Heather Eisthen - US grants

Evolutionary changes in the nervous system offer a tremendous
opportunity to understand the relationship between the nervous system and
behavior. When new brain structures are created, what does this do to an
organism's behavior? How do these new structures interact with other
portions of the nervous system? We are using
the vertebrate olfactory system (the system responsible for the sense of
smell) as a model for examining evolutionary changes in the nervous system.
To this end, we are exploring the organization of the olfactory system in
a variety of vertebrate animals.
Recently, scientists have learned that receptor cells in the nose
of some fishes and amphibians are connected to the a region in the base of
the forebrain, instead of to the olfactory bulbs, as is usual for these
cells. The structure of this newly-discovered subsystem, the "extra-bulbar
pathway," is poorly understood, and its function has not been investigated.
In the proposed experiments, the location of the receptor cells of the
extra-bulbar pathway within the nose will be examined in adult axolotls
(Ambystoma mexicanum), a salamander species that lives in water throughout
its life. Single-cell electrophysiological recording methods will be used
to study the odorant response properties of the receptor cells of the
extra-bulbar pathway in axolotls. These experiments will allow us to
examine the function of the extra-bulbar pathway, and
to test an interesting hypothesis that suggests that the extra-bulbar
pathway functions in detection of pheromones. Pheromones are chemicals
produced by animals that carry information to other members of the same
species to tell them about the producer's sex or reproductive status.
These experiments will provide the first clear description of the
organization and function of the extra-bulbar pathway. This research has
important implications for our understanding of the ways in which odorant
information is converted into electrical signals in olfactory receptor
cells, as well as the way odorant information is processed in the olfactory
bulbs, potentially shedding light on the neural mechanisms underlying odor
perception.

DESCRIPTION (provided by applicant): The terminal nerve is an anterior cranial nerve that extends between the nasal cavity and hypothalamic/ preoptic area in all classes of jawed vertebrates, including humans. The function of the terminal nerve is poorly understood, but recent research indicates that this peptide-rich nerve serves a neuromodulatory function. The research described in this proposal will elucidate the neuromodulatory effects of terminal nerve-derived peptides on olfactory receptor neurons. The current project has four Specific Aims: (1) to identify and sequence NPY-superfamily and/or FMRFamide-like peptides present in the terminal nerve; (2) to localize terminal nerve cells and fibers containing NPY-superfamily and/or FMRFamide-like peptides relative to those containing another peptide, gonadotropin releasing hormone (GnRH); (3) to characterize the effects of terminal nerve-derived peptides on electrophysiological properties and odorant responses from olfactory receptor neurons; and (4) to investigate the relationships among modulatory effects produced by different terminal nerve-derived peptides. A combination of molecular, biochemical, anatomical, and electrophysiological techniques will be used. This research will contribute to two interrelated goals. The primary goal is to understand the nature of peripheral processing of odorant information. People suffer from olfactory system dysfunction from a variety of causes, including metabolic disorders, hormonal imbalance, and exposure to toxic chemicals. Medications can also alter olfactory system function. Some of the olfactory aberrations associated with these factors may be due to interference with neuromodulatory mechanisms. The secondary goal is to contribute to a general understanding of the physiological function and mechanisms of action of neuropeptide Y (NPY) and other members of this superfamily of peptides. Although NPY is the most prevalent peptide in the brains of mammals, including humans, the actions of this peptide at a cellular and system level are virtually unexplored. NPY has been implicated in the control of feeding and anorexia, as well as seizures, memory disorders, anxiety and depression, and heart problems. In the research proposed here, olfactory receptor neurons will be used as a model system for studying the function and mechanisms of action of NPY.

