Name: a simple and inexpensive method for determining cold sensitivity and adaptation in mice
Uploaded: 2015-03-17T15:00:00
Description: Watch this Scientific Journal Video about A Simple and Inexpensive Method for Determining Cold Sensitivity and Adaptation in Mice at JoVE.com

The authors would like to acknowledge contributions from the entire Gereau Lab for manuscript editing. This work is supported by NINDS funds 1F31NS078852 to DSB and NINDS fund NS42595 to RWG.

Tous les protocoles de souris étaient conformes aux Instituts nationaux de la Santé et des lignes directrices ont été approuvées par le Comité des études animales de l&#39;École de médecine de l&#39;Université de Washington (St. Louis, MO). 1. Préparation de la plaque d&#39;essais et boîtiers <ol><li> Nettoyer la surface du verre. </li><li> Fixez la sonde filament thermocouple de type T à la surface dans le milieu de la plaque de verre avec du ruban de laboratoire. </li><li> P...

<ol>
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The CPA can be used to assess cold sensitivity and cold adaptation in mice. It provides an affordable, efficient way to measure cold responses in unrestrained, acclimated animals at a wide variety of temperature ranges. It also provides an unambiguous behavioral response with an easily quantified and analyzed output variable. It has already been used to assess changes in cold sensitivity induced by inflammation1, neuropathic injury1, analgesics1, genetic knockouts20, and ge...

Mesure de la réactivité à froid chez les rongeurs est important pour améliorer la compréhension des mécanismes potentiels de sensibilité au froid chez les humains dans des conditions normales et pathologiques. The Cold plantaire Assay (CPA), développé à l&#39;origine il ya plusieurs années une, est conçu pour générer des réponses claires murins comportement reproductibles à un stimulus froid amené à RT. Améliorations plus récentes de cet essai ont permis la mesure reproductible de sensibilité au froid à une large gamme de températures 2. Les deux versions sont également conçus pour être relativement à haut débit, et peu coûteux à utiliser. A beaucoup de progrès ont été réalisés dans la compréhension des mécanismes de sensibilité au froid en utilisant d&#39;autres méthodes comportementales. Un procédé est le test de l&#39;évaporation de l&#39;acétone, ce qui implique tamponnant ou en pulvérisant de l&#39;acétone sur la patte de la souris et en mesurant la quantité de temps que la souris consacre effleurant la patte de 3,4. Malheureusement,les réponses à l&#39;acétone évaporation sont confondus par la sensation d&#39;humidité et l&#39;odeur de l&#39;acétone. En outre, le stimulus froid qui est appliquée dans le test d&#39;évaporation de l&#39;acétone peuvent varier en fonction de la quantité d&#39;acétone appliqué, et est difficile à quantifier. Enfin, les souris non lésés ont des réponses minimales au niveau de référence à l&#39;acétone, ce qui rend impossible la mesure de l&#39;analgésie en l&#39;absence d&#39;une hypersensibilité à la présente méthode. Un autre essai classique pour les réponses à froid est le test du Tail Flick, où la latence de retrait est mesuré après la queue est immergée dans 5,6 d&#39;eau froide. Alors que les réponses comportementales dans cet essai sont sans ambiguïté et l&#39;essai mesure les réponses à une température spécifique, les animaux doivent être retenus pendant les essais, qui peut altérer la réactivité à froid grâce à des mécanismes analgésiques induits par le stress bien décrits 7. Un autre outil couramment utilisé est le test de la plaque froide, qui mesure le comportementles réponses des souris après leur mise sur une plaque refroidie par effet Peltier 10/08. Bien que cet outil fournit des informations sur les réponses des animaux à des températures spécifiques, il a également été utilisé d&#39;une manière incompatible; différents groupes ont mesuré différents types de réponses, y compris nombre de sauts 8,11, la latence de la première réponse 8,11- 13, et le nombre de patte ascenseurs 11,13,14 avec des résultats très différents. Le dosage de la plaque froide est également relativement faible débit comme un seul animal peut être testée à la fois, et il nécessite un dispositif de Peltier coûteux et fragile. Le test de préférence de la température de 2 plaques est un dérivé communément utilisé pour le test de la plaque froide qui mesure la quantité relative de temps que les animaux passent sur ​​deux plaques reliées à des températures différentes 9,15- 17. Un autre test couramment utilisé est le même dosage de gradient thermique, où la quantité de temps que les souris passent en différentes zones de températurecomprise entre 5 ° C et 45 ° C sur une plaque longue métallique 16 est mesurée. Bien que ces tests permettent de comparer les températures, il est difficile de savoir si le comportement représente l&#39;aversion de la température ou à la préférence de température. Enfin, l&#39;analyse dynamique de plaque froide a été utilisé pour mesurer la souris répondent à l&#39;évolution des températures ambiantes 18. Cette méthode consiste à placer des souris sur un dispositif à effet Peltier et RT rampe vers le bas à 1 ° C tout en mesurant combien les souris sautent ou lèchent les pattes de la plaque à des températures différentes. Bien que cette façon teste les souris se adapter à un environnement de refroidissement, il ne fournit pas un moyen pour tester la souris réagissent à un stimulus froid dans le cadre d&#39;une température ambiante froide. En outre, il nécessite un équipement coûteux à réaliser et ne offre pas un moyen de souris se acclimater à l&#39;équipement de test avant de mesurer leur sensibilité au froid. Pour compléter ces essais, le CPA teste la ACCLIMATED réponses à un stimulus froid bien défini à une variété de gammes de température, ou pendant le processus d&#39;adaptation à des températures ambiantes froides. Il peut tester jusqu&#39;à 14 souris à la fois avec notre appareil actuel, avec le potentiel d&#39;être économiquement plus grande échelle pour les tests à haut débit.

