The following research projects will be presented at the Robert B. Glassman Memorial Brain, Mind, and Behavior Symposium during Brain Awareness Week on Wednesday, November 13 at 5:45 p.m.
Creation of α-synuclein truncation variants to test Parkinson’s disease relevance in yeast.
Sarah Chiren ’16, Charles Alvarado ’16, Jyothis James ’16, Alex Roman ’16
Parkinson’s disease (PD) is a hypokinetic neurodegenerative disorder characterized by the death of midbrain dopaminergic neurons. This selective cell death is linked to the misfolding and aggregation of the brain protein α-synuclein that accumulates as Lewy bodies. The full-length α-synuclein (140 amino acids long) associates with membranes and its carboxyl-terminus keeps it soluble. In PD, this full-length form is the major component of Lewy bodies, although several carboxyl-terminal truncation variants (α-syn 103, 110, 120, and 123) were also recently found in them (Lewis et al., 2010). While these variants can increase the aggregation of the full-length α-synuclein in vitro and enhance toxicity in specialized cell cultures (Li et al., 2005; Liu et al., 2005), the individual properties of each variant towards aggregation, membrane association, and toxicity in free living organisms is not well studied. In this first-year Richter Scholar project, we sought to test the hypothesis that the larger the truncations, the more the variants would reduce α-synuclein solubility and membrane association, and increase toxicity in organisms. Our goal was to create these four variants of α-synuclein (in both wild-type and two familial PD mutant versions- A30P and A53T) and characterize their properties in a budding yeast (Saccharomyces cerevisiae) model for PD. In this poster, we report the creation of all twelve variants and their successful transformation into yeast. The next goal is to evaluate several properties of these variants by comparing them to the full-length form: their cellular localization (GFP imaging), expression/accumulation (Western blotting), and toxicity (serially diluted growth on plates).
Stress, Sex, & the Brain: Functional Neuroanatomy of the Basolateral Amygdala
Anhar Mohamed ’14
Females are at a higher risk for developing stress and anxiety disorders than males. In previous studies, female rats showed a heightened fear response, which is linked primarily to the basolateral amygdala (BLA), the brain structure associated with regulation of emotion. Despite a trend of sex differences in stress disorders and the knowledge that BLA activity is modulated via small-conductance calcium-activated potassium (KCa) channels, it is not clear what influence gender has on KCa channels in the BLA. The purpose of this thesis was to examine the hypothesis that neurons in female rat amygdalae are more excitable than those of males due to decreased expression of KCa channels. To test this hypothesis, we characterized KCa channel expression in the BLA, using immunohistochemistry for protein expression and in-situ hybridization for gene expression. We compared results between females at varying stages of their estrous cycle to males. Our preliminary data, not yet statistically analyzed, shows a trend consistent with our hypothesis that females with decreased estrogen exhibit similar KCa channel expression to males, and both exhibit less than females with elevated levels of estrogen. Once completed, these studies will aid in understanding mechanisms for sex differences in anxiety disorders.
Investigation of Spartin in Troyer Syndrome Using a Worm Model
Natalie Zemela ’15
Hereditary spastic paraplegias (or Troyer syndrome) is characterized by the spasticity and muscle weakness of the lower limbs. It is an autosomal recessive genetic disorder caused by a mutation in the SPG20 gene, which codes for production of a protein called spartin. This protein is thought to have diverse functions (endosomal trafficking, microtubule assisting), multiple organelle locations (mitochondria, lipid droplets) and is expressed in different cells (neurons). To test the hypothesis that loss of gene function leads to Troyer syndrome, we studied a worm model that expressed the knockout of the SPG20 gene. Specifically, we tested movement by behavioral methods (reversal intervals) and stress by oxidative conditions (survival rate). Our preliminary data indicates that spartin knockout worms had less frequent reversal intervals than the wild-type worms, and the spartin knockout worms lived shorter than their wild-type counterparts.
