Name: sampling and analysis of animal scent signals
Uploaded: 2021-02-13T12:30:00
Description: Watch this Scientific Journal Video about Sampling and Analysis of Animal Scent Signals at JoVE.com

Мы благодарим Keith Holding за его помощь в химическом анализе в Научном центре Розалинд Франклин, Вулверхэмптон, и Бена Мантла за производство видео. Мы также благодарны профессору Глориано Монети, доктору Джузеппе Пьераччини и членам Центра масс-спектрометрии Университета Флоренции во Флоренции, а также профессору Луке Каламаю и доктору Марко Микелоцци из лаборатории ARCA CNR, Флоренция, за их помощь в создании этой методологии. Исследовательские проекты, которые включали методы отбора проб и анализа, описанные в рукописи, были поддержаны двумя внутриевропейскими стипендиями Марии Склодовской-Кюри (идентификаторы грантового соглашения: 327083, 703611), небольшим грантом(«Сенсорно обогащенный примат»)от Общества приматов Великобритании и небольшим исследовательским грантом(«Имеют ли охотники-собиратели особое обоняние?»)от Британской академии / The Leverhulme Trust для S.V. Лабораторные работы, необходимые для создания этой методологии, также получили финансирование от Ежегодного конкурса финансирования факультета науки и техники (Вулверхэмптон) для S.V.

1. Сбор образцов
<ol><li>Образцы запахов, которые являются одним из следующих:<ol>
<li>Собирать спонтанно высвобождаемые привычными субъектами исследования (например, приматами зоопарка) через ароматизаторы на стерильной фильтровальной бумаге (например, выделения запаха паховой железы) или...

<ol>
	<li>Wyatt, T. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Pheromones and Animal Behavior: Chemical Signals and Signatures. , (2014).</li><li>Heymann, E. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+neglected+sense-olfaction+in+primate+behavior,+ecology,+and+evolution.">The neglected sense-olfaction in primate behavior, ecology, and evolution.</a> American Journal of Primatology. 68 (6), 519-524 (2006).</li><li>Drea, C. M., Boulet, M., DelBarco-Trillo, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+"secret"+in+secretions:+Methodological+considerations+in+deciphering+primate+olfactory+communication.">The "secret" in secretions: Methodological considerations in deciphering primate olfactory communication.</a> American Journal of Primatology. 75 (7), 621-642 (2013).</li><li>Vaglio, 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=Sternal+gland+scent-marking+signals+sex,+age,+rank+and+group+identity+in+captive+mandrills.">Sternal gland scent-marking signals sex, age, rank and group identity in captive mandrills.</a> Chemical Senses. 41 (2), 177-186 (2016).</li><li>Marneweck, C., J&#252;rgens, A., Shrader, A. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dung+odours+signal+sex,+age,+territorial+and+oestrous+state+in+white+rhinos.">Dung odours signal sex, age, territorial and oestrous state in white rhinos.</a> Proceedings of the Royal Society of London B: Biological Sciences. 284 (1846), (2016).</li><li>Shear, W. A., Jones, T. H., Miras, H. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+possible+phylogenetic+signal+in+milliped+chemical+defenses.">A possible phylogenetic signal in milliped chemical defenses.</a> Biochemical Systematics and Ecology. 35, 838-842 (2007).</li><li>Kimura, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Volatile+substances+in+feces,+urine+and+urine-marked+feces+of+feral+horses.">Volatile substances in feces, urine and urine-marked feces of feral horses.</a> Canadian Journal of Animal Science. 81 (3), 411-420 (2001).</li><li>Vaglio, S., Minicozzi, P., Bonometti, E., Mello, G., Chiarelli, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Volatile+signals+during+pregnancy:+a+possible+chemical+basis+for+mother-infant+recognition.">Volatile signals during pregnancy: a possible chemical basis for mother-infant recognition.</a> Journal of Chemical Ecology. 35 (1), 131-139 (2009).</li><li>Setchell, J. 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=Chemical+composition+of+scent-gland+secretions+in+an+Old+World+monkey+(Mandrillus+sphinx):+influence+of+sex,+male+status,+and+individual+identity.">Chemical composition of scent-gland secretions in an Old World monkey (Mandrillus sphinx): influence of sex, male status, and individual identity.</a> Chemical Senses. 35 (3), 205-220 (2010).</li><li>Setchell, J. 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=Odour+signals+MHC+genotype+in+an+Old+World+monkey.">Odour signals MHC genotype in an Old World monkey.</a> Proceedings of the Royal Society of London B: Biological Sciences. 278 (1703), 274-280 (2011).</li><li>Pawliszyn, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Solid phase microextraction: theory and practice. , (1997).</li><li>Janda, E. D., Perry, K., Hankinson, E., Walker, D., Vaglio, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sex+differences+in+scent-marking+in+captive+red-ruffed+lemurs.">Sex differences in scent-marking in captive red-ruffed lemurs.</a> American Journal of Primatology. 81 (1), 22951 (2019).</li></ol>

