We thank Petra Grutzik for construction of the behavioral cage, Dr. Thomas Borowski for help designing the experiment, and Dr. Stephanie White for her generous support. Funding from an HHMI undergraduate summer research grant (HHMI #30052007536 to D.S. and L.H).

All experiments were approved by the Institutional Care and Use Committee of the W.M. Keck Science Department, in accordance to NIH guidelines. All animals used were adults (&#62;90 days post hatch).
1. Construction of Operant Conditioning Chamber
<ol>
	<li>Build a testing cage composed of two identical chambers, 15 x 15 x 17 inch3 each (Figure 1A). Make a small door (as an entrance to each chamber) on the side of the cage near the perches by cutting out a small 4.5 x 6 inch2 window that is 2.5 inchs away from the center of the cage between the chambers. Use a separate piece of the shelving to make a door that covers the window. 
	NOTE: The cage is made from wire shelving for closets that can be obtained from any home improvement store. The hinge of the door can be made with zip-ties.</li>
	<li>Cut the shelving with wire cutters to make a 4 x 6 inch2 opening between the two chambers to allow the bird to move between chambers.</li>
	<li>In each chamber, place a perch 11 inches from the center of the cage and 6 in from the bottom of the cage. Use any perch that is suitable for the type of bird being tested and that will span the width of the cage.</li>
	<li>Place the emitter and receiver of the photoelectric sensor on the cage directly above each end of the perch. Tether the sensors to the wire above the perch using zip-ties. 
	NOTE: Placing the sensors onto a rigid backing (e.g., a tongue depressor) can help keep the sensors from twisting on the wire cage and becoming misaligned. It is important to test the placement of the sensors prior to each experiment by manually breaking the infrared (IR) beam. If the sensors are not correctly aligned, a bird will not trigger a song, or landing on the perch may be counted as multiple events.</li>
	<li>Place under-cabinet LED lights on the top of the cage to provide enough illumination so that the animals will move freely between the two side chambers.</li>
	<li>Place bird seed and water next to each of the perches.</li>
	<li>Place a speaker at each end of each chamber.</li>
	<li>To reduce sound diffraction and audio and visual distractions, place anechoic foam on the perimeter of the entire cage, with holes left for the cage doors and water bottles.</li>
	<li>Connect the speaker to an audio amplifier and connect the audio amplifier to the sound output on a computer.</li>
</ol>
2. Connection of Sensors
NOTE: There are two sets of IR sensors, each with an emitter and receiver. The emitter has a cable with four wires (brown, blue, black, and white). The receiver has a cable with three wires (brown, blue, and black). The white wire of one emitter is connected directly to input #1 on the digital I/O board. The white wire of the second emitter is connected directly to input #9. The AC power input has two wires (usually red and black).
<ol>
	<li>Install a digital I/O card (Table of Materials), its drivers, and the associated program &#39;Measurement and Automation&#39;. 
	NOTE: The digital I/O card used here requires a connector block and multifunction ribbon cable to complete installation. Alternatively, a USB I/O device requires no PCI card (or the required connector block and ribbon cable accessories), and can be run on a laptop.</li>
	<li>Connect the two blue wires of each emitter and receiver set together. Attach two small wires to the black wire of the power cable so that the black power wire can connect to the blue wires of each sensor set.</li>
	<li>Connect the black power wire and two blue wires to a common wire. Connect this common wire to either input #2 or # 9 on the connector block. 
	NOTE: One set of sensors should be connected to input #1 and #2 and the other set of sensors should be connected to input #9 and #10.</li>
	<li>For both sensor sets, connect the brown wire from the emitter and the receiver together. For each sensor set, connect the two brown wires to the red AC power wire. 
	NOTE: All four brown wires are connected to the red AC power wire. The photoelectric sensors are powered by a 10&#8211;30 V input and must be connected to an AC converter (e.g., a 120 V to 12 V converter).</li>
	<li>Within the Measurement and Automation program associated with the digital I/O card, determine the Device ID for the I/O card by selecting the Devices and Interfaces menu.</li>
	<li>Select &#39;Device Pinouts&#39; option to determine the port and line number that correspond with the inputs in 2.3.</li>
	<li>Verify that the I/O detects when the IR beam is broken using the Test Panels&#8230; option in the program. Note the channel and port IDs for each set of sensors. 
	NOTE: Green indicators will change color when the status of the beam is altered.</li>
</ol>
3. Install Software and Hardware to Count Perch Landings
<ol>
