Support provided by grant AA019462 to DAH and AA019884 to DDS.

All procedures described here and in the accompanying video have been approved by the Institutional Animal Care and Use Committees of the Health Sciences Center and the main campus of the University of New Mexico.
1. Prenatal Ethanol Exposure
<ol>
	<li>Obtain all required materials and chemicals: Saccharin sodium salt hydrate, 190 proof ethanol (95% alcohol by volume), beaded glass drinking tubes , natural rubber white #4 stopper one hole , 1&#34; bend tubes-ball point, paper rulers printed from www.vendian.org/mncharity/dir3/paper_rulers/.</li>
	<li>Obtain proven adult breeder rats from a vendor or alternative source. Use Long-Evans breeders for the methods and representative data presented in this protocol. Females weigh 125-150 g and 6-7 weeks of age upon arrival such that they are approximately 9-10 weeks old at the time of breeding (step 1.5). Ensure that all prospective dams are first-time mothers. Make sure that males are 12 weeks of age at arrival and 15 weeks old at the start of the breeding protocol.</li>
	<li>House the animals individually in plastic cages at 22 &#176;C on a reverse 12 hr light/dark schedule (lights on from 21:00 to 09:00 hr). Provide access to food and tap water ad libitum throughout the study, including during all drinking sessions. Allow at least 1 week for acclimation to the facility before proceeding to step 1.4.</li>
	<li>Obtain baseline body weights for each female rat.</li>
	<li>Evaluate pre-pregnancy drinking levels in female rats for 2 weeks. Follow steps 1.5.1&#8211;1.5.6 in order.
	<ol>
		<li>On days 1 and 2 fill a drinking tube containing 0.066% saccharin solution (no ethanol) in tap water. To quantify consumption affix a paper ruler with mm precision to the drinking tube prior to filling it. Use this method as it reduces error in measurement associated with weighing tubes to quantify ethanol consumption. Fill the tube to the 20 mm mark on the ruler. Begin four-hour drinking sessions (steps 1.5.1-1.5.6 and 1.7) 1 hr after the onset of the dark phase (10:00 hr) when activity and drinking levels are highest.</li>
		<li>At the end of each drinking session quantify the volume of the saccharin solution that was consumed. To facilitate measurement, determine the volume of solution per mm on the ruler in advance and convert mm to volumes. For the tubes recommended here each mm corresponds to 0.366 ml.</li>
		<li>On days 3 and 4 fill a tube with 2.5% ethanol (v/v) and 0.066% saccharin solution in tap water. At the end of each session quantify the amount of ethanol solution (weight per kg of body weight) that was consumed.</li>
		<li>On day 5 and thereafter fill a tube with 5% ethanol (v/v) and 0.066% saccharin solution in tap water. At the end of each session quantify the amount of ethanol solution (weight per kg of body weight) that was consumed.</li>
		<li>Upon completion of the pre-pregnancy drinking phase weigh the rats and calculate the mean ethanol consumption and standard deviation for the entire group. Remove rats for which mean consumption is greater than 1 standard deviation above or below the group mean from the study to reduce the variability in voluntary drinking during pregnancy.</li>
		<li>Assign the remaining rats to either the saccharin control or PAE conditions such that the pre-pregnancy drinking levels are matched as closely as possible for the two groups.</li>
	</ol>
	</li>
	<li>Within 1-14 days pair each female rat with a proven male breeder. Female rats do not consume ethanol during the breeding phase. Confirm pregnancy by the presence of a vaginal plug, weigh the female rat, and house her individually. This is defined as gestational day 1. Optionally, leave the female rats with a male breeder for up to 5 days, after which remove the females from the study. 
	NOTE: Because ethanol consumption during pregnancy begins at 10:00 hr on gestational day 1 inspection for the presence of a vaginal plug should be performed prior to this time.</li>
	<li>Provide ethanol or saccharin solutions for 4 hr per day (1,000 to 1,400 hr) for the duration of pregnancy.
	<ol>
		<li>Beginning on gestational day 1 provide the female rat with either 0% or 5% ethanol and 0.066% saccharin solution in tap water based on the group assignment. Ensure that the volume of 0% ethanol solution provided to saccharin control rats is matched to the mean volume of 5% ethanol solution consumed by ethanol rats. At the end of each session quantify the amount of ethanol solution (weight per kg of body weight) or saccharin that was consumed.</li>
		<li>Weigh rat dams weekly to assess maternal weight gain.</li>
		<li>Provide food and tap water at all times including the drinking sessions.</li>
		<li>Cease the ethanol exposure procedures when the offspring are born. Record the number of live pups and pup weights at birth. Designate the day of birth as postnatal day 0.</li>
		<li>Cull the litter to 10 pups around postnatal day 2-3. Attempt to maintain an equal ratio of male to female animals in each litter. If this is not possible, cull to 10 pups with an unequal number of males and females.</li>
		<li>Record pup weights 2-3 days after birth.</li>
		<li>Wean the animals at approximately postnatal day 21-24 and house in same-sex pairs with an animal from the same prenatal treatment condition. Do not use more than 1-2 rats from each litter per experiment to limit potential litter effects.</li>
	</ol>
	</li>
</ol>
2. Social Behavior
<ol>
	<li>Obtain and prepare all required materials and equipment.
	<ol>
		<li>Obtain an apparatus for social interaction. Use a chamber with an open top constructed of material that is easy to clean and sanitize. The front of the apparatus should be covered with rigid transparent plastic (~2 mm thickness) for filming. Line the interior walls and floor with transparent plastic (~2 mm thickness) to aid in cleaning and odor control. Place mirrors along the back interior wall to aid analysis. 
