Data from: Medial prefrontal cortex encodes but is not required to generate goal-directed actions under threat
Abstract
Adaptive behavior under threat requires deciding when to act and when to withhold action to avoid harm, often under conditions where movement, arousal, and task demand covary. Medial prefrontal cortex (mPFC) activity is widely associated with such control, yet it remains unclear whether this activity reflects causal action generation or broader evaluative processes shaped by behavioral state. Here, we combined fiber photometry, single-cell calcium imaging, mixed-effects modeling, and optogenetic inhibition to examine how GABAergic neurons in mouse mPFC represent cues, actions, and outcomes during a series of learned avoidance tasks of increasing complexity that promote cautious responding. By explicitly controlling for baseline activity and movement, we show that much apparent task-related activity in mPFC reflects movement and cue-evoked signals that are also present in a control cortical region, the visual cortex. mPFC GABAergic neurons showed little encoding of simple avoidance contingencies but broadly encoded punished outcomes. A small subset of neurons with strong movement sensitivity encoded more demanding avoidance contingencies requiring selection between action generation and deferment. For equivalent avoidance actions, distinct neuronal populations preferentially encoded either cue onset or the action. Despite this encoding, optogenetic inhibition of mPFC had minimal effects on the learning or performance of the different contingencies. These findings reveal a dissociation between neural encoding and causal necessity, indicating that mPFC GABAergic activity primarily reflects evaluative and contextual aspects of cautious avoidance behavior rather than direct control of action execution.
Each file is named by referencing the figure panel from which the data comes. Here we list, in bold, each figure panel, followed by the Parameter-Row or Column names within each file worksheet (the parentheses further define the term). Each name is defined by a brief description (e.g., File/getFile: filename defines File or getFile as the filename). Please use the figures in the paper as a reference.
Description of the data and file structure
File/getFile: filename (the filename field is always an identifier)
Figure 1. Calcium imaging fiber photometry reveals that both mPFC and VI GABAergic neurons activate during spontaneous exploratory movement.
Fig1 B ΔF/F calcium imaging time extracted around detected spontaneous movements recorded in mPFC or VI
Fig1 C Cross-correlation between movement and ΔF/F for the overall, translational, and rotational (right) components in mPFC and VI
Fig1 D Per-session and mean±SEM linear fit between overall movement and ΔF/F in mPFC and VI, including the rotational and translational components
Fig1-Sup1A-B ΔF/F calcium imaging, overall movement, rotational movement, and angle of turning direction for detected movements classified by the turning direction (ipsiversive and contraversive) versus the side of the recording in mPFC
Fig1-Sup1C-D Population measures (mean, peak amplitude, and time to peak for traces 3 s around the detected peaks) of ΔF/F and overall movement for the different classified peaks in mPFC.
Units: 0, 4, and 5 (integer indicates the peak type)
p=contraversive turn, n=ipsiversive turn
Area: measured area under the curve
Pk: measured peak value
xPk: time to Pk
Event: peak types
- Cxp=All peaks contraversive
- Cxp4=No peaks 3 sec prior contraversive
- Cxp5=1 peak every>5 sec contraversive
- Cxn=All peaks ipsiversive
- Cxn4=No peaks 3 sec prior ipsiversive
- Cxn5=1 peak every>5 sec ipsiversive
Figure 2. Behavioral performance in the cue-driven action tasks.
Fig2 B-C Behavioral performance across three sequential procedures: noUS, AA19, and AA39. Shows percentage of actions, action latency, and intertrial crossings.
File: filename
Tone Int: Tone intensities (Lo, Me, and Hi)
Task: Tasks (noUS, AA1=AA19, AA3=AA39)
Avoids%: percentage of avoids
Avoid Latency: latency of avoid responses
ITCs/trial: number of intertrial crossings per trial
Fig2-Sup1A-C Movement traces (overall speed) across task phases (noUS, AA19, AA39), cues (CS1–CS3), and cue intensities (low, medium, high), separated by Action and No-Action outcomes for all mPFC and VI mice.
File: filename
MPC1: Tone intensity-Tone frequency
Task1: Trial type - Tone frequency
Fig2-Sup2A-B Marginal means of movement from mixed-effects models. Population-level marginal means of movement speed estimated from linear mixed-effects models across all mice. A, compares trial types across task phases. B, compares Action versus No-Action outcomes within each task and CS.
