advanced startle - Med Associates Inc.
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1. 0 SB 50 75 10 125 150 175 20 25 250 275 MO MS 0 375 0 425 0 45 w Time msec Cus A 19 0 0000 lula Tene mone RVers BE Chamber 1 Trial J 5 Cursor A LI nina Sl la Pot Coke mam d Ana 45 ziam The primary feature of Startle Viewer is the voltage graph Figure 0 1 Select the particular trial to view on the graph by either double clicking on the trial number in the table window or by using the toggle switch in the lower right corner of the screen labeled Trial Use Cursors A and B to pinpoint and quantify certain locations on the graph For instance to quantify the peak to peak frequency in milliseconds place cursor A on one peak and cursor B on the subsequent peak Doing this also allows the user to quantify maximum peak amplitude within a specified period of time Another benefit of Startle Viewer is that it determines the slope of a line in volts per millisecond for any selected peak amplitude The x axis and y axis can be modified by clicking on the icons in the lower portion of the screen The precision and notation of each dimension can be altered and the overall scale of the graph can be modified using the magnifying glass icon and the zoom function Startle Viewer can be used to generate data regarding startle latency a dependent measurement taken in milliseconds which is defined as the time period in between the white noise startle stimulus presentation and the maximal startle respon2. 0 O O O O O DOC 005 Rev 2 6 Copyright 2012 MED Associates Inc APPENDIX C CUSTOMIZING TABLES IN MICROSOFT EXCEL Modifying experimental parameters to create experiments is easy with Advanced Startle All experimental tables conclude with txt but it is recommended that tables be created using a spreadsheet program such as Microsoft Excel and then saved as a tab delimited text format txt file To modify the value for any parameter in the table enter the desired value in the correct row column location and save the file as described Then open Advanced Startle and select the new experiment file using the Main Menu Load Experiment menu choice Extracting Data from a text file into Microsoft Excel A spreadsheet program can be used to open data saved as an ASCII text file txt This guideline illustrates an example of how to open ASCII data using Microsoft Excel2000 1 In Excel select File Open Locate where the file has been saved Select Text Files txt in the file type drop down menu Choose the file to open and click Open This will automatically open the first of three windows for the Text Import Wizard The data file is tab delimited so make sure to select Delimited Start the import at row 1 the file origin Windows ANSI Click Next to bring up the second of the three step process Select tab delimiters and click Next The final stage of the import wizard should now be open Make sure
3. Chamber Ctrl R Saves all the trial information for that chamber Trial Ctrl T Saves only the data for that trial Opening a Data File A window automatically appears upon opening Startle Viewer that prompts the user to select a data file Find the appropriate directory and select the data file to view All readable data files conclude with RAW Another window will appear after opening the desired data file and select the appropriate chamber to import the data from that chamber 18 DOC 005 Rev 2 6 Copyright 2012 MED Associates Inc The Table Display Startle Viewer allows the user to view the startle output in volts for any particular trial The Startle Viewer table window lists the maximum startle output by trial number and the corresponding stimuli us used in that particular trial Note that RAW data files are matched to txt files with the same name and the txt versions are the data files used to display data in the Excel spreadsheet Viewing a Particular Startle Response Figure 0 1 Startle Viewer Window P agy e Co bgt ib Ted Te ci c2 VoRsPP Commeritsi Comments2 Cormenersts3 Date Time D 0019 CSUS STRNIL 19995 J xvas 2 012 BAC STRNIO 161 82 29 AM 3 00 s TIMK18 179 4 02 BK TIAK101 5 OW MAIO T KI 14 131 6 0520 BA T K10210 V 7 05 TRAIN TEST RAW 5 File JEMALI 09 17 05 LTE o 10 0 A B Cursor Date veks soj 01465 604 Time 1 mox 40 20 Cursor B
4. 5 Click Finish to close this screen Figure 0 4 Advanced Startle Installation Checklist Advanced Startle Install Wizard m pd associates inc Welcome to the install wizard for Advanced Startle The wizard will install Advanced Startle on your computer To continue click Install DIG 729 Driver ANL 929A PC Driver NI DAQ Software and Driver Advanced Startle neu Figure 0 5 Advanced Startle Driver and Software Installation Complete 7 Advanced Startle Install Wizard med A550CITES INE Install Wizard Complete The Install Wizard has successfully installed Advanced Startle Click Finish to exit the wizard V DIG 729 Driver of ANL 929A PC Driver y NI DAQ Software and Driver Advanced Startle 21 DOC 005 Rev 2 6 Copyright 2012 MED Associates Inc APPENDIX F REFERENCES 1 10 Falls W A Carlson S Turner J G amp Willott J F 1997 Fear potentiated startle in two strains of inbred mice Behavioral Neuroscience 111 855 61 Josselyn S A Shi C Carlezon W A Jr Neve R L Nestler E J amp Davis M 2001 Long term memory is facilitated by cAMP response element binding protein overexpression in the amygdala Journal of Neuroscience 21 2404 2412 Antioniadis E A Winslow J W Freedman L Davis M amp Amaral D G Submitted Role of the primate amygdala in fear potentiated startle Effe