	<li>Download and install Sound Analysis Pro 2011 (SAP2011) and MySQL from http://soundanalysispro.com/. 
	NOTE: This is freeware and comes with installation instructions and a user manual.</li>
	<li>Open SAP recorder.exe.</li>
	<li>To configure the sensor input so that SAP can detect perch landings and initiate song playbacks, click the Operant Devices tab in the SAP Recorder control window. Check the box Enable Operant Training (NI Card Installed).</li>
	<li>Select the appropriate Device ID (the Ports slider can be set to 3). For each detector, indicate the appropriate port and line from the information collected in step 2.6. 
	NOTE: When the port and line are set correctly for each detector, the light will change from yellow to red.</li>
	<li>In the Main window for the SAP recorder, hit Train to activate the sensors for the appropriate channels. 
	NOTE: A yellow button should appear.</li>
	<li>Record the name of the bird by selecting the channel number of interest (Press channel 1 or 2 tab on the left side of the screen), and in the Identification and Mode window, type in the bird&#39;s identification in the Name field.</li>
	<li>To ensure that sound is played to the speakers, manipulate the settings in the Output Selection tab to select the appropriate device connected to the computer (i.e., speakers) and the channel connected to the speaker. 
	NOTE: Additional setup information can be found in the Sound Analysis Pro user manual.</li>
</ol>
4. Isolate Pairs of Birds and Collect Male Song
<ol>
	<li>Isolate a male and a female finch in a sound-attenuation chamber for 24&#8211;48 h. During this cohabitation, collect the male song, which is referred to as the &#39;partner&#39;s song&#39;. 
	NOTE: Paired birds are a male and female that have been housed together for at least 2 weeks. Unpaired females are placed with a male for only 24&#8211;48 h.
	<ol>
		<li>Use a microphone connected to an audio amplifier to capture the partner&#39;s song.</li>
		<li>Acquire the song using Sound Analysis Pro 2011 and store the song on a hard drive.</li>
		<li>To test for the effect of the neurotransmitter systems on female song preference, give the female a 50 &#181;L subcutaneous injection of saline (vehicle control) or quinpirole (1 mg/mL in 0.9% saline) when the pair is first put into the chamber. Repeat the injection after 6&#8211;24 h (see Figure 1B).</li>
	</ol>
	</li>
	<li>Create songs for playback in the operant conditioning cage.
	<ol>
		<li>Create a .wav file that plays &#39;silence&#39; approximately the same length as the song playbacks. 
		NOTE: This can be done using any sound-editing software (e.g., Audacity).</li>
		<li>Cut a representative song (2&#8211;3 motifs) from the partner male.</li>
		<li>Cut a representative song (2&#8211;3 motifs) from an unfamiliar male. 
		NOTE: Use the same unfamiliar male song for all experiments.</li>
		<li>Filter the songs from 300 Hz&#8211;10 kHz.</li>
		<li>Adjust the volume so that the songs play at ~70 dB (average amplitude over the song duration from the speakers in the behavior chamber). 
		NOTE: The volume of the song is tested using a sound pressure meter that is placed at the perch.</li>
	</ol>
	</li>
</ol>
5. Testing Song Preference in Paired Females
<ol>
	<li>Determine the female&#39;s side chamber preference.
	<ol>
		<li>Prior to the behavioral testing, place the female in the testing cage to allow her time to adjust to the cage for at least 1 h. Ensure the finch explores both side chambers by using an object to cause the bird to move from one chamber to the other through the opening.</li>
		<li>In Sound Analysis Pro, select the &#39;Playbacks&#39; tab and then the &#39;Sounds&#39; button in the main window.</li>
		<li>Select the audio .wav file to play &#39;Silence&#39; from chamber 1 and chamber 2.</li>
		<li>Go back to &#39;Main&#39; and hit &#39;Reset&#39; on the top of the boxes on the right.</li>
		<li>After the acclimation period, press &#39;Start&#39;.</li>
		<li>At the end of the session, press &#39;Stop&#39;. Write down the number of triggers at each perch, which is displayed in the blue boxes. 
		NOTE: The data are saved to a MySQL database. Have a minimum number of triggers to count the trial to ensure the female is active enough (e.g., 12 or more perches total).</li>
	</ol>
	</li>
	<li>Determine song preference.
	<ol>
		<li>Repeat the steps in section 5.1, but choose the partner&#39;s song to play from the side chamber with the fewest perch triggers (the non-preferred side), and an unfamiliar song from the side chamber with the most perch triggers (the preferred side). 
		NOTE: The partner&#39;s song is played from the side chamber with the fewest triggers during the playback of silence (the non-preferred side).</li>
		<li>Press &#39;reset&#39; prior to starting the 1 h with song playback.</li>
		<li>Calculate the chamber and song preference by dividing the number of triggers on the side chamber playing the partner&#39;s song by the total number of triggers.</li>
	</ol>
	</li>
</ol>

