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Socially Transmitted Food Preference (STFP) Task Protocol
可通过社会交往传播的食物偏好实验方案   

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Abstract

Temporally-graded retrograde amnesia (TGRA) refers to a phenomenon of premorbid memory loss whereby information acquired recently is more impaired than information acquired more remotely. Studies of human amnesia have illuminated this phenomenon (Hodges, 1994; Squire and Alvarez, 1995), but such studies necessarily rely on retrospective methods and imperfect tests. Studies in experimental animals have the advantage that retrograde amnesia can be studied prospectively, the locus and extent of brain lesions can be determined accurately, and the timing and strength of original learning can be precisely controlled. The socially transmitted food preference (STFP) task has been one of the most productive rodent behavioral tasks to study TGRA (Clark et al., 2002).

Materials and Reagents

  1. Rats
  2. Cinnamon
  3. Cocoa
  4. Ethanol
  5. Powder
  6. Flavored rat chow meal (see Recipes)

Equipment

  1. Glass jars and the jar holders
  2. Fisher Scientific scale
  3. Feeding apparatus
  4. Cages

Procedure

The Task:
The STFP task is separated into four distinct phases. Phase I consists of food deprivation and habituation of the demonstrator rats to the powder rat chow (“tuning of the demonstrator rats”). Phase II consists of the feeding of the demonstrator with a specified flavored meal, and the interaction of a demonstrator rat with an observer rat. Phase III is a specified delay interval following the interaction between the rats. Finally, phase IV is a testing period in which the observer rat is given a choice between a familiar (the flavored meal that the demonstrator rat was fed) and a novel (a flavor that neither rat has been exposed to) meal chow sample to test the memory of the interaction.

  1. Phase I: Food deprivation and habituation to meal chow
    In order to ensure that the demonstrator rats eat the desired flavored rat chow at the specified time, the rats must be habituation to the meal chow (finely ground rat chow pellets that the rats are normally fed). To do this the rats must become accustomed to a routine of 23 h of food deprivation, followed by one hour of feeding. This trains the rats to eat its fill within the hour ensuring the consistent transmission of the flavor from the demonstrator to the observer. The following steps are taken for this training.
    1. Day 1
      1. Cull the rats into individual cages so that all of the demonstrators and all of the observers are in their own cages.
      2. Remove the pellet rat chow from the demonstrators rats’ cages for 23 h, and place a “do not feed” sign on the cages.
    2. Day 2
      1. After the 23-h food deprivation, feed the demonstrator rats non-flavored meal chow for 1 h using the glass jars and the jar holders. Using a sharpie, label the glass jars with the rat number and the side (left or right) so the jars can be easily distinguished from each other.
      2. Using the Fisher Scientific scale, weigh the mass of one empty jar (massE) for each rat. Fill the jars approximately ¾ of the way full (45-50 g), and weigh the initial mass of each sample and jar (mass).
      3. Administer the samples to the rats using the jar holders in the rats' home cages.
        Place the sample holders and the samples in the demonstrator’s cage (opposite from the food and water bottle allowing for the maximum amount of room for the rat to eat) and bury the base of the holder in the bedding to stabilize the apparatus.
      4. After one hour has elapsed, remove the feeding apparatus and resume the 23-h food deprivation.
      5. Weigh the final mass of the sample and jar (massF) for each rat, and record the data.
    3. Day 3- same as day 2.
      NOTE: If a demonstrator rat fails to eat 5 g of powder chow during the one hour feeding session, he should not be used the following day in the interaction phase of the task. Additional days of training may be accrued until the rats reach the suggested criteria.
      1. After two days of 23 h food deprivation and one hour feeding, the rats should be accustomed to the eating schedule and the interaction with the observer rat will take place the following day.
      2. Using the large weigh boats, the two flavored meal chows should be prepared at this time. They are approximately 1% cinnamon (3 g of cinnamon to 300 g of meal chow) and 2% cocoa (6 g cocoa to 300 g of meal chow). Precautions should be taken so that cross contamination doesn’t occur during the preparation of the samples (i.e. change gloves after weighing out cinnamon or cocoa). Combine the meal and the given flavor in the same zip-lock bag and mix until evenly dispersed. It is recommended that the flavored meal should be prepared at least a day in advance and shouldn’t be stored for longer than a couple of days.

