Mechanism Regulating ActivityDependent Neural Plasticity

Child Health And Human Development

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Abstract

A central question in neurobiology is to understand how nature and nurture contribute to remodeling of the nervous system during development, and in response to experience. The general goal of our laboratory is to investigate molecular mechanisms that regulate neuronal and muscle plasticity during development. A. NEUREGULIN REGULATES NEURONAL PLASTICITY: POSSIBLE RELEVANCE TO SCHIZOPHRENIA. We are studying how Neuregulins (NRG 1-3), a family of growth/differentiation factors, regulate synaptic transmission at interneuronal synapses. These factors and their receptors (ErbB tyrosine kinases) are important for regulating behavior in rodents. Numerous genetic studies recently implicated single nucleotide polymorphisms in the Neuregulin gene with schizophrenia in different populations. Initially, we found that the co-activation of ErbB receptors by NRG-1 and of NMDA receptors by glutamate is necessary to modify the expression of an NMDA receptor subunit gene, suggesting a cross-talk between these two types of signaling pathways (Ozaki et al. [1997] Nature 390:691-694). The subsequent demonstration that ErbB4 and NMDA receptors co-localize at glutamatergic synapses with PSD-95, a PDZ protein that couples postsynaptic receptors to signaling complexes, lead us to hypothesize that the NRG/ErbB signaling pathway may acutely modify synaptic properties (Garcia et al. [2000], PNAS, 97:3596-3601). We recently found that NRG-1 alters synaptic transmission. The implications of these findings for basic and clinical science may be extremely important in light of recent evidence that associates single nucleotide polymorphisms in the NRG-1 gene with higher risk for schizophrenia in seven different cohorts of patients from Finland, Scotland and China. Mutant mice with decreased levels of NRG-1 and ErbB receptors develop normally but have a reduction in NMDA receptors, and manifest ?schizophrenia-like? behaviors that are reversed with clozapine, a pharmacological agent used to treat schizophrenia. Future studies on the NRG/ErbB signaling pathway should contribute to understanding distinct mechanisms that regulate synaptic plasticity and that may be associated with neurological disorders. To understand how NRGs contribute to distinct aspects of neural development and function, we initially characterized their regional and subcellular expression patterns in developing brain. We found that NRG-1 expression is highest at birth, while NRG-2 mRNA levels increase with development; expression of both genes is restricted to distinct brain regions. In contrast, NRG-3 transcripts are abundant in most brain regions throughout development. NRG-2 antibodies were generated to analyze the processing, expression and subcellular distribution of this factor in central neurons. Despite the structural similarities between NRG-1 and NRG-2, we found that both factors are targeted to distinct subcellular compartments. NRG-2 accumulates in proximal primary dendrites of hippocampal neurons in culture and in vivo, while it is not detectable in axons or synaptic terminals. In contrast, NRG-1 is highly expressed in axons of dissociated hippocampal neurons, as well as in somas and dendrites. The distinct temporal, regional and subcellular expression of NRGs suggests their unique and non-redundant roles in neural function. B. DEVELOPMENTAL AND ACTIVITY-DEPENDENT REGULATION OF MUSCLE TYPES. The developmental and neuronal regulation of skeletal muscles provides an excellent model to study how different patterns of electrical impulses differentially regulate the properties of slow- and fast-twitch muscles. This type of plasticity is necessary for the adaptation of muscles to exercise, and underlies the changes in muscle properties that result from disease. Our long-term objective is to identify the signaling pathways that regulate the properties of slow- and fast-twitch muscles in response to activity. The troponin I slow (TnIs) and fast (TnIf) genes serve as our experimental model because expression of both genes is muscle-type-specific and regulated by distinct patterns of electrical impulses that mimic motor neuron activity. We found that the transcription factor GTF3 (General Transcription Factor 3), which binds to the SURE enhancer of the TnIs gene, is highly expressed in most tissues during early fetal development when muscle types emerge. This factor has 6 helix-loop-helix (HLH) motifs. Using SELIX, a method that selects specific DNA binding sites from random pools of sequences, indicate that several of the HLH motifs exhibited different preferences for DNA sequence. We found that HLH motif 4 has the highest avidity for DNA and is necessary for binding to the TnI SURE. A leucine zipper domain located at the N-terminus promotes GTF3 homodimerization but not heterodimerization with GTF2i, a protein closely related to GTF3. We speculate that the other HLH motifs may interact with a series of other transcription factors. Proteomic approaches are being utilized to identify proteins that may form larger transcriptional complexes with GTF3. The genes encoding GTF3 and GTF2i are lost in a ~2.0 Mb micro-deletion of chromosome 7q11.2 in individuals with Williams Syndrome (WS). Persons with WS have distinctive physical, cognitive and behavior abnormalities that include impaired spatial cognitive skills and myopathies. Our studies using ectopically transfected GTF3 constructs in adult muscles and GTF3 knock-out mice support a possible role for this factor in regulating muscle contractile properties. The observation that GTF3 and GTF2i are highly expressed in developing musculature and neurons, raises the possibility that reduction of these nuclear factors during embryogenesis may affect the expression of target genes later in development. Firing patterns typical of slow and fast motor units activate genes for slow and fast isoforms of contractile proteins, respectively. The mechanisms responsible for sensing and decoding distinct patterns of action potentials, and converting them into specific changes in gene expression, remain unknown. We have used a combination of in vivo muscle transfection, live imaging and fluorescence quantification to investigate the transcriptional control of the TnIs and TnIf genes in muscles stimulated with activity patterns that mimic either slow or fast motor neurons. Transcription was measured in adult muscles by following the fluorescence of the green fluorescent protein (GFP) expressed under the control of the TnIs and TnIf regulatory sequences. We found that transcription from the TnIs and TnIf enhancers was increased only when matched with its corresponding slow or fast pattern, respectively. Removal of nerve-evoked activity by denervation, or stimulation with a mismatching pattern, reduced transcriptional activity of both enhancers. These results indicate that the TnI slow and fast enhancers we have isolated can sense, and respond to, distinct patterns of neuronal activity. Future experiments will focus on identifying signal transduction pathways and transcription factors that mediate these responses.