<ol>
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J., Wang, Z. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+neurobiology+of+pair+bonding.">The neurobiology of pair bonding.</a> Nature Neuroscience. 7 (10), 1048-1054 (2004).</li><li>Aragona, B. J., Liu, Y., Curtis, J. T., Stephan, F. K., Wang, Z. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+critical+role+for+nucleus+accumbens+dopamine+in+partner-preference+formation+in+male+prairie+voles.">A critical role for nucleus accumbens dopamine in partner-preference formation in male prairie voles.</a> Journal of Neuroscience. 23 (8), 3483-3490 (2003).</li><li>Lippert, R. N., 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=Time-dependent+assessment+of+stimulus-evoked+regional+dopamine+release.">Time-dependent assessment of stimulus-evoked regional dopamine release.</a> Nature Communications. 10 (1), 336 (2019).</li><li>Lack, D. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Ecological adaptations for breeding in birds. , (1968).</li><li>O'Connell, L. A., Hofmann, H. 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M., Vyas, A., Woolley, S. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Automated+auditory+recognition+training+and+testing.">Automated auditory recognition training and testing.</a> Animal Behavior. 82 (2), 285-293 (2011).</li><li>Woolley, S. C., Doupe, A. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Social+context-induced+song+variation+affects+female+behavior+and+gene+expression.">Social context-induced song variation affects female behavior and gene expression.</a> PLoS Biology. 6 (3), 62 (2008).</li><li>Day, N. 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The authors have nothing to disclose.

We describe a method to test the song preference of zebra finches. We used this assay to test the preference of a paired female for her partner's song. In addition, the assay was used to test the effect of dopamine on induction of song preference in naive females. This method is relatively inexpensive and was designed and used by undergraduate students, making it an excellent training tool for students. Several other studies have tested song preference in several song species, making this method useful to many investigat...

An erratum was issued for:&#160;<a href="https://www.jove.com/video/60590/operant-conditioning-task-to-measure-song-preference-in-zebra-finches">Operant Conditioning Task to Measure Song Preference in Zebra Finches</a>. An affiliation&#160;was updated.
One of the affiliations was updated from:
W.M. Keck Science Department, Clarmont McKenna College &#8211; Pitzer College &#8211; Scripps College
to:
W.M. Keck Science Department, Claremont McKenna College &#8211; Pitzer College &#8211; Scripps College

An erratum was issued for:&#160;<a href="https://www.jove.com/video/60590/operant-conditioning-task-to-measure-song-preference-in-zebra-finches">Operant Conditioning Task to Measure Song Preference in Zebra Finches</a>. An affiliation&#160;was updated.