Использование контрольных образцов, как экологических контрольных образцов, созданных во время отбора проб, так и системных заготовок, имеет решающее значение для интерпретации образцов с запаховым метками. Любые пики, приписываемые среде выборки или инструментальной системе, должны...

Очень мало известно о химических изменениях, лежащих в основе обонятельных сигналов у животных1,в том же время из-за методологических проблем в регистрации и количественной оценке летучих химических профилей запахов2. Существует несколько потенциальных ловушек при работе с очень сложными химическими матрицами; к ним относятся при отборе проб и анализе запаха образцов3.
В Научном центре Розалинд Франклин Университета Вулверхэмптона мы проводим анализ запахов и запахов, чтобы понять, как они могут использоваться животными. Мы объединяем семиохимию с поведенческой экологией, эндокринологией и цитологией, чтобы улучшить наше понимание роли обонятельных сигналов в общении животных.
Мы разработали методологию, а затем проанализировали запахи и маркировку различных видов, включая нескольких нечеловеческих приматов (то есть коронованных лемуров, красно-взъерошенные лемуры, японские макаки, оливковые бабуины, шимпанзе) и других млекопитающих (то есть кошек, коров). Мы собрали и проанализировали различные образцы, включая мочу, фекалии, волосы и выделения запаха пахлочек. Эти запахи и запаховые метки состоят из сложных смесей соединений, и поэтому любая методология, используемая для их анализа, должна включать некоторую форму сепараторного метода. Как показано на рисунке, они также встречаются в ряде матриц, что обусловливает необходимость использования методов извлечения интересующих компонентов.
Предыдущие исследования Vaglio et al.4 и других авторов5 использовали динамическую экстракцию в пространстве над головой (DHS) с помощью газовой хроматографии-масс-спектрометрии (GC-MS), в то время как прямая экстракция растворителем6 и комплексная экстракция растворителем7 также использовались. В частности, динамический отбор проб в пространстве над головой включает очистку пространства над головой известным объемом инертного газа, который в конечном итоге удаляет все летучие соединения, за исключением тех, которые демонстрируют сильное сродство к матрице образца (например, полярные соединения в водных образцах).
Для текущей методологии мы приняли метод твердофазной микроэкстракции пространства над головой (HS-SPME) в сочетании с GC-MS. В частности, мы разработали и усовершенствовали методологию, уже используемую Vaglio et al. в своей предыдущей лаборатории GC-MS8,9,10.
Методы экстракции без растворителя очень эффективны для анализа небольших, высоколетучих соединений (которые в противном случае могут быть легко потеряны из образца), поскольку эти методы иммобилизуют соединения на стабильной твердофазной опоре. HS-SPME использует волокно, покрытое адсорбентным полимером, для захвата летучих соединений в пространстве образца или для извлечения растворенных соединений путем погружения в водную биологическую жидкость11. Полимерное покрытие не связывает соединения сильно, поэтому нагреванием в инжекционном порту ГК их можно удалить. Этот метод является более мощным, чем методы экстракции растворителем, а также более эффективным, чем DHS.
При нынешнем подходе образцы содержатся в стеклянных флаконах. Эти флаконы нагревают до температуры 40 °C для имитации температуры тела животного, чтобы способствовать тому, чтобы летучие компоненты аромата занимали пространство головы флакона. Волокно SPME, покрытое 65 мкм сорбентного материала полидиметилсилоксана/дивинилбензола (PDMS/DVB), подвергается воздействию среды пространства над головой, и летучие компоненты из образца адсорбируются на волокно. При нагревании волокна во входном порту GC-MS летучие компоненты десорбируются от волокна, а затем отделяются GC. Паттерны масс-спектральной фрагментации получены для каждого компонента с использованием MS. Сравнивая эти масс-спектры с масс-спектральными базами данных, можно предварительно определить компоненты ароматического знака. Благодаря использованию автоматического пробоотборщика мы можем последовательно анализировать несколько образцов партиями.
Учитывая, что каждый тип волокна SPME имеет различное сродство с полярными химическими веществами, волокно обычно выбирают в зависимости от полярности и/или молекулярной массы целевых химических соединений. Кроме того, условия ГК изменяются в зависимости от типа колонки ГК и характеристик целевых химических соединений.
Этот метод позволяет проводить полуциплиматический анализ летучих компонентов запаховой маркировки, позволяя разделять и предварительно идентифицировать компоненты в образце с последующим анализом коэффициентов пиковой площади для поиска тенденций, которые могли бы означать компоненты ароматовой маркировки, которые могут быть вовлечены в сигнализацию.
Ключевыми сильными сторонами этого нынешнего подхода являются:
<ul style="list-style-type: square;"><li>Диапазон типов образцов, которые могут быть проанализированы.</li>
	<li>Не требуется сложная пробоподготовка или экстракция.</li>
	<li>Возможность анализа летучих компонентов.</li>
	<li>Возможность разделения компонентов смеси.</li>
	<li>Чтобы иметь возможность идентифицировать обнаруженные компоненты.</li>
	<li>Способность предоставлять полу количественное и потенциально количественное представление об обнаруженных компонентах.</li>
</ul>

