We would like to thank Joy Bergmann, Attilio Ceretti, Sarah Cohen, Nina Duncan, Arron Hall, Hanna Lefebo, and Madeleine Salvatore for their assistance with the behavior, and Dr. Vinay Parikh, Dr. Brittney Yegla, and Rob Cole for their advice on the SAT parameters. We would like to thank Dr. Jill McGaughy for tips on the training procedure. This work was supported by the National Institutes of Health grant NIMH 092438 and National Science Foundation grant IOS 1552416 to D.A.B.

All experiments were conducted in accordance with the National Institutes of Health guidelines and were approved by the Temple University Institutional Animal Use and Care Committee. This protocol was developed with adult (~60 days of age when starting the study) male and female Sprague-Dawley rats (Charles River).
1. Materials for Touchscreen SAT
<ol>
	<li>Ensure that a sound attenuation cubicle surrounds each touchscreen chamber and that chambers are equipped with a house light, tone generator, camera positioned above the chamber, and a pellet dispenser with feeder reward area (that can be illuminated) on the wall opposite the touchscreen.</li>
	<li>Use a feeder reward area with a large aperture if cannula or electrodes implants are planned. Use 45 mg reward pellets that have been sifted with a flour sifter to reduce dust, which can clog the feeder.</li>
	<li>Fit each chamber with a black acrylic mask (3.175 mm thick, matte) to limit response areas on the touchscreen. Prepare the mask with a central circle opening (28.58 mm diameter, 107.95 mm from the bottom, centered between the left and right side) for the light stimulus and two square response areas (28.57 mm x 28.57 mm) each positioned below and off center (one 85.73 mm from the left side and one 85.73 mm from the right side, and both 34.93 mm from the bottom). A depiction of the mask can be found in the publication, Wicks, et al.19.</li>
	<li>Place the mask in front of the touchscreen with matte finish positioned towards the chamber.</li>
</ol>
2. Schedule Design for Touchscreen SAT
NOTE: The below protocol details how to set the parameters for the shaping and testing schedules for touchscreen SAT. All these schedules are also available upon request. Table 1 depicts the schedule parameters.
<ol>
	<li>Nose Poke Shaping parameters.
	<ol>
		<li>Set program so that no signals are presented and the houselight remains off.</li>
		<li>Set the feeder reward area to illuminate to indicate the presence of a pellet and for this light to turn off following pellet retrieval as determined by the feeder port infrared beam.</li>
		<li>Set program so that nose-pokes in either response area are rewarded with food pellets under a fixed-ratio 1 (FR1) reinforcement schedule (i.e., each appropriate response is rewarded). However, set the program so that if the side differential for nose-pokes is five or greater, the rat must poke the non-favored side to receive a reward to prevent the development of a side bias.</li>
		<li>Set the session to end after 40 min or 120 rewarded pokes. Advance subjects to Training Schedule 1 if they complete 120 nose-pokes in 40 min for two consecutive days.</li>
	</ol>
	</li>
	<li>Description of shared parameters for Training Schedule 1, Training Schedule 2, and SAT.
	<ol>
		<li>Program training schedules so that the houselight remains on to increase attentional demands.</li>
		<li>Program each session so that it consists of 162 trials that can be divided into three blocks of 54 trials per block with the intertrial interval (ITI) set at 9 &#177; 3 s.</li>
		<li>Ensure that signal and non-signal trials are pseudorandomized such that the same number of signal and non-signal trials occur in each of the three blocks.</li>
		<li>Set the program to deliver a food pellet and turn on the food port light automatically at the start of each session. When an infrared beam break indicates that the rat has removed its head from the reward area, set the program to turn off the food port light and begin the first ITI.</li>
		<li>Following each trial (i.e., signal or non-signal), set the program to indicate the 4-s response window with a 200 ms, 70 dB tone (3 kHz).</li>
		<li>If the rat correctly responds, set the program to initiate the ITI when the rat removes its head from the reward area. If the rat incorrectly responds, set the ITI to begin following the response. If the rat omits (i.e., makes no response), set the ITI to begin after the termination of the response window.</li>
		<li>For counterbalancing, develop two versions of these schedules, the first with the right response area designated for hits (Version A) and the second with the left response area designated for hits (Version B).</li>
	</ol>
	</li>
	<li>Training Schedule 1 parameters.
	<ol>
		<li>Set all signal durations to 500 ms.</li>
		<li>On signal trials, set the program to simultaneously illuminate the response area designated for hits at 50% intensity, along with the signal, to draw the rat&#39;s attention to the correct response area.</li>
		<li>Keep the designated area for correct rejections unlit on non-signal trials.</li>
		<li>Set the program so that incorrect responses trigger correction trials in which the previous trial type (i.e., signal or non-signal) is presented again. Ensure that a correct response or four consecutive incorrect responses ends the series of correction trials. Do not count correction trials towards the 162 total trials required for task completion.</li>
		<li>Advance subjects to Training Schedule 2 if they have at least 70% hits, at least 70% correct rejections, and less than 20% omissions for three consecutive days.</li>
	</ol>
	</li>
	<li>Training Schedule 2 parameters.
	<ol>
		<li>Set all signal durations to 500 ms.</li>
		<li>Unlike Training Schedule 1, set the program to keep the response area for hits unlit on signal trials.</li>
		<li>Unlike Training Schedule 1, do not have the program execute correction trials.</li>
