Bruce Trapp, PhD - US grants

Alzheimer's disease (AD), the most prevalent cause of dementia, affects over 4.0 million individuals in the USA alone. Characteristic and diagnostic features of AD brains include degeneration of neurons, the accumulation of amyloid in neuritic (senile) plaques and perivascular regions, and the presence of neurofibrillary tangles. A major component of senile plaques is a 4 kD protein referred to as beta/A4. Beta/A4 is derived from one or more forms of a larger amyloid precursor protein (APP) which is constitutively expressed in brain. Although beta/A4 deposition is an early event in the pathogenesis of AD, the role that beta/A4 has in the neurodegenerative process is unknown, as are the normal function of APP and the mechanism by which beta/A4 is derived from APP. The long-term goals of this proposal are to determine the function of APP in normal brain and the mechanisms by which beta/A4 is derived from APP. Elucidating the cellular and subcellular distribution of APP and beta/A4 in normal brain and AD brain are essential for obtaining these goals. In vitro studies indicate that the APP extracellular domain is cleaved shortly after APP is inserted into plasma membranes. Specific Aim 1 of this proposal will test the hypothesis that APP is abundantly expressed in fetal mammalian brain, where it forms a matrix for the histotypic organization of neurons and neurites. Extracellular and cytoplasmic APP epitopes will be co-localized in tissue sections of fetal brain. Preliminary studies detect abundant APP in fetal brain and support this hypothesis. Specific Aim 2 will localize APP in light and electron micrographs of mammalian brain during postnatal development and ageing. It is predicted that little APP is expressed in normal postnatal brain. As ageing occurs, however, APP expression is upregulated to provide a matrix for neurite extension and neuropil rearrangement that accompanies neuronal death or atrophy. Studies in Specific Aim 3 will correlate the distribution of APP epitopes and beta/A4 in the CNS of individuals with Alzheimer's disease at the light and electron microscopic level. These studies will help elucidate the source of beta/A4 and the mechanisms involved in APP processing in AD brains. Collectively, the studies outlined in this proposal will provide valuable insights into the potential function of APP and the pathogenesis of amyloid formation.

Based on location, morphology, and molecular phenotype, we have identified three distinct stages of oligodendrocyte lineage in developing rodent brain: oligodendrocyte progenitors, premyelinating oligodendrocytes, and myelinating oligodendrocytes. The objective of this proposal is to obtain a better understanding of cellular and molecular events that regulate oligodendrocyte differentiation (Specific Aim 1) and CNS myelination (Specific Aim 2). Specific Aim 1A will determine the appearance and distribution of oligodendrocyte progenitors, premyelinating oligodendrocytes, and myelinating oligodendrocyte in the mouse optic nerve. These data will serve as a baseline for those in Specific Aim 1C which investigate how axonal transection ad ablation of action potentials affect oligodendrocyte lineage in optic nerve. Specific Aim 1B will phenotype optic nerve cells undergoing programmed cell death and determine the average life span of a premyelinating oligodendrocyte. Studies in Specific Aim 1D will determine how overexpression of the most studied oligodendrocyte trophic factor, PDGF, affects oligodendrocyte lineage during normal development and after optic nerve transection or ablation of optic nerve action potentials. The second part of the proposal will characterize changes in myelin protein expression and microtubule organization as oligodendrocytes mature from a premyelinating (nonpolarized) to a myelinating (polarized) cell. Studies in Specific Aim 2B will identify regions (MAG and MOG) polypeptides that are responsible for their delivery to myelin and nonmyelin surface membranes. The studies outlined in the proposal will provide new and valuable insights into cellular and molecular mechanisms of oligodendrocyte differentiation and myelination that will prove valuable in understanding and treating diseases of myelin.

