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. Estudio de Animales <ol><li> escarabajos traseros individuales Mecynorrhina torquata (6 cm, 8 g) en envases de plástico por separado con ropa de cama de pellets de madera. </li><li> Alimentar cada escarabajo de una taza de gelatina de azúcar (12 ml) cada 3 días. </li><li> Mantener la temperatura y humedad de la habitación de crianza, a 25 ° C y 60%, respectivamente. </li><li> Prueba de la capacidad de vuelo de cada escarabajo antes de implantar electrodos de alambre fino. <ol><li> tirar suavemente un escarabajo en el aire. Si el escarabajo puede volar durante más de 10 segundos durante 5 ensayos consecutivos, la conclusión de que el escarabajo tiene capacidad de vuelo regulares y emplearla para experimentos posteriores de vuelo. Para recuperar el escarabajo, apagar todas las luces de la habitación para que sea oscuro. Esto hace que el escarabajo de interrumpir el vuelo. Nota: Un escarabajo comienza espontáneamente a volar cuando se liberan en el aire. Es mejor para llevar a cabo los experimentos de vuelo en una habitación grande cerrado tal como el que se muestra en la Figura 1 (16 x 8 x 4 m <sup&gt; 3 con un espacio de captura de movimiento de 12,5 x 8 x 4 m 3), como un escarabajo volador mueve muy rápido (aproximadamente 3-5 m / seg) y atrae a grandes arcos al girar en el aire. </li></ol></li></ol> 2. Electrodo de implantación <ol><li> Anestesiar al escarabajo colocándolo en un recipiente de plástico llena de CO 2 durante 1 min 13,16,20-24. </li><li> Ablandar la cera dental sumergiéndola en agua caliente durante 10 seg. Coloque el escarabajo anestesiado en un bloque de madera e inmovilizarlo con la cera dental ablandada. La cera dental natural se enfría y se solidifica en unos pocos minutos. </li><li> Corte los cables aislados de plata (127 micras de diámetro desnudo, 178 m de diámetro cuando se recubre con perfluoroalkoxy) en longitudes de 25 mm para utilizar como electrodos de alambre fino para la implantación. </li><li> Exponer a 3 mm de plata desnuda por flameado el aislante en ambos extremos de cada cable. </li><li> Diseccionar la superficie superior de la cutícula del escarabajo utilizando una tijera de punta fina para crear un smaventana ll de aproximadamente 4 x 4 mm en la metepisternum (Figura 2c). Nota: una cutícula de color marrón suave se expone entonces, como se muestra en las figuras 2c - e. El músculo 3AX se encuentra debajo de la cutícula suave. </li><li> Pierce dos agujeros en la cutícula marrón expuesto utilizando un PIN de insectos (tamaño 00) con una distancia de 2 mm entre los dos agujeros (Figura 2D). </li><li> Insertar dos electrodos de alambre (incluyendo uno electrodos activos y uno de retorno preparados en la etapa 2.4) con cuidado a través de los agujeros y implantarlos en cada músculo 3AX a una profundidad de 3 mm. </li><li> Asegurar los electrodos implantados y mantenerlos en su lugar para evitar el contacto y cortocircuitos dejando caer cera derretida en los agujeros. Si es necesario, ajustar el flujo de cera de abeja sobre la cutícula tocando la cera de abeja con la punta de un soldador caliente. La cera de abejas se solidifica rápidamente y refuerza la implantación. Nota: Para comprobar si el implante es correcta, los élitros de la remolachaLe puede ser levantado para observar el movimiento del músculo 3AX durante la estimulación eléctrica. </li></ol> 3. Asamblea Mochila inalámbrica Nota: La mochila consistía en un microcontrolador incorporado de radio en una tabla de capas 4 FR-4 (1,6 x 1,6 cm 2). La mochila fue impulsado por un MicroBattery de polímero de litio (3,7 V, 350 mg, 10 mAh). La masa total de la mochila incluyendo la batería fue de 1,2 ± 0,26 g, que es menor que la capacidad de carga útil del escarabajo (30% de 10 g de peso corporal). La mochila se pre-programados para recibir comunicaciones inalámbricas y tenía dos canales de salida. <ol><li> Limpiar la superficie pronoto (quitar la capa de cera sobre la cutícula) con cinta adhesiva de doble cara. A continuación, coloque la mochila en el pronoto del escarabajo con un trozo de cinta de doble cara. </li><li> Conectar los extremos de los electrodos implantados a las salidas de la mochila. </li><li> Envuelva la cinta retrorreflectante alrededor del MicroBattery para producir un marcador for cámaras de captura de movimiento para detectar. </li><li> Coloque la microbatería a la parte superior de la mochila con un trozo de cinta de doble cara para que la cinta retro-reflectante puede ser detectada por las cámaras de captura de movimiento. </li></ol> 4. Sistema de control inalámbrico Nota: En este caso, el sistema de control inalámbrico término incluye un receptor para el mando a distancia, un ordenador portátil para ejecutar el software de control de vuelo a medida, una estación base, la mochila, y el sistema de captura de movimiento. <ol><li> Conectar la estación base y el receptor del mando a distancia para el ordenador portátil a través de puertos USB. </li><li> Encienda el sistema de captura de movimiento y conectarlo a la computadora portátil a través de un puerto Ethernet. </li><li> Realizar