Willott1 project protocol

Startle and prepulse inhibition survey of 40 inbred strains of mice   (2002)

Willott JF, Johnson KR
With: Gagnon L, Reid MC, Tanner L, O'Steen J

See also: Willott1 animal documentation


See Willott et al., 2003 for detailed description.

Assays in order of testing:

1. PPI (defined frequencies of 70-dB SPL)
8 trials: S1(4 kHz) + S2
8 trials: S1(12 kHz) + S2
8 trials: S1(20 kHz) + S2

2. ASRbaseline
16 trials: startle stimulus (S2) only, 100-dB white noise

3. ASR (startle intensity function)
15 trials of each tone:
90 dB SPL
80 dB SPL
70 dB SPL

Equipment

Activity Monitoring / Startle Measuring System and screening software (Med Associates, Inc., Georgia, VT). This is a sound-attenuated test chamber on a movement-sensitive load cell (amplifier gain = 2.5).

Testing apparatus and calibration

Holding container: disposable 32 ounce plastic cup placed in test chamber. Acoustic stimuli generated by tweeter fit above rim of cup. No background noise applied, but ambient noise level in chamber was monitored and was < 50 dB SPL.

Acoustic calibration: 1/8 inch Bruel & Kjaer condenser microphone through hole in cup to measure intensity in various locations. Mean attenuation values to produce target intensities at each frequency were determined and programmed.

Standard startle stimulus (S2):
100 dB SPL white noise burst
Abrupt onset: 0 ms rise time setting
Duration: 10 ms

Prepulse Stimulus (S1):
70 dB SPL tone
Onset: 3 ms rise-fall time
Duration: 10 ms
Frequency: 4 kHz, 12 kHz, or 20 kHz
Timing: 100 ms after S1

Testing Schedule

Mice acclimated to holding cup for at least 1 min.

Trials contained an equal number of each stimulus delivered in a pseudo-random order; the number of trials for each test are noted above. Each mouse was tested in two rounds (second round immediately following the first).

Investigators notes:

1. Most mice were tested at The Jackson Laboratory but some mice from nine strains were tested at the University of South Florida. PPI and ASR were not significantly different as a result of the two test sites (shown by t-tests).

2. Inter-stimulus presentation rate, variable at 3-8 s.

3. Background noise was not generated in order to eliminate confounding effects on ASR amplitude; ambient noise was monitored.

4. To prevent inclusion of possible outlier data points, data for all mice were trimmed by discarding the 2 largest and 2 smallest baseline ASRs (S2-only). Not done for habituation index data.



Definitions:

ASR amplitude   peak voltage deflection 5-50 ms post startle stimulus (S2)

ASRbaseline   peak voltage deflection evoked by startle stimulus (S2) at 100 dB SPL

Habituation index   index of single session ASR test: ratio of ASR-trials #13-16/ASR-trials #1-4

Peak latency   time to peak amplitude after startle stimulus (S2)

PPI = S1+S2trial / ASRbaseline

PPI interval =100 ms (time between prepulse and S2)

PPIt = PPI total, sum of three PPI scores for each mouse