<p class="jove_content"> Gli autori sono grati per i contributi artistici di Hyewon Cooper. </p>

<ol>
	<li>Whitear, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Chordotonal+organs+in+Crustacea.">Chordotonal organs in Crustacea.</a> <em style='font-style: italic'>Nature</em>. <b>187</b>, 522-523 (1038).</li><li>Alexandrowicz, J. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+comparative+anatomy+of+leg+propriocetors+in+some+decapod+Crustacea.">The comparative anatomy of leg propriocetors in some decapod Crustacea.</a> <em style='font-style: italic'>J. Mar. Biol. Assoc. U.K</em>. <b>52</b> (3), 605-634 (1972).</li><li>Hartman, H. B., Boettiger, E. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+functional+organization+of+the+propys-dactylus+organ+in+Cancer+irroratus+Say.">The functional organization of the propys-dactylus organ in Cancer irroratus Say.</a> <em style='font-style: italic'>Comp. Biochem. Physiol</em>. <b>22</b>, 651-663 (1967).</li><li>Cooper, R. L., Hartman, H. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Responses+of+the+bender+apodeme+tension+receptors+in+the+Dungeness+crab,+Cancer+magister.">Responses of the bender apodeme tension receptors in the Dungeness crab, Cancer magister.</a> <em style='font-style: italic'>Comp. Biochem. Physiol. A Physiol</em>. <b>109</b> (2), 479-486 (1994).</li><li>Cooper, R. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mapping+proprioceptive+neurons+on+chordotonal+organs+in+the+crab,+Cancer+magister.">Mapping proprioceptive neurons on chordotonal organs in the crab, Cancer magister.</a> <em style='font-style: italic'>Crustaceana</em>. <b>81</b> (4), 447-475 (2008).</li><li>Burke, W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=An+organ+for+proprioception+and+vibration+sense+in+Carcinus+maenas.">An organ for proprioception and vibration sense in Carcinus maenas.</a> <em style='font-style: italic'>J. Exp. Biol</em>. <b>31</b>, 127-138 (1953).</li><li>Hill, A. V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> <em style='font-style: italic'>First and Last Experiments in Muscle Mechanics</em>. , (1970).</li><li>Stuart, D. G., Mosher, C. C., Gerlach, R. L., Reinking, R. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mechanical+arrangement+and+transducing+properties+of+Golgi+tendon+organs.">Mechanical arrangement and transducing properties of Golgi tendon organs.</a> <em style='font-style: italic'>Exp. Brain Res</em>. <b>14</b> (3), 274-292 (1972).</li><li>Macmillan, D. L., Dando, M. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Tension+receptors+on+the+apodemes+of+muscles+in+the+walking+legs+of+the+crab,+Cancer+magister.">Tension receptors on the apodemes of muscles in the walking legs of the crab, Cancer magister.</a> <em style='font-style: italic'>Mar. Behav. Physiol</em>. <b>1</b> (1-4), 185-208 (1972).</li><li>B&#233;vengut, M., Simmers, A. J., Clarac, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Central+neuronal+projections+and+neuromuscular+organization+of+the+basal+region+of+the+shore+crab+leg.">Central neuronal projections and neuromuscular organization of the basal region of the shore crab leg.</a> <em style='font-style: italic'>J. Comp. Neurol</em>. <b>221</b> (2), 185-198 (1983).</li><li>El Manira, A., Cattaert, D., Clarac, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Monosynaptic+connections+mediate+resistance+reflex+in+crayfish+(Procambarus+clarkii)+legs.">Monosynaptic connections mediate resistance reflex in crayfish (Procambarus clarkii) legs.</a> <em style='font-style: italic'>J. Comp. Physiol. A</em>. <b>168</b> (3), 337-349 (1991).</li><li>Le Bon-Jego, M., Cattaert, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Inhibitory+component+of+the+resistance+reflex+in+the+locomotor+network+of+the+crayfish.">Inhibitory component of the resistance reflex in the locomotor network of the crayfish.</a> <em style='font-style: italic'>J. Neurophysiol</em>. <b>88</b> (5), 2575-2588 (2002).</li><li>Ray, D. L., Clarac, F., Cattaert, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Functional+analysis+of+the+sensory+motor+pathway+of+resistance+reflex+in+crayfish.+I.+Multisensory+coding+and+motor+neuron+monosynaptic+responses.">Functional analysis of the sensory motor pathway of resistance reflex in crayfish. I. Multisensory coding and motor neuron monosynaptic responses.</a> <em style='font-style: italic'>J. Neurophysiol</em>. <b>78</b> (6), 3133-3143 (1997).</li><li>Wiersma, C. A. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Vergleichende+Untersuchungen&#252;ber+das+periphere+Nerve-muskelsystem+von+Crustaceen+(Comparative+studies+on+the+peripheral+nerve-muscle+system+of+crustaceans).">Vergleichende Untersuchungen&#252;ber das periphere Nerve-muskelsystem von Crustaceen (Comparative studies on the peripheral nerve-muscle system of crustaceans).</a> <em style='font-style: italic'>J. Comp. Physiol</em>. <b>19</b>, 349-385 (1933).</li><li>Wiersma, C. A. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Movement+receptors+in+decapod+crustacea.">Movement receptors in decapod crustacea.</a> <em style='font-style: italic'>J. Mar. Biol. Assoc. U.K</em>. <b>38</b> (01), 01-143 (1959).</li><li>Hartman, H. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Tension+receptors+on+the+closer+muscle+apodeme+in+the+walking+legs+of+the+blue+crab+Callinectes+sapidus.">Tension receptors on the closer muscle apodeme in the walking legs of the blue crab Callinectes sapidus.</a> <em style='font-style: italic'>J. Comp. Physiol. A</em>. <b>157</b> (3), 355-362 (1985).</li><li>Holsinger, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> <em style='font-style: italic'>The effect of regional phenotypic differences of Procambarus clarkii opener muscle on sarcomere length, fiber diameter, and force development MSc thesis [dissertation]</em>. , (2013).</li><li>Parsons, D. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+morphology+and+ultrastructure+of+tension+receptors+in+the+walking+legs+of+the+crab,+Carcinus+maenas.">The morphology and ultrastructure of tension receptors in the walking legs of the crab, Carcinus maenas.</a> <em style='font-style: italic'>Cell Tissue Res</em>. <b>211</b> (1), 139-149 (1980).</li><li>Parsons, D. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Responses+and+central+interactions+of+tension+receptors+in+the+leg+muscle+of+Carcinus.">Responses and central interactions of tension receptors in the leg muscle of Carcinus.</a> <em style='font-style: italic'>Comp. Biochem. Physiol. A Physiol</em>. <b>72</b> (2), 391-399 (1982).</li><li>Tryba, A. K., Hartman, H. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dynamic+responses+of+series+force+receptors+innervating+the+opener+muscle+apodeme+in+the+blue+crab,+Callinectes+sapidus.">Dynamic responses of series force receptors innervating the opener muscle apodeme in the blue crab, Callinectes sapidus.</a> <em style='font-style: italic'>J. Comp. Physiol. A</em>. <b>180</b> (3), 215-221 (1997).</li><li>Whitear, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+fine+structure+of+crustacean+proprioceptors.+I.+The+chordotonal+organs+in+the+legs+of+the+shore+crab,+Carcinus+meanas.">The fine structure of crustacean proprioceptors. I. The chordotonal organs in the legs of the shore crab, Carcinus meanas.</a> <em style='font-style: italic'>Philos. Trans. R. Soc. Lond. B Biol. Sci</em>. <b>245</b> (725), 291-325 (1962).</li><li>Whitear, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+fine+structure+of+crustacean+proprioceptors.+II.+The+thoracico-coxal+organs+in+Carcinus,+Pagurus+and+Astacus.">The fine structure of crustacean proprioceptors. II. The thoracico-coxal organs in Carcinus, Pagurus and Astacus.</a> <em style='font-style: italic'>Philos. Trans. R. Soc. Lond. B Biol. Sci</em>. <b>248</b> (752), 437-456 (1965).</li><li>Leksrisawat, B., Cooper, A. S., Gilberts, A. B., Cooper, R. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Muscle+Receptor+Organs+in+the+Crayfish+Abdomen:+A+Student+Laboratory+Exercise+in+Proprioception.">Muscle Receptor Organs in the Crayfish Abdomen: A Student Laboratory Exercise in Proprioception.</a> <em style='font-style: italic'>J. Vis. Exp</em>.  (45), e2323 (2010).</li><li>Baierlein, B., Thurow, A. L., Atwood, H. L., Cooper, R. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Membrane+Potentials,+Synaptic+Responses,+Neuronal+Circuitry,+Neuromodulation+and+Muscle+Histology+Using+the+Crayfish:+Student+Laboratory+Exercises.">Membrane Potentials, Synaptic Responses, Neuronal Circuitry, Neuromodulation and Muscle Histology Using the Crayfish: Student Laboratory Exercises.</a> <em style='font-style: italic'>J. Vis. Exp</em>.  (47), e2322 (2011).</li><li>Delaney, K., Gelperin, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cerebral+interneurons+controlling+fictive+feeding+in+Limaxmaximus;+I.+Anatomy+and+criteria+for+re-identification.">Cerebral interneurons controlling fictive feeding in Limaxmaximus; I. Anatomy and criteria for re-identification.</a> <em style='font-style: italic'>J. Comp. Physiol. A</em>. <b>166</b>, 297-310 (1990).</li><li>Hartman, H. B., Cooper, R. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Regeneration+and+molting+effects+on+a+proprioceptor+organ+in+the+Dungeness+crab,+Cancer+magister.">Regeneration and molting effects on a proprioceptor organ in the Dungeness crab, Cancer magister.</a> <em style='font-style: italic'>J. Neurobiol</em>. <b>25</b> (5), 461-471 (1994).</li><li>Hartman, H. B., Austin, W. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Proprioceptor+organs+in+the+antennae+of+Decapoda+Crustacea.">Proprioceptor organs in the antennae of Decapoda Crustacea.</a> <em style='font-style: italic'>J. Comp. Physiol</em>. <b>81</b> (2), 187-202 (1972).</li><li>Cooper, R. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Development+of+sensory+processes+during+limb+regeneration+in+adult+crayfish.">Development of sensory processes during limb regeneration in adult crayfish.</a> <em style='font-style: italic'>J. Exp. Biol</em>. <b>201</b> (Pt 11), 1745-1752 (1998).</li><li>Chung, Y. -. S., Cooper, R. M., Graff, J., Cooper, R. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+acute+and+chronic+effect+of+low+temperature+on+survival,+heart+rate+and+neural+function+in+crayfish+(Procambarus+clarkii)+and+prawn+(Macrobrachium+rosenbergii)+species.">The acute and chronic effect of low temperature on survival, heart rate and neural function in crayfish (Procambarus clarkii) and prawn (Macrobrachium rosenbergii) species.</a> <em style='font-style: italic'>Open J. Mol. Int. Physiol</em>. <b>2</b> (3), 75-86 (2012).</li><li>Blundon, J. A. <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+temperature+and+thermal+history+on+neuromuscular+properties+of+two+crustacean+species.">Effects of temperature and thermal history on neuromuscular properties of two crustacean species.</a> <em style='font-style: italic'>J. Comp. Physiol. B</em>. <b>158</b> (6), 689-696 (1989).</li><li>Cooper, R. M., Schapker, H., Adami, H., Cooper, R. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Heart+and+ventilatory+measures+in+crayfish+during+copulation.">Heart and ventilatory measures in crayfish during copulation.</a> <em style='font-style: italic'>Open J. Mol. Int. Physiol</em>. <b>1</b> (3), 36-42 (2011).</li><li>Bierbower, S. M., Cooper, R. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+mechanistic+action+of+carbon+dioxide+on+a+neural+circuit+and+NMJ+communication.">The mechanistic action of carbon dioxide on a neural circuit and NMJ communication.</a> <em style='font-style: italic'>Journal of Experimental Zoology</em>. , (2013).</li><li>Bierbower, S. M., Shuranova, Z. P., Viele, K., Cooper, R. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sensory+systems+and+environmental+change+on+behavior+during+social+interactions.">Sensory systems and environmental change on behavior during social interactions.</a> <em style='font-style: italic'>Brain Behav</em>. <b>3</b> (1), 4-13 (2013).</li></ol>