Regulation of Cell Volume by Alligator Red Blood Cells
Kayla Huber ’16, Rahul B. Thakuri ’16, Rachel Granberg ’16
The ability to regulate volume is a fundamental process of cells necessary to maintain homeostasis. In this study, we examined regulated volume decrease (RVD), a process by which cells compensate for changes that cause cell size to inappropriately increase. This was accomplished by using alligator red blood cells as a model system and measuring the volume of cells under different conditions and in the presence of specific pharmacological agents. We found that potassium efflux from cells was necessary for RVD. In fact, potassium efflux was a rate limiting step and it was stimulated by a volume-sensitive signal transduction system relying on the calcium-activated enzyme phospholipase A2.
Social Fear Learning: The Importance of Prior Interactions
Kayla Sarkis ’14
Social learning is important for coping with threatening environments and developing social abilities. Prior individual experiences effects the transmittance of fear through social observation and influences the way the observer reacts to future situations . However, it was unclear how much social interaction must occur in order for that learning to take place. In this study, we sought to test whether rats who engage in more social interaction are capable of learning more easily than those who do not.
Focus, Breath, Relax: How Not to Get the Flu!
Jessica Dudley ’14
What is meditation: Meditation is a practice of concentrated focus upon a sound, object, visualization, the breath, or attention itself in order to increase awareness about the present moment, reduce stress, promote relaxation, and enhance personal and spiritual growth. When most humans hear the word meditation, usually it denotes only spiritual and religious practices. However, much research is underway to investigate the physiological effects and underlying mechanisms of the practice of meditation. Because of this, mediation has been incorporated into an increasing number of therapeutic interventions to help in the physical and mental well-being of the individual. The present study reviews the neurophysiological effects of meditation on stress and the results can relieve stress and ultimately keep you from getting the flu.
The Effects of the C-terminus on Alpha-Synuclein
Katrina Campbell ’14 and Maribel Munoz ’16
Parkinson’s disease (PD) is the second most common neurodegenerative disease, second only to Alzheimer’s disease. In patients with PD, dopaminergic neurons in the substantia nigra die. These neurons are an integral part of the system that helps coordinate movement. Because of the loss of these neurons, PD patients often experience hypokinesia, an overall slowing down of movement. Within a PD affected substantia nigra, there are proteinatious aggregates called Lewy bodies. These Lewy bodies are comprised mainly of alpha-synuclein. Alpha synuclein is a small protein, 140 amino acids, and has three main domains, the N-terminus, the NAC domain, and the C terminus. The N terminus allows the protein to bind to membranes. The NAC domain is hydrophobic and prone to aggregation. The C terminus; however, is acidic and is proposed to keep alpha-synuclein soluble. My project involves truncating alpha-synuclein at the c-terminus to make versions of alpha-synuclein 103, 110, 120, or 123 amino acids long. I hypothesize that as more of the C terminus is truncated, the more alpha-synuclein will be prone to aggregation.
Identifying Specific Amino Acids in Parkinson’s Disease Protein Alpha-Synuclein that Control its Toxic Properties
Galina Lipkin ’15
Parkinson’s disease (PD) is caused by the death of midbrain neurons linked to the aggregation of a membrane-associated protein called alpha-synuclein. Recent studies have suggested that certain amino acids within alpha-synuclein mediate its membrane-association and aggregation, but more evidence is needed. In this study, we first mutated each amino acid to either block or enhance its proposed contribution. Using two yeast models, we then studied each mutant’s ability to associate with membranes, aggregate, and be toxic. So far, we found that each amino acid regulates membrane association and aggregation, but effects on toxicity are less clear and need more analysis.