		NOTE : For the representative data reported here a custom chamber (95 cm long &#215; 47 cm wide &#215; 43 cm tall) with an open top and rigid transparent plastic (2 mm thickness) covering the interior sides and floor of the apparatus was used. Mirrors were placed along the back wall. The specific dimensions and materials are not critical for measurement of social behavior, however, it is recommended that the dimensions be sufficiently large to ensure that genuine social interaction can be distinguished from other behaviors. That is, the apparatus should be large enough so that animals are not always in close proximity to one another.</li>
		<li>Obtain video cameras capable of recording under low or no-light conditions as all filming is conducted with little or no ambient lighting within the visible spectrum. Ensure that the camera has a high resolution within the infrared spectrum, however, any camera capable of recording under dark conditions in its native night mode or with additional infrared illuminators should be sufficient.</li>
		<li>Position infrared illuminators around the apparatus to improve lighting of the apparatus in the video recording.</li>
		<li>Obtain laboratory grade wood chips (aspen chip).</li>
		<li>Obtain a brush, dustpan, chlorine dioxide and isopropyl alcohol (70%) for cleaning the apparatus between sessions and odor control.</li>
	</ol>
	</li>
	<li>Apparatus acclimation and social behavior
	<ol>
		<li>For this phase of the experiment, make measurements in adult rats that are at least 90 days of age.</li>
		<li>Prior to each session remove any woods chips, wipe the apparatus clean with isopropyl alcohol to control odors between sessions and provide fresh wood chips. Ensure that the wood chips entirely cover the bottom of the apparatus. Clean and sanitize the apparatus with an appropriate agent, such as chlorine dioxide, as necessary.</li>
		<li>For three consecutive days place an animal and its cage-mate into the chamber for 30 minutes to acclimate the animals to the apparatus. During the acclimation sessions all room lights are turned off.</li>
		<li>At the end of the third acclimation session house the animals individually in new cages with fresh bedding, food, and water for 24 hr to motivate social interaction.</li>
		<li>Record social interaction on the following day 24 hr after the animals were separated.
		<ol>
			<li>Remove wood chips from the apparatus, clean and sanitize with chlorine dioxide, wipe with isopropyl alcohol to control odors, and replace the wood chips prior to the session.</li>
			<li>Position one or more cameras to record the interaction. Position at least one camera in front of the apparatus so that the mirrors on the back wall of the apparatus can provide an additional perspective for analysis.</li>
			<li>Retrieve animals one at a time and hold the animal in front of the camera so that the unique features of the fur pattern can be noted. These identifying features of the fur can be used to distinguish rats during analysis rather than artificially marking the animals. 
			NOTE: Because many aspects of rodent social interaction involve olfactory signals and smelling the partner, wherever possible introducing foreign odors should be avoided. Long-Evans rats typically have some feature of the fur pattern that can be utilized to distinguish any given pair of animals. For other strains (e.g., Sprague-Dawley rats) alternative approaches such as marking the tail with an unscented dye could be used. It is important to recognize that many behaviors of interest are directed toward specific targets (e.g., anogenital sniffing directed near the base of the tail, playful attacks directed at the nape of the neck, or aggressive attacks directed toward the flanks or belly). Marking the animals closer to the tip of the tail far from these targets of interest is recommended.</li>
			<li>Record video of the social interaction for at least 12 min.</li>
			<li>Monitor animals for fighting throughout the session. If possible, watch the animals via a monitor or window so that the experimenter is not in the room during the session. 
			NOTE: Fighting has been only rarely observed in studies with adult rats, however, animals should be monitored throughout the session. The session should be ceased if there is excessive fighting or there are signs of harm or injury to an animal.</li>
		</ol>
		</li>
	</ol>
	</li>
	<li>Behavioral coding and analysis.
	<ol>
		<li>Identify the following behaviors of interest as per previous work with PAE1,18. Quantify the duration, frequency and latency to first occurrence of the following behaviors; wrestling (including pinning), boxing, crawling (crossing) over/under the partner, anogenital sniffing, other sniffing of the partner&#8217;s body (body sniffing), allogrooming (grooming of the partner), rearing, and sniffing/digging in the wood chips. Examples of each behavior are illustrated in the video component of this article.</li>
		<li>Quantify the social behaviors of interest from the video. Obtain the frequency, total duration and latency to first instance for each behavior of interest. 
		NOTE: Obtaining these measures can be achieved manually, however, quantification of these measures using computerized analyses of digitized video is recommended. A Matlab (www.mathworks.com) script for playback of the video and quantification of behaviors is provided as a supplement to this article.</li>
		<li>After coding is completed for all rats the resulting duration, frequency and latency data are analyzed with a statistical package.</li>
	</ol>
	</li>
</ol>

