Kv7.4 Channel Contribute to Projection-Specific Auto-Inhibition of Dopamine Neurons in the Ventral Tegmental Area

Figures

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  Figure 1. Kv7.4 contribute to the dopