Financial support for this study was provided by the Intramural Research Program of the National Institute of Mental Health, ZIAMH002798 (ClinicalTrial.gov Identifier: NCT00026559: Protocol ID 01-M-0185).

All participants gave written informed consent approved by the National Institute of Mental Health (NIMH) Combined Neuroscience Institutional Review Board (IRB) and were compensated for participating.
1. Set Up the Equipment
NOTE: Set up the equipment as described below (see Figure 1A)3.
<ol>
	<li>In the control room, set up two computers, one to administer the experiment, and one to record the physiological data.</li>
	<li>In the subject room, set up a standard 19 in liquid crystal display monitor and keyboard (or button box) to display stimuli to the participant and to record participant responses, respectively.</li>
	<li>To record the psychophysiology, connect the recording computer to the psychophysiology monitoring hardware using an Ethernet to USB adapter.</li>
	<li>To split the Transistor-Transistor Logic (TTL) signals between the recording and stimulus delivery hardware, connect the parallel port of the experiment computer to the breakout box using a ribbon cable.</li>
	<li>To pass the TTL signals to the psychophysiology monitoring hardware, connect the breakout box to the hardware using a ribbon cable.</li>
	<li>To pass the TTL pulses to the stimulus delivery hardware, connect the breakout box to the signal generator using a Bayonet Neill-Concelman (BNC) cable.</li>
	<li>To generate a control signal for the shock device, connect the signal generator to the shock device using a BNC cable.</li>
	<li>Set the signal generator and shock device to deliver a 100 ms, 200 Hz shock. See Figure 1B and C for all settings.</li>
</ol>
2. Program the Experiment Using Available Software
NOTE: A neurobehavioral systems software (here referred to as the experimental software; see the Table of Materials) was used. Other equivalent software can be used.
<ol>
	<li>Program four testing phases using the parameters described below and the supplementary code files provided (see the supplemental code for details).
	<ol>
		<li>For each phase, program 26 trials.</li>
		<li>Divide the trials into 4 alternating blocks of threat and safety, with 6 trials per block.</li>
		<li>At the beginning of each trial, present a cue indicating how many letters will be presented for 2,000 ms each.</li>
		<li>After the cue, present the encoding letter sequence for 2,500 &#177;1,000 ms.</li>
		<li>On low-load trials, present 5 letters sequentially, one after the other.</li>
		<li>On high-load trials, present 8 letters sequentially, one after the other.</li>
		<li>Program a maintenance period following the encoding phase for 9,000 &#177;1,000 ms.</li>
		<li>At the end of the maintenance period, present a response prompt for 2,000 ms.</li>
		<li>Program the response prompt to display a letter on the left side and a number on the right side of the monitor, with the letter representing a letter from the encoding sequence, and the number referring to a position in the sequence.</li>
		<li>Underneath the letter and number, display the words &#34;match/mismatch,&#34; referring to if the letter matched or mismatched the position number.</li>
		<li>Program the experiment so that half of the trials match and half mismatch.</li>
		<li>Use a keyboard or button box to record responses.</li>
		<li>Separate the trials by a variable-duration intertrial interval (ITI) that is dependent upon the timing of the within-trial events such that each trial is 23 s long.</li>
		<li>Vary the durations of the encoding, maintenance, and ITI periods across trials by selecting a random duration (in ms) between the ceiling and floor values for each period.</li>
		<li>Counterbalance the experiments to have half of the participants start in a safe block and half of the participants start in a threat block.</li>
		<li>In each run, present between 0 and 2 pseudorandom shocks during each of the threat blocks for a total of 3 shock presentations per run. Make sure to include an extra (dummy) trial for each shock to ensure that the same number of trials are included in the safe and threat blocks.</li>
		<li>At the beginning of each run, present five 40 ms bursts of 103 dB white noise (near-instantaneous rise/fall times) over headphones to habituate the startle response.</li>
		<li>During each run, present 3 presentations of the white noise during the following conditions to probe the startle response (see Figure 3): safe versus threat, low load versus high load, and maintenance period versus ITI.</li>
		<li>Space the probes so that they occur with a minimum interprobe interval of at least 17 s to avoid the short-term habituation of the startle response.</li>
		<li>For maintenance period trials, present probes no less than 1 s after the offset of the letter series.</li>
		<li>For ITI trials, present probes no less than 4 s after the offset of the response prompt.</li>
		<li>Set up the equipment for physiological monitoring using the associated software package, as per the manufacturer&#39;s instructions.</li>
	</ol>
	</li>
</ol>
3. Run the Experiment
<ol>
	<li>Escort the participants to the study room.</li>
	<li>Administer informed consent.</li>
	<li>Give the participants the State-Trait Anxiety Inventory Y-1 (STAI-Y1)25, the Beck Anxiety Inventory (BAI)26, the Beck Depression Inventory (BDI)27, and the Anxiety Sensitivity Index (ASI)28 to fill out prior to task instructions and set up.</li>
	<li>Inform the participants that they will see 2 types of trials and will respond to those trials based on the following details.</li>
	<li>During the low-load trials, instruct the participants to maintain a series of 5 letters in their memory in the order in which they are presented.</li>
	<li>During the high-load trials, instruct the participants to maintain a series of 8 letters in their memory in the order in which they are presented.</li>
	<li>Inform the participants that, after a delay, they will be prompted with a letter and a number that refers to the position in the sequence.</li>
	<li>Instruct the participants to indicate whether the letter and position number match or mismatch the trial sequence using the left or right arrow key, respectively.</li>
	<li>Inform the participants that trials will occur during periods of safety and periods of threat, when they are at risk of receiving unpredictable mild electrical shocks to the wrist.</li>
	<li>Inform the participants that they will hear acoustic startle probes throughout the experiment, in both the safe and threat conditions.</li>
	<li>Clean and attach electrodes to each participant, based on the diagram in Figure 2.
	<ol>
		<li>Place two disposable 11 mm silver-silver chloride (Ag-AgCl) electrodes on the palm of the left hand, about 2 cm apart, to monitor skin conductance.</li>
		<li>Place two disposable 11 mm Ag-AgCl electrodes on the inner wrist of the left hand, about 3 cm apart, to administer the electrical stimulation.</li>
		<li>Place one disposable 11 mm Ag-AgCl electrode on the inside of the left arm, just above the elbow, and one disposable electrode just under the right clavicle to monitor the heart rate.</li>