5. 70 dB Tone Neutral Stimulus Noise Burst Onset Il Startle Response Noise Burst Startle Stimulus Fi Onset of 70 dB Tone 70 dB Tone Conditioned Stimulus 0 4 mA Aversive Stimulus Pavlovian Fear Conditioning gt Neutral Stimulus Becomes Fear Provoking Conditioned Stimulus 90 100 sec 70 dB Tone Conditioned Stimulus Noise Burst Onset NA Potentiated Startle Response a Noise Burst Startle Stimulus f Onset of 70 dB Tone Figure created by modifying previous figure from W A Falls PhD 14 DOC 005 Rev 2 6 Copyright 2012 MED Associates Inc MED ASSOCIATES INC SOF 828 ADVANCED STARTLE Experiment Initiated by the FPStrain txt Procedure The Fear Potentiated Experiment protocol describes two separate procedures The first procedure is used to create Pavlovian conditioning in a test subject This experiment is described in the FPStrain txt Experiment Table shown in Figure 3 13 This experiment consists of 10 trials each trial in its own column in the table columns D through M Inspection of the table reveals the fact that the only Trial Variable that changes from trial to trial is the ACC ITI variable it ranges from 150 to 300 seconds Figure 3 13 FPStrain txt Experiment Table A B C D E F G H l J K IL M 1 STL DISPLAY 0 5 ACCIITI 300 210 210 240 120 210 180 150 180 150 2 CS DISPLAY 0 5 AUX1 CS 0 O O O 1 D 0 O O O 3 CS FREEZE O AUX2 CS 0 O O 0
6. ACC ITI for a particular trial is 30 seconds then the Event Marker will occur 30 seconds after the previous trial s Event Marker Trial Variables include either the suffix CS condition stimulus or STL startle such as AUX1 CS or AUX STL All stimuli present PRIOR to the Event Marker include the suffix CS Therefore if AUX1 CS is defined as 2 5 then this stimulus will turn on 2 5 seconds prior to the Event Marker and stay on for 2 5 seconds All CS stimuli will terminate at the Event Marker unless CS STL DELAY is set to a value other than 0 see the upcoming discussion on CS STL DELAY If a stimulus is to remain on past the Event Marker then include a value for that stimulus in the group of variables that have the suffix STL For instance if the AUX1 stimulus is to remain on for 1 7 seconds past the Event Marker define AUX1 STL as 1 7 In this example the AUX1 stimulus would remain on for a total of 4 2 seconds 2 5 seconds before the Event Marker and 1 7 seconds afterward Figure 2 1 Event Marker and Trial Timeline EVENT MARKER Time Ligat US Noise gt Intertrial Interval ACC ITI Next Intertrial Interval CS DISPLAY STL DISPLAY CS Defined Variables STL Defined Variables Rev 2 5 Copyright 2012 MED Associates Inc SOF 828 ADVANCED STARTLE CS DISPLAY amp STL DISPLAY These two variables determine the time window
7. AUX1 1 Always on O NOISE CS dB 0 O O 9 AUX2 1 Always on O TONE CSidB O 70 70 10 AUX3 1 Always on O TONE CSiHz O 12000 4000 11 AUX4 1 Always on O TONE STL Hz 0 0 O 12 Reserved O TONE STL dB 0 O O 13 Reserved 3 AUX1 STL O O O 14 Reserved O AUX2 STL 0 O O 15 Reserved O AUX3 STL O 0 O 16 Reserved O AUX4 STL O O O 17 Reserved O TONE STL O O O 18 Reserved O NOISE STL 0 01 0 01 0 01 19 DAQ Gain olts 10 NOISE STL dB 100 100 100 20 DAQ Freguencyi Hz 1000 Trial Comment 1 NB 70 12 70 04 21 Reserved O Trial Comment 2 100 100 100 22 Reserved O CS STL DELAY 0 1 0 1 0 1 Column A lists the titles of Control Variables that stay constant throughout the experiment The values of these variables are found in the corresponding row in column B for instance STL DISPLAY is set to 0 3 in seconds and this means that the startle response will be recorded for 300 milliseconds for every trial Column C lists the Trial Variables and these are the parameters that define the stimulus presentations and inter trial interval durations their values are found in the corresponding row in Columns D and greater E F etc which describe individual trials for the experiment The other Control Variables for this experiment are as follows RISE TIME TONE is set at 3 milliseconds which is the rise time of the tone stimulus LATENCY is also set to 3 milliseconds Essentially Latency is the amount of time set between the onset of a response and when a peak
