Development and maintenance of synaptic integrity: The Colon-Ramos lab is interested in studying how synapses form. We use the nematode C. elegans as our model system, focusing specifically on the development of synapses between two neurons, the AIY and RIA interneurons, in the head of the animal. The AIY interneurons act as the presynaptic partners to the RIA interneurons, forming synapses at specific points within a portion of the AIY neurite we refer to as zone 2. Traditionally, synapse formation is thought to take place after neurite outgrowth has occurred, but we recently observed that Zone 2 is the first part of the AIY neurite to form. This observation raises the possibility that synapse formation between AIY and RIA takes place concurrently with neurite outgrowth. We are currently testing this hypothesis by visualizing synapse formation in AIY during embryogenesis.
In addition to observing how synapses form in the developing embryo, we are also interested in understanding the pathways and molecules involved in directing where synapses form between AIY and RIA. Previous research has demonstrated that the UNC-6/Netrin pathway is important for directing synapse formation in AIY and axon guidance in RIA. UNC-6 is produced by a glial cell which ensheathes one side of Zone 2, and UNC-6 signals through its receptor UNC-40 to promote synapse localization in AIY. Interestingly, RIA (the AIY postsynaptic partner) is located on the opposite side of the AIY neurite as the glial cell, suggesting that there might be a secondary mechanism directing synapse formation between AIY and RIA besides glia-derived UNC-6/Netrin. We are currently examining whether other molecules known to signal through the UNC-40 receptor may play a role in directing AIY synapse formation.
New Microscopy approaches for imaging neuronal development: One of the exciting new frontiers in studying C. elegans neuronal development is the examination of neural circuit formation in living C. elegans embryos. For example, axon outgrowth in the AIY interneurons takes place between comma-stage and two-fold stages of embryogenesis. However, our ability to examine developmental events during embryogenesis is limited by a couple of factors, both those intrinsic to the worm, and technical limitations of current microscopy techniques. Visualization of the C. elegans embryo is limited by the fact that about halfway through embryogenesis, the embryo has developed to the point that it is capable of moving within its eggshell; this movement, called twitching, occurs quickly enough that it can interfere with microscopic imaging of the embryo. In addition, current microscopic techniques, such as confocal imaging, have the capability to damage specimens if the specimens undergo long-term imaging. These factors, when combined, can severely limit our ability to study neurodevelopmental events during embryogenesis in C. elegans.
Recently, development of new microscopy techniques raises the possibility of bypassing the difficulties in studying C. elegans embryos described above. iSPIM (inverted selective plane illumination microscopy) is a new microscopy technique which allows extremely high imaging rates (~30x per minutes) with very low associated photodamage. Basically, iSPIM works by rapidly scanning a thin “sheet” of light over the specimen to be imaged many times a minute. The rapidity of imaging and decreased photodamage of this technique allows a worm embryo to be imaged continuously from egg-laying to hatching, and greatly increase our ability to examine neurodevelopmental events during embryogenesis. We are currently using iSPIM to investigate developmental events occurring during embryogenesis.
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