		<li>Attach two 4 mm Ag-AgCl cup electrodes to the lower side of the left orbicularis oculi muscle to measure the startle response.</li>
	</ol>
	</li>
	<li>Secure all electrodes with biomedical tape.</li>
	<li>Attach leads to the electrodes on the palm and plug them into the EDA channel of the psychophysiology monitoring hardware.</li>
	<li>Attach leads to the electrodes on the wrist and plug them into the shock device.</li>
	<li>Attach leads to the electrodes on the arm and clavicle and plug them into the ECG channel of the psychophysiology monitoring hardware.</li>
	<li>Plug the cup electrodes attached to the orbicularis oculi muscle into the electromyography (EMG) channel of the psychophysiology monitoring hardware.</li>
	<li>Shock calibration.
	<ol>
		<li>Prior to the start of the experiment, have the participants rate a series of 100-ms sample electrical stimulations to identify an intensity level that is unpleasant and uncomfortable, but not painful.
		<ol>
			<li>Administer a series presentation (~5-10) of the 100 ms shock stimulus to the wrist using the experimental software package (see supplemental code files and the Table of Materials).</li>
			<li>After each presentation, have participants verbally rate each presentation on a scale from 1 (not uncomfortable at all) to 10 (uncomfortable but not painful).</li>
			<li>Using the mA scale on the shock device, gradually increase the intensity of the shock and continue the series of stimulations the until subject rates the stimulation as a &#34;10.&#34;</li>
			<li>Record the intensity value on the participant detail packet. 
			NOTE: During the study, present the shocks at the determined intensity.</li>
		</ol>
		</li>
	</ol>
	</li>
	<li>To start the experiment, enter the participant ID number, counterbalance condition, and run number into the run box, as prompted by the experiment software. 
	NOTE: Create two counterbalanced conditions. The first counterbalance will start the experiment in a threat block, and the second counterbalance will start the experiment in a safe block. See section 2.</li>
	<li>Click &#34;start&#34; on the psychophysiology monitoring recording.</li>
	<li>Press &#34;enter&#34; on the experimental software prompt box to start the experiment.</li>
	<li>Allow the subject to complete 4 runs of the experiment. Have the participant select the left or right arrow key if the letter and position number match or mismatch the trial sequence, respectively (steps 3.7 and 3.8). 
	NOTE: Program each run length to last between 6 and 7 min. Program the shocks to be delivered pseudo-randomly between 0-2 times/run. See section 2.</li>
	<li>After each run, have the subject verbally rate their anxiety level on a scale from 0 (not anxious) - 10 (extremely anxious) during the safe and threat blocks of the run they just completed.</li>
	<li>Have the subjects verbally rate the intensity of the shocks presented during the previous run on the same 0-10 scale used in the initial calibration procedure (section 3.17).</li>
</ol>
4. Analyze the Performance
NOTE: Analyze the performance data for a single participant using the following instructions.
<ol>
	<li>Open the output file created from the experiment software.
	<ol>
		<li>To average the correct responses across the different conditions, first separate the data into safe versus threat and low load versus high load to yield 4 unique conditions of response data.</li>
		<li>Count the correct trials for each of the 4 conditions and divide this number by the total number of trials in each condition.</li>
		<li>To average the reaction time across the different conditions, separate the data as in step 4.1.1.</li>
		<li>Sum all the reaction times for each condition and divide this number by the number of trials in each condition. 
		NOTE: Omit trials that include a shock presentation, as indicated in the experimental software output.</li>
	</ol>
	</li>
	<li>At the group level, perform a 2 (safe versus threat) x 2 (low load versus high load) ANOVA across subjects to identify differences in behavioral performance and reaction times29.</li>
</ol>
5. Analyze Startle
<ol>
	<li>Prepare the raw EMG data for analysis using psychophysiology analysis software30. See Figure 4A.
	<ol>
		<li>Select &#34;Transform&#34; &#62;&#62; Digital Filters &#62;&#62; FIR &#62;&#62; Bandpass from the psychophysiology analysis software to apply a digital bandpass filter (30-300 Hz passband), smoothing the raw EMG channel (see Figure 4B).</li>
	</ol>
	</li>
	<li>Select Analysis &#62;&#62; Electromyography &#62;&#62; Derive Average Rectified EMG from the psychophysiology analysis software to rectify the smoothed EMG signal using a time-windowed mean of 20 ms (see Figure 4C).</li>
	<li>Select Analysis &#62;&#62; Stim-Response &#62;&#62; Digital Input to Stim Events from the psychophysiology analysis software to label the stimulus events that correspond to the digital inputs for each trial type. 
	NOTE: For example, trial types include safe versus threat, low load versus high load, and maintenance period versus ITI period.</li>
	<li>Extract the blink magnitude around each stimulus event30.
	<ol>
		<li>Select Analysis &#62;&#62; Stim-Response &#62;&#62; Stim-Response Analysis and specify the Mean of Channel (i.e., the channel number corresponding to the processed EMG) from the psychophysiology analysis software to extract the mean baseline activity in a fixed window of -50 to 0 ms preceding the onset of the white noise.</li>
		<li>Select Analysis &#62;&#62; Stim-Response &#62;&#62; Stim-Response Analysis and specify the Max of Channel (i.e., the channel number corresponding to the processed EMG) from the psychophysiology analysis software to identify the blink onset and peak in a fixed window of 20 to 100 ms following the onset of the white noise.</li>
	</ol>
	</li>
	<li>Exclude trials with excessive noise on the EMG channel30. 
	NOTE: The acoustic startle responses must be reliably distinguished from excessive background EMG activity or other sources of contamination (e.g., movement artifacts or voluntary and spontaneous blinks immediately preceding auditory probes; see Figure 4D).</li>
	<li>Analyze the trial-by-trial blink responses using a standard spreadsheet software.
	<ol>
		<li>Normalize the blink magnitudes into z-scores (optional).</li>
		<li>Convert the z-scores to t-scores for further analysis (t = 10x + 50; optional).</li>
		<li>Average the t-scores and/or raw scores across trials for each trial type and calculate the APS (threat versus safe) for each condition (e.g., low load versus high load and maintenance period versus ITI period).</li>
		<li>At the group level, perform a 2 (safe versus threat) x 2 (maintenance period versus ITI) ANOVA across subjects to identify the effect of WM maintenance on APS.</li>
	</ol>
	</li>
</ol>
6. Analyze the Self-report Data
<ol>
	<li>Average the anxiety ratings across runs for the safe and threat conditions.</li>
	<li>At the group level, perform a threat versus safe t-test to determine the effectiveness of the threat manipulation.</li>
</ol>