Erratum: Operant Conditioning Task to Measure Song Preference in Zebra Finches

One of the fundamental questions in biology is how animals form affiliative bonds. In particular, what are the neural mechanisms by which these bonds are made and are these basic mechanisms conserved across vertebrate species? The prairie vole has given clues to some of the neurotransmitter systems that are important for pair-bond formation1,2,3. In particular, dopamine acting through receptors in the nucleus accumbens can induce partner preference in both male and female voles4,5. It is unclear whether the general principles underlying partner preference formation and maintenance are evolutionarily conserved.
Monogamy is more common in birds than in mammals6. Therefore, comparing mechanisms of affiliative behavior in birds to other species is critical to understanding conserved neural foundations7,8,9. In general, across many species of songbirds, male song is thought to serve as an honest indicator of male fitness10,11. Male zebra finch songbirds sing to attract mates and influence the formation of monogamous pairs12,13. Thus, song can be used to determine partner preference in these birds.
The mechanisms by which females form a preference for their partner's song is unknown. Mesotocin, the avian homologue of oxytocin, appears to play a role in pair-bond formation in finches14,15. In addition, dopamine has also been shown to play a role in pair-bond formation9,16,17,18. For example, dopamine levels are higher in the nucleus accumbens of paired versus non-paired finches9.
To study the role of neurotransmitter systems on pair-bond formation and maintenance, we measured song preference using an operant testing paradigm equipped with infrared sensors to trigger song playbacks19. The ratio of song playbacks identified the female's preference for a male's song. Prior to the operant conditioning task, each female was isolated for up to 48 h in an anechoic chamber with her partner for 'paired females' or with an unfamiliar male, for 'unpaired females'. To test the effect of dopamine on song preference, unpaired females were treated twice with a D2R dopamine agonist while isolated with the unfamiliar male. This behavioral paradigm is based on previous studies19,20 and is amenable to research by undergraduate students.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>(-)-Quinpirole</td>
			<td>sigmaaldrich</td>
			<td>Q102</td>
			<td></td>
		</tr>
		<tr>
			<td>acoustic foam</td>
			<td>Coulbourn Instruments, Allentown, PA</td>
			<td>H10-24A</td>
			<td></td>
		</tr>
		<tr>
			<td>audio amplifier - 2 channel</td>
			<td>Amazon</td>
			<td>Pyle PCAU46A</td>
			<td>any small audio amplifier should work</td>
		</tr>
		<tr>
			<td>Banner Engineering Q08 Series, emitter. SO60-Q08</td>
			<td><a href="https://www.alliedelec.com" target="_blank">https://www.alliedelec.com</a></td>
			<td>Stock #:70659809</td>
			<td>photoelectric sensor</td>
		</tr>
		<tr>
			<td>Banner Engineering Q08 series, receiver. EO60-Q08-AN6X</td>
			<td><a href="https://www.alliedelec.com" target="_blank">https://www.alliedelec.com</a></td>
			<td>Stock #:70699384</td>
			<td>photoelectric sensor</td>
		</tr>
		<tr>
			<td>Car stereo speakers</td>
			<td>Amazon</td>
			<td></td>
			<td>Pioneer TS - F1643R</td>
		</tr>
		<tr>
			<td>Digital I/O card</td>
			<td>National Instruments</td>
			<td>PCI-6503 or USB-6501</td>
			<td></td>
		</tr>
		<tr>
			<td>LED lights - under-counter</td>
			<td>Amazon</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>multifunctional ribbon cable</td>
			<td>National Instruments</td>
			<td>180524-20</td>
			<td></td>
		</tr>
		<tr>
			<td>sound pressure level meter</td>
			<td>Amazon</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>terminal block</td>
			<td>National Instruments</td>
			<td>777101-1</td>
			<td></td>
		</tr>
	</tbody>
</table>

operant conditioning task to measure song preference in zebra finches

An operant conditioning paradigm is used to test the song preference of female zebra finches. Finches are placed in a two-chambered cage with a connecting opening and indicate their preference for a song by landing on a perch within each chamber. By interrupting the infrared beam from a photoelectric sensor above each perch, the bird activates the playback of a song through a speaker located on each side of the cage. Freely available software is used to trigger the song playback from each perch. To determine the song preference of each animal, her chamber preference is first identified by triggering no song playback when she lands on each perch. This chamber preference is then compared to her song preference. A minimum activity threshold is set to ensure the preference is real. Using this method, we show that paired females prefer the song of their partner. This method was used to understand the contribution of dopamine to the formation and maintenance of song preference.

