Articles by Michelle M. Giarmarco in JoVE
Preparing Fresh Retinal Slices from Adult Zebrafish for Ex Vivo Imaging Experiments Michelle M. Giarmarco1, Whitney M. Cleghorn1, James B. Hurley1,2, Susan E. Brockerhoff1,2 1Department of Biochemistry, University of Washington, 2Department of Ophthalmology, University of Washington Imaging retinal tissue can provide single-cell information that cannot be gathered from traditional biochemical methods. This protocol describes preparation of retinal slices from zebrafish for confocal imaging. Fluorescent genetically encoded sensors or indicator dyes allow visualization of numerous biological processes in distinct retinal cell types.
Other articles by Michelle M. Giarmarco on PubMed
Divalent Cation-mediated Polysaccharide Interactions with Zwitterionic Surfaces Biomaterials. | Pubmed ID: 22177617 One popular postulation in the design of a nonfouling surface is that a surface capable of resisting nonspecific protein adsorption should also resist bacterial adhesion and subsequent biofilm formation. Such a hypothesis, though valid in certain cases, oversimplifies complex biological systems, since they contain not only proteins but also other biomacromolecules, such as polysaccharides. This work aims to re-examine this postulation by testing the biofouling of polysaccharides onto protein-resisting zwitterionic surfaces in the presence of a multivalent cation. Our results show that Mg(2+) plays an important role in mediating alginate adsorption onto zwitterionic surfaces through ion-bridged interactions from surface plasmon resonance (SPR) experiments. Three zwitterionic polymers tested in this work have clearly different responses to changes in Mg(2+) concentration, indicating that such ion-bridged adsorption is strongly dependent on cation-zwitterionic polymer binding affinities and is dictated by the specific chemical structure of the polymer betaine side chain. This work underlines the necessity to go beyond current nonfouling criteria at the protein level and to take into account polysaccharides when it comes to complex environments.
Mitochondria Maintain Distinct Ca Pools in Cone Photoreceptors The Journal of Neuroscience : the Official Journal of the Society for Neuroscience. | Pubmed ID: 28115482 Ca ions have distinct roles in the outer segment, cell body, and synaptic terminal of photoreceptors. We tested the hypothesis that distinct Ca domains are maintained by Ca uptake into mitochondria. Serial block face scanning electron microscopy of zebrafish cones revealed that nearly 100 mitochondria cluster at the apical side of the inner segment, directly below the outer segment. The endoplasmic reticulum surrounds the basal and lateral surfaces of this cluster, but does not reach the apical surface or penetrate into the cluster. Using genetically encoded Ca sensors, we found that mitochondria take up Ca when it accumulates either in the cone cell body or outer segment. Blocking mitochondrial Ca uniporter activity compromises the ability of mitochondria to maintain distinct Ca domains. Together, our findings indicate that mitochondria can modulate subcellular functional specialization in photoreceptors. Ca homeostasis is essential for the survival and function of retinal photoreceptors. Separate pools of Ca regulate phototransduction in the outer segment, metabolism in the cell body, and neurotransmitter release at the synaptic terminal. We investigated the role of mitochondria in compartmentalization of Ca We found that mitochondria form a dense cluster that acts as a diffusion barrier between the outer segment and cell body. The cluster is surprisingly only partially surrounded by the endoplasmic reticulum, a key mediator of mitochondrial Ca uptake. Blocking the uptake of Ca by mitochondria causes redistribution of Ca throughout the cell. Our results show that mitochondrial Ca uptake in photoreceptors is complex and plays an essential role in normal function.
Biochemical Adaptations of the Retina and Retinal Pigment Epithelium Support a Metabolic Ecosystem in the Vertebrate Eye ELife. | Pubmed ID: 28901286 Here we report multiple lines of evidence for a comprehensive model of energy metabolism in the vertebrate eye. Metabolic flux, locations of key enzymes, and our finding that glucose enters mouse and zebrafish retinas mostly through photoreceptors support a conceptually new model for retinal metabolism. In this model, glucose from the choroidal blood passes through the retinal pigment epithelium to the retina where photoreceptors convert it to lactate. Photoreceptors then export the lactate as fuel for the retinal pigment epithelium and for neighboring Müller glial cells. We used human retinal epithelial cells to show that lactate can suppress consumption of glucose by the retinal pigment epithelium. Suppression of glucose consumption in the retinal pigment epithelium can increase the amount of glucose that reaches the retina. This framework for understanding metabolic relationships in the vertebrate retina provides new insights into the underlying causes of retinal disease and age-related vision loss.