This material is based on the works supported by Nanyang Assistant Professorship (NAP, M4080740), Agency for Science, Technology and Research (A*STAR) Public Sector Research Funding (PSF, M4070190), A*STAR-JST (The Japan Science and Technology Agency) joint grant (M4070198), and Singapore Ministry of Education (MOE2013-T2-2-049). The authors would like to thank Mr. Roger Tan Kay Chia, Prof. Low Kin Huat, Mr. Poon Kee Chun, Mr. Chew Hock See, Mr. Lam Kim Kheong and Dr. Mao Shixin at School of MAE for their support in setting up and maintaining the research facilities. The authors thank Prof. Michel Maharbiz (U.C. Berkeley) his advice and discussion, Prof. Kris Pister and his group (U.C. Berkeley) for their support in providing the GINA used in this study.

1. Animal Studio <ol><li> Posteriori singoli coleotteri torquata Mecynorrhina (6 cm, 8 g) in contenitori di plastica separati con biancheria pellet. </li><li> Alimentare ogni coleottero una tazza di gelatina di zucchero (12 ml) ogni 3 giorni. </li><li> Mantenere la temperatura e l&#39;umidità della stanza allevamento a 25 ° C e 60%, rispettivamente. </li><li> Testare la capacità di volo di ogni coleottero prima di impiantare elettrodi a filo sottile. <ol><li> gettare delicatamente un coleottero in aria. Se lo scarafaggio può volare per più di 10 secondi per 5 prove consecutive, concludere che il coleottero ha capacità di volo regolari e impiegare per esperimenti di volo successivo. Per riprendere il coleottero, spegnere tutte le luci in sala per renderlo scuro. Questo fa sì che il coleottero di interrompere volo. Nota: Un coleottero inizia spontaneamente a volare via quando rilasciato nell&#39;aria. E &#39;meglio condurre gli esperimenti di volo in una grande camera chiusa come quella mostrata in Figura 1 (16 x 8 x 4 m <sup&gt; 3 con uno spazio di motion capture di 12,5 x 8 x 4 m 3), un coleottero volanti si muove molto velocemente (circa 3-5 m / sec) e attira grandi archi quando si gira in aria. </li></ol></li></ol> 2. impianto di elettrodi <ol><li> Anestetizzare il coleottero ponendolo in un contenitore di plastica pieno di CO 2 per 1 min 13,16,20-24. </li><li> Ammorbidire impronte dentarie immergendolo in acqua calda per 10 secondi. Posizionare il coleottero anestetizzato su un blocco di legno e immobilizzarlo con la cera dentale ammorbidita. La cera dentale raffredda naturalmente e solidifica in pochi minuti. </li><li> Tagliare i fili d&#39;argento isolati (127 micron di diametro nuda, 178 micron di diametro, quando rivestito con perfluoroalkoxy) in lunghezze di 25 mm da utilizzare come elettrodi a filo sottile per l&#39;impianto. </li><li> Esporre 3 mm di argento nuda sfiammando l&#39;isolatore a entrambe le estremità di ogni filo. </li><li> Sezionare la superficie superiore della cuticola del coleottero utilizzando una forbice a punta fine di creare un small finestra di circa 4 x 4 mm sulla metepisternum (figura 2c). Nota: Un cuticola morbida color marrone viene quindi esposto, come illustrato nelle figure 2c - e. Il muscolo 3AX si trova sotto la cuticola morbida. </li><li> Pierce due fori sulla cuticola marrone esposto usando un perno insetto (dimensione 00) con una distanza di 2 mm tra i due fori (figura 2d). </li><li> Inserire due elettrodi a filo (di cui uno elettrodi attivi e uno di ritorno previste al punto 2.4) con attenzione attraverso i fori e impiantare in ogni muscolo 3AX ad una profondità di 3 mm. </li><li> Fissare gli elettrodi impiantati e tenerli in atto per evitare il contatto e cortocircuiti facendo cadere la cera d&#39;api fusa sui fori. Se necessario, ridisporre la cera d&#39;api sopra la cuticola toccando la cera con la punta di un saldatore. La cera d&#39;api solidifica rapidamente e rafforza l&#39;impianto. Nota: Per verificare se l&#39;impianto è corretta, elitre della barbabietolale può essere sollevato per osservare il movimento del muscolo 3AX durante la stimolazione elettrica. </li></ol> 3. Montaggio zaino senza fili Nota: Lo zaino consisteva in un armadio a microcontrollore radio una stratificata FR-4 scheda 4 (1,6 x 1,6 cm 2). Lo zaino è stato guidato da un microbattery ai polimeri di litio (3,7 V, 350 mg, 10 mAh). La massa totale dello zaino compresa la batteria era 1,2 ± 0,26 g, che è inferiore alla capacità di carico utile del coleottero (30% di 10 g di peso corporeo). Lo zaino è stato pre-programmato per ricevere le comunicazioni wireless e aveva due canali di uscita. <ol><li> Pulire la superficie pronoto (rimuovere lo strato di cera sulla cuticola) usando del nastro biadesivo. Quindi, fissare lo zaino sul pronoto del coleottero con un pezzo di nastro biadesivo. </li><li> Collegare le estremità degli elettrodi impiantati alle uscite dello zaino. </li><li> Avvolgere nastro retro-riflettente intorno al microbattery per produrre un marcatore FOr telecamere motion capture per rilevare. </li><li> Fissare la microbattery all&#39;inizio dello zaino con un pezzo di nastro biadesivo in modo che il nastro retroriflettente può essere rilevato da telecamere motion capture. </li></ol> 4. Wireless Control System Nota: In questo caso, il sistema di controllo senza fili termine comprende un ricevitore per il telecomando, un computer portatile per eseguire il software di controllo di volo personalizzato, una stazione