<p class="jove_content"> Gli autori dichiarano di non avere interessi finanziari in competizione. </p>

<p class="jove_content"> Lo scopo di questa serie di esperimenti è 1) insegnare ed esporre i principi fondamentali di registrazioni extracellulari da un organo propriocettiva identificabile e la tensione nervosa e 2) per sottolineare l&#39;importanza della mappatura anatomica in relazione alla funzione fisiologica di particolari sistemi sensoriali. Questo approccio sperimentale e modelli animali utilizzati sono poco costoso e relativamente facile da condurre in laboratori didattici neurofisiologia. </p><p class="jove_content"> I neuroni di organi cordotonali sono di due tipi funzionali specifici, quelli che rispondono al movimento e coloro che rispondono a posizioni statiche. Registrazioni di cellule singole a partire da una varietà di organi cordotonali, non importa quale viene esaminato congiunta, hanno dimostrato che questo sia il caso <sup>di 3,5.</sup> Infatti, gli organi cordotonali associati alle articolazioni antennali di aragoste rivelano gli stessi due tipi sensoriali e anatomia di base <sup>27.</sup> Oltre a ci siano due tipi di neuroni (movimento e position), i neuroni condividono la stessa disposizione anatomica sui rispettivi fili elastici. Il grande somata trova prossimale sul filamento tendono ad appartenere ai neuroni sensibili movimento dinamico. I neuroni che segnalano posizioni statiche hanno piccole somata e si trovano distale. Queste cellule sono tonico attivo. Il giunto PD contiene solo un singolo organo cordotonali mentre ci sono due organi cordotonali nel carpo-propodus (CP) e merus-carpo (MC) articolazioni. </p><p class="jove_content"> La dissezione di esporre le strutture propriocettivi in granchi blu <em>(C. sapidus)</em> per la registrazione elettrofisiologica richiede una strategia che permette movimenti articolari che si terrà nelle posizioni naturali, mentre la registrazione da neuroni sensoriali. Il nervo tensione per il muscolo opener nella gamba a piedi è un bellissimo nervo composto da diversi neuroni. A meno che non si fa attenzione, il nervo tensione nonché il nervo motore che innervano il muscolo da stimolare, possono essere danneggiati durante questa dissezione. Perregistrazioni ottimali gli elettrodi di aspirazione devono essere adattati alle dimensioni del nervo. Le registrazioni sono facilmente accessibili in un laboratorio studente utilizzando un microscopio ingrandimento di dissezione 30-40X e micromanipolatori di fascia bassa. </p><p class="jove_content"> Futuri esperimenti che sarebbe interessante proseguire con gli organi cordotonali comune sarebbe quello di esaminare i profili strutturali e fisiologiche durante la rigenerazione gamba in varie specie nelle diverse fasi del ciclo di vita come un follow-up a un primo studio che ha utilizzato <em>Cancer magister</em> <sup>19,26</sup> . Domande ancora da affrontare sono: 1) non effettua la distribuzione e l&#39;organizzazione dei neuroni artificiali dipendono dall&#39;età dell&#39;animale quando rigenerare un arto, 2) sono le proiezioni assonale al CNS (cordone nervoso ventrale) in un arto rigenerante funzionale o lo fa prendere tempo e l&#39;uso congiunto di stabilire connessioni funzionali, e 3) che cosa accade agli assoni mozzate prossimale al piano autotomy quando l&#39;arto è autotomized? <sup>28</sup> </p><p class="jove_content"> Crostacei conformi alle condizioni ambientali e la loro temperatura circostante, ma non è chiaro come si mantengono coordinamento all&#39;interno di un circuito neurale come neuroni alterano la loro attività in risposta alle variazioni di temperatura. Un basso tasso di cambiamento potrebbe permettere all&#39;animale po &#39;di tempo per acclimatarsi, mentre un rapido cambiamento può not29, 30. Cambiamenti fisiologici di pH o di osmolarità a causa di metabolismo, il comportamento <sup>31,</sup> o l&#39;impatto ambientale, possono presentare problemi simili ai circuiti neurali coinvolti nella propriocezione. Queste preparazioni crostacei sono ideali per affrontare questi tipi di problemi perché la loro funzione è ben caratterizzata a livello di singola cellula. </p><p class="jove_content"> In questo protocollo abbiamo dimostrato l&#39;importanza fisiologica dei neuroni tensione in forza generata dal muscolo apri monitoraggio. Questi recettori di tensione possono essere ricondotte alla loro posizione all&#39;interno della apodeme utilizzando procedure di colorazione. Questeneuroni, come nei mammiferi, rilevano forza a vari livelli e reclutare neuroni addizionali come la forza aumenta. La frequenza di attività è correlata alla frequenza di stimolazione del motoneurone fino a saturazione in ricezione. Utilizzando un protocollo sgancio rapido con il giunto dactylus flessa, attività tensione sparisce rapidamente, ma poi torna sui riconquistare la tensione in un giunto completamente esteso. Questa è una procedura sperimentale classico per illustrare la forza misurata dai recettori di tensione. Vari neuromodulatori possono essere applicati alla preparazione di vedere come effettua lo sviluppo della forza e della risposta neuronale. Uno degli aspetti importanti è come le risposte neurali vengono elaborati e integrati nel sistema nervoso centrale e loro impatto sull&#39;attività dei neuroni motori. Le tecniche che abbiamo mostrato permettono di iniziare ad affrontare ulteriori informazioni sulla (sensoriale) Funzione circuito neurone nervo motore tensione, <em>cioè</em> il segnale in una gamba intatta al ganglio e ritornoal muscolo. </p><p class="jove_content"> Le procedure di colorazione sono dimostrati chiave per comprendere la fisiologia dei neuroni sensoriali che innervano organi propriocettivi. Disposizione anatomica dei neuroni in base alla funzione e la dimensione del soma sono simili nei vari organi cordotonali all&#39;interno delle gambe di granchio. Non è noto se accordi neuronali simili si trovano anche in altre specie di insetti o crostacei. Combinando le registrazioni fisiologiche da singole cellule e la mappatura della posizione permette relazioni funzionali struttura diretta. La conservazione a lungo termine del regime anatomica con CoCl <sub>2</sub> colorazione e dal fissaggio permette di fare ripetutamente le misure e valutare la disposizione strutturale. </p><p class="jove_content"> Propriocezione e ricezione tensione dei muscoli scheletrici sono modalità sensoriali che consentono comportamenti coordinati e risposte ambiente esterno ed interno per animali articolati in una varietà di configurazioni di muscolo scheletricos. L&#39;organo recettore muscolare nell&#39;addome del gambero è un altro preparato ben documentata (vedi il Progetto Crawdad, <a href="http://www.crawdad.cornell.edu/">http://www.crawdad.cornell.edu/</a> ) a fini didattici di propriocezione con soli due neuroni per addominali emi-segmento <sup>23</sup> . Essere in grado di registrare da singoli neuroni a fasci nervosi sensoriali fornisce ulteriori dettagli che aiuti a comprendere i principi fondamentali della percezione sensoriale. Questi relativamente semplici preparazioni di crostacei permettono di affrontare gli aspetti fondamentali della propriocezione e controllo tensione, con il potenziale per determinare i circuiti neurali che consentono l&#39;integrazione centrale di ingressi propriocettivi e altri sensoriali <sup>9-12, 32, 33.</sup> </p>

