previously been shown to be important in regulating lipoproteinmetabolism. Using a floxed allele ofmesd, we are examining the effectsof tissue-specific loss of MESD on plasma lipoprotein levels and arterialplaque formation. This work was supported by GM5396407 to BCH.

doi:10.1016/j.ydbio.2008.05.334

Program/Abstract # 314A zebrafish genetic model of spinal muscular atrophy andfunctional analysis of the Smn-binding protein, Gemin2Michelle L. McWhorter a, Kum-loong Boon b, Shu Xiao b, JessicaMullenberg c, Thomas Donn c, Cecilia Moens c, Christine E. Beattie b

a Biology Department, Wittenberg University, Springfield, OH, USAb Center for Molecular Neurobiology, Department of Neuroscience,The Ohio State University, Columbus, OH, USAcHoward Hughes Medical Institute, Division of Basic Science,

Fred Hutchinson Cancer Research Center, Seattle, WA, USA

Spinal muscular atrophy (SMA) is an autosomal recessive disordercharacterized by a loss of α-motoneurons in the spinal cord. SMA iscausedby low levels of theubiquitouslyexpressed SurvivalMotorNeuron(Smn) protein. Inprevious studies,we have shown thatmorpholino (MO)knockdown of smn in zebrafish embryos results in motor axon-specificoutgrowthdefects. To obtain a genetic zebrafishmodel of this disease,wehave identified 3 smn mutations by TILLING. smn−/− zebrafish have highmaternal Smnprotein; at 11dpf, however, protein levels are undetectableand the larvae die at 11–12 dpf. Synapse analysis reveals NMJ changesconsistent with denervation inmutants (andmorphants). To address thefunction of Smn, we knocked down Gemin2, a Smn-binding proteininvolved in snRNP assembly. gemin2 MO knockdown in the entireembryo showed overall abnormal development. However, when weknocked downGemin2 specifically inmotoneurons (either by iontophor-esis or blastula transplantation), their axons developed normally. Thesedata show that reduction of Gemin2, unlike Smn, does not directly causemotor axon outgrowth defects, which is consistent with an snRNP-independent role for Smn in motoneurons. Funding sources include:Families of SMA grants MCW2006 (MLM) and BOO2008 (KLB); NIHgrants RO1NS050414 (CEB) and P30-NS045758.

doi:10.1016/j.ydbio.2008.05.335

Program/Abstract # 315Discovery and characterization of novel synuclein genes inzebrafishZhihui Sun, Aaron D. GitlerDepartment of Cell and Developmental Biology, University ofPennsylvania School of Medicine, Philadelphia, PA, USA

The presynaptic protein, α-synuclein, appears to play a key role inthe pathogenesis of Parkinson's disease but the normal cellularfunctions of synuclein proteins remain a mystery. Understanding therole of α-synuclein during normal development will provide insightinto its later role in pathophysiology. The zebrafish is a powerfulvertebrate model system for studying many important developmentalprocesses. We hypothesized that the simpler nervous system of zeb-rafish might allow us to readily explore the functions of synucleins.We cloned three novel synuclein genes from zebrafish and have foundthat they are expressed strongly in the nervous system. We areperforming loss-of-function experiments using morpholino-modifiedantisense oligonucleotides to determine the effect of losing one, two,

or three synucleins. The phenotypes we observe will suggest potentialcellular functions for these zebrafish synucleins and will providehypotheses to test in higher systems, such as mouse and rat.

doi:10.1016/j.ydbio.2008.05.336

Program/Abstract # 316Seeking the biochemical basis of type III 3-methylglutaconicaciduria through zebrafish modelsWuhong Pei a, Isa Bernardini a, Christopher Wassif a, Forbes Porter a,Yair Anikster b, Marjan Huizing a, Benjamin Feldman a

a National Institute of Health, Bethesda, MD, USAb Sheba Medical Centre, Sackle Medical School, Tel Aviv University,Tel-Hashomer, Israel