		<li>Keep all other parameters the same as in Training Schedule 1.</li>
		<li>Advance subjects to SAT if they have at least 70% hits, at least 70% correct rejections, and less than 20% omissions for three consecutive days.</li>
	</ol>
	</li>
	<li>SAT parameters.
	<ol>
		<li>Set three signal durations: 500 ms, 50 ms, and 25 ms.</li>
		<li>Set the program to present the variable signals in a pseudorandom fashion, such that the three signal durations are presented an equal number of times during each one of the three trial blocks.</li>
		<li>Define criteria performance as at least 70% hits at the 500 ms signal duration, at least 70% correct rejections, and less than 20% omissions for three consecutive days.</li>
	</ol>
	</li>
</ol>
<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>Training Stage</td>
			<td>Max. Trials</td>
			<td>Signal Duration (ms)</td>
			<td>Special Conditions</td>
			<td>Criteria</td>
			<td>Median Days per Stage</td>
		</tr>
		<tr>
			<td>Nose Poke Shape</td>
			<td>120</td>
			<td>No events</td>
			<td>Houselight on</td>
			<td>120 trials in 40 minutes</td>
			<td>Males: 2, Females: 2</td>
		</tr>
		<tr>
			<td>Training Schedule 1</td>
			<td>162</td>
			<td>500</td>
			<td>Houselight on; Correction trials; Response window lit at 50% during signal trials</td>
			<td>&#62; 70% Hits, &#62; 70% CR, &#60; 20% Omissions for 3 consecutive days</td>
			<td>Males: 19, Females: 25</td>
		</tr>
		<tr>
			<td>Training Schedule 2</td>
			<td>162</td>
			<td>500</td>
			<td>Houselight on</td>
			<td>&#62; 70% Hits, &#62; 70% CR, &#60; 20% Omissions for 3 consecutive days</td>
			<td>Males: 13, Females: 21</td>
		</tr>
		<tr>
			<td>SAT</td>
			<td>162</td>
			<td>500, 50, 25</td>
			<td>Houselight on</td>
			<td>&#62; 70% Hits at 500 ms, &#62; 70% CR, &#60; 20% Omissions for 3 consecutive days</td>
			<td>Males: 18, Females: 12</td>
		</tr>
	</tbody>
</table>
Table 1: Training Schedules and Average Acquisition Time for Each Schedule.
<ol start="6">
	<li>Distractor SAT.
	<ol>
		<li>Keep parameters for Blocks 1 and 3 identical to SAT.</li>
		<li>Program the houselight so that it flashes at a frequency of 0.5 Hz during Block 2.</li>
		<li>When analyzing data, compare performance between Blocks 1, 2, and 3 to analyze performance before, during, and after the distractor within the same session.</li>
	</ol>
	</li>
	<li>Data extraction program.
	<ol>
		<li>Extract the following data for SAT and distractor SAT broken down by trial block and each signal duration (for VI and percent hits):
		<ol>
			<li>Calculate the percentage of hits by dividing the number of hits by the combined number of hits and misses and then multiplying by 100.</li>
			<li>Calculate the percentage of correct rejections by dividing the number of correct rejections by the combined number of correct rejections and false alarms and then multiplying by 100.</li>
			<li>Calculate the vigilance index (VI), an overall measure of attentional performance, using the following formula: <img alt="Equation" src="/files/ftp_upload/56219/56219eq1.jpg" /> where h and f are the proportion of hits on signal trials and false alarms on non-signal trials, respectively. The values of VI range from -1 to +1 with a value of 0 indicating that the rat is unable to distinguish between signal and non-signal trials. Do not include omitted trials in the VI measure.</li>
			<li>Calculate the percentage of omissions by determining the percentage of trials where no response occurred.</li>
		</ol>
		</li>
	</ol>
	</li>
</ol>
3. Procedure for Running Rats in Touchscreen SAT
<ol>
	<li>Reverse light/dark cycle considerations 
	NOTE: We test our rats during their dark cycle, but see the discussion section for considerations regarding testing in the light cycle.
	<ol>
		<li>For testing during the dark cycle, begin behavior testing at least 30 min after the start of the dark cycle.</li>
		<li>Once the equipment is set up, keep the behavioral testing room in darkness during all training and testing procedures to avoid interfering with the rats&#39; sleep/wake cycle.</li>
		<li>Cover rat cages with black towels during transport to minimize light exposure.</li>
		<li>Use headlights with red lights turned on for hands-free work in the dark without disrupting the light/dark cycle.</li>
	</ol>
	</li>
	<li>Food restriction procedures and schedule assignment
	<ol>
		<li>Don appropriate personal protective equipment (PPE; e.g., gloves, mask, bonnet, lab coat, shoe covers, etc.) prior to handling the rats.</li>
		<li>When rats arrive to the colony room, house rats individually or in groups. Most rats in our studies are eventually implanted with cannulae, which necessitates individual housing.</li>
		<li>Handle rats daily for 5 min per day, 5 days prior to the start of the study.</li>
		<li>Prior to food restriction, record each rat&#39;s free feeding weight and calculate their target weight at 85% of their free feeding weight.</li>
		<li>Food restrict rats for 3 days before beginning shaping.</li>
		<li>Prior to shaping, pre-expose rats to 15 food reward pellets in their home cage.</li>
		<li>After the initial food restriction, adjust daily food amounts as needed to maintain performance on the attention task and ~85% of each rat&#39;s free-feeding body weight allowing for slight weight gain over time (see guidelines25). Prior to starting a new schedule, restrict rats more than usual to increase motivation.</li>
		<li>On the day(s) of a week when rats do not perform behavior, increase their food by at least 1.0 g to make-up for the calories that they would have received while being rewarded during the behavioral task.</li>
		<li>Record each rat&#39;s weight after their session is completed.</li>
		<li>Do not feed rats until at least 30 min after completing their session to avoid an association between behavioral testing cessation and feeding time. If rats are group housed, feed each rat separately for 1 h, then put rats back into the group housing condition.</li>