8. PPI has been observed in mice rats Rhesus Macagues and humans Rodent testing of PPI is used in the testing of therapeutic compounds and has been used as a screening technique for transgenic knock out mice Figure 3 10 Prepulse Inhibition of Startle PPI Startle Stimulus Alone Noise Burst Onset Startle Response Prepulse and Startle Stimulus Noise Burst Onset Inhibited Startle Response 100 ms Prepulse Onset Figure courtesy of William A Falls PhD University of Vermont 11 DOC 005 Rev 2 6 Copyright 2012 MED Associates Inc Experiment Initiated by the PPinhibition txt Procedure Earlier it was stated that all experiment files contain data to set the values for both Control and Trial Variables while the program is in execution As noted earlier these files are in ASCII format so they can be viewed in spreadsheet software like Microsoft Excel Shown in Figure 3 11 is the PPinhibition txt experiment file as it appears when it is opened in Microsoft Excel The experiment is controlled by the values entered in the PPinhibition txt file A description of the variables in the experiment follows Figure 3 11 PPinhibition txt Experiment Table A B Cc D E F 1 STL DISPLAY 0 3 ACC ITI 60 5 Hi 2 CS DISPLAY 0 AUX1 CS O O O 3 CS FREEZE O AUX2 CS 0 O O 4 RISE TIME NOISE ms O AUXx3 CS 0 O O 5 RISE TIME TONE ms 3 AUX4 CS O 0 O 6 LATENCY ms 3 TONE CS O 0 11 0 11 7 BACKGROUND dB O NOISE CS O 0 O 8
9. Qa D O 0 a O 4 RISE TIME NOISE ms 1 AUX3 CS 0 O O O D O 0 O 0 O 5 RISE TIME TONE ms 50 AUX4 CS 0 a O O 0 Oo D O 0 O 6 LATENCY ms O TONE CS 29 5 29 5 29 5 29 5 29 5 29 5 29 5 29 5 29 5 29 5 7 BACKGROUND dB O NOISE CS 0 O O O D O Oo 0 O O 8 AUX1 1zAlways on O NOISE CS dB 0 O a O Oo D D O O O 9 AUX2 15Always on O TONE CS dB 70 70 70 70 70 70 70 70 70 70 10 AUX3 15Always on O TONE CS Hz 12000 12000 12000 12000 12000 12000 12000 12000 12000 12000 11 AUX4 1 Always on O TONE STL Hz 12000 12000 12000 12000 12000 12000 12000 12000 12000 12000 12 Reserved O TONE STL dB 70 70 70 70 70 70 70 70 70 70 13 Reserved 3 AUX1 STL 0 5 0 5 0 5 0 5 0 5 0 5 0 5 0 5 0 5 0 5 14 Reserved O AUX2 STL O O a Oo 0 D D O O O 15 Reserved O AUX3 STL 0 O O O D D QO O O O 16 Reserved O AUX4 STL 0 O O O 0 0 0 O 0 O 17 Reserved O TONE STL 0 5 0 5 0 5 0 5 0 5 0 5 0 5 0 5 0 5 0 5 18 Reserved O NOISE STL 0 O O O D D Qa 0 O O 19 DAQ Gain Volts 10 NOISE STL dB 0 O O O 0 D 0 0 O O 20 DAQ Freguency Hz 1000 Trial Comment 1 21 Reserved O Trial Comment 2 22 Reserved 0 CS STL DELAY 0 O 0 O 0 O 0 O 0 O 15 DOC 005 Rev 2 6 Copyright 2012 MED Associates Inc MED ASSOCIATES INC SOF 828 ADVANCED STARTLE Experiment Initiated by the FPStest txt Procedure The second procedure is used to test a subject that has been trained using the first procedures This experiment is described in the FPStest txt Experiment Table shown in Figure 3 14 The procedure be
10. during which the behavioral data will be recorded and is displayed on the voltage graph in Advanced Startle or Startle Viewer CS DISPLAY records behavioral activity PRIOR to the Event Marker and STL DISPLAY records activity for a specified duration of time AFTER the Event Marker STL DISPLAY also includes the time data used to calculate peak to peak intervals The total duration of behavioral recording CS DISPLAY STL DISPLAY should not exceed 2 seconds CS STL DELAY The CS STL DELAY sets the Controlled Stimulus Uncontrolled Stimulus CS US trace interval and overrides the values set for CS durations If a value of CS STL DELAY is set to 1 5 in the table for any given trial all CS defined variables will terminate 1 5 seconds PRIOR to the Event Marker If for instance the AUX1 CS is set to 2 5 and the CS STL DELAY is set to 1 5 then the AUX1 CS stimulus will turn on for 1 second and the turn off 1 5 seconds PRIOR to the Event Marker In Figure 2 2 the AUX1 CS is turned off at the commencement of the CS STL DELAY and no stimulus is present until the Event Marker at which time the AUX2 STL stimulus turns on Figure 2 2 Event Marker and AUX1CS AUX2 STL Timeline EVENT MARKER A AUX1 CS AUX2 STL CS OFF CS STL DELAY Session Duration Session Duration is the sum of ACC ITI values for all the trials plus the STL DISPLAY value There is a minimum 2 second requirement between the offset of any post Event Marker variable
11. e g STL defined stimuli or STL DISPLAY and the onset of any pre Event Marker variable e g CS defined stimuli or CS DISPLAY Rev 2 5 Copyright 2012 MED Associates Inc SOF 828 ADVANCED STARTLE CHAPTER 3 HARDWARE CONFIGURATION AND CALIBRATION Refer to the MED ASR Startle Hardware manual for hardware configuration instructions For Audio Calibration and Input load cell platform Calibration procedures we recommend that you refer to calibration procedures described in the Startle Reflex User s Manual The Hardware menu in the Startle Reflex software runs utilities for complete testing and calibration of all Startle hardware Rev 2 5 Copyright 2012 MED Associates Inc SOF 828 ADVANCED STARTLE APPENDIX A DEFINITIONS OF CONTROL AND TRIAL VARIABLES Figure 3 1 contains definitions of the Control Variables that are used to initialize an experiment in Advanced Startle Please note that the Control Variables will not change during execution of the program Figure 3 1 Advanced Startle Control Variable Definitions A Control Variables Defined 1 STL DISPLAY The time during which activity baseline or startle is recorded Enter the time value in seconds 2 CS DISPLAY Records the subject s activity immediately prior to STL DISPLAY Enter a time value here in seconds Records data but does not display it This option is good for recording data to be stored 3 CS FREEZE in the RAW data file 3 This value sets