la calibración de volumen agitando la varita de calibración (proporcionado por la empresa proveedora del sistema de captura de movimiento) para cubrir la totalidad del espacio de captura de movimiento. <ol><li> Abra el software de captura de movimiento desde el escritorio del ordenador portátil. Haga clic y drag para seleccionar todas las cámaras en el menú &quot;Sistema&quot; del panel &quot;Recursos&quot;. </li><li> Haga clic en el menú &quot;Perspectiva 3D&quot; y seleccione &quot;Cámara&quot; para cambiar a la vista de la cámara. Haga clic en la pestaña &quot;Cámara&quot; en el panel &quot;Herramientas&quot; para mostrar la configuración de calibración. Haga clic en &quot;Inicio&quot; en el menú &quot;Crear máscaras de la cámara&quot; para eliminar el ruido de las cámaras y luego &quot;Stop&quot; después de que el ruido se enmascara en azul. </li><li> Haga clic y seleccione &quot;Marcador de 5 Wand &amp; L-Frame&quot; en el menú &quot;Wand&quot; y el menú &quot;L-Frame&quot; en la pestaña &quot;Cámara&quot;. Ajuste el &quot;Conde Wand&quot; de 2500, haga clic en &quot;Inicio&quot; en el menú &quot;Cámaras Calibrar&quot;, y agitar la varita de calibración a través de todo el espacio de captura de movimiento. El proceso de calibración se detiene cuando el recuento de varita alcanza 2.500. </li><li> Repetir el proceso de calibración de error si la imagen (en la parte inferior de la pestaña &quot;Cámara&quot; del panel &quot;Herramientas&quot;) es superior a 0,3 fo cualquier cámara. Después de la calibración, poner la varita en el suelo en el centro del espacio de captura de movimiento y haga clic en &quot;Inicio&quot; en el menú &quot;Configuración de Origen Volumen&quot; para establecer el origen del espacio de captura de movimiento. </li></ol></li><li> Compruebe la cobertura del sistema de captura de movimiento utilizando una prueba simulada para registrar la trayectoria de movimiento de un marcador ondulado por un usuario en el espacio de captura de movimiento y confirmar si se detecta el marcador y de seguimiento. Si el marcador es frecuentemente pierde durante la detección, repetir la calibración volumen hasta que el maniquí de ensayo tiene éxito. <ol><li> Haga clic en la pestaña de &quot;captura&quot; en el panel &quot;Herramientas&quot; y luego &quot;Inicio&quot; en el menú &quot;Captura&quot; antes de agitar el marcador de la muestra a través de todo el espacio de captura de movimiento para grabar su trayectoria. </li><li> Después de la grabación, haga clic en &quot;Ejecuta la tubería Reconstruir&quot; para reconstruir las posiciones del marcador y comprobar la calidad de la grabación. </li></ol></li><li> Conecte los terminales de la Microbialattery (que se adjunta a la mochila en el paso 3.4) para los pines de alimentación de la mochila. </li><li> Prueba de la comunicación inalámbrica entre el ordenador portátil y la mochila con el software de control de vuelo personalizado. Haga clic en el comando &quot;Inicio&quot; en el software y comprobar el estado de la conexión mostrada. </li></ol> Experimento 5. Vuelo Libre <ol><li> Llevar a cabo el experimento de vuelo libre en una arena de vuelo midiendo 16 x 8 x 4 m 3. </li><li> Introducir los parámetros adecuados en el software de control de vuelo (voltaje, anchura de pulso, frecuencia, y duración de estimulación). Nota: Para la demostración, se fijó la tensión de 3 V, ancho de pulso a 3 mseg, y la duración de estimulación a 1 seg y variada la frecuencia de 60 a 100 Hz. <ol><li> En la pantalla de software, tipo 3 de 3 V en el cuadro de &quot;tensión&quot;, 1.000 por 1.000 mseg en la casilla &quot;Duración de estimulación&quot;, 3 por 3 m en el cuadro de &quot;ancho de pulso&quot;, y una frecuencia deseada en Hz en el &quot; frecuencia &quot;o cuadron la ventana de comandos. </li></ol></li><li> Suelte el escarabajo de la mochila montado en el aire que le permite volar libremente dentro de la arena de vuelo. activar manualmente la estimulación cuando el escarabajo entra en el espacio de captura de movimiento. Pulse el botón correspondiente de comandos (izquierda o derecha) en el control remoto para estimular el músculo deseado en el lado izquierdo o derecho del escarabajo. Nota: Una vez que se pulsa el botón, el software de control de vuelo que se ejecuta en el ordenador portátil genera el comando y lo envía a la mochila. La mochila a continuación, emite el estímulo eléctrico al músculo de interés (en el lado izquierdo o derecho). </li><li> Observar la reacción del escarabajo en tiempo real durante la estimulación y la reconstrucción de los datos utilizando el software de gráficos 3D. <ol><li> Seleccione uno de los ensayos registrados en la lista de datos de la ventana &quot;Display Escarabajo&quot; y haga clic en &quot;Exportar Panda&quot; para copiar los datos de dicho ensayo a la carpeta de análisis y ejecutar el módulo de gráficos 3D. </li><li> Pulse la tecla &quot;N&quot; en lael teclado para combinar la señal de estímulo con la trayectoria grabada. Pulse I para mostrar la trayectoria del escarabajo con los periodos de estimulación resaltados. </li></ol></li></ol>