<p class="jove_content"> Propriocezione è la sensazione di posizione dell&#39;arto e movimento che consente comportamento coordinato motore. Propriocettori consistono in posizione (statico) e il movimento recettori (cinestetica). In insetti e crostacei, gli organi cordotonali sono le strutture che forniscono tali informazioni al sistema nervoso centrale <sup>1.</sup> Non tutti gli organi cordotonali abbracciano una comune, ma possono ancora controllare i movimenti articolari a causa del loro attaccamento alle apodemes (come le strutture tendinee), che coprono il giunto e si muovono in associazione con il muscolo scheletrico e l&#39;articolazione congiunta. Granchio gambe hanno sei giunti, ciascuno con uno o due organi cordotonali <sup>2.</sup> Tipicamente un organo cordotonali ha 60-100 o più neuroni sensoriali incorporati all&#39;interno di un filo elastico, i neuroni che segnalano la posizione comune statica, direzione e velocità di movimento <sup>3-6.</sup> L&#39;ingresso da organi cordotonali ad ogni articolazione e la gamba viene poi elaborato centralmente permettendo movimenti coordinati da parte degli animali. </p><p class="jove_content"&gt; Le forze che muscoli delle gambe producono durante contrazioni isometriche e isotoniche vengono rilevati dai recettori di tensione associati alle fibre muscolari e dei loro allegati ai apodemes7-9. Nei protocolli gambe piedi crostacei che seguono vi presentiamo metodologia per registrazioni da neuroni sensoriali primari che controllano la propriocezione e neuroni che rispondono alle forze generate da fibre muscolari. Una tecnica per l&#39;attivazione movimenti delle gambe e quantificare generazione della forza è presentato anche, così come le tecniche anatomiche che possono essere utilizzati per caratterizzare la disposizione di queste strutture del sistema nervoso periferico. </p><p class="jove_content"> Le procedure illustrate in dettaglio successivamente consentire l&#39;analisi strutturale e funzionale dei neuroni che innervano entrambi i tipi di recettori relativi alla loro posizione su un filo elastico cordotonali e apodeme. Per illustrare, usiamo il propodite-dactylopodite (PD) organo cordotonali, l&#39;organo che attraversa il segmento più distale della gamba granchio <sup>3.</sup>Anche se studi elettrofisiologici dettagliati iniziarono nel 1930 e sono ancora in corso oggi, alcuni aspetti sono diventati noti per le connessioni segmentale dei propriocettori nelle varie articolazioni e il loro ruolo nel controllo coordinato di muscles10-16. Stabilire il rapporto struttura-funzione tra gli organi propriocettivi, i muscoli e il sistema nervoso ulteriormente aiutare a definire questi ruoli. Ad esempio, l&#39;etichettatura del somata e terminazioni distali dei neuroni tensione inserite nel apodeme rivelerà la loro posizione rispetto alle fibre muscolari <sup>8,17-21.</sup> </p><p class="jove_content"> Presentiamo tre tecniche di colorazione per le gambe dei crostacei che possono essere utilizzati nella ricerca o laboratori accademici. Metilene colorazione blu offre un contrasto adeguato per muscoli e nervi ed è raccomandato come una semplice tecnica per gli studenti di imparare l&#39;anatomia. Labs che hanno configurazioni microscopia a fluorescenza possono realizzare più selettivo colorazione neuronale esponendo brevemente il nervos al colorante vitale a 4-di-2-ASP. La terza alternativa è CoCl <sub>2</sub> di recupero, che colora e fissa i neuroni, e non necessita di imaging di fluorescenza. Anche se è lavoro e tempo intenso, questo processo di colorazione conferisce un elevato contrasto e specificità per i nervi che sono pieni. Insieme, queste tecniche possono essere utilizzate per la comparazione dei vari organi cordotonali, non solo all&#39;interno di un arto o tra gli arti, ma anche tra le altre specie di crostacei e insetti <sup>20-22.</sup> Granchi blu <em>(C. sapidus)</em> utilizzati nelle registrazioni fisiologiche e anatomiche per la colorazione sono facilmente disponibili in tutto il confine meridionale e sud-est degli Stati Uniti. Questa specie funge da rappresentante del regime nervose cordotonali e la tensione si trovano nella maggior parte dei granchi. Laboratori sulla costa occidentale preferiscono utilizzare il molto più grande granchio Dungeness <em>(Cancer magister)</em> per questi esperimenti. </p>

<table><tbody><tr><td>Sylgard</td><td>Dow Corning</td><td>182 silicone kit</td><td></td></tr><tr><td>NaCl</td><td>Sigma-Aldrich</td><td>S7653</td><td></td></tr><tr><td>KCl</td><td>Sigma-Aldrich</td><td>P9333</td><td></td></tr><tr><td>CaCl&lt;sub&gt;2&lt;/sub&gt;</td><td>Sigma-Aldrich</td><td>C5670</td><td></td></tr><tr><td>HEPES acid</td><td>Sigma</td><td>3375</td><td>Free acid, crystalline</td></tr><tr><td>HEPES base</td><td>Sigma</td><td></td><td></td></tr><tr><td>D-Glucose</td><td>Sigma</td><td>G7021</td><td></td></tr><tr><td>MgSO&lt;sub&gt;4&lt;/sub&gt;&amp;bull;7H&lt;sub&gt;2&lt;/sub&gt;O</td><td>Sigma</td><td>M2643</td><td></td></tr><tr><td>Na&lt;sub&gt;2&lt;/sub&gt;SO&lt;sub&gt;4&lt;/sub&gt;&amp;bull;10H&lt;sub&gt;2&lt;/sub&gt;O</td><td>Sigma</td><td>246980</td><td></td></tr><tr><td>Bouin&amp;rsquo;s solution fixative</td><td>Sigma</td><td>HT10-1-32</td><td>Caution: Hazardous material (Special shipping cost required)</td></tr><tr><td>CoCl&lt;sub&gt;2&lt;/sub&gt;</td><td>Sigma</td><td></td><td>Caution: Hazardous material. Please follow proper disposal according to local and federal regulations.</td></tr><tr><td>Methylene blue chloride</td><td>Matheson Co., Inc</td><td>Basic Blue 9, C.I. 52015</td><td></td></tr><tr><td>4-Di-2-ASP</td><td>Molecular Probes</td><td></td><td>4-(4-diethylaminostyryl)-N-methylpyridinium iodide</td></tr><tr><td>Bleach</td><td>Sigma-Aldrich</td><td></td><td>To chloride silver wire</td></tr><tr><td>NaOH</td><td>Sigma-Aldrich</td><td>221465</td><td>To adjust pH</td></tr><tr><td>HCl</td><td>Sigma-Aldrich</td><td>H1758</td><td>To adjust pH</td></tr><tr><td></td><td></td><td></td><td></td></tr><tr><td>Materials</td><td></td><td></td><td></td></tr><tr><td>Dissecting tools</td><td>World Precision Instruments</td><td>assortment</td><td></td></tr><tr><td>Intracellular electrode probe</td><td></td><td></td><td></td></tr><tr><td>Faraday cage</td><td></td><td></td><td></td></tr><tr><td>Insect Pins</td><td>Fine Science Tools, Inc</td><td>26001-60</td><td></td></tr><tr><td>Dissecting microscope (100X)</td><td></td><td></td><td></td></tr><tr><td>Fiber optic lamp</td><td></td><td></td><td></td></tr><tr><td>Small adjustable mirror</td><td></td><td></td><td>To direct light beam toward the preparation.</td></tr><tr><td>Glass electrodes</td><td>Sigma-Aldrich</td><td>CLS7095B5X</td><td>Box of 200, suction electrodes</td></tr><tr><td>Micromanipulator</td><td>World Precision Instruments</td><td>MD4-M3-R</td><td>Can fix for base or on a metal rod</td></tr><tr><td>Raised preparation stand</td><td></td><td></td><td></td></tr><tr><td>Silver wire (10/1,000 inch)</td><td>A-M Systems</td><td>782500</td><td></td></tr><tr><td>Computer</td><td>Any company</td><td></td><td></td></tr><tr><td>AC/DC differential amplifier</td><td>A-M Systems</td><td>Model 3000</td><td></td></tr><tr><td>PowerLab 26T</td><td>AD Instruments</td><td>27T</td><td></td></tr><tr><td>Force transducer</td><td>AD Instruments</td><td>0-50g</td><td>MLTF050/ST</td></tr><tr><td>Head stage</td><td>AD Instruments</td><td></td><td>Comes with AC/DC amplifier</td></tr><tr><td>LabChart7</td><td>AD Instruments</td><td></td><td></td></tr><tr><td>Electrical leads</td><td>Any company</td><td></td><td></td></tr><tr><td>Glass tools</td><td>Make yourself</td><td></td><td>For manipulating nerves</td></tr><tr><td>Cable and connectors</td><td>Any company</td><td></td><td></td></tr><tr><td>Pipettes with bulbs</td><td>Fisher Scientific</td><td>13-711-7</td><td>Box of 500</td></tr><tr><td>Beakers</td><td>Any company</td><td></td><td></td></tr><tr><td>Wax or modeling clay</td><td>Any company or local stores</td><td></td><td></td></tr><tr><td>Stimulator</td><td>Grass Instruments</td><td>SD9 or S88</td><td></td></tr><tr><td>Plastic tip for suction electrode</td><td>Local hardware store (Watt&amp;rsquo;s brand)</td><td>&amp;frac14; inch OD x 0.170 inch ID</td><td>Cut in small pieces. Pull out over a flame and cut back the tip to the correct size</td></tr></tbody></table>

proprioception and tension receptors in crab limbs: student laboratory exercises

<p class="jove_content"> Lo scopo principale di queste procedure è di dimostrare per fini didattici e di ricerca come registrare l&#39;attività dei neuroni sensoriali primari vivere responsabili della propriocezione come sono Rilevazione della posizione comune e il movimento, e la tensione muscolare. Attività elettrica da propriocettori crostacei e recettori di tensione viene registrata dalla strumentazione neurofisiologico di base, e un trasduttore viene utilizzato per misurare simultaneamente forza generata stimolando un nervo motore. Inoltre, dimostriamo come macchiare i neuroni per una rapida valutazione della loro organizzazione anatomica o per il fissaggio permanente. La colorazione rivela organizzazione anatomica che è rappresentativo di organi cordotonali nella maggior parte dei crostacei. Confrontando le risposte tensione nervose verso le risposte propriocettive è uno strumento didattico efficace nel determinare come questi neuroni sensoriali sono definiti funzionalmente e come l&#39;anatomia è correlata alla funzione. Tre tecniche di colorazionesono presentati consentendo ai ricercatori e istruttori di scegliere un metodo che è ideale per il loro laboratorio. </p>

