Chronic social-stress is known to lead to a number of aberrant behaviors, including anxiety and depression. Using a mouse model of social-defeat stress, this project aims to better understand the relationship between the kappa-opioid receptor (KOR), a G-protein coupled receptor (GPCR) previously shown to be activated by multiple forms of stress, and its role in generating anxiety-like and aversion behaviors in mice. Different forms of stress lead to the release of the endogenous KOR agonist, dynorphin, which activates an intracellular signal transduction cascade resulting in anxiety-like and aversion behaviors. Prior studies have demonstrated that the long-acting KOR antagonist, norbinaltorphimine (norBNI), is able to reduce social-defeat induced social avoidance behaviors when administered prior to social defeat stress exposure. Although these studies have implicated the KOR system in social-defeat induced behaviors, pretreatment of pharmacotherapies prior to a stress event is limited in therapeutic benefit.  In this research, the ability of norBNI to reverse repeated social-defeat induced changes in social behaviors was tested by administering norBNI following exposure to the social-defeat stress. The results have so far shown that post treatment with norBNI is able to effectively counter the altered social interaction behaviors caused by a repeated social-defeat stress; moreover, the reversal of the stress-induced social interaction deficits also correlates with a reduction in anxiety-like behaviors. These data suggest that social interaction deficits are due to increased anxiety in previously benign social settings, and norBNI has the therapeutic potential to reverse these social-stress induced behavior alterations.

The effects of amphetamine on learning and memory are important areas of research due to the widespread use of amphetamine among people, ranging from prescription to recreational. It has been shown in animals that repeated treatment of amphetamine will result in sensitization to the drug. Sensitization is a process through which treated subjects develop an elevated response to a drug. Long term potentiation (LTP) is a physical change strengthening the connection between synapses that is thought to underlie learning and memory. Enhancement of LTP in the striatum has been shown to occur as a result of amphetamine use. The striatum is a brain region important in mediating a large number of behaviors including motor control and learning. For this reason it is possible that sensitization to amphetamine may affect learning in mice. To investigate this possibility, wild-type mice sensitized to amphetamine were tested in behavioral paradigms designed to measure learning ability. It was found that they do not show enhanced performance in motor learning, appetitive learning, or spatial learning, but they may show an improvement over controls when they are required to alter their strategy to escape from a water-maze. In order to explore the neurological pathway through which sensitization to amphetamine may affect learning, we also tested knockout mice that lack N-methyl-D–aspartate-type glutamate receptors (NMDARs) in medium spiny neurons, the primary neuron type in the striatum. In many brain regions, NMDARs are important for various forms of learning and memory, and it has been shown that these knockout mice display impaired motor and appetitive learning and yet are still able to sensitize to amphetamine. Using the same learning paradigms as with our wild-type mice, we found that knockout mice did not show any enhancement in motor or appetitive learning as a result of amphetamine sensitization.

Cameras have become a fundamental part of our daily lives because of the powerful ability of images to communicate, preserve, and educate. Computational photography is an emerging field of study that considers sensory information from a scene to be intermediate data, and uses computation to form the final photo. The goal is to improve images by combining computing power with digital imaging to create new images that no digital camera on the market today could capture. Techniques that represent computational photography include high-dynamic-range (HDR) imaging, panoramic stitching, and light-field imaging. Last year, Andrew Adams, a leading Stanford researcher behind computational photography, released a fully programmable digital camera system to aid other researchers in computational photography. Named FCam, it is an open-source C++ application programmable interface (API) for easy and precise control of digital cameras. My research will focus on using the FCam API to develop faster, smarter algorithms to capture images for accessibility purposes. Computational photography can be directly applied to several accessibility research projects. The project I am currently working on improves the visibility of street signs so that another person or a computer can recognize the text. By quickly capturing multiple images with different levels of luminosity, an optical character recognition engine can provide better textual results to the user. This research demonstrates the importance of using computational photography to make information more accessible to people with disabilities.