<ol>
	<li>Kutsch, W., Schwarz, G., Fischer, H., Kautz, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Wireless+Transmission+of+Muscle+Potentials+During+Free+Flight+of+a+Locust.">Wireless Transmission of Muscle Potentials During Free Flight of a Locust.</a> J. Exp. Biol. 185 (1), 367-373 (1993).</li><li>Fischer, H., Kautz, H., Kutsch, W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+Radiotelemetric+2-Channel+Unit+for+Transmission+of+Muscle+Potentials+During+Free+Flight+of+the+Desert+Locust,+Schistocerca+Gregaria.">A Radiotelemetric 2-Channel Unit for Transmission of Muscle Potentials During Free Flight of the Desert Locust, Schistocerca Gregaria.</a> J. Neurosci. Methods. 64 (1), 39-45 (1996).</li><li>Ando, N., Shimoyama, I., Kanzaki, 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+Dual-Channel+FM+Transmitter+for+Acquisition+of+Flight+Muscle+Activities+from+the+Freely+Flying+Hawkmoth,+Agrius+Convolvuli.">A Dual-Channel FM Transmitter for Acquisition of Flight Muscle Activities from the Freely Flying Hawkmoth, Agrius Convolvuli.</a> J. Neurosci. Methods. 115 (2), 181-187 (2002).</li><li>Sanchez, C. J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Locomotion+control+of+hybrid+cockroach+robots.">Locomotion control of hybrid cockroach robots.</a> J. R. Soc. Interface. 12 (105), (2015).</li><li>Sato, H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+cyborg+beetle:+insect+flight+control+through+an+implantable,+tetherless+microsystem.">A cyborg beetle: insect flight control through an implantable, tetherless microsystem.</a> , 164-167 (2008).</li><li>Bozkurt, A., Gilmour, R. F., Lal, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Balloon-Assisted+Flight+of+Radio-Controlled+Insect+Biobots.">Balloon-Assisted Flight of Radio-Controlled Insect Biobots.</a> IEEE Trans. Biomed. Eng. 56 (9), 2304-2307 (2009).</li><li>Sato, H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Remote+Radio+Control+of+Insect+Flight.">Remote Radio Control of Insect Flight.</a> Front. Neurosci. 3, (2009).</li><li>Daly, D. C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+Pulsed+UWB+Receiver+SoC+for+Insect+Motion+Control.">A Pulsed UWB Receiver SoC for Insect Motion Control.</a> IEEE J. Solid-State Circuits. 45 (1), 153-166 (2010).</li><li>Maharbiz, M. M., Sato, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cyborg+Beetles.">Cyborg Beetles.</a> Sci. Am. 303 (6), 94-99 (2010).</li><li>Tsang, W. M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Remote+control+of+a+cyborg+moth+using+carbon+nanotube-enhanced+flexible+neuroprosthetic+probe.">Remote control of a cyborg moth using carbon nanotube-enhanced flexible neuroprosthetic probe.</a> , 39-42 (2010).</li><li>Hinterwirth, A. J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Wireless+Stimulation+of+Antennal+Muscles+in+Freely+Flying+Hawkmoths+Leads+to+Flight+Path+Changes.">Wireless Stimulation of Antennal Muscles in Freely Flying Hawkmoths Leads to Flight Path Changes.</a> PloS ONE. 7 (12), (2012).</li><li>Whitmire, E., Latif, T., Bozkurt, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Kinect-based+system+for+automated+control+of+terrestrial+insect+biobots.">Kinect-based system for automated control of terrestrial insect biobots.</a> , 1470-1473 (2013).</li><li>Sato, H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Deciphering+the+Role+of+a+Coleopteran+Steering+Muscle+via+Free+Flight+Stimulation.">Deciphering the Role of a Coleopteran Steering Muscle via Free Flight Stimulation.</a> Curr. Biol. 25 (6), 798-803 (2015).</li><li>Erickson, J. C., Herrera, M., Bustamante, M., Shingiro, A., Bowen, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effective+Stimulus+Parameters+for+Directed+Locomotion+in+Madagascar+Hissing+Cockroach+Biobot.">Effective Stimulus Parameters for Directed Locomotion in Madagascar Hissing Cockroach Biobot.</a> PLoS ONE. 10 (8), e0134348 (2015).</li><li>Zhaolin, Y., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+preliminary+study+of+motion+control+patterns+for+biorobotic+spiders.">A preliminary study of motion control patterns for biorobotic spiders.</a> , 128-132 (2014).</li><li>Feng, C., Chao, Z., Hao Yu, C., Sato, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Insect-machine+hybrid+robot:+Insect+walking+control+by+sequential+electrical+stimulation+of+leg+muscles.">Insect-machine hybrid robot: Insect walking control by sequential electrical stimulation of leg muscles.