Name: multimodal imaging of stem cell implantation in the central nervous system of mice
Uploaded: 2012-06-13T13:00:00
Description: Watch this Scientific Journal Video about Multimodal Imaging of Stem Cell Implantation in the Central Nervous System of Mice at JoVE.com

<p class="jove_content"&gt; Авторы Работа выполнена при поддержке исследовательского гранта ID-конвертерного 2006 году в Университете Антверпена (предоставляется РРО и AVDL), на исследовательский грант G.0136.11 и G.0130.11 (предоставляется AVDL, ZB и ППО) и 1.5.021.09. N.00 (предоставляется РРО) Фонда научных исследований, Фландрии (FWO-Vlaanderen, Бельгия), по SBO грант ИВТ-60838: BRAINSTIM из Фламандского института по науке и технике (предоставляется ZB и AVDL), в частично Methusalem исследовательский грант от правительства Фландрии (предоставляется ZB), в частности, ИС-FP6-NoE Дими (LSHB-CT-2005-512146), EC-FP6-NoE EMIL (LSHC-CT-2004-503569) , а Интер университета Привлечение поляков IUAP-NIMI-P6/38 (предоставляется AVDL). Натали Де Vocht степень доктора наук, стипендия от FWO-Vlaanderen. Петр Ponsaerts это после защиты докторской диссертации в FWO-Vlaanderen.</p

1. Сотовые подготовка <ol><li> Эксперимент следует начать использование бывших естественных культурных стволовых клеточных популяций генетически модифицированных выразить люциферазы и белков EGFP репортера. Здесь мы используем люциферазы / EGFP экспрессией мышиных костномозгового происх...