Mitochondria are organelles vital to all cells in the body, generating energy and regulating metabolism, calcium signaling and apoptosis. These diverse functions are made possible partly through constant remodeling of mitochondria via fission and fusion. Neurons are especially vulnerable to mitochondrial defects, and this is true for mutations arising in mitochondrial fusion/fission proteins. Mitofusins (Mfn) are key proteins for mitochondrial fusion. Presently, we do not have clinically useful drugs to modulate fission and fusion in order to prevent neuronal damage due to disease and injury. In order to characterize how mitochondrial phenotype affects the metabolic rate in mammalian cells and disease models, we genetically engineered a chemically inducible system that allowed us to manipulate the mitochondrial fusion machineries. Specifically, I helped produce a ligand-induced heterodimerization system to initiate assembly and activation of fusion protein complexes by a specific chemical cue or ligand. Rapamycin, an antibiotic, complexes with immunophilin FKBP-12 (FKB) and binds to the FKB-rapamycin-binding domain (FRB) of the serine/threonine protein kinase mTORC1. I assisted in genetically engineering FKB and FRB onto the functional ends of the Mfn proteins. These Mfn proteins were expressed in mutant cells lacking all Mfn proteins. Since they lack fusion proteins their mitochondria are highly fragmented and we hypothesized that they could be rescued by inducing fusion with this inducible system. When we express the modified Mfn proteins containing FKB and FRB in cells, they will only dimerize and express fusion activity in the presence of rapamycin, which acts as a chemical ligand uniting the two modified forms of Mfn. We used a Seahorse analyzer to measure the oxygen consumption rate of FRB/FKB Mfn cells exposed to rapamycin for different periods of time. This system will help to identify new drugs that regulate mitochondrial dynamics in disease and injury.

Wind is highly variable and unpredictable, unlike conventional generation resources such as coal. The U.S. grid system is ill-equipped to accommodate resources that are stochastic in nature. Policies exist at the federal, state, and local levels that attempt to subsidize the production cost of wind generation, but how will these incentives impact system operations? We explore how the power generation mixture will change due to policies that incentivize wind. Our approach involves an analysis of a case study of three different incentive schemes: a base case (un-incentivized), the federal production tax credit (PTC) for wind, and a feed-in tariff (FIT) system modeled on the FIT proposed in California. Computer simulations will be developed to qualitatively model how these policies impact system operations using the economic dispatch algorithm, which is used in practice to deploy generation resources to serve the load and meet all power system constraints at the lowest cost. Comparing these cases will show which of these incentives have an impact on the total energy cost over a given time frame, as well as give an idea about the resulting generation profile.

Pedaliaceae (Sesame Family) are a small family of plants, about 14 genera with approximately 70 species that occur mainly in arid regions and deserts of the Old World, with their center of distribution in Southern Africa. Contained in Pedaliaceae are two widely known species: Sesamum indicum, which is cultivated and provides sesame seed and Harpagophytum procumbens, Devils Claw, an endangered species which is harvested wild and used for medicinal purposes. Evolutionary relationships (phylogeny) within the family are currently not very well understood. Previous classification systems have included the Martyniaceae, a small New World family, which occurs in similar environments and exhibits similar fruit morphology, with the Pedaliaceae but this inclusion of has since been questioned. In order to infer a phylogeny for the Pedaliaceae I am using the chloroplast loci ndhF and tnrL-tnrF which generally provide good resolution to determine the relationships between families and major lineages within a family of plants. Preliminary analyses based on these two loci indicate that the genus Sesamum is not monophyletic. To achieve resolution of the relationships of the species within Sesamum, I am sequencing two commonly used nuclear loci: ETS and ITS, the external and internal transcribed spacers of the nuclear ribosomal repeat, and one of the many loci in the pentatricopeptide repeat (PPR) which have only recently been developed for phylogenetic study in the Olmstead Lab. The goal of this project is to infer a phylogenetic tree of the Pedaliaceae on the species level, resolve their relationship to Martyniaceae and the genus Trapella, as well as the placement of these groups within the larger tree of the Lamiales.