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>10 mL autosampler vials</td>
			<td>Agilent</td>
			<td>5188-5392</td>
			<td>10 ml screwtop vials with</td>
		</tr>
		<tr>
			<td>18 mm vial caps</td>
			<td>Agilent</td>
			<td>8010-0139</td>
			<td>Magnetic with PTFE/silicone septa</td>
		</tr>
		<tr>
			<td>Autosampler</td>
			<td>Agilent</td>
			<td>GC120 PAL autosampler</td>
			<td></td>
		</tr>
		<tr>
			<td>Capillary column</td>
			<td>Agilent</td>
			<td>HP5-MS</td>
			<td>30 m x 0.25 mm; 0.25 &#181;m</td>
		</tr>
		<tr>
			<td>Data analysis software</td>
			<td>Agilent</td>
			<td>-</td>
			<td>ChemStation</td>
		</tr>
		<tr>
			<td>Gas Chromatograph</td>
			<td>Agilent</td>
			<td>7890B</td>
			<td></td>
		</tr>
		<tr>
			<td>Inlet septa</td>
			<td>Agilent</td>
			<td>5182-3442</td>
			<td>Merlin microseal</td>
		</tr>
		<tr>
			<td>Mass Selective Detector</td>
			<td>Agilent</td>
			<td>5977A</td>
			<td></td>
		</tr>
		<tr>
			<td>Reporting software</td>
			<td>Microsoft</td>
			<td>-</td>
			<td>Excel</td>
		</tr>
		<tr>
			<td>Spectral library</td>
			<td>NIST</td>
			<td>-</td>
			<td>NIST/EPA/NIH Mass Spectral Library</td>
		</tr>
		<tr>
			<td>Spectral library search program</td>
			<td>NIST</td>
			<td>-</td>
			<td>MS Search v.2.2</td>
		</tr>
		<tr>
			<td>Splitless Inlet liner</td>
			<td>Agilent</td>
			<td>5190-4048</td>
			<td></td>
		</tr>
		<tr>
			<td>SPME fibres</td>
			<td>Agilent</td>
			<td>SU57345U</td>
			<td>65 &#181;m PDMS/DVB fibre</td>
		</tr>
	</tbody>
</table>

