug resistance. These genes were not those typically studied for drug resistance, such as those in neurotransmitter systems and drug metabolism, but were involved in the development of the CNS and neuronal differentiation. Furthermore, an association study between nicotine and caffeine resistance and single nucleotide polymorphisms in three serotonin receptor genes, 5-HT1A, 5-HT1B and 5-HT2 detected significant associations between a SNP and nicotine resistance in the 5-HT1A gene. This suggests that although drug resistance is a complicated trait involving the interaction of many genes and environmental effects, mutations in individual genes and naturally occurring polymorphisms affecting survival time upon chronic exposure to nicotine and caffeine can be detected in Drosophila.