Fac1: CS (8k=CS1, 4k=CS2, 12k=CS3)
Fac2: Task (noUS, AA1=AA19, AA3=AA39)
Fac5: Behavioral outcome (act=Action, noact=NoAction)
Subj: Animal
Figure 3. Simple (AA19) and challenging (AA39) avoidance contingencies mixed-effects models of ΔF/F activity controlling for movement and baseline activity.
Fig3 A-D Population estimates of marginal means from ΔF/F fiber-photometry mixed-effects models for AA19 (A,B ) and AA39 (C,D ) tasks, shown separately for mPFC neurons (A,C ) and VI neurons (B,D ). Datasets contain four rows corresponding to the baseline, orienting, action (CS-aligned), and from-action model windows.
Fac1: Ig, AA, PA (Ignore, Active Avoid, Passive Avoid)
Fac3: act=Action, noact=NoAction
Subj: Animal
Fig3 E Mean ΔF/F and movement traces for mPFC and VI neurons during the unsignaled escape task, showing interleaved trials of footshock alone, white noise alone, and the full US, aligned to US onset.
Fig3 F Population estimates of marginal means from ΔF/F mixed-effects models (including movement covariates) during the escape window (0-4 s from US onset).
Fac1: WNshk=US, shk=Shock, WN
Subj: Animal
Fig3-Sup1A-C ΔF/F fiber photometry traces and corresponding movement traces for mPFC mice across task phases (noUS, AA19, AA39) and cues (CS1–CS3), separated by Action (A) and No-Action (B) outcomes. (C) has the traces for the Action trials in A, replotted aligned to the action occurrence (From-Action is defined as the time at which the animal crosses the door). Otherwise, same as Fig2-Sup1A-C.
Fig3-Sup2A-D Full mixed-effects models of ΔF/F activity across task phases controlling for movement and baseline activity. Population estimates of marginal means from full mixed-effects models of ΔF/F fiber-photometry signals for mPFC (A, B) and VI (C, D) GABAergic neurons. Models include all task phases (noUS, AA19, AA39) and incorporate covariates for window-specific movement, baseline movement, and baseline ΔF/F (excluded from the baseline window). Otherwise, same as Fig2-Sup2A-B.
Figure 4. Movement-related neuron classes during a simple avoidance context (AA19 task).
Fig4 A-B (inset a) Mean ΔF/F traces and corresponding movement traces for five mPFC GABAergic neuron classes identified by k-means clustering of the cross-correlation functions between speed and ΔF/F (inset a). Class 1 and Class 2 exhibited moderate positive correlations with movement, Class 3 showed negative correlations, and Classes 4 and 5 exhibited strong positive correlations, with Class 5 centered near zero lag.
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MPC1: Tone intensities - Tone frequency
Fig4 C-D Estimated marginal means of ΔF/F from linear mixed-effects models for the baseline, orienting, action, and from-action windows. Models compared contingencies (Ig vs AA) and outcomes (action vs No-Action) across neuron classes. C, contrasts across contingencies within outcomes. D, contrasts between outcomes within each contingency and neuron class.
Fac1: Contingencies (Ig, AA)
Fac2: act=Action, noact=NoAction
Fac3: classes, (cl1=class2, cl2=class1, cl3=class4, cl4=class3, cl5=class5)
Subj: Animal
Fig4-Sup1 A-B (inset a) Movement-related neuron classes during a difficult avoidance context (AA39 task). A, ΔF/F traces and movement traces for five mPFC GABAergic neuron classes identified by k-means clustering of cross-correlation functions between speed and ΔF/F (inset a). Class 1 and Class 2 neurons show moderate positive correlations with movement. Class 3 neurons show negative correlations with movement. Class 4 and Class 5 neurons show strong positive correlations, with Class 5 activity preceding Class 4 around zero lag. Traces are aligned to CS onset and shown separately for CS1–CS3 and behavioral outcome (action vs. No-Action), which may reflect correct responses, errors, or spurious actions depending on contingency. In AA39, CS1 signals active avoidance and an action is the correct response, whereas CS2 signals passive avoidance and an action is an error. B, Same as A, aligned from-action (defined as the time at which the animal crosses the door) for Action outcomes. Otherwise, same as Fig4 A-B
Fig4-Sup1 C-D C, Marginal mean ΔF/F estimates from linear mixed-effects models for the baseline, orienting, action, and from-action windows, comparing Ignore (Ig), Active Avoid (AA), and Passive Avoid (PA) contingencies across outcomes (Action vs. No-Action) for each neuron class. D, Same as C, but comparing outcomes within each contingency for each neuron class.