Following the protocol, we found that paired females preferred their partner's song (Figure 2A). There was a significant difference between the side chamber preference during silence to that during song playback (t-test corrected for multiple comparisons; p = 0.004; t = 3.35, df = 16). Thus, the female preferentially triggered the song of her partner in comparison to the song of an unfamiliar male.
Females that were paired with an unfamiliar male ...

Watch this Scientific Journal Video about Operant Conditioning Task to Measure Song Preference in Zebra Finches at JoVE.com

Operant Conditioning Task to Measure Song Preference in Zebra Finches

We describe a technique to evaluate song preference in zebra finches. Females are placed in a two-chambered cage and song preference is measured by the number of times she triggers the playback of one song by landing on a perch within one chamber, compared with triggering a different song in the second chamber. Perch landings are counted using infrared sensors.

An operant conditioning paradigm is used to test the song preference of female zebra finches. Finches are placed in a two-chambered cage with a connecting ...

operant-conditioning-task-to-measure-song-preference-in-zebra-finches

Research

JoVE Journal

Behavior

6.3K Views.  Claremont McKenna College – Pitzer College – Scripps College. We describe a technique to evaluate song preference in zebra finches. Females are placed in a two-chambered cage and song preference is measured by the number of times she triggers the playback of one song by landing on a perch within one chamber, compared with triggering a different song in the second chamber. Perch landings are counted using infrared sensors.

Video: Operant Conditioning Task to Measure Song Preference in Zebra Finches

Use of the Operant Orofacial Pain Assessment Device (OPAD) to Measure Changes in Nociceptive Behavior

We present an operant system for the detection of pain in awake, conscious rodents. The Orofacial Pain Assessment Device (OPAD) assesses pain behaviors in a more clinically relevant way by not relying on reflex-based measures of nociception. Food fasted, hairless (or shaved) rodents are placed into a Plexiglas chamber which has two Peltier-based thermodes that can be programmed to any temperature between 7 &deg;C and 60 &deg;C. The rodent is trained to make contact with these in order to access a reward bottle. During a session, a number of behavioral pain outcomes are automatically recorded and saved. These measures include the number of reward bottle activations (licks) and facial contact stimuli (face contacts), but custom measures like the lick/face ratio (total number of licks per session/total number of contacts) can also be created. The stimulus temperature can be set to a single temperature or multiple temperatures within a session. The OPAD is a high-throughput, easy to use operant assay which will lead to better translation of pain research in the future as it includes cortical input instead of relying on spinal reflex-based nociceptive assays.

Use of the Operant Orofacial Pain Assessment Device OPAD to Measure Changes in Nociceptive Behavior

We present a user-friendly, high-throughput operant system for the evaluation of pain behaviors in awake, conscious rodents. The Orofacial Pain Assessment Device (OPAD) can assess pain through a reward/conflict paradigm thus providing a more humane way of testing. This protocol will yield more clinically relevant and translational data from rodents.

We present an operant  system for the detection of pain in awake, conscious rodents. The Orofacial  Pain Assessment Device (OPAD) assesses pain behaviors ...

The Crossmodal Congruency Task as a Means to Obtain an Objective Behavioral Measure in the Rubber Hand Illusion Paradigm

The rubber hand illusion (RHI) is a popular experimental paradigm. Participants view touch on an artificial rubber hand while the participants' own hidden hand is touched. If the viewed and felt touches are given at the same time then this is sufficient to induce the compelling experience that the rubber hand is one's own hand. The RHI can be used to investigate exactly how the brain constructs distinct body representations for one's own body. Such representations are crucial for successful interactions with the external world. To obtain a subjective measure of the RHI, researchers typically ask participants to rate statements such as "I felt as if the rubber hand were my hand". Here we demonstrate how the crossmodal congruency task can be used to obtain an objective behavioral measure within this paradigm.
The variant of the crossmodal congruency task we employ involves the presentation of tactile targets and visual distractors. Targets and distractors are spatially congruent (i.e. same finger) on some trials and incongruent (i.e. different finger) on others. The difference in performance between incongruent and congruent trials - the crossmodal congruency effect (CCE) - indexes multisensory interactions. Importantly, the CCE is modulated both by viewing a hand as well as the synchrony of viewed and felt touch which are both crucial factors for the RHI.
The use of the crossmodal congruency task within the RHI paradigm has several advantages. It is a simple behavioral measure which can be repeated many times and which can be obtained during the illusion while participants view the artificial hand. Furthermore, this measure is not susceptible to observer and experimenter biases. The combination of the RHI paradigm with the crossmodal congruency task allows in particular for the investigation of multisensory processes which are critical for modulations of body representations as in the RHI.