</a> , 4576-4582 (2015).</li><li>Cao, F., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+Biological+Micro+Actuator:+Graded+and+Closed-Loop+Control+of+Insect+Leg+Motion+by+Electrical+Stimulation+of+Muscles.">A Biological Micro Actuator: Graded and Closed-Loop Control of Insect Leg Motion by Electrical Stimulation of Muscles.</a> PLoS ONE. 9 (8), e105389 (2014).</li><li>Zhao, H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Neuromechanism+Study+of+Insect-Machine+Interface:+Flight+Control+by+Neural+Electrical+Stimulation.">Neuromechanism Study of Insect-Machine Interface: Flight Control by Neural Electrical Stimulation.</a> PLoS ONE. 9 (11), e113012 (2014).</li><li>Tsang, W. M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Flexible+Split-Ring+Electrode+for+Insect+Flight+Biasing+Using+Multisite+Neural+Stimulation.">Flexible Split-Ring Electrode for Insect Flight Biasing Using Multisite Neural Stimulation.</a> IEEE Trans. Biomed. Eng. 57 (7), 1757-1764 (2010).</li><li>Barron, A. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Anaesthetising+Drosophila+for+behavioural+studies.">Anaesthetising Drosophila for behavioural studies.</a> J. Insect Physiol. 46 (4), 439-442 (2000).</li><li>Cooper, J. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Anesthesia,+Analgesia,+and+Euthanasia+of+Invertebrates.">Anesthesia, Analgesia, and Euthanasia of Invertebrates.</a> ILAR Journal. 52 (2), 196-204 (2011).</li><li>Miller, T. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Insect neurophysiological techniques. , (2012).</li><li>Leary, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> AVMA guidelines for the euthanasia of animals. , (2013).</li><li>Heath, B., West, G., Heard, D., Caulkett, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mobile+Inhalant+Anesthesia+Techniques.">Mobile Inhalant Anesthesia Techniques.</a> in Zoo Animal and Wildlife Immobilization and Anesthesia. , 75-80 (2008).</li><li>Mischiati, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Internal+models+direct+dragonfly+interception+steering.">Internal models direct dragonfly interception steering.</a> Nature. 517 (7534), 333-338 (2015).</li><li>Kutsch, W., Berger, S., Kautz, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Turning+Manoeuvres+in+Free-Flying+Locusts:+Two-Channel+Radio-Telemetric+Transmission+of+Muscle+Activity.">Turning Manoeuvres in Free-Flying Locusts: Two-Channel Radio-Telemetric Transmission of Muscle Activity.</a> J. Exp. Zoolog. Part A Comp. Exp. Biol. 299 (2), 139-150 (2003).</li><li>Wang, H., Ando, N., Kanzaki, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Active+Control+of+Free+Flight+Manoeuvres+in+a+Hawkmoth,+Agrius+Convolvuli.">Active Control of Free Flight Manoeuvres in a Hawkmoth, Agrius Convolvuli.</a> J. Exp. Biol. 211 (3), 423-432 (2008).</li><li>Sato, H., Maharbiz, M. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Recent+developments+in+the+remote+radio+control+of+insect+flight.">Recent developments in the remote radio control of insect flight.</a> Front. Neurosci. 4, (2010).</li><li>Tien Van, T., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Flight+behavior+of+the+rhinoceros+beetle+Trypoxylus+dichotomus+during+electrical+nerve+stimulation.">Flight behavior of the rhinoceros beetle Trypoxylus dichotomus during electrical nerve stimulation.</a> Bioinsp. Biomim. 7 (3), 036021 (2012).</li><li>Sane, S. P., Dickinson, M. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+control+of+flight+force+by+a+flapping+wing:+lift+and+drag+production.">The control of flight force by a flapping wing: lift and drag production.</a> J. Exp. Biol. 204 (15), 2607-2626 (2001).</li><li>de Croon, G. C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Design,+aerodynamics+and+autonomy+of+the+DelFly.">Design, aerodynamics and autonomy of the DelFly.</a> Bioinsp. Biomim. 7 (2), 025003 (2012).</li><li>Ma, K. Y., Chirarattananon, P., Fuller, S. B., Wood, R. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Controlled+Flight+of+a+Biologically+Inspired,+Insect-Scale+Robot.">Controlled Flight of a Biologically Inspired, Insect-Scale Robot.</a> Science. 340 (6132), 603-607 (2013).</li></ol>