		<li>Prior to the start of the study, randomly assign each rat to one of the two versions of the training schedules (A or B) and maintain them on their designated version throughout the training and SAT procedures.</li>
	</ol>
	</li>
	<li>Daily running procedure
	<ol>
		<li>Don appropriate PPE.</li>
		<li>Power up operant chambers and the computer.</li>
		<li>Check that the pellet dispensers are filled with pellets and that the mask is inserted in front of the touchscreen.</li>
		<li>Ensure that each box is functioning properly by verifying that the feeder is properly dispensing food and the screen, lights, and tone are working. A simple box test schedule can be written for this purpose and ours is available upon request.</li>
		<li>Load the correct schedule in each box, paying close attention to the appropriate version (i.e., A or B). Input relevant information regarding animal ID, sex, experimental condition, user initials, and other notes if needed (e.g., infusion day 1).</li>
		<li>Retrieve rats from colony room and put them in the appropriate box.</li>
		<li>Start the session.</li>
		<li>Watch performance via the video camera set-up at the start of the session for several minutes to ensure that everything is functioning properly. Check performance periodically.</li>
		<li>Once a session is complete, record summary data (e.g., percent hits, percent CRs, percent omissions) and any issues in the notebook.</li>
		<li>Wipe each chamber with 10% ethanol and replace the bedding in between sets of animals.</li>
		<li>After running the final set of rats for the day, clean the chambers thoroughly with 10% ethanol, ensuring that the floor grid is as clean as possible, and replace the bedding.</li>
		<li>Back up data on a cloud server and extract any critical data.</li>
		<li>Power down equipment.</li>
		<li>Run animals at the same time in the same boxes each day.</li>
		<li>Run rats 5 - 6 days a week (we run our rats 6 days a week).</li>
	</ol>
	</li>
</ol>
4. Troubleshooting the Behavior
NOTE: It is normal for performance to drop when rats are transitioned to the next shaping phase (i.e., Nose Poke Shape to Training Schedule 1, Training Schedule 1 to Training Schedule 2, Training Schedule 2 to SAT). However, if performance does not improve over time or if a rat was performing well and then suddenly stops performing well, most often the issue is with food restriction.
<ol>
	<li>Troubleshooting food restriction issues
	<ol>
		<li>If food restriction is a suspected issue, watch the rat perform the task to help diagnose where the problem lies and check the omission numbers, particularly across the session.</li>
		<li>If a rat has high omissions, especially if they increase in Block 3, and sits calmly in the box not performing, reduce daily food amount by 1 g and monitor the situation to determine if performance is improved.</li>
		<li>If a rat has high omissions, impatiently pokes the screen, and runs back and forth to the food port throughout the session, increase daily food amount by 1 g and monitor the situation to determine if performance is improved.</li>
		<li>In cases where it is difficult to tell whether a rat is over- or underfed, increase food amount for 2 days and monitor performance. If performance does not improve or gets worse, decrease food amount for 2 days and monitor performance</li>
		<li>If performance does not improve after adjusting food restriction, move the rat to an easier scheduled as detailed next.</li>
	</ol>
	</li>
	<li>Troubleshooting performance at different schedules of training 
	NOTE: Drop a poorly performing rat if its acquisition time exceeds two standard deviations above the mean number of days that it takes an animal of its same sex to reach criteria on that schedule. We have made suggestions based on our data for cut-offs for Sprague-Dawley rats, but these numbers may need to be adjusted for different strains.
	<ol>
		<li>Training Schedule 1: For Training Schedule 1, drop Sprague-Dawley males if they cannot acquire after 36 days and drop females if they have not acquired after 48 days.</li>
		<li>Training Schedule 2: If a rat&#39;s performance on Training Schedule 2 stagnates below criteria for 12 days in a row and a food restriction issue has been ruled out, move the rat back down to Training Schedule 1 for a minimum of 5 days or until it reaches criteria. If the rat does not reach criteria on training Schedule 1 after 12 days, drop the rat from the study. If the rat passes Training Schedule 1 and is moved back to Training Schedule 2, but does not improve after 36 and 42 days for males and females, respectively, after initially starting Training Schedule 2, then drop the rat from the study.</li>
		<li>SAT: If a rat&#39;s performance on SAT stagnates below criteria for 12 days in a row and a food restriction issue has been ruled out, move the rat back down to Training Schedule 1 for a minimum of 5 days or until they reach criteria. If the rat does not reach criteria on Schedule 1 in 12 days, drop the rat from the study. If the rat passes Training Schedule 1 and is moved back to SAT, but does not improve after 41 and 28 days for males and females, respectively, after initially starting SAT, drop the rat from the study.</li>
	</ol>
	</li>
	<li>Troubleshooting other issues
	<ol>
		<li>If a rat develops a side bias (i.e., consistently scoring &#62;70% hits and &#60;40% on correct rejections, or vice versa) while on Training Schedule 2 or SAT, move the rat to Training Phase 1, which has correction trials that encourage the rat to respond to both response areas. Keep the rat at Training Schedule 1 for a minimum of 5 days or until they reach criteria. Drop the rat from the study if criteria on training Schedule 1 is not met after 12 days.</li>
		<li>Consult with the veterinary staff immediately if a rat stops eating, develops porphyrin, or otherwise is showing signs of pain and distress.</li>
	</ol>
	</li>
</ol>

