Imaging Microtubule Dynamics in Cortical and Hippocampal Dendritic Spines

Erik W. Dent, Ph.D.

University of Wisconsin

Funded in June, 2008: $200000 for 3 years
LAY SUMMARY . ABSTRACT . BIOGRAPHY .

LAY SUMMARY

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Cellular Imaging may Help to Reveal how Structure Affects Function in Brain Plasticity

University of Wisconsin investigators will use cellular imaging in laboratory cultures of neurons to explore how regulation of certain neuronal structures may affect brain plasticity,” the development of new neural network connections in the brain.   

Brain cells (neurons) communicate by passing neurotransmitters from one cell to another.  When a neurotransmitter travels down one neuron’s axon, it is released into a minute space (called the “synaptic junction”) and binds to receptors on a neighboring neuron’s dendrite. Axons and dendrites branch out extensively, creating an estimated 10 trillion synaptic junctions in the human brain.  These synaptic junctions hold the potential for creating myriad neural networks. Dendrites have tiny protrusions, called dendritic spines. Spine shapes vary and change in response to activation. Spine shape directly affects the efficiency of neuronal synaptic communication. Large spines are associated with stronger synaptic connections. A stronger response by dendritic spines, termed “long-term potentiation,” is an experimental model of memory formation.  Understanding how spine shapes are regulated, according to the University of Wisconsin researchers, may provide fundamental insight into how memory-related brain plasticity occurs and how plasticity may be altered in developmental disorders and cognitive diseases, such as Alzheimer’s.

Their initial evidence suggests that regulation of spine shapes occurs through a direct route from the neuronalcell body to the synapse via “microtubules.” These are long, hollow tubes that transport material within neurons.  The researchers hypothesize that microtubules remodel dendritic spines in response to changes in neuronal activity.  They will test this hypothesis using total internal reflection fluorescence microscopy (TIRFM) in laboratory cultures of cortical and hippocampal neurons, while manipulating neuronal activity.  They also will use two-photon confocal microscopy in cortical and hippocampal brain slices to determine if microtubule invasion of dendritic spines occurs in an intact brain tissue preparation.

Significance: The results eventually may lead to a new understanding of how memory-related changes in brain plasticity occur and reveal targets for genetic and pharmacologic treatments for developmental and cognitive disorders.  

 

ABSTRACT

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Imaging Microtubule Dynamics in Cortical and Hippocampal Dendritic Spines

Excitatory cortical and hippocampal neurons communicate with other neurons through micron-sized protrusions on their dendrites termed spines.  Evidence suggests that the morphology of these spines directly affects the functional communication between neurons by changes in synaptic efficacy.  Enlarged spine heads correlate with an increased size of the post-synaptic density and the strength of the spine.  Long term potentiation, an experimental model of memory formation, can induce such changes in spines.  Conversely, simplification of spine morphology and loss of entire dendritic spines can occur after long term depression and in many neurological diseases.  Therefore, knowing how spine morphology is regulated may provide fundamental insight into synaptogenesis and synaptic plasticity.

To date there have been thousands of studies on dendritic spines of excitatory cortical and hippocampal neurons. Some of these studies have focused on the neuronal cytoskeleton, which is composed of actin filaments, microtubules and neurofilaments. Recent studies have shown that actin filament dynamics play a prominent role in the formation and maintenance of dendritic spines.  However, microtubules, which are numerous in dendrite shafts, are assumed to be stable and to not play a direct role in sculpting spine form or function.  Surprisingly, by imaging fluorescently-labeled microtubules we have discovered that they are very dynamic and capable of rapidly extending into and out of dendritic spines in mature cultured cortical and hippocampal neurons.  Preliminary evidence indicates that microtubule invasion into dendritic protrusions is associated with rapid morphological changes of spines.  These findings suggest that many of the components that are either transported on microtubules or are associated with their growing tips are capable of directly entering spines in a regulated fashion.

In this application we propose to: 1) Determine how microtubule invasion affects spine morphology, 2) investigate how neuronal activity affects microtubule dynamics in dendritic protrusions, 3) determine if microtubule dynamics occur in dendritic spines in situ.  This work will provide fundamental insights into synaptic function.  Furthermore, because dendritic spines are the sites that are affected in numerous psychiatric and neurological diseases, these studies hold promise for novel cytoskeletal-based therapies for synaptic dysfunction.

INVESTIGATOR BIOGRAPHIES

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Erik W. Dent, Ph.D.

Dr. Erik W. Dent is an Assistant Professor in the Department of Anatomy at the University of Wisconsin, Madison.  He received his Ph.D. in Neuroscience from the University of Wisconsin, Madison, with Dr. Katherine Kalil and conducted postdoctoral studies at the Massachusetts Institute of Technology in the Biology Department with Dr. Frank Gertler.  He began his present faculty position in September 2006.  His lab focuses on understanding the cellular basis of neuronal development.  Specifically, his lab is interested in understanding the cross-talk between the actin and microtubule cytoskeleton and the proteins involved in the coordination of these two important polymers.  Several developmental (i.e. mental retardation) and age-related  (i.e. Alzheimer’s disease) diseases  affect the neuronal cytoskeleton.  We specialize in high resolution live-cell fluorescent imaging (TIRF, confocal and widefield microscopy) of the cytoskeleton in developing and mature cortical and hippocampal neurons.  By imaging the neuronal cytoskeleton in time-lapse, we have discovered that both microtubules and actin filaments remain dynamic throughout the life of the neuron.  Therefore, we are uniquely positioned to make discoveries that could lead to development of effective strategies to ameliorate diseases affecting the neuronal cytoskeleton.