base, lo zaino, e il sistema di cattura di movimento. <ol><li> Collegare la stazione base e il ricevitore del telecomando al computer portatile tramite porte USB. </li><li> Accendere il sistema di motion capture e collegarlo al computer portatile tramite una porta Ethernet. </li><li> Eseguire la taratura del volume agitando la bacchetta di calibrazione (fornito dalla società fornitore del sistema di motion capture) per coprire completamente lo spazio motion capture. <ol><li> Aprire il software di motion capture dal desktop del portatile. Fare clic e drAG per selezionare tutte le telecamere sul menu &quot;Sistema&quot; del pannello &quot;Risorse&quot;. </li><li> Fare clic sul menu &quot;Prospettiva 3D&quot; e selezionare &quot;Camera&quot; per passare alla visualizzazione della telecamera. Fare clic sulla scheda &quot;Camera&quot; sul pannello &quot;Strumenti&quot; per mostrare l&#39;impostazione di calibrazione. Fai clic su &quot;Start&quot; sul menu &quot;creare maschere telecamera&quot; per eliminare il rumore dalle telecamere e poi &quot;Stop&quot; dopo il rumore è mascherato in blu. </li><li> Fare clic e selezionare &quot;5 Marker Wand &amp; L-Frame&quot; dal menu &quot;Wand&quot; e il menu &quot;L-Frame&quot; sulla scheda &quot;Camera&quot;. Impostare il &quot;conte Wand&quot; a 2.500, fai clic su &quot;Start&quot; sul menu &quot;Calibrate Telecamere&quot;, e sventolare la bacchetta di calibrazione attraverso l&#39;intero spazio motion capture. Il processo di calibrazione si arresta quando il conteggio raggiunge bacchetta 2.500. </li><li> Ripetere il processo di calibrazione se l&#39;errore di immagine (nella parte inferiore della scheda &quot;Camera&quot; del pannello &quot;Strumenti&quot;) è superiore a 0,3 fo di qualsiasi fotocamera. Dopo aver calibrato, mettere la bacchetta sul pavimento al centro dello spazio motion capture e fare clic su &quot;Start&quot; sul menu &quot;Imposta volume d&#39;origine&quot; per impostare l&#39;origine dello spazio motion capture. </li></ol></li><li> Verificare la copertura del sistema di motion capture con una prova senza per registrare il tracciato del movimento di un marcatore sventolato da un utente nello spazio motion capture e confermare se il marcatore viene rilevato e monitorato. Se l&#39;indicatore è spesso persa durante il rilevamento, ripetere la calibrazione del volume fino a quando il manichino di prova ha esito positivo. <ol><li> Fare clic sulla scheda &quot;Capture&quot; sul pannello &quot;Strumenti&quot; e poi &quot;Start&quot; sul menu &quot;Capture&quot; prima agitando il marcatore campione attraverso l&#39;intero spazio motion capture per registrare la sua traiettoria. </li><li> Dopo la registrazione, clicca su &quot;Esegue il gasdotto Ricostruire&quot; per ricostruire le posizioni del marcatore e controllare la qualità della registrazione. </li></ol></li><li> Collegare i terminali del MicroBattery (allegato al zaino nel passaggio 3.4) per i piedini di alimentazione dello zaino. </li><li> Testare la comunicazione wireless tra il computer portatile e lo zaino con il software di controllo di volo personalizzato. Fare clic sul comando &quot;Start&quot; sul software e verificare lo stato della connessione visualizzato. </li></ol> Esperimento 5. Volo Libero <ol><li> Effettuare l&#39;esperimento volo libero in un&#39;arena volo misura 16 x 8 x 4 m 3. </li><li> Inserire i parametri appropriati per il software di controllo di volo (tensione, larghezza d&#39;impulso, la frequenza, la durata e la stimolazione). Nota: Per la dimostrazione, abbiamo risolto la tensione di 3 V, durata dell&#39;impulso di 3 msec, e durata stimolo per 1 sec e varia la frequenza da 60 a 100 Hz. <ol><li> Nella schermata del software, tipo 3 per 3 V nella casella &quot;tensione&quot;, 1.000 per 1.000 msec nel box &quot;Stimolazione Durata&quot;, 3 per 3 ms nella casella &quot;Pulse Width&quot;, e una frequenza desiderata in Hz nel &quot; Frequenza &quot;scatola on la finestra di comando. </li></ol></li><li> Rilasciare il coleottero zaino montato in aria permettendo di volare liberamente all&#39;interno dell&#39;arena di volo. attivare manualmente la stimolazione quando il coleottero entra nello spazio motion capture. Premere il pulsante appropriato di comando (sinistra o destra) sul telecomando per stimolare il muscolo bersaglio sul lato sinistro o destro del coleottero. Nota: Una volta che il pulsante viene premuto, il software di controllo di volo in esecuzione sul computer portatile genera il comando e lo invia allo zaino. Lo zaino irradia il stimolo elettrico al muscolo di interesse (sul lato sinistro o destro). </li><li> Osservare la reazione del coleottero in tempo reale durante la stimolazione e ricostruire i dati utilizzando il software rappresentazione grafica 3D. <ol><li> Selezionare una delle prove registrate nella lista di dati della finestra &quot;Maggiolino Display&quot; e fare clic su &quot;Esporta Panda&quot; per copiare i dati di quel processo nella cartella di analisi ed eseguire il modulo di rappresentazione grafica 3D. </li><li> Premere il tasto &quot;N&quot; sula tastiera per combinare il segnale di stimolo con la traiettoria registrata. Premere I per mostrare la traiettoria del coleottero con i periodi di stimolazione evidenziati. </li></ol></li></ol>