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C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+touchscreen+operant+platform+for+assessing+executive+function+in+rats+and+mice.">The touchscreen operant platform for assessing executive function in rats and mice.</a> Nat Protoc. 8 (10), 1985-2005 (2013).</li><li>Gill, T. M., Sarter, M., Givens, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sustained+visual+attention+performance-associated+prefrontal+neuronal+activity:+evidence+for+cholinergic+modulation.">Sustained visual attention performance-associated prefrontal neuronal activity: evidence for cholinergic modulation.</a> J Neurosci. 20 (12), 4745-4757 (2000).</li><li>Mazure, C. M., Jones, D. 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J., Lustig, C., Sarter, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=CNTRICS+Final+Task+Selection:+Control+of+Attention.">CNTRICS Final Task Selection: Control of Attention.</a> Schizo Bull. 35 (1), 182-196 (2009).</li><li>Burk, J. A., Herzog, C. D., Porter, M. C., Sarter, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Interactions+between+aging+and+cortical+cholinergic+deafferentation+on+attention.">Interactions between aging and cortical cholinergic deafferentation on attention.</a> Neurobiol Aging. 23 (3), 467-477 (2002).</li><li>St Peters, M., Demeter, E., Lustig, C., Bruno, J. P., Sarter, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Enhanced+control+of+attention+by+stimulating+mesolimbic-corticopetal+cholinergic+circuitry.">Enhanced control of attention by stimulating mesolimbic-corticopetal cholinergic circuitry.</a> J Neurosci. 31 (26), 9760-9771 (2011).</li><li>Turchi, J., Holley, L. A., Sarter, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+of+nicotinic+acetylcholine+receptor+ligands+on+behavioral+vigilance+in+rats.">Effects of nicotinic acetylcholine receptor ligands on behavioral vigilance in rats.</a> Psychopharmacology. 118 (2), 195-205 (1995).</li><li>Newman, L. 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A., McGaughy, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cholinergic+deafferentation+of+prefrontal+cortex+increases+sensitivity+to+cross-modal+distractors+during+a+sustained+attention+task.">Cholinergic deafferentation of prefrontal cortex increases sensitivity to cross-modal distractors during a sustained attention task.</a> J Neurosci. 28 (10), 2642-2650 (2008).</li><li>Yu, A., Dayan, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Uncertainty,+Neuromodulation,+and+Attention.">Uncertainty, Neuromodulation, and Attention.</a> Neuron. 46 (4), 681-692 (2005).</li><li>Bari, A., Dalley, J. W., Robbins, T. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+application+of+the+5-choice+serial+reaction+time+task+for+the+assessment+of+visual+attentional+processes+and+impulse+control+in+rats.">The application of the 5-choice serial reaction time task for the assessment of visual attentional processes and impulse control in rats.</a> Nat Protoc. 3 (5), 759-767 (2008).</li><li>Bhandari, J., Daya, R., Mishra, R. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Improvements+and+important+considerations+for+the+5-choice+serial+reaction+time+task-An+effective+measurement+of+visual+attention+in+rats.">Improvements and important considerations for the 5-choice serial reaction time task-An effective measurement of visual attention in rats.</a> J Neurosci Meth. 270, 17-29 (2016).</li><li>Bangasser, D. A., Shors, T. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+hippocampus+is+necessary+for+enhancements+and+impairments+of+learning+following+stress.">The hippocampus is necessary for enhancements and impairments of learning following stress.</a> Nat Neurosci. 10 (11), 1401-1403 (2007).</li><li>Wiersielis, K. 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Q. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Variation+in+visual+acuity+within+pigmented,+and+between+pigmented+and+albino+rat+strains.">Variation in visual acuity within pigmented, and between pigmented and albino rat strains.</a> Behav Brain Res. 136 (2), 339-348 (2002).</li><li>Peters, M., Cherian, A. K., Bradshaw, M., Sarter, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sustained+attention+in+mice:+Expanding+the+translational+utility+of+the+SAT+by+incorporating+the+Michigan+Controlled+Access+Response+Port+(MICARP).">Sustained attention in mice: Expanding the translational utility of the SAT by incorporating the Michigan Controlled Access Response Port (MICARP).</a> Behav Brain Research. 225 (2), 574-583 (2011).</li><li>Romberg, C., Mattson, M. P., Mughal, M. R., Bussey, T. J., Saksida, L. 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The authors have nothing to disclose.