12. that general is listed in the data format box and click Finish The desired data file table should appear in an Excel spreadsheet upon completing the Text Import Wizard 17 DOC 005 Rev 2 6 Copyright 2012 MED Associates Inc APPENDIX D USING STARTLE VIEWER Advanced Startle Software is equipped with Med Associates Startle Viewer This is a brand new feature of the Advanced Startle Software that allows the researcher to examine the details of the startle reflex and manually adjust the scale of the primary dependent measure Use Startle Viewer to quantify the amplitudes and frequencies of the startle response Menu Selections After opening Startle Viewer three options appear in the upper left corner of the screen These options and corresponding keystroke commands are File Alt F Open Ctrl O New File Ctrl F Opens data file Readable data files conclude with RAW Chamber Ctrl H Selects a specific chamber to view Save Ctrl S Saves the Startle Viewer file to a specified directory Quit Ctrl Q Exits Startle Viewer About Ctrl A Provides information regarding the Startle Viewer program Edit Alt E Cut Ctrl X Copy Ctrl C Paste Ctrl V Export Alt P Saves the RAW file in ASCII tabbed file format Compression Ctrl M Select how the file is compressed Compressing using 2X 5X or 10X saves every second fifth or tenth data point None means the file will not compress Range Ctrl R
13. the rise time of the white noise stimulus in milliseconds The 4 Rise Time Noise ms A recommended value is 1 K This value sets the rise time of the tone stimulus The recommended minimum value is 5 Rise Time Tone ms 3 6 LATENCY ms Peak to peak values are recorded after specified value in milliseconds This value should remain at 0 to exclude this option Constant background noise If a value is entered here a noise will be present during the 7 BACKGROUND dB entire experimental session at the dB value entered here and this variable should range from 70 to 115 db 8 AUX1 1 Always on If 1 is entered here then the stimulus corresponding to AUX1 on PHM 255C will be ON throughout the experiment 9 AUX2 1 Always on If 1 is entered here then the stimulus corresponding to AUX2 on PHM 255C will be ON throughout the experiment 10 AUX3 1 Always on If 1 is entered here then the stimulus corresponding to AUX3 on PHM 255C will be ON throughout the experiment 11 AUX4 1 Always on If 1 is entered here then the stimulus corresponding to AUX4 on PHM 255C will be ON throughout the experiment 12 Reserved N A keep this value at 0 13 Reserved N A keep this value at 0 14 Reserved N A keep this value at 0 15 Reserved N A keep this value at 0 16 Reserved N A keep this value at 0 17 Reserved N A keep this value at 0 18 Reserved N A keep this value at 0 19 DAQ Gain Volts This represents the maxi
14. ae MEO associates inc instrumentation and software for research ADVANCED STARTLE SOF 828 USER S MANUAL DOC 005 Rev 2 5 Copyright 2012 All Rights Reserved Med Associates Inc P O Box 319 St Albans Vermont 05478 Phone 802 527 2343 Fax 802 527 5095 www med associates com SOF 828 ADVANCED STARTLE SOF 828 ADVANCED STARTLE Table of Contents Chapter 1 IMErOAUCUHON sisi iscsnsccscncscccesscesnasessnsassancesssssadsensessecbesdensssestsnsonacshsesesnsncssnades 1 Chapter 2 Description of Advanced Startle Software 2 Timeline for Creating a Trial Zg o rane ii a ea ben 2 CS DISPLAY St STLADISPLAV onina aa a ataata ae aaah aiieieo iaa 3 CSS TH DELAY EA E T E E E ES A 3 SESSION Durations en e a anton ies 3 Chapter 3 Hardware Configuration and Calibration ccsssecsssessseeeeeeees 4 Appendix A Definitions of Control and Trial Variables 5 Appendix B Pre Programmed Experimental Procedures 7 Loading and Executing a Pre Programmed File ccescccssessesssesseseescsesesseeecsesesseeessesaseees 7 Prepulse Inhibition of Startle PPI e nana 11 Experiment Initiated by the PPinhibition txt Procedure cecccecsseeeecscsssseeseseseeseeessesees 12 Fear Potentiated Startle FPS sni eth sitet a ots tte EN ae re ae te ue eli a al 14 Experiment Initiated by
15. ckgrounds motor tests Genes Brain amp Behavior 3 206 15 APPENDIX G CONTACT INFORMATION Please contact MED Associates Inc for information regarding any of our products Visit our website at www med associates com for contact information For technical questions email support med associates com 22 DOC 005 Rev 2 6 Copyright 2012 MED Associates Inc
