Biological Sciences Student Research Showcase 2010

Neurobiology Abstracts


Development of Hippocampal Mossy Fiber Connectivity using Serial-Blockface Scanning Electron Microscopy

Joseph Antonios
Dr. Anirvan Ghosh

The circuitry of the hippocampus is highly organized, yet its assembly is less understood.  The mossy fiber (MF) pathway mediates the connectivity between the dentate gyrus (DG) granule cells and CA3 pyramidal neurons, and is characterized by unique and powerful synapses.  Mossy fiber axons send out massive MF boutons (MFBs) to interact with complex, multi-headed dendritic spines known as thorny excrescences (TEs) located on the proximal aspect of CA3 apical dendrites.  Light level analysis of the development of these synapses is limited by low spatial resolution and an inability to visualize multiple interacting components.  In this study, a new technique of electron microscopy is used to investigate the formation of mossy fiber microcircuitry.  The use of Serial-Blockface Scanning Electron Microscopy (SBSEM) allows for the reconstruction of large volumes of the developing pathway and a very high-resolution study of the synaptic ultrastructure.  The volume of reconstructed pre and post-synaptic structures allowed for a more complete understanding of the diversity of synapse morphology at different ages, and a detailed analysis of MF connectivity. The study has provided insight previously unattainable through other methods into developmentally regulated refinements of connectivity.  Future plans include applying this new technique to study the development of other synapses in the central nervous system and to study the role of select molecules in synapse formation and maturation.


Hippocampal network encoding of time-of-day in an episodic memory task

Slayyeh Begum
Dr. Jill Leutgeb

Episodic memories consist of the what, when, and where components of an experience.  They are unique in that they are formed rapidly and can require only a single exposure to an event for later recollection. The medial temporal lobe, including the hippocampus has shown in behavioral studies, to be critical for encoding ‘where’ and ‘what’ components of an event, but there is very little known about how the hippocampus encodes ‘when’ an event occurred. The goal of our study is to see how the hippocampal networks represent temporal information from events distinguishable only by the time of day in which they occur.  In our study, male Long Evans rats were implanted with a hyperdrive consisting of 12 tetrodes, in order to record action potentials from large populations of hippocampal neurons in subareas CA1 and CA3.  Rats were trained over a period of approximately 20 days in a random foraging task performed twice a day (9 am and 3pm), in the same room/location within a square and circle environment containing a single spatial cue.  The rats were allowed to forage for four sessions of 10 minutes within each environment. It has been established that CA1 and CA3 will form stable place fields and pattern separate using alterations in their firing rates to distinguish between distinct spatial contexts (Leutgeb et al., 2007).  We observed strong rate coding between the square and circle environments in both CA1 and CA3 within each time block. We next asked if there was a significant alteration in the firing rates between the morning and afternoon blocks for congruent spatial environments.  There was a subpopulation of neurons that exhibited this strong alteration in firing rates.  This decorrelation of the firing rates between time blocks suggests that the hippocampal network activity represents time-of-day information alongside the ‘what’ and ‘where’ components of an event. To test whether this activity represents time-of-day recognition or temporal sequencing, a shift day was introduced on subsequent rats where the session times were shifted to 3:00 pm and 9:00 pm.  Evidence for time of day encoding by the hippocampal networks could be important for further episodic memory studies where a temporal context is integrated alongside ‘where’ and ‘what’ information.


A functional Magnetic Resonance Imaging study of amygdalar activity in Depressed Adolescents.