Deficits in sustained attention are reported in neuropsychiatric, neurodevelopmental, and neurodegenerative disorders1,2,3, but the processes that contribute to optimal versus disrupted attention are understudied. This is, in part, because specialized equipment limits the easy adaptation of rodent attention tasks. However, touchscreen chambers are addressing this issue by providing the flexibility to run a variety of cognitive t...

Disorders ranging from attention deficit hyperactivity disorder (ADHD) to schizophrenia to Alzheimer&#39;s disease, share attentional impairments as a feature1,2,3. Deficits in sustained attention&#8212;the ability to monitor continuously a situation for intermittent and unpredictable events&#8212;are particularly disruptive, because sustained attention is crucial for selective and divided attention, as well as other cognitive processes4,5. Even in healthy people, difficulty with sustaining attention negatively affects cognition, impairing daily function6. Thus, understanding the neurobiological basis of sustained attention and how it becomes dysregulated could lead to interventions to improve cognition that would benefit many people.
In order to delineate circuits and molecular processes that contribute to proper versus disrupted attention, many researchers have turned to non-human animal models, where the manipulation of specific cell populations and molecular processes during attention tasks is possible. The effort has led to the development of a variety of operant attention tasks that can assess the ability to sustain attention7,8,9,10. One such paradigm, developed by McGaughy and Sarter (1995), is known as the sustained attention task (SAT). Rodents trained in SAT must distinguish signal trials, in which a signal light briefly flashes, from non-signal trials. Attentional demands can be increased by introducing a visual distractor (flashing houselight)10. The parameters critical for assessing attention in SAT are well documented and this task has been validated in male and female rats and mice10,11,12. A version has even been adapted for humans, highlighting the translational utility of SAT13. Importantly, the use of this task helped implicate the basal forebrain corticopetal system in sustained attention4. Specifically, cholinergic neurons in the nucleus basalis of Meynert (NBM)/ substantia inominata (SI) region of the basal forebrain that project to the prefrontal cortex are critical for hits, which are accurate responses on signal trials14,15. In contrast, GABAergic neurons in this region are thought to mediate performance during correct rejections (CRs), which are accurate responses on non-signal trials16. Once the basic circuit for this task was established, factors that modulate this circuit to impair attention ranging from stress hormones to neurotoxic proteins have been identified17,18. Collectively, these studies highlight the utility of SAT.
One limitation to implementing SAT in the laboratory is that the original procedure requires operant chambers permanently configured for the task, with a center panel light and two extendable levers, one designated for hits and one for CRs10. On a signal trial, the panel light briefly illuminates and then levers extend to indicate a 4 s response window. During that window, if the designated hit lever is pressed, then the rat receives a reward (either food or water). Incorrect lever presses on signal trials, known as misses, are not rewarded. On non-signaled trials, the panel light remains off, then levers are extended. While a CR is rewarded, incorrect lever presses on non-signaled trials, known as false alarms, are not rewarded. A failure to make any response during the response window counts as an omission. To expand the availability of SAT, we recently modified the task for touchscreen operant chambers, in which one wall is a touchscreen that can both display visual stimuli and record nose-poke responses19. Touchscreen chambers are becoming popular because of their versatility to run a variety of tasks (e.g., paired associate learning, reversal learning, etc.) with one piece of equipment20. Similar to other touchscreen procedures20,21,22, our adaptation of SAT requires the use of an opaque plastic touchscreen cover, called a mask, with holes cut into it. A central, circular hole allows for the presentation of a signal by presenting a white stimulus on the screen behind it, and two square response areas, one designated for hits and one for CRs, allow the rat to make touch responses19. In the traditional chambers, the 4-s response window is typically indicated with the presentation of the levers, but a previous study demonstrated that a tone can also be used to signal the response window23. In touchscreen SAT, we similarly signal the response window with a brief tone. Other training parameters (e.g., number of trials, intertrial interval, etc.) are kept similar between traditional and touchscreen SAT. We previously assessed performance between the traditional and touchscreen SAT versions and found that it was comparable, suggesting that touchscreen SAT is a valid way to measure sustained attention19. The touchscreen modification makes SAT more versatile because it can now be adapted for widespread use in touchscreen operant boxes.
The present protocol details how to run touchscreen SAT in the laboratory. The training schedules have been slightly altered from our previous report of the task19 to optimize acquisition time in female rats, which, unlike males, had difficulty advancing through the previous schedules24. Results illustrating typical performance in male and female rats are included, along with tips for troubleshooting issues with the task.