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J., Krimsky, M., Grillon, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+impact+of+induced+anxiety+on+response+inhibition.">The impact of induced anxiety on response inhibition.</a> Front. Hum. Neurosci. 7, 69 (2013).</li><li>Owen, A. M., McMillan, K. M., Laird, A. R., Bullmore, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=N-back+working+memory+paradigm:+A+meta-analysis+of+normative+functional+neuroimaging+studies.">N-back working memory paradigm: A meta-analysis of normative functional neuroimaging studies.</a> Hum. Brain Mapp. 25 (1), 46-59 (2005).</li><li>Bailey, J. E., Argyropoulos, S. V., Kendrick, A. H., Nutt, D. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Behavioral+and+cardiovascular+effects+of+7.5%+CO2+in+human+volunteers.">Behavioral and cardiovascular effects of 7.5% CO2 in human volunteers.</a> Depress. Anxiety. 21 (1), 18-25 (2005).</li><li>Thibodeau, M. A., Welch, P. G., Katz, J., Asmundson, G. J. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Pain-related+anxiety+influences+pain+perception+differently+in+men+and+women:+A+quantitative+sensory+test+across+thermal+pain+modalities.">Pain-related anxiety influences pain perception differently in men and women: A quantitative sensory test across thermal pain modalities.</a> Pain. 154 (3), 419-426 (2013).</li><li>Lamm, C., Pine, D. S., Fox, N. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Impact+of+negative+affectively+charged+stimuli+and+response+style+on+cognitive-control-related+neural+activation:+An+ERP+study.">Impact of negative affectively charged stimuli and response style on cognitive-control-related neural activation: An ERP study.</a> Brain Cogn. 83 (2), 234-243 (2013).</li></ol>

The authors report no conflicts of interest.

This paper demonstrates how to administer the Sternberg WM task during threat of shock. Using this protocol, it was possible to show that WM maintenance is sufficient to reduce anxiety, as measured by the potentiation of the acoustic startle reflex3. These results suggest that the relationship between cognition and anxiety is bidirectional3-5 and that models of anxiety (e.g., the attentional control theory)1 must explain the effect of cognition on anxiety in addition to the effect of anxiety on cognition. Although the current protocol describes the integration of the Sternberg WM task and the threat of shock paradigm, it can also serve as a framework for studying the relationship between cognition and anxiety more generally21.
By redesigning existing cognitive tasks to take place during alternating periods of safety and threat, it is possible to study the effect of anxiety on specific cognitive processes, like WM and sustained attention2,31,32. For instance, in previous work, the N-back working memory task was integrated with the threat of shock paradigm, demonstrating that anxiety interferes with WM at a low load but not a high load4,5. These results suggest that anxiety interferes with WM, but also that healthy individuals are capable of overcoming anxiety when task demands are high. The Sustained Attention to Response Task (SART) was also integrated with the threat of shock paradigm; subjects had to inhibit their responses to infrequent target stimuli. This demonstrated that the threat of shock increases the accuracy on NoGo trials during the task31,32. Together with the N-back studies, these results suggest that anxiety can both impair and facilitate performance, and that the direction of the effect depends on the specific cognitive processes engaged by the task.
Likewise, by adding precisely timed startle probes to an existing cognitive task that has been adapted to the threat of shock paradigm, it is possible to study the effect of specific cognitive tasks on anxiety. The relationship between the WM load and anxiety was initially observed during the N-back WM tasks, where increasing the number of to-be-maintained items reduced APS4,5. However, because this task requires both maintenance and manipulation, it was difficult to determine which WM components were necessary for the observed reduction in anxiety23,33. By following up these studies with the simpler Sternberg WM paradigm, it was possible to show that central executive processing was not necessary for anxiety reduction3.
This technique can be used to study both the effect of anxiety on cognition, as well as the effect of cognition on anxiety. Accordingly, it is important to manipulate both anxiety and cognitive load in this paradigm and to take reliable measures of each. When applying this method to novel cognitive paradigms, it is important to ensure that the cognitive paradigm has distinguishable levels of difficulty based on performance. If pilot testing does not show differences in performance across experimental conditions, check for ceiling/floor effects and adjust the difficulty of the task accordingly. Likewise, it is important to design the threat of shock manipulation such that it is possible to observe APS during conditions of low cognitive load. If pilot testing does not show differences in startle during conditions of low cognitive load, try checking the signal-to-noise ratio in the EMG channel.
There are 3 critical steps to ensuring the effectiveness of this protocol. First, it is important to ensure that the subject understands the cognitive task being implemented. If necessary, design a practice version of the task to ensure that the subjects understand the instructions. Second, it is important to ensure that the electrical stimulation used is of a sufficient intensity to induce anxiety in the subject. If necessary, recalibrate the intensity of the electrical stimulation after each run. Third, it is important to ensure that the signal-to-noise ratio of the EMG channel is sufficient to recover the acoustic startle response. If the channel is noisy or the impedance is too high, thoroughly clean the skin under the eye and reapply the EMG electrodes.
Although there are a number of strengths to this paradigm, there are also limitations that should be addressed. For instance, the use of aversive electrical shock can raise concern among some IRBs, especially when dealing with vulnerable populations. It should be noted that there are alternative approaches to induce anxiety besides using electric shock. These include breathing elevated levels of CO2 (7.5%) for extended periods (8-20 min)34, using the threat of an aversive thermal stimulus35, presenting negatively valenced pictures36, etc. However, it should be noted that electrical stimuli are safe (when used properly), widely used, and effective. Although this protocol recommends a standardization approach to analyze potentiated startle, raw scores may be more reliable in some cases9,10. If standardized scores are used, it is recommended to examine the raw scores as well.
The strength of this protocol is that it allows the researcher to flexibly manipulate state anxiety within-subject in a single session and to test the relationship between anxiety and specific cognitive processes. There are three potential future applications of this protocol. First, it is important to understand how cognitive and emotional systems interact at the level of neural processes. Future studies should examine the relationship between anxiety and WM maintenance-related neural activity, using this paradigm while recording BOLD activity. Second, it is important to generalize these findings to other cognitive processes, such as sustained attention and reward processing. Future studies using this protocol should manipulate these processes during periods of threat and safety. Third, it is important to understand the relationship between cognition and anxiety, in both healthy individuals and in patient populations. Future studies using this protocol should include individuals from these special populations.
In conclusion, this work presents a protocol for studying the relationship between WM load and induced anxiety. Studies using this paradigm have shown that WM maintenance is sufficient to reduce anxiety but that anxiety does not interfere with the WM load itself. Although the findings presented here are specific to the Sternberg WM paradigm, this protocol can be adapted to study the bidirectional relationship between cognition and anxiety in general.