DESCRIPTION (provided by applicant): Myelin surrounds many of the axons in the central and peripheral nervous systems where it facilitates the rapid conduction of nerve impulses and provides an extrinsic trophic effect that promotes axonal maturation and survival. Dysmyelination and demyelination are major causes of neurological disability in humans and can be fatal. Historically, neurological deficits in these primary myelin diseases were thought to result from myelin pathology. However, recent studies have identified axonal degeneration in a number of primary myelin diseases. The most common causes of genetic myelin disease in humans are gene duplications that alter the dosage of myelin proteins. Much of what is known about the cellular and molecular aspects of normal myelination and the pathogenesis of inherited myelin diseases has been obtained from studies of rodents in which myelin protein genes are mutated, deleted or overexpressed. We have developed transgenic mouse models of PNS and CNS dysmyelination by overexpressing P0 protein, the major structural protein of PNS myelin in Schwann cells, and by expressing high levels of P0 protein in myelinating oligodendrocytes. The overall goal of this application is to understand how P0 overexpression causes myelin and axonal pathology. Schwann cells that overexpress P0 protein ensheath but fail to myelinate axons because they mistarget P0 to non-myelin surface membranes. Studies in Specific Aim 1 will investigate mechanisms responsible for P0 and MAG targeting in MDCK cells in vitro. Specific Aim 2 will investigate how dysmyelination in P0 overexpressing mice causes alteration in ion channel distribution in PNS axons and a distal axonopathy that consists of axonal withdrawal from the neuromuscular junction and subsequent axonal sprouting and neuromuscular junction reinnervation. We have also established that P0 expression by oligodendrocytes results in CNS dysmyelination and axonal degeneration. Specific Aim 3 will investigate molecular mechanisms responsible for these pathologies and determine if the phenotype is rescued by their breeding to PLP null mice. Collectively, these studies should provide novel information about the pathogenesis of dysmyelination, molecular mechanisms of normal myelination, and the mechanisms by which myelin-forming cells modulate the development and survival of axons.

Multiple sclerosis is an inflammatory demyelinating disease of the human central nervous system. The primary cause or causes of MS are unknown. The purpose of this proposal is to obtain a better understanding of the mechanisms of inflammation and myelin and oligodendrocyte destruction in MS lesions by analyzing tissue from MS patients. The underlying hypothesis of this proposal is that inflammation and breakdown of the blood-brain barrier activate parenchymal microglia cells. These activated microglia cells then attack and destroy myelin and oligodendrocyte. The proposal is divided into three Specific Aims. The first Specific Aim will collect, process and characterize tissue from multiple sclerosis, other neurological diseases, and control brains. MS lesions will be identified and the extent of demyelination, remyelination, microglia and astrocyte activation, regions of blood-brain barrier breakdown, and extent of inflammation will be determined. The second Specific Aim will investigate the role of activated microglia in myelin and oligodendrocyte destruction. We will determine the three-dimensional relationship between activated microglia and myelin internodes and oligodendrocytes in MS brains. Preliminary studies indicate that microglia attack and destroy normal appearing myelin and that a select population of VCAM-1-positive microglia/monocytes attack oligodendrocytes. An important question raised by these studies is the mechanism by which microglia become activated in MS brain. This questions will be addressed by studies in Specific Aim 3, which investigate mechanisms of leukocyte trafficking in MS lesions. Leukocyte extravasation into tissue occurs by a series of interactions between the Beta2 integrins on leukocytes and the Ig-like molecules on endothelial cells. We will determine if the Beta2 integrins (LFA-1, MAC-1, P150-95, (Beta2alphad) and their punitive receptors (ICAM-1, ICAM-2, ICAM-3, ICAM-4, PECAM, VCAM-1) are induced or upregulated in MS lesions. Studies in Specific Aim 3 will also determine the distribution of the Beta1 integrin, VLA-4 and its receptor, the CS1 form of fibronectin. VLA-4 is expressed by leukocytes and is thought to play an essential role in their entry into active demyelinating lesions in MS brain. VLA-4 is a current therapeutic target in MS patients. We anticipate that these studies will identify adhesion molecules that are induced or upregulated in MS lesions. If it can be demonstrated that these molecules play an essential role in leukocyte trafficking, they become attractive targets for therapeutic intervention in MS.

neuroprotectants; axon; multiple sclerosis; histopathology; stem cells; myelination; experimental allergic encephalomyelitis; aspartate; neural degeneration; spinal cord injury; oligodendroglia; nervous system regeneration; human tissue; postmortem; laboratory rat; laboratory mouse;