<table><tbody><tr><td>Bussey-Saksida Rat Touch Screen Chambers Easy-Install System</td><td>Lafayette Instrument</td><td>80604-20</td><td>Includes: Touch Screen, Chamber base with perforated floors, Large Feeder Reward Area (44.5 mm wide x 92.5 mm high), Speaker, Houselight, Tone Generator, SAC; Trapezoidal animal working area is 126 mm wide at the Feeder, 240 mm wide and 332 mm deep at the Screen, and 300 mm in height</td></tr><tr><td>Sound Attenuation Cubicles (SAC)</td><td>Lafayette Instrument</td><td>80604-20</td><td>Attenuation to approximately 35 dB; 38 kg in weight; External dimensions: 600 mm wide x 670 mm tall x 352 mm deep; Internal dimensions: 540 mm wide x 610 mm tall x 532 mm deep</td></tr><tr><td>Abet II Software for Touch Screens</td><td>Lafayette Instrument</td><td>89505</td><td>Version 2.19</td></tr><tr><td>Whisker Multimedia</td><td>Lafayette Instrument</td><td>80698-1</td><td>Required for Abet II Touch Operation</td></tr><tr><td>Pellet Dispenser</td><td>Lafayette Instrument</td><td>80209-45</td><td>45 mg Interchangeable Pellet Size Wheel</td></tr><tr><td>Camera</td><td>Lafayette Instrument</td><td>80600-CAM</td><td>Filtered and focused for IR light</td></tr><tr><td>Microsoft Windows</td><td>Microsoft</td><td></td><td>Windows 7-64 bit recommended</td></tr><tr><td>Controller PC-Touch Screen Chambers</td><td>Lafayette Instrument</td><td>88530</td><td>Dual Core Pentinum Processor; 2.5 GHz or greater; keyboard, mouse, and monitor; Installed PCI card, cable and expansion; Four RS232 ports; Four VGA ports</td></tr><tr><td>Dustless Precision Pellets</td><td>BioServe</td><td>F0165</td><td>45 mg</td></tr><tr><td>Black Acrylic Mask</td><td>Everything Plastic</td><td>Custom Product</td><td>Black polycarbonate with one matte finish side, 3.18 mm thick; Central circular opening, 28.58 mm diameter, 107.95 mm from the bottom, centered between the left and right side; Two square response areas, 28.57 mm x 28.57 mm each positioned below and off center, one 85.725 mm from the left and one 85.725 mm from right side, and both 34.925 mm from the bottom.</td></tr><tr><td>Utility Cart</td><td>Fisher Scientific</td><td>11-954-754</td><td>30.75 in. x 18.38 in x 33 in</td></tr><tr><td>Black towel</td><td></td><td></td><td>Large enough to cover cages for animal transport</td></tr><tr><td>Headlights with red lights</td><td>Energizer</td><td>HDL33A2E</td><td>LED white and red light</td></tr></tbody></table>

touchscreen sustained attention task (sat) for rats

Sustained attention is the ability to monitor intermittent and unpredictable events over a prolonged period of time. This attentional process subserves other aspects of cognition and is disrupted in certain neurodevelopmental, neuropsychiatric, and neurodegenerative disorders. Thus, it is clinically important to identify mechanisms that impair and improve sustained attention. Such mechanisms are often first discovered using rodent models. Therefore, several behavior procedures for testing aspects of sustained attention have been developed for rodents. One, first described by McGaughy and Sarter (1995), called the sustained attention task (SAT), trains rats to distinguish between signal (i.e., brief light presentation) and non-signal trials. The signals are short and thus require careful attention to be perceived. Attentional demands can be increased further by introducing a distractor (e.g., flashing houselight). We have modified this task for touchscreen operant chambers, which are configured with a touchscreen on one wall that can present stimuli and record responses. Here we detail our protocol for SAT in touchscreen chambers. Additionally, we present standard measures of performance in male and female Sprague-Dawley rats. Comparable performance on this task in both sexes highlights its use for attention studies, especially as more researchers are including female rodents in their experimental design. Moreover, the easy implementation of SAT for the increasingly popular touchscreen chambers increases its utility.

For most attention studies, rats are trained to criteria on SAT and then manipulations to disrupt or improve attention are performed. Acquisition data is not typically presented, but we do so here to illustrate rates of acquisition in male and female Sprague-Dawley rats. To this end, we quantified the median number of days to reach criteria on SAT (starting on the Nose Poke Phase). Note that a few rats never made it to criteria (n = 5 males, and n = 1 female). The median days that it took...

Watch this Scientific Journal Video about Touchscreen Sustained Attention Task SAT for Rats at JoVE.com

Touchscreen Sustained Attention Task SAT for Rats

Sustained attention, or continuously monitoring situations for intermittent and unpredictable events, is a critical aspect of cognition. Here we detail how to test sustained attention in rats using touchscreen operant chambers. We demonstrate comparable performance in male and female rats, making this task useful for studying attention in both sexes.

Touchscreen Sustained Attention Task (SAT) for Rats

Sustained attention is the ability to monitor intermittent and unpredictable events over a prolonged period of time. This attentional process subserves ...

touchscreen-sustained-attention-task-sat-for-rats

Research

JoVE Journal

Neuroscience

9.7K Views.  Temple University. Sustained attention, or continuously monitoring situations for intermittent and unpredictable events, is a critical aspect of cognition. Here we detail how to test sustained attention in rats using touchscreen operant chambers. We demonstrate comparable performance in male and female rats, making this task useful for studying attention in both sexes.