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(71), e4288-e4288 (2013).</li><li>Panksepp, J., Siviy, S., Normansell, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+psychobiology+of+play+:+Theoretical+and+methodological+perspectives.">The psychobiology of play : Theoretical and methodological perspectives.</a> Neurosci. Biobehav. Rev. 8 (4), 465-492 (1984).</li><li>Siviy, S. M., Panksepp, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sensory+modulation+of+juvenile+play+in+rats.">Sensory modulation of juvenile play in rats.</a> Dev. Psychobiol. 20 (1), 39-55 (1987).</li><li>Pellis, S. 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=The+effects+of+orbital+frontal+cortex+damage+on+the+modulation+of+defensive+responses+by+rats+in+playful+and+nonplayful+social+contexts.">The effects of orbital frontal cortex damage on the modulation of defensive responses by rats in playful and nonplayful social contexts.</a> Behav. Neurosci. 120 (1), 72-84 (2006).</li><li>Blanchard, R. J., Blanchard, D. C., Takahashi, T., Kelley, M. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Attack+and+defensive+behavior+in+albino-rat.">Attack and defensive behavior in albino-rat.</a> Anim. Behav. 25 (3), 622-634 (1977).</li></ol>

The authors have no conflicts of interest to disclose.

The prenatal alcohol exposure paradigm described here involves voluntary consumption of ethanol (5% v/v) by rat dams during pregnancy. There are a number of protocols for exposing non-human animals to ethanol during brain development represented in the literature, which differ with respect to the timing, dose, duration and route of ethanol administration as well as the species under investigation. Although a thorough treatment of the advantages of various exposure protocols is not provided here, several advantages of the...