  2. Phase II: Interaction between the observer rats and the demonstrator rats.
    After the demonstrator rats have been trained to eat the meal chow, they will now be exposed to a flavored meal chow. This will be followed by an interaction with an observer rat allowing for the transmission of the scent of the specific flavor to the observer. The interaction is accomplished through a wire screen that is constructed in a cylindrical shape allowing the demonstrator rat to be placed into the empty space of the cylinder. This allows the rats to interact, the experimenter to differentiate between the two rats, and also prevents the rats from fighting.
    1. Day 4
      1. Weigh out the samples of the flavored powder chow, one jar each rat, (prepared the previous day) to be administered to the demonstrator rats. Record the empty mass of the jar (massE) and the initial mass of the sample (masso) on the data sheet.
        Note: It is important to always feed the same demonstrators the same flavor of meal. It is unclear if previous feedings of the demonstrator can influence or confuse the observer rats, therefore, feeding the demonstrators two different types of flavored meal chow for two different testing periods should always be avoided.
      2. After the 23-h food deprivation, feed the demonstrator rats flavored meal chow (cinnamon or cocoa) for 1 h.
      3. While the demonstrator rats are eating, remove the corresponding observer rats’ food.
      4. After the demonstrator rats have eaten the flavored powder chow for 1 h, place the cylindrical screen partition in the observer’s home cage and transfer the demonstrator rat into the center space of the screen partition. This separates the demonstrator and the observer rats. Replace the wire cage top to secure the partition and remove the water bottle. Allow the demonstrator and the observer rats to interact for the given exposure time. Typically 30 min, but this can be used as an independent variable.
        This is the only time that either rat is not allowed to have water present in their cages. It is unclear if drinking water can dilute the scent. It is also desirable to have the rats interacting as much as possible during this time so any distractions (e.g. a water bottle) should be avoided.
      5. While the demonstrator and the observer rats are interacting, determine the final mass of the demonstrator’s flavored powder chow samples and record the data.
      6. After the interaction time is complete, return the demonstrator rat to his home cage. It is easiest to remove both the rat and the screen partition concurrently since the demonstrator will often grab onto the screen once he is lifted out of the cage.
      7. If no further testing of the demonstrator rats will be done the following day, return the rat chow pellets and remove the do not feed sign from the demonstrator’s cage. If the demonstrators are going to be used in an interaction the following day, do not return their food.  Instead, continue with the 23 h. food deprivation schedule and return the caged demonstrator to the appropriate shelf.
      8. If a longer delay is used, replace the rat chow pellets and the water to the observer rat’s cage and return the caged observer to the shelf. Do not return the food if the observer rats are going to be tested immediately.  Instead, go directly to phase IV.

  3. Phase III: Delay interval.
    At this time, a given delay is allowed to elapse before the observer rats are given a choice between the food that their demonstrator rat was exposed to, the familiar food, and a novel flavored sample that neither rat has ever been expose to before. The length of this delay is variable and determined by the experimenter. This delay is crucial for post-operative animal recovery or to test the retention of the memory over longer delays.