Type III 3-methylglutaconic aciduria (MGA-III) is a rare disorderwithneuro-opthamalgicmanifestations and increased urinary excretion of 3-methylglutaric acid (3MGA). Two familial mutations have been found inthe OPA3 gene associated with MGA-III. We found that the zebrafishOpa3 orthologue is expressed ubiquitously during embryogenesis and isenriched in the brain from the pharyngula stage until at least 120 hpf.Antisense-based depletion of zebrafish Opa3 causes the signatureincrease in 3MGA, but also a more severe eye defect than seen inMGA-III patients. To explore whether Opa3 acts in the leucine catabolicpathway,wedelivered exogenous leucine toOpa3-deficientembryos.Asa comparison, leucine delivery to an MGA-I model deficient for theleucine catabolic pathway enzyme 3-methylglutaconyl-CoA hydratasecauses nomorphological defects, but leads to a sharp increase in leucine,and 3MGA. In contrast, leucine-treated Opa3-deficient embryos displaysevere brain dysmorphology but no accumulation of leucine or 3MGA.We also examined the effects on Opa3-deficient embryos of mevalo-nate-depletion via simvastatin treatment, and found that simvastatincauses additional braindefects inOpa3-deficient embryos.Wehave thusuncovered two classes of metabolic sensitivity that are specific to thebrain of our zebrafish MGA-III model, indicating that zebrafish Opa3interacts with both the leucine and mevalonate pathways.

doi:10.1016/j.ydbio.2008.05.337

Program/Abstract # 317Imaging of intestinal lipid absorption and processing in a livezebrafishJames Walters, Steven FarberDepartment of Embryology, Carnegie Institution of Washington, USA

The larval zebrafish (Danio rerio) is an ideal model of vertebrateintestinal physiology because of its rapid development and opticalclarity. This systemallows for thedirect observationoffluorescent lipids,fusion proteins, and reporter gene constructs within the developinganimal. Data fromavarietyof vertebrates indicates thatdietary lipids areinitially absorbed by intestinal enterocytes. Since energy homeostasisrelies on dietary lipids, how enterocytes process lipids profoundlyimpacts whole animal lipidmetabolism.While several studies implicatevarious subcellular compartments and proteins in the absorptiveprocess, many questions remain. The highly dynamic nature of theabsorptive process and the subcellular structures are difficult to image inmammals. An accurate model of intestinal physiology will reflect thecombined action of both organelles as well as many cell types. We havebegun the characterization of lipid uptake by zebrafish intestinalenterocytes. The ability to perform real-time in vivo lipid absorption

562 ABSTRACTS / Developmental Biology 319 (2008) 561–563

studieswill be essential for developing an understanding of dietary lipidprocessing and facilitate the identification of new therapeutic targets.We have developed an assay to observe lipid absorption at the singleenterocyte level using lipophilic dyes in zebrafish larvae that expressfluorescent fusion proteins. When larvae are fed a high fat diet,enterocytes accumulate lipid droplets that can be visualized in real-time in live animals. The intracellular fate of these newly absorbed lipidswas examined by co-localization of well-characterized fluorescentfusion proteins important for endocytosis and trafficking of lipids.

doi:10.1016/j.ydbio.2008.05.338

Program/Abstract # 318Valproic acid, an HDAC inhibitor, disrupts primitive hematopoiesisin Xenopus laevisRishita Shah, Peter KleinSchool of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA

Valproic acid (VPA) is a short-chain fatty acid that is widely usedto treat epilepsy and bipolar disorder. However, VPA treatment duringpregnancy is associated with neural tube defects, cardiac malforma-

tions, craniofacial abnormalities, and limb defects. The mechanism bywhich these developmental defects occur is poorly understood. Ourlab has shown that VPA directly inhibits histone deacetylases(HDACs). We are using Xenopus laevis as a model system to studythe consequences of VPA mediated HDAC inhibition in earlydevelopment. Administering a therapeutically relevant dose of VPA(1–2 mM) to gastrulating Xenopus embryos causes heart loopingabnormalities, pericardial edema, and a significant reduction incirculating red blood cells. However, overall development appearsessentially normal, and markers of dorsal and lateral mesoderm aregrossly normal, suggesting a relatively restricted effect of VPA underthis treatment regimen. We have found that VPA significantly reducesexpression of the hematopoietic marker Xenopus acute myeloidleukemia (Xaml/runx1) specifically in the ventral blood islands. Thereduction of Xaml expression can be recapitulated by another HDACinhibitor, Trichostatin A (TSA), but not by valpromide (VPM), astructural analog of VPA that does not inhibit HDACs, suggesting thatthe effects of VPA are mediated through HDAC inhibition. Our futurestudies will focus on understanding the mechanism by which VPAand HDAC inhibitions disrupt erythropoiesis during earlydevelopment.

doi:10.1016/j.ydbio.2008.05.339

563ABSTRACTS / Developmental Biology 319 (2008) 561–563