An estimated 1-5% of children are diagnosed with Fetal Alcohol Spectrum Disorders (FASDs)2, which include Fetal Alcohol Syndrome (FAS), partial FAS (pFAS), and Alcohol-Related Neurodevelopmental Disorders (ARNDs)3. Deficits in social behavior and cognition are among the most common adverse outcomes observed in children with FASDs4-7. Negative consequences are not limited to heavy prenatal alcohol exposure (PAE), as moderate PAE that does not lead to the conspicuous morphological, behavioral and cognitive deficits characteristic of FAS can cause comparatively subtle, but nonetheless persistent, deficits in humans with FASDs8-10 and non-human animals exposed to ethanol during brain development11. The importance of understanding the behavioral and corresponding neurobiological consequences of moderate PAE is underscored by current estimates indicating that the large majority of FASD cases fall within the less severe range of the spectrum12.
Several independent laboratories have reported alterations in rodent social behavior related to ethanol exposure during brain development, including decreased investigation and interaction1,13-15, altered play14,16,17, increased aggressive interactions17,18, alterations in responsiveness to social stimuli19-21, and deficits in socially acquired food preferences and social recognition memory22. Social behavior deficits have been observed following exposure to heavy (blood ethanol concentrations (BECs) ~300mg/dl)22,23 or more moderate levels of ethanol (BECs ~80mg/dl)1, and across a broad range of parameters for other significant factors including exposure timing, duration of exposure, and age at the time of behavioral measurement.
Previous research has demonstrated that alterations in specific aspects of social interaction in adulthood discriminate rats exposed to moderate levels of alcohol from control animals exposed to saccharin1,18. In particular, moderate PAE has consistently been associated with robust increases in wrestling, which suggests increases in aggressive behavior, and lower levels of social investigation (e.g., sniffing of the partner) in adulthood. Because alterations in social behavior are reliable consequences of PAE, the quantification of social behavior following PAE may hold importance for progressing toward a more complete understanding of the neural bases of PAE-related alterations in social behavior and the development of interventional approaches. The goal of this paper and the associated video is to provide instruction on the moderate PAE protocol and methods for quantification of social behavior in adult offspring that have reliably distinguished prenatal alcohol-exposed from non-exposed rat offspring.

<table border="0" cellpadding="0" cellspacing="0">
	<tbody>
		<tr>
			<td>Saccharin sodium salt hydrate</td>
			<td>Sigma</td>
			<td>S1002</td>
			<td></td>
		</tr>
		<tr>
			<td>190 proof ethanol</td>
			<td>Sigma</td>
			<td>493538</td>
			<td></td>
		</tr>
		<tr>
			<td>Beaded glass drinking tubes</td>
			<td>Fisher</td>
			<td>14-955K</td>
			<td></td>
		</tr>
		<tr>
			<td>Natural rubber white #4 stopper one hole</td>
			<td>Plasticoid</td>
			<td>LSG4M181</td>
			<td></td>
		</tr>
		<tr>
			<td>1&#34; bend tubes-ball point</td>
			<td>Ancare</td>
			<td>TD-199-3&#34;</td>
			<td></td>
		</tr>
		<tr>
			<td>Paper rulers</td>
			<td>N/A</td>
			<td>N/A</td>
			<td><a href="http://www.vendian.org/mncharity/dir3/paper_rulers">www.vendian.org/mncharity/dir3/paper_rulers</a></td>
		</tr>
		<tr>
			<td>Apparatus for social interaction</td>
			<td>Custom built</td>
			<td>N/A</td>
			<td>95 cm X 47 cm X 43 cm</td>
		</tr>
		<tr>
			<td>Video cameras</td>
			<td>N/A</td>
			<td>N/A</td>
			<td>Capable of recording low/no light conditions</td>
		</tr>
		<tr>
			<td>Infrared illuminators</td>
			<td>Vitek</td>
			<td>VT-IR1-12</td>
			<td></td>
		</tr>
		<tr>
			<td>Teklad laboratory grade sani-chips</td>
			<td>Harlan</td>
			<td>7090A</td>
			<td></td>
		</tr>
		<tr>
			<td>Brush and dustpan</td>
			<td>N/A</td>
			<td>N/A</td>
			<td></td>
		</tr>
		<tr>
			<td>Isopropyl alcohol</td>
			<td>Sigma</td>
			<td>W292907</td>
			<td></td>
		</tr>
		<tr>
			<td>Chlorine Dioxide (1.5 mg Tablets)</td>
			<td>Quiplabs</td>
			<td>N/A</td>
			<td>Prepare per manufacturer&#39;s recommendation</td>
		</tr>
	</tbody>
</table>

moderate prenatal alcohol exposure and quantification of social behavior in adult rats

Alterations in social behavior are among the major negative consequences observed in children with Fetal Alcohol Spectrum Disorders (FASDs). Several independent laboratories have demonstrated robust alterations in the social behavior of rodents exposed to alcohol during brain development across a wide range of exposure durations, timing, doses, and ages at the time of behavioral quantification. Prior work from this laboratory has identified reliable alterations in specific forms of social interaction following moderate prenatal alcohol exposure (PAE) in the rat that persist well into adulthood, including increased wrestling and decreased investigation. These behavioral alterations have been useful in identifying neural circuits altered by moderate PAE1, and may hold importance for progressing toward a more complete understanding of the neural bases of PAE-related alterations in social behavior. This paper describes procedures for performing moderate PAE in which rat dams voluntarily consume ethanol or saccharin (control) throughout gestation, and measurement of social behaviors in adult offspring.