  4. Phase IV: Testing phase of the observer rats.
    After the delay has elapsed, the observer rats are tested on the retention of the memory created during the interaction with the demonstrators. The test consists of exposing the observer rats to two samples, the familiar and the novel food samples, and determining by mass how much the observer rat eats of both samples. Measurements are made at one, two, and twenty four hour intervals and the percent preference for the familiar and novel food is calculated.
    1. Day 1
      1. After the given delay has elapsed, the observer rat’s food is removed and a “do not feed” sign is placed on the cage, one hour prior to administering the two samples.
      2. After removing the rat chow pellets, label two of the jars, weigh them empty (massE), and add the flavored rat chow meal  (1% cin and 2% cocoa) to approximately ¾ full (45-50 g) and determine the initial mass (masso).
      3. The two samples, novel and familiar, are taped (any kind will do) to the plastic sample holder and placed into a new fresh cage containing a minimal (just enough to cover the bottom of the cage) amount of bedding. The minimum bedding decreases the chances of the rat burying or contaminating the samples.  If a group of animals is being tested, alternate the familiar food on the left and the right sides so that an equal distribution between the two sides is obtained to compensate for any natural side preferences that the rats may have.
      4. Place the samples, one cinnamon and the other cocoa, into the new cage and move the observer rat, along with his water and wire cage top.
      5. Secure the cage and return it to the shelf, and allow the rat to eat from the two samples for one hour.
      6. After one hour has expired, both food samples are removed and weighed (massF1).
        Wipe away any bedding from the exterior of the jar and only attempt to remove contamination from the inside (by sifting) if there is an excess present.
      7. The samples are then returned to the test cage and the rat is allowed to eat for another hour.
      8. After the second hour, the samples are again removed, weighed, and the masses are recorded (massF2). The samples are placed back into the observers’ testing cage for another 22 h.
      9. After 24 h has elapsed, the samples and the do not feed sign are removed and the rat chow pellets are given back to the observer rat.
      10. The final samples are sifted (if necessary) through a screen to remove any bedding or other contaminating material and the final mass is measured (massF24).
        1. Use the small green handled sifting tool to sift away any contaminating material. It is easiest to sift the entire sample through the screen into a weigh boat and then transfer the entire sample back into the original glass container to make the final mass measurement.
        2. Often it is not necessary to sift the samples and any minor contamination can be removed using forceps. However, if there is contamination after the 24-hour exposure and if one sample of a pair is sifted, be sure to sift the other sample of the same pair. This ensures that if any of the samples mass is lost in the sifting process, it is proportionally lost in both samples so that the relative masses are roughly unchanged.

  5. Miscellaneous information
    1. Cleaning of glassware
    2. Organization of data
      All primary data should be kept in the black STFP binder. In addition to this, all data should be entered into the computer so that two copies of all the data are kept. The data sheets can be found in the Excel file under the C drive, Clark folder, Rat folder, and STFP folder. The data sheets used in the notebook are the “Data sheets for the STFP Task” and the data sheets used for the computer are the “Cal. data sheets for the STFP Task.” The calculation data sheets do the minor calculations needed to determine the mass consumed by the rats and can be used to save the experimenter time or as a check of any primary calculations. After all primary data has been recorded and stored; the data should be transferred to the “all data” sheet found in the same folder. This data file calculates both of the percent preferences and allows for quick and easy interpretation.
    3. Sample calculation
      Data is often interpreted as a percent preference for the familiar or novel food. This calculation is simply the mass of familiar/novel food consumed divided by the total mass of food eaten multiplied by 100%. Commonly, the percent preference for the familiar food is determined. For example,
                                       % preference =   mass of familiar food (g)      (100%)
                                                              Total mass of food eaten (g)

Recipes

  1. Flavored rat chow meal
    1% cinnamon
    2% cocoa

Acknowledgments

This protocol is adapted from Clark et al. (2002).

References

  1. Clark, R. E., Broadbent, N. J., Zola, S. M. and Squire, L. R. (2002). Anterograde amnesia and temporally graded retrograde amnesia for a nonspatial memory task after lesions of hippocampus and subiculum. J Neurosci 22(11): 4663-4669.