16. cts of transient inactivation or chronic lesions in the Rhesus Monkey Maccaca mulatta Winslow J T Parr L A amp Davis M 2002 Acoustic startle prepulse inhibition and fear potentiated startle measured in rhesus monkeys Biological Psychiatry 51 859 866 Baas J M Grillon C Bocker K B Brack A A Morgan C A Kenemans J L amp Verbaten M N 2002 Benzodiazepines have no effect on fear potentiated startle in humans Psychopharmacology 161 233 47 Schmidt L A Fox N A Goldberg M C Smith C C amp Schulkin J 1999 Effects of acute prednisone administration on memory attention and emotion in healthy human adults Psychoneuroendocrinology 24 461 83 Guitar B 2003 Acoustic startle responses and temperament in individuals who stutter Journal of Speech Language amp Hearing Research 46 233 240 Depoortere R Perrault G amp Sanger D J 1997 Potentiation of prepulse inhibition of the startle reflex in rats pharmacological evaluation of the procedure as a model for detecting antipsychotic activity Psychopharmacology 132 366 374 Olivier B Leahy C Mullen T Paylor R Groppi V E Sarnyai Z amp Brunner D 2001 The DBA 2J strain and prepulse inhibition of startle a model system to test antipsychotics Psychopharmacology 156 284 290 Brooks S P Pask T Jones L amp Dunnett S B 2004 Behavioural profiles of inbred mouse strains used as transgenic ba
17. e amplitude of startle on these two trial types that is our operational measure of fear Rev 2 5 Copyright 2012 MED Associates Inc SOF 828 ADVANCED STARTLE CHAPTER 2 DESCRIPTION OF ADVANCED STARTLE SOFTWARE The Advanced Startle Software allows investigators to use both pre programmed research protocols for performing both Prepulse Inhibition of Startle PPI and Fear Potentiated Startle FPS experiments and user defined research protocols based on the researcher s present needs The software reads files that have been written in a tabular format to execute a particular experimental procedure These files are in ASCII format so they can be viewed in spreadsheet software like Microsoft Excel The purpose of the files is to initialize both Control and Trial Variables that the software uses to execute an experiment To understand what these variables represent one should be familiar with the definition of an Event Marker Understanding the Event Marker is critical to understanding what data has been recorded from a trial and when it was recorded The Event Marker is anchor for the timeline in a trial Timeline for Creating a Trial Figure 2 1 illustrates the events occurring within any given trial The Event Marker divides the trial into two parts and initiates STL DISPLAY one of two time periods during which the subject s activity will be captured The value set for ACC ITI determines when the Event Marker occurs therefore if
18. g data by block or stimulus condition Enter any descriptor here Turns off all CS durations prior to Startle Onset Normally set to 0 for no delay Rev 2 5 Copyright 2012 MED Associates Inc SOF 828 ADVANCED STARTLE APPENDIX B PRE PROGRAMMED EXPERIMENTAL PROCEDURES Loading and Executing a Pre Programmed File The installation of the Advance Startle program included a folder called Program Files Advanced Startle Tables The following experimental programs tables are listed in the folder These are the files used to execute the following pre programmed experiments Open the Advanced Startle software and the main screen shown in Figure 3 3 will appear Figure 3 3 Advanced Startle Main Screen 2 MED Advanced Startle Start Pause gt 00 00 C Acclimation 00 00 si 00 00 0 cs 00 00 10 04 10 0 1 1 O Chamber 1 O Chamber 2 Startle o Trial O of 0 10 0 10 0 Experiment File 5 0 5 0 0 0 0 0 Data File 50 50 10 0 10 0 O Chamber 3 O Chamber 4 Comments 10 0 moa 5 0 5 0 0 0 0 0 Review Trial gdo ooo aux wy B 0 000 au oo 10 0 O Chamber 5 0 chi ber 6 AUX amber AUX4 JI pet Tone i 5 04 Nose Wo 0 04 Back Ground 5 0 10 0 4 10 0 O Chamber 7 i O Chamber 8 Select Menu Load Experiment as shown below and a File Name dialog box as shown in Figure 3 5 will open Figure 3 4 Menu Load Experiment Edit L
19. gins with 9 noise burst only trials described in columns D through L that are followed by randomly selected noise burst and pre pulse trials for the remainder of the experiment Oo Om wy ajo 12 13 14 15 16 dizi 18 19 20 21 2 Figure 3 14 FPStest txt Experiment Table A STL DISPLAY CS DISPLAY CS FREEZE RISE TIME NOISE ms RISE TIME TONE ms LATENCY ms BACKGROUND dB AUX1 1 Always on AUX2 1 Always on AUX3 1 Always on AUX4 1 Always on Peak Calc Method Reserved Reserved Reserved Reserved Reserved Reserved DAQ Gain Volts DAQ Freguency Hz Reserved Reserved B C 0 5 ACCITI 0 5 AUX1 CS O AUX2 CS 1 AUX3 CS 1 AUX4 CS O TONE CS O NOISE CS O NOISE CS dB O TONE CS dB O TONE CS Hz O TONE STL Hz O TONE STL dB 3 AUX1 STL O AUX2 STL O AUX3 STL O AUX4 STL O TONE STL O NOISE STL 10 NOISE STL dB 1000 Trial Comment 1 NB O Trial Comment 2 O CS STL DELAY D 300 0 O 0 O 0 0 03 105 70 12000 12000 70 O Ooooocoo 105 O NB 16 m H J K E M 60 60 60 60 60 60 Oo 0 0 O 0 O 0 0 O 0 O 0 Oo 0 O O 0 O 0 0 O 0 O 0 0 0 O 0 O 29 03 0 03 0 03 0 03 0 03 0 03 0 03 110 105 110 105 100 105 70 70 70 70 70 70 12000 12000 12000 12000 12000 12000 12000 12000 12000 12000 12000 12000 70 70 70 70 70 70 0 Oo O O O O 0 0 O 0 O 0 0 0 O 0 O 0 0 Oo 0 O O 0 O 0 O 0 O 0 47 0 0 O O 0 O 0 Oo O O O O NB NB NB NB NB 12k Tone 110 105 110 105 100 105
