Bridge balance was maintained and Rseries monitored in all experiments. Recordings were stopped if Rseries became >25 MΩ. CN-SO slices were prepared by cutting 1,000- to 1,500-μm-thick coronal brainstem sections that were tilted slightly in the rostral-caudal Selleck Galunisertib axis so that the auditory nerves, cochlear nuclei, and superior olivary complexes were contained within one slice. Slices were incubated in a custom interface chamber at 35°C for 30–60 min and then held at room temperature for up to 30 min before transfer to the recording chamber. Survival of the input circuitry

to the MSO was presumably enhanced by the fact that most of the input circuitry is not far from the ventral surface of the slice. We also found that slice health was much better at the age range used (P15–P20) than at older ages, presumably due to enhanced ability of younger tissue to withstand hypoxia. Recordings were made at 35°C while slices were perfused with ACSF at 8–10 ml/min. Auditory nerve stumps were stimulated with suction electrodes. MSO neurons were patched under visual control at a depth of ∼25–200 μm

below the slice surface. In cells where nerve stimulation evoked both EPSPs and IPSPs, these events were typically evoked together only over a narrow range of stimulus amplitudes, and the minimal stimulus required for evoking each type of event was similar but not identical. Increases in stimulus current beyond this narrow range caused the EPSP or IPSP to fail, possibly due to depolarization block of individual axons in the auditory nerve stump. Thus, the lowest stimulation current that reliably evoked both EPSPs and IPSPs was used. Recordings were made from 200 μm NVP-BKM120 solubility dmso horizontal slices prepared from P21–P32 gerbils. Slices were perfused with 37°C ACSF at ∼2 ml/min. In experiments involving stimulation of excitatory

afferents, ACSF contained 1 μM strychnine and 5 μM SR-95531 (gabazine) to block glycine and GABAA receptors. Inhibitory afferents were isolated by ACSF containing 20 μM CNQX and 50 μM D-APV to block AMPA and NMDA receptors. When afferent stimulation was not used, ACSF contained 1 μM strychnine, 20 μM CNQX, and 50 μM D-APV. For dynamic-clamp recordings, cells were patched with two somatic electrodes, one to measure Vm and the other Oxymatrine to inject current. The dynamic clamp used SM2 software from Cambridge Conductance to control a Toro 8 DSP circuit board operating at 33–50 kHz. EPSGs were simulated with a double exponential waveform (time constants = 0.1 ms rise, 0.18 ms decay) and reversal potential of 0 mV. IPSGs were simulated with a double exponential waveform (time constants = 0.28 ms rise, 1.85 ms decay) and reversal potential of −85 mV (physiological inhibition) or equal to Vrest (∼−58 mV, purely shunting inhibition). The peak conductance of IPSGs was adjusted so that an individual event elicited a 3 mV hyperpolarization from Vrest. Inhibitory step conductances used the same reversal potentials as IPSGs.