Poonam Manwani
Dr. Tony Yang

Major Depressive Disorder or commonly known as MDD has been found to be a very relevant and vital problem that affects individuals across all age groups, ethnicities, and social classes. However, there is a significant lack of neuroimaging studies with depressed adolescents. Specifically literature on adults has shown that amygdala has significant influence on the autonomic nervous system (ANS), however no neuroimaging studies have been published in adolescents that examine amygdala activation and ANS activity.  The aim of this functional magnetic resonance imaging (fMRI) study was to examine the relationship between amygdala activation and heart rate variability in normal adolescents. Nineteen 12- to 17-year old adolescent subjects participated in this study. Heart rate and fMRI data were simultaneously collected while subjects performed an emotion face assessment task that reliably activates the amygdala. The emotion face assessment task produced significant amygdala activation for all facial expressions relative to the baseline task.  Activation in the left amygdala was significantly correlated with an increased sympathovagal balance (r = 0.41, P < 0.05). This data suggests that there is a direct correlation between increased amygdalar activation and depression in adolescents.


Nicotinic Receptors and the Acetylcholine Binding Protein

Phuong Thoi
Dr. Palmer Taylor

Nicotinic acetylcholine receptors (nAChRs) are ion channels which primarily modulate chemical synaptic transmission. Nicotinic receptors have nine α (alpha) type subunits and three β (beta) type subunits that are used in different sequences to regulate expression, stability, and ion channel conductance. It has been shown that acetylcholine binding proteins (AChBPs) are a valuable model for the ligand-binding domain of ligand gated ion channels. These studies have shown that the AChBPs have the cis-loop that is a definitive structural feature of nicotinic and related receptors.  Since the AChBPs are homologous to nicotinic receptors they offer the best template for obtaining high resolution structures of the ligand binding domain and are commonly used in research. Scientists generate mutant AChBPs to reveal the determining factors of ligand specificity and selectivity. These newly created AChBPs will help in the study of ligand gated ion channel function and will also be used for X-ray crystallography to generate information for the design of new drugs. In my part of this research, I will take an AChBP construct where the C loop that has already been modified to resemble α-6 and α-4 rat nicotinic receptors and transform them to resemble α-6 and α-4 human receptors using molecular biology. Once this is completed I will inject the DNA into human cells grown in culture to generate proteins which can then be used to conduct pharmacological studies on the ligand recognition and specificity.  If we find that these constructs possess interesting binding properties they will be used to grow crystals that will hopefully provide new insight into how these proteins function and aid in the development of new drugs.

POSTER # 49:

Establishing vertebrate model systems for the study of Gle1-mediated motor neuron disease

Joseph Tsai
Dr. Samuel Pfaff

Gle1 is a protein most heavily studied in yeast and is known to be involved in both mRNA export and translation in yeast and humans.  A recent study has also suggested that a mutation in Gle1 is the cause of lethal congenital contracture syndrome 1 (LCCS1) and lethal arthrogryposis with anterior horn cell disease (LAAHD), recessive fatal motor neuron diseases characterized by ventral horn motor neuron degeneration before birth.  This is particularly interesting in light of a growing pool of evidence indicating that a common denominator in many motor neuron disorders is defects in mRNA regulation.  To investigate the role of Gle1 in motor neuron development, we are using chick embryos, mouse, and mouse embryonic stem (ES) cells as model systems.  Through RT-PCR, we have shown that endogenous Gle1 transcripts are present in each species in a wide variety of cell types and stages of development, including during motor neuron development.  Expression of the mutant Gle1 protein responsible for motor neuron disease in humans failed to kill or noticeably affect the development of motor neurons in chick, suggesting that the mutant human transcript does not cause disease through a gain of function.  To study the effects of loss of Gle1 on development, we have generated a Gene-trap mouse line in which Gle1 is created as a truncated protein linked to β-Galactosidase.  Heterozygous animals have so far not shown any immediately apparent phenotype, and additional experiments are required to determine whether homozygous animals are viable.  Following failure of commercial antibodies to reliably detect Gle1 through immunohistochemistry and immunoblotting, we created a Gle1-Flag fusion construct.  Overexpression of the Gle1-Flag proteins during chicken development and in mouse embryonic stem cells undergoing differentiation into motor neurons will serve as a useful tool in future studies to localize the protein and immunoprecipitate Gle1's binding partners.  The Gene-trap animal will be further analyzed, and strategies to conditionally knock out or down the gene to examine its specific effects on motor neuron survival and function may be considered.