Balanced excitation and inhibition determine spike timing during frequency adaptation

Abstract

In layer 4 (L4) of the rat barrel cortex, a single whisker deflection evokes a stereotyped sequence of excitation followed by inhibition, hypothesized to result in a narrow temporal window for spike output. However, awake rats sweep their whiskers across objects, activating the cortex at frequencies known to induce short-term depression at both excitatory and inhibitory synapses within L4. Although periodic whisker deflection causes a frequency-dependent reduction of the cortical response magnitude, whether this adaptation involves changes in the relative balance of excitation and inhibition and how these changes might impact the proposed narrow window of spike timing in L4 is unknown. Here, we demonstrate for the first time that spike output in L4 is determined precisely by the dynamic interaction of excitatory and inhibitory conductances. Furthermore, we show that periodic whisker deflection results in balanced adaptation of the magnitude and timing of excitatory and inhibitory input to L4 neurons. This balanced adaptation mediates a reduction in spike output while preserving the narrow time window of spike generation, suggesting that L4 circuits are calibrated to maintain relative levels of excitation and inhibition across varying magnitudes of input.

Figure 1.

Adaptation of synaptic and spike responses to 10 Hz PW deflection in L4 barrel cortex neurons. A, Single-unit recording (Extra), example trace, and corresponding PSTH (1 ms bins) are shown. Stimulus times are indicated above (Wh). The depth of the recording was 830 μm. B, Intracellular recording (Intra) from a different animal, example trace, and corresponding PSTH are shown. Resting V_m was –72 mV. The depth of the cell was 510 μm. Details below, indicated by arrows, are 30 superimposed traces for the first and 10th deflection in the train. Stimulus times are indicated by arrowheads. C, Average (Avg) synaptic responses to the first through 10th deflection for the cell in B. First (#1) and 10th (#10) responses are highlighted in black. D, Mean PSP amplitude (Amp) for all intracellular recordings (n = 31) and mean spike output for all cells with suprathreshold responses (intracellular plus single unit recordings; n = 32), normalized to the magnitude of the first response. Error bars indicate ±SEM. E, Reconstruction of a L4 (500–850 μm) spiny stellate and pyramidal neuron recorded from a single micropipette penetration. An additional layer 3 pyramid, recorded in the same track, is shown. The inset shows a lower magnification image of the contralateral hemisphere that was processed for cytochrome oxidase; dashed lines indicate depth of darkly stained barrels in L4.

Figure 2.

Temporal changes in the synaptic and spike responses during frequency adaptation. A, Synaptic response of an L4 neuron(660 μm) to the first (Stim #1) and 10th (Stim #10) PW deflection in a 10 Hz train. The top traces are 10 sequential synaptic responses, and raster plots illustrate corresponding spike output. The bottom traces are the average synaptic responses from 30 deflections. Resting V_m was –73 mV. Stimulus time is indicated by an arrowhead. Dashed lines are the 10–90% slope (dV/dt) of the PSPs. B, Population spike output for the first (Stim #1) and 10th (Stim #10) PW deflection. Overlaid traces are the PSTHs (1 ms bins) for all cells with suprathreshold responses. Stimulus time is indicated by arrowheads. C, Mean onset latency, peak latency, and dV/dt for all synaptic responses. Error bars indicate ±SEM. D, Mean latency, SD (σ), and vector strength (VS) for all spike responses.

Figure 3.

Contribution of excitatory and inhibitory conductances to the PW-evoked synaptic response. A, Response to PW deflection of an L4 neuron (680 μm) filled with QX-314 and recorded at multiple V_m levels via current injection through the recording pipette. Stimulustime is indicated by an arrowhead. B, Plot of V_m versus injected current corrected for capacitative current (I_inj – I_cap; see Materials and Methods) for the time points in the response indicated in A. Inverse slope of the best-fit line through each set of points gives the total membrane conductance. Intersection of the line calculated during the response (gray squares) with the line calculated from baseline V_m (black squares) gives the apparent synaptic reversal potential (V_rev). C, Calculated apparent reversal potential (blue trace) and total synaptic (black trace), excitatory (green trace), and inhibitory (red trace) conductances for the PW-evoked response from A. The inset highlights the changes in reversal potential and conductances early in the response.

Figure 4.

Balanced frequency adaptation of excitatory and inhibitory conductances. A, Response to 10 Hz PW deflection of an L4 neuron (800 μm) filled with QX-314 and recorded at multiple V_m levels. B, Response to the first and 10th deflection from A. Traces are vertically offset to highlight the V_m dependence of PSP shape. Rasters and corresponding PSTHs (1 ms bins) illustrate spike output. Excitatory (green trace), inhibitory (red trace), and total synaptic (black trace) conductances underlying the responses are shown below. Color-coded circles indicate onset latencies. C, Left, Mean peak magnitude of excitatory (green filled triangles) and inhibitory (red filled triangles) conductances during 10 Hz train, normalized to the magnitude of the first response, for all QX-314-filled cells (n = 10). Right, Mean onset latency (green and red open squares) and peak latency (green and red filled circles) of excitatory and inhibitory conductances, respectively, during a 10 Hz train. D, Left, Plot of the normalized peak magnitude, for the 10th response, of excitation [g_E(10)/g_E(1)] versus inhibition [g_I(10)/g_I(1)] across all cells. Dashed lines indicate unity ± 20%. Right, Plot of PSP width, measured from –50 mV, versus the excitation dominance window, measured as the interval from excitation onset to peak inhibition.

Figure 5.

Simulated responses of an L4 neuron to balanced and unbalanced adaptation of excitation and inhibition. A, Parameters for simulated excitatory and inhibitory conductances were obtained from the data. Thin traces are individual traces of excitatory (green) and inhibitory (red) conductances to the first (Stim #1) and 10th (Stim #10) deflections in a train, taken from the data. Traces are normalized to the average peak amplitude. Overlaid thick traces are the simulated conductances used in the model. B, Simulated synaptic and spike responses of the model cell to the first (Stim #1, left panel) and 10th (Stim #10, 3 right panels) PW deflections in a 10 Hz train. The top histograms illustrate spike output for 100 stimuli, middle traces illustrate 20 example synaptic responses, and bottom traces illustrate g_E and g_I used to generate the responses. Responses to the 10th deflection were simulated under three conditions: (1) balanced adaptation of both excitatory and inhibitory conductances to 50% of the first stimulus magnitude, (2) increased adaptation of inhibition to 25% of the first stimulus magnitude, and (3) reduced adaptation of inhibition to 75% of the first stimulus magnitude. C, Plot of spikes/stimulus (filled circles), SD (filled triangles), and vector strength (open squares) for responses to the simulated 10th stimulus under varying conditions of adaptation of inhibition, expressed as a percentage of first stimulus magnitude. In all cases, adaptation of excitation was 50% of first stimulus magnitude.