According to the Attention Control Theory (ACT), anxiety interferes with cognitive processing by competing for access to limited Working Memory (WM) resources1. However, the ACT does not address the inverse of this relationship (i.e.&#160;the effect of cognitive processing on anxiety). By manipulating anxiety during cognitive tasks using the threat of shock paradigm, it is possible to assess both the effect of anxiety on cognition and the effect of cognition on anxiety2,3,4,5. The purpose of this protocol is to demonstrate how to administer the Sternberg WM paradigm during threat of shock paradigm to probe the bi-directional relationship between anxiety and WM maintenance.
The threat of shock paradigm is widely used in the laboratory to manipulate state anxiety6,7,8,9,10,11 and can be implemented in healthy subjects2,3,4,5 and patients12,13,14,15 alike (see Bradford et al.16 for an example). The paradigm consists of alternating blocks of threat and safety17. Subjects are at risk of receiving unpredictable electrical stimulations during the threat blocks, but not during the safe blocks. Subjects&#39; anxiety can be probed periodically using the acoustic startle reflex18,19. Subjects typically show larger startle responses during the threat blocks compared to the safe blocks, and this Anxiety-Potentiated Startle (APS) can be used as a peripheral index of change in the ongoing anxiety during the test17,18. Potentiated startle in the threat of shock paradigm is recognized by the National Institute of Mental Health (NIMH) as a physiological index of anxiety in their Research Domain Criteria matrix20. However, it is also possible to probe an individual&#39;s anxiety using a self-report Likert-type scale. Because threat of shock is a passive paradigm, other cognitive tasks can be conducted simultaneously21. By combining the threat of shock with the Sternberg WM task, it is possible to probe anxiety during WM maintenance3.
During the Sternberg WM paradigm, subjects are required to encode a series of letters in WM and respond after a brief interval3,22. Unlike more complex WM tasks (e.g., the N-back task)4,5,23, the Sternberg task does not require the manipulation of information in the WM3,22. In addition, subjects encode, maintain, and respond to items during distinct intervals. Together, these features make it possible to dissociate WM maintenance from other, more complex cognitive processes24. By probing APS during the WM maintenance interval, it is possible to determine the effect of WM maintenance on anxiety. Likewise, by comparing WM accuracy and Reaction Time (RT) between the threat and safe blocks, it is possible to determine the effect of anxiety on WM maintenance. This protocol will detail the procedural steps necessary to conduct the Sternberg WM paradigm during threat of shock, as well as the analytical steps necessary to assess APS, accuracy, and reaction time during the task.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>Biopac System</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>System</td>
			<td>Biopac Systems Inc.</td>
			<td>MP150</td>
			<td>1, Psychophysiology monitoring hardware</td>
		</tr>
		<tr>
			<td>TTL integration</td>
			<td>Biopac Systems Inc.</td>
			<td>STP100C</td>
			<td>1</td>
		</tr>
		<tr>
			<td>EDA</td>
			<td>Biopac Systems Inc.</td>
			<td>EDA100C</td>
			<td>1</td>
		</tr>
		<tr>
			<td>ECG</td>
			<td>Biopac Systems Inc.</td>
			<td>ECG100C</td>
			<td>1</td>
		</tr>
		<tr>
			<td>EMG</td>
			<td>Biopac Systems Inc.</td>
			<td>EMG100C</td>
			<td>1</td>
		</tr>
		<tr>
			<td>Name</td>
			<td>Company</td>
			<td>Catalog Number</td>
			<td>Comments</td>
		</tr>
		<tr>
			<td>Other Equipment</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Breakout box</td>
			<td>See Alternatives</td>
			<td>Custom</td>
			<td>1</td>
		</tr>
		<tr>
			<td>Grass Signal Generator</td>
			<td>Grass Instruments</td>
			<td>SD9</td>
			<td>1</td>
		</tr>
		<tr>
			<td>Shock device</td>
			<td>Digitimer North America, LLC</td>
			<td>DS7A</td>
			<td>1</td>
		</tr>
		<tr>
			<td>Name</td>
			<td>Company</td>
			<td>Catalog Number</td>
			<td>Comments</td>
		</tr>
		<tr>
			<td>Alternatives</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Alternative to Breakout box</td>
			<td>Cortech Solutions</td>
			<td>SD-MS-TCPBNC</td>
			<td>1</td>
		</tr>
		<tr>
			<td>Alternative Grass Signal Generator</td>
			<td>Digitimer North America, LLC</td>
			<td>DG2A</td>
			<td>1</td>
		</tr>
		<tr>
			<td>Name</td>
			<td>Company</td>
			<td>Catalog Number</td>
			<td>Comments</td>
		</tr>
		<tr>
			<td>Audio Equipment</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Headphones</td>
			<td>Sennheiser Electronic GMBH &#38; CO</td>
			<td>HD-280</td>
			<td>1</td>
		</tr>
		<tr>
			<td>Headphone Amplifier</td>
			<td>Applied Research and Technology</td>
			<td>AMP4</td>
			<td>1</td>
		</tr>
		<tr>
			<td>Sound Pressure Level Meter</td>
			<td>Hisgadget Inc</td>
			<td>MS10</td>
			<td>1</td>
		</tr>
		<tr>
			<td>Name</td>
			<td>Company</td>
			<td>Catalog Number</td>
			<td>Comments</td>
		</tr>
		<tr>
			<td>Electrodes and Leads from Biopac</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>EMG</td>
			<td>Biopac Systems Inc.</td>
			<td>EL254S</td>
			<td>2</td>
		</tr>
		<tr>
			<td>EMG stickers</td>
			<td>Biopac Systems Inc.</td>
			<td>ADD204</td>
			<td>2</td>
		</tr>
		<tr>
			<td>Gel for EMG</td>
			<td>Biopac Systems Inc.</td>
			<td>GEL100</td>
			<td>1</td>
		</tr>
		<tr>
			<td>ECG</td>
			<td>Biopac Systems Inc.</td>
			<td>LEAD110</td>
			<td>2</td>
		</tr>
		<tr>
			<td>Shock</td>
			<td>Biopac Systems Inc.</td>
			<td>LEAD110</td>
			<td>2</td>
		</tr>
		<tr>
			<td>ECG</td>
			<td>Biopac Systems Inc.</td>
			<td>LEAD110S-W</td>
			<td>1</td>
		</tr>
		<tr>
			<td>ECG</td>
			<td>Biopac Systems Inc.</td>
			<td>LEAD110S-R</td>
			<td>1</td>
		</tr>
		<tr>
			<td>Disposable electrodes</td>
			<td>Biopac Systems Inc.</td>
			<td>EL508</td>
			<td>6</td>
		</tr>
		<tr>
			<td>Name</td>
			<td>Company</td>
			<td>Catalog Number</td>
			<td>Comments</td>
		</tr>
		<tr>
			<td>Software</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Presentation</td>
			<td>Neurobehavioral Systems</td>
			<td>Version 18</td>
			<td>Referred to here as experimental software</td>
		</tr>
		<tr>
			<td>Acknowledge</td>
			<td>Biopac Systems Inc.</td>
			<td>Version 4.2</td>
			<td>Referred to here as psychophysiology analysis software</td>
		</tr>
	</tbody>
</table>