Video: Touchscreen Sustained Attention Task SAT for Rats

A Fully Automated Rodent Conditioning Protocol for Sensorimotor Integration and Cognitive Control Experiments

Rodents have been traditionally used as a standard animal model in laboratory experiments involving a myriad of sensory, cognitive, and motor tasks. Higher cognitive functions that require precise control over sensorimotor responses such as decision-making and attentional modulation, however, are typically assessed in nonhuman primates. Despite the richness of primate behavior that allows multiple variants of these functions to be studied, the rodent model remains an attractive, cost-effective alternative to primate models. Furthermore, the ability to fully automate operant conditioning in rodents adds unique advantages over the labor intensive training of nonhuman primates while studying a broad range of these complex functions.
Here, we introduce a protocol for operantly conditioning rats on performing working memory tasks. During critical epochs of the task, the protocol ensures that the animal's overt movement is minimized by requiring the animal to 'fixate' until a Go cue is delivered, akin to nonhuman primate experimental design. A simple two alternative forced choice task is implemented to demonstrate the performance. We discuss the application of this paradigm to other tasks.

A fully automated protocol for rodent operant conditioning is proposed. The protocol relies on precise temporal control of behavioral events to investigate the extent to which this control influences neural activity underlying sensorimotor integration and cognitive control experiments.

Rodents have been traditionally used as a standard animal model in laboratory experiments involving a myriad of sensory, cognitive, and motor tasks. ...

Behavior

Automated Visual Cognitive Tasks for Recording Neural Activity Using a Floor Projection Maze

Neuropsychological tasks used in primates to investigate mechanisms of learning and memory are typically visually guided cognitive tasks. We have developed visual cognitive tasks for rats using the Floor Projection Maze1,2&#160;that are optimized for visual abilities of rats permitting stronger comparisons of experimental findings with other species.
In order to investigate neural correlates of learning and memory, we have integrated electrophysiological recordings into fully automated cognitive tasks on the Floor Projection Maze1,2. Behavioral software interfaced with an animal tracking system allows monitoring of the animal's behavior with precise control of image presentation and reward contingencies for better trained animals. Integration with an in vivo electrophysiological recording system enables examination of behavioral correlates of neural activity at selected epochs of a given cognitive task.
We describe protocols for a model system that combines automated visual presentation of information to rodents and intracranial reward with electrophysiological approaches. Our model system offers a sophisticated set of tools as a framework for other cognitive tasks to better isolate and identify specific mechanisms contributing to particular cognitive processes.

We describe protocols for training rats for chronic electrophysiological recordings in fully automated cognitive tasks on a Floor Projection Maze.

Neuropsychological tasks used in primates to investigate mechanisms of learning and memory are typically visually guided cognitive tasks. We have ...

The 5-Choice Serial Reaction Time Task: A Task of Attention and Impulse Control for Rodents

This protocol describes the 5-choice serial reaction time task, which is an operant based task used to study attention and impulse control in rodents. Test day challenges, modifications to the standard task, can be used to systematically tax the neural systems controlling either attention or impulse control. Importantly, these challenges have consistent effects on behavior across laboratories in intact animals and can reveal either enhancements or deficits in cognitive function that are not apparent when rats are only tested on the standard task. The variety of behavioral measures that are collected can be used to determine if other factors (i.e., sedation, motivation deficits, locomotor impairments) are contributing to changes in performance. The versatility of the 5CSRTT is further enhanced because it is amenable to combination with pharmacological, molecular, and genetic techniques.

This protocol describes the 5-choice serial reaction time task, which is an operant based task used to study attention and impulse control in rodents. Test day challenges, which are modifications of the standard task, increase flexibility of the task and can be combined with other manipulations to more fully characterize behavior.

This protocol describes the 5-choice serial reaction time task, which is an operant based task used to study attention and impulse control in rodents. ...

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 ...

A Fully Automated and Highly Versatile System for Testing Multi-cognitive Functions and Recording Neuronal Activities in Rodents

We have developed a fully automated system for operant behavior testing and neuronal activity recording by which multiple cognitive brain functions can be investigated in a single task sequence. The unique feature of this system is a custom-made, acoustically transparent chamber that eliminates many of the issues associated with auditory cue control in most commercially available chambers. The ease with which operant devices can be added or replaced makes this system quite versatile, allowing for the implementation of a variety of auditory, visual, and olfactory behavioral tasks. Automation of the system allows fine temporal (10 ms) control and precise time-stamping of each event in a predesigned behavioral sequence. When combined with a multi-channel electrophysiology recording system, multiple cognitive brain functions, such as motivation, attention, decision-making, patience, and rewards, can be examined sequentially or independently.

In this report, we present a fully automated and highly versatile system capable of simultaneously testing multi-cognitive behaviors and recording neuronal activities for rodents.

We have developed a fully automated system for operant behavior testing and neuronal activity recording by which multiple cognitive brain functions can be ...