<ol>
	<li>Rodriguez-Porcel, M., Wu, J. C., Gambhir, S. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Molecular imaging of stem cells. , (2008).</li><li>Bergwerf, I., De Vocht, N., Tambuyzer, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Reporter+gene-expressing+bone+marrow-derived+stromal+cells+are+immune-tolerated+following+implantation+in+the+central+nervous+system+of+syngeneic+immunocompetent+mice.">Reporter gene-expressing bone marrow-derived stromal cells are immune-tolerated following implantation in the central nervous system of syngeneic immunocompetent mice.</a> BMC Biotechnol. 9, 1 (2009).</li><li>Bergwerf, I., Tambuyzer, B., De Vocht, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Recognition+of+cellular+implants+by+the+brain's+innate+immune+system.">Recognition of cellular implants by the brain's innate immune system.</a> Immunol. Cell Biol. 89, 511-516 (2011).</li><li>Bradbury, M. S., Panagiotakos, G., Chan, B. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Optical+bioluminescence+imaging+of+human+ES+cell+progeny+in+the+rodent+CNS.">Optical bioluminescence imaging of human ES cell progeny in the rodent CNS.</a> J. Neurochem. 102, 2029-2039 (2007).</li><li>De Vocht, N., Bergwerf, I., Vanhoutte, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Labeling+of+Luciferase/eGFP-Expressing+Bone+Marrow-Derived+Stromal+Cells+with+Fluorescent+Micron-Sized+Iron+Oxide+Particles+Improves+Quantitative+and+Qualitative+Multimodal+Imaging+of+Cellular+Grafts+In+Vivo.">Labeling of Luciferase/eGFP-Expressing Bone Marrow-Derived Stromal Cells with Fluorescent Micron-Sized Iron Oxide Particles Improves Quantitative and Qualitative Multimodal Imaging of Cellular Grafts In Vivo.</a> Mol Imaging Biol. , (2011).</li><li>Reekmans, K., Praet, J., Daans, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Current+Challenges+for+the+Advancement+of+Neural+Stem+Cell+Biology+and+Transplantation+Research.">Current Challenges for the Advancement of Neural Stem Cell Biology and Transplantation Research.</a> Stem Cell Rev. , (2011).</li><li>Reekmans, K. P., Praet, J., De Vocht, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Clinical+potential+of+intravenous+neural+stem+cell+delivery+for+treatment+of+neuroinflammatory+disease+in+mice.">Clinical potential of intravenous neural stem cell delivery for treatment of neuroinflammatory disease in mice.</a> Cell Transplant. 20, 851-869 (2011).</li><li>Contag, C. H., Bachmann, M. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Advances+in+in+vivo+bioluminescence+imaging+of+gene+expression.">Advances in in vivo bioluminescence imaging of gene expression.</a> Annu. Rev. Biomed. Eng. 4, 235-260 (2002).</li><li>Sadikot, R. T., Blackwell, T. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bioluminescence+imaging.">Bioluminescence imaging.</a> Proc. Am. Thorac. Soc. 2, 511-540 (2005).</li><li>Sykova, E., Jendelova, P., Herynek, V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=MR+tracking+of+stem+cells+in+living+recipients.">MR tracking of stem cells in living recipients.</a> Methods Mol. Biol. 549, 197-215 (2009).</li><li>Daadi, M. M., Li, Z., Arac, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Molecular+and+magnetic+resonance+imaging+of+human+embryonic+stem+cell-derived+neural+stem+cell+grafts+in+ischemic+rat.">Molecular and magnetic resonance imaging of human embryonic stem cell-derived neural stem cell grafts in ischemic rat.</a> 17, 1282-1291 (2009).</li><li>Heyn, C., Ronald, J. A., Ramadan, S. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=In+vivo+MRI+of+cancer+cell+fate+at+the+single-cell+level+in+a+mouse+model+of+breast+cancer+metastasis+to+the+brain.">In vivo MRI of cancer cell fate at the single-cell level in a mouse model of breast cancer metastasis to the brain.</a> Magn. Reson. Med. 56, 1001-1010 (2006).</li><li>Hoehn, M., Kustermann, E., Blunk, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Monitoring+of+implanted+stem+cell+migration+in+vivo:+a+highly+resolved+in+vivo+magnetic+resonance+imaging+investigation+of+experimental+stroke+in+rat.">Monitoring of implanted stem cell migration in vivo: a highly resolved in vivo magnetic resonance imaging investigation of experimental stroke in rat.</a> Proc. Natl. Acad. Sci. U.S.A. 99, 16267-16272 (2002).</li><li>Jendelova, P., Herynek, V., DeCroos, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Imaging+the+fate+of+implanted+bone+marrow+stromal+cells+labeled+with+superparamagnetic+nanoparticles.">Imaging the fate of implanted bone marrow stromal cells labeled with superparamagnetic nanoparticles.</a> Magn. Reson. Med. 50, 767-776 (2003).</li><li>Modo, M., Mellodew, K., Cash, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mapping+transplanted+stem+cell+migration+after+a+stroke:+a+serial,+in+vivo+magnetic+resonance+imaging+study.">Mapping transplanted stem cell migration after a stroke: a serial, in vivo magnetic resonance imaging study.</a> Neuroimage. 21, 311-317 (2004).