Axonal degeneration is the major cause irreversible neurological disability patients multiple of in with sclerosis (MS). Previous examination of postmortem MS brains and spinal cords has identified three settings of axonal and neuronal degeneration. 1) Axons are transected during inflammatory demyelination. 2) Demyelination of the cerebral cortex transects axons and dendrites and causes neuronal apoptosis. 3) Chronically demyelinated axons degenerate. The overall goal of this project is to extend our previous studies by gaining insight, into the molecular mechanisms responsible for degeneration of chronically demyelinated axons. Such knowledge is essential for the development of neuroprotective therapeutics that will stop or delay the relentless progression of neurological disability in MS patients. Our studies are based upon the hypothesis that alterations in the molecular composition of the axolemma and reduced mitochrondrial function initiate a vicious cycle of axonal changes that cannot be controlled by the neuron or chronically demyelinated axon. Reduced energy inhibits axonal transport and mitochrondrial renewal which further decreases energy production. The increased energy demands of nerve conduction through demyelinated axons cannot be met resulting in a series of ionic imbalances that increases axonal calcium and destroys the axon. Our studies are divided into two Specific Aims. The first will continue to examine postmortem MS brains and spinal cords by characterizing the composition and distribution of proteins in the chemically demyelinated axon, determining the distribution and functional status of axonal mitochrondria and by characterizing axonal organelles that reflect or contribute to altered axon transport and to axonal degeneration. The second aim is designed to directly test various aspects of our hypothesis in an animal model of chronic axonal degeneration. Collectively, these studies should identify therapeutic targets that could stop or delay neurodegeneration in MS patients.

DESCRIPTION (provided by applicant): Multiple sclerosis (MS) is an inflammatory demyelinating disease of the CNS and the major cause of non-traumatic neurological disability in young adults in North America and Europe. The best described and most extensive repair that occurs in the adult human brain is the production of new oligodendrocytes that remyelinate axons in MS lesions. Although this repair can be extensive, most lesions in chronic MS patients are not remyelinated. Remyelination requires the production of new oligodendrocytes from oligodendrocyte progenitor cells (OPC) that are present in adult brain, but dramatically reduced in chronically demyelinated MS lesions. Enhancement of endogenous OPC production is a viable therapeutic target in MS patients. The major goal of this competitive renewal is to obtain a better understanding of how multipotent CNS stem cells located in the subventricular zone (SVZ) of the telencephalon produce OPCs in response to demyelination. This renewal focuses on a SVZ cell, the betaT4 cell (identified by betaT4 tubulin antibodies) which is 1) abundant in developing human SVZ, 2) detected at low densities in mature brain, 3) increased in MS brains, and 4) displays many of the characteristics of a multipotential CNS stem cell in vitro. We hypothesize that betaT4 cells are a major source of OPCs during late fetal and early postnatal telencephalon development, remain at low levels in adult brain, and can produce new OPCs in response to demyelination. Our studies are divided into two Aims. The first will investigate the fate of betaT4 SVZ cells during rodent brain development and in response to demyelination. These studies will 1) directly follow progeny of betaT4 cells in vivo using Cre/lox P technology, 2) further characterize molecules expressed by betaT4 cells using a genomic approach, and 3) investigate molecules that affect betaT4 cell development in neurospheres assays. The second Aim will continue to characterize betaT4 cells in MS lesions and developing human brains. Studies in humans are essential for accomplishing our long-term goals of understanding the role of betaT4 cells in repair of the adult brain. Collectively, these studies should identify therapeutic targets that enhance remyelination and reduce the progression of neurological disability in MS patients.