reducing state anxiety using working memory maintenance

The purpose of this protocol is to explain how to examine the relationship between working memory processes and anxiety by combining the Sternberg Working Memory (WM) and the threat of shock paradigms. In the Sternberg WM paradigm, subjects are required to maintain a series of letters in the WM for a brief interval and respond by identifying whether the position of a given letter in the series matches a numerical prompt. In the threat of shock paradigm, subjects are exposed to alternating blocks where they are either at risk of receiving unpredictable presentations of a mild electric shock or are safe from the shock. Anxiety is probed throughout the safe and threat blocks using the acoustic startle reflex, which is potentiated under threat (Anxiety-Potentiated Startle (APS)). By conducting the Sternberg WM paradigm during the threat of shock and probing the startle response during either the WM maintenance interval or the intertrial interval, it is possible to determine the effect of WM maintenance on APS.

This protocol yields three primary data types: accuracy, RT, and APS. For accuracy and RT, this protocol involves two experimental manipulations, threat and load. For accuracy, typical results show a main effect of load but no main effect of threat and no load-by-threat interaction (trials (F(1,18) = 84.34; p &#60;0.01; see Figure 5). Subjects are typically more accurate on the low-load than the high-load trials. For RT, typical results show a main effect of both load (F(1,18) = 19.49; p &#60;0.01) and threat (F(1,18) = 8.03; p = 0.01) but no load-by-threat interaction (see Figure 6). Subjects typically show faster RTs during low-load trials than during high-load trials and faster RTs during threat blocks than during safe blocks.
This protocol also involves two experimental manipulations for APS: load and startle timing. Typical results show a load-by-timing interaction (F(1,18) = 16.63; p &#60;0.01; see Figure 7). Subjects typically show significantly larger APS during low load vs. high load trials, but only when the startle probe is delivered during the maintenance interval (MNT; maintenance period: t(18) = 3.92; p &#60;0.01; ITI: p &#62;0.05; d = 0.72). It should be noted that, because inferential statistics can vary from study to study, it is important to replicate these effects. After this experiment, a consistent decrease in APS as a function of task difficulty was found. This finding was observed in a verbal N-back task (3-back &#62; 0-back d(25) = 2.2)4, the Sternberg WM paradigm (see above, d(18) = 0.72; for replication, see Experiment 1 in Balderston et al. 20163; high load &#62; low load, d(18) = 0.44), and a complex picture recognition task (retrieval &#62; encoding, d(21) = 0.47)2. However, it should be noted that the final result may be driven in part by habituation.
Although it is difficult to determine an individual&#39;s subjective affective state during each trial, self-report data can be used to determine the effectiveness of the anxiety manipulation and as an individual difference measure. Therefore, it is important to assess the subject&#39;s affective state prior to the experiment using standardized questionnaires and to probe the subject&#39;s anxiety during the experiment. Typical results show significantly higher anxiety ratings during the threat blocks than during the safe blocks; t(18) = 8.85; p &#60;0.001.
<img alt="Figure 1" class="xfigimg" src="/files/ftp_upload/55727/55727fig1.jpg" /> 
Figure 1: Schematic of a Typical Equipment Setup. (A) Use separate computers to administer the task and record physiological signals from the subject. Synchronize events with the psychophysiology monitoring hardware and the shock device via the parallel port of the experiment computer. Relay the physiological signals from the psychophysiology monitoring hardware to the acquisition computer via the Ethernet cable. Deliver the shock to the subject using the shock device, which is controlled by a signal generator and triggered by the task computer. Deliver the white noise to the subject via the sound card of the task computer and record the trace using the psychophysiology monitoring hardware. (B) Required settings for the signal generator. (C) Required settings for the shock device. <a href="http://cloudflare.jove.com/files/ftp_upload/55727/55727fig1large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 2" class="xfigimg" src="/files/ftp_upload/55727/55727fig2.jpg" /> 
Figure 2: Schematic of a Typical Subject Setup. Attach electrodes to deliver the shock to the subject&#39;s non-dominant wrist. Attach electrodes to measure skin conductance on the subject&#39;s non-dominant palm. Attach electrodes to measure electromyography below the right eye, over the orbicularis oculi muscle. Attach electrodes to measure electrocardiography on the subject&#39;s left bicep and right clavicle. <a href="http://cloudflare.jove.com/files/ftp_upload/55727/55727fig2large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 3" class="xfigimg" src="/files/ftp_upload/55727/55727fig3.jpg" /> 
Figure 3: Schematic of Typical Experimental Design. Present the subjects with a series of letters followed by a brief maintenance period and a response prompt. During the response prompt, present the subjects with a letter (from the series) and a number. Instruct the subjects to indicate whether the number matches the position of the target letter in the previous series. Present startle probes during each trial, either during the maintenance period or the intertrial interval (ITI). <a href="http://cloudflare.jove.com/files/ftp_upload/55727/55727fig3large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 4" class="xfigimg" src="/files/ftp_upload/55727/55727fig4.jpg" /> 
