Fig. 3: Social stress potentiates LHAglu-VTADA synapses via AMPA receptor modifications.
a Timeline and schematic for electrophysiological patch clamp experiments. b Representative image of an LHA slice with ChR2-mCherry in red. Scale bar: 100 µm. Anatomical reference point: f fornix. Right: Image of green fluorescent VTADA neurons (Pitx3-GFP) with patch clamp electrode. Scale bar: 10 µm. c AMPAR-NMDAR ratios at LHAglu-VTADA synapses increase post stress. Left: representative I-AMPAR and I-NMDAR traces, Blue rectangle: start of LED stimulation. Scale bar: 20 ms, 20 pA. I-AMPAR measured at −65 mV, I-NMDAR at +40 mV at 100 ms from stimulation onset (black arrows). Right quantification of AMPAR/NMDARs +SEMs (n cells control = 15, n cells stress = 10, One-Way ANOVA, F(1,23) = 14.72, p = 0.001). d AMPAR Rectification index increases at LHAglu-VTADA synapses after stress. Left: example traces, scale bars: 20 ms, 10 pA. Middle: AMPAR current at −60 mV, 0 mV and +40 mV normalized to AMPAR current at −60 mV (n cells control = 18, n cells stress = 17). Right: Averages + SEMs (n cells control = 18, n cells stress = 17, One-Way ANOVA, F(1,33) = 5.81, p = 0.02). e Paired pulse ratio of LHAglu-VTADA neurons does not differ due to stress. Left: example traces. Scale bars: 25 pA, 25 ms. Right: Averages + SEMs (n cells = 31 per group, One-Way ANOVA, F(1,60) = 0.02, p = 0.89). f Left: Schematic for Array Tomography experiment. Right: Representative image of VTA slice stained for LHA fibers (YFP: green), DA neurons (TH: blue) and glutamatergic fibers (Vglut2: red). White arrows: puncta overlap between YFP, Vglut2 and TH. Yellow arrows: puncta with overlap between YFP and Vglut2. Scale bar: 10 µm. g Representative images of reconstructed VTA slices with array tomography. On the left side the white arrows indicate overlap between Vglut2+ glutamatergic terminals (red) with GluA1 subunits (blue). On the right side the same slice is shown, where the white arrows indicate overlap between LHA terminals (YFP) and dopamine neurons (TH: blue). In both cases DAPI-stained nuclei are shown (white). The top row shows an example for a control mouse, and the bottom row for a stressed mouse. Scale bar: 5 µm. h Quantification of overlap of LHA terminals, synapsin, GluA1 AMPAR subunits and dopamine neurons, for Control and Stress conditions (averages + SEMs, mice per group n = 4, One-Way ANOVA, F(1,6) = 18.26, p = 0.005). i As h but for the overlap of LHA terminals, Vglut2, GluA1-AMPAR subunits and dopamine neurons for Control and Stress conditions (averages + SEMs, mice per group n = 4, One-Way ANOVA, F(1,6) = 14.9, p = 0.008). All statistical tests were performed two-sided *p < 0.05, **p < 0.01. Source data are provided as a Source Data file.