</li><li>Boutry, S., Brunin, S., Mahieu, I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Magnetic+labeling+of+non-phagocytic+adherent+cells+with+iron+oxide+nanoparticles:+a+comprehensive+study.">Magnetic labeling of non-phagocytic adherent cells with iron oxide nanoparticles: a comprehensive study.</a> Contrast Media Mol. Imaging. 3, 223-232 (2008).</li><li>Mailander, V., Lorenz, M. R., Holzapfel, V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Carboxylated+superparamagnetic+iron+oxide+particles+label+cells+intracellularly+without+transfection+agents.">Carboxylated superparamagnetic iron oxide particles label cells intracellularly without transfection agents.</a> Mol Imaging Biol. 10, 138-146 (2008).</li><li>Modo, M., Hoehn, M., Bulte, J. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cellular+MR+imaging.">Cellular MR imaging.</a> Mol. Imaging. 4, 143-164 (2005).</li><li>Hinds, K. A., Hill, J. M., Shapiro, E. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Highly+efficient+endosomal+labeling+of+progenitor+and+stem+cells+with+large+magnetic+particles+allows+magnetic+resonance+imaging+of+single+cells.">Highly efficient endosomal labeling of progenitor and stem cells with large magnetic particles allows magnetic resonance imaging of single cells.</a> Blood. 102, 867-872 (2003).</li><li>Shapiro, E. M., Sharer, K., Skrtic, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=In+vivo+detection+of+single+cells+by+MRI.">In vivo detection of single cells by MRI.</a> Magn. Reson. Med. 55, 242-249 (2006).</li><li>Shapiro, E. M., Skrtic, S., Sharer, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=MRI+detection+of+single+particles+for+cellular+imaging.">MRI detection of single particles for cellular imaging.</a> Proc. Natl. Acad. Sci. U.S.A. 101, 10901-10906 (2004).</li><li>Crabbe, A., Vandeputte, C., Dresselaers, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+of+MRI+contrast+agents+on+the+stem+cell+phenotype.">Effects of MRI contrast agents on the stem cell phenotype.</a> Cell Transplant. 19, 919-936 (2010).</li><li>Szarecka, A., Xu, Y., Tang, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dynamics+of+firefly+luciferase+inhibition+by+general+anesthetics:+Gaussian+and+anisotropic+network+analyses.">Dynamics of firefly luciferase inhibition by general anesthetics: Gaussian and anisotropic network analyses.</a> Biophys. J. 93, 1895-1905 (2007).</li><li>Keyaerts, M., Heneweer, C., Gainkam, L. O. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Plasma+protein+binding+of+luciferase+substrates+influences+sensitivity+and+accuracy+of+bioluminescence+imaging.">Plasma protein binding of luciferase substrates influences sensitivity and accuracy of bioluminescence imaging.</a> Mol. Imaging. Biol. 13, 59-66 (2011).</li><li>Keyaerts, M., Verschueren, J., Bos, T. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dynamic+bioluminescence+imaging+for+quantitative+tumour+burden+assessment+using+IV+or+IP+administration+of+D:+-luciferin:+effect+on+intensity,+time+kinetics+and+repeatability+of+photon+emission.">Dynamic bioluminescence imaging for quantitative tumour burden assessment using IV or IP administration of D: -luciferin: effect on intensity, time kinetics and repeatability of photon emission.</a> Eur. J. Nucl. Med. Mol. Imaging. 35, 999-1007 (2008).</li><li>Zhang, Y., Bressler, J. P., Neal, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=ABCG2/BCRP+expression+modulates+D-Luciferin+based+bioluminescence+imaging.">ABCG2/BCRP expression modulates D-Luciferin based bioluminescence imaging.</a> Cancer Res. 67, 9389-9397 (2007).</li><li>Brightwell, G., Poirier, V., Cole, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Serum-dependent+and+cell+cycle-dependent+expression+from+a+cytomegalovirus-based+mammalian+expression+vector.">Serum-dependent and cell cycle-dependent expression from a cytomegalovirus-based mammalian expression vector.</a> Gene. 194, 115-123 (1997).</li><li>Grassi, G., Maccaroni, P., Meyer, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Inhibitors+of+DNA+methylation+and+histone+deacetylation+activate+cytomegalovirus+promoter-controlled+reporter+gene+expression+in+human+glioblastoma+cell+line+U87.">Inhibitors of DNA methylation and histone deacetylation activate cytomegalovirus promoter-controlled reporter gene expression in human glioblastoma cell line U87.</a> Carcinogenesis. 24, 1625-1635 (2003).</li><li>Krishnan, M., Park, J. M., Cao, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+of+epigenetic+modulation+on+reporter+gene+expression:+implications+for+stem+cell+imaging.">Effects of epigenetic modulation on reporter gene expression: implications for stem cell imaging.</a> FASEB J. 20, 106-108 (2006).</li><li>Svensson, R. U., Barnes, J. M., Rokhlin, O. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Chemotherapeutic+agents+up-regulate+the+cytomegalovirus+promoter:+implications+for+bioluminescence+imaging+of+tumor+response+to+therapy.">Chemotherapeutic agents up-regulate the cytomegalovirus promoter: implications for bioluminescence imaging of tumor response to therapy.</a> Cancer Res. 67, 10445-10454 (2007).</li></ol>