<p class="jove_content"> Quando l&#39;organo PD è allungato estendendo completamente il giunto, attività nel nervo PD è robusto durante il movimento come illustrato per il primo secondo in <strong>Figura 10.</strong> Alcune attività rimane mentre è tenuto fermo nella posizione di apertura. Questa attività è dai neuroni sensibili statici posizione (seconda metà di registrazione mostrato nella <strong>Figura 10).</strong> Movimento evoca una risposta durante lo spostamento, e la cottura è presente soprattutto durante l&#39;allungamento del filamento cordotonali <strong>(Figura 11).</strong> </p><p class="jove_content"> Ulteriori analisi delle punte può essere facilmente avvicinato di classificare le relative ampiezze. Questo è un approccio per dimostrare le diverse popolazioni di neuroni sensoriali essere reclutati per posizioni o tipi di movimenti <sup>5.</sup> Ampiezze tipiche vanno 0,25-1,5 mV, ma questi valori dipendono dalla resistenza <em>(cioè</em> tenuta) della tenuta elettrodo di aspirazione. Frequenza dei picchi di tha varie gamme di dimensioni può anche essere rappresentato graficamente per l&#39;analisi. </p><p class="jove_content"> Forze generate dal muscolo apri rispetto alla frequenza di stimolazione possono essere confrontati sovrapponendo le rispettive tracce tensione-tempo sopra l&#39;altra <strong>(Figure 13A</strong> e <strong>B).</strong> Questo può anche essere eseguita per ogni posizione collettiva a ogni data frequenza di stimolazione. Attività del nervo tensione può poi essere correlato con la quantità di forza relativa generato per ciascuna frequenza di stimolazione, e per ogni posizione comune. Come nel nervo PD, una varietà di ampiezze picco sono considerati in risposta alla contrazione del muscolo apri <strong>(Figura 13C).</strong> </p><p class="jove_content"> Disposizione anatomica dei neuroni nella gamba a piedi è chiaramente osservata con colorazione blu di metilene <strong>(Figura 14).</strong> Notare il cordotonali filo e la tensione neurone elastico che si trova vicino al apodeme. Diversi somata con differenti diametri did località specifiche sono visibili anche in questa figura. L&#39;intero corso del nervo tensione e barella nervo motore sono mostrati in <strong>Figura 15.</strong> Neuroni individuali del nervo PD sono mostrati con maggiore contrasto utilizzando 4-Di-2-ASP <strong>(Figura 16)</strong> e CoCl <sub>2</sub> <strong>(Figura 17)</strong> di recupero informazioni tecniche. Ad alto ingrandimento le terminazioni sensoriali possono essere visti all&#39;interno delle scolopales di supporto <strong>(Figure 16B)</strong> <sup>21,22,26.</sup> </p><p class="jove_content" fo:keep-together.within-page="always"><img alt="Figura 10" fo:content-width="5in" src="/files/ftp_upload/51050/51050fig10.jpg" width="500px" /><br /> <strong>Figura 10. Spostare e tenere a 0 °.</strong> I neuroni dinamici sono robusti in cottura durante il movimento e picchi di neuroni sensibili all&#39;elettricità statica sono presenti mentre il comune è tenuto aperto. <a href="https://jove.unicartagenaproxy.elogim.com/files/ftp_upload/51050/51050fig10highres.jpg" target="_blank">Clicca qui per vedere l&#39;immagine ingrandita &lt;</a>/ A&gt;. </p><p class="jove_content" fo:keep-together.within-page="always"><img alt="Figura 11" fo:content-width="5in" src="/files/ftp_upload/51050/51050fig11.jpg" width="500px" /><br /> <strong>Figura 11. Rapid aperto e chiusura da completamente flessa in posizione estesa (90-0 °).</strong> <a href="https://jove.unicartagenaproxy.elogim.com/files/ftp_upload/51050/51050fig11highres.jpg" target="_blank">Clicca qui per vedere l&#39;immagine ingrandita</a> . </p><p class="jove_content" fo:keep-together.within-page="always"><img alt="Figura 12" fo:content-width="5in" src="/files/ftp_upload/51050/51050fig12.jpg" width="500px" /><br /> <strong>Figura 12. Extracellulari picchi registrati dal nervo PD.</strong> Il giunto è completamente estesa e poi rapidamente spostato in una ½ posizione flessa e poi tenendola ferma. Si noti l&#39;attività durante il movimento e la diminuzione dell&#39;attività quando statica. <a href="https://jove.unicartagenaproxy.elogim.com/files/ftp_upload/51050/51050fig12highres.jpg" target="_blank">Clicca qui per ingrandire ima</a>ge. </p><p class="jove_content" fo:keep-together.within-page="always"><img alt="Figura 13" fo:content-width="5in" src="/files/ftp_upload/51050/51050fig13.jpg" width="500px" /><br /> <strong>Figura 13. Le forze relative che si sviluppano con il giunto completamente flesse e stimolati alle varie frequenze. (A)</strong> tracce Voltage-time dal trasduttore di forza sono indicati con ogni frequenza di stimolazione. <strong>(B)</strong> Le tracce nel pannello A si sovrappongono in diversi colori per la facilità di confronto. <strong>(C)</strong> Tensione in tempo le tracce di attività elettrica registrate dal nervo tensione quando il nervo motore è stato stimolato a 80 Hz. Si noti la struttura regolare di manufatti come stimolo rispetto all&#39;attività neurale. Inoltre, notare le diverse ampiezze delle risposte neurali. <a href="https://jove.unicartagenaproxy.elogim.com/files/ftp_upload/51050/51050fig13highres.jpg" target="_blank">Clicca qui per vedere l&#39;immagine ingrandita</a> . </p><p class="jove_content" fo:keep-together.wNEGLI-page = &quot;always&quot;&gt; <img alt="Figura 14" fo:content-width="5in" src="/files/ftp_upload/51050/51050fig14.jpg" width="500px" /><br /> <strong>Figura 14. Blu di metilene macchia della preparazione gamba a piedi.</strong> Somas individuali vengono mostrati con le loro terminazioni sensoriali proiettando nel filo elastico. Vicino alla apodeme un neurone tensione è mostrato. <a href="https://jove.unicartagenaproxy.elogim.com/files/ftp_upload/51050/51050fig14highres.jpg" target="_blank">Clicca qui per vedere l&#39;immagine ingrandita</a> . </p><p class="jove_content" fo:keep-together.within-page="always"><img alt="Figura 15" fo:content-width="5in" src="/files/ftp_upload/51050/51050fig15.jpg" width="500px" /><br /> <strong>Figura 15. Il nervo tensione derivante dalla estremità distale (frecce rosse) e unire il nervo motore (freccia verde).</strong> <a href="https://jove.unicartagenaproxy.elogim.com/files/ftp_upload/51050/51050fig15highres.jpg" target="_blank">Clicca qui per vedere l&#39;immagine ingrandita</a> . </p><p class="jove_content" fo:keep-together.within-page = &quot;always&quot;&gt; <img alt="Figura 16" fo:content-width="5in" src="/files/ftp_upload/51050/51050fig16.jpg" width="500px" /><br /> <strong>Figura 16. (A) Un back-riempimento del nervo PD in <em>Cancer magister</em> con 4-Di-2-ASP. (B)</strong> Maggiore ingrandimento delle terminazioni sensitive. <a href="https://jove.unicartagenaproxy.elogim.com/files/ftp_upload/51050/51050fig16highres.jpg" target="_blank">Clicca qui per vedere l&#39;immagine ingrandita</a> . </p><p class="jove_content" fo:keep-together.within-page="always"><img alt="Figura 17" fo:content-width="5in" src="/files/ftp_upload/51050/51050fig17.jpg" width="500px" /><br /> <strong>Figura 17. I neuroni che sono stati riempiti con CoCl <sub>2</sub> e trattati (A).</strong> Contorno Tracciato della preparazione macchiato mostrato <strong>(B).</strong> <a href="https://jove.unicartagenaproxy.elogim.com/files/ftp_upload/51050/51050fig17highres.jpg" target="_blank">Clicca qui per vedere l&#39;immagine ingrandita</a> . </p><table border="0" cellpadding="0" cellspacing = &quot;0&quot; width = &quot;392&quot;&gt; <colgroup><col /><col /></colgroup><tbody><tr height="21"><td height="21" style="height:21px;width:235px;"> <strong>Salino</strong> </td><td style="width:157px;"> <strong>g / L</strong> </td></tr><tr height="20"><td height="20" style="height:20px;width:235px;"> NaCl </td><td style="width:157px;"> 27.29 </td></tr><tr height="20"><td height="20" style="height:20px;width:235px;"> KCl </td><td style="width:157px;"> 0.81 </td></tr><tr height="26"><td height="26" style="height:26px;width:235px;"> MgSO <sub>4</sub> • 7H <sub>2</sub> O </td><td style="width:157px;"> 4.81 </td></tr><tr height="26"><td height="26" style="height:26px;width:235px;"> CaCl <sub>2</sub> • 2H <sub>2</sub> O </td><td style="width:157px;"> 1.85 </td></tr><tr height="26"><td height="26" style="height:26px;width:235px;"> Na <sub>2</sub> SO <sub>4</sub> • 10H <sub>2</sub> O </td><td style="width:157px;"> 0.97 </td></tr><tr height=&quot;20&quot;&gt; <td height="20" style="height:20px;width:235px;"> Destrosio (D-glucosio) </td><td style="width:157px;"> 1.982 </td></tr><tr height="20"><td height="20" style="height:20px;width:235px;"> L&#39;acido HEPES </td><td style="width:157px;"> 0,476 </td></tr><tr height="20"><td height="20" style="height:20px;width:235px;"> HEPES sale </td><td style="width:157px;"> 2.08 </td></tr><tr height="40"><td height="40" style="height:40px;width:235px;"> Regolare a pH 8.1 con NaOH o HCl </td><td style="width:157px;"></td></tr></tbody></table><p> <strong>Tabella 1. Ricette per granchi salina.</strong> </p>

Watch this Scientific Journal Video about Proprioception and Tension Receptors in Crab Limbs: Student Laboratory Exercises at JoVE.com

Recettori della propriocezione e della tensione negli arti del granchio | Neuroscienze | Video

Proprioception and Tension Receptors in Crab Limbs: Student Laboratory Exercises

<p class="jove_content"> Tecniche fisiologiche e anatomiche sono dimostrati per affrontare funzione e la struttura per propriocettori articolari e recettori tensione muscolare degli arti crostacei piedi. </p>

Propriocezione e la tensione recettori in Crab Arti: Esercizi di laboratorio per studenti

The primary purpose of these procedures is to demonstrate for teaching and research purposes how to record the activity of living primary sensory neurons ...

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25.1K Views.  University of Kentucky.  Tecniche fisiologiche e anatomiche sono dimostrati per affrontare funzione e la struttura per propriocettori articolari e recettori tensione muscolare degli arti crostacei piedi. 

Video: Proprioception and Tension Receptors in Crab Limbs: Student Laboratory Exercises