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The authors declare that there are no conflicts of interest.

Il processo di impianto è importante, in quanto compromettono l&#39;affidabilità dell&#39;esperimento. Gli elettrodi devono essere inseriti nel muscolo ad una profondità pari o inferiore a seconda delle dimensioni del coleottero (evitando il contatto con muscoli adiacenti) 3 mm. Se gli elettrodi si toccano i muscoli circostanti, azioni motorie indesiderate e comportamenti si possono verificare a causa della contrazione dei muscoli vicini. I due elettrodi devono essere ben allineati per garantire che non si verifichin...

An insect-machine hybrid system, often referred to as a cyborg insect or biobot, is the fusion of a living insect platform with a miniature mounted electronic device. The electronic device, which is wirelessly commanded by a remote user, outputs an electrical signal to electrically stimulate neuromuscular sites in the insect via implanted wire electrodes to induce user desired motor actions and behaviors. In the early stages of this research field, researchers were limited to conducting wireless recording of the muscular action of an insect, using simple analog circuits comprised of surface-mounted components1-3. The development of system-on-a-chip technology with radio frequency functionality enabled not only the wireless recording of neuromuscular signals but also the electrical stimulation of the neuromuscular sites in living insects. At present, a built-in radio microcontroller is small enough to be mounted on living insects without causing any obstructions to their locomotion4-13.
The development of the built-in radio microcontroller allows researchers to determine electrical stimulation protocols to induce desired motor actions to control the locomotion of the insect of interest. On the ground, researchers have demonstrated walking control by stimulating the neuromuscular sites of cockroaches4,12,14, spiders15, and beetles16,17. In the air, the initiation and cessation of flight were achieved using different methods such as the stimulation of the optic lobes (the massive neural cluster of a compound eye) in beetles7,9 and brain sub-regions in bees18, whereas turning control has been demonstrated by stimulating the antennae muscles and nervous system of the abdomens in moths11,19 and the flight muscles of beetles7,9,13. In most cases, a built-in radio microcontroller was integrated on a custom-designed printed circuit board to produce a miniature wireless stimulator (backpack), which was mounted on the insect of interest. This allows wireless electrical stimulation to be applied to a freely walking or flying insect. Such a microcontroller-mounted insect is what is referred to as an insect-machine hybrid system. 
This study describes the experimental protocols for building an insect-machine hybrid system, wherein a living beetle is employed as the insect platform, and instructs on how to operate the robot and test its flight control systems. The third axillary sclerite (3Ax) muscle was chosen as the muscle of interest for electrical stimulation and demonstration of left or right turning control13. A pair of thin silver wire electrodes was implanted in both the left and right 3Ax muscles. Moreover, a backpack was mounted on the living beetle. The other ends of the wire electrode were connected to the output pins of the microcontroller. The backpack was small enough for the beetle to carry in flight. Thus, this allows an experimentalist to remotely stimulate the muscle of interest of an insect in free flight and investigate its reactions to the stimulations.