The authors declare that there are no conflicts of interest.

El proceso de implantación es importante, ya que afecta a la fiabilidad del experimento. Los electrodos deben ser insertados en el músculo a una profundidad de 3 mm o menos, dependiendo del tamaño del escarabajo (evitando el contacto con los músculos cercanos). Si los electrodos tocan los músculos cercanos, acciones motoras y comportamientos indeseables pueden ocurrir debido a la contracción de los músculos cercanos. Los dos electrodos deben estar bien alineados para asegurar que no se produzcan cortocircuitos. C...

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 border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<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 
			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 
			The board is custom made based on the GINA board from Prof. Kris Pister&#8217;s lab. The layout of GINA board can be found at&#160;&#160;&#160; 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 x 8 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>&#160;201013HS10C&#160;</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&#160;</td>
			<td>A-M systems</td>
			<td>786000</td>
			<td>127 &#181;m bare, 177.8 &#181;m coated, 3 mm bare silver flame exposed at tips</td>
		</tr>
		<tr>
			<td>SMT Micro Header&#160;</td>
			<td>SAMTEC</td>
			<td>FTSH-110-01-L-DV</td>
			<td>0.3 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&#160; 00; 0.3mm Rod diameter; 0.03 mm tip width; 38 mm Length&#160; 
			Make electrode guiding holes on cuticle</td>
		</tr>
		<tr>
			<td>Tweezers</td>
			<td>DUMONT</td>
			<td>RS-5015</td>
			<td>Pattern #5; .05 X .01mm Tip Size; 110mm Length 
			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; 0.1mm Tip Width; 3&#34; Overall Length&#160; 
			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&#160;</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 cm2). Each panel has 3 lamps. 
			14 W, 549 mm x 17 mm&#160;</td>
		</tr>
	</tbody>
</table>

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

El aumento de los dispositivos electrónicos digitales de radio habilitado ha impulsado el uso de pequeñas grabadoras neuromusculares inalámbricas y estimuladores para el estudio de comportamiento de los insectos en vuelo. Esta tecnología permite el desarrollo de un sistema híbrido de insectos-máquina mediante una plataforma de insectos que viven descrito en este protocolo. Además, este protocolo presenta la configuración del sistema y los procedimientos experimentales de vuelo libre para la evaluación de la función de los músculos de vuelo en un insecto sin ataduras. Para la demostración, nos centramos en el tercer músculo axilar sclerite (3AX) para controlar y lograr girando a la izquierda oa la derecha de un escarabajo volador. Un electrodo de alambre de plata fina fue implantado en el músculo 3AX en cada lado del escarabajo. Estos se conectan a las salidas de una mochila inalámbrica (es decir, un estimulador neuromuscular eléctrica) montado en el pronoto del escarabajo. El músculo se estimuló en vuelo libre, alternando el lado de estimulación (izquierda o derecha) o variando la stimulation frecuencia. El escarabajo se volvió hacia el lado ipsilateral cuando el músculo se estimuló y exhibió una respuesta gradual a una frecuencia creciente. El proceso de implantación y calibración del volumen del sistema de cámara de captura de movimiento dimensional 3 deben llevarse a cabo con cuidado para evitar dañar el músculo y perder la pista del marcador, respectivamente. Este método es altamente beneficioso para estudiar el vuelo de insectos, ya que ayuda a revelar las funciones del músculo de vuelo de interés en vuelo libre.

El procedimiento de implantación de electrodos se presenta en la Figura 2 electrodos de hilo de plata fina se implantaron en el músculo 3AX del escarabajo través de pequeños agujeros perforados en la cutícula suave en el músculo (figuras 2d - e).. Esta cutícula suave se encuentra justo por encima del apodema del músculo basalar después de la eliminación de la parte anterior de la metepisternum (Figuras 2d - <st...

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.

Sistema híbrido de insectos-máquina: radio control remoto de un escarabajo que vuela libremente (<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 vistas.  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

Localizing Protein in 3D Neural Stem Cell Culture: a Hybrid Visualization Methodology

The importance of 3-dimensional (3D) topography in influencing neural stem and progenitor cell (NPC) phenotype is widely acknowledged yet challenging to study. When dissociated from embryonic or post-natal brain, single NPCs will proliferate in suspension to form neurospheres. Daughter cells within these cultures spontaneously adopt distinct developmental lineages (neurons, oligodendrocytes, and astrocytes) over the course of expansion despite being exposed to the same extracellular milieu. This progression recapitulates many of the stages observed over the course of neurogenesis and gliogenesis in post-natal brain and is often used to study basic NPC biology within a controlled environment. Assessing the full impact of 3D topography and cellular positioning within these cultures on NPC fate is, however, difficult. To localize target proteins and identify NPC lineages by immunocytochemistry, free-floating neurospheres must be plated on a substrate or serially sectioned. This processing is required to ensure equivalent cell permeabilization and antibody access throughout the sphere. As a result, 2D epifluorescent images of cryosections or confocal reconstructions of 3D Z-stacks can only provide spatial information about cell position within discrete physical or digital 3D slices and do not visualize cellular position in the intact sphere. Here, to reiterate the topography of the neurosphere culture and permit spatial analysis of protein expression throughout the entire culture, we present a protocol for isolation, expansion, and serial sectioning of post-natal hippocampal neurospheres suitable for epifluorescent or confocal immunodetection of target proteins. Connexin29 (Cx29) is analyzed as an example. Next, using a hybrid of graphic editing and 3D modelling softwares rigorously applied to maintain biological detail, we describe how to re-assemble the 3D structural positioning of these images and digitally map labelled cells within the complete neurosphere. This methodology enables visualization and analysis of the cellular position of target proteins and cells throughout the entire 3D culture topography and will facilitate a more detailed analysis of the spatial relationships between cells over the course of neurogenesis and gliogenesis in vitro.
Both Imbeault and Valenzuela contributed equally and should be considered joint first authors.