We demonstrate how an objective measure can be employed in the widely employed rubber hand illusion paradigm. This measure is obtained by modifying the well-established crossmodal congruency task. This task allows the investigation of multisensory processes which are critical for modulations of body representations as in the rubber hand illusion.

The rubber hand illusion (RHI) is a popular experimental paradigm. Participants view touch on an artificial rubber hand while the participants' own hidden ...

Stereotaxic Microinjection of Viral Vectors Expressing Cre Recombinase to Study the Role of Target Genes in Cocaine Conditioned Place Preference

Microinjecting recombinant adenoassociated viral (rAAV) vectors expressing Cre recombinase into distinct mouse brain regions to selectively knockout genes of interest allows for enhanced temporally- and regionally-specific control of gene deletion, compared to existing methods. While conditional deletion can also be achieved by mating mice that express Cre recombinase under the control of specific gene promoters with mice carrying a floxed gene, stereotaxic microinjection allows for targeting of discrete brain areas at experimenter-determined time points of interest. In the context of cocaine conditioned place preference, and other cocaine behavioral paradigms such as self-administration or psychomotor sensitization that can involve withdrawal, extinction and/or reinstatement phases, this technique is particularly useful in exploring the unique contribution of target genes to these distinct phases of behavioral models of cocaine-induced plasticity. Specifically, this technique allows for selective ablation of target genes during discrete phases of a behavior to test their contribution to the behavior across time. Ultimately, this understanding allows for more targeted therapeutics that are best able to address the most potent risk factors that present themselves during each phase of addictive behavior.

This article describes how to microinject viral vectors into mouse brain and then test in a conditioned place preference paradigm that includes an acquisition, extinction and reinstatement phase.

Microinjecting recombinant adenoassociated viral (rAAV) vectors expressing Cre recombinase into distinct mouse brain regions to selectively knockout genes ...

Trace Fear Conditioning in Mice

In this experiment we present a technique to measure learning and memory. In the trace fear conditioning protocol presented here there are five pairings between a neutral stimulus and an unconditioned stimulus. There is a 20 sec trace period that separates each conditioning trial. On the following day freezing is measured during presentation of the conditioned stimulus (CS) and trace period. On the third day there is an 8 min test to measure contextual memory. The representative results are from mice that were presented with the aversive unconditioned stimulus (shock) compared to mice that received the tone presentations without the unconditioned stimulus. Trace fear conditioning has been successfully used to detect subtle learning and memory deficits and enhancements in mice that are not found with other fear conditioning methods. This type of fear conditioning is believed to be dependent upon connections between the medial prefrontal cortex and the hippocampus. One current controversy is whether this method is believed to be amygdala-independent. Therefore, other fear conditioning testing is needed to examine amygdala-dependent learning and memory effects, such as through the delay fear conditioning.

In the following experiment we describe a protocol for trace fear conditioning in mice. This type of associative memory includes a trace period that separates the neutral stimulus and the unconditioned stimulus.

In this experiment we present a technique to measure learning and memory. In the trace fear conditioning protocol presented here there are five pairings ...

Fat Preference: A Novel Model of Eating Behavior in Rats

Obesity is a growing problem in the United States of America, with more than a third of the population classified as obese. One factor contributing to this multifactorial disorder is the consumption of a high fat diet, a behavior that has been shown to increase both caloric intake and body fat content. However, the elements regulating preference for high fat food over other foods remain understudied.
To overcome this deficit, a model to quickly and easily test changes in the preference for dietary fat was developed. The Fat Preference model presents rats with a series of choices between foods with differing fat content. Like humans, rats have a natural bias toward consuming high fat food, making the rat model ideal for translational studies. Changes in preference can be ascribed to the effect of either genetic differences or pharmacological interventions. This model allows for the exploration of determinates of fat preference and screening pharmacotherapeutic agents that influence acquisition of obesity.