Lab Techniques

<h4>Measurement Accuracy</h4>
<p>The ability to repeat an experiment and get the same results, or <em>reproducibility</em>, is essential in scientific research. However, it&#8217;s impossible to repeat an experiment if you don&#8217;t know how you did it. Thus, scientists keep detailed records of their experiments in lab notebooks. These records include important information like the amount of each material used and descriptions of each step of the procedure.</p>
<p>Equipment, like balances and volumetric or graduated glassware, is used to measure solid and liquid compounds for experiments. The precision of each measurement is limited by the equipment used. Balances and other equipment with digital readouts automatically display values at the maximum precision that they can support. The precision of graduated glassware and rulers depends on the increment, which is the amount represented by the smallest distance between two marks.</p>
<p>The surface of most liquids curves upwards at the edges in glass containers. This type of curved liquid surface is called a concave meniscus. When the edges of the liquid curve down, it is called a convex meniscus. When measuring liquid volumes, the reference point for the top of the liquid is the bottom of a concave meniscus or the top of a convex meniscus. To accurately identify the position of the meniscus, liquid volumes are read by looking at the liquid&#8217;s surface from the side at eye-level. Looking at the liquid from above or below will cause the liquid surface level to appear higher or lower than it actually is.</p>
<p>When the reference point of the meniscus touches a mark on graduated or volumetric glassware, the glassware is holding the volume defined for that mark. Volumetric glassware is designed to measure a specific volume, so it only has one mark. Graduated glassware is designed to measure a range of volumes, so it has many marks, or <em>graduations</em>, which are labeled at regular intervals. Both graduated and volumetric glassware may have additional manufacturer-provided information about their precision.</p>
<h4>Calibration and Uncertainty</h4>
<p>Manufacturers calibrate equipment to ensure that the measurements are accurate to within a specified range of uncertainty. For example, a high-precision analytical balance may read out to four decimal places (0.0000 g) with an uncertainty of &#177; 0.0001 g. A reading of 0.0345 g indicates that the true value is between 0.0344 g and 0.0346 g.</p>
<p>Glassware typically has its measurement uncertainty printed on it. To accurately record volume measurements, you must consider both the increment and the uncertainty. For example, a precise volume measurement in a 100 mL graduated cylinder with 1 mL increments and an uncertainty of &#177; 0.5 mL would be recorded to the tenths place (000.0 mL &#177; 0.5 mL). If the reference point of the meniscus falls between two marks, you should estimate a value for the tenths place; otherwise, leave it as 0. Since temperature affects volume, the temperature for which the glassware is calibrated will be printed on it.</p>
<p>Glassware is calibrated either <em>to contain</em> (TC) or <em>to deliver</em> (TD). TC glassware holds the specified volume of liquid when it is filled to the mark, but a small amount of liquid will be left behind when it is poured into another container. Solutions are often prepared in TC glassware because the accuracy of the solution volume affects the accuracy of the concentration. Volumetric flasks are typically calibrated to contain.</p>
<p>TD glassware holds a little more than the amount of liquid specified when it is filled to the mark, but it dispenses only the specified volume. Thus, TD glassware should not be emptied completely when dispensing liquids. This type of glassware is useful for transferring a precise volume of liquid to another container. Both graduated and volumetric pipettes are typically calibrated to deliver.</p>
<p>Unless otherwise specified, the uncertainty of a number is assumed to be on the scale of the last digit. Thus, it&#8217;s particularly important to report the uncertainty of a measurement if the uncertainty is on a larger scale. For rulers and graduated glassware without a known uncertainty, the uncertainty is estimated as half of the smallest increment. The uncertain digit is based on where the meniscus falls between two tick marks.</p>
<h4>Significant Figures</h4>
<p>Significant figures are the numbers in a value that are <em>meaningful</em>, or essential for expressing that value with the appropriate precision. All numbers other than leading zeroes (<strong>0.00</strong>1), trailing zeroes (1,<strong>000</strong>), or scientific notation multipliers (<strong>10</strong><sup>x</sup>) are always significant.</p>
<p>Leading zeroes are never significant because they can be removed by rewriting the number either in scientific notation or, if it is an SI unit, in smaller units. For example, there are only three significant figures in the value 0.00123 m because it can be rewritten as either 1.23 &#215; 10<sup>-3</sup> m or 1.23 mm without losing any information. Note that the scaling factor between different multiples of the same SI unit does <em>not</em> affect the number of significant figures.</p>
<p>Trailing zeroes are significant when they are before or after a decimal point. For example, the measurement 100.110 mL has six significant figures and 100.0 mL has four. Trailing zeroes in a number written without a decimal point are assumed to be insignificant unless they are marked otherwise. There are several different conventions for marking significant zeroes in those cases, but to avoid ambiguity it is advisable to rewrite those values in scientific notation. If the measurement is in SI units, use larger units.</p>
<p>The precision of a measurement limits the precision of any values calculated from it. A calculated value has the same number of significant figures as the <em>least</em> precise measurement or value used in its calculation. If the result has too many significant figures, it is rounded to the appropriate precision. If the result has too few significant figures, it is extended with significant trailing zeroes to preserve the precision. The significant figures of experimentally derived values, like molar mass, density, or some unit conversion factors, must also be considered when they are used in calculations.</p>
<p>The number of significant figures in a calculated value is <em>not</em> limited by physical or mathematical constants in formulas or exact numbers, like the number of data points acquired in an experiment. For example, the formula for calculating the volume of a sphere is V = 4/3&#960;r<sup>3</sup>. The number of significant figures in the calculated volume is <em>only</em> affected by the number of significant figures in the radius measurement <strong>r</strong>. 4/3&#8201; and &#960; are constants, and <sup>3</sup> is simply notation for a mathematical operation.</p>
<p>Defined conversion factors are also treated as constants. For instance, the inch is defined as exactly 25.4 millimeters; therefore, the number of significant figures of the converted value depends only on the original measurement. Always check whether the conversion factor is exactly defined or an experimental value when converting between units.</p>
<p>For simple addition and subtraction of measurements, we use significance arithmetic to determine the number of significant figures. In this case, the answer has as many decimal places as the measurement with the fewest decimal places <em>regardless of the number of significant figures</em>. Thus, 15,643.7 mL + 0.613 mL = 15,644.3 mL.</p>
<p>For multi-step calculations or complex equations that you break into smaller parts to solve, you should keep at least one or two insignificant figures on the intermediate values. For example, if you calculate the values of the numerator and denominator of a fraction separately before dividing them, you should retain the insignificant figures of the numerator and denominator when you divide them. This minimizes <em>rounding error</em>, which is the difference from the number you would get if you entered the entire formula into your calculator at once. If you write down these intermediate values, you should note which digits are insignificant.</p>
<h4>References</h4>
<p>Harris, D.C. (2015). <em>Quantitative Chemical Analysis</em>. New York, NY: W.H. Freeman and Company.</p>

Accuracy, Precision, Uncertainty, and Significant Figures - Concept

Tecniche di laboratorio

Measurement Accuracy
The ability to repeat an experiment and get the same results, or reproducibility, is essential in scientific research. However, ...

Didattica

JoVE Lab Manual

Chimica

Extraction

<p><strong>Extraction</strong></p>
<p>Extraction is a common technique used in organic chemistry to isolate a target compound. In the extraction process, a solute is transferred from one phase to another to separate it from unreacted starting materials or impurities. Extraction is also used to facilitate the isolation of a solute from a reaction solvent that is difficult to remove by evaporation, such as a solvent with a high boiling point.<span style="color: rgb(0, 0, 0); font-family: Roboto, RobotoDraft, Helvetica, Arial, sans-serif; font-size: 13px; font-style: normal; font-variant-ligatures: normal; font-variant-caps: normal; font-weight: 400; letter-spacing: normal; orphans: 2; text-align: start; text-indent: 0px; text-transform: none; white-space: pre-wrap; widows: 2; word-spacing: 0px; -webkit-text-stroke-width: 0px; background-color: rgb(255, 255, 255); text-decoration-style: initial; text-decoration-color: initial; display: inline !important; float: none;"> </span>Generally, there are three types of extractions. First, in solid-liquid extraction, the solute is transferred from a solid phase to a liquid phase. In liquid-liquid extraction, a solute is transferred from one liquid to another. In acid-base extraction, a solute is transformed into an ionic compound and transferred from an organic phase to an aqueous phase. A common example of extraction is the brewing coffee or tea. The solid phase, which contains the caffeine, plant flavors, and odors, is extracted by the hot water into the liquid phase.</p>
<p><strong>Liquid-Liquid Extraction</strong></p>
<p>A liquid-liquid extraction either transfers an organic compound that is dissolved in an aqueous phase to an organic solvent, or it is used to transfer unreacted reactants, salts, and other water-soluble impurities to the aqueous phase while leaving the organic compound of interest in the organic phase. Immiscible liquids are liquids that never form a homogenous solution, even when thoroughly mixed. Instead, immiscible liquids separate into different phases, like oil and water.</p>
<p>A liquid-liquid extraction transfers an organic compound that is dissolved in an aqueous phase to an organic solvent. To perform a liquid-liquid extraction, first, the aqueous solution containing the solute is added to a separatory funnel. Then, a non-water-soluble organic solvent is added to the separatory funnel. When the contents of the funnel are mixed well, the organic compound partitions into the organic phase based on its higher solubility in the organic phase than the aqueous phase.</p>
<p>Since the two solvents are immiscible, the two liquids form discrete layers, with the dense liquid on the bottom and the less dense liquid on the top. Once the two phases settle back into two layers, they are separated by opening the stopcock at the bottom of the separatory funnel and allowing one layer to flow out. The liquid that had the solute removed is called the raffinate, while the liquid that gained the solute is called the extract.</p>
<p>The organic compound is partitioned between the two layers based on its solubility in each phase. Equilibrium is reached when the chemical potential of the solute is the same in the two phases. The partition coefficient for a solute, K, is the ratio of the concentration of the sample in the organic layer divided by the concentration in the aqueous phase. The partition coefficient is a constant dependent on both the solute and the pair of solvents used in the extraction. The partition coefficient is an expression of preference of the solute for each of the two solvents. Solutes with a large partition coefficient have a higher tendency to be extracted into the organic solvent layer. Solutes with small partition coefficients prefer to transition into the aqueous phase.</p>
<p>Determining which solvent pair to use for a liquid-liquid extraction is a vital step. Consider the following when choosing which solvents to use: First, the solute must be more soluble in the solvent than in water. Therefore, knowing the partition coefficient of the solute in a potential solvent pair is necessary. Second, the solvent-pair must be immiscible in water and not form a homogeneous solution when mixed. Third, the solvents must be inert and not react with the solute. The solvent should also be volatile so that it can be removed from the solute easily. The water-immiscible organic solvent generally possesses a non-polar or low polarity.</p>
<p>It is also critical to know the densities of the solvents to determine the identities of the top and bottom layers. Most organic liquids have a lower density than water, with the exception of chlorinated organic solvents, and will settle to the bottom of the separatory funnel.</p>
<p><strong>Acid-Base Extraction</strong></p>
<p>Acid-base extraction is a type of liquid-liquid extraction that separates organic compounds based on their acid-base properties. If a solute is an acid or base, its charge changes as the pH is changed. Generally, most organic compounds are neutral, and therefore more soluble in organic solvents than they are in water. However, if the organic compound becomes ionic, then it becomes more soluble in water. This is useful in extracting an organic acid or base compound from an organic phase to an aqueous phase.</p>
<p>Acid-base extraction harnesses this property by transforming the solute into its water-soluble salt form, thereby changing its solubility. The solubility of the organic compound and its salt must be dramatically different in order for the technique to be effective.</p>
<p>For example, consider a mixture containing an organic carboxylic acid, an amine, and a neutral compound. Carboxylic acids consisting of six carbons or more are insoluble in water and entirely soluble in organic solvents. However, their conjugate bases (an ionic compound) are water-soluble and insoluble in organic solvents. An amine consisting of at least seven carbons is insoluble in water but soluble in organic solvents. The conjugate acid of that amine (an ionic compound) is water-soluble and insoluble in organic solvents.</p>
<p>When reacted with a base, the carboxylic acid is neutralized to its salt form. The other compounds in the mixture remain neutral. Once the carboxylic acid is transformed into a salt, it will partition to the aqueous phase, while the neutral compounds remain in the organic phase.</p>
<p>Acid-base extraction is also used to separate two weak acids or two weak bases with a significant difference in their pK<sub>a</sub>. In the case of the acids, the relatively stronger acid, having a small pKa value, is neutralized to a salt using a weak base. The weak base does not efficiently react with the weaker acid, and only the stronger acid is transformed to a salt. Then, the salt is extracted into the aqueous phase during extraction. The process follows similarly for weak bases, where the relatively stronger base is neutralized to a salt using a weak acid.</p>
<p><strong>References</strong></p>
<ol>
	<li style="font-size:17px">Harris, D.C. (2015). <em>Quantitative Chemical Analysis.</em> New York, NY: W.H. Freeman and Company.</li>
</ol>

Extraction: Liquid-Liquid and Acid-Base Extraction - Concept

Estrazione

Extraction
Extraction is a common technique used in organic chemistry to isolate a target compound. In the extraction process, a solute is transferred ...