Here, we describe how to produce, expand, and immunolabel postnatal hippocampal neural progenitor cells (NPCs) in three-dimensional (3D) culture. Next, using hybrid visualization technologies, we demonstrate how digital images of immunolabelled cryosections can be used to reconstruct and map the spatial position of immunopositive cells throughout the entire 3D neurosphere.

La localización de la proteína en 3D Cultura de células madre neurales: una metodología de visualización híbrida

The importance of 3-dimensional (3D) topography in influencing neural stem and progenitor cell (NPC) phenotype is widely acknowledged yet challenging to ...

Investigación

Neurociencias

Long-term Behavioral Tracking of Freely Swimming Weakly Electric Fish

Long-term behavioral tracking can capture and quantify natural animal behaviors, including those occurring infrequently. Behaviors such as exploration and social interactions can be best studied by observing unrestrained, freely behaving animals. Weakly electric fish (WEF) display readily observable exploratory and social behaviors by emitting electric organ discharge (EOD). Here, we describe three effective techniques to synchronously measure the EOD, body position, and posture of a free-swimming WEF for an extended period of time. First, we describe the construction of an experimental tank inside of an isolation chamber designed to block external sources of sensory stimuli such as light, sound, and vibration. The aquarium was partitioned to accommodate four test specimens, and automated gates remotely control the animals&#39; access to the central arena. Second, we describe a precise and reliable real-time EOD timing measurement method from freely swimming WEF. Signal distortions caused by the animal&#39;s body movements are corrected by spatial averaging and temporal processing stages. Third, we describe an underwater near-infrared imaging setup to observe unperturbed nocturnal animal behaviors. Infrared light pulses were used to synchronize the timing between the video and the physiological signal over a long recording duration. Our automated tracking software measures the animal&#39;s body position and posture reliably in an aquatic scene. In combination, these techniques enable long term observation of spontaneous behavior of freely swimming weakly electric fish in a reliable and precise manner. We believe our method can be similarly applied to the study of other aquatic animals by relating their physiological signals with exploratory or social behaviors.

We describe a set of techniques for studying spontaneous behavior of freely swimming weakly electric fish over an extended period of time, by synchronously measuring the animal&#39;s electric organ discharge timing, body position and posture both accurately and reliably in a specially designed aquarium tank inside a sensory isolation chamber.

Seguimiento del comportamiento a largo plazo de Libremente Natación débilmente eléctrico Fish

Long-term behavioral tracking can capture and quantify natural animal behaviors, including those occurring infrequently. Behaviors such as exploration and ...

A Simplified Method for Ultra-Low Density, Long-Term Primary Hippocampal Neuron Culture

Culturing primary hippocampal neurons in vitro facilitates mechanistic interrogation of many aspects of neuronal development. Dissociated embryonic hippocampal neurons can often grow successfully on glass coverslips at high density under serum-free conditions, but low density cultures typically require a supply of trophic factors by co-culturing them with a glia feeder layer, preparation of which can be time-consuming and laborious. In addition, the presence of glia may confound interpretation of results and preclude studies on neuron-specific mechanisms. Here, a simplified method is presented for ultra-low density (~2,000 neurons/cm2), long-term (&#62;3 months) primary hippocampal neuron culture that is under serum free conditions and without glia cell support. Low density neurons are grown on poly-D-lysine coated coverslips, and flipped on high density neurons grown in a 24-well plate. Instead of using paraffin dots to create a space between the two neuronal layers, the experimenters can simply etch the plastic bottom of the well, on which the high density neurons reside, to create a microspace conducive to low density neuron growth. The co-culture can be easily maintained for &#62;3 months without significant loss of low density neurons, thus facilitating the morphological and physiological study of these neurons. To illustrate this successful culture condition, data are provided to show profuse synapse formation in low density cells after prolonged culture. This co-culture system also facilitates the survival of sparse individual neurons grown in islands of poly-D-lysine substrates and thus the formation of autaptic connections.

Low density cultures of primary hippocampal neurons usually require glia feeder layer to supply neurotrophic factors and sustain longevity. We describe here a simplified method to culture ultra-low density neurons on glass coverslips in the presence of a high density neuronal feeder layer, which facilitates investigation of specific neuronal-autonomous mechanisms.

Un método simplificado para determinar la densidad ultra-baja, a largo plazo del hipocampo primaria Neurona Cultura

Culturing primary hippocampal neurons in vitro facilitates mechanistic interrogation of many aspects of neuronal development. Dissociated embryonic ...

