Paul J. Yarowsky - US grants

Although astrocytes constitute the major cell type in the mammalian central nervous system (CNS), their functional properties in maintaining homeostatis during neuronal activity have been reconsidered, with the ability to prepare homogeneous cell cultures. Using these cultures, recent studies indicate that this previously considered electrically silent cell possesses voltage-sensitive channels for sodium, calcium, potassium and chloride. Voltage-sensitive channels meditate changes in membrane permeability to selected ions in response to a change in membrane voltage. This project deals specifically with types of voltage-gated sodium channels in astrocytes. In electrically excitable tissue, two distinct types of sodium channels have been found. These types of voltage-gated channels have separate binding sites for saxitoxin and tetrodotoxin. Rat brain astrocytes also have these types of sodium channels. They are present in differing proportions at early and late stages of astrocyte maturity. Using neurotoxin-activated 22Na+ flux, intracellular and whole-cell recording techniques and toxin binding methods, physiological and pharmacological properties of both types of astrocyte sodium channels will be compared to properties in excitable membranes; the density of sodium channels will be discovered at different stages of morphological differentiation; electrical excitability will be determined in early and late astrocyte cultures; and factors that trigger or retard the expression of toxin-insensitive channels and toxin-sensitive channels will be explored. In addition, an astrocyte membrane preparation will be incorporated into planar lipid bilayers in order to study the biophysical characteristics of single toxin-insensitive and toxin-sensitive sodium channels and to compare their properties with those from electrically excitable tissue. The results of this proposal should provide a new basis for understanding some of the complex interactions between neurons and glia and whether astrocyte sodium channels are similar to those in nerve and muscle.

DESCRIPTION: (adapted from applicant's abstract) The goal of this research program is to investigate the development of the cerebral cortex of the segmental trisomy 16 (Ts65Dn) mouse, a potential animal model of Down syndrome (DS, trisomy 21). Many of the same chromosome 21 genes triplicated in DS are also triplicated in Ts65Dn and therefore, DS and Ts65Dn share a common genetic abnormality. Both DS and Ts65Dn have deficits in spatial learning and memory that are consistent with functional impairment in the frontal cortex. However, there is little information available about the development of the frontal cortex in Ts65Dn. The investigators hypothesize that the spatial learning and memory deficits in Ts65Dn are due to abnormalities in the generation, maturation, and survival of neurons during prenatal neuronogenesis and/or early postnatal neocortical development. The proposed experiments probe the behavior of neuroblasts and young postmitotic neurons in the developing neocortex. During the neuronogenic period, neurons are born and migrate to their final locations and the formation of connections begins. Abnormalities occurring during this period could lead to delayed formation of the pallial layers and cortical connections. During early postnatal life, synapses are formed, interneuronal connections are firmly established, and extra neurons are removed by programmed cell death. Abnormalities during this period could lead to abnormal dendritic organization and synaptogenesis, loss of connections and inappropriate programmed cell death. Defects in neuroblast proliferation, synaptogenesis, connectivity, and programmed cell death during neocortical development could lead to permanent cortical abnormalities responsible for the cognitive deficits in Ts65Dn. Similar defects in human brain development could lead to cognitive deficits in DS.

DESCRIPTION (provided by applicant): The objective of the 10th International Meeting of Chromosome 21 & Down Syndrome is to bring together investigators from the international community with a broad spectrum of research interests to discuss the current state of knowledge regarding Down syndrome and human chromosome 21. Topics for discussion are designed to encompass all relevant areas of biology, molecular biology, and medicine, including functional genomics and neurobiology. Invited speakers will present an overview highlighting topical issues. These introductions will be followed by platform presentations from submitted abstracts. The organizers anticipate a small but highly interactive meeting to facilitate the exchange of the latest ideas between senior and junior scientists, including postdoctoral fellows. Following the sequencing of chromosome 21, researchers are putting the results of these molecular analyses into biological context. This can best be done with a multidisciplinary group of researchers including those specializing in the mental retardation and cognitive deficits of Down syndrome, the neurobiology of Down syndrome, congenital heart defects, immune deficits, leukemia, craniofacial abnormalities, communicative and digestive disorders, and premature aging, as well as genomic research, transgenic mouse construction and analysis, and mouse behavioral studies. It is anticipated that this conference will foster interaction across disciplines, help to focus and redirect research efforts for maximal use of developing data, and establish new collaborations and avenues of investigation.

