Extended Data Fig. 5: Optogenetic manipulation of DA neurons in the VTA.
From: Dopamine D2 receptors in discrimination learning and spine enlargement
a, Schematic of experimental setup. b, Representative confocal images of virus expression and fibre position. The locations of fibre tips were plotted on the reference atlas. TH, tyrosine hydroxylase immunostaining. White bars indicate traces of optical fibres. Grey areas in the atlas indicate the NAc core. c, Photometry of DA neurons stimulated with red LED. Top, schema for stimulus presentation; middle, photometry responses at NAc; bottom, plots of peak photometry responses. Friedman’s test for mCherry (n = 8 mice, χ2(3) = 15.8, P = 0.001; post hoc two-sided Wilcoxon signed rank test with Bonferroni correction, no stimulation versus 5 Hz, W = 10, P = 0.26; no stimulation versus 20 Hz, W = 17, P = 0.89; no stimulation versus sucrose, W = 0, **P = 0.008) and for CsChrimsonR (n = 7 mice, χ2(3) = 18.9, P = 0.0003; post hoc two-sided Wilcoxon signed rank test with Bonferroni correction, no stimulation versus 5 Hz, W = 0, P = 0.093; no stimulation versus 20 Hz, W = 0, *P = 0.016; no stimulation versus sucrose, W = 0, *P = 0.016). d, Schematic of reinforcement of licking behaviour by optogenetic self-stimulation of DA neurons. Licking behaviour triggered optogenetic self-stimulation of DA neurons in the VTA without water delivery. e, Cumulative lick number against time (top) and plots across the conditions (bottom). Friedman’s test for mCherry (n = 8 mice, χ2(2) = 1, P = 0.60) and for CsChrimson (n = 7 mice, χ2(2) = 11.1, P = 0.004; post hoc two-sided Wilcoxon signed rank test with Bonferroni correction, no stimulation versus 5 Hz, W = 8, P = 0.31; no stimulation versus 20 Hz in CsChrimsonR, W = 0, *P = 0.016). f, Schematic of experimental setup for optogenetic enhancement of DA dips during discrimination. AAVs were injected to express eNpHR3.0 and GCaMP6f in VTA-DA neurons. Optical fibres were implanted at the VTA for stimulation of eNpHR3.0 and at the right NAc for photometry of GCaMP. g, Locations of the fibre tips plotted on the reference atlas. h, Averaged z-scores of photometry responses during CS− trials in the discrimination period for eNpHR3.0 (red) and mCherry (blue). i, Lick scores plotted against time. Red bar indicates the period of optogenetic stimulation. n = 6 (h, i, mCherry) and 5 (h, i, eNpHR3.0) mice. Light green bar indicates sampling periods for the fibre photometry data shown in h. j, Locations of the tips of fibres plotted on the reference atlas. k, Averaged z-scores of photometry responses with 20-Hz optogenetic stimulation during extinction. l, Lick scores plotted against time with 20-Hz DA stimulation during extinction. Red bar indicates the period of optogenetic stimulation. Green bar indicates sampling periods for the fibre photometry data shown in k. m, Averaged lick traces during CT and extinction test (top) and lick score at CT and extinction test (bottom). n = 5 mice (k–m). n, Plots of extinction indices across conditions. n = 6 mice for mCherry 5 Hz, n = 4 mice for CsChrimsonR 5 Hz, n = 4 mice for CsChrimsonR 20 Hz. Kruskal–Wallis test (χ2(2) = 9.5, P = 0.009; post hoc Steel’s test, CsChrimsonR 5 Hz versus mCherry, t = 1.28, P = 0.34; CsChrimsonR 20 Hz versus mCherry, t = −2.6, *P = 0.02). Data for CsChrimsionR 5 Hz and mCherry 5 Hz are from Fig. 2m. Scale bars in b indicate 500 μm. Scale bars in h, k indicate 1 s and 1 z-score. Scale bars in m indicate 1 s and 2 Hz. Error bars and shading indicate s.e.m. Vertical grey shading in h, k indicates CS periods. Samples of mice are biological replicates.