Optical Control of a Neuronal Protein Using a Genetically Encoded Unnatural Amino Acid in Neurons

Photostimulation is a noninvasive way to control biological events with excellent spatial and temporal resolution. New methods are desired to photo-regulate endogenous proteins expressed in their native environment. Here, we present an approach to optically control the function of a neuronal protein directly in neurons using a genetically encoded unnatural amino acid (Uaa). By using an orthogonal tRNA/aminoacyl-tRNA synthetase pair to suppress the amber codon, a photo-reactive Uaa 4,5-dimethoxy-2-nitrobenzyl-cysteine (Cmn) is site-specifically incorporated in the pore of a neuronal protein Kir2.1, an inwardly rectifying potassium channel. The bulky Cmn physically blocks the channel pore, rendering Kir2.1 non-conducting. Light illumination instantaneously converts Cmn into a smaller natural amino acid Cys, activating Kir2.1 channel function. We express these photo-inducible inwardly rectifying potassium (PIRK) channels in rat hippocampal primary neurons, and demonstrate that light-activation of PIRK ceases the neuronal firing due to the outflux of K+ current through the activated Kir2.1 channels. Using in utero electroporation, we also express PIRK in the embryonic mouse neocortex in vivo, showing the light-activation of PIRK in neocortical neurons. Genetically encoding Uaa imposes no restrictions on target protein type or cellular location, and a family of photoreactive Uaas is available for modulating different natural amino acid residues. This technique thus has the potential to be generally applied to many neuronal proteins to achieve optical regulation of different processes in brains. The current protocol presents an accessible procedure for intricate Uaa incorporation in neurons in vitro and in vivo to achieve photo control of neuronal protein activity on the molecular level.

Here, a procedure to selectively activate a neuronal protein with a short pulse of light by genetically encoding a photo-reactive unnatural amino acid into a target neuronal protein expressed in neurons in culture or in vivo is presented.

Control óptico de una proteína neuronal utilizando un ácido Unnatural Amino genéticamente codificados en las neuronas

Photostimulation is a noninvasive way to control biological events with excellent spatial and temporal resolution. New methods are desired to ...

Neuropharmacological Manipulation of Restrained and Free-flying Honey Bees, Apis mellifera

Honey bees demonstrate astonishing learning abilities and advanced social behavior and communication. In addition, their brain is small, easy to visualize and to study. Therefore, bees have long been a favored model amongst neurobiologists and neuroethologists for studying the neural basis of social and natural behavior. It is important, however, that the experimental techniques used to study bees do not interfere with the behaviors being studied. Because of this, it has been necessary to develop a range of techniques for pharmacological manipulation of honey bees. In this paper we demonstrate methods for treating restrained or free-flying honey bees with a wide range of pharmacological agents. These include both noninvasive methods such as oral and topical treatments, as well as more invasive methods that allow for precise drug delivery in either systemic or localized fashion. Finally, we discuss the advantages and disadvantages of each method and&#160;describe common hurdles and how to best overcome them. We conclude with a discussion on the importance of adapting the experimental method to the biological questions rather than the other way around.

Neuropharmacological Manipulation of Restrained and Free-flying Honey Bees, Apis mellifera

This manuscript describes several protocols for administering pharmacological agents to honey bees, including simple noninvasive methods for free-flying bees, as well as more invasive variants that allow precise localized treatment of restrained bees.

La manipulación de neurofarmacológico restringida y de vuelo libre abejas de la miel,<em&gt; Apis mellifera</em

Honey bees demonstrate astonishing learning abilities and advanced social behavior and communication. In addition, their brain is small, easy to visualize ...

Recording EEG in Freely Moving Neonatal Rats Using a Novel Method

EEG is a useful method to detect electrical activity in the brain. Moreover, it is a widely used diagnostic tool for various neurological conditions, such as epilepsy and neurodegenerative disorders. However, it is technically difficult to obtain EEG recordings in neonates as it requires specialized handling and great care. Here, we present a novel method to record EEG in neonatal rat pups (P8-P15). We designed a simple and reliable electrode using computer pin loci; it can be easily implanted into the skull of a rat pup to record high-quality EEG signals in the normal and epileptic brain. Pups were given an intraperitoneal (i.p.) injection of the neurotoxin kainic acid (KA) to induce epileptic seizures. The surgical implantation performed in this procedure is less expensive than other EEG procedures for neonates. This method allows one to record high-quality and stable EEG signals for more than 1 week. Furthermore, this procedure can also be applied to adult rats and mice to study epilepsy or other neurological disorders.

Here, we introduce a novel technique designed to record electroencephalography (EEG) in freely moving neonatal epileptic pups and describe its procedures, features, and applications. This method allows one to record EEG for more than 1 week.

Grabación de EEG en Ratas Neonatales que se mueven libremente usando un método nuevo

EEG is a useful method to detect electrical activity in the brain. Moreover, it is a widely used diagnostic tool for various neurological conditions, such ...

Noninvasive EEG Recordings from Freely Moving Piglets

The method allows the recording of high-quality electroencephalograms (EEGs) from freely moving piglets directly in the pigpen. We use a one-channel telemetric electroencephalography system in combination with standard self-adhesive hydrogel electrodes. The piglets are calmed down without the use of sedatives. After their release into the pigpen, the piglets behave normally&#8212;they drink and sleep in the same cycle as their siblings. Their sleep phases are used for the EEG recordings.