Dietary fat content influences both energy intake and body fat composition in mammals. By examining rats&#8217; preference for high fat food in a series of choice experiments, it is possible to test genetic differences and pharmacological interventions on their preference for high fat food.

Obesity is a growing problem in the United States of America, with more than a third of the population classified as obese. One factor contributing to ...

The Attentional Set Shifting Task: A Measure of Cognitive Flexibility in Mice

Cognitive impairment, particularly involving dysfunction of circuitry within the prefrontal cortex (PFC), represents a core feature of many neuropsychiatric and neurodevelopmental disorders, including depression, post-traumatic stress disorder, schizophrenia and autism spectrum disorder. Deficits in cognitive function also represent the most difficult symptom domain&#160;to successfully treat, as serotonin reuptake inhibitors and tricyclic antidepressants have only modest effects. Functional neuroimaging studies and postmortem analysis of human brain tissue implicate the PFC as being a primary region of dysregulation in patients with these disorders. However, preclinical behavioral assays used to assess these deficits in mouse models which can be readily manipulated genetically and could provide the basis for studies of new treatment avenues have been underutilized. Here we describe the adaptation of a behavioral assay, the attentional set shifting task (AST), to be performed in mice to assess prefrontal cortex mediated cognitive deficits. The neural circuits underlying behavior during the AST are highly conserved across humans, nonhuman primates and rodents, providing excellent face, construct and predictive validity.

The goal of this protocol is to perform a behavioral assay such as the attentional set shifting task (AST) to assess prefrontal cortex-mediated cognitive flexibility in mice.

Cognitive impairment, particularly involving dysfunction of circuitry within the prefrontal cortex (PFC), represents a core feature of many ...

Operant Procedures for Assessing Behavioral Flexibility in Rats

Executive functions consist of multiple high-level cognitive processes that drive rule generation and behavioral selection. An emergent property of these processes is the ability to adjust behavior in response to changes in one&#8217;s environment (i.e., behavioral flexibility). These processes are essential to normal human behavior, and may be disrupted in diverse neuropsychiatric conditions, including schizophrenia, alcoholism, depression, stroke, and Alzheimer&#8217;s disease. Understanding of the neurobiology of executive functions has been greatly advanced by the availability of animal tasks for assessing discrete components of behavioral flexibility, particularly strategy shifting and reversal learning. While several types of tasks have been developed, most are non-automated, labor intensive, and allow testing of only one animal at a time. The recent development of automated, operant-based tasks for assessing behavioral flexibility streamlines testing, standardizes stimulus presentation and data recording, and dramatically improves throughput. Here, we describe automated strategy shifting and reversal tasks, using operant chambers controlled by custom written software programs. Using these tasks, we have shown that the medial prefrontal cortex governs strategy shifting but not reversal learning in the rat, similar to the dissociation observed in humans. Moreover, animals with a neonatal hippocampal lesion, a neurodevelopmental model of schizophrenia, are selectively impaired on the strategy shifting task but not the reversal task. The strategy shifting task also allows the identification of separate types of performance errors, each of which is attributable to distinct neural substrates. The availability of these automated tasks, and the evidence supporting the dissociable contributions of separate prefrontal areas, makes them particularly well-suited assays for the investigation of basic neurobiological processes as well as drug discovery and screening in disease models.

The ability to assess executive functions such as behavioral flexibility in rats is useful for investigating the neurobiology of cognition in both intact animals and disease models. Here we describe automated tasks for assessing strategy shifting and reversal learning, which are particularly sensitive to disruptions in prefrontal cortical networks.

Executive functions consist of multiple high-level cognitive processes that drive rule generation and behavioral selection. An emergent property of these ...