<table><tbody><tr><td>Mecynorrhina torquata beetle</td><td>Kingdom of Beetle Taiwan</td><td></td><td>10 g, 8 cm, pay load capacity is 30% of the body mass&lt;br/&gt; Aproval of importing and using by Agri-Food and Veterinary Authority of Singapore (AVA; HS code: 01069000, product code: ALV002).</td></tr><tr><td>Wireless backpack stimulator</td><td>Custom</td><td></td><td>TI CC2431 micocontroler&lt;br/&gt; The board is custom made based on the GINA board from Prof. Kris Pister&amp;rsquo;s lab. The layout of GINA board can be found at&amp;nbsp;&amp;nbsp;&amp;nbsp; https://openwsn.atlassian.net/wiki/display/OW/GINA</td></tr><tr><td>Wii Remote control</td><td>Nintendo</td><td></td><td>Bluetooth remote control to send the command to the operator laptop</td></tr><tr><td>BeetleCommander v1.8</td><td>Custom. Maharbiz group at UC Berkeley and Sato group at NTU</td><td></td><td>Establish the wireless communication of the backpack and the operator laptop. Configure the stimulus parameters and log the positional data. Visualize the flight data.</td></tr><tr><td>GINA base station</td><td>Kris Pister group at UC Berkeley</td><td></td><td>TI MSP430F2618 and AT86RF231</td></tr><tr><td>Motion capture system</td><td>VICON</td><td>T160</td><td>8 cameras for a flight arena of 12.5 m x 8 m x 4 m</td></tr><tr><td>Motion capture system</td><td>VICON</td><td>T40s</td><td>12 cameras for a flight arena of 12.5 x 8 x 4 m</td></tr><tr><td>Micro battery</td><td>Fullriver</td><td>&amp;nbsp;201013HS10C&amp;nbsp;</td><td>3.7V, 10 mAh</td></tr><tr><td>Retro reflective tape</td><td>Reflexite</td><td>V92-1549-010150</td><td>V92 reflective tape, silver color</td></tr><tr><td>PFA-Insulated Silver Wire&amp;nbsp;</td><td>A-M systems</td><td>786000</td><td>127 &amp;micro;m bare, 177.8 &amp;micro;m coated, 3 mm bare silver flame exposed at tips</td></tr><tr><td>SMT Micro Header&amp;nbsp;</td><td>SAMTEC</td><td>FTSH-110-01-L-DV</td><td>0.3 mm x 6 mm, bend to make a 3 mm long slider to secure the electrode into the PCB header.</td></tr><tr><td>Beeswax</td><td></td><td></td><td>Secure the electrodes</td></tr><tr><td>Dental Wax</td><td>Vertex</td><td></td><td>Immobilize the beetle</td></tr><tr><td>Insect pin</td><td>ROBOZ</td><td>RS-6082-30</td><td>Size&amp;nbsp; 00; 0.3 mm Rod diameter; 0.03 mm tip width; 38 mm Length&amp;nbsp;&lt;br/&gt; Make electrode guiding holes on cuticle</td></tr><tr><td>Tweezers</td><td>DUMONT</td><td>RS-5015</td><td>Pattern #5; .05 mm x .01 mm Tip Size; 110 mm Length&lt;br/&gt; Dissecting and implantation</td></tr><tr><td>Scissors</td><td>ROBOZ</td><td>RS-5620</td><td>Vannas Micro Dissecting Spring Scissors; Straight; 3mm Cutting Edge; 0.1 mm Tip Width; 3&quot; Overall Length&amp;nbsp;&lt;br/&gt; Dissecting and implantation</td></tr><tr><td>Potable soldering iron</td><td>DAIYO</td><td>DS241</td><td>Reflow beeswax</td></tr><tr><td>Hotplate&amp;nbsp;</td><td>CORNING</td><td>PC-400D</td><td>Melting beeswax and dental wax</td></tr><tr><td>Flourescent lamp</td><td>Philips</td><td>TL5 14W</td><td>Light the entire flight arena with 30 panels (60 x 60 cm&lt;sup&gt;2&lt;/sup&gt;). Each panel has 3 lamps.&lt;br/&gt; 14 W, 549 mm x 17 mm&amp;nbsp;</td></tr></tbody></table>

insect-machine hybrid system: remote radio control of a freely flying beetle (mercynorrhina torquata)

L&#39;aumento dei dispositivi elettronici digitali di radio-enabled ha spinto l&#39;uso di piccoli registratori neuromuscolari wireless e stimolatori per studiare il comportamento degli insetti in volo. Questa tecnologia consente lo sviluppo di un sistema ibrido insetto-macchina utilizzando una piattaforma di insetto che vive descritto in questo protocollo. Inoltre, questo protocollo presenta la configurazione del sistema e ponti procedure sperimentali gratuiti per valutare la funzione dei muscoli di volo in un insetto untethered. Per la dimostrazione, abbiamo mirato al terzo muscolo ascellare sclerite (3AX) per controllare e raggiungere svolta a sinistra oa destra di un coleottero che vola. Un sottile elettrodo filo d&#39;argento è stato impiantato sul muscolo 3AX su ciascun lato del coleottero. Questi sono stati collegati alle uscite di uno zaino wireless (cioè, uno stimolatore elettrico neuromuscolare) montato sul pronoto del coleottero. Il muscolo è stata stimolata in volo libero, alternando il lato stimolazione (sinistra o destra) oppure variando la stimulatiofrequenza n. Il coleottero si voltò verso il lato ipsilaterale quando il muscolo è stata stimolata ed esposto una risposta graduata ad una frequenza crescente. Il processo di impiantazione e calibrazione del volume del sistema di telecamere capture 3 dimensionale motion devono essere effettuate con cura per evitare di danneggiare il muscolo e perdere traccia del marcatore, rispettivamente. Questo metodo è estremamente utile per studiare volo degli insetti, in quanto contribuisce a rivelare le funzioni del muscolo volo di interesse volo libero.