sampling and analysis of animal scent signals

Мы разработали эффективную методологию отбора проб и анализа сигналов запаха, используя твердофазную микроэкстракцию пространства над головой в сочетании с газовой хроматографией-масс-спектрометрией, чтобы понять, как они могут быть использованы в общении животных. Этот метод позволяет проводить полуциплиматический анализ летучих компонентов секреций запаха, позволяя разделять и предварительно идентифицировать компоненты в образце с последующим анализом соотношений пиковой площади для поиска тенденций, которые могут означать соединения, которые могут быть вовлечены в передачу сигналов. Ключевыми сильными сторонами этого нынешнего подхода являются диапазон типов образцов, которые могут быть проанализированы; отсутствие необходимости в какой-либо сложной пробоподготовке или извлечении; возможность разделения и анализа компонентов смеси; идентификация обнаруженных компонентов; и способность предоставлять полу количественно-количественную и потенциально количественную информацию об обнаруженных компонентах. Основное ограничение методологии касается самих образцов. Поскольку компоненты, представляющие особый интерес, являются летучими, и они могут быть легко потеряны или их концентрации изменены, важно, чтобы образцы хранились и транспортировались надлежащим образом после их сбора. Это также означает, что условия хранения и транспортировки образцов являются относительно дорогостоящими. Этот метод может быть применен к различным образцам (включая мочу, кал, волосы и выделения запаха ароматических желез). Эти запахи состоят из сложных смесей, встречающихся в ряде матриц, и, таким образом, требуют использования методов для разделения отдельных компонентов и извлечения соединений, представляющих биологический интерес.

Следуя этому протоколу, мы предварительно идентифицировали в общей сложности 32 летучих химических соединения из анализа 14 аногелитических запаховых меток, спонтанно высвобождаемых на фильтровальной бумаге красно-взъерошенными лемурами (Varecia variegata rubra) и сравнили профили запаха с...

Watch this Scientific Journal Video about Sampling and Analysis of Animal Scent Signals at JoVE.com

Sampling and Analysis of Animal Scent Signals

Мы разработали эффективную методологию отбора проб и анализа сигналов запаха, чтобы понять, как они могут быть использованы в общении с животными. В частности, мы используем твердофазную микроэкстракцию пространства над головой в сочетании с газовой хроматографией-масс-спектрометрией для анализа летучих компонентов запахов животных и запаховых меток.

Отбор проб и анализ сигналов запаха животных

We have developed an effective methodology for sampling and analysis of odor signals, by using headspace solid-phase microextraction coupled with gas ...

sampling-and-analysis-of-animal-scent-signals

Research

JoVE Journal

Behavior

Automated, Quantitative Cognitive/Behavioral Screening of Mice: For Genetics, Pharmacology, Animal Cognition and Undergraduate Instruction

We describe a high-throughput, high-volume, fully automated, live-in 24/7 behavioral testing system for assessing the effects of genetic and pharmacological manipulations on basic mechanisms of cognition and learning in mice. A standard polypropylene mouse housing tub is connected through an acrylic tube to a standard commercial mouse test box. The test box has 3 hoppers, 2 of which are connected to pellet feeders. All are internally illuminable with an LED and monitored for head entries by infrared (IR) beams. Mice live in the environment, which eliminates handling during screening. They obtain their food during two or more daily feeding periods by performing in operant (instrumental) and Pavlovian (classical) protocols, for which we have written protocol-control software and quasi-real-time data analysis and graphing software. The data analysis and graphing routines are written in a MATLAB-based language created to simplify greatly the analysis of large time-stamped behavioral and physiological event records and to preserve a full data trail from raw data through all intermediate analyses to the published graphs and statistics within a single data structure. The data-analysis code harvests the data several times a day and subjects it to statistical and graphical analyses, which are automatically stored in the &#34;cloud&#34; and on in-lab computers. Thus, the progress of individual mice is visualized and quantified daily. The data-analysis code talks to the protocol-control code, permitting the automated advance from protocol to protocol of individual subjects. The behavioral protocols implemented are matching, autoshaping, timed hopper-switching, risk assessment in timed hopper-switching, impulsivity measurement, and the circadian anticipation of food availability. Open-source protocol-control and data-analysis code makes the addition of new protocols simple. Eight test environments fit in a 48 in x 24 in x 78 in&#160;cabinet; two such cabinets (16 environments) may be controlled by one computer.

