Timeline of drug application and analysis of firing and release rates

30–60 s are typically needed for a drug to wash into the tissue, and this was the minimal wait time before analyzing dopamine effects. For cochlear afferent recordings, the rate of EPSC/EPSP and AP fluctuates in a power-law fashion throughout the recording, even in control condition, as described in Wu et al. (2016). Therefore, it is essential, to record for long enough in control, in DA, and after washout, to be able to separate random fluctuations from drug effects. This is why at least 30 s of recording were analyzed before, in and after DA.

To study the effects of DA, segments of recordings were selected for analysis. For extracellular loose-patch recordings of ANFs, 2 min before DA application, 2 min before the end of DA application, and 2 min after 2 min of washout, were taken as the ‘before’, ‘DA’ and ‘after’ time windows, respectively. The control experiments were performed and analyzed in the same way. The total application time for DA or Control solution was 3–5 min.

For whole-cell recordings of ANFs, 30 s to 2 min of recording before DA application was selected for ‘before’, 1 or 2 min of recording after at least 30 s of DA application was selected for ‘DA’, 30 s to 2 min of recording after at least 1.5 min of wash-out was selected for ‘after’. The total application time of DA varied between 1.5–6 min.