Scientific Measurement and Lab Skills

<p>Source: Smaa Koraym at Johns Hopkins University, MD, USA</p>
<ol class="note-items">
	<li data-note-item="1" data-parent-collapse="">Preparation of the Laboratory<button class="expand">Expand</button>
	<p><em>Here, we show the laboratory preparation for 10 students working individually, with some excess. Please adjust quantities as needed.</em></p>
	<ul class="lab-items">
		<li data-sub-note-item="1-2">
		<div class="lab-step">To set up for this lab experiment, wear the appropriate personal protective equipment, including a lab coat, chemical splash goggles, and gloves.</div>
		</li>
		<li data-sub-note-item="1-3">
		<div class="lab-step">Prepare a 1.5 x 10<sup>-5</sup> M erythrosin B red dye solution. To prepare 500 mL of this solution, weigh 0.0066 g of erythrosin B on an analytical balance.</div>
		</li>
		<li data-sub-note-item="1-4">
		<div class="lab-step">Transfer the erythrosin B to a 250-mL Erlenmeyer flask. Then, measure 200 mL of deionized water and add it to the flask. Place a magnetic stir bar in the flask, set the flask on a stir plate, and mix the solution until it appears homogeneous.</div>
		</li>
		<li data-sub-note-item="1-5">
		<div class="lab-step">Use a funnel to pour the solution into a 500-mL volumetric flask, and carefully fill the flask to the line using deionized water. Seal the top of the flask with paraffin film and invert it several times until the solution is well-mixed.</div>
		</li>
		<li data-sub-note-item="1-6">
		<div class="lab-step">Label a 500-mL squeeze bottle with the concentration and transfer the solution into the bottle. Store the bottle on a mat in the hood.<strong> Note: </strong>Take significant figures into account.</div>
		</li>
		<li data-sub-note-item="1-7">
		<div class="lab-step">Place acetone in an area so that students have access to it during the lab. <strong>Note: </strong>Each student will need about 40 mL.</div>
		</li>
		<li data-sub-note-item="1-8">
		<div class="lab-step">Set weigh boats, scoops, and a container of sand next to a balance. <strong>Note: </strong>Each student will need about 3 g.</div>
		</li>
		<li data-sub-note-item="1-9">
		<div class="lab-step">Place an organic waste container in a central hood.</div>
		</li>
		<li data-sub-note-item="1-10">
		<div class="lab-step">Set out the following glassware and equipment at each student lab station (we suggest students work independently during this lab):
		<table align="center" style="border-collapse: collapse; font-size: 17px; line-height: 24px;">
			<tbody>
				<tr>
					<td style="border: 1px solid; padding: 0px; ">&#160;1&#160;&#160;&#160;&#160;50-mL beaker</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px; ">&#160;1&#160;&#160;&#160;&#160;100-mL beaker</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px; ">&#160;1&#160;&#160;&#160;&#160;250-mL beaker</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px; ">&#160;1&#160;&#160;&#160;&#160;400-mL beaker</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px; ">&#160;1&#160;&#160;&#160;&#160;600-mL beaker</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px; ">&#160;1&#160;&#160;&#160;&#160;500-mL filter flask</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px; ">&#160;1&#160;&#160;&#160;&#160;B&#252;chner funnel</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px; ">&#160;1&#160;&#160;&#160;&#160;10-mL graduated cylinder</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px; ">&#160;1&#160;&#160;&#160;&#160;Watch glass</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px; ">&#160;1&#160;&#160;&#160;&#160;5-mL volumetric pipette</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px; ">&#160;1&#160;&#160;&#160;&#160;10-mL volumetric pipette</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px; ">&#160;1&#160;&#160;&#160;&#160;10-mL capacity pipetter</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px; ">&#160;1&#160;&#160;&#160;&#160;10-mL volumetric flask</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px; ">&#160;1&#160;&#160;&#160;&#160;String (18 in)</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px; ">&#160;1&#160;&#160;&#160;&#160;Silicone vacuum tubing (18 in)</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px; ">&#160;1&#160;&#160;&#160;&#160;12-in ruler</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px; ">&#160;1&#160;&#160;&#160;&#160;Rubber adapter</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px; ">&#160;1&#160;&#160;&#160;&#160;Rubber stopper</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px; ">&#160;1&#160;&#160;&#160;&#160;Glass stirring rod</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px; ">&#160;1&#160;&#160;&#160;&#160;Rubber policeman</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px; ">&#160;1&#160;&#160;&#160;&#160;Disposable pipette</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px; ">&#160;1&#160;&#160;&#160;&#160;Bottle of deionized water</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px; ">&#160;1&#160;&#160;&#160;&#160;Lab mat</td>
				</tr>
				<tr>
					<td style="border: 1px solid; padding: 0px; ">&#160;1&#160;&#160;&#160;&#160;Box of filter papers (1 filter paper per student)</td>
				</tr>
			</tbody>
		</table>
		</div>
		</li>
	</ul>
	</li>
</ol>

Two-Dimensional Graphs, Curve Fitting, and Standard Deviation - Prep

Misure scientifiche e competenze di laboratorio

Source: Smaa Koraym at Johns Hopkins University, MD, USA

	Preparation of the LaboratoryExpand
	Here, we show the laboratory preparation for 10 students ...

Lab Manual Sub Articles

Energy Dynamics

<ol class="note-items"><li data-note-item="1" data-parent-collapse="">Caterpillar Energy Dynamics
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="1-1"><div class="lab-step">NOTE: In this lab you will grow caterpillars in conditions with and without food to examine the transfer of biomass in a typical trophic system.
</div></li><li data-sub-note-item="1-2"><div class="lab-step">Weigh and record the wet mass of enough leaves of cabbage to line the bottom of a plastic tub.
</div></li><li data-sub-note-item="1-3"><div class="lab-step">Then, collect two batches of five caterpillars from their cabbage habitat, and weigh and record the wet mass of each batch separately in individual weigh boats. Make sure to tare the balance before each measurement. HYPOTHESES: The experimental hypothesis is that only a fraction of energy will be transferred between the trophic levels of the cabbage and the caterpillars and a significant amount will be lost as waste and heat energy. The null hypothesis is that there will be no significant difference between the energy input from the cabbage, and the obtained energy stored as biomass in the caterpillars.
</div></li><li data-sub-note-item="1-4"><div class="lab-step">Place the weighed cabbage and five caterpillars in a plastic tub lined with damp paper towels, leaving the lid slightly cracked open for ventilation, but not wide enough for the animals to escape.
</div></li><li data-sub-note-item="1-5"><div class="lab-step">Place the other five caterpillars in a second clean container prepared the same way, but with no cabbage.
</div></li><li data-sub-note-item="1-6"><div class="lab-step">After three days, transfer the caterpillars to new weigh boats, and weigh and record the final wet mass of both groups of caterpillars and the cabbage.
</div></li><li data-sub-note-item="1-7"><div class="lab-step">Then place the caterpillar waste, or frass, and cabbage into two separate dishes, and put both dishes into a 37 &#176;C incubator to dry for 48 hours.
</div></li><li data-sub-note-item="1-8"><div class="lab-step">Euthanize the weighed caterpillars by putting them into a minus 20 &#176;C freezer for 24 hours.
</div></li><li data-sub-note-item="1-9"><div class="lab-step">Then, transfer the dishes to a 37 &#176;C incubator for 48 hours.
</div></li><li data-sub-note-item="1-10"><div class="lab-step">After drying, weigh and record the dry mass of the uneaten cabbage, the two groups of caterpillars, and the frass, and use these data to calculate the percent biomass of the caterpillars.
</div></li><li data-sub-note-item="1-11"><div class="lab-step">Finally, discard all of the dried organisms and plates into the hazardous waste bin, and wipe down your workspace with 70% ethanol.
</div></li></ul></li><li data-note-item="2" data-parent-collapse="">Results
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="2-1"><div class="lab-step">Take a look at the data you collected. Ensure the initial wet mass, the final wet mass, and the final dry mass columns are accurately filled out in your table. <a href="https://jove.unicartagenaproxy.elogim.com/CDNSource/lm/labs/13/Table_1_Energy_Dynamics.xlsx" style="display:block;margin:20px;" target="__blank">Click Here to download Table 1</a>
</div></li><li data-sub-note-item="2-2"><div class="lab-step">NOTE: Remember an object&#39;s biomass is equal to its dry mass. Therefore, percent biomass for each group is the final dry mass divided by the final wet mass times 100. For example, obtain the percent biomass of the cabbage by dividing the final dry mass by the final wet mass and multiplying by 100. This means that approximately 15% of the cabbage&#39;s total weight is biomass.
</div></li><li data-sub-note-item="2-3"><div class="lab-step">After this, estimate the initial biomass of the cabbage and caterpillars by multiplying the percent biomass by the initial wet mass, and put the values in this new table. NOTE: Remember, biomass equals dry mass. Therefore, the final dry mass is equal to the final biomass of each group.
</div></li><li data-sub-note-item="2-4"><div class="lab-step">To calculate the approximate initial and final energy in the form of kilocalories within the cabbage, caterpillars, and frass, multiply the biomass values recorded in your table by the conversions, and record the calculated values in the table. <a href="https://jove.unicartagenaproxy.elogim.com/CDNSource/lm/labs/13/Table_2_Energy_Dynamics.xlsx" style="display:block;margin:20px;" target="__blank">Click Here to download Table 2</a>
</div></li><li data-sub-note-item="2-5"><div class="lab-step">After this, subtract the initial energy from the final energy to obtain the change in energy for each group.
</div></li><li data-sub-note-item="2-6"><div class="lab-step">NOTE: The negative sign denotes loss in energy, and positive indicates gain.
</div></li><li data-sub-note-item="2-7"><div class="lab-step">Using your data table, calculate the energy in kilocalories that the fed caterpillars lost to respiration. The formula is the energy gained from cabbage, which is the positive of the energy lost by cabbage, minus the biomass gained from cabbage, minus the energy lost via frass. <a href="https://jove.unicartagenaproxy.elogim.com/CDNSource/lm/labs/13/Table_3_Energy_Dynamics.xlsx" style="display:block;margin:20px;" target="__blank">Click Here to download Table 3</a>
</div></li><li data-sub-note-item="2-8"><div class="lab-step">Then, repeat the same calculation for the unfed group of caterpillars. Make another column in the table and note this down.
</div></li><li data-sub-note-item="2-9"><div class="lab-step">Finally, add up all of the energy gained via biomass and cabbage consumption, and the energy lost via frass and respiration for each group of caterpillars. NOTE: For both groups of caterpillars the net energy is zero, meaning that this experiment obeyed the first law of thermodynamics.
</div></li></ul></li></ol>

Studying Energy Dynamics and Productivity - Procedure

Dinamica dell'energia

Caterpillar Energy Dynamics
ExpandNOTE: In this lab you will grow caterpillars in conditions with and without food to examine the transfer of biomass in a ...