Using antisense oligonucleotides as a therapy for batten disease (Juvenile neuronal ceroid lipofuscinoses)
Maria Ruiz ’16
Who needs a brain? Multiple behavioural strategies exhibited by an animal without a centralized nervous system
Yusuf Ismail ’15
Defensive behaviour in the sea anemone Stomphia coccinea was observed in order to test how rich a behavioural repertoire the organism possesses despite lacking a centralized nervous system. Like other anemones, Stomphia have no brains – they make do with a primitive nervous system consisting of a simple, distributed nerve net. We tested Stomphia’s responses to contact with two of its natural predators, the nudibranch sea slug Aeolidia papillosa and the sea star Dermasterias imbricata. Studies were conducted at Friday Harbor Laboratories, Friday Harbor, WA which is the natural habitat of Stomphia. Because Stomphia are famous for being able to swim to get away from predators, they were able to be collected and studied here. Numerous Stomphia were exposed to each predator over multiple trials and at different orientations with respect to a marker on the columns of each anemone. Results consistently showed an attack and hide strategy in response to Aeolidia involving repeated cycles of directed attack sometimes involving 360o tentacle whirl, tentacle withdrawal, re-emergence and directed attack, and withdrawal again. Conversely, a defensive escape strategy involving tentacle withdrawal, column extension, detachment and swimming was exhibited in response to Dermasterias. This diversity of behavioural strategies and response components is remarkable for an animal with no centralized nervous system. Our findings have broad implications in neurobiology and evolutionary biology as they pose important considerations of what centralized brains are needed for.
Manipulation of RNA Splicing of APOER2 with Antisense Oligonucleotides to Rescue Spatial Memory in Alzheimer’s Disease Mice
Rida Khan ’14
Alzheimer’s disease (AD) is a devastating neurodegenerative disease afflicting every 1 in 8 individuals above the age of 65. This highlights the need to identify novel targets and develop more efficient therapies. I aim to use cellular and mouse models of AD to validate therapeutic targets and develop treatments for AD based on modulation of gene expression using ASOs. APOER2, which is one of the targets, is a post-synaptic APOE and Reelin receptor that is part of the signaling that controls neuronal migration during brain development and induces long-term potentiation in the adult brain. Reelin signaling through APOER2 requires amino acids encoded by the alternatively spliced APOER2 exon 19. Preliminary studies in our lab found that decrease in splicing of this alternative APOER2 exon is associated with AD. We designed and tested ASOs targeted to the regions flanking exon 19 splicing in vivo when injected into the brains of mice. My project aimed at determining the effects of ASOs on memory in mice with the transgenic human AD gene. These mice were injected with the specific ASOs and memory was analyzed using open-field habituation, novel object recognition test and Morris water maze. Preliminary results show promising spatial memory rescue in AD-like mice treated with the ASO compared to AD-like mice treated with control ASO. This accentuates the role of ASOs as therapeutic agents for AD.
Steals clock, faces time: Examining the relationship between humor and lexical ambiguity in newspaper headlines
Krista Miller ’09
This study explored whether the perception of humor in newspaper headlines is influenced by the presence of lexical ambiguity and framing narratives (satirical or non-satirical newspaper). Participants made humor and ambiguity judgments for the headlines. Headlines containing a lexical ambiguity were rated as more humorous than disambiguated headlines and control headlines (no ambiguity). Framing information did not influence ratings. It appears that lexical ambiguity influenced the perception of humor whereas framing narratives did not.
Immersion in Elementary School Enhances Inhibitory Control
Joanna Bovee ’06
Several studies provide evidence of enhanced executive control for early bilinguals, especially on tasks requiring inhibitory control (e.g., Martin-Rhee & Bialystok, 2008). Whether such effects are evidenced in second-language learners and what type of experience is necessary to develop such effects are open questions. We administered an executive control task, the Simon task, to native English high school students who were enrolled in a Spanish class. A group of participants had also been enrolled in a partial Spanish immersion program (approximately 10 hours a week) throughout elementary school. When compared to non-immersion students, the immersion students evidenced a smaller Simon reaction time cost (difference between incongruent and congruent trials). This finding suggests that early exposure to a second language in an immersion environment can provide the benefits typically found in early bilinguals.
The Joint Influence of Collaboration and Part-Set Cuing
Thomas Estruth ’14
The purpose of this study was to investigate the phenomenon of Part-Set Cuing. Part-Set Cuing is described as newly encoded information that is part of a set of to-be-remembered information. There are two main effects of part-set cuing: (a) part-set cuing inhibition, which refers to the finding that hints, specifically part of the set of to-be-remembered information, often impair memory performance in free recall tasks, and (b) part-set cuing facilitation, the finding that these hints enhance memory. Participants viewed the construction on snap circuit objects and then attempted to reconstruct the objects either in the presence or absence of part-set cues. Subjects participated in at least one of two experiments in which they watched a video on the construction of snap circuit objects. Once the video was completed, subjects used a snap circuit board to re-construct what was shown to them in the video. Results included that robust part-set cuing facilitation on the spatial memory tasks. These results support predictions that retrieval strategy disruption hypothesis and the two and three-mechanism accounts of part-set cuing.