Over the course of many breeding rounds female rats in the ethanol condition consistently drink an average of about 2.1 g/kg of ethanol per 4 hr drinking session. Rat dams consume approximately one-half of the four hr total during the first 15 to 30 min after the introduction of the drinking tubes, resulting in a peak maternal serum ethanol concentration of about 60 mg/dl, measured at the 45 min time point. Over the remaining 3.5 hr of the drinking period, they continue to consume 5% ethanol at a lower, but relatively st...

Watch this Scientific Journal Video about Moderate Prenatal Alcohol Exposure and Quantification of Social Behavior in Adult Rats at JoVE.com

Moderate Prenatal Alcohol Exposure and Quantification of Social Behavior in Adult Rats

The goal of the protocol presented here is to describe procedures to expose rats to moderate levels of alcohol during prenatal brain development and to quantify resulting alterations in social behavior during adulthood.

Alterations in social behavior are among the major negative consequences observed in children with Fetal Alcohol Spectrum Disorders (FASDs). Several ...

moderate-prenatal-alcohol-exposure-quantification-social-behavior

Research

JoVE Journal

Behavior

9.6K Views.  University of New Mexico. The goal of the protocol presented here is to describe procedures to expose rats to moderate levels of alcohol during prenatal brain development and to quantify resulting alterations in social behavior during adulthood. 

Video: Moderate Prenatal Alcohol Exposure and Quantification of Social Behavior in Adult Rats

A Procedure to Observe Context-induced Renewal of Pavlovian-conditioned Alcohol-seeking Behavior in Rats

Environmental contexts in which drugs of abuse are consumed can trigger craving, a subjective Pavlovian-conditioned response that can facilitate drug-seeking behavior and prompt relapse in abstinent drug users. We have developed a procedure to study the behavioral and neural processes that mediate the impact of context on alcohol-seeking behavior in rats. Following acclimation to the taste and pharmacological effects of 15% ethanol in the home cage, male Long-Evans rats receive Pavlovian discrimination training (PDT) in conditioning chambers. In each daily (Mon-Fri) PDT session, 16 trials each of two different 10 sec auditory conditioned stimuli occur. During one stimulus, the CS+, 0.2 ml of 15% ethanol is delivered into a fluid port for oral consumption. The second stimulus, the CS-, is not paired with ethanol. Across sessions, entries into the fluid port during the CS+ increase, whereas entries during the CS- stabilize at a lower level, indicating that a predictive association between the CS+ and ethanol is acquired. During PDT each chamber is equipped with a specific configuration of visual, olfactory and tactile contextual stimuli. Following PDT, extinction training is conducted in the same chamber that is now equipped with a different configuration of contextual stimuli. The CS+ and CS- are presented as before, but ethanol is withheld, which causes a gradual decline in port entries during the CS+. At test, rats are placed back into the PDT context and presented with the CS+ and CS- as before, but without ethanol. This manipulation triggers a robust and selective increase in the number of port entries made during the alcohol predictive CS+, with no change in responding during the CS-. This effect, referred to as context-induced renewal, illustrates the powerful capacity of contexts associated with alcohol consumption to stimulate alcohol-seeking behavior in response to Pavlovian alcohol cues.

A procedure to study the capacity of an alcohol associated environmental context to trigger the renewal of alcohol-seeking behavior in rats is described.

Environmental contexts in which drugs of abuse are consumed can trigger craving, a subjective Pavlovian-conditioned response that can facilitate ...