简介

短暂性的逆行性遗忘(TFRA)是指依靠获取近期信息的暂时性失忆比依靠获取较久远的信息的暂时性失忆更有害的现象。人健忘症的研究已经解释了这一现象(Hodges, 1994; Squire and Alvarez, 1995), 但是这种研究必须依赖回忆法和不完全测试。利用实验动物来研究更加方便,因为逆行性遗忘可以被预测,脑损伤的位置和范围可以精确的确定,还可以精确的控制初始学习的时间和强度。 社会转交食物偏好(STFP)任务可以很好的研究啮齿类动物的暂时性逆行性失忆症TGRA(Clark et al., 2002)。

材料和试剂

  1. 大鼠
  2. 肉桂
  3. Cocoa
  4. 乙醇
  5. 粉末
  6. 风味老鼠膳食(见食谱)

设备

  1. 玻璃罐和罐支架
  2. 费雪科学仪器公司
  3. 喂料装置
  4. 笼子

程序

任务:
STFP任务分为四个不同的阶段。阶段I包括食物剥夺和示范大鼠习惯于粉末大鼠食物("示范大鼠的调整")。阶段II包括给予示范者特定调味膳食以及示范大鼠与观察者大鼠的相互作用。阶段III是在大鼠之间的相互作用之后的指定延迟间隔。最后,阶段IV是测试期,其中观察者大鼠在熟悉的(示范大鼠被喂食的风味膳食)和新鲜(没有大鼠已经暴露的风味)膳食食物样品之间进行选择记忆的互动。

  1. 第一阶段:食物缺乏和习惯习惯食物
    为了确保示范大鼠在规定的时间吃所需的调味大鼠食物,大鼠必须习惯于膳食(大鼠正常食用的精细研磨的大鼠食物颗粒)。为了做到这一点,老鼠必须习惯于23小时的食物剥夺的程序,然后是1小时的喂养。这训练大鼠在一小时内吃其填充,以确保从示威者到观察者的味道的一致传输。本培训采取以下步骤。
    1. 第1天
      1. 将大鼠分成单独的笼子,以便所有的示威者和所有的观察者都在自己的笼子里。
      2. 从示威者大鼠笼中取出小鼠大鼠食物23小时,并在笼子上放置一个"不喂"标志。
    2. 第2天
      1. 在23小时的食物剥夺后,喂食示威者大鼠非调味膳食1小时,使用玻璃瓶和瓶架。 使用锐利,用大鼠号和侧面(左或右)标记玻璃瓶,以便容器可以容易地彼此区分。
      2. 使用Fisher Scientific scale,称量每个大鼠的一个空罐的质量(质量 E)。 填充罐的大约3/4的方式(45-50克),并称量的初始质量 每个样品和罐(质量)。
      3. 使用大鼠家笼中的罐支架将样品给予大鼠。
        将样品架和样品放置在示范者的笼子中(与食物和水瓶相对,允许最大量的空间供大鼠吃),并将支架的底部埋在床上以稳定装置。 >
      4. 过了一小时后,取出喂食器,恢复23小时的食物剥夺
      5. 称量每只大鼠的样品和罐的质量(质量),并记录数据。
    3. 第3天与第2天相同。
      注意:如果示威者大鼠在一小时喂食期间未能吃5克粉末食物,则不应在第二天的任务的相互作用阶段使用。可能会累积额外的培训天数,直到大鼠达到建议的标准。
      1. 在23天的食物缺乏和1小时喂食的两天后,大鼠应该习惯于进食时间表,并且与观察者大鼠的相互作用将在第二天发生。
      2. 使用大型称重船,这两个风味的膳食食物应该准备在这个时候。它们是大约1%肉桂(3g肉桂至300g膳食)和2%可可(6g可可至300g膳食)。应该采取预防措施,以免在样品制备过程中发生交叉污染(即在称出肉桂或可可之​​后更换手套)。将膳食和给定的风味在相同的拉链袋中混合并混合直到均匀分散。建议至少提前一天准备调味餐,且不应储存超过两天。

