Diane W. Lee - US grants

Advances in our understanding of the biological bases of learning and memory from evolutionary influences down to molecular mechanisms will inevitably lead to practical applications. Understanding how memory systems evolved in response to environmental pressures can add to our understanding of how human memory systems have taken form, and thus aid in our attempt to elevate some human memory deficits. The aim of the proposed research is to increase our understanding of the ways in which the hippocampus mediates memory formation through the study of a naturalistic model of memory formation: the storage and recovery of food (caching) in birds. Results from proposed behavioral and neural lines of inquiry will address the possibility that certain memory features have become modified during evolution in response to the selective advantage of retrieval of stored food. The comparative work proposed looks at the differences between the formation of spatial memory and memory for visual cues (1) within a food-storing species and (2) between closely related species that do and do not store food. This includes an analysis of memory formation at both the behavioral and neurobiological level by investigating (1) the effects of photoperiod on short- and long-term memory (STM, LTM) for spatial and visual cues; (2) the effect of photoperiod on hippocampal size; and (3) hippocampal involvement in STM and LTM for spatial and visual cues through the use of lesions. Results will add to our current models of the biological bases of learning and memory, hippocampal structure and function, and season-mediated brain plasticity and contribute to several areas of neuroscience including neuroethology, biological psychology, and psychopharmacology.

This proposal represents a novel, integrative approach to the study of hormone-mediated and memory-induced morphological change in the avian hippocampus. In large part, birth of cortical neurons occurs only in the young developing brain and not in adulthood such that the brain is thought to be in a constant state of age-related degeneration and neuron loss. This does not appear to be the case in the hippocampus of adult birds and mammals which retains the capacity for substantial plasticity including the genesis of new neurons in response to (1) enriched environments, (2) changes in season, (3) performing specific behaviors such as storing and retrieving food, and (4) performing spatial learning tasks. The ability of the brain to respond to environmental influences, both internal (e.g., hormones) and external (e.g., learning experiences), through the process of neurogenesis has exciting scientific potential and profound implications for mental health. A recent intriguing finding is that new hippocampal cells are born as a result of damage. Since the hippocampus has retained the ability to recruit new neurons, we can use this result to investigate those factors influencing cytogenesis and differentiation of these new cells into neurons and glia. Using this model, we have the potential to start unraveling some of the factors mediating hippocampal plasticity especially neurogenesis. Our aim is to exploit the ability of the avian hippocampus to respond to environmental influences by investigating ways in which memory formation and hormones influence neural proliferation, migration, and differentiation by asking 3 basic questions: (1) Where and when are new hippocampal cells born and how long do they take to reach their destination?; (2) Does memory formation on a spatial learning task alter patterns of neuron or glia proliferation, migration, or differentiation and do estrogens change these patterns?; and (3) Is the damaged hippocampus capable of showing prolonged effects of neurogenesis resulting in recovery of function and can estrogens aid in this repair and recovery? A strong, integrative approach will be taken that exploits Dr. Lee's skills in avian neurobiology of learning and memory, and Dr. Schlinger's expertise in avian neurobiology and neuroendocrinology specifically hormone-mediated changes in behavior and neural growth. Zebra finches will be trained on a spatial learning task known to depend upon an intact hippocampus. Birds will be given lesions and the impact of these lesions on behavior as well as brain plasticity will be investigated. Effects of ovariectomy and estrogen-replacement will also be determined by (1) training ovariectomized females, both estrogen-replaced and non- replaced, on the spatial learning task, (2) lesioning the hippocampus, then (3) re-training the birds over a long period of time to determine how estrogens and behavior influence plastic changes and recovery of function in the hippocampus.

A grant has been awarded to Drs. Lee and Underwood at California State
University Long Beach to acquire an imaging system comprised of a fluorescence
-capable microscope with motorized stage, CCD camera, computer, and software.
This system will provide unbiased stereology capabilities, as well as the ability to
1) visualize immunofluorescent labeled samples; 2) produce 3-dimensional (3-D)
reconstructions; and 3) establish image databases accessible via the internet.
The goal of Dr. Lee's research is to understand the effects of memory
formation and hormones on neurogenesis in the zebra finch hippocampus. The
hippocampus of adult birds and mammals retains the capacity for substantial
plasticity including the birth of new cells (cytogenesis) and neurons
(neurogenesis) in response to a number of environmental influences including
learning and memory formation. Dr. Lee recently found that hippocampal cells
are also born as a result of injury and collect around the damaged site in the
zebra finch brain. Injured brain areas become rich in aromatase as well, an
enzyme that converts testosterone to estrogen. Estrogens, in turn, are known
to promote differentiation and survival of neurons. These intriguing results
are being used to investigate factors influencing cytogenesis and
differentiation of new cells into neurons and glia. The aim then is to exploit
the ability of the avian hippocampus to respond to environmental influences by
training birds on learning tasks, lesioning the hippocampus, injecting them
with BrdU (a mitotic marker), and assessing the patterns of cell proliferation,
migration, and/or differentiation through the use of immunohistochemical
techniques. New BrdU-positive cells will be visualized and counted using the
MicroBrightField StereoInvestigator imaging system; double labeling procedures
will be used to determine whether new cells are neurons or glia.
The goal of Dr. Underwood's research is to understand the relationship
between chromosomal abnormalities, a biased sex ratio, and the evolution of
"excess" males in the madrone butterfly. Dr. Underwood's research led to the
discovery of abnormalities in chromosome pairing during germ cell formation in
the madrone butterfly. One subspecies shows a normal complement of 26
bivalents; however, another subspecies shows extreme variations from cell to
cell within the same individual. The primary sex ratio of the abnormal
subspecies is 75% male while the normal subspecies appears to have a normal sex
ratio of 50% males. Butterflies have chromosomal sex determination.
Spermatogenesis is extremely abnormal with multivalent chromosomal associations
and lagging chromosomes formed during metaphase and anaphase, and the
production of micronuclei associated with sperm nuclei.
NSF funding will also make it possible to digitize samples obtained from a
variety of research projects. This will result in public databases to be used
by interested scientists with access to the internet and thus offers an
unparalleled opportunity to share information both locally and globally.
Individuals from multiple labs will perform procedures remotely thus broadening
the scope of research and increasing the opportunity for publication and
dissemination of results, while minimizing labor and the number of animals.
Beyond the immediate research applications, instructors and their students will
be empowered through accessibility to data previously reserved for only a
select few. Image databases, accessed as classroom exercises, will expose
students to topical and exciting new research as well as train them in the use
of 21st century technology thus advancing science and giving them a competitive
edge.

An award has been made to California State University-Long Beach under the direction of Dr. Bruno Pernet for the acquisition of a laser scanning confocal microscope for research and teaching. The new microscope will support studies of cells and their internal structures at a finer resolution than currently possible. The microscope will support a dozen faculty members in biology, chemistry, and psychology, including several newly hired faculty. Research that will benefit includes studies of growth and development of fungal and mammalian cells, development of grass plants, gene expression in muscle cells, and neural structure and function in birds. The institution has a large Hispanic population, who can be expected to participate in research projects and to benefit from the instrument in their classes. Graduate students will use the instrument in their Masters Degree research.