The Modified Hole Board - Measuring Behavior, Cognition and Social Interaction in Mice and Rats

This protocol describes the modified hole board (mHB), which combines features from a traditional hole board and open field and is designed to measure multiple dimensions of unconditioned behavior in small laboratory mammals (e.g., mice, rats, tree shrews and small primates). This paradigm is a valuable alternative for the use of a behavioral test battery, since a broad behavioral spectrum of an animal&#8217;s behavioral profile can be investigated in one single test.
The apparatus consists of a box, representing the &#8216;protected&#8217; area, separated from a group compartment. A board, on which small cylinders are staggered in three lines, is placed in the center of the box, representing the &#8216;unprotected&#8217; area of the set-up. The cognitive abilities of the animals can be measured by baiting some cylinders on the board and measuring the working and reference memory. Other unconditioned behavior, such as activity-related-, anxiety-related- and social behavior, can be observed using this paradigm. Behavioral flexibility and the ability to habituate to a novel environment can additionally be observed by subjecting the animals to multiple trials in the mHB, revealing insight into the animals&#8217; adaptive capacities.
Due to testing order effects in a behavioral test battery, na&#239;ve animals should be used for each individual experiment. By testing multiple behavioral dimensions in a single paradigm and thereby circumventing this issue, the number of experimental animals used is reduced. Furthermore, by avoiding social isolation during testing and without the need to food deprive the animals, the mHB represents a behavioral test system, inducing if any, very low amount of stress.

This protocol describes the modified hole board, which is a behavioral test set-up that comprises the characteristics of an open field and a traditional hole board. This set-up enables the differential analysis of unconditioned behavior of small laboratory mammals as well as the analysis of cognitive abilities.

This protocol describes the modified hole board (mHB), which combines features from a traditional hole board and open field and is designed to measure ...

Rodent Brain Microinjection to Study Molecular Substrates of Motivated Behavior

Brain microinjection can aid elucidation of the molecular substrates of complex behaviors, such as motivation. For this purpose rodents can serve as appropriate models, partly because the response to behaviorally relevant stimuli and the circuitry parsing stimulus-action outcomes is astonishingly similar between humans and rodents. In studying molecular substrates of complex behaviors, the microinjection of reagents that modify, augment, or silence specific systems is an invaluable technique. However, it is crucial that the microinjection site is precisely targeted in order to aid interpretation of the results. We present a method for the manufacture of surgical implements and microinjection needles that enables accurate microinjection and unlimited customizability with minimal cost. Importantly, this technique can be successfully completed in awake rodents if conducted in conjunction with other JoVE articles that covered requisite surgical procedures. Additionally, there are many behavioral paradigms that are well suited for measuring motivation. The progressive ratio is a commonly used method that quantifies the efficacy of a reinforcer to maintain responding despite an (often exponentially) increasing work requirement. This assay is sensitive to reinforcer magnitude and pharmacological manipulations, which allows reinforcing efficacy and/ or motivation to be determined. We also present a straightforward approach to program operant software to accommodate a progressive ratio reinforcement schedule.

Rodents are an appropriate model to investigate the molecular substrates of behavior and complex psychiatric disorders. Brain microinjection in awake rodents can be used to elucidate disease substrates. An efficient and customizable brain microinjection method as well as the execution of an operant paradigm that quantifies motivation is presented.

Brain microinjection can aid elucidation of the molecular substrates of complex behaviors, such as motivation. For this purpose rodents can serve as ...

The Social Dimension of Stress: Experimental Manipulations of Social Support and Social Identity in the Trier Social Stress Test

In many situations humans are influenced by the behavior of other people and their relationships with them. For example, in stressful situations supportive behavior of other people as well as positive social relationships can act as powerful resources to cope with stress. In order to study the interplay between these variables, this protocol describes two effective experimental manipulations of social relationships and supportive behavior in the laboratory. In the present article, these two manipulations are implemented in the Trier Social Stress Test (TSST)&#8212;a standard stress induction paradigm in which participants are subjected to a simulated job interview. More precisely, we propose (a) a manipulation of the relationship between different protagonists in the TSST by making a shared social identity salient and (b) a manipulation of the behavior of the TSST-selection committee, which acts either supportively or unsupportively. These two experimental manipulations are designed in a modular fashion and can be applied independently of each other but can also be combined. Moreover, these two manipulations can also be integrated into other stress protocols and into other standardized social interactions such as trust games, negotiation tasks, or other group tasks.

Previous research on the social dimension of stress has focused on two important variables: social identity and social support. This protocol introduces an effective experimental manipulation of these two social variables and describes their implementation in a standard stress induction paradigm (Trier Social Stress Test).

