The commissure of the inferior colliculi (CoIC) reciprocally innervates the inferior colliculi (ICs), the principal midbrain auditory nuclei. The CoIC interconnects the ordered representation of sound frequency in the ICs, but its function is not understood. We investigated the role of these connections in the processing of sound by reversibly deactivating spiking activity in one IC using a cryoloop while recording sound driven responses in the other. Experiments were performed in guinea pig anaesthetised with urethane (1.0 g/kg, 20% solution, i.p.) and Hypnorm (1mg/kg i.m.). Thermocouple measurements (Hyp-0, Omega) determined that cooling was restricted to the directly cooled IC. Single and multi-unit recordings revealed a gradient of deactivation through the cooled IC with deactivation maximal dorsally, adjacent to the cryoloop, while more ventral areas were less affected by cooling. A linear regression fitted the maximal change in firing rate as a function of characteristic frequency (r2 = 0.82, P < 0.001). Averaged auditory evoked potentials to clicks showed that the amplitude of the afferent volley to the IC was unchanged by cooling (Wilcoxon matched-pairs test; P = 0.0825) suggesting that lower auditory nuclei were unaffected. This was corroborated by thermocouple measurements in the cochlear nucleus ipsilateral to cooling. Together these observations suggest that the effects of cooling on the contralateral IC are mediated via the CoIC and not by descending connections from the IC. The frequency response areas (FRAs) of most single units (57/94) in the IC contralateral to cooling were modulated (> 20% area change) by deactivation. A one sample Wilcoxon signed rank test showed that cooling produced a significant reduction in the area of FRAs (P = 0.0017). Cooling changed the areas of 39/72 V-shaped FRAs and across the population there was a significant reduction in area (Wilcoxon matched-pairs; P = 0.036). The change in area of non-V-shaped FRAs did not reach significance for the population (Wilcoxon matched-pairs; P = 0.08) despite a change in 18/23 FRAs. Of the 18 FRAs that changed with cooling, 8 increased and 10 decreased in area. When the direction of change in the FRA area was ignored, there was a significant difference between the change in V-shaped and non-V-shaped FRAs (Mann Whitney test; P = 0.0052), with non-V-shaped FRAs showing a bigger area change with cooling. These results show that cryoloop cooling offers a focal, reversible deactivation technique, suitable for IC deactivation in guinea pig. Recordings contralateral to the deactivated IC indicate that projections via the CoIC can modulate the encoding of spectral cues in IC neurons. Differential changes in V-shaped and non-V-shaped units suggest that the CoIC selectively modulates frequency receptive fields, possibly sub serving intercollicular communication between cells possessing similar properties.