1993 — 1997 |
Williams, Diana L |
R29Activity Code Description: Undocumented code - click on the grant title for more information. |
Dna Diagnostic Assay For Rifampin-Resistant Tuberculosis @ National Hansen's Disease Program
The development of a DNA diagnostic assay for the rapid detection of rifampin-resistant (Rif) Mycobacterium tuberculosis strains directly from patient specimens could result in a decrease in the amount of time from sample acquisition to diagnosis of resistance in tuberculosis patients. This could lead to implementation of appropriate drug therapy much earlier in the course of disease, and, thereby, increase patient's chances of survival, potentially decrease the spread of drug-resistant tuberculosis and decrease the cost of treatment of affected individuals. The identification of the molecular mechanism of rifampin resistance in M. tuberculosis is an important step in the development of a DNA diagnostic assay for the rapid detection of these organisms directly from patient samples and in choosing rational approaches for alternative drug design. The work outlined in this proposed research is designed to determine the molecular basis of rifampin resistance in M. tuberculosis and to use this information to design a DNA diagnostic assay for the rapid and specific detection of Rif strains of M. tuberculosis directly from clinical specimens. It is anticipated that use of this assay will make the identification of these drug-resistant organisms possible much earlier in the course of disease. To reach the goals of this research the following objectives will be accomplished: The mutation(s) which are associated with the development of rifampin resistance in M. tuberculosis will be determined using PCR amplification and a direct DNA sequencing methodology. The stability of these mutations will be examined using characterized clinical isolates of Rib M. tuberculosis. These isolates have been characterized using drug- susceptibility testing and restriction fragment length polymorphism analysis to differentiate strains. A DNA diagnostic assay for the detection of Rif M. tuberculosis strains using characterized clinical isolates will be developed. This assay is based on PCR amplification of a target sequence which contains all known Rif alleles and subsequent hybridization with allele-specific probes and nonisotopic detection. The feasibility of detecting Rif M. tuberculosis strains directly from clinical specimens using the DNA diagnostic assay will be determined initially using sediments from highly-characterized sputum samples. Finally, the molecular basis of rifampin resistance in M. tuberculosis as a function of an altered beta-subunit of the DNA-dependent RNA polymerase will be characterized by purifying RNA polymerase holoenzymes from Rif and Rif strains of M. tuberculosis, purifying the subunits of these enzymes and reconstituting Rif beta-subunit into Rif holoenzyme. The hybrid holoenzyme will be analyzed for the production of mRNA in the presence and absence of rifampin.
|
0.903 |
2005 — 2007 |
Williams, Diana L |
F32Activity Code Description: To provide postdoctoral research training to individuals to broaden their scientific background and extend their potential for research in specified health-related areas. |
Hindbrain Integration of Adiposity and Satiety Signals @ University of Washington
[unreadable] DESCRIPTION (provided by applicant): This proposal investigates the neural circuitry that integrates input from the adiposity hormone, leptin, and the gut-derived satiety signals, cholecystokinin (CCK), in the control of food intake. Available data provides compelling support for the hypothesis that leptin reduces food intake in part by enhancing satiety and hindbrain responses to gastrointestinal signals, including CCK. Here, we propose to identify the specific subset of hindbrain neurons whose responses to CCK are stimulated by leptin. Second, we will determine whether the interaction between leptin and CCK, clearly demonstrated with pharmacological doses of leptin, occurs with physiological changes in leptin levels. Finally, we will determine whether leptin regulates hindbrain cell responsiveness to CCK through a direct action within the caudal brainstem or through an indirect route, perhaps involving a descending projection from the forebrain. Together, these studies will help to clarify how changes in body fat stores lead to compensatory adjustments of meal size and thus have the potential to significantly advance our understanding of the neural circuitry that regulates energy balance. [unreadable] [unreadable]
|
0.943 |
2007 — 2011 |
Williams, Diana L |
K99Activity Code Description: To support the initial phase of a Career/Research Transition award program that provides 1-2 years of mentored support for highly motivated, advanced postdoctoral research scientists. R00Activity Code Description: To support the second phase of a Career/Research Transition award program that provides 1 -3 years of independent research support (R00) contingent on securing an independent research position. Award recipients will be expected to compete successfully for independent R01 support from the NIH during the R00 research transition award period. |