In many situations humans are influenced by the behavior of other people and their relationships with them. For example, in stressful situations ...

A Method for Evaluating the Reinforcing Properties of Ethanol in Rats without Water Deprivation, Saccharin Fading or Extended Access Training

Operant oral self-administration methods are commonly used to study the reinforcing properties of ethanol in animals. However, the standard methods require saccharin/sucrose fading, water deprivation and/or extended training to initiate operant responding in rats. This paper describes a novel and efficient method to quickly initiate operant responding for ethanol that is convenient for experimenters and does not require water deprivation or saccharin/sucrose fading, thus eliminating the potential confound of using sweeteners in ethanol operant self-administration studies. With this method, Wistar rats typically acquire and maintain self-administration of a 20% ethanol solution in less than two weeks of training. Furthermore, blood ethanol concentrations and rewards are positively correlated for a 30 min self-administration session. Moreover, naltrexone, an FDA-approved medication for alcohol dependence that has been shown to suppress ethanol self-administration in rodents, dose-dependently decreases alcohol intake and motivation to consume alcohol for rats self-administering 20% ethanol, thus validating the use of this new method to study the reinforcing properties of alcohol in rats.

This protocol describes a novel and efficient method to quickly initiate operant responding for ethanol in rats that, contrary to standard methods, does not require water deprivation or saccharin/sucrose fading to initiate responding.

Operant oral self-administration methods are commonly used to study the reinforcing properties of ethanol in animals. However, the standard methods ...

Examining Recall Memory in Infancy and Early Childhood Using the Elicited Imitation Paradigm

The ability to recall the past allows us to report on details of previous experiences, from the everyday to the significant. Because recall memory is commonly assessed using verbal report paradigms in adults, studying the development of this ability in preverbal infants and children proved challenging. Over the past 30 years, researchers have developed a non-verbal means of assessing recall memory known as the elicited or deferred imitation paradigm. In one variant of the procedure, participants are presented with novel three-dimensional stimuli for a brief baseline period before a researcher demonstrates a series of actions that culminate in an end- or goal-state. The participant is allowed to imitate the demonstrated actions immediately, after a delay, or both. Recall performance is then compared to baseline or to performance on novel control sequences presented at the same session; memory can be assessed for the individual target actions and the order in which they were completed. This procedure is an accepted analogue to the verbal report techniques used with adults, and it has served to establish a solid foundation of the nature of recall memory in infancy and early childhood. In addition, the elicited or deferred imitation procedure has been modified and adapted to answer questions relevant to other aspects of cognitive functioning. The broad utility and application of imitation paradigms is discussed, along with limitations of the approach and directions for future research.

The elicited imitation procedure was established to examine the development of recall memory in infancy and early childhood. This procedure has been widely used to establish a solid foundation of the nature of recall memory in infancy and early childhood.

The ability to recall the past allows us to report on details of previous experiences, from the everyday to the significant. Because recall memory is ...

The Rodent Psychomotor Vigilance Test (rPVT): A Method for Assessing Neurobehavioral Performance in Rats and Mice

The human Psychomotor Vigilance Test (PVT) is a widely used procedure for measuring changes in fatigue and sustained attention. The present article describes a rodent version of the PVT&#8212;termed the &#34;rPVT&#34;&#8212;that measures similar aspects of attention (i.e., performance accuracy, motor speed, premature responding, and lapses in attention). Data are presented that demonstrate both the short- and long-term usefulness of the rPVT when employed with laboratory rats. Rats easily learn the rPVT, and learning to perform the basic procedure takes less than two weeks of training. Once acquired, rat performances in the rPVT show a high degree of similarity to these same performance measures in the human PVT, including similarities in, lapses in attention, reaction times, vigilance decrements across session time (i.e., the human &#34;time-on-task&#34; effects), and the response-stimulus interval (RSI) effect described for humans. Thus the rPVT can be an extremely valuable tool for assessing the effects of a wide range of variables on sustained attention quite similar to human PVT performances, and thus can be useful for developing novel treatments for neurobehavioral dysfunctions.

The Rodent Psychomotor Vigilance Test rPVT: A Method for Assessing Neurobehavioral Performance in Rats and Mice

A rat version of the human Psychomotor Vigilance Test (PVT) is described that measures aspects of attention similar to those measured with the human PVT, including aspects of human vigilance such as performance accuracy, motor speed, premature responding, and lapses in attention.