Heather L. Eisthen
0817785
Centrifugal Modulation in the Vertebrate Olfactory Epithelium

The brain sends signals to the sense organs to focus attention on the stimuli that are most important at any given time, depending on the animal's behavioral context or physiological state. Although this type of activity has been described in the eye and ear, it has received little attention in the nose, or olfactory organ. Research by Dr. Eisthen and her collaborators suggests that the terminal nerve, a nerve that extends between the brain and nose, releases chemicals into the olfactory organ that change its activity. Specifically, the terminal nerve contains two chemicals, gonadotropin releasing hormone (GnRH) and neuropeptide Y (NPY), that change odorant responses and excitability of olfactory neurons in the nose. The effects appear to depend on the animal's overall physiological condition: GnRH is more likely to affect olfactory receptor neurons during the breeding season, and NPY?s effects are limited to hungry animals. This is interesting, because in other parts of the brain, GnRH functions to control release of hormones involved in reproduction, whereas NPY is involved in controlling appetite and hunger. The project will involve a combination of anatomical, electrophysiological, molecular, and biochemical techniques, and are expected to reveal how specific these effects are, as well as the cellular and molecular changes that underlie these effects. The research will contribute to the training of students and researchers at Michigan State University, and involves cooperation with scientists in other parts of the United States as well as in Japan and Sweden.

A Research Experience for Undergraduates (REU) Site award has been made to Michigan State University, providing research training for 10 students during the summers of 2013-2015 focusing on the Integrative Biology of Social Behaviors. Participating students will receive 10 weeks of hands-on research experience with projects involving a wide range of model organisms (rodents, carnivores, birds, lizards, salamanders, fish, insects, and digital organisms) and levels of inquiry (molecular, cellular, systems, individuals, populations). Twelve faculty members in the departments of Fisheries and Wildlife, Neuroscience, Psychology, and Zoology will serve as mentors. In addition to their research work, students will benefit from weekly journal clubs and research presentations from the mentoring faculty; will participate in modules related to research ethics, statistics, laboratory safety, applying to graduate school, and careers in the biological sciences; and will participate in outreach activities at a local nature center. At the end of the program, students will present their activities at a university-wide undergraduate research symposium. These activities will sharpen the students' critical and scientific thinking skills, provide a unique opportunity to gain experience with laboratory and field research methods, and help prepare the next generation of researchers studying the biology of social behaviors. Undergraduates entering their junior or senior year will be recruited through personal contacts and by mailings and emails targeted to institutions that can provide the most diverse pool of candidates. Students will be selected based on academic record, any previous research experience, letters of recommendations, and potential for outstanding research in behavioral biology. Students will be tracked in part via Facebook to determine their continued interest in their academic field of study, their career paths, and the lasting influences of the research experience. Assessments of interns and participating faculty members will be comprised of a REU common assessment tool (Undergraduate Research Student Self-Assessment) and other questionnaires, as well as one-on-one interviews that assess interns' topical knowledge in the field, their sense of community, and their knowledge about professional development. More information is available at www.msu.edu/~ibsb, or by contacting the PI (Dr. Joseph Lonstein at lonstein@msu.edu) or the co-PI (Dr. Heather Eisthen at eisthen@msu.edu).

Leading researchers in the field of Neuroethology from around the world meet at the International Congress of Neuroethology to share their research findings and forge research collaborations that are critical in moving the discipline forward. Research in neuroethology is at the cutting edge of neuroscience and behavioral research with basic biological and health related implications. For U.S. researchers to remain current, it is essential that they exchange ideas directly with their international colleagues. The Congress will emphasis comparative evolutionary approaches which meets the biodiversity initiatives of NSF as well as fostering development of scientists that may be involved in the Brain Initiative. The exchange of information among scientists during oral and poster presentations is of great value, but equally important are the personal exchanges and resulting collaborations established in the hallways between sessions and during the numerous topic-based workshops and symposia scheduled throughout the congress.