Brain Integration of Adiposity and Satiety Signals in the Control of Food Intake @ Florida State University
Obesity and related metabolic disorders have become a tremendous public health problem. This proposal Investigates the neural circuitry that Integrates input from the adiposity hormone leptin and the gut-derived satiety signals cholecystokinin (CCK) and glucagon-like peptlde-1 (GLP-1), and the physiological relevance ofthe interactions between these signals for the control of food Intake and body weight. Available data support the hypothesis that the adiposity hormone leptin reduces food Intake in part by enhancing satiety responses to gastrointestinal signals, Includingj CCK. We have recently determined that leptin Interacts In a sirnilar rrianrier with GLP-1, but our data suggest that the leptin-CCK and leptin-GLP-1 interactions are mediated through different neuronal pathways, Here, we propose to Identify and compare brain areas In which leptin detection Is sufficient to enhance the responses to GLP-1 and CCK, using site-speclfic microinjections and gene therapy In normal rats and rats that lack leptin receptors due to genetic mutation. Next, we will Investigate whether leptin reduces food Intake and enhances the satiety response to gastrointestinal nutrients in part by modulating rats'sensitivity to endogenously released GLP-1 or CCK. Finally, we will determine whether altered sensitivity to GLP-i or CCK plays a role in the pathogenesis of diet-Induced obesity when rats are maintained on a high-fat diet, which produces leptin resistance, together, these studies will help to clarify how changes In body fat stores lead to compensatory adjustments in meal size, as well as how defects In this process may lead to obesity and metabolic disorders, and thus have the potential to significantly advance our understanding ofthe neural control of feeding and body weight.
|
1 |
2013 — 2021 |
Williams, Diana L |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Neuroendocrine Integration of Satiety and Food Reward @ Florida State University
Overconsumption of highly palatable calorically dense food is a major contributor to obesity and related metabolic disorders. This proposal investigates one of the neuropeptide systems thought to underlie neural integration of the rewarding value of food with input from gut-derived satiety signals, Glucagon-like peptide 1 (GLP-1). Preproglucagon (PPG, the precursor to GLP-1) neurons project to many brain areas where activation of GLP-1 receptors (GLP-1R) promotes satiety and reduces motivation for food. Most research has focused on one or another individual PPG neuron projection and GLP-1R population at a time, and although this has informed us about brain GLP-1 action, this approach does not provide broad insight into the functional organization of the central GLP-1 network. Here we will take advantage of transgenic mouse models to investigate GLP-1R neuron projections that mediate behavioral effects. We hypothesize that: 1) activation of some, and inhibition of other GLP-1R neuron projections reduce feeding; 2) that GLP-1R neurons in different brain nuclei receive synaptic input from unique brain regions; and 3) that GLP-1R neurons communicate with one another across brain regions. Based on our data implicating GLP-1R neurons of the Lateral Septum (LS) and Bed Nucleus of the Stria Terminalis (BNST) in feeding control, we focus on two exemplar cell populations: the LS GLP-1R neuron projection to Lateral Hypothalamus (LH); and the BNST GLP-1R neuron projection to the LH. Aim 1 focuses on the GLP-1R LS-to-LH pathway, which we hypothesize promotes satiety and suppresses food reward when activated. Aim 2 examines the GLP-1R BNST-to-LH projection, which we hypothesize works in the opposite direction, such that inhibition of these neurons promotes satiety and suppresses food reward. Experiments will test these hypotheses using a combination of cell type-specific chemogenetic and pharmacologic approaches to manipulate the activity of each of these GLP-1R neuron projections to LH. We will conduct detailed behavioral analyses to distinguish effects on satiation, satiety, motivation, and stress or malaise that can alter feeding, and we will use slice electrophysiology to characterize the underlying neuronal signaling pathways. Aim 3 will determine sources of synaptic input to GLP-1R neurons in each location, testing the hypothesis that they receive distinct sources of input from PPG and other neurons, including GLP-1R+ neurons in other nuclei. Studies in this aim will apply a combination of traditional retrograde tracing and cutting edge cell type- and anatomic pathway-specific mono- and transsynaptic viral tracing methods. Together, our results will elucidate new mechanisms for GLP-1's hypophagic effects, identify new cell type-specific neuronal pathways that play a role in brain control of feeding, and provide a more complete picture of how PPG neurons and GLP-1-receptive cells throughout the brain coordinate to influence behavior. We propose that central GLP-1 signaling pathways are not unique in their integrated network organization, and expect that our findings will serve as a template for assessing these same questions for other circuits.
|
1 |