La procedura di impianto di elettrodi è presentato in figura 2 elettrodi a filo d&#39;argento sottile sono stati impiantati nel muscolo 3AX del coleottero attraverso piccoli fori trafitto sulla cuticola morbida sul muscolo (figure 2d - e).. Questo cuticola morbida si trova appena sopra il apodema del muscolo basalar dopo aver rimosso la parte anteriore del metepisternum (figure 2d - c). Gli elettrodi so...

Watch this Scientific Journal Video about Insect-machine Hybrid System: Remote Radio Control of a Freely Flying Beetle Mercynorrhina torquata at JoVE.com

Insect-machine Hybrid System: Remote Radio Control of a Freely Flying Beetle Mercynorrhina torquata

This protocol describes the process of constructing an insect-machine hybrid system and carrying out wireless electrical stimulation of the flight muscles required to control the turning motion of a flying insect.

Insect-macchina Sistema ibrido: radiocomandato di un coleottero liberamente Volare (<em&gt; Torquata Mercynorrhina</em&gt;)

The rise of radio-enabled digital electronic devices has prompted the use of small wireless neuromuscular recorders and stimulators for studying in-flight ...

insect-machine-hybrid-system-remote-radio-control-freely-flying

Research

JoVE Journal

Neuroscience

Insect-machine Hybrid System: Remote Radio Control of a Freely Flying Beetle (Mercynorrhina torquata)

12.1K Views.  Nanyang Technological University. This protocol describes the process of constructing an insect-machine hybrid system and carrying out wireless electrical stimulation of the flight muscles required to control the turning motion of a flying insect.

Video: Insect-machine Hybrid System: Remote Radio Control of a Freely Flying Beetle Mercynorrhina torquata

Early Metamorphic Insertion Technology for Insect Flight Behavior Monitoring

Early Metamorphosis Insertion Technology (EMIT) is a novel methodology for integrating microfabricated neuromuscular recording and actuation platforms on insects during their metamorphic development. Here, the implants are fused within the structure and function of the neuromuscular system as a result of metamorphic tissue remaking. The implants emerge with the insect where the development of tissue around the electronics during pupal development results in a bioelectrically and biomechanically enhanced tissue interface. This relatively more reliable and stable interface would be beneficial for many researchers exploring the neural basis of the insect locomotion with alleviated traumatic effects caused during adult stage insertions. In this article, we implant our electrodes into the indirect flight muscles of Manduca sexta. Located in the dorsal-thorax, these main flight powering dorsoventral and dorsolongitudinal muscles actuate the wings and supply the mechanical power for up and down strokes. Relative contraction of these two muscle groups has been under investigation to explore how the yaw maneuver is neurophysiologically coordinated. To characterize the flight dynamics, insects are often tethered with wires and their flight is recorded with digital cameras. We also developed a novel way to tether Manduca sexta on a magnetically levitating frame where the insect is connected to a commercially available wireless neural amplifier. This set up can be used to limit the degree of freedom to yawing &#8220;only&#8221; while transmitting the related electromyography signals from dorsoventral and dorsolongitudinal muscle groups.

We present a novel surgical procedure to implant electrodes in Manduca sexta during its early metamorphic stages. This technique allows mechanically stable and electrically reliable coupling with the neuromuscular tissue to study flight neurophysiology dynamics. We also present a novel magnetic levitation platform for tethered studies of insect yaw.

All&#39;inizio metamorfica Tecnologia Inserzione per Insect Volo monitoraggio del comportamento

Early Metamorphosis Insertion Technology (EMIT) is a novel methodology for integrating microfabricated neuromuscular recording and actuation platforms on ...

Ricerca

Comportamento

Rearing and Double-stranded RNA-mediated Gene Knockdown in the Hide Beetle, Dermestes maculatus

Advances in genomics have raised the possibility of probing biodiversity at an unprecedented scale. However, sequence alone will not be informative without tools to study gene function. The development and sharing of detailed protocols for the establishment of new model systems in laboratories, and for tools to carry out functional studies, is thus crucial for leveraging the power of genomics. Coleoptera (beetles) are the largest clade of insects and occupy virtually all types of habitats on the planet. In addition to providing ideal models for fundamental research, studies of beetles can have impacts on pest control as they are often pests of households, agriculture, and food industries. Detailed protocols for rearing and maintenance of D. maculatus laboratory colonies and for carrying out dsRNA-mediated interference in D. maculatus are presented. Both embryonic and parental RNAi procedures-including apparatus set up, preparation, injection, and post-injection recovery-are described. Methods are also presented for analyzing embryonic phenotypes, including viability, patterning defects in hatched larvae, and cuticle preparations for unhatched larvae. These assays, together with in situ hybridization and immunostaining for molecular markers, make D. maculatus an accessible model system for basic and applied research. They further provide useful information for establishing procedures in other emerging insect model systems.