20. have been given values In the case of the second trial a 70 dB tone set in TONE CS dB will be emitted from the speaker at 12000 Hz set in TONE CS Hz for duration of 0 01 seconds set value of TONE CS 0 11 seconds set value in CS STL DELAY 0 10 seconds 0 01 seconds This is a functional description of how the software will reference an executable txt file during the execution of the experiment Let us now consider the Fear Potentiated Startle experiment and its experiment tables for training and testing of a subject 13 DOC 005 Rev 2 6 Copyright 2012 MED Associates Inc MED ASSOCIATES INC SOF 828 ADVANCED STARTLE Fear Potentiated Startle FPS Fear potentiated startle FPS occurs when the startle response to an acoustic stimulus is increased relative to baseline startle responding via Pavlovian conditioning The general procedure for producing FPS involves preceding an aversive stimulus such as a shock or air puff with a tone or some other neutral stimulus After several presentations of this stimulus pairing the presentation of the tone elicits a fear response Once conditioning has occurred the tone is then presented before an acoustic startle stimulus increasing the intensity of the startle response FPS is a typical reflex that has been observed in a variety of species thus making it a useful tool for investigating many learning and memory processes Figure 3 12 Fear Potentiated Startle FPS
21. igure 3 7 Enter a Data File Name Save in EEE A E 061107 E3 txt E vT3txt 61107 E3 pre2 txt E VT4 txt My Recent 61107 F3 txt Documents 061107 F3 pre2 txt 061107 H3 txt 3 061107 H3 pre2 txt 61107 N3 txt 161107 N3 pre2 txt Desktop My Documents RE TEST TEST txt est txt rain O62107 txt E vT2 txt My Computer a File name test txt My Network Saveastype Custom Pattemn txt During the experiment the program file selected is listed in the Experiment File box and the data file to save data in is listed in the Data File box as shown in Figure 3 8 Data will be saved to the data file specified even if the experiment is aborted before the session has expired Figure 3 8 Experiment and Data File Names 2 MED Advanced Startle Abort Pause Sp 229 Cs Acclimation 04 48 Is 00 00 cs 00 29 Starte Trial 1 of 10 Experiment File C Program Files Advanced Startle a Tables FPStrain txt Data File C Program Files Advanced Startle data test txt Advanced Startle includes a total of eight startle response display windows one window corresponds to each of eight chambers The startle response in volts for each trial is displayed in the corresponding chamber window DOC 005 Rev 2 6 Copyright 2012 MED Associates Inc MED ASSOCIATES INC SOF 828 ADVANCED STARTLE Figure 3 9 Advanced Startle Screen with Response in D
22. isplay Window af MED Advanced Startle Menu Edit Abort Pause gt 2X CD sccimatori 04 55 si 00 00 cs 00 29 10 0 State trali of 10 0 Chamber 2_ 10 0 Experiment File C Program Files Advanced Startle Tables FPStrain txt o0 DataFile C Program Files Advanced Startle detal test tet Ji 10 0 ___O Chamber 4 __ 10 0 5 0 5 0 Comments _ 5 0 0 0 4 Review lu lu HE D 0 Trial 000 000 O oa 8 v N Please note that only the first trial will be displayed initially To see the results of the current trial its number is shown in the Trial XX of XX boxes in the top left part of the display toggle the Review Trial control in the lower left corner to find the trial of interest Alternatively the trial number can be entered into the text box provided then hit Enter to bring up the data recorded for that particular trial The Review Trial feature shows the maximum startle output for a selected trial per chamber The data column to the right displays the data for the most recent trial 10 DOC 005 Rev 2 6 Copyright 2012 MED Associates Inc Prepulse Inhibition of Startle PPI PPI of the startle response modeled in Figure 3 10 is a behavioral phenomenon in which a short tone presented just prior to an acoustic startle stimulus a sound presented at a fairly high amplitude will serve to dampen the subsequent startle response
23. load value will be considered as the result of the startle stimulus If the amount of time between the onset of a response and a peak is less than the Latency value the peak is not counted as a response 12 DOC 005 Rev 2 6 Copyright 2012 MED Associates Inc DAQ Gain Volts is the maximum voltage recorded from the load cell and typically should be left at 10 volts The DAQ Frequency Hz is set to 1000 this is the sample frequency for the program and is generally left at 1000 samples second A trial is an event during which a response is recorded whether it is a baseline measurement of activity or a startle response Columns D and E F etc above list the corresponding parameter values for every trial In other words these columns are user defined and each column identifies the stimulus properties used during a single trial Therefore the number of columns that contain values from columns D E etc determines the total number of trials in a particular experiment The protocol for this experiment has randomly intermixed trials that receive a noise burst only with those that receive a prepulse tone before the delivery of the noise burst the experiment shows 10 trials columns D through M The first trial shown in column D is a trial that receives a noise burst only This is true because only the NOISE STL and NOISE STL dB Trial Variables have been given values The Trial Comment cells allow the user to define a particula