<table border="0" cellpadding="0" cellspacing="0" width="1209">
	<tbody>
		<tr height="21">
			<td height="21" style="height:21px;width:233px;">Name</td>
			<td style="width:320px;">Company</td>
			<td style="width:353px;">Catalog Number</td>
			<td style="width:303px;">Comments</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">T-type thermocouple probe</td>
			<td>Physitemp</td>
			<td>IT-24p</td>
			<td>Used to measure the surface temperature of the glass (http://www.physitemp.com/products/probesandwire/)</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Glass plate</td>
			<td>Local glass company (in St. Louis, Stemmerich Inc)</td>
			<td>N/A</td>
			<td>We use pyrex glass (borosilicate float). Our lab generally uses 1/4&#39;&#39;, but 3/16&#39;&#39; and 1/8&#39;&#39; are also useful</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Thermal Data logger</td>
			<td>Extech</td>
			<td>EA15</td>
			<td>Thermologger to keep track of glass temperature (http://www.extech.com/instruments/product.asp?catid=64&#38;prodid=408)</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">3mL Syringe</td>
			<td>BD</td>
			<td align="right">309657</td>
			<td>The top is cut off, and dry ice is compressed in the syringe to generate a cold probe</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Computer</td>
			<td>If using Extech logger, any Pcwill work</td>
			<td>N/A</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Aluminum boxes</td>
			<td>Washington University in St. Louis machine shop</td>
			<td>N/A</td>
			<td>boxes are 3&#39; long, 4.5&#39;&#39; wide, and 3&#39;&#39; tall with a sealed lid.&#160; There is a 1/2&#39;&#39; hole drilled into one short side of each box, near the bottom. These holes are filled with rubber stopcocks when the boxes are filled with wet ice or hot water.</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Heated water circulator</td>
			<td>VWR</td>
			<td></td>
			<td>Any water circulator model with a pump will work</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">21 gauge needle</td>
			<td>BD</td>
			<td align="right">305165</td>
			<td>The exact needle size is not important</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Hand timer</td>
			<td>N/A</td>
			<td></td>
			<td>Any hand timer will work</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Mirror</td>
			<td>N/A</td>
			<td></td>
			<td>Any flat mirror will work</td>
		</tr>
	</tbody>
</table>

a simple and inexpensive method for determining cold sensitivity and adaptation in mice