Automated, Long-term Behavioral Assay for Cognitive Functions in Multiple Genetic Models of Alzheimer's Disease, Using IntelliCage

Multiple factors&#8212;such as aging and genes&#8212;are frequently associated with cognitive decline. Genetically modified mouse models of cognitive decline, such as Alzheimer&#39;s disease (AD), have become a promising tool to elucidate the underlying mechanisms and promote the therapeutic advances. An important step is the validation and characterization of expected behavioral abnormality in the models, in the case of AD, cognitive decline. The long-term behavioral investigations of laboratory animals to study the effect of aging demand substantial efforts from researchers. The IntelliCage system is a high-throughput and cost-effective test battery for mice that eliminates the need for daily human handling. Here, we describe how the system is utilized in the long-term phenotyping of a genetic Alzheimer&#39;s disease model, specifically focusing on the cognitive functions. The experiment employs repeated battery of tests that assess spatial learning and executive functions. This cost-effective age-dependent phenotyping allows us to identify the transient and/or permanent effects of genes on various cognitive aspects.

This paper describes a protocol for cognitive assessments for genetic models of the Alzheimer&#39;s disease using the IntelliCage system, which is a high throughput automated behavioral monitoring system with operant conditioning.

Multiple factors&#8212;such as aging and genes&#8212;are frequently associated with cognitive decline. Genetically modified mouse models of cognitive ...

Automated Rat Single-Pellet Reaching with 3-Dimensional Reconstruction of Paw and Digit Trajectories

Rodent skilled reaching is commonly used to study dexterous skills, but requires significant time and effort to implement the task and analyze the behavior. Several automated versions of skilled reaching have been developed recently. Here, we describe a version that automatically presents pellets to rats while recording high-definition video from multiple angles at high frame rates (300 fps). The paw and individual digits are tracked with DeepLabCut, a machine learning algorithm for markerless pose estimation. This system can also be synchronized with physiological recordings, or be used to trigger physiologic interventions (e.g., electrical or optical stimulation).

Rodent skilled reaching is commonly used to study dexterous skills, but requires significant time and effort to implement the task and analyze the behavior. We describe an automated version of skilled reaching with motion tracking and three-dimensional reconstruction of reach trajectories.

Rodent skilled reaching is commonly used to study dexterous skills, but requires significant time and effort to implement the task and analyze the ...

Tracking Rats in Operant Conditioning Chambers Using a Versatile Homemade Video Camera and DeepLabCut

Operant conditioning chambers are used to perform a wide range of behavioral tests in the field of neuroscience. The recorded data is typically based on the triggering of lever and nose-poke sensors present inside the chambers. While this provides a detailed view of when and how animals perform certain responses, it cannot be used to evaluate behaviors that do not trigger any sensors. As such, assessing how animals position themselves and move inside the chamber is rarely possible. To obtain this information, researchers generally have to record and analyze videos. Manufacturers of operant conditioning chambers can typically supply their customers with high-quality camera setups. However, these can be very costly and do not necessarily fit chambers from other manufacturers or other behavioral test setups. The current protocol describes how to build an inexpensive and versatile video camera using hobby electronics components. It further describes how to use the image analysis software package DeepLabCut to track the status of a strong light signal, as well as the position of a rat, in videos gathered from an operant conditioning chamber. The former is a great aid when selecting short segments of interest in videos that cover entire test sessions, and the latter enables analysis of parameters that cannot be obtained from the data logs produced by the operant chambers.

This protocol describes how to build a small and versatile video camera, and how to use videos obtained from it to train a neural network to track the position of an animal inside operant conditioning chambers. This is a valuable complement to standard analyses of data logs obtained from operant conditioning tests.

Operant conditioning chambers are used to perform a wide range of behavioral tests in the field of neuroscience. The recorded data is typically based on ...

An Open-Source, Fully Customizable 5-Choice Serial Reaction Time Task Toolbox for Automated Behavioral Training of Rodents

The 5-choice serial reaction time task (5-CSRTT) is a behavioral test often used to study visuospatial attention and impulsiveness in rodents. The task requires animals to allocate attention to a horizontal array of five small apertures equipped with light sources and, within a limited time window, nose-poke one illuminated target aperture to get a food reward at the food magazine located in the opposite wall of the chamber. The task considers behavioral control measures such as response accuracy and reaction times and allows to infer selective attention and impulsivity. Task difficulty can be controlled by modifying the stimulus duration and task design in general. Commercially available apparatus usually consists of an experimental chamber and particular software to specify task parameters, but due to fixed hard- and software, they pose many limitations on changes in the general experimental design and specific task requirements and the related data output. This article explains a fully customizable alternative based on an easy-to-use single-board microcontroller and standard electrotechnical components, an open-access Arduino script, and a Matlab-toolbox for hardware control and behavioral task specifications, respectively. The toolbox includes an optional staircase procedure, enabling automated behavioral training. The complete hardware setup, which can be installed in customized chambers, and the freely adaptable software encourage non-standardized task and chamber design. The design of the system and the open-source code for hardware control and experimental setup are described.

The present protocol describes the development of an open-source 5-choice serial reaction time task toolbox for rodent animal models, using Arduino and related hardware and a versatile Matlab toolbox, including an optional script for automated behavioral training. The scripts are customizable and facilitate the implementation of different trial-and-test designs.

The 5-choice serial reaction time task (5-CSRTT) is a behavioral test often used to study visuospatial attention and impulsiveness in rodents. The task ...