Fully automated system for measuring physiologically meaningful properties of the mechanisms mediating spatial localization, temporal localization, duration, rate&#160;and probability estimation, risk assessment, impulsivity, and the accuracy and precision of memory, in order to assess the effects of genetic and pharmacological manipulations on foundational mechanisms of cognition in mice.

Автоматизированная, Количественный Когнитивная / Поведенческие Скрининг мышей: Для генетики, фармакологии, животных познания и Бакалавриат Инструкции

We describe a high-throughput, high-volume, fully automated, live-in 24/7 behavioral testing system for assessing the effects of genetic and ...

Measurement of Neurophysiological Signals of Ignoring and Attending Processes in Attention Control

Attention control is the ability to selectively attend to some sensory signals while ignoring others. This ability is thought to involve two processes: enhancement of sensory signals that are to be attended and the attenuation of sensory signals that are to be ignored. The overall strength of attentional modulation is often measured by comparing the amplitude of a sensory neural response to an external input when attended versus when ignored. This method is robust for detecting attentional modulation, but precludes the ability to assess the separate dynamics of attending and ignoring processes. Here, we describe methodology to measure independently the neurophysiological signals of attending and ignoring using the intermodal attention task (IMAT). This task, when combined with electroencephalography, isolates neurophysiological sensory responses in auditory and visual modalities, when either attending or ignoring, with respect to a passive control. As a result, independent dynamics of attending and of a ignoring can be assessed in either modality. Our results using this task indicate that the timing and cortical sources of attending and ignoring effects differ, as do their contributions to the attention modulation effect, pointing to unique neural trajectories and demonstrating sample utility of measuring them separately.

Attention control comprises enhancement of target signals and attenuation of distractor signals. We describe an approach to measure separately but concurrently, the neurophysiology of attending and ignoring in sustained intermodal attention, utilizing a passive control condition during which neither process is continuously engaged.

Измерение нейрофизиологических сигналах Игнорирование и приду процессов во внимание контролю

Attention control is the ability to selectively attend to some sensory signals while ignoring others. This ability is thought to involve two processes: ...

Wheel Running and Environmental Complexity as a Therapeutic Intervention in an Animal Model of FASD

Aerobic exercise (e.g., wheel running (WR) extensively used in animal research) positively impacts many measures of neuroplastic potential in the brain, such as rates of adult neurogenesis, angiogenesis, and expression of neurotrophic factors in rodents. This intervention has also been shown to mitigate behavioral and neuroanatomical aspects of the negative impacts of teratogens (i.e., developmental exposure to alcohol) and age-related neurodegeneration in rodents. Environmental complexity (EC) has been shown to produce numerous neuroplastic benefits in cortical and subcortical structures and can be coupled with wheel running to increase the proliferation and survival of new cells in the adult hippocampus. The combination of these two interventions provides a robust &#34;superintervention&#34; (WR-EC) that can be implemented in a range of rodent models of neurological disorders. We will discuss the implementation of WR/EC and its constituent interventions for use as a more powerful therapeutic intervention in rats using the animal model of prenatal exposure to alcohol in humans. We will also discuss which elements of the procedures are absolutely necessary for the interventions and which ones may be altered depending on the experimenter&#39;s question or facilities.

Cardiovascular exercise and stimulating experiences in a complex environment have positive benefits on multiple measures of neuroplasticity within the rodent brain. This article will discuss the implementation of these interventions as a &#34;superintervention&#34; which combines wheel running and environmental complexity and will address the limitations of these interventions.

Колеса Запуск и окружающей среды Сложность в качестве терапевтического вмешательства в животную модель FASD

Aerobic exercise (e.g., wheel running (WR) extensively used in animal research) positively impacts many measures of neuroplastic potential in the brain, ...