Here, we present a protocol to record telemetric electroencephalograms (EEGs) from freely moving piglets directly in the pigpen without the use of a sedative, making it possible to record typical EEG patterns during non-REM sleep, like spindle bursts.

Grabaciones de EEG no invasivo se mueva libremente lechones

The method allows the recording of high-quality electroencephalograms (EEGs) from freely moving piglets directly in the pigpen. We use a one-channel ...

Long-term Sensory Conflict in Freely Behaving Mice

Long-term sensory conflict protocols are a valuable means of studying motor learning. The presented protocol produces a persistent sensory conflict for experiments aimed at studying long-term learning in mice. By permanently wearing a device fixed on their heads, mice are continuously exposed to a sensory mismatch between visual and vestibular inputs while freely moving in home cages. Therefore, this protocol readily enables the study of the visual system and multisensory interactions over an extended timeframe that would not be accessible otherwise. In addition to lowering the experimental costs of long-term sensory learning in naturally behaving mice, this approach accommodates the combination of in vivo and in vitro experiments. In the reported example, video-oculography is performed to quantify the vestibulo-ocular reflex (VOR) and optokinetic reflex (OKR) before and after learning. Mice exposed to this long-term sensory conflict between visual and vestibular inputs presented a strong VOR gain decrease but exhibited few OKR changes. Detailed steps of device assembly, animal care, and reflex measurements are hereby reported.

The presented protocol produces a persistent sensory conflict for experiments aimed at studying long-term learning. By permanently wearing a fixed device on their heads, mice are continuously exposed to a sensory mismatch between visual and vestibular inputs while freely moving in home cages.

A largo plazo conflicto sensorial en comportarse libremente ratones

Long-term sensory conflict protocols are a valuable means of studying motor learning. The presented protocol produces a persistent sensory conflict for ...

Hybrid Microdrive System with Recoverable Opto-Silicon Probe and Tetrode for Dual-Site High Density Recording in Freely Moving Mice

Multi-regional neural recordings can provide crucial information to understanding fine-timescale interactions between multiple brain regions. However, conventional microdrive designs often only allow use of one type of electrode to record from single or multiple regions, limiting the yield of single-unit or depth profile recordings. It also often limits the ability to combine electrode recordings with optogenetic tools to target pathway and/or cell type specific activity. Presented here is a hybrid microdrive array for freely moving mice to optimize yield and a description of its fabrication and reuse of the microdrive array. The current design employs nine tetrodes and one opto-silicon probe implanted in two different brain areas simultaneously in freely moving mice. The tetrodes and the opto-silicon probe are independently adjustable along the dorsoventral axis in the brain to maximize the yield of unit and oscillatory activities. This microdrive array also incorporates a set-up for light, mediating optogenetic manipulation to investigate the regional- or cell type-specific responses and functions of long-range neural circuits. In addition, the opto-silicon probe can be safely recovered and reused after each experiment. Because the microdrive array consists of 3D-printed parts, the design of microdrives can be easily modified to accommodate various settings. First described is the design of the microdrive array and how to attach the optical fiber to a silicon probe for optogenetics experiments, followed by fabrication of the tetrode bundle and implantation of the array into a mouse brain. The recording of local field potentials and unit spiking combined with optogenetic stimulation also demonstrate feasibility of the microdrive array system in freely moving mice.

This protocol describes the construction of a hybrid microdrive array that allows implantation of nine independently adjustable tetrodes and one adjustable opto-silicon probe in two brain regions in freely moving mice. Also demonstrated is a method for safely recovering and reusing the opto-silicon probe for multiple purposes.

Sistema microdrive híbrido con sonda de optosilicio recuperable y tetroda para grabación de alta densidad de doble sitio en ratones en movimiento libre

Multi-regional neural recordings can provide crucial information to understanding fine-timescale interactions between multiple brain regions. However, ...

Wireless Electrophysiological Recording of Neurons by Movable Tetrodes in Freely Swimming Fish

The neural mechanisms governing fish behavior remain mostly unknown, although fish constitute the majority of all vertebrates. The ability to record brain activity from freely moving fish would advance research on the neural basis of fish behavior considerably. Moreover, precise control of the recording location in the brain is critical to studying coordinated neural activity across regions in fish brain. Here, we present a technique that records wirelessly from the brain of freely swimming fish while controlling for the depth of the recording location. The system is based on a neural logger associated with a novel water-compatible implant that can adjust the recording location by microdrive-controlled tetrodes. The capabilities of the system are illustrated through recordings from the telencephalon of goldfish.

A novel wireless technique for recording extracellular neural signals from the brain of freely swimming goldfish is presented. The recording device is composed of two tetrodes, a microdrive, a neural data logger, and a waterproof case. All parts are custom-made except for the data logger and its connector.

Grabación electrofisiológica inalámbrica de las neuronas por tetrodes móviles en la natación libre de peces

The neural mechanisms governing fish behavior remain mostly unknown, although fish constitute the majority of all vertebrates. The ability to record brain ...

Sistema híbrido de insectos-máquina: radio control remoto de un escarabajo que vuela libremente (

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