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. Author manuscript; available in PMC: 2023 Mar 16.
Published in final edited form as: Neuron. 2022 Jan 18;110(6):1068–1083.e5. doi: 10.1016/j.neuron.2021.12.027

Figure 5. BLA inputs recruit SST-INs to shunt MDT-PFC transmission.

Figure 5.

(A) ChR2 was expressed in the BLA and electrical (el)-EPSCs were evoked by stimulating distal synapses in Layer 1 (L1). (B) Top, Representative traces depicting BLA op-EPSCs and L1 el-EPSCs evoked in the same pyramidal cell. Bottom, Recent prior stimulation of BLA terminals inhibited the amplitude of subsequent el-EPSCs (10ms ISI, left; 30ms ISI, right). Scale bars 100pA, 10ms. (C) At short pre-pulse interstimulus intervals (ISIs), BLA terminal stimulation inhibits the amplitude of L1 el-EPSCs. Including the GABAA receptor antagonist picrotoxin in the patch pipette blocked the effect at the 10-ms disynaptic ISI. (RM Two-way ANOVA ISI x picrotoxin interaction: F4,60=4.4, p<0.01; ***:p<0.001, Sidak test, n/N=8-9/4 Cells/mice). (D) Drugs acutely restricted by tethering (DART) was deployed by viral-mediated expression of the HaloTag protein (HT+), or an inactive mutant variant (HT−), selectively in PFC SST-INs. Both constructs also expressed tdTomato to allow for visualization and cellular targeting. (E) The AMPA receptor antagonist YM90K DART dose-dependently inhibited EPSCs on SST-INs following expression of HT+ but not HT−. (RM Two-way ANOVA YM90K x HaloTag interaction: F2,14=7.2, p<0.01; main effect of YM90K: F1,7=11.4, p<0.05; 56±9 vs 109±12% (1μM), *:p<0.05, Sidak test). n/N=4-5/3-4. (F) Following YM90K DART washout, HT+ SST-INs displayed decreased sEPSC amplitude. (Two-way ANOVA YM90K x HaloTag interaction: F1,37=2.9, p<0.1; *:p<0.04, Sidak test). n/N=9-13/3-4. Scale bars 5pA, 2ms. (G) HT+ was expressed in PFC SST-INs and ChR2 was expressed in the BLA to evaluate heterosynaptic inhibition of L1 terminals. (H) Representative recordings showing isolated (top) and summated (bottom) EPSCs following YM-90K DART application. (10ms ISI, left; 30ms ISI, right). Scale bars 100pA, 10ms. (I) SST-IN-directed YM90K-DART blocked BLA-driven heterosynaptic inhibition of EPSCs evoked by L1 stimulation. (RM Two-way ANOVA ISI x YM90K interaction: F4,36=3.1, p<0.03; ***:p<0.001; $:p<0.1, Sidak test). n/N=5-6/3. (J) The red-shifted opsin Chrimson was expressed in the BLA and ChR2 was expressed in the MDT. Red light stimulation preceded blue light stimulation to mitigate concerns related to spectral overlap (see Figure S5). (K) At short pre-pulse ISIs, BLA stimulation inhibited MDT op-EPSCs in control conditions but not if picrotoxin was included in the patch pipette. (Two-way ANOVA picrotoxin x ISI interaction: F4,72=5.7, p<0.001; **:p<0.01, Sidak test). n/N=5-16/2-5. (L) A single exposure to 20-minute restraint stress disrupted disynaptic inhibition from the BLA to MDT input to PFC. (RM Two-way ANOVA stress x ISI: F4,92=2.5, p<0.05; *:p<0.04, Sidak test). n/N=10-16/3-5.