DESCRIPTION (provided by applicant): Down syndrome (DS) is the most common genetic form of mental retardation. One of the specific hallmarks of DS is anomalous cerebral cortical development. Studies have found cortical hypoplasia, abnormal lamination, reduced synaptogenesis, aberrant dendritic development, and a delay in myelination. These abnormalities lead to abnormal cortical organization and circuitry. Because of the lack of adequate DS tissue, it has been difficult to determine when abnormalities in cortical development first occur. Newer methods of brain imaging especially MRI have allowed studies of brain development to be done 3-dimensionally, without the invasiveness of histological methods. The present experiments will investigate development of the cortex during fetal development in DS compared to normal human development and also compare cortical formation and growth in DS with an animal model of DS, the segmental trisomy 16 mice (Ts65Dn). Many of the same chromosome 21 genes triplicated in DS are also triplicated in Ts65Dn and therefore Ts65Dn and DS share a common genetic abnormality. Using high resolution NM and the new method of resolving water diffusion in situ, diffusion tensor imaging (DTI) fiber bundle architecture and cortical organization will be resolved. We will characterize the volume of cortical regions, delineate the microstructure of the cortex and the architecture of major fiber bundles present during the fetal period. We have a unique collection of postmortem fetal DS brains (18-26 weeks gestation), a period of formation of future cortical organization. We will directly address the hypothesis that cortical volume and the size and shape of specific fiber bundles is abnormal in DS. In Aim 1, we will characterize the volume of cortical regions in fetal DS and Ts65Dn using MRI. In Aim 2, we will characterize cortical microstructure and fiber bundle architecture in DS and Ts65Dn using DTI. Thus we will not only characterize fetal development in DS and Ts65Dn, but also determine how similar they are. Defects during fetal neocortical development could lead to permanent cortical abnormalities responsible for cognitive deficits.

Parkinson's disease (PD) is a progressive incurable neurological disease that affects 1-2 million people in the U.S. The cardinal motor symptoms of PD are believed to be due to the degeneration of dopaminergic neurons in the pars compacta of the substantia nigra. However, patients with PD suffer from a number of symptoms such as hyposmia, sleep disturbances, depression, hypotonia, and constipation that cannot be due only to the degeneration of nigral dopaminergic neurons. Intracytoplasmic (Lewy bodies) and intraneuritic (Lewy neurites) inclusions that contain alpha-synuclein are the pathological hallmarks of PD. alpha-Synuclein plays a critical role in the etiology of PD. In addition to the brain, Lewy body and neurite pathologies appear in the peripheral autonomic nervous system early in the disease process. The systemic degenerations that occur in PD start in peripheral neuronal systems and progress centripetally and then caudal to rostral within the CNS. Our hypothesis is that pituitary adenylate cyclase-activating polypeptide (PACAP) administered after the onset of motor dysfunction will halt or slow the progression of PD. PACAP is a pleiotropic peptide with both potent anti-inflammatory and potent cytoprotective properties. PACAP significantly reduces the degeneration of nigral dopaminergic neurons caused by either 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) or 6- hydroxydopamine. However, both of these neurotoxins cause PD-like symptoms in hours while PD usually takes decades for symptoms to develop. It is exceedingly unlikely that the nervous system responds in the same way to extremely rapid and slowly progressive diseases. The predictive value of these acute models for a progressive chronic disease is questionable. Therefore, there is a huge unmet need for an animal model that better mimics the human conditions. The alpha-synuclein over-expressing transgenic (A53T) mice seem to meet these expectations. The first autonomic dysfunctions in the urinary and intestinal tracts appear at 4 month of age in these mice while the motor dysfunction appears at 10-11 months of age. The slow progression and the organ-selective appearance of the symptoms in the A53T mice are similar to those in humans with PD. Although PACAP has remarkable cytoprotective and anti-inflammatory properties, its half-life in the bloodstream is short and, therefore, its use for chronic diseases is not practical. Therefore, we will evaluate our proprietary, metabolically stable PACAP analogs for neuroprotection in the A53T mouse model of PD. Specific Aim 1: Provide evidence that PACAP administered after the onset of motor dysfunction will halt or slow the progression of PD-like symptoms. Specific Aim 2: Provide evidence that the metabolically stable and receptor subtype-specific PACAP analogs administered as described in Specific Aim 1 will also halt or slow the progression of PD-like symptoms. Motor dysfunction (cylinder test, paw-print analysis, and grip strength) will be tested biweekly before and during treatment, and then the brains of the mice will be processed for PD pathology (alpha-synuclein, Lewy bodies, tyrosine hydroxylase, and apoptotic and inflammatory markers).