Biologia

Ecologia

Animal Behavior

<ol class="note-items"><li data-note-item="1" data-parent-collapse="">Making Choice Chambers
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="1-1"><div class="lab-step">Before beginning the experiments, use one of the Sharpies from the lab table to trace a line around the circumference of the bottom of both bottles. One line should be 1 cm from the base of the bottle and the other should be 1.5 cm from the base of the bottle.
</div></li><li data-sub-note-item="1-2"><div class="lab-step">Use a razor blade to make a small cut in each bottle at the mark. 
</div></li><li data-sub-note-item="1-3"><div class="lab-step">Then, insert the scissors and carefully cut along the line until the bottoms of the bottles are removed completely.
</div></li><li data-sub-note-item="1-4"><div class="lab-step">Fit one bottle bottom into the other.
</div></li><li data-sub-note-item="1-5"><div class="lab-step">Tape the bottles together to make a secure chamber.
</div></li><li data-sub-note-item="1-6"><div class="lab-step">Use the razor blade and scissors to poke a hole into the center of the chamber. 
</div></li><li data-sub-note-item="1-7"><div class="lab-step">Take a piece of tape and fold the edge onto itself to make a tab and place the tape over the hole.  Note: The tab helps with easy removal and reattachment of the tape to the chamber.
</div></li><li data-sub-note-item="1-8"><div class="lab-step">Use the Sharpie to label one side of the chamber A and the other side B.
</div></li><li data-sub-note-item="1-9"><div class="lab-step">Upon receiving the colony of <em>Drosophila</em>, perform a count to make sure there are 40 living individuals in the vial.
</div></li></ul></li><li data-note-item="2" data-parent-collapse="">Geotaxis
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="2-1"><div class="lab-step">To perform a geotaxis assay, first attach a clean, empty bottle cap to each end of the choice chamber and turn the fly vial upside down onto its lid.
</div></li><li data-sub-note-item="2-2"><div class="lab-step">Open the tape flap in the center of the choice chamber.
</div></li><li data-sub-note-item="2-3"><div class="lab-step">Insert the funnel into the hole and tap all of the flies into the chamber. HYPOTHESES: The alternate hypothesis is that more flies will be moving against gravity, or up, regardless of the orientation of the choice chamber. Alternatively, if the flies are positively geotactic, then more flies will move down, or with gravity. The null hypothesis is that the flies will have no preference with regard to gravity and will move randomly within the chamber. 
</div></li><li data-sub-note-item="2-4"><div class="lab-step">Once the <em>Drosophila</em> are in the chamber, close the tape flap, rotate the chamber so that it stands vertically with side A on the bottom, and start the timer.
</div></li><li data-sub-note-item="2-5"><div class="lab-step">Immediately, count the number of <em>Drosophila</em> that are moving up and the number that are moving down. Record these data for time point 0 in the table. 
</div></li><li data-sub-note-item="2-6"><div class="lab-step">After 30 seconds, rotate the chamber so that side B is on the bottom and count the number of <em>Drosophila</em> that are moving up or down. Record these data for time point 30 in the table.
</div></li><li data-sub-note-item="2-7"><div class="lab-step">After 60 seconds, rotate the chamber so that side A is on the bottom again and record the number of moving flies for time point 60.  <a href="https://jove.unicartagenaproxy.elogim.com/CDNSource/lm/labs/15/Table_1_Animal_behavior.xlsx" style="display:block;margin:20px;" target="__blank">Click Here to download Table 1</a>
</div></li><li data-sub-note-item="2-8"><div class="lab-step">Hold the chamber over the fly vial and carefully open the cap to return all of the <em>Drosophila</em> back to their empty vial. 
</div></li></ul></li><li data-note-item="3" data-parent-collapse="">Phototaxis
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="3-1"><div class="lab-step">To perform a phototaxis analysis, first cover side A of the choice chamber with tin foil, taking care that the foil reaches to the exact center between the two sides. 
</div></li><li data-sub-note-item="3-2"><div class="lab-step">Ensure the caps are secured on both ends of the chamber.  HYPOTHESES: In this exercise, the alternative hypothesis is that more flies will be found in the light side of the choice chamber, if the flues are attracted to light, or on the dark side if they are repelled by light. The null hypothesis is that <em>Drosophila</em> will have no preference with regard to either stimulus. Therefore, equal numbers of flies will be found on either side of the choice chamber.
</div></li><li data-sub-note-item="3-3"><div class="lab-step">Open the tape flap in the center of the choice chamber, insert the funnel into the hole, and tap all 40 flies into the chamber as previously described.
</div></li><li data-sub-note-item="3-4"><div class="lab-step">Set the timer to sound an alarm at 10 minutes.
</div></li><li data-sub-note-item="3-5"><div class="lab-step">When the alarm sounds, stop the timer, and count the number of flies in the open side B of the chamber.
</div></li><li data-sub-note-item="3-6"><div class="lab-step">Subtract that number from the total number of flies added to the choice chamber to calculate the number of flies in the covered side A.
</div></li><li data-sub-note-item="3-7"><div class="lab-step">Record these data in the table for time point 10.  <a href="https://jove.unicartagenaproxy.elogim.com/CDNSource/lm/labs/15/Table_2_Animal_behavior.xlsx" style="display:block;margin:20px;" target="__blank">Click Here to download Table 2</a>
</div></li><li data-sub-note-item="3-8"><div class="lab-step">Carefully, return all of the flies back to their empty vial.
</div></li></ul></li><li data-note-item="4" data-parent-collapse="">Chemotaxis
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="4-1"><div class="lab-step">HYPOTHESES: In this exercise, the alternative hypothesis is that more flies will be found on the side of the chamber with the apple cider vinegar if the flies are attracted to the odor. Or, if the flies are repelled by the vinegar, then more of them will be found on the side of the chamber presenting the odorless water. The null hypothesis is that <em>Drosophila</em> will have no preference with regard to either stimulus and will move randomly in the chamber.  Use an individual disposable plastic pipette to dispense two drops of the apple cider vinegar onto one cotton ball for side A of the chamber. 
</div></li><li data-sub-note-item="4-2"><div class="lab-step">Use an individual disposable plastic pipette to dispense two drops of the distilled water onto a cotton ball for side B of the chamber. 
</div></li><li data-sub-note-item="4-3"><div class="lab-step">Fold two pieces of tape over themselves to create individual pieces of double-sided tape and use the tape to attach each cotton ball to the inside of the single bottle cap.
</div></li><li data-sub-note-item="4-4"><div class="lab-step">Screw the bottle caps with the cotton balls onto their corresponding sides of the chamber.
</div></li><li data-sub-note-item="4-5"><div class="lab-step">Transfer all of the flies into the chamber and set the timer to sound an alarm after 10 minutes. 
</div></li><li data-sub-note-item="4-6"><div class="lab-step">When the alarm sounds, stop the timer and record the number of flies in each side of the chamber. <a href="https://jove.unicartagenaproxy.elogim.com/CDNSource/lm/labs/15/Table_3_Animal_behavior.xlsx" style="display:block;margin:20px;" target="__blank">Click Here to download Table 3</a>
</div></li><li data-sub-note-item="4-7"><div class="lab-step">At the end of the final experiment, funnel the flies into a morgue tube containing 70% ethanol. 
</div></li><li data-sub-note-item="4-8"><div class="lab-step">Dismantle the choice chambers for recycling.  NOTE: Since all of the chemicals used in these experiments are lab safe, the liquids can be disposed of down the sink, the chambers rinsed, and the solids can be thrown away in the trash. 
</div></li></ul></li><li data-note-item="5" data-parent-collapse="">Results
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="5-1"><div class="lab-step">Gather all of the data collected for the geotaxis, phototaxis, and chemotaxis experiments.
</div></li><li data-sub-note-item="5-2"><div class="lab-step">Plot these values as bar graphs for different conditions and different time points.
</div></li><li data-sub-note-item="5-3"><div class="lab-step">Use these data to draw inferences about the behavior of <em>drosophila melanogaster</em>. 
</div></li></ul></li></ol>

Animal Behavior: Studying Taxis Behavior of Drosophila - Procedure

Comportamento animale

Making Choice Chambers
ExpandBefore beginning the experiments, use one of the Sharpies from the lab table to trace a line around the circumference of the ...

Natural Selection

<ol class="note-items"><li data-note-item="1" data-parent-collapse="">Simulating Natural Selection
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="1-1"><div class="lab-step">To prepare for the lab, set out workstations for each student pair containing 6 white, 6 black, and 13 gray 12 inch pipe cleaners.
</div></li><li data-sub-note-item="1-2"><div class="lab-step">Then, add 62 pipe cleaners in any color but black, white or gray, a roll of tape, and a pair of scissors.
</div></li><li data-sub-note-item="1-3"><div class="lab-step">Also, place one piece of black construction paper and one piece of white construction paper at each station.
</div></li><li data-sub-note-item="1-4"><div class="lab-step">Finally, print out tables so that there are enough for each set of partners in the class.
</div></li><li data-sub-note-item="1-5"><div class="lab-step">To save materials for subsequent labs, inform students to save the materials and to avoid damaging them as much as possible.
</div></li></ul></li></ol>

Natural Selection: Calculating Fitness and Selection - Prep

Selezione naturale

Simulating Natural Selection
ExpandTo prepare for the lab, set out workstations for each student pair containing 6 white, 6 black, and 13 gray 12 inch ...