DESCRIPTION (provided by applicant): Multiple Sclerosis (MS) is an inflammatory demyelinating and neurodegenerative disease of the human central nervous system. MS affects over 2.0 million individuals worldwide and is the major cause of non-traumatic neurological disability in young adults in the United States. The classical view of MS as a myelin and white matter disease has been extended to include axonal transection, cortical demyelination and neuronal degeneration. While the clinical impact of gray matter pathology in MS brains is unknown, greater than 50% of MS patients are cognitively impaired. There is minimal or no correlation between physical disability and cognitive dysfunction in MS patients. Although focal hippocampal pathology causes cognitive dysfunction in several CNS diseases, the hippocampus has received little attention in MS research. The overall goal of the proposal is to determine if hippocampal changes contribute to episodic memory loss and cognitive decline in MS patients. Our studies are based upon the hypothesis that hippocampal demyelination reduces synaptic plasticity and causes episodic memory dysfunction. Preliminary studies identify significant hippocampal demyelination and alterations in hippocampal gene transcripts that facilitate memory and learning. Our studies are translational, MS patient based and divided into three specific aims. The first aim will determine if memory impaired MS patients have characteristic hippocampal pathology as determined by MRI. Preliminary studies suggest that diffusion tensor imaging (DTI) measures of the fornix will provide reliable and non-invasive surrogate markers for the episodic memory impairment in MS patients. The second aim will characterize hippocampal demyelination and neuronal pathology in MS hippocampi and establish pathological correlates of MRI alterations with hippocampus and fornix obtained by rapid autopsy of MS patients. The third aim will define molecular mechanisms of hippocampal dysfunction in MS patients. Preliminary studies support the hypothesis that hippocampal demyelination decreases delivery of synaptic vesicles to presynaptic terminals. This will reduce the glutamate and Ca entry into post synaptic terminals and decrease second messenger signally that facilitates hippocampal memory function in MS patients. Collectively, this proposal presents an integrated multi-disciplinary approach designed to elucidate causes of memory dysfunction in MS patients. PUBLIC HEALTH RELEVANCE: Multiple sclerosis (MS) afflicts over 2 million individuals worldwide and is a major cause of neurological disability in the USA. Most MS research investigates the cause of physical disability in MS patients. Despite the fact that over half of MS patients have reduced memory function, very little research has been performed on the cause of memory loss in MS patients. This is surprising as memory loss is a major contributor to unemployment of MS patients. Research in this proposal will determine the causes of reduced memory function in MS patients. We will study MS patients with reduced memory, develop brain imaging techniques to identify MS patients at risk for memory problems and identify ways to treat MS patients with memory loss.

The purpose of this project is to characterize cortical remyelination in postmortem multiple sclerosis (MS) brains. In contrast to white matter lesions, our preliminary studies establish that new oligodendrocyte production and active remyelination are prominent features of many chronic cortical MS lesions. Two major patterns of cortical demyelination have been described: leukocortical lesions (Type I) and subpial bands of demyelination which can extend over several gyri (Type III). Two specific aims will compare and contrast successful and failed remyelination in chronic Type I and III cortical lesions. Aim I will determine if remyelination is more prevalent in Type I or III lesions, and if there is a difference in repair of grey and white matter regions of Type I lesions. Remyelination requires the production of new oligodendrocytes. The second goal of Aim 1 is to quantify and characterize pre-myelinating and remyelinating oligodendrocytes in type I and III cortical lesions. We will determine the relationship between oligodendrocyte processes and remyelinating internodes and establish whether functional nodes of Ranvier are formed during cortical remyelination. Specific Aim 2 will perform a similar characterization of oligodendrocyte progenitor cell (OPC) density and phenotype in cortical lesions. OPCs are the only source of new oligodendrocytes, and their density and phenotype can regulate the success of myelin repair. Preliminary studies indicate that OPC densities are not altered in cortical lesions. Studies will investigate whether OPC molecular phenotype, OPC and pre-myelinating oligodendrocyte programmed cell death, or OPC mitosis varies in cortical lesions. We will also investigate whether inhibitors of oligodendrogenesis or myelination modulate myelin repair in MS brains. While anti-inflammatory therapies delay the progression of MS, they do not stop the disease process. The future of MS therapeutics lies in the identification of additional therapeutic targets and combinatorial therapies. By characterizing successful and failed myelin repair in MS brains, the studies confined in this proposal should identify novel targets that will enhance repair of the MS brain and quality of life of MS patients.

