obtained during the final three days and the best day were taken as a measure of practiced detection. Behavioral data analysis. A performance value, d9 = z(false alarm) - z(hit), was obtained for z scores that corresponded to the right-tail p values [34,35], and was calculated for each MD. Thresholds were defined as the sAM depth at which performance reached a d9 = 1 and only sessions in which animal performed a minimum of 25 trials were included in the analysis. Psychometric functions of d9 across MD were constructed from the initial assessment with random stimuli and throughout each day of additional testing. The best or average performance during the final 3 days of testing served as the assessment of practiced sAM detection thresholds. Performance during training to criterion (when animals received warning trials of 100% MD) was quantified across all training trials (excluding the first 10) as average #hits - average #FAs. All values are given as mean 6 standard error (SEM). Neurophysiology Some of the control neurons presented here were used in a previous study [36]. That work included neurons with a range of best modulation frequencies (BMF; defined below). This study analyzes the subset of those cells with BMFs of 5 Hz, to match the behaviorally-tested MF (with the exception of responses across multiple modulation frequencies and for static tones). Surgical preparation and chronic recordings. Gerbils were premedicated with ketoprophen (1.5 mg/kg, i.n.) and dexamethasone (0.35 mg/kg, i.p.) and hydrated with normosol (1.5 ml, s.c.). The animals were anesthetized with isoflurane and held in a stereotaxic apparatus. A small headpost was positioned along the midline and secured with dental acrylic, and a silver ground wire was implanted into the posterior contralateral skull. A craniotomy was made over the left temporal cortex caudal to the bregma suture using stereotaxic coordinates [37], and the dura was left intact. A thin well of dental acrylic was built along the perimeter of the craniotomy, the cortical surface was covered with silicone oil, and the craniotomized area was covered with a disposable cap of silicone elastomer (Sammons Preston Rolyan, Bolingbrook, IL). Animals were placed in a soundproof chamber (Industrial Acoustics Company, Bronx, NY), and the head was stabilized using the headpost. During chronic recording sessions, animals stood comfortably on a platform and were free to move their limbs while the head position remained fixed. If an animal exhibited heightened anxiety or restlessness, medetomidine (a relaxant that acts on 2-adrenoceptors) was administered intranasally (0.1– 0.3 mg/kg). Gerbils trained to detect amplitude modulation are able to perform the task after receiving similar doses of medetomidine (D. H. Sanes, unpublished observations), and medetomidine does not produce an observable effect on neuronal activity [38]. Thus the results for cells recorded with and without medetomidine are presented together. The silicone elastomer cap was removed at the beginning of each session and a fresh cap applied at the end. The dura was covered with saline during recording to maintain moisture. Platinum-plated tungsten electrodes (1.5–2.5 MV; MicroProbe, Gaithersburg, MD) were advanced ventrally through the craniot- omy with an electrode tip angle of 14u lateral to vertical to isolate neurons in primary ACx. Single-unit recording procedures were identical to those described previously [36]. Acoustic stimulation for neurophysiology. The system used for stimulus generation and sound delivery (MALab, Kaiser Instruments) as well as the calibration procedure have been described previously [39]. Calibrated acoustic stimuli were Hearing Loss Disrupts AM Processing presented to each ear through an electrostatic speaker coupled to a custom-made ear insert. All stimuli were presented monaurally to the right ear except in cases in which a monaural stimulus failed to elicit reliable responses, in which case a binaural stimulus was used. To assess responses to static tones, neurons were presented with tone pips (200 ms, 5 ms cosine-ramped rise/fall time). First, the frequency range over which the neuron was responsive was obtained with an iso-intensity function at 10–30 dB above threshold (threshold criteria are described below). This was followed by a rate-level function (RLF) at the BF of the unit, measured at increments of 5 dB SPL, for 10–20 trials (with a 1 sec intertrial interval). To assess responses to sAM, each cell was presented with sAM tones (2 s duration with 10 ms rise/fall) at BF, at the SPL that produced the strongest response. This was typically 40 dB above the cell’s threshold. Modulation transfer functions (MTFs) were obtained by presenting five or ten trials of multiple modulation frequencies (1, 2, 5, 10, 20, 50, and 100 Hz) at 100% modulation depth with an intertrial interval of 1 sec. An unmodulated tone at the same frequency and SPL and of the same duration as the SAM tones was included in the stimulus sequence as a control. The best modulation frequency (BMF) was determined from the MTF as the modulation frequency eliciting the highest response strength (i.e., the product of firing rate and vector strength). Modulation depth functions (MDFs) were then obtained by presenting ten trials of multiple modulation depths at BF and BMF with an intertrial interval of 1 sec. The following modulation depths were typically presented to each ACx neuron: 5, 10, 20, 50, 70, 80 and 100% MD. Neurophysiological data analysis: RLFs and sAM responses. From RLFS, mean first-spike latency over 10 or more trials was calculated at BF and the SPL that produced the maximum response. Firing rates to tones were calculated over a time window equal to the stimulus duration. Maximum firing rates were calculated at the sound level that elicited the highest firing rate. Threshold, dynamic range and monotonicity were deter- mined from the RLF. Threshold was defined as the dB SPL level below which there was at least a 20% increase in firing rate, stepping up from one dB SPL level to the next; threshold firing rates were calculated at this sound level. Dynamic range was defined as the range between the dB SPL levels where each cell responded at 10% and 90% of its maximum firing rate, calculated by interpolation. To allow valid comparisons, RLF data were collected using equivalent ranges of levels across the two groups (0 to 90 dB SPL for CTRs, 30 to 120 dB SPL for CHLs). Group differences for each of these were assessed with t-tests. Non- monotonic neurons were defined as those whose firing rates at the highest dB SPL tested dropped below 50% of their maximum firing rate. Group differences for monotonicity were assessed using chi-square. Responses to sAM tones were analyzed using three measures: phase-locking to the modulation period (vector strength), firing rate, and power at the modulation frequency (referred to hereafter simply as power). These three measures were used to construct MTFs and MDFs based on phase locking, firing rate, and power. The first modulation period was excluded to eliminate contam- ination by the onset response [40]. First, vector strength [41] was calculated by treating every spike as a unit vector in a polar coordinate system, where the angle was the instantaneous phase, and the mean phase of the response was the direction of the sum of those vectors, normalized to the firing rate. Vector strength (the length of that mean vector) depends on how tightly the spikes cluster about that one point in the stimulus cycle, varying between PLoS ONE | www.plosone.org 3 July 2012 | Volume 7 | Issue 7 | e41514