Rearing and Double-stranded RNA-mediated Gene Knockdown in the Hide Beetle, Dermestes maculatus

Here, we present protocols for rearing an intermediate-germ beetle, Dermestes maculatus (D. maculatus) in the lab. We also share protocols for embryonic and parental RNAi and methods for analyzing embryonic phenotypes to study gene function in this species.

Allevamento e RNA a doppio filamento mediata silenziamento genico nel Hide Beetle,<em&gt; Dermestes maculatus</em

Advances in genomics have raised the possibility of probing biodiversity at an unprecedented scale. However, sequence alone will not be informative ...

Genetica

Operant Learning of Drosophila at the Torque Meter

For experiments at the torque meter, flies are kept on standard fly medium at 25&deg;C and 60% humidity with a 12hr light/12hr dark regime. A standardized breeding regime assures proper larval density and age-matched cohorts. Cold-anesthetized flies are glued with head and thorax to a triangle-shaped hook the day before the experiment. Attached to the torque meter via a clamp, the fly's intended flight maneuvers are measured as the angular momentum around its vertical body axis. The fly is placed in the center of a cylindrical panorama to accomplish stationary flight. An analog to digital converter card feeds the yaw torque signal into a computer which stores the trace for later analysis. The computer also controls a variety of stimuli which can be brought under the fly's control by closing the feedback loop between these stimuli and the yaw torque trace. Punishment is achieved by applying heat from an adjustable infrared laser.

Measuring the yaw torque of tethered Drosophila with the torque meter allows the neuroscientist exquisite control of the stimulus situation of the experimental animal. Together with the unique genetic tools available in the fruit fly, this paradigm is used for a wide variety of neurobiological research.

L&#39;apprendimento della Drosophila operante presso il misuratore di coppia

For experiments at the torque meter, flies are kept on standard fly medium at 25&deg;C and 60% humidity with a 12hr light/12hr dark regime. A standardized ...

Biologia

Measuring the Flight Ability of the Ambrosia Beetle, Platypus Quercivorus (Murayama), Using a Low-Cost, Small, and Easily Constructed Flight Mill

The ambrosia beetle, Platypus quercivorus (Murayama), is the vector of a fungal pathogen that causes mass mortality of Fagaceae trees (Japanese oak wilt). Therefore, knowing the dispersal capacity may help inform trapping/tree removal efforts to prevent this disease more effectively. In this study, we measured the flight velocity and duration and estimated the flight distance of the beetle using a newly developed flight mill. The flight mill is low cost, small, and constructed using commonly available items. Both the flight mill arm and its vertical axis comprise a thin needle. A beetle specimen is glued to one tip of the arm using instant glue. The other tip is thick due to being covered with plastic, thus it facilitates the detection of rotations of the arm. The revolution of the arm is detected by a photo sensor mounted on an infrared LED, and is indicated by a change in the output voltage when the arm passed above the LED. The photo sensor is connected to a personal computer and the output voltage data are stored at a sampling rate of 1 kHz. By conducting experiments using this flight mill, we found that P. quercivorus can fly at least 27 km. Because our flight mill comprises cheap and small ordinary items, many flight mills can be prepared and used simultaneously in a small laboratory space. This enables experimenters to obtain a sufficient amount of data within a short period.

Measuring the Flight Ability of the Ambrosia Beetle, Platypus Quercivorus Murayama, Using a Low-Cost, Small, and Easily Constructed Flight Mill

We developed a low cost and small flight mill, constructed with commonly available items and easily used in experimentation. Using this apparatus, we measured the flight ability of an ambrosia beetle, Platypus quercivorus.

Misura la capacità di volo del Scarabeo dell'Ambrosia, Platypus Quercivorus (Murayama), utilizzando un basso costo, piccole e facilmente costruito volo Mill

The ambrosia beetle, Platypus quercivorus (Murayama), is the vector of a fungal pathogen that causes mass mortality of Fagaceae trees (Japanese oak wilt). ...

Ambiente

Using Flight Mills to Measure Flight Propensity and Performance of Western Corn Rootworm, Diabrotica virgifera virgifera (LeConte)

The western corn rootworm, Diabrotica virgifera virgifera (LeConte) (Coleoptera: Chrysomelidae), is an economically important pest of corn in the northern United States. Some populations have developed resistance to management strategies including transgenic corn that produces insecticidal toxins derived from the bacterium Bacillus thuringiensis (Bt). Knowledge of western corn rootworm dispersal is of critical importance for models of resistance evolution, spread, and mitigation. Flight behavior of an insect, especially over a long distance, is inherently difficult to observe and characterize. Flight mills provide a means to directly test developmental and physiological impacts and consequences of flight in the laboratory that cannot be obtained in field studies. In this study, flight mills were used to measure the timing of flight activity, total number of flights, and the distance, duration, and speed of flights taken by female rootworms during a 22-h test period. Sixteen flight mills were housed in an environmental chamber with programmable lighting, temperature, and humidity control. The flight mill described is of a typical design, where a flight arm is free to rotate about a central pivot. Rotation is caused by flight of an insect tethered to one end of the flight arm, and each rotation is recorded by a sensor with a time-stamp. Raw data are compiled by software, which are subsequently processed to provide summary statistics for flight parameters of interest. The most difficult task for any flight mill study is attachment of the tether to the insect with an adhesive, and the method used must be tailored to each species. The attachment must be strong enough to hold the insect in a rigid orientation and to prevent detachment during movement, while not interfering with natural wing motion during flight. The attachment process requires dexterity, finesse, and speed, making video footage of the process for rootworms of value.