В этом докладе мы описываем оптимизированный протокол для комбинации из трех дополнительных методов визуализации (BLI, МРТ и гистология) подробные характеристики клеточных имплантантов в ЦНС иммунокомпетентных мышей. Сочетание гена-репортера маркировки клеток, основанный на генетиче...

<table border="1"><tbody><tr><td> Название реагента </td><td> Компания </td><td> Номер по каталогу </td><td> Комментарии </td></tr><tr><td> IMDM </td><td> Lonza </td><td> Be12-722F </td><td> Компонент среды CEM рост клеток </td></tr><tr><td> Эмбриональной телячьей сыворотки </td><td> Гибко </td><td> 10270-106 </td><td> Компонент среды CEM рост клеток </td></tr><tr><td> Лошадиная сыворотка </td><td> Гибко </td><td> 1605-122 </td><td> Компонент среды CEM рост клеток </td></tr><tr><td> Пенициллин, стрептомицин </td><td> Гибко </td><td> 15140 </td><td> Компонент среды CEM рост клеток </td></tr><tr><td> Fungizone </td><td> Гибко </td><td> 15290-018 </td><td> Компонент среды CEM рост клеток </td></tr><tr><td> PBS </td><td> Гибко </td><td> 14190 </td><td></td></tr><tr><td> Puromycine </td><td> Invivogen </td><td> Муравей-пр-1 </td><td></td></tr><tr><td> трипсин </tг&gt; <td> Гибко </td><td> 25300 </td><td></td></tr><tr><td> ГБ MPIO </td><td> Удары лаборатории </td><td> ME04F/7833 </td><td></td></tr><tr><td> D-люциферин </td><td> Promega </td><td> E1601 </td><td></td></tr><tr><td> Кетамин (Ketalar) </td><td> Pfizer </td><td></td><td></td></tr><tr><td> Ксилазин (Rompun) </td><td> Bayer здравоохранения </td><td></td><td></td></tr><tr><td> Изофлюран </td><td> Isoflo </td><td> 05260-05 </td><td></td></tr><tr><td> 0,9% раствором хлористого натрия </td><td> Бакстер </td><td></td><td></td></tr><tr><td> параформальдегид </td><td> Merck </td><td> 1.04005.1000 </td><td></td></tr><tr><td> сахароза </td><td> AppliChem </td><td> A1125 </td><td></td></tr><tr><td> Micro-топливного насоса </td><td> KD научных </td><td> KDS100 </td><td></td></tr><tr><td> Фотон тепловизор </td><td> Biospace лаборатории </td><td></td><td></td></tr><tr&gt; <td> 9.4T МР-сканер </td><td> Bruker BioSpin </td><td> Биоспек 94/20 USR </td><td></td></tr><tr><td> BX51 микроскоп </td><td> Олимп </td><td> BX51 </td><td></td></tr><tr><td> Mycrom HM криостат </td><td> Prosan </td><td> HM525 </td><td></td></tr><tr><td> шприц </td><td> Гамильтон </td><td> 7635-01 </td><td></td></tr><tr><td> 30 иглы </td><td> Гамильтон </td><td> 7762-03 </td><td></td></tr><tr><td> Фотография видение программного обеспечения </td><td> Biospace лаборатории </td><td></td><td></td></tr><tr><td> M3vision программного обеспечения </td><td> Biospace лаборатории </td><td></td><td></td></tr><tr><td> Paravision 5,1 программного обеспечения </td><td> Bruker BioSpin </td><td></td><td></td></tr><tr><td> Амира 4,0 программного обеспечения </td><td> Visage изображений </td><td></td><td></td></tr></tbody></table>

multimodal imaging of stem cell implantation in the central nervous system of mice

