An acoustic startle-based method of assessing frequency discrimination in mice

Abstract

The acoustic startle response (ASR) is a reflexive contraction of skeletal muscles in response to a loud, abrupt acoustic stimulus. ASR magnitude is reduced if the startle stimulus is preceded by a weaker acoustic or non-acoustic stimulus, a phenomenon known as prepulse inhibition (PPI). PPI has been used to test various aspects of sensory discrimination in both animals and humans. Here we show that PPI of the ASR is an advantageous method of assessing frequency discrimination. We describe the apparatus and its performance testing frequency discrimination in young CD1 mice. Compared to classical conditioning paradigms, PPI of the ASR is less time consuming, produces robust results, and can be used without training even in young animals. This approach can be used to investigate the neuronal mechanisms underlying frequency discrimination, its maturation during development, and its relationship to tonotopic organization.

Fig. 1

Acoustic startle apparatus for frequency discrimination. (A) Schematic of apparatus. (B-D) Frequency spectra measured at the animal’s ear inside the housing within the anechoic chamber. (B) Ambient noise, (C) 16 kHz tone at 70 dB SPL used as the background frequency, and (D) white noise at 100 dB SPL. To protect the speaker during the 10 s sound delivery required for acquisition of the frequency spectra, white noise was delivered at 100 dB SPL, rather than the 120 dB SPL used to elicit the ASR. Arrow in (C) indicates the 70 dB SPL peak at 16 kHz. (E) Trial schematic. ITI, inter-trial interval; ISI, inter-stimulus interval; f₁, background frequency; f₂, prepulse frequency; Δf, frequency change.

Fig. 2

Inhibition of the ASR by a preceding frequency change. Traces represent the force measured on the platform during the first recording period, “baseline,” and second recording period, “ASR”. Examples of (A) startle only and (B) prepulse trials. For each trial, the maximum force recorded during the second recording period, positive or negative, was reported as the ASR. Scale bars, 0.4 arbitrary units of force, 100 ms. (C) Mean ASR for each trial type. Asterisks indicate that ASR magnitude differed significantly from that of the startle only ASR (p < 0.05, corrected for multiple comparisons). (D) Inhibition of the ASR by a frequency change of various magnitudes. Asterisks indicate frequency changes that caused significant inhibition of the ASR (p < 0.05, one-sample t-test against zero). Line represents a three-parameter sigmoid fit to the data (R² = 0.99). Dotted lines represent the 95% confidence interval of the sigmoidal fit. Dashed line indicates discrimination threshold. Data show mean ± s.e.m; n = 10 animals.