Using Flight Mills to Measure Flight Propensity and Performance of Western Corn Rootworm, Diabrotica virgifera virgifera LeConte

Flight mills are important tools for comparing how age, sex, mating status, temperature, or various other factors may influence an insect&#8217;s flight behavior. Here we describe protocols to tether and measure the flight propensity and performance of western corn rootworm under different treatments.

Utilizzando Flight Mills per misurare la propensione di volo e le prestazioni di Western Corn Rootworm, Diabrotica virgifera virgifera (LeConte)

The western corn rootworm, Diabrotica virgifera virgifera (LeConte) (Coleoptera: Chrysomelidae), is an economically important pest of corn in the northern ...

Extracellular Wire Tetrode Recording in Brain of Freely Walking Insects

Increasing interest in the role of brain activity in insect motor control requires that we be able to monitor neural activity while insects perform natural behavior. We previously developed a technique for implanting tetrode wires into the central complex of cockroach brains that allowed us to record activity from multiple neurons simultaneously while a tethered cockroach turned or altered walking speed. While a major advance, tethered preparations provide access to limited behaviors and often lack feedback processes that occur in freely moving animals. We now present a modified version of that technique that allows us to record from the central complex of freely moving cockroaches as they walk in an arena and deal with barriers by turning, climbing or tunneling. Coupled with high speed video and cluster cutting, we can now relate brain activity to various parameters of the movement of freely behaving insects.

We previously developed a technique for implanting tetrode wires into the central complex of cockroach brains that allows us to monitor activity in individual units of tethered cockroaches. Here we present a modified version of that technique that allows us to also record brain activity in freely moving insects.

Extracellulare Wire Tetrode registrazione in Brain di Liberamente Walking Insetti

Increasing interest in the role of brain activity in insect motor control requires that we be able to monitor neural activity while insects perform ...

Neuroscienze

Building an Enhanced Flight Mill for the Study of Tethered Insect Flight

Makerspaces have a high potential of enabling researchers to develop new techniques and to work with novel species in ecological research. This protocol demonstrates how to take advantage of the technology found in makerspaces in order to build a more versatile flight mill for a relatively low cost. Given that this study extracted its&#160;prototype from flight mills built in the last decade, this protocol focuses more on outlining divergences made from the simple, modern flight mill. Previous studies have already shown how advantageous flight mills are to measuring flight parameters such as speed, distance, or periodicity. Such mills have allowed researchers to associate these parameters with morphological, physiological, or genetic factors. In addition to these advantages, this study discusses the benefits of using the technology in makerspaces, like 3D printers and laser cutters, in order to build a more flexible, sturdy, and collapsible flight mill design. Most notably, the 3D printed components of this design allow the user to test insects of various sizes by making the heights of the mill arm and infrared (IR) sensors adjustable. The 3D prints also enable the user to easily disassemble the machine for quick storage or transportation to the field. Moreover, this study makes greater use of magnets and magnetic paint to tether insects with minimal stress. Lastly, this protocol details a versatile analysis of flight data through computer scripts that efficiently separate and analyze differentiable flight trials within a single recording. Although more labor-intensive, applying the tools available in makerspaces and on online 3D modeling programs facilitates multidisciplinary and process-orientated practices and helps researchers avoid costly, premade products with narrowly adjustable dimensions. By taking advantage of the flexibility and reproducibility of technology in makerspaces, this protocol promotes creative flight mill design and inspires open science.

This protocol uses three-dimensional (3D) printers and laser cutters found in makerspaces in order to create a more flexible flight mill design. By using this technology, researchers can reduce costs, enhance design flexibility, and generate reproducible work when constructing their flight mills for tethered insect flight studies.

Costruire un mulino di volo potenziato per lo studio del volo degli insetti legati

Makerspaces have a high potential of enabling researchers to develop new techniques and to work with novel species in ecological research. This protocol ...