During the past decade, stem cell transplantation has gained increasing interest as primary or secondary therapeutic modality for a variety of diseases, both in preclinical and clinical studies. However, to date results regarding functional outcome and/or tissue regeneration following stem cell transplantation are quite diverse. Generally, a clinical benefit is observed without profound understanding of the underlying mechanism(s)1. Therefore, multiple efforts have led to the development of different molecular imaging modalities to monitor stem cell grafting with the ultimate aim to accurately evaluate survival, fate and physiology of grafted stem cells and/or their micro-environment. Changes observed in one or more parameters determined by molecular imaging might be related to the observed clinical effect. In this context, our studies focus on the combined use of bioluminescence imaging (BLI), magnetic resonance imaging (MRI) and histological analysis to evaluate stem cell grafting.
BLI is commonly used to non-invasively perform cell tracking and monitor cell survival in time following transplantation2-7, based on a biochemical reaction where cells expressing the Luciferase-reporter gene are able to emit light following interaction with its substrate (e.g. D-luciferin)8, 9. MRI on the other hand is a non-invasive technique which is clinically applicable10 and can be used to precisely locate cellular grafts with very high resolution11-15, although its sensitivity highly depends on the contrast generated after cell labeling with an MRI contrast agent. Finally, post-mortem histological analysis is the method of choice to validate research results obtained with non-invasive techniques with highest resolution and sensitivity. Moreover end-point histological analysis allows us to perform detailed phenotypic analysis of grafted cells and/or the surrounding tissue, based on the use of fluorescent reporter proteins and/or direct cell labeling with specific antibodies.
In summary, we here visually demonstrate the complementarities of BLI, MRI and histology to unravel different stem cell- and/or environment-associated characteristics following stem cell grafting in the CNS of mice. As an example, bone marrow-derived stromal cells, genetically engineered to express the enhanced Green Fluorescent Protein (eGFP) and firefly Luciferase (fLuc), and labeled with blue fluorescent micron-sized iron oxide particles (MPIOs), will be grafted in the CNS of immune-competent mice and outcome will be monitored by BLI, MRI and histology (Figure 1).

Watch this Scientific Journal Video about Multimodal Imaging of Stem Cell Implantation in the Central Nervous System of Mice at JoVE.com

Multimodal Imaging of Stem Cell Implantation in the Central Nervous System of Mice

В этой статье описываются оптимизированные последовательности событий для мультимодальных изображений клеточных трансплантатов у грызунов мозга с помощью: (I) в естественных условиях биолюминесценции и магнитно-резонансной томографии, и (II) после смерти гистологический анализ. Сочетание этих условий изображение на одного животного позволяет сотовой трансплантата оценки с высоким разрешением, чувствительностью и специфичностью.

Мультимодальные изображений имплантации стволовых клеток в центральной нервной системе мышей

During the past decade, stem cell transplantation has gained increasing interest as primary or secondary therapeutic modality for a variety of diseases, ...

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In Situ Ca2+ Imaging of the Enteric Nervous System

В Ситу Ca 2+ изображений нервной системы кишечной

Reflex behaviors of the intestine are controlled by the enteric nervous system (ENS). The ENS is an integrative network of neurons and glia in two ...

Optical Clearing of the Mouse Central Nervous System Using Passive CLARITY

Оптический Клиринговый нервной системы мыши Центральной использованием пассивных ЯСНОСТИ

Traditionally, tissue visualization has required that the tissue of interest be serially sectioned and imaged, subjecting each tissue section to unique ...

Electrophysiological Recording of The Central Nervous System Activity of Third-Instar Drosophila Melanogaster

Electrophysiological Recording of The Central Nervous System Activity of Third-Instar  Drosophila Melanogaster

Электрофизиологические запись третьего Instar Drosophila Melanogaster деятельности центральной нервной системы

The majority of the currently available insecticides target the nervous system and genetic mutations of invertebrate neural proteins oftentimes yield ...