Evoluzione

Scientific Method

<ol class="note-items"><li data-note-item="1" data-parent-collapse="">Demonstrating the Scientific Method with Tree Branches
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="1-1"><div class="lab-step">Obtain branches containing a few dozen typical leaves from five different local tree species. The branches should represent a range of leaf and tree types. 
</div></li><li data-sub-note-item="1-2"><div class="lab-step">Label each branch and leaf with their species name. 
</div></li><li data-sub-note-item="1-3"><div class="lab-step">Distribute the branches to the students, so that each student has access to all five species. 
</div></li><li data-sub-note-item="1-4"><div class="lab-step">Provide some background information about the environment from which each branch was collected. 
</div></li></ul></li></ol>

Scientific Method: Steps and Applications - Prep

Metodo scientifico

Demonstrating the Scientific Method with Tree Branches
ExpandObtain branches containing a few dozen typical leaves from five different local tree species. ...

Fondamenti

Measuring Biodiversity

<ol class="note-items"><li data-note-item="1" data-parent-collapse="">Quadrats and Transects Preparation
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="1-1"><div class="lab-step">Start by tying a knot at the beginning, end and every 5 feet of six 20-foot pieces of rope.
</div></li><li data-sub-note-item="1-2"><div class="lab-step">Then, identify three outdoor areas near the classroom, each containing a variety of at least four plant species.  NOTE: Surprisingly, grassy areas often contain at least four plant species. Although most plants in these areas will be of a single grass species, there are usually many weeds, such as clovers or dandelions mixed in. Therefore, a well-kept grassy area without shade, a well-kept grassy area with shade, for example under a tree, and a weedy field could be used for the three different habitats that they students will be moving between.
</div></li><li data-sub-note-item="1-3"><div class="lab-step">Place two ropes parallel to each other within each of the habitats. It does not matter which direction the ropes face as long as the entire length of rope is within each area and a hula hoop can be placed on either side of the ropes within the habitat.
</div></li><li data-sub-note-item="1-4"><div class="lab-step">Finally, back in the lab set out enough plant identification guides for each student group to have at least one.
</div></li><li data-sub-note-item="1-5"><div class="lab-step">Also set out enough hula hoops so that each group has one to use as a sampling quadrat.
</div></li></ul></li><li data-note-item="2" data-parent-collapse="">Diversity Lab Simulation Preparation
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="2-1"><div class="lab-step">Before beginning the species accumulation curves and richness evenness activity, place 50 colored beads into one paper bag. The bag should contain 15 black beads, 10 purple beads, 8 red beads, 5 green beads, 3 blue beads, 3 yellow beads, 2 pink beads, 2 brown beads, 1 orange bead and 1 white bead.
</div></li><li data-sub-note-item="2-2"><div class="lab-step">Label this bag Community A.
</div></li><li data-sub-note-item="2-3"><div class="lab-step">Then, add five beads of each color to the second bag and mark this bag Community B.
</div></li></ul></li></ol>

Misurare la biodiversità

Quadrats and Transects Preparation
ExpandStart by tying a knot at the beginning, end and every 5 feet of six 20-foot pieces of rope.
Then, identify three ...

Animal Diversity

<ol class="note-items"><li data-note-item="1" data-parent-collapse="">Form and Function of Crickets and Crayfish
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="1-1"><div class="lab-step">This lab involves dissection of both crickets and crayfish. You will need one cricket and one crayfish for each pair of students. IMPORTANT: Purchase the largest crickets available and place them in the freezer the night before.
</div></li><li data-sub-note-item="1-2"><div class="lab-step">On the day of the lab if they are frozen, allow the crickets to thaw for at least three hours before the dissections begin.
</div></li><li data-sub-note-item="1-3"><div class="lab-step">While waiting for the crickets to thaw, melt dissecting wax in a microwave or on a hotplate until it is completely liquefied.
</div></li><li data-sub-note-item="1-4"><div class="lab-step">Fill enough Petri dishes to have one per student pair to around halfway. Allow these plates to harden at room temperature.
</div></li><li data-sub-note-item="1-5"><div class="lab-step">Next set out a dissecting tray, 12 pins, scissors and a needle probe for each pair of students.
</div></li><li data-sub-note-item="1-6"><div class="lab-step">Also set up a few dissecting microscopes around the room and turn them on. NOTE: Generally, one microscope for four students is sufficient.
</div></li><li data-sub-note-item="1-7"><div class="lab-step">Then, set out a pitcher or beaker of water for students to use during the cricket dissection.
</div></li></ul></li></ol>

Animal Diversity: Form and Function of Crickets and Crayfish - Prep

Diversità animale

Form and Function of Crickets and Crayfish
ExpandThis lab involves dissection of both crickets and crayfish. You will need one cricket and one crayfish ...

Evolutionary Relationships

<ol class="note-items"><li data-note-item="1" data-parent-collapse="">Forming an Evolutionary Hypothesis
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="1-1"><div class="lab-step">NOTE: A cladogram is an important tool for forming an evolutionary hypothesis. A cladogram is a tree-shaped chart used to depict the hypothetical genealogical relationships between species. The tips or leaves of the chart represent specific species and the branches of the tree are different lengths. The different lengths represent the degree of change between each of the species. The common ancestor of all of the species that a specific line branches from is located at a node, where the branches intersect. Species that share a node are called a sister group.
</div></li><li data-sub-note-item="1-2"><div class="lab-step">To begin, look at the provided blank cladogram and make a hypothesis about where the assigned animals should be placed on the tree. If a blank cladogram has not been provided please <a target="_blank" href="/files/ftp_upload/10562/Blank_Cladogram.png">download one here</a>.
</div></li><li data-sub-note-item="1-3"><div class="lab-step">Next, observe the image of the fossilized animal.
</div></li><li data-sub-note-item="1-4"><div class="lab-step">Using the morphology, or physical characteristics, of the fossil, make a hypothesis about where on your cladogram it should be placed. Add a new line or mark to indicate your proposed position for the fossil.
</div></li></ul></li><li data-note-item="2" data-parent-collapse="">Comparing Gene Sequences Using BLAST
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="2-1"><div class="lab-step">NOTE: Historically, cladograms have been constructed through the comparison of morphology. Now, genetic sequences can also be compared between species of interest to construct cladograms. DNA sequences are not usually preserved in fossils, so for this lab you will use the BLAST database to compare the DNA sequences of living species that are closely related to the fossil to thousands of other living species.
</div></li><li data-sub-note-item="2-2"><div class="lab-step">Before beginning this lab, your instructor will have downloaded the three preserved gene sequences to a folder on your desktop.
</div></li><li data-sub-note-item="2-3"><div class="lab-step">Navigate to the BLAST homepage, which the instructor should have left open in your browser, and click on Saved Strategies from the menu at the top of the page.
</div></li><li data-sub-note-item="2-4"><div class="lab-step">Click Open on the Evolutionary_Relationships_Gene1.txt file.
</div></li><li data-sub-note-item="2-5"><div class="lab-step">Then, click View to go to the Standard Nucleotide BLAST search page. You should be able to see the DNA sequence in the Query Sequence box at the top-right of the screen.
</div></li><li data-sub-note-item="2-6"><div class="lab-step">Scroll to the bottom of the screen and click the BLAST button. Note: It will take a few moments to process and analyze the data. 
</div></li><li data-sub-note-item="2-7"><div class="lab-step">When the data has been processed, a graphic summary will appear.  NOTE: Your query sequence is represented by the blue line at the top of the box. Each of the lines below the blue line represents another sequence in the BLAST database that matches your query. The length and location of these bars represent where and how much of the sequence match up to the query. The color of the bar represents its score, or how identical the sequence is to the query. Below the graphic summary is the Sequences producing significant alignments list, which contains descriptions of the DNA sequences retrieved from the database that most closely align with the query sentence. These are listed in descending order, from the most similar at the top to the least similar at the bottom. On the right, there are several statistics on how closely related each database sequence is to the experimental sequence. The higher the Max, Total, Query Coverage, and Identity scores are, the more similar the query and retrieved sequences are to each other. Similarly, the lower the E value number, the less likely the sequences match was found by random chance and more significantly the alignment is accurate. An E value of 0 indicates a highly significant match.
</div></li><li data-sub-note-item="2-8"><div class="lab-step">Click on the name of the description of the most similar alignment listed. This will take you further down the page to the exact DNA sequence alignment between the retrieved sequence and the query sequence.
</div></li><li data-sub-note-item="2-9"><div class="lab-step">Click on the Sequence ID number. This will open a new tab with more specific information on the retrieved gene sequence.
</div></li><li data-sub-note-item="2-10"><div class="lab-step">Identify and record the scientific and common names of the organism, which should be listed next to Source.
</div></li><li data-sub-note-item="2-11"><div class="lab-step">Identify and record the protein that the gene codes for, which should be listed next to Definition.
</div></li><li data-sub-note-item="2-12"><div class="lab-step">Close the tab and hit Back to return to the Sequences producing significant alignments list.
</div></li><li data-sub-note-item="2-13"><div class="lab-step">Then, repeat steps 7 &#8211; 9 for the next most similar sequences.
</div></li><li data-sub-note-item="2-14"><div class="lab-step">After collecting several species names, return to the Sequences producing significant alignments list and click Select: All to check all of the boxes next to the names of each listed alignment sequence.
</div></li><li data-sub-note-item="2-15"><div class="lab-step">Click on Distance of tree results to create a phylogram based on the similarity of all of the BLAST result gene sequences to the query gene sequence.  Note: Your query fossil relative sequence will be highlighted in yellow.
</div></li><li data-sub-note-item="2-16"><div class="lab-step">Record your sequence&#8217;s location in relation to the other taxon groups shown in the tree.
</div></li><li data-sub-note-item="2-17"><div class="lab-step">Finally, return to the Saved Strategies tab and upload the next saved gene sequence.
</div></li><li data-sub-note-item="2-18"><div class="lab-step">Query the remaining two fossil relative DNA sequences as previously demonstrated (steps 1 &#8211; 16).
</div></li></ul></li><li data-note-item="3" data-parent-collapse="">Data Analysis
<button class="expand">Expand</button><ul class="lab-items"><li data-sub-note-item="3-1"><div class="lab-step">Based on your gene sequence results, come up with a second hypothesis about where your fossil specimen fit in the original cladogram.
</div></li><li data-sub-note-item="3-2"><div class="lab-step">Compare this placement with your first hypothesis.
</div></li><li data-sub-note-item="3-3"><div class="lab-step">Compare your tree with your classmates&#8217;. If any of your classmates put the fossil on a different branch, ask them how they came to that decision.
</div></li></ul></li></ol>

Evolutionary Relationships: Using BLAST to Test Evolutionary Hypotheses - Procedure

Relazioni evolutive

Forming an Evolutionary Hypothesis
ExpandNOTE: A cladogram is an important tool for forming an evolutionary hypothesis. A cladogram is a tree-shaped chart ...