24. mum voltage recorded from the load cell Keep this at 10 20 DAOFreguency Hz The number of samples taken per second leave this at 1 000 21 Reserved N A keep this value at 0 22 Reserved N A keep this value at 0 Likewise Figure 3 2 contains definition of the Trial Variables that are used to initialize an experiment Please note that the nature of these variables is to allow the control of the behavior of the program in any given trial and due to this at any given moment in the dynamic Rev 2 5 Copyright 2012 MED Associates Inc SOF 828 ADVANCED STARTLE execution of the program the values may be different then they were at the beginning of the session o on o 11 12 13 14 15 16 17 18 19 20 21 22 Figure 3 2 Advanced Startle Trial Variable Definitions ACC ITI AUX1 CS AUX2 CS AUX3 CS AUX4 CS TONE CS NOISE CS NOISE CS dB TONE CS dB TONE CS Hz TONE STL Hz TONE STL dB AUX1 STL AUX2 STL AUX3 STL AUX4 STL TONE STL NOISE STL NOISE STL dB Trial Comment 1 Trial Comment 2 CS STL DELAY Trial Variables Defined Interval of time between the previous trial s Event Marker and this current trial s Event Marker The duration in seconds that the stimulus corresponding to AUX1 on PHM 255C is on prior to the Event Marker The duration in seconds that the stimulus corresponding to AUX2 on PHM 255C is on prior to the Event Marker The duration in seconds that the
25. oad Experiment Ctri L p Hardware Quit About Rev 2 5 Copyright 2012 MED Associates Inc MED ASSOCIATES INC SOF 828 ADVANCED STARTLE Choose the directory and file to execute and click OK all executable files will have the file extension txt Figure 3 5 Select the File to Execute Look in My Recent Documents 3 Desktop My Documents My Computer My Network ek FpStest txt FPStrain txt E MouseFPS PreTest txt E PPinhibition txt E Rat FPS Exercise 3 txt E Rat Train txt E tone fear conditioning txt File name Files of type JENV 230 Tone Test tt Custom Pattern txt Cancel The name of the file selected shown with its full path will appear in the Experiment File box just under the Trial X of X boxes in the left part of the display Figure 3 6 Experiment File Box Experiment File C Program Files Advanced Startle Tables FPStrain txt DOC 005 Rev 2 6 Copyright 2012 MED Associates Inc MED ASSOCIATES INC SOF 828 ADVANCED STARTLE Click the Start button and another File Name dialog box will appear as shown in Figure 3 7 Enter a name for the data file a folder in the C Program Files Advanced Startle directory called data has been provided to store data files The file will get a txt extension by default Clicking the OK button will automatically begin the experiment F
26. r trial using an identifier The identifier is a sequence of alphanumeric characters no more than 6 digits and or letters and is intended to define a unique tag to help sort the data once it has been imported into a spreadsheet program like Microsoft Excel The Trial Comment cells will be present in the table of the Startle Viewer and in the txt data files that can be used for data analysis Therefore use identifiers that will allow the data to be sorted easily for analysis The computer program does not read the Trial Comment cells during the execution of the experiment so values entered here will not impact the procedure Note that Trial Comment 1 entry for the first trial column D is NB and Trial Comment 2 entry is 100 These values can be used to sort the trials in the experiment that receive only a noise burst at 100 dB The first trial column D occurs 60 seconds 1 minute after the experiment is initiated the next trial column E occurs 5 seconds later the third trial column F occurs 7 seconds after that etc These values are defining the ACC ITI Trial Variable which sets the inter trial interval i e the amount of time between event markers The ACC ITI value set in the first trial is used to define the length of the acclimation period for the test subject The second trial column E is one in which the noise burst will be preceded by a prepulse tone since the TONE CS TONE CS dB and the TONE CS Hz Trial Variables
