Rolf Karlstrom Ph.D.
Chair, Department of Biology
University of Massachusetts
PROJECT TITLE: Fate Mapping the Zebrafish Pituitary Gland
The vertebrate pituitary gland and lens originate as cranial placodes, being induced from a pre-placodal region at the anterior margin of the developing nervous system. We previously showed that graded. Hedgehog signaling from neural ectoderm is necessary and sufficient for the induction of pituitary fates, for suppressing lens fates, and for patterning the pituitary gland (Sbrogna et al, 2003, Guner et al, 2008, Devine et al, 2009). Our work suggested that the pituitary placode is patterned very early in development, with distinct regions of the adenohypophysis being established soon after induction of the pituitary placode at the anterior margin of the neural plate/keel. Here I propose to collaborate with Dr. Jonathan Gitlin at the MBL to investigate the morphogenetic movements that give rise to the pituitary gland and lens (Aims 1 and 2), and to use 4-dimensional celltracking methods to create a fate map of the anterior margin of the CNS, the region that gives rise to pituitary and lens tissue (Aim 3). These studies will contribute to our understanding of the early origin of the pituitary and lens placodes, and will provide insight into the molecular and cellular mechanisms that lead to a functional pituitary gland and lens. Given the conservation across vertebrate species, this work in zebrafish can inform similar studies in mammals and provide insight into the mechanisms underlying human birth defects that affect the pituitary and ventral forebrain.
Eduardo Rosa-Molinar Ph.D
University of Puerto Rico
PROJECT TITLE: Connexin composition in apposed gap junction hemiplaques at mixed synapses between identifiable dye-coupled spinal neurons revealed by super-resolution fluorescence imaging
The goal of this proposal is the first step toward developing a correlative technique integrating direct stochastic optical reconstruction microscopy (dSTORM), grid-mapped freeze-fracture-matched-double-replica-immunogold-labeling, combined with gallium focused ion beam low voltage field emission scanning electron microscopy. By combining these imaging modalities/techniques we will be able to unambiguously show the connexin composition in apposed gap junction hemiplaques at “mixed synapses” (conjoined electrical and chemical synaptic components) between identifiable dye-coupled spinal neurons.
Paul Selvin Ph.D.
University of Illinois
PROJECT TITLE: Small quantum dots for live super-resolution imaging of synaptic proteins
Synapses are sub-micron structures that mediate nerve-nerve interactions, underlie memory formation, and deteriorate in neurodegenerative diseases. Glutamate and other neurotransmitters are released into the synaptic cleft and bind to receptors embedded in the postsynaptic density (PSD) that mediate rapid synaptic responses. Changes in the number of postsynaptic receptors with repeated excitation are central to the synaptic plasticity changes that underlie memory formation. The details about what regulates these receptor levels are largely uncharacterized, but many mechanisms have been implicated including diffusion at the synapse, receptor recycling (endocytosis and exocytosis), and receptor delivery from the cell body (where they are synthesized and assembled) to a particular synapse. We propose to visualize the dynamics of postsynaptic receptors, by using and improving super-resolution fluorescence microscopy at the single-molecule level. We will do so by merging two new technologies that we partially or completely developed: small (<8 nm) and chemically stable fluorescent quantum dots (sQD) with very long photostability (> 1 hr), and three-dimensional imaging with ~10 nm resolution. We have preliminary results on AMPA receptors and various post-synaptic proteins (Homer1, PSD-95). NMDA receptors are also of interest. To understand their 3-dimensional configuration, as a function of LTP and LTD, is the primary focus. No other technique(s) can resolve dynamically changing structures such as synaptic receptors where the dimensions are typically much smaller than the diffraction limit.