Ingegneria

Using Insect Electroantennogram Sensors on Autonomous Robots for Olfactory Searches

Robots designed to track chemical leaks in hazardous industrial facilities1 or explosive traces in landmine fields2 face the same problem as insects foraging for food or searching for mates3: the olfactory search is constrained by the physics of turbulent transport4. The concentration landscape of wind borne odors is discontinuous and consists of sporadically located patches. A pre-requisite to olfactory search is that intermittent odor patches are detected. Because of its high speed and sensitivity5-6, the olfactory organ of insects provides a unique opportunity for detection. Insect antennae have been used in the past to detect not only sex pheromones7 but also chemicals that are relevant to humans, e.g., volatile compounds emanating from cancer cells8 or toxic and illicit substances9-11. We describe here a protocol for using insect antennae on autonomous robots and present a proof of concept for tracking odor plumes to their source. The global response of olfactory neurons is recorded in situ in the form of electroantennograms (EAGs). Our experimental design, based on a whole insect preparation, allows stable recordings within a working day. In comparison, EAGs on excised antennae have a lifetime of 2 hr. A custom hardware/software interface was developed between the EAG electrodes and a robot. The measurement system resolves individual odor patches up to 10 Hz, which exceeds the time scale of artificial chemical sensors12. The efficiency of EAG sensors for olfactory searches is further demonstrated in driving the robot toward a source of pheromone. By using identical olfactory stimuli and sensors as in real animals, our robotic platform provides a direct means for testing biological hypotheses about olfactory coding and search strategies13. It may also prove beneficial for detecting other odorants of interests by combining EAGs from different insect species in a bioelectronic nose configuration14 or using nanostructured gas sensors that mimic insect antennae15.

We describe a protocol for using insect antennae in the form of electroantennograms (EAGs) on autonomous robots. Our experimental design allows stable recordings within a day and resolves individual odor patches up to 10 Hz. The efficiency of EAG sensors for olfactory searches is demonstrated in driving a robot toward an odor source.

Utilizzando Insetto Electroantennogram sensori sul robot autonomi per le ricerche olfattive

Robots designed to track chemical leaks in hazardous industrial facilities1 or explosive traces in landmine fields2 face the same problem as insects ...

Radio Frequency Identification and Motion-sensitive Video Efficiently Automate Recording of Unrewarded Choice Behavior by Bumblebees

We present two methods for observing bumblebee choice behavior in an enclosed testing space. The first method consists of Radio Frequency Identification (RFID) readers built into artificial flowers that display various visual cues, and RFID tags (i.e., passive transponders) glued to the thorax of bumblebee workers. The novelty in our implementation is that RFID readers are built directly into artificial flowers that are capable of displaying several distinct visual properties such as color, pattern type, spatial frequency (i.e., &#8220;busyness&#8221; of the pattern), and symmetry (spatial frequency and symmetry were not manipulated in this experiment). Additionally, these visual displays in conjunction with the automated systems are capable of recording unrewarded and untrained choice behavior. The second method consists of recording choice behavior at artificial flowers using motion-sensitive high-definition camcorders. Bumblebees have number tags glued to their thoraces for unique identification. The advantage in this implementation over RFID is that in addition to observing landing behavior, alternate measures of preference such as hovering and antennation may also be observed. Both automation methods increase experimental control, and internal validity by allowing larger scale studies that take into account individual differences. External validity is also improved because bees can freely enter and exit the testing environment without constraints such as the availability of a research assistant on-site. Compared to human observation in real time, the automated methods are more cost-effective and possibly less error-prone.

This video describes Radio-Frequency Identification (RFID) and motion-sensitive video recording methods to monitor choice behavior by bumblebees.

Radio Frequency Identification e video sensibile al movimento in modo efficiente Automatizzare registrazione di ricompensa Scelta comportamento di bombi

We present two methods for observing bumblebee choice behavior in an enclosed testing space. The first method consists of Radio Frequency Identification ...

A Simple Flight Mill for the Study of Tethered Flight in Insects

Flight in insects can be long-range migratory flights, intermediate-range dispersal flights, or short-range host-seeking flights. Previous studies have shown that flight mills are valuable tools for the experimental study of insect flight behavior, allowing researchers to examine how factors such as age, host plants, or population source can influence an insects' propensity to disperse. Flight mills allow researchers to measure components of flight such as speed and distance flown. Lack of detailed information about how to build such a device can make their construction appear to be prohibitively complex. We present a simple and relatively inexpensive flight mill for the study of tethered flight in insects. Experimental insects can be tethered with non-toxic adhesives and revolve around an axis by means of a very low friction magnetic bearing. The mill is designed for the study of flight in controlled conditions as it can be used inside an incubator or environmental chamber. The strongest points are the very simple electronic circuitry, the design that allows sixteen insects to fly simultaneously allowing the collection and analysis of a large number of samples in a short time and the potential to use the device in a very limited workspace. This design is extremely flexible, and we have adjusted the mill to accommodate different species of insects of various sizes.

Flight in insects is influenced by a number of factors and the propensity to disperse is an important variable in understanding insect ecology and biological control strategies. We describe the construction and use of a simple, relatively inexpensive, and flexible flight mill for measuring parameters of tethered flight in insects.

Un semplice volo Mill per lo Studio del Tethered Volo in Insetti

Flight in insects can be long-range migratory flights, intermediate-range dispersal flights, or short-range host-seeking flights. Previous studies have ...

Insect-macchina Sistema ibrido: radiocomandato di un coleottero liberamente Volare (

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Capitoli in questo video

All'inizio metamorfica Tecnologia Inserzione per Insect Volo monitoraggio del comportamento

Allevamento e RNA a doppio filamento mediata silenziamento genico nel Hide Beetle,

L'apprendimento della Drosophila operante presso il misuratore di coppia