The human Psychomotor Vigilance Test (PVT) is a widely used procedure for measuring changes in fatigue and sustained attention. The present article ...

Using Clicker Training and Social Observation to Teach Rats to Voluntarily Change Cages

Cage cleaning is a routinely performed husbandry procedure and is known to induce stress in laboratory rats. As stress can have a negative impact on well-being and can affect the comparability and reproducibility of research results, the amount of stress experienced by laboratory animals should be minimized and avoided when possible. Further, the direct contact between the rat and animal caretaker during the cage change bears hygiene risks and therefore possibly negatively impacts the well-being of the rats and the quality of the research. 
Our protocol aims to improve the routinely performed cage changing procedure. For this reason, we present a feasible protocol that enables rats to learn via clicker training and observation to voluntarily change to a clean cage. This training helps to reduce stress caused by the physical disturbance and handling associated with the cage changes and concurrently enables a reduction in direct contact between animal and animal caretaker after the training phase is completed.
The implementation of clicker training to rats is fast and easy. Rats are generally interested in the training and efficiently learn the desired behavior, which entails changing cages through a pipe. Even without training, the rats learn to perform the desired behavior by observation, as 80% of the observational learning group successfully changed cages when tested. The training further helps to establish a relationship of trust between trainer and animal. As hygiene and well-being are both very important in animal experiments, this protocol might also help to improve high-quality research.

This protocol introduces a method of cage change for rats via clicker training. Rats learn the desired behavior not only by direct training but also by observational learning. The implementation of this fast and easy protocol might help to improve well-being and hygiene in rodent facilities.

Cage cleaning is a routinely performed husbandry procedure and is known to induce stress in laboratory rats. As stress can have a negative impact on ...

Integrating Computerized Linguistic and Social Network Analyses to Capture Addiction Recovery Capital in an Online Community

The article describes a new methodology designed with the aim of finding a comprehensive, unobtrusive, and accurate way of capturing social recovery capital development in online communities of recovery from alcohol and drug (AOD) addiction. Recovery capital was conceptualised as both engagement in the online recovery community and identification with the community. To measure recovery capital development, naturally occurring data were extracted from the social media page of a specific recovery program, with the page being set up as a resource for a face-to-face recovery program. To map engagement with the online community, social network analysis (SNA) capturing online social interaction was performed. Social interaction was measured through the linkages between the online contributors/members of the online community as represented by program clients, staff, and supporters from the broader community. To capture markers of social identification with the online community, computerised linguistic analysis of the textual data (content from posts and comments) was conducted. Recovery capital captured in this way was analysed against retention data (a proxy outcome indicator), as days spent in the (face-to-face) recovery program. The online data extracted was linked to participant data in regards to program retention to test prediction of a key recovery outcome. This approach allowed the examination of the role of online support communities and assessment of the association between recovery capital (developed via the online community of recovery) and recovery outcomes.

The article describes a novel approach for analyzing dynamic online social interactions (in an online context) exemplified by a study of an online community of recovery from alcohol and drug addiction.

The article describes a new methodology designed with the aim of finding a comprehensive, unobtrusive, and accurate way of capturing social recovery ...

A Complex Diving-For-Food Task to Investigate Social Organization and Interactions in Rats

For many species, where status is a vital motivator that can affect health, social hierarchies influence behavior. Social hierarchies that include dominant-submissive relationships are common in both animal and human societies. These relationships can be affected by interactions with others and with their environment, making them difficult to analyze in a controlled study. Rather than a simple dominance hierarchy, this formation has a complicated presentation that allows rats to avoid aggression. Status can be stagnant or mutable, and results in complex societal stratifications. Here we describe a complex diving-for-food task to investigate rodent social hierarchy and behavioral interactions. This animal model may allow us to assess the relationship between a wide range of mental illnesses and social organization, as well as to study the effectiveness of therapy on social dysfunction.

This protocol describes a method of examining social hierarchy in a rat model. Rats perform a complex diving-for-food task in which they form a distinct hierarchy according to their willingness to dive underwater and swim to obtain a food pellet. This method is used to understand decision making and social relationships among highly social animals in small groups.