Quantifying Social Motivation in Mice Using Operant Conditioning

In this protocol, social motivation is measured in mice through a pair of operant conditioning paradigms. To conduct the experiments, two-chambered shuttle boxes were equipped with two operant levers (left and right) and a food receptacle in one chamber, which was then divided from the second chamber by an automated guillotine door covered by a wire grid. Different stimulus mice, rotated across testing days, served as a social stimulus behind the wire grid, and were only visible following the opening of the guillotine door. Test mice were trained to lever press in order to open the door and gain access to the stimulus partner for 15 sec. The number of lever presses required to obtain the social reward progressively increased on a fixed schedule of 3. Testing sessions ended after test mice stopped lever pressing for 5 consecutive minutes. The last reinforced ratio or breakpoint can be used as a quantitative measure of social motivation. For the second paradigm, test mice were trained to discriminate between left and right lever presses in order to obtain either a food reward or the social reward. Mice were rewarded for every 3 presses of each respective lever. The number of food and social rewards can be compared as a measurement of the value placed upon each reward. The ratio of each reward type can also be compared between mouse strains and the change in this ratio can be monitored within testing sessions to measure satiation with a given reward type. Both of these operant conditioning paradigms are highly useful for the quantification of social motivation in mouse models of autism and other disorders of social behavior.

This article describes a new protocol for the quantitative measurement of social motivation in mice using operant conditioning for a social reward. The protocol is useful for the measurement of social motivation in mouse models of autism and other disorders of social behavior.

In this protocol, social motivation is measured in mice through a pair of operant conditioning paradigms. To conduct the experiments, two-chambered ...

Whisker-signaled Eyeblink Classical Conditioning in Head-fixed Mice

Eyeblink conditioning is a common paradigm for investigating the neural mechanisms underlying learning and memory. To better utilize the extensive repertoire of scientific techniques available to study learning and memory at the cellular level, it is ideal to have a stable cranial platform. Because mice do not readily tolerate restraint, they are usually trained while moving about freely in a chamber. Conditioned stimulus (CS) and unconditioned stimulus (US) information are delivered and eyeblink responses recorded via a tether connected to the mouse's head. In the head-fixed apparatus presented here, mice are allowed to run as they desire while their heads are secured to facilitate experimentation. Reliable conditioning of the eyeblink response is obtained with this training apparatus, which allows for the delivery of whisker stimulation as the CS, a periorbital electrical shock as the US, and analysis of electromyographic (EMG) activity from the eyelid to detect blink responses.

The preparation presented here for whisker-signaled eyeblink conditioning in head-fixed mice precisely stimulates specific whiskers while allowing mice to ambulate on a cylindrical treadmill. A whisker stimulation conditioned stimulus (CS) paired with a periorbital shock unconditioned stimulus (US) results in reliable associative learning on this apparatus.

Eyeblink conditioning is a common paradigm for investigating the neural mechanisms underlying learning and memory. To better utilize the extensive ...

An Operant Intra-/Extra-dimensional Set-shift Task for Mice

Alterations in executive control and cognitive flexibility, such as attentional set-shifting abilities, are core features of several neuropsychiatric diseases. The most widely used neuropsychological tests for the evaluation of attentional set-shifting in human subjects are the Wisconsin Card Sorting Test (WCST) and the CANTAB Intra-/Extra-dimensional set shift task (ID/ED). These tasks have proven clinical relevance and have been modified and successfully adapted for research in animal models. However, currently available tasks for rodents present several limitations, mainly due to their manual-based testing procedures, which are hampering translational advances in psychiatric medicine. To overcome these limitations and to better mimic the original version in primates, we present the development of a novel operant-based two-chamber ID/ED "Operon" task for rodents. We demonstrated the effectiveness of this novel task to measure different facets of cognitive flexibility in mice including attentional set formation and shifting, and reversal learning. Moreover, we show the high flexibility of this task in which three different perceptual dimensions can be manipulated with a high number of stimuli cues for each dimension. This novel ID/ED Operon task can be an effective preclinical tool for drug testing and/or large genetic screening relevant to the study of executive dysfunction and cognitive symptoms found in psychiatric disorders.

Attentional set-shifting cognitive abnormalities are a core feature of many psychiatric diseases. Here, we present the development of a novel automatic system for the effective study of attentional set-shifting abilities, executive functions and other cognitive abilities in mice with high translational validity to human studies.

Alterations in executive control and cognitive flexibility, such as attentional set-shifting abilities, are core features of several neuropsychiatric ...