DESCRIPTION (provided by applicant): Multiple sclerosis (MS) remains the leading cause of neurological nontraumatic disability for young adults in North America and outcomes of current therapy are often unsatisfactory. This P50 competing renewal application directly extends and amplifies our research during the previous two cycles of support. We focus here on gray matter pathology, extending our previous work addressing interlinked hypotheses regarding the destructive inflammatory and neurodegenerative processes in MS patients. The P50 encompasses several unique resources including a rapid-autopsy program which has yielded insights into MRI-pathology correlations; a longitudinal MRI cohort which has illuminated the mechanisms underlying brain atrophy in MS; and newly-recruited, a series of MS biopsy cases which can be studied to understand cortical pathology early in MS. Core A: Tissue acquisition, imaging, biostatistics. administration (R Ransohoff) will establish, maintain and distribute a unique resource of MS autopsy tissue with dedicated postmortem imaging, as well as coordinate database management and provide administrative and biostatistical support for all projects. Project 1: Cortical demyelination and leukocyte trafficking early in MS (R Ransohoff; C Lucchinetti) will address the hypothesis that meningeal inflammation and cortical demyelination early in the MS process are implicated in facilitating white matter demyelination. This project will also address mechanisms of selective leukocyte trafficking to meninges and cortex. Project 2: Cellular and molecular mechanisms of cortical remyelination in MS patients (B Trapp) will examine cortical demyelination and remyelination in MS brains, and characterize both the cells mediating repair and the factors that inhibit repair. Project 3: Gray matter atrophy in multiple sclerosis (E Fisher) will use novel quantification techniques, developed as part of the P50 research, address in a longitudinal MRI cohort, the relationship between gray matter atrophy and white matter lesions as well as normal-appearing white matter changes. This project will also define relationships between gray matter atrophy and neuropsychological changes. Project 4: Gray matter atrophy in multiple sclerosis: Clinical implications (R Rudick) will extend the use of our new MRI analytic software to define the time course of gray matter atrophy, and evaluate the effects of immunomodulatory therapy, using files from completed clinical trials of a diversity of agents Extending our established research approaches to address gray matter pathology in MS will provide new insights into disease mechanisms and identify novel therapeutic targets.

DESCRIPTION (provided by applicant): Multiple sclerosis (MS) is an inflammatory demyelinating disease of the human central nervous system (CNS) and the major cause of non-traumatic neurological disability in young adults in the USA. The classical view of MS as a white matter disease has recently been extended to include gray matter demyelination. The clinical impact of gray matter demyelination may explain why white matter lesions detected by brain imaging poorly correlate with neurological disability in MS patients. Greater than 50% of MS patients are cognitively impaired and their cognitive dysfunction has a greater impact on quality of life than physical disability. Recent studies of postmortem MS brains have established that hippocampal demyelination disrupts the maintenance of excitatory synapses and activation of neuronal signaling cascades that modulate memory and learning. Hippocampal demyelination is a likely cause of cognitive decline in MS patients. To confirm and extend this hypothesis, we have successfully developed a mouse model of hippocampal demyelination and established that these mice have reduced memory/learning, reduced synaptic densities, reduced neuronal proteins that modulate memory/learning and reduced long term potentiation (LTP). Remyelination reversed these changes. The studies outlined in this proposal are designed to unravel the sequence of neuronal changes in demyelinated hippocampi that contribute to neuronal dysfunction and memory impairment. We utilize a multidisciplinary approach to correlate behavioral, electrophysiological, biochemical, morphological, and in vivo imaging changes in demyelinated hippocampi and will determine if remyelination reverses documented changes. Our studies are translational and should identify potential therapeutic targets that could reduce or delay cognitive decline in MS patients. Our mouse model should also provide a platform for proof of principle testing of therapies designed to improve cognition in MS patients.