Introducing Clicker Training as a Cognitive Enrichment for Laboratory Mice

Establishing new refinement strategies in laboratory animal science is a central goal in fulfilling the requirements of Directive 2010/63/EU. Previous research determined a profound impact of gentle handling protocols on the well-being of laboratory mice. By introducing clicker training to the keeping of mice, not only do we promote the amicable treatment of mice, but we also enable them to experience cognitive enrichment. Clicker training is a form of positive reinforcement training using a conditioned secondary reinforcer, the "click" sound of a clicker, which serves as a time bridge between the strengthened behavior and an upcoming reward. The effective implementation of the clicker training protocol with a cohort of 12 BALB/c inbred mice of each sex proved to be uncomplicated. The mice learned rather quickly when challenged with tasks of the clicker training protocol, and almost all trained mice overcame the challenges they were given (100% of female mice and 83% of male mice). This study has identified that clicker training for mice strongly correlates with reduced fear in the mice during human-mice interactions, as shown by reduced anxiety-related behaviors (e.g., defecation, vocalization, and urination) and fewer depression-like behaviors (e.g., floating). By developing a reliable protocol that can be easily integrated into the daily routine of the keeping of laboratory mice, the lifetime experience of welfare in the mice can be improved substantially.

The development of new refinement strategies for laboratory mice is a challenging task that contributes towards fulfilling the 3R principle. This protocol introduces clicker training as a cognitive enrichment program for laboratory mice.

Введение в Clicker подготовки в качестве когнитивного обогащения для лабораторных мышей

Establishing new refinement strategies in laboratory animal science is a central goal in fulfilling the requirements of Directive 2010/63/EU. Previous ...

Systematic Assessment of Well-Being in Mice for Procedures Using General Anesthesia

In keeping with the 3R Principle (Replacement, Reduction, Refinement) developed by Russel and Burch, scientific research should use alternatives to animal experimentation whenever possible. When there is no alternative to animal experimentation, the total number of laboratory animals used should be the minimum needed to obtain valuable data. Moreover, appropriate refinement measures should be applied to minimize pain, suffering, and distress accompanying the experimental procedure. The categories used to classify the degree of pain, suffering, and distress are non-recovery, mild, moderate, or severe (EU Directive 2010/63). To determine which categories apply in individual cases, it is crucial to use scientifically sound tools.
The well-being-assessment protocol presented here is designed for procedures during which general anesthesia is used. The protocol focuses on home cage activity, the Mouse Grimace Scale, and luxury behaviors such as burrowing and nest building behavior as indicators of well-being. It also uses the free exploratory paradigm for trait anxiety-related behavior. Fecal corticosterone metabolites as indicators of acute stress are measured over the 24-h post-anesthetic period.
The protocol provides scientifically solid information on the well-being of mice following general anesthesia. Due to its simplicity, the protocol can easily be adapted and integrated in a planned study. Although it does not provide a scale to classify distress in categories according to the EU Directive 2010/63, it can help researchers estimate the degree of severity of a procedure using scientifically sound data. It provides a way to improve the assessment of well-being in a scientific, animal-centered manner.

We developed a protocol to assess well-being in mice during procedures using general anesthesia. A series of behavioral parameters indicating levels of well-being as well as glucocorticoid metabolites were analyzed. The protocol can serve as a general aid to estimate the degree of severity in a scientific, animal-centered manner.

Систематическая оценка благосостояния в мышах для процедур с использованием общего наркоза

In keeping with the 3R Principle (Replacement, Reduction, Refinement) developed by Russel and Burch, scientific research should use alternatives to animal ...

Tickling, a Technique for Inducing Positive Affect When Handling Rats

Handling small animals such as rats can lead to several adverse effects. These include the fear of humans, resistance to handling, increased injury risk for both the animals and the hands of their handlers, decreased animal welfare, and less valid research data. To minimize negative effects on experimental results and human-animal relationships, research animals are often habituated to being handled. However, the methods of habituation are highly variable and often of limited effectiveness. More potently, it is possible for humans to mimic aspects of the animals&#39; playful rough-and-tumble behavior during handling. When applied to laboratory rats in a systematic manner, this playful handling, referred to as tickling, consistently gives rise to positive behavioral responses. This article provides a detailed description of a standardized rat tickling technique. This method can contribute to future investigations into positive affective states in animals, make it easier to handle rats for common husbandry activities such as cage changing or medical/research procedures such as injection, and be implemented as a source of social enrichment. It is concluded that this method can be used to efficiently and practicably reduce rats&#39; fearfulness of humans and improve their welfare, as well as reliably model positive affective states.