Figure 4: Example EMG Traces Following a White Noise Probe. (A) Raw EMG trace. (B) EMG trace bandpass filtered at 30 to 500 Hz. (C) EMG trace that has been both filtered and rectified using a 20 ms constant. (D) Raw EMG trace from a trial contaminated by baseline noise. <a href="http://cloudflare.jove.com/files/ftp_upload/55727/55727fig4large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 5" class="xfigimg" src="/files/ftp_upload/55727/55727fig5.jpg" /> 
Figure 5: Typical Reaction Time (RT) Results. Subjects are typically faster during low-load trials than during high-load trials. Subjects are also typically faster under threat of shock. Bars represent the mean &#177; SEM. <a href="http://cloudflare.jove.com/files/ftp_upload/55727/55727fig5large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 6" class="xfigimg" src="/files/ftp_upload/55727/55727fig6.jpg" /> 
Figure 6: Typical Anxiety-potentiated Startle (APS) Results. When startle is probed during the maintenance period (MNT), subjects typically show larger startle potentiation on low-load compared to high-load trials. However, this effect does not hold when startle is probed during the ITI. Bars represent the mean &#177;SEM. <a href="http://cloudflare.jove.com/files/ftp_upload/55727/55727fig6large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 7" class="xfigimg" src="/files/ftp_upload/55727/55727fig7.jpg" /> 
Figure 7: Typical Accuracy (percent (%) correct) Results. Subjects are typically more accurate during low load trials than high load trials; however, performance does not tend to vary as a function of threat of shock. Bars represent the mean &#177; SEM. <a href="http://cloudflare.jove.com/files/ftp_upload/55727/55727fig7large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
Supplemental Code Files: Wav file for white noise presentation (40ms_wn.wav.) <a href="http://cloudflare.jove.com/files/ftp_upload/55727/40ms_wn.wav">Please click here to download this file.</a> Code necessary to setup hardware parameters for experimental software (Sternberg_threat_v5.exp.) <a href="http://cloudflare.jove.com/files/ftp_upload/55727/sternberg_threat_v5_run.exp">Please click here to download this file.</a> Code necessary to run experiment (Sternberg_threat_v5.sce.). <a href="http://cloudflare.jove.com/files/ftp_upload/55727/sternberg_threat_v5.sce">Please click here to download this file.</a>

Watch this Scientific Journal Video about Reducing State Anxiety Using Working Memory Maintenance at JoVE.com

Reducing State Anxiety Using Working Memory Maintenance

This protocol demonstrates how to measure anxiety-potentiated startle during the Sternberg Working Memory paradigm.

The purpose of this protocol is to explain how to examine the relationship between working memory processes and anxiety by combining the Sternberg Working ...

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7.4K Views.  National Institutes of Health (NIH). This protocol demonstrates how to measure anxiety-potentiated startle during the Sternberg Working Memory paradigm. 

Video: Reducing State Anxiety Using Working Memory Maintenance

The Successive Alleys Test of Anxiety in Mice and Rats

The plus-maze was derived from the early work of Montgomery. He observed that rats tended to avoid the open arms of a maze, preferring the enclosed ones. Handley, Mithani and File et al. performed the first studies on the plus-maze design we use today, and in 1987 Lister published a design for use with mice. 
Time spent on, and entries into, the open arms are an index of anxiety; the lower these indices, the more anxious the mouse is. Alternatively, a mouse that spends most of its time in the closed arms is classed as anxious. 
One of the problems of the plus-maze is that, while time spent on, and entries into, the open arms is a fairly unambiguous measure of anxiety, time in the central area is more difficult to interpret, although time spent here has been classified as &#x201C;decision making&#x201D;. In many tests central area time is a considerable part of the total test time.
Shepherd et al. produced an ingenious design to eliminate the central area, which they called the &#x201C;zero maze&#x201D;. However, although used by several groups, it has never been as widely adopted as the plus-maze. 
In the present article I describe a modification of the plus-maze design that not only eliminates the central area but also incorporates elements from other anxiety tests, such as the light-dark box and emergence tests. It is a linear series of four alleys, each having increasing anxiogenic properties. It has given similar results to the plus-maze in general. Although it may not be more sensitive than the plus-maze (more data is needed before a firm conclusion can be reached on this point), it provides a useful confirmation of plus-maze results which would be useful when, for example, only a single example of a mutant mouse was available, as, for example, in ENU-based mutagenesis programs.

The plus-maze measures anxiety-like behaviour in rodents. There are two opposite closed and two opposite open arms; anxious rodents avoid the open arms. The central area is neither completely open nor closed, so time spent here is ambiguous and difficult to interpret. Here a modification of the plus-maze protocol eliminating this area is described.