DESCRIPTION (provided by applicant): Autism spectrum disorders (ASDs) are psychiatric disorders highlighted by social and communicative dysfunction. These complex behavioral changes are caused by altered synaptic functioning in select brain regions. Studies have previously focused on altered neuronal development and maturation. In addition to these neuronal changes, more recent studies have implicated astrocytes, the major glial cell in the brain, as major contributors to the pathogenesis of ASDs. Astrocytes are integral components of the tripartite synapse. In this model astrocyte processes surround and/or associate with pre- and post-synaptic components and regulate neurotransmitter homeostasis and recycling, provide basic substrates for neuronal metabolism, sequester Ca2+ ions and promote synaptogenesis and synaptic remodeling. Astrocytes have also been implicated in the pathogenesis of animal models of inherited human ASDs; these include mice lacking methyl-CpG-binding protein 2 (MeCP2) or the fragile X mental retardation 1 (FMR1) gene. While astrocyte diversity has been recognized for over a century, the molecular and cellular mechanisms underpinning this diversity in vivo and the consequences for psychiatric disorders remain poorly understood. The purpose of this proposal is to characterize astrocyte diversity in subpopulations of gray matter astrocytes in two mouse models of ASDs. Our experiments are facilitated by two technical advances that permit the identification and molecular characterization of astrocytes in animal models of ASDs. First, we have produced a novel transgenic mouse line (BT4-mEGFP) in which astrocyte surface membranes are fluorescently tagged from embryonic development onward. Breeding of these fluorescently tagged astrocytes into genetic models of ASDs will permit astrocyte isolation by fluorescent activated cell sorting and subsequent gene profiling. We will establish how these gene defects alter the development of astrocytes, the association of astrocytes with synapses and the expression of astrocyte proteins identified in the gene profiling studies. These data will be extended by determining the three dimensional associations between astrocytes and synapses using automated serial electron microscopy. Our studies are based upon the overall hypotheses that astrocytes play two key roles in ASDs. First, they have abnormal associations with synapses, which results in altered neurotransmitter homeostasis and abnormal neuronal electrical activity. Second they have altered mitochondrial functions, deficient ATP production and Ca2+ buffering, which reduces their ability to provide basic nutrition to neurons. These studies will elucidate previousl unidentified roles of astrocytes in ASDs and will provide a critical genetic and ultrastructural framework for the development of future therapeutic strategies that target astrocyte function in treating or preventing ASDs.

DESCRIPTION (provided by applicant): Autism spectrum disorders (ASDs) are psychiatric disorders highlighted by social and communicative dysfunction. These complex behavioral changes are caused by altered synaptic functioning in select brain regions. Studies have previously focused on altered neuronal development and maturation. In addition to these neuronal changes, more recent studies have implicated astrocytes, the major glial cell in the brain, as major contributors to the pathogenesis of ASDs. Astrocytes are integral components of the tripartite synapse. In this model astrocyte processes surround and/or associate with pre- and post-synaptic components and regulate neurotransmitter homeostasis and recycling, provide basic substrates for neuronal metabolism, sequester Ca2+ ions and promote synaptogenesis and synaptic remodeling. Astrocytes have also been implicated in the pathogenesis of animal models of inherited human ASDs; these include mice lacking methyl-CpG-binding protein 2 (MeCP2) or the fragile X mental retardation 1 (FMR1) gene. While astrocyte diversity has been recognized for over a century, the molecular and cellular mechanisms underpinning this diversity in vivo and the consequences for psychiatric disorders remain poorly understood. The purpose of this proposal is to characterize astrocyte diversity in subpopulations of gray matter astrocytes in two mouse models of ASDs. Our experiments are facilitated by two technical advances that permit the identification and molecular characterization of astrocytes in animal models of ASDs. First, we have produced a novel transgenic mouse line (BT4-mEGFP) in which astrocyte surface membranes are fluorescently tagged from embryonic development onward. Breeding of these fluorescently tagged astrocytes into genetic models of ASDs will permit astrocyte isolation by fluorescent activated cell sorting and subsequent gene profiling. We will establish how these gene defects alter the development of astrocytes, the association of astrocytes with synapses and the expression of astrocyte proteins identified in the gene profiling studies. These data will be extended by determining the three dimensional associations between astrocytes and synapses using automated serial electron microscopy. Our studies are based upon the overall hypotheses that astrocytes play two key roles in ASDs. First, they have abnormal associations with synapses, which results in altered neurotransmitter homeostasis and abnormal neuronal electrical activity. Second they have altered mitochondrial functions, deficient ATP production and Ca2+ buffering, which reduces their ability to provide basic nutrition to neurons. These studies will elucidate previousl unidentified roles of astrocytes in ASDs and will provide a critical genetic and ultrastructural framework for the development of future therapeutic strategies that target astrocyte function in treating or preventing ASDs.