  2. 阶段II:观察鼠和示范大鼠之间的相互作用 在示威者大鼠被训练吃饭饭后,他们现在将暴露于风味的膳食。这之后将是与观察者大鼠的相互作用,从而允许特定香味的气味传递给观察者。这种相互作用是通过金属丝筛网来实现的,该金属丝筛网被构造成圆柱形,允许示范老鼠被放置在圆柱体的空的空间中。这允许大鼠交互, 实验者区分两只大鼠,也防止大鼠战斗。
    1. 第4天
      1. 称量调味粉末食品的样品,每只大鼠一个瓶子(前一天制备)给予示范性大鼠。在数据表上记录瓶子的空质量(质量)和样品的初始质量(质量)。
        注意:重要的是给同样的示威者提供相同的口味。不清楚示范者的先前喂食是否会影响或混淆观察者的大鼠,因此,为两个不同的测试期间喂食示范者两种不同类型的风味膳食食物应始终避免。
      2. 在23小时的食物剥夺后,喂食示威者大鼠风味膳食(肉桂或可可)1小时。
      3. 当示威者大鼠吃饭时,取出相应的观察者大鼠的食物
      4. 示威者大鼠吃了风味的粉末食物1小时后,将圆柱屏幕分区在观察员的家笼,并将示威者大鼠转移到 屏幕分区的中心空间。这分开示威者和观察者大鼠。更换电线罩顶部以固定隔板,并取下加水瓶。允许示范者和观察者大鼠交互给定的暴露时间。通常为30分钟,但这可以用作一个独立变量。
        这是唯一一次大鼠不允许有水存在于他们的笼子里。目前尚不清楚饮用水是否能稀释香味。还希望在这段时间内使大鼠尽可能相互作用,以避免任何分心(例如水瓶)。
      5. 当示范者和观察者大鼠相互作用时,确定示范者风味粉末样品的最终质量并记录数据。
      6. 在相互作用时间完成后,将示范大鼠返回他的家笼。最容易同时移除鼠标和屏幕分区,因为示威者在被提出笼子后将经常抓住屏幕。
      7. 如果第二天没有进行示范性大鼠的进一步测试,返回大鼠食物颗粒并从示范器的笼子中取出不进食的标志。如果示威者要在第二天在互动中使用,不要退回他们的食物。相反,继续23 h。食物剥夺时间表,并将笼子示威者返回适当的架子
      8. 如果使用更长的延迟,将大鼠食物颗粒和水更换到观察者大鼠的笼子,并将笼子观察者返回架子。如果观察鼠要立即进行测试,不要返回食物。而是直接进入第四阶段。

  3. 阶段III:延迟间隔。
    此时,在观察者大鼠被给予他们的示威者大鼠暴露于的食物,熟悉的食物和新的风味样品之间的选择之前,允许给定的延迟,所述新的风味样品既没有大鼠曾经暴露过。该延迟的长度是可变的并且由实验者确定。这种延迟对于手术后动物恢复或测试在较长时间内保留记忆是至关重要的。