The plus-maze was derived from the early work of Montgomery. He observed that rats tended to avoid the open arms of a maze, preferring the enclosed ones.  ...

Use of an Eight-arm Radial Water Maze to Assess Working and Reference Memory Following Neonatal Brain Injury

Working and reference memory are commonly assessed using the land based radial arm maze.&#160;However, this paradigm requires pretraining, food deprivation, and may introduce scent cue confounds. The eight-arm radial water maze is designed to evaluate reference and working memory performance simultaneously by requiring subjects to use extra-maze cues to locate escape platforms and remedies the limitations observed in land based radial arm maze designs. Specifically, subjects are required to avoid the arms previously used for escape during each testing day (working memory) as well as avoid the fixed arms, which never contain escape platforms (reference memory). Re-entries into arms that have already been used for escape during a testing session (and thus the escape platform has been removed) and re-entries into reference memory arms are indicative of working memory deficits. Alternatively, first entries into reference memory arms are indicative of reference memory deficits. We used this maze to compare performance of rats with neonatal brain injury and sham controls following induction of hypoxia-ischemia and show significant deficits in both working and reference memory after eleven days of testing. This protocol could be easily modified to examine many other models of learning impairment.

The eight-arm radial water maze is designed to evaluate reference and working memory performance simultaneously by requiring subjects to use extra-maze cues to locate escape platforms and remedies the limitations observed in land based radial arm maze designs.

Working and reference memory are commonly assessed using the land based radial arm maze.&#160;However, this paradigm requires pretraining, food ...

Using the Threat Probability Task to Assess Anxiety and Fear During Uncertain and Certain Threat

Fear of certain threat and anxiety about uncertain threat are distinct emotions with unique behavioral, cognitive-attentional, and neuroanatomical components. Both anxiety and fear can be studied in the laboratory by measuring the potentiation of the startle reflex. The startle reflex is a defensive reflex that is potentiated when an organism is threatened and the need for defense is high. The startle reflex is assessed via electromyography (EMG) in the orbicularis oculi muscle elicited by brief, intense, bursts of acoustic white noise (i.e., &#8220;startle probes&#8221;). Startle potentiation is calculated as the increase in startle response magnitude during presentation of sets of visual threat cues that signal delivery of mild electric shock relative to sets of matched cues that signal the absence of shock (no-threat cues). In the Threat Probability Task, fear is measured via startle potentiation to high probability (100% cue-contingent shock; certain) threat cues whereas anxiety is measured via startle potentiation to low probability (20% cue-contingent shock; uncertain) threat cues. Measurement of startle potentiation during the Threat Probability Task provides an objective and easily implemented alternative to assessment of negative affect via self-report or other methods (e.g., neuroimaging) that may be inappropriate or impractical for some researchers. Startle potentiation has been studied rigorously in both animals (e.g., rodents, non-human primates) and humans which facilitates animal-to-human translational research. Startle potentiation during certain and uncertain threat provides an objective measure of negative affective and distinct emotional states (fear, anxiety) to use in research on psychopathology, substance use/abuse and broadly in affective science. As such, it has been used extensively by clinical scientists interested in psychopathology etiology and by affective scientists interested in individual differences in emotion.

Potentiation of the startle reflex is measured via electromyography of the orbicularis oculi muscle during low (uncertain) and high (certain) probability electric shock threat in the Threat Probability Task. This provides an objective measure of distinct negative emotional states (fear/anxiety) for research on psychopathology, substance use/abuse, and broad affective science.

Fear of certain threat and anxiety about uncertain threat are distinct emotions with unique behavioral, cognitive-attentional, and neuroanatomical ...

A Cognitive Paradigm to Investigate Interference in Working Memory by Distractions and Interruptions

Goal-directed behavior is often impaired by interference from the external environment, either in the form of distraction by irrelevant information that one attempts to ignore, or by interrupting information that demands attention as part of another (secondary) task goal. Both forms of external interference have been shown to detrimentally impact the ability to maintain information in working memory (WM). Emerging evidence suggests that these different types of external interference exert different effects on behavior and may be mediated by distinct neural mechanisms. Better characterizing the distinct neuro-behavioral impact of irrelevant distractions versus attended interruptions is essential for advancing an understanding of top-down attention, resolution of external interference, and how these abilities become degraded in healthy aging and in neuropsychiatric conditions. This manuscript describes a novel cognitive paradigm developed the Gazzaley lab that has now been modified into several distinct versions used to elucidate behavioral and neural correlates of interference, by to-be-ignored distractors versus to-be-attended interruptors. Details are provided on variants of this paradigm for investigating interference in visual and auditory modalities, at multiple levels of stimulus complexity, and with experimental timing optimized for electroencephalography (EEG) or functional magnetic resonance imaging (fMRI) studies. In addition, data from younger and older adult participants obtained using this paradigm is reviewed and discussed in the context of its relationship with the broader literatures on external interference and age-related neuro-behavioral changes in resolving interference in working memory.

A novel cognitive paradigm is developed to elucidate behavioral and neural correlates of interference by to-be-ignored distractors versus interference by to-be-attended interruptors during a working memory task. In this manuscript, several variants of this paradigm are detailed, and data obtained with this paradigm in younger/older adult participants is reviewed.

Goal-directed behavior is often impaired by interference from the external environment, either in the form of distraction by irrelevant information that ...

Morris Water Maze Test: Optimization for Mouse Strain and Testing Environment

The Morris water maze (MWM) is a commonly used task to assess hippocampal-dependent spatial learning and memory in transgenic mouse models of disease, including neurocognitive disorders such as Alzheimer&#8217;s disease. However, the background strain of the mouse model used can have a substantial effect on the observed behavioral phenotype, with some strains exhibiting superior learning ability relative to others. To ensure differences between transgene negative and transgene positive mice can be detected, identification of a training procedure sensitive to the background strain is essential. Failure to tailor the MWM protocol to the background strain of the mouse model may lead to under- or over- training, thereby masking group differences in probe trials. Here, a MWM protocol tailored for use with the F1 FVB/N x 129S6 background is described. This is a frequently used background strain to study the age-dependent effects of mutant P301L tau (rTg(TauP301L)4510 mice) on the memory deficits associated with Alzheimer&#8217;s disease. Also described is a strategy to re-optimize, as dictated by the particular testing environment utilized.