Part-Set Cuing Facilitation for Spatial Information
Sydni Cole ’12
Part-set cuing inhibition refers to the counterintuitive finding that hints—specifically, part of the set of to-be-remembered information—often impair memory performance in free recall tasks. Although inhibition is the most commonly reported result, part-set cuing facilitation has been shown with serial order tasks. The present study examined the influence of part-set cuing for spatial locations using novel methods and materials. Participants viewed the construction of Snap Circuit objects and then attempted to reconstruct the objects either in the presence or absence of part-set cues. Two experiments revealed robust part-set cuing facilitation on the spatial memory tasks. Generally, these results are consistent with the predictions of the retrieval strategy disruption hypothesis (e.g., Basden & Basden, 1995) and the two- and three-mechanism accounts of part-set cuing (Bäuml and Aslan, 2006; Bäuml & Samenieh, 2012).
Repeated adolescent CB1 cannabinoid receptor activation impairs the maturation of prefrontal corticalGABAergic function in adulthood
Daryn Cass ’10
Converging epidemiological studies indicate that cannabis abuse during adolescence significantly increases the risk of developing psychosis and prefrontal cortex (PFC)-dependent cognitive impairments later in life. However, the mechanisms underlying the adolescent susceptibility to chronic cannabis exposure are poorly understood. Given that the CB1 cannabinoid receptor mediates the psychoactive action of cannabis, the present study was designed to determine how repeated exposure to the CB1 receptor agonist WIN during adolescence impacts the functional maturation of the PFC network. By means of local field potential (LFP) recordings and ventral hippocampal stimulation in vivo, we found that a history of WIN exposure during early (P35-40) or mid-(P40-45) adolescence, but not in late adolescence (P50-55) or adulthood (P75-80), is sufficient to yield a state of frequency-dependent prefrontal disinhibition in adulthood comparable to that seen in the juvenile PFC. Remarkably, this prefrontal disinhibition could be normalized following a single acute local infusion of the GABA-Aα1 positive allosteric modulator Indiplon, suggesting that adolescent exposure to WIN causes a functional downregulation of GABAergic transmission in the PFC. Accordingly, in vitro recordings from adult PFC slices obtained from rats exposed to WIN during adolescence further demonstrate that local GABAergic transmission onto layer V pyramidal neurons is markedly reduced to the levels seen in the P30-35 PFC. Together, these results indicate that early and mid-adolescence constitute a critical period during which repeated CB1 receptor stimulation is sufficient to elicit a state of PFC network disinhibition resulting from a developmental impairment of local prefrontal GABAergic transmission that endures through adulthood.
The Influence of Cuing on Spatial Memory for Chess Positions
Jennifer Salgado-Benz ’16
In our study we explored the sequential effects of part-set cuing and chunk type in spatial memory. We built upon previous studies on part-set cuing and spatial memory using chess boards, by Watkins et al. (1984), and snap circuits, by Kelley (2013). Snap circuits were more successful in showing part-set cuing facilitation, while it was not shown in studies with chess boards. We hypothesized that showing participants a chess board with chess pieces close together will facilitate spatial memory retrieval. To test our hypothesis we set up four chess boards with either densely and sparsely placed pieces and cued and uncued pieces when the participants reconstructed the boards. Our results showed that there was a significant interaction of part-set cuing between densely placed pieces and cued reconstruction but showed no significance between sparse and cued reconstruction. This could be due to participant unfamiliarity with chess pieces which resulted in them having more difficulty recalling chess piece positions.