This article demonstrates the standardized application of playful handling, a tickling technique designed to mimic rat rough-and-tumble play. This technique is effective at reducing fearful reactions to humans and generating positive affect when rats are handled for common husbandry activities and medical and research procedures such as injection.

Щекотка, техника для стимулирования положительное влияние при обработке крысы

Handling small animals such as rats can lead to several adverse effects. These include the fear of humans, resistance to handling, increased injury risk ...

Methods of Pairing and Pair Maintenance of New Zealand White Rabbits (Oryctolagus Cuniculus) Via Behavioral Ethogram, Monitoring, and Interventions

New Zealand White (NZW) laboratory rabbits (Oryctolagus cuniculus), as well as their ancestors the European Rabbit, are a social species that exhibit numerous benefits to being housed accordingly. Although these rabbits are innately gregarious, certain behaviors can still arise when kept in captivity, which if left unchecked, can confound research results or lead to wounding, which in extreme cases can be severe. To prevent these issues, there must be a well-structured plan for the monitoring and maintenance of paired laboratory rabbits. The purpose of this protocol is to present effective procedures for establishing newly paired NZW rabbits as well as methods for successful maintenance. Multiple methods have been tested for the creation of newly paired female rabbits from the vendor, but the most efficacious technique emphasizes capitalizing on the stress bonding from transport, urine marking, pairing in a neutral cage with no forced sharing of resources and a system of monitoring and intervention. To determine the best method of housing paired rabbits in a standard caging environment, data were collected to generate a behavioral ethogram. Behaviors were then quantified as positive, neutral or negative and were tracked across the lifespan of the pair to determine which behaviors indicated pair success or failure. With the newfound knowledge of socially housed laboratory NZW rabbit behavior, enrichment intervention was applied to alleviate aggression and prevent wounding, thus resulting in a higher percentage of successful pairs. Through several years of trialing different pairing methods, the development of the ethogram and the resulting enrichment interventions, understanding of the highly complex social constructs that dominate pair housed rabbit behavior has dramatically increased and allowed for the provision of more species-specific care and increased standards of welfare.

Methods of Pairing and Pair Maintenance of New Zealand White Rabbits Oryctolagus Cuniculus Via Behavioral Ethogram, Monitoring, and Interventions

Though European rabbits are a social species, socially housing them can be challenging. Therefore, there must be a thorough understanding of behaviors and social structures of pair-housed laboratory rabbits. Here we present a protocol to identify pairing methods, species-typical hierarchy establishment behaviors and behaviors that warrant appropriate intervention.

Методы спаривания и пара обслуживание Новой Зеландии белый кролик (ареала Cuniculus) через поведенческие этограмма, мониторинг и вмешательства

New Zealand White (NZW) laboratory rabbits (Oryctolagus cuniculus), as well as their ancestors the European Rabbit, are a social species that exhibit ...

Key Elements of Photo Attraction Bioassay for Insect Studies or Monitoring Programs

Optimized visual attractants will increase insect trapping efficiency by using the target insect&#39;s innate behaviors (positive photo-taxis) as a means to lure the insect into a population control or monitoring trap. Light emitting diodes (LEDs) have created customizable lighting options with specific wavelengths (colors), intensities, and bandwidths, all of which can be customized to the target insects. Photo-attraction behavioral bioassays can use LEDs to optimize the attractive color(s) for an insect species down to specific life history stages or behaviors (mating, feeding, or seeking shelter). Researchers must then confirm the bioassay results in the field and understand the limited attractive distance of the visual attractants.
The cloverleaf bioassay arena is a flexible method to assess photo attraction while also assessing a range of natural insect behaviors such as escape and feeding responses. The arena can be used for terrestrial or aerial insect experiments, as well as diurnal, and nocturnal insects. Data collection techniques with the arena are videotaping, counting contact with the lights, or physically collecting the insects as they are attracted towards the lights. The assay accounts for insects that make no-choice and the arenas can be single (noncompetitive) color or multiple (competitive) colors. The cloverleaf design causes insects with strong thigmotaxis to return to the center of the arena where they can view all the options in a competitive LED tests. The cloverleaf arena presented here has been used with mosquitoes, bed bugs, Hessian fly, house flies, biting midges, red flour beetles, and psocids. Bioassays are used to develop accurate and effective insect traps to guide the development and optimization of insect traps used to monitor pest population fluctuations for disease vector risk assessments, the introduction of invasive species, and/or be used for population suppression.