Fac1: Contingencies (Ig, AA, PA)
Fac2: act=Action, noact=NoAction
Fac3: classes (cl1=class3, cl2=class4, cl3=class5, cl4=class1, cl5=class2)
Subj: Animal
Figure 5. Distinct mPFC GABAergic neuron activation types during active avoidance modes.
Fig5 A-B A, k-means clustering of ΔF/F time series from individual mPFC GABAergic neurons within each active avoidance mode revealed three activation types (a–c). B, Same as A, aligned from action.
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MPC1: Tone intensities - Tone frequency
Fig5 C Population estimates of marginal means (ΔF/F) from linear mixed-effects models for the baseline, orienting, action, and from-action windows, comparing activation Types (a–c) across the three active avoidance modes.
Fac1: Active Modes (cl1=Mode3, cl2=Mode2, cl3=Mode1)
Fac2: Type (a1=Type1, a2=Type2, a3=Type3)
Subj: Animal
Fig5-Sup1 A-B A, k-means clustering of movement speed time-series from CS onset revealed three distinct active avoidance modes. Mode 1 avoids were initiated earlier after CS onset, whereas Mode 2 and Mode 3 avoids were delayed and faster, reflecting increasingly cautious responding. B, Same as A, aligned from-action (avoid) occurrence.
File: filename
MPC1: Tone intensities - Tone frequency
Fig5-Sup1 C Marginal means (ΔF/F) from the linear mixed-effect models for the baseline, orienting, avoidance, and from-action windows for the three active avoidance Modes.
Fac1: Active Modes (cl1=Mode3, cl2=Mode2, cl3=Mode1)
Subj: Animal
Figure 6. Optogenetic inhibition of mPFC neurons has little effect on signaled avoidance.
Fig6 A-B A, Effects of continuous green light at three different powers (10-30 mW) on learning AA1 (black circles) followed by AA2 (red circles) in naïve mice expressing eArch3.0 in mPFC neurons. In AA1, the CS signals the active avoidance contingency. In AA2, ITCs are punished. On every trial, light was delivered during the CS and US to inhibit mPFC activity. During AA2, performance remained unchanged, although mice exhibited the typical shift toward longer avoidance latencies. Catch trials (NoCs) consisted of blank trials without light or consequence for crossing. B, Same as A, but during subsequent AA3 task, in which CS1, CS1+Light, CS2, and CS2+Light trials were presented randomly.
CondA: Trial Condition (AA16=AA1, AA916=AA2, AA=AA3-CS1(AA), PA=AA3-CS2(PA))
CondB: Light intensity (i0=No-Light, i15=10mW, i25=20mW, i35=30mW)
Ses+Subses: Session+Subsession
Avoids%: percentage of avoids
Avoid Latency: latency of avoid responses
ITCs/trial: number of intertrial crossings per trial
Fig6 C Example PSTH shows the effect of blue light in Vgat-Cre mice expressing ChR2 in GABAergic neurons recorded in freely behaving mice with an optrode implanted in mPFC.
getFile: filename
Fig6 D Effects of continuous blue light in mPFC on learning AA1 followed by AA2 in naïve mice expressing ChR2 in GABAergic neurons. The data represent groups of mice with optical cannulas in mPFC or VI, with a third No Opsin group with optical fibers in mPFC that did not express ChR2. Otherwise, the same as Fig6A.
CondA: group (PFC, VI, NOP)
CondB: Trial Condition (AAL=CS+Light, NoCS)
CondC: Task (L16=AA1, L916=AA2)
Fig6 E Effect of blue light in the same three groups during AA3. Otherwise, same as Fig6A.
CondA: group (PFC, VI, NOP)
CondB: Trial Condition (AA=AA3-CS1(AA), PA=AA3-CS2(PA))
CondC: Light Condition, (L=AA3-Light, NL=AA3-NoLight)
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