This manuscript describes a Morris water maze (MWM) protocol tailored for use with a commonly used mouse model of Alzheimer's disease. The MWM is widely used in transgenic mouse models. Implementation of a procedure sensitive to the background strain of the mouse model is essential for detecting group differences.

 The Morris water maze (MWM) is a commonly used task to assess hippocampal-dependent spatial learning and memory in transgenic mouse models of disease, ...

Errors as a Means of Reducing Impulsive Food Choice

Nowadays, the increasing incidence of eating disorders due to poor self-control has given rise to increased obesity and other chronic weight problems, and ultimately, to reduced life expectancy. The capacity to refrain from automatic responses is usually high in situations in which making errors is highly likely. The protocol described here aims at reducing imprudent preference in women during hypothetical intertemporal choices about appetitive food by associating it with errors. First, participants undergo an error task where two different edible stimuli are associated with two different error likelihoods (high and low). Second, they make intertemporal choices about the two edible stimuli, separately. As a result, this method decreases the discount rate for future amounts of the edible reward that cued higher error likelihood, selectively. This effect is under the influence of the self-reported hunger level. The present protocol demonstrates that errors, well known as motivationally salient events, can induce the recruitment of cognitive control, thus being ultimately useful in reducing impatient choices for edible commodities.

Giving in to temptation of tasty food may result in long-term overweight problems. This protocol describes how to reduce imprudent preference for edible commodities during hypothetical intertemporal choices in women by associating them with errors.

Nowadays, the increasing incidence of eating disorders due to poor self-control has given rise to increased obesity and other chronic weight problems, and ...

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 ...

Assessing Working Memory in Children: The Comprehensive Assessment Battery for Children &#8211; Working Memory (CABC-WM)

The Comprehensive Assessment Battery for Children - Working Memory (CABC-WM) is a computer-based battery designed to assess different components of working memory in young school-age children. Working memory deficits have been identified in children with language-based learning disabilities, including dyslexia1,2 and language impairment3,4, but it is not clear whether these children exhibit deficits in subcomponents of working memory, such as visuospatial or phonological working memory. The CABC-WM is administered on a desktop computer with a touchscreen interface and was specifically developed to be engaging and motivating for children. Although the long-term goal of the CABC-WM is to provide individualized working memory profiles in children, the present study focuses on the initial success and utility of the CABC-WM for measuring central executive, visuospatial, phonological loop, and binding constructs in children with typical development. Immediate next steps are to administer the CABC-WM to children with specific language impairment, dyslexia, and comorbid specific language impairment and dyslexia.

Working memory predicts a significant amount of variance for a variety of cognitive tasks, including speaking, reading, and writing. However, few tools are available to assess working memory in children. We present an innovative, computer-based battery that comprehensively assesses different components of working memory in school-age children.

The Comprehensive Assessment Battery for Children - Working Memory (CABC-WM) is a computer-based battery designed to assess different components of ...

A Real-world What-Where-When Memory Test

Episodic memory is a complex memory system which allows recall and mental re-experience of previous episodes from one&#39;s own life. Real-life episodic memories are about events in their spatiotemporal context and are typically visuospatial, rather than verbal. Yet often, tests of episodic memory use verbal material to be recalled (word lists, stories). The Real-World What-Where-When memory test requires participants to hide a total of 16 different objects in 16 different locations over two temporal occasions, 2 h apart. Another two hours later, they are then asked to recall which objects (What) they had hidden in which locations (Where) and on which of the two occasions (When). In addition to counting the number of correctly recalled complete what-where-when combinations, this task can also be used to test real-world spatial memory and object memory. This task is sensitive to normal cognitive aging, and correlates well with performance on other episodic memory tasks, while at the same time providing more ecological validity and being cheap and easy to run.

A Real-world What-Where-When Memory Test

The Real-World What-Where-When memory test is a novel episodic memory test, in which participants need to recall which objects have been hidden in which locations on which of two distinct occasions. It is easy to run and is sensitive to normal cognitive aging.

Episodic memory is a complex memory system which allows recall and mental re-experience of previous episodes from one&#39;s own life. Real-life episodic ...

Utilizing the Modified T-Maze to Assess Functional Memory Outcomes After Cardiac Arrest

Background: Evaluating mild to moderate cognitive impairment in a global cerebral ischemia (i.e. cardiac arrest) model can be difficult due to poor locomotion after surgery. For example, rats who undergo surgical procedures and are subjected to the Morris water maze may not be able to swim, thus voiding the experiment.
New Method: We established a modified behavioral spontaneous alternation T-maze test. The major advantage of the modified T-maze protocol is its relatively simple design that is powerful enough to assess functional learning/memory after ischemia. Additionally, the data analysis is simple and straightforward. We used the T-maze to determine the rats&#39; learning/memory deficits both in the presence or absence of mild to moderate (6 min) asphyxial cardiac arrest (ACA). Rats have a natural tendency for exploration and will explore the alternate arms in the T-maze, whereas hippocampal-lesioned rats tend to adopt a side-preference resulting in decreased spontaneous alternation ratios, revealing the hippocampal-related functional learning/memory in the presence or absence of ACA.
Results: ACA groups have higher side-preference ratios and lower alternations as compared to control.
Comparison with Existing Method(s): The Morris water and Barnes maze are more prominent for assessing learning/memory function. However, the Morris water maze is more stressful than other mazes. The Barnes maze is widely used to measure reference (long-term) memory, while ACA-induced neurocognitive deficits are more closely related to working (short-term) memory.
Conclusions: We have developed a simple, yet effective strategy to delineate working (short-term) memory via the T-maze in our global cerebral ischemia model (ACA).

This protocol describes the use of a modified T-maze to evaluate functional learning/memory in asphyxia cardiac arrest-induced cerebral ischemia.

Background: Evaluating mild to moderate cognitive impairment in a global cerebral ischemia (i.e. cardiac arrest) model can be difficult due to poor ...