Funds provided by NSF will help support the attendance of future leaders of in neuroethology and behavioral neuroscience. Specifically, recipients will be at an early stage in their careers (e.g., graduate students, post-doctoral fellows, and beginning investigators) and will be from U.S. institutions. The selection committee will also strive to support individuals from groups that are historically underrepresented. Assistance of such scientists will help forge new, international collaborations, thereby creating long-term benefits for all involved.

Understanding how organisms communicate with one other and how communication systems arise and change over time is a major challenge in biology. This project provides a rare opportunity to investigate the origin and diversification of a chemical communication system at the molecular level in amphibians (salamanders and frogs). In addition, this research can provide a novel framework for applications in computer science, where current research is focused on building systems that can evolve and use communication to accomplish jobs and then adapt as needs change. The project will also contribute to the training of a postdoctoral researcher and numerous undergraduate students in collaboration with MSU's BEACON Center and an ongoing REU site award.

Specifically, the objectives of this work are to trace the evolutionary link between compounds that amphibians produce in the skin as antimicrobials and their later use as pheromones; to purify and test the efficacy of those compounds as antimicrobials and pheromones; and to identify the molecular mechanism used to smell those compounds. The results will be obtained through the use of bioinformatics and genomics to trace the genetic history of genes, protein biochemistry to isolate and test the gene products, behavioral experiments to determine which compounds function as pheromones, and electrophysiology to identify the neuronal mechanisms responsible for sensing the pheromones. Results from these studies will be disseminated through peer-reviewed publications and through presentations at scientific conferences. All data will be available upon request and deposited at Dryad and the NCBI Trace archive.

Non-Technical Paragraph

Rough-skinned newts (Taricha granulosa) are among the most toxic animals known: some individuals possess enormous quantities of tetrodotoxin (TTX), which prevents brain cells (neurons) from signaling to each other. Scientists know a great deal about how TTX normally blocks neural activity, but not much about how animals that possess TTX are able to resist its effects. The ability of newts to resist TTX is particularly puzzling because several of their genes must be mutated in concert, as a mutation in only one affected gene would leave the newt vulnerable to TTX's affects, and likely dead. Strangely, newts are not simply resistant to TTX's effects, but are actually attracted to the smell of TTX. In the proposed work, the team will use well-established methods to quantify the distribution of TTX inside newts' bodies, identify mutations that are likely involved in TTX resistance, examine the effects of TTX on their neurons, and discover how the neurons involved in smelling are activated by TTX. This basic research will help scientists understand how neurons work at a molecular level, which will increase or understanding of the "nuts and bolts" of healthy brain function, as well as how animals adapt to the presence of toxins. Undergraduate students from under-represented groups, including students from Native American populations in Oregon, will be recruited to participate in summer research programs associated with the work. These experiences will help prepare a diverse population of students for future careers in STEM related fields.

Technical Paragraph

TTX is toxic because it blocks voltage-gated sodium channels (NaVs), essential for the generation and propagation of action potentials. The proposed work will identify where the different forms of NaVs are expressed in neurons and muscles in newts, identify specific mutations in these channels that allow them to resist the toxic effects of TTX, and determine how these structural changes alter neuron function. Using analytical chemistry, histochemistry, and molecular biology, levels of TTX in different tissues as well as the location and structure of the six different NaVs will be examined in both highly toxic and non-toxic newts to quantify levels of TTX resistance and identify mutations that confer resistance. The electrophysiological properties of NaVs in a heterologous expression system and of neurons in the brain of newts will be characterized to determine whether and how TTX resistance alters channel and neuron function. In addition, using a combination of techniques, the adaptations underlying TTX detection in the olfactory epithelium will be identified to understand how this unusual ability evolved. The proposed work will contribute to understanding a physiologically vital class of ion channels, as well as the ways in which evolution at the molecular level shapes nervous system function and animal behavior, thereby deepening our understanding of how neurons function and change over evolutionary time. In addition, the proposed work will immerse undergraduate students in the nature and practice of science, particularly through the involvement of Williamette University students in an REU program at Michigan State University.