Interspecific hybridization is common in flowering plants.  Hybrids tend to exhibit characteristics intermediate between both parents, and to grow more vigorously.  A novel specimen of the genus Heuchera (alumroot) found in a garden setting in Seattle is thought to be a hybrid between the commonly cultivated Heuchera sanguinea and the PNW native H. cylindrica.  In nature, H. cylindrica and H. sanguinea have non-overlapping geographic distributions.  Heuchera cylindrica is native to the western United States and Canada, whereas H. sanguinea is native to New Mexico and Arizona and it is not known if interbreeding between the two species is possible.  The unknown specimen in question shows intermediate characteristics of both species, which include flower color, inflorescence architecture and more vigorous growth.  To determine the relationship of the putative hybrid specimen with the two suspected parents, fixed differences in the Internal Transcribed Spacer (ITS) regions of nuclear ribosomal DNA of H. cylindrica and H. sanguinea were compared .  ITS regions were amplified using PCR and were then sequenced.  The results were edited, assembled and aligned to look for differences in the parents' ITS regions.  The differences in the parents' ITS regions are present in combination in the unknown specimen's DNA and shows that it is the hybrid of H. cylindrica and H. sanguinea.  A further test, using the maternally inherited chloroplast genome (cpDNA), was conducted in a similar fashion, with the expectation that the putative hybrid should contain the cpDNA of the species that is the maternal parent.

Ubiquitin is a protein modifier that is essential for many eukaryotic cellular processes. Attachment of ubiquitin to substrate proteins (ubiquitination) occurs in a three-step cascade that involves a ubiquitin activase, ubiquitin conjugase, and ubiquitin ligase; it is the ligase that targets substrate proteins. One challenge for the ubiquitin field is identification of substrate cohorts for ubiquitin ligases. Although ubiquitination only occurs naturally in eukaryotic organisms, we have previously reconstituted the ubiqutination cascade in E. coli. Isolation of the ubiquitin pathway within a prokaryotic cell, such as E. coli, allows for the investigation and modification of individual components of the pathway and the measurement of immediate effects of these changes upon the overall process of ubiquitination. This project, which consists of the construction of a split protein assay, aims to address the abovementioned challenge by building on our previous work reconstituting ubiquitination in E. coli. The assay uses two fragments of the protein dihydrofolate reductase (DHFR) to detect the ubiquitination of proteins in E. coli, outside of the natural eukaryotic environment of ubiquitin. All of the essential components of the ubiquitination pathway, including individual fragments of DHFR fused to ubiquitin and a substrate protein of interest, are transformed into the bacterial cell. The assay tests for the covalent attachment of ubiquitin to the substrate protein through growth on selective medium, which is possible only through the combination of the separated fragments of DHFR. This reporter system allows for experimentation regarding the interaction and selectivity of the ubiquitin ligase with the substrate protein of interest. Thus, a functional reporter system will allow for the streamlined analysis and determination of the substrate cohort for a given ubiquitin ligase. Changes to the components of the system may include using different ubiquitin ligases and substrate proteins.

Stroke is the leading cause of adult disability in the United States. Motor impairments often remain even after extensive physical therapy. To improve rehabilitation outcomes in severely affected individuals, we hypothesize that electrical stimulation could rewire the brain such that non-lesioned brain areas take over the function of lesioned areas. Mechanisms that modify connections in the brain rely on temporal associations on the order of tens of milliseconds. For targeted rewiring, we propose stimulation of the non-lesioned areas be triggered from activation of muscles weakened by stroke. This will provide a tight temporal association between muscle activity and brain activation and should boost the neural drive to the affected muscles. As an initial test of the proposed stimulation therapy, we trained neurologically-intact pigtailed macaques to make bimanual hand movements in response to visual cues. The macaques were implanted with electrodes at the surface of the brain that stimulated and recorded neural activity. Electrodes were located over left and right motor areas and over areas with less direct involvement in hand movements. Experimental sessions began with baseline neural activity recordings while the animal performed bimanual hand movements in the absence of stimulation. Next, movement-triggered stimulation was applied for 2-6 hours. Across sessions, a number of parameters were explored in this phase including the stimulus timing relative to movement, the stimulus intensity, and whether a motor or non-motor electrode was being stimulated. Each session ended with another recording in the absence of stimulation. The stimulation effect was quantified by changes in neural activity and behavior (i.e. reaction time between visual cue and movement onset) between pre-stimulus and post-stimulus recordings in each session. Significant stimulus-induced changes in reaction time were found to be a function of both stimulus timing and stimulus location. The results indicate that movement-triggered stimulation can powerfully modulate motor output.