Photo-attraction bioassay arenas are used to determine the optimal light color(s) to maximize insect attraction; however bioassays and methods are specific to target insect behaviors and habitats. Customizable equipment and modifications are explained for nocturnal or diurnal and terrestrial or aerial insects.

Ключевые элементы Фото Аттракцион биопроб для насекомых исследования и мониторинг программ

Optimized visual attractants will increase insect trapping efficiency by using the target insect&#39;s innate behaviors (positive photo-taxis) as a means ...

Multi-Modal Signals for Analyzing Pain Responses to Thermal and Electrical Stimuli

The assessment of pain relies mostly on methods that require a person to communicate. However, for people with cognitive and verbal impairments, existing methods are not sufficient as they lack reliability and validity. To approach this problem, recent research focuses on an objective pain assessment facilitated by parameters of responses derived from physiology, and video and audio signals. To develop reliable automated pain recognition systems, efforts have been made in creating multimodal databases in order to analyze pain and detect valid pain patterns. While the results are promising, they only focus on discriminating pain or pain intensities versus no pain. In order to advance this, research should also consider the quality and duration of pain as they provide additional valuable information for more advanced pain management. To complement existing databases and the analysis of pain regarding quality and length, this paper proposes a psychophysiological experiment to elicit, measure, and collect valid pain reactions. Participants are subjected to painful stimuli that differ in intensity (low, medium, and high), duration (5 s / 1 min), and modality (heat / electric pain) while audio, video (e.g., facial expressions, body gestures, facial skin temperature), and physiological signals (e.g., electrocardiogram [ECG], skin conductance level [SCL], facial electromyography [EMG], and EMG of M. trapezius) are being recorded. The study consists of a calibration phase to determine a subject&#8217;s individual pain range (from low to intolerable pain) and a stimulation phase in which pain stimuli, depending on the calibrated range, are applied. The obtained data may allow refining, improving, and evaluating automated recognition systems in terms of an objective pain assessment. For further development of such systems and to investigate pain reactions in more detail, additional pain modalities such as pressure, chemical, or cold pain should be included in future studies. Recorded data of this study will be released as the &#8220;X-ITE Pain Database&#8221;.

This article focuses on the experimental elicitation of pain through heat (thermal) and electrical stimulation while recording physiological, visual, and paralinguistic responses. It aims at collecting valid multimodal data for analyzing pain based on its intensity, quality, and duration.&#160;

Мульти-модальных сигналы для анализа боли реакции на раздражители, тепловой и электрической

The assessment of pain relies mostly on methods that require a person to communicate. However, for people with cognitive and verbal impairments, existing ...

Evaluating Skilled Prehension in Mice Using an Auto-Trainer

We describe a method to introduce na&#239;ve mice to a novel prehension (reach-to-grasp) task. Mice are housed singly in cages with a frontal slot that permits the mouse to reach out of its cage and retrieve food pellets. Minimal food restriction is employed to encourage the mice to perform the food retrieval from the slot. As the mice begin to associate coming to the slot for food, the pellets are manually pulled away to stimulate extension and pronation of their paw to grasp and retrieve the pellet through the frontal slot. When the mice begin to reach for the pellets as they arrive at the slot, the behavioral assay can be performed by measuring the rate at which they successfully grasp and retrieve the desired pellet. They are then introduced to an auto-trainer that automates both the process of providing food pellets for the mouse to grasp, and the recording of successful and failed reaching and grasping attempts. This allows for the collection of reaching data for multiple mice with minimal effort, to be used in experimental analysis as appropriate.

Method to assess the impact of training on motor skills is a useful tool. Unfortunately, most behavioral assessments can be labor intensive and/or expensive.We describe here a robotic method of assessing prehension (reach-to-grasp) skill in mice.

Оценка квалифицированных Предениза в мышах с помощью авто-тренер

We describe a method to introduce na&#239;ve mice to a novel prehension (reach-to-grasp) task. Mice are housed singly in cages with a frontal slot that ...