A, O-LM cell and (inset) voltage response to a 1 s, ± 90 pA current step from −60 mV. B, voltage response of O-LM cell in A in control (a) and in the presence of 10 μm muscarine (b). C–G, sinusoidal currents at frequencies 0.5–100 Hz were applied in control (grey) and after application of 10 μm muscarine (black). Bias current was applied such that the cell was maintained at a similar voltage just below threshold in all conditions. C, in response to an 8-Hz sinusoidal current injection (4 s, 120 pA peak to peak), the cell fired more regularly in the presence of 10 μm muscarine than in control. D, plot of AP reliability versus frequency of sinusoidal current for cell in C before () and after application of 10 μm muscarine (•) shows this cell was maximally reliable (0.57) at 4 Hz in control (grey arrow) and maximally reliable (0.76) at 6 Hz in 10 μm muscarine (black arrow). E, plot of AP reliability versus frequency for a population of 8 cells (excluding 1 cell, which was done at select frequencies) shows that mAChR activation significantly increased (P < 0.05) reliability at each frequency between 5 and 12 Hz (P > 0.05 at frequencies < 5 Hz and > 12 Hz). **P = 6 × 10⁻⁵, averaging across 5–12 Hz. F, overlay of traces from dotted box in C, illustrating that mAChR activation induced phase shift. F, lower, overlay of traces from dotted box in the above panel, illustrating that mAChR activation was associated with an increased slope (dV/dt) on the upswing of the sinusoidal cycle. G, plot of slope prior to the AP versus frequency, as measured in a 3-ms window 2 ms prior to the onset of the AP in control (F, grey bar) and in the presence of 10 μm muscarine (F, black bar). *Slope significantly increased (P < 0.05) at each frequency between 6 and 11 Hz. H, plot of slope versus frequency with slope measured at a fixed point in the sinusoidal cycle (in a 74–79 degree window; between dotted lines in F); *slope significantly increased (P < 0.05) at each frequency between 4 and 12 Hz.

A and B, overlaying each cycle (9 Hz shown) illustrates spikes arriving earlier and more precisely in the presence of 10 μm muscarine (black traces) compared with control (grey traces). The upper trace indicates the input current over a single cycle. Changes in AP amplitude between conditions were not consistently observed. C, for the representative cell in A, plot of phase (a), and increase in precision, as measured in degrees (b) and milliseconds (c) versus frequency (○: control; •: 10 μm muscarine). Phase was defined as the time from the trough of the input current to the time of the first spike, expressed as degrees (t_{trough-to-spike} (seconds) × frequency (Hz) × 360 degrees). Precision was quantified as the standard deviation (s.d.) of the first spike time per cycle, expressed both in degrees (phase error) and seconds (jitter). D, in a population of 9 cells, averaging over the theta band (5–12 Hz), APs arrived significantly earlier (a; 170 ± 16 degrees versus 154 ± 12 degrees; P = 0.03) and more precisely (b, phase s.d.: 9.8 ± 1.3 degrees versus 6.2 ± 1.5 degrees, P = 0.005; c, jitter: 3.5 ± 0.5 ms versus 2.2 ± 0.6 ms, P = 0.005).

A, inset, in a representative cell, rotation number (average spikes per cycle; R#) gradually declines from 12.5 at 0.5 Hz to 0 at 100 Hz in control (○), and 10.7 at 0.5 Hz to 0.016 at 100 Hz after application of 10 μm muscarine (•). A, plot of R#versus frequency shows that the cell fires approximately every cycle (R#= 1) from 5 to 12 Hz in control (width = 7 Hz) and from 7 to 17 Hz after application of 10 μm muscarine (width = 10 Hz). B, inset, mAChR activation increases the average R# in the theta band from 0.79 ± 0.07 (open symbols) to 1.11 ± 0.08 (filled symbols; n = 9, P = 0.007). C, representative cells for frequency bands at which R# is 1 ± 0.4 (vertical grey bar in A) in control and 10 μm muscarine (black bars). D, averaging across cells shows this bandwidth is significantly wider after application of 10 μm muscarine (8.4 ± 2.4 versus 15.3 ± 2.0 Hz; P < 0.05).

A, representative voltage output in response to a 4 s, 90 pA sinusoidal current in the absence (a) or presence (b) of 10 μm muscarine. B, AP phase over the course of the 4 s train for 3 trials in both control (grey) and 10 μm muscarine (black). Expanded, overlaid traces during the first (C; 0–1 s in A) or last (D; 3–4 s in A) second of the sinusoidal current injection. While the APs occur at a similar time in the cycle initially, mAChR activation shifts the timing of AP generation to earlier in the cycle. E, phase for control (grey; n = 7) and upon mAChR activation (black; n = 8) in the first (0–1) or last (3–4) second. In control conditions, the phase increased from 212 ± 3 to 220 ± 3 degrees (6 of 7 cells, P = 0.03; P = 0.13 for all 7 cells). Concomitant with the development of a muscarinic ADP (by 1.7 ± 0.5 mV, P = 0.01, n = 10), mAChR activation shifted phase from 218 ± 6 to 208 ± 6 degrees (n = 8, P = 0.026). The ADP was measured by comparing the difference in average voltage in seconds 0–1 versus 3–4 of the sinusoidal stimulation; the sinusoidal stimulation itself generated no net voltage difference in control conditions (see Methods).