? DESCRIPTION (provided by applicant): Multiple sclerosis (MS) is an inflammatory-mediated demyelinating disease of the central nervous system (CNS). While historically considered to be a disease of white matter (WM), recent studies have estimated that cortical demyelination exceeds WM demyelination in many MS patients. While cortical demyelination is a major aspect of MS pathogenesis, little is known about the dynamics and mechanisms of cortical myelin loss. This is especially true for subpial lesions, which are the most abundant type of cortical demyelination. Subpial lesions fail to show many of the pathological hallmarks of WM lesions, with little infiltration of peripheral immune cells or breakdown of the blood- brain barrier. Data also support the concept that cortical atrophy/demyelination correlates more strongly with clinical and cognitive disability than WM atrophy/demyelination. Studies outlined in this proposal will investigate how subpial demyelination occurs in MS and investigate the influence of brain WM demyelination on WM MRI alterations and brain atrophy. Specific Aim 1 will test the hypotheses that a dying-back oligodendrogliopathy occurs in upper layers of the cerebral cortex with age and activates microglia, which subsequently remove defective myelin. Loss of oligodendrocytes and myelin induces new oligodendrocyte production and remyelination. In MS brains, this oligodendrogliopathy is increased and an imbalance between demyelination and remyelination results in subpial lesions. Studies in Specific Aim 1 will also establish the turnove of oligodendrocyte lineage cells by measuring the level of `nuclear bomb' 14C in oligodendrocyte lineage cells. Specific Aim 2 will establish the molecular phenotype of microglia isolated from MS cortex and determine whether microglia cells are removing defective subpial myelin in MS brains. Since current anti-inflammatory therapies may not stop subpial demyelination, these studies are highly significant because they may identify novel therapeutic targets that will reduce or eliminate subpial demyelination. Specific Aim 3 investigates a cohort of 18 postmortem MS cases that have significant brain WM MRI lesion loads, but virtually no brain WM demyelination. Preliminary studies detected significant spinal cord and subpial cortical demyelination and provided pathological confirmation of MS. We will ask whether brain WM demyelination is essential for brain WM MRI abnormalities, brain WM atrophy, and cortical atrophy and identify other pathological changes that may contribute to these surrogate markers of MS disease progression. These studies have the potential to refine dogmas of MS pathogenesis.

ABSTRACT Acquired and inherited diseases of myelin are the major cause of non-traumatic neurological disability in young adults in the USA. Studies have also described myelin pathology in brains from individuals with amyotrophic lateral sclerosis (ALS) and Alzheimer's disease (AD), and myelin protein gene alleles are risk factors for schizophrenia, depression, and autism. In addition to its insulating properties, therefore, myelin has multiple effects on neuronal function. It is now accepted that axonal and neuronal degeneration cause permanent neurological disability in individuals with primary myelin disease. My laboratory played a significant role in identifying axonal and neuronal degeneration in brains from individuals with multiple sclerosis (MS), an inflammatory demyelinating disease of the human central nervous system (CNS). We leveraged these data to develop animal models that recapitulate mechanistic aspects of myelin-induced axonal and neuronal degeneration. The purpose of the present proposal is to consolidate three NINDS R01s that investigate mechanisms of myelin-induced neurodegeneration. We will address three key questions. 1) How does myelin provide trophic support to axons? We propose that transfer of ATP substrates is the major mechanism by which myelin provides trophic support to axons. Disruption of this metabolic coupling in inherited myelin diseases induces mitochondrial pathology/degeneration in paranodal axoplasm, which causes axonal degeneration. 2) How does demyelination affect neurons and their synaptic connections? We propose that demyelination alters neuronal gene expression, modulates dendritic structure, and reduces neuronal viability; we further propose that remyelination will reverse these changes. 3) How does subpial cortical demyelination occur? We propose that subpial demyelination occurs by mechanisms that differ from immune-mediated mechanisms that demyelinate white matter and that novel therapies are needed to prevent subpial demyelination. The biggest challenge facing the myelin research community is the development of neuroprotective therapies. R35 funding will consolidate our efforts to identify new therapeutic targets that cause axonal and neuronal degeneration in myelin diseases. This is essential for the development of neuroprotective therapies that delay and possibly reverse permanent neurological disability in individuals with myelin disease.