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In JoVE (1)
Other Publications (8)
Articles by Karen Zito in JoVE
Preparation of Gene Gun Bullets and Biolistic Transfection of Neurons in Slice Culture
Georgia Woods, Karen Zito
Center for Neuroscience, University of California, Davis
We describe a method for preparing DNA coated gold bullets and demonstrate the use of such bullets to biolistically transfect neurons in cultured hippocampal slices.
Other articles by Karen Zito on PubMed
Neural Computation. Jun, 2002 | Pubmed ID: 12020447
The structure of neuronal dendrites and their spines underlie the connectivity of neural networks. Dendrites, spines, and their dynamics are shaped by genetic programs as well as sensory experience. Dendritic structures and dynamics may therefore be important predictors of the function of neural networks. Based on new imaging approaches and increases in the speed of computation, it has become possible to acquire large sets of high-resolution optical micrographs of neuron structure at length scales small enough to resolve spines. This advance in data acquisition has not been accompanied by comparable advances in data analysis techniques; the analysis of dendritic and spine morphology is still accomplished largely manually. In addition to being extremely time intensive, manual analysis also introduces systematic and hard-to-characterize biases. We present a geometric approach for automatically detecting and quantifying the three-dimensional structure of dendritic spines from stacks of image data acquired using laser scanning microscopy. We present results on the measurement of dendritic spine length, volume, density, and shape classification for both static and time-lapse images of dendrites of hippocampal pyramidal neurons. For spine length and density, the automated measurements in static images are compared with manual measurements. Comparisons are also made between automated and manual spine length measurements for a time-series data set. The algorithm performs well compared to a human analyzer, especially on time-series data. Automated analysis of dendritic spine morphology will enable objective analysis of large morphological data sets. The approaches presented here are generalizable to other aspects of neuronal morphology.
Neuron. Sep, 2002 | Pubmed ID: 12354392
Recent electron microscopic studies provide evidence that the adult cortex generates new synapses in response to sensory activity and that these structural changes can occur rapidly, within 24 hr of sensory stimulation. Together with progress imaging synapses in vivo, the stage appears set for advances in understanding the dynamics and mechanisms of experience-dependent synaptogenesis.
Neuron. Jan, 2003 | Pubmed ID: 12526764
The fine tuning of synaptic circuits often requires an activity-dependent phase in which appropriate connections are strengthened and inappropriate connections are eliminated. In this issue of Neuron, Walsh and Lichtman propose a novel "synaptic takeover" mechanism for synapse elimination at the vertebrate NMJ, where withdrawal of one axon is accompanied by expansion of a competing axon into the newly vacated territory.
Current Opinion in Neurobiology. Oct, 2003 | Pubmed ID: 14630220
The impact of microarray studies on neurobiology has been limited because, with the exception of a few outstanding papers, most reports provide little more than lists of genes, often leaving the reader at a loss to understand which and how many of the identified transcripts will be true positives with significant biological impact. However, some recent papers have offered considerable biological insight by providing independent in vivo confirmation of the roles of candidate genes, offering a glimpse of the potential power of microarrays in neurobiological research.
Neuron. Oct, 2004 | Pubmed ID: 15473970
We explored the relationship between regulation of the spine actin cytoskeleton, spine morphogenesis, and synapse formation by manipulating expression of the actin binding protein NrbI and its deletion mutants. In pyramidal neurons of cultured rat hippocampal slices, NrbI is concentrated in dendritic spines by binding to the actin cytoskeleton. Expression of one NrbI deletion mutant, containing the actin binding domain, dramatically increased the density and length of dendritic spines with synapses. This hyperspinogenesis was accompanied by enhanced actin polymerization and spine motility. Synaptic strengths were reduced to compensate for extra synapses, keeping total synaptic input per neuron constant. Our data support a model in which synapse formation is promoted by actin-powered motility.
Neuron. Jan, 2009 | Pubmed ID: 19186167
Spine growth and retraction with synapse formation and elimination plays an important role in shaping brain circuits during development and in the adult brain, yet the temporal relationship between spine morphogenesis and the formation of functional synapses remains poorly defined. We imaged hippocampal pyramidal neurons to identify spines of different ages. We then used two-photon glutamate uncaging, whole-cell recording, and Ca(2+) imaging to analyze the properties of nascent spines and their older neighbors. New spines expressed glutamate-sensitive currents that were indistinguishable from mature spines of comparable volumes. Some spines exhibited negligible AMPA receptor-mediated responses, but the occurrence of these "silent" spines was uncorrelated with spine age. In contrast, NMDA receptor-mediated Ca(2+) accumulations were significantly lower in new spines. New spines reconstructed using electron microscopy made synapses. Our data support a model in which outgrowth and enlargement of nascent spines is tightly coupled to formation and maturation of glutamatergic synapses.
The Journal of Neuroscience : the Official Journal of the Society for Neuroscience. Aug, 2011 | Pubmed ID: 21865455
Changes in neuronal structure are thought to underlie long-term behavioral modifications associated with learning and memory. In particular, considerable evidence implicates the destabilization and retraction of dendritic spines along with the loss of spine synapses as an important cellular mechanism for refining brain circuits, yet the molecular mechanisms regulating spine elimination remain ill-defined. The postsynaptic density protein, PSD-95, is highly enriched in dendritic spines and has been associated with spine stability. Because spines with low levels of PSD-95 are more dynamic, and the recruitment of PSD-95 to nascent spines has been associated with spine stabilization, we hypothesized that loss of PSD-95 enrichment would be a prerequisite for spine retraction. To test this hypothesis, we used dual-color time-lapse two-photon microscopy to monitor rat hippocampal pyramidal neurons cotransfected with PSD-95-GFP and DsRed-Express, and we analyzed the relationship between PSD-95-GFP enrichment and spine morphological changes. Consistent with our hypothesis, we found that the majority of spines that retracted were relatively unenriched for PSD-95-GFP. However, in the subset of PSD-95-GFP-enriched spines that retracted, spine shrinkage and loss of PSD-95-GFP were tightly coupled, suggesting that loss of PSD-95-GFP enrichment did not precede spine retraction. Moreover, we found that, in some instances, spine retraction resulted in a significant enrichment of PSD-95-GFP on the dendritic shaft. Our data support a model of spine retraction in which loss of PSD-95 enrichment is not required prior to the destabilization of spines.