  4. 第四阶段:观察鼠的测试阶段 在延迟过去之后,对观察者大鼠在与演示者的相互作用期间产生的记忆的保留进行测试。该测试包括将观察者大鼠暴露于两个样品,熟悉的和新的食物样品,并且通过质量确定观察者大鼠吃多少两种样品。以一,二和二十四小时的间隔进行测量,计算熟悉的和新颖的食物的百分比偏好。
    1. 第1天
      1. 在给定的延迟过去之后,在施用两个样品前一小时,去除观察者大鼠的食物并且在笼子上放置"不进食"标志。
      2. 除去大鼠食物颗粒后,标记两个瓶子,称其为空(质量),并加入调味的大鼠食物膳食(1%cin和2%可可)至约3/4全(45-50g),并测定初始质量(质量%)。
      3. 两个样品,新颖和熟悉的,被胶带(任何种类将做)到塑料样品架,并放置在一个新的笼子里,包含一个最小(刚好足以覆盖笼子的底部)的床上用品。最小的床铺减少了大鼠埋藏或污染样品的机会。如果正在测试一组动物,则在左侧和右侧交替熟悉的食物,以获得两侧之间的平均分布,以补偿大鼠可能具有的任何自然侧偏好。
      4. 将样品,一个肉桂和其他可可,放入新的笼子,并移动观察员大鼠,连同他的水和金属丝笼顶。
      5. 固定笼子并将其放回架子,让老鼠从两个样品中吃一小时。
      6. 一小时过后,取出两个食物样品并称重(质量 F1)。
        擦拭罐外部的任何床上用品,如果有过量的存在,只试图从内部去除污染物(通过筛选)。
      7. 然后将样品返回测试笼,使大鼠再吃一小时。
      8. 第二小时后,再次取出样品,称重,记录质量(质量 F2)。将样品放回观察者的测试笼中另外22小时
      9. 24小时后,取出样品和不进食标志,将大鼠食物颗粒送回观察者大鼠。
      10. 最终样品通过筛网过筛(如果需要)以除去任何垫料或其它污染物质,并测量最终质量(质量F 24)。
        1. 使用小的绿色处理筛分工具筛选任何污染物质。最容易的是将整个样品通过筛网筛入称重船中 将整个样品转移回原来的玻璃容器中进行最终的质量测量
        2. 通常不需要筛选样品,并且可以使用镊子去除任何微小的污染物。 然而,如果在24小时暴露后存在污染,并且如果一对样品中的一个样品被筛分,则一定要筛选同一对的另一个样品。 这确保了如果任何样品质量在筛分过程中损失,则在两个样品中成比例地损失,使得相对质量大致不变。

  5. 其他信息
    1. 清洗玻璃器皿
    2. 数据组织
      所有主数据应保存在黑色STFP绑定程序中。除此之外,所有数据都应输入计算机,以便保存所有数据的两个副本。数据表可以在C驱动器,Clark文件夹,Rat文件夹和STFP文件夹下的Excel文件中找到。笔记本中使用的数据表是"STFP任务的数据表",用于计算机的数据表是"Cal。 STFP任务的数据表"。计算数据表执行确定大鼠消耗的质量所需的次要计算,并且可用于保存实验者时间或作为任何主要计算的检查。在所有主数据被记录和存储之后;数据应该传输到在同一文件夹中找到的"所有数据"表。此数据文件计算偏好百分比,并允许快速简单的解释。
    3. 示例计算
      数据通常被解释为熟悉或新颖食物的偏好百分比。这个计算只是熟悉的/新的食物消耗的质量除以食物的总质量乘以100%。通常,确定熟悉的食物的偏好百分比。例如,
                                       %preference =  熟食量(g)      (100%)
                                                             食物总质量(g)

食谱

  1. 风味大鼠膳食
    1%肉桂
    2%可可

致谢

该协议改编自Clark等人(2002)。

参考文献

  1. Clark,R.E.,Broadbent,N.J.,Zola,S.M.and Squire,L.R。(2002)。 顺行性遗忘和暂时分级的逆行性遗忘,用于在海马和下颌骨损伤后进行非空间记忆任务。 a> J Neurosci 22(11):4663-4669。
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免责声明 × 为了向广大用户提供经翻译的内容,www.bio-protocol.org 采用人工翻译与计算机翻译结合的技术翻译了本文章。基于计算机的翻译质量再高,也不及 100% 的人工翻译的质量。为此,我们始终建议用户参考原始英文版本。 Bio-protocol., LLC对翻译版本的准确性不承担任何责任。
Copyright: © 2012 The Authors; exclusive licensee Bio-protocol LLC.
引用: Readers should cite both the Bio-protocol article and the original research article where this protocol was used:
  1. Clark, R. E. (2012). Socially Transmitted Food Preference (STFP) Task Protocol. Bio-protocol 2(11): e224. DOI: 10.21769/BioProtoc.224.
  2. Clark, R. E., Broadbent, N. J., Zola, S. M. and Squire, L. R. (2002). Anterograde amnesia and temporally graded retrograde amnesia for a nonspatial memory task after lesions of hippocampus and subiculum. J Neurosci 22(11): 4663-4669.
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