27. se Select the specific trial to examine and the startle response will appear in the main display Fix cursor A on time point zero and move cursor B to the peak positive amplitude for that trial Note the difference between cursor positions along the x axis 19 DOC 005 Rev 2 6 Copyright 2012 MED Associates Inc MED ASSOCIATES INC SOF 828 ADVANCED STARTLE APPENDIX E DRIVER AND SOFTWARE INSTALLATION Insert the Advanced Startle CD into the CD ROM drive and the screen shown in Figure 0 2 will appear Click Install Advance Startle and the screen shown in Figure 0 3 will appear Figure 0 2 Advanced Startle CD Main Screen uni SA Advanced Startle m m med JSSOCIATES ING _ Welcome to Advanced Startle Install Advanced Startle Extras Browse the CD Enter the Advanced Startle password that was issued by MED Associates when the software was registered and click OK Figure 0 3 Enter Password Advanced Startle Please enter your password 20 DOC 005 Rev 2 6 Copyright 2012 MED Associates Inc MED ASSOCIATES INC SOF 828 ADVANCED STARTLE The checklist shown on the screen below indicates the items that will be installed Click Install to begin A green check mark will appear next to each item on the list once it has been successfully installed Complete the steps on each screen and once all of the drivers and software are installed the screen shown in Figure 0
28. stimulus corresponding to AUX3 on PHM 255C is turned on prior to the Event Marker The duration in seconds that the stimulus corresponding to AUX4 on PHM 255C is turned on prior to the Event Marker The duration of the tone stimulus prior to the Event Marker The duration of the noise stimulus prior to the Event Marker The dB of the NOISE CS and this variable should range from 70 to 115 db The dB of the TONE CS and this variable should range from 70 to 115 db The frequency of the TONE CS this variable should range from 4 000 to 20 000 The frequency of the TONE STL see below this variable should range from 4 000 to 20 000 The dB of the TONE STL and this variable should range from 70 to 115 db The duration of the AUX1 stimulus in seconds that is turned on after the Event Marker The duration of the AUX2 stimulus in seconds that is turned on after the Event Marker The duration of the AUX3 stimulus in seconds that is turned on after the Event Marker The duration of the AUX4 stimulus in seconds that is turned on after the Event Marker The duration of the tone stimulus occurring after the Event Marker The duration of the noise stimulus occurring after the Event Marker The dB of the NOISE STL this variable should range from 70 to 115 db A description of the particular trial use this to sort data for analysis Enter any descriptor here because this cell is not read during execution A secondary descriptor for sortin
29. the FPStrain txt Procedure cccccssesscescsssseseeecsseseeeeecsesseeeseeeeeas 15 Experiment Initiated by the FPStest txt ProCeCure cccccccssesssscscssesssececsssseeecessessserseseeeas 16 Appendix C Customizing Tables in Microsoft Excel ee 17 Extracting Data from a text file into Microsoft Excel 17 Appendix D Using Startle Viewer cesscsssecssseeessecesssseeesseeecsseceessscessaeeees 18 M nu SelectioN Seenen nn a A A ea a E E nama etiaty 18 Opening a Data FINS reac tin nak vaste a acted e re aa A diab ve Sea a i 18 The Table DS Ua cet hits Ail le Ne S et A ul A a nal Lt 19 Viewing a Particular Startle RESPONSE x decree cen elas uted Aa do an odno 19 Appendix E Driver and Software Installation ne 20 Appendix F REfCrences cee ccssscssssecesssesssceessseesssseeessecessssesessecesseesessececsaeeess 22 Appendix G Contact Information nnnneca 22 SOF 828 ADVANCED STARTLE Rev 2 5 Copyright 2012 MED Associates Inc SOF 828 ADVANCED STARTLE CHAPTER 1 INTRODUCTION The Startle paradigm is used to study fear and anxiety in laboratory animals Although the underlying circuits and systems controlling fear and anxiety are complex the Startle paradigm itself is relatively simple it is a form of Pavlovian conditioning where a central state of fear is inferred from an animal s behavioral response This is done by measuring the amplit
30. ude of a simple brainstem reflex the acoustic startle reflex in the presence of a cue the conditioned stimulus CS previously paired with a mild footshock the unconditioned stimulus US For example when animals are repeatedly presented with a neutral stimulus one that does not elicit any behavioral effect on its own such as a light flash a tone or an odor followed by a brief footshock the animal comes to associate the presentation of the cue with an impending aversive event the shock Eventually presentation of the cue in the absence of the shock comes to elicit a constellation of behaviors that are typically used to define a state of fear in animals Using SOF 828 Advanced Startle to accomplish this animals are placed in cages in an isolation chamber the same chamber in which animals received the prior fear conditioning and presented with a number of startle eliciting white noise bursts The cages are positioned on a sensitive load cell device that transduces movement of the cage produced by the animal s startle response into an electrical signal that is amplified and quantified in arbitrary startle units On some trials the startle eliciting stimulus is preceded by the cue previously paired with shock In this case the startle response is much larger i e potentiated than on trials without presentation of the cue and it is the difference expressed either as an absolute value or a percent change in different cases between th
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