Cold hypersensitivity is a serious clinical problem, affecting a broad subset of patients and causing significant decreases in quality of life. The cold plantar assay allows the objective and inexpensive assessment of cold sensitivity in mice, and can quantify both analgesia and hypersensitivity. Mice are acclimated on a glass plate, and a compressed dry ice pellet is held against the glass surface underneath the hindpaw. The latency to withdrawal from the cooling glass is used as a measure of cold sensitivity.
Cold sensation is also important for survival in regions with seasonal temperature shifts, and in order to maintain sensitivity animals must be able to adjust their thermal response thresholds to match the ambient temperature. The Cold Plantar Assay (CPA) also allows the study of adaptation to changes in ambient temperature by testing the cold sensitivity of mice at temperatures ranging from 30 &#176;C to 5 &#176;C. Mice are acclimated as described above, but the glass plate is cooled to the desired starting temperature using aluminum boxes (or aluminum foil packets) filled with hot water, wet ice, or dry ice. The temperature of the plate is measured at the center using a filament T-type thermocouple probe. Once the plate has reached the desired starting temperature, the animals are tested as described above.
This assay allows testing of mice at temperatures ranging from innocuous to noxious. The CPA yields unambiguous and consistent behavioral responses in uninjured mice and can be used to quantify both hypersensitivity and analgesia. This protocol describes how to use the CPA to measure cold hypersensitivity, analgesia, and adaptation in mice.

Les réponses comportementales induites chez des souris à partir de 30 ° C, 23 ° C, 17 ° C et 12 ° C sont très reproductible (figure 4A) 20. Afin de mesurer le stimulus froid étant généré sous la patte arrière, les souris ont été anesthésiés avec un kétamine / xylazine / acépromazine cocktails et leurs pattes ont été fixés sur le verre sur le dessus d&#39;un thermocouple de type T filament (figure 4B) 20. Le verre est refroidi ou réchauffé à ...

Watch this Scientific Journal Video about A Simple and Inexpensive Method for Determining Cold Sensitivity and Adaptation in Mice at JoVE.com

A Simple and Inexpensive Method for Determining Cold Sensitivity and Adaptation in Mice

The Cold Plantar Assay (CPA) measures cold responsiveness between 30 &#176;C and 5 &#176;C, and can also measure cold adaptation. This protocol describes how to use the CPA to measure cold hypersensitivity, analgesia, and adaptation in mice.

Une méthode simple et peu coûteuse pour déterminer Sensibilité au froid et adaptation chez la souris

Cold hypersensitivity is a serious clinical problem, affecting a broad subset of patients and causing significant decreases in quality of life.  The cold ...

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Cortical development involves complex interactions between neurons and non-neuronal elements including precursor cells, blood vessels, meninges and ...

An Inexpensive, Scalable Behavioral Assay for Measuring Ethanol Sedation Sensitivity and Rapid Tolerance in Drosophila

An Inexpensive, Scalable Behavioral Assay for Measuring Ethanol Sedation Sensitivity and Rapid Tolerance in Drosophila

Un, test de comportement évolutives et économiques pour mesurer l&#39;éthanol sédation sensibilité et la tolérance rapide dans<em&gt; Drosophila</em

Alcohol use disorder (AUD) is a serious health challenge. Despite a large hereditary component to AUD, few genes have been unambiguously implicated in ...

A Method to Target and Isolate Airway-innervating Sensory Neurons in Mice

Une méthode pour cibler et Isoler Airway-innervant neurones sensoriels chez la souris

Somatosensory nerves transduce thermal, mechanical, chemical, and noxious stimuli caused by both endogenous and environmental agents. The cell bodies of ...

A Simple One-step Dissection Protocol for Whole-mount Preparation of Adult Drosophila Brains

A Simple One-step Dissection Protocol for Whole-mount Preparation of Adult Drosophila Brains

Une seule étape simple protocole Dissection pour Whole-mount Préparation des adultes<em&gt; Drosophila</em&gt; Brains

There is an increasing interest in using Drosophila to model human brain degenerative diseases, map neuronal circuitries in adult brains, and study the ...

Novel Assay for Cold Nociception in Drosophila Larvae

Novel Assay for Cold Nociception in Drosophila Larvae

Nouveau test pour froid dans nociception<em&gt; drosophile</em&gt; larves

How organisms sense and respond to noxious temperatures is still poorly understood. Further, the mechanisms underlying sensitization of the sensory ...

Culturing In Vivo-like Murine Astrocytes Using the Fast, Simple, and Inexpensive AWESAM Protocol

Culturing In Vivo-like Murine Astrocytes Using the Fast, Simple, and Inexpensive AWESAM Protocol

Culture In Vivo-comme les Astrocytes Murine en utilisant le protocole AWESAM rapide, Simple et peu coûteux

The AWESAM (a low-cost easy stellate astrocyte method) protocol entails a fast, simple, and inexpensive way to generate large quantities of in vivo-like ...