Chronic downregulation of the sGC-cGMP signaling pathway as a therapeutic target in Parkinson’s Disease
John Vinkavich ’15
Parkinson’s Disease (PD) is a degenerative disorder of the central nervous system marked by severe motor deficits (akinesias) and loss of dopaminergic cells in the subtantia nigra (SN) area of the brain. The current primary treatment for PD is levodopa which replaces missing dopamine in order to alleviate motor symptoms. Unfortunately, levodopa has many severe side effects; namely uncontrollable motor movements called dyskinesias. There is a necessity to identify novel non-dopaminergic mechanisms as new therapeutic targets for Parkinson’s disease. In the indirect pathway of PD, the soluable guanylyl cyclase -cyclic GMP- protein kinase A (sGC-cGMP-PKA) portion of the Nitrous Oxide (NO) signaling cascade has emerged as one promising candidate for pharmaceutical intervention. Using selective sGC inhibitor 1H-[1,2,4] oxadiazolo-[4,3-a]quinuxalin-1-one (ODQ) as a way to reverse basal ganglia dysfunction, akinesia in a 6OHDA rat model of PD was examined using electrophysiological, behavioral, and histological analyses.
Evaluation of alpha-synuclein splice variants in yeast to better understand Parkinson’s Disease
Saul Bello Rojas ’16 and Khadijah Hamid ’16
Parkinson’s disease (PD) is a hypokinetic neurodegenerative disorder characterized by the death of midbrain dopaminergic neurons. Misfolded proteins and aggregated α-synuclein leads to the accumulation of Lewy Bodies. While the full-length α-synuclein (which is 140 amino acids long) is the major Lewy body component, alternatively smaller versions of α-synuclein include splice variants (α-syn 126, α-syn 112, and α-syn 98; Beyer et al, 2008, McLean et al., 2011). The individual contributions of each such splice variant towards α-synuclein-based PD pathology is not known. We hypothesized that splice variants likely contribute to PD as they lack regions that keep α-synuclein soluble or bound to membranes. In this study, we created the three splice variants for yeast expression, attached with and without a localization tag - green fluorescent protein (GFP). Here, we will describe the steps and data that led to creating the variants. Next summer, we plan to evaluate their aggregation, localization, and toxicity properties, compared with the full length α-synuclein.
Medical Mysteries of Sleep and Wakefulness
Mallory Burney ’17, James Haney ’17, Kevin Kupiec ’17, Danielle Shaw ’17, Lily Veldran ’17, Grant Zimmerman ’17
Medical Mysteries of Sex and Sexuality
Alexandra Begoun ’17, Milan Bozic ’17, Cam Gudmundson ’17, Lauren Lyon ’17, Marina Rawlings ’17, Alex Silvola ’17
Medical Mysteries of Emotions
Pooja Acharya ’17, Joseph Bortollotti ’17, Logan Graham ’17, Peyton Schrag ’17
Does Linear or Circular DNA Work Better?
Fatima Hooda ’16 and Tyler Kaplan ’16
Telomeres are nuceloprotein complexes that protect the ends of eukaryotic chromosomes. During DNA replication, chromosomes cannot be completely copied causing telomeres to shorten overtime. Shortened telomeres are characteristic in cellular abnormalities, such as cancer. However, the enzyme telomerase, lengthens telomeres to prevent crucial genetic material from being deleted. Telomerase is composed of a protein (TERT) and a RNA template (TER), which contains the sequence that is complementary to the telomeric repeats. In our lab, the model organism is Aspergillus nidulans because of its short, highly regulated telomere length. Currently, the lab is attempting to perform transformation in A. nidulans. PyrG serves as a nutritional marker because it produces uridine and uracil. PyrG89 is a mutation where uridine and uracil cannot be produced, thus it must be provided. First, PyrG, will be swapped with PyrG89 to A. nidulans and it will be grown on a plate without uridine and uracil. This transformation will be completed using a DNA plasmid (circular DNA), but we hope to eventually perform transformation with linear DNA. This summer, we will isolate an A. nidulans plasmid from Escherichia coli and isolate the wild-type PyrG gene from the plasmid into linear DNA for future projects. In the future, PyrG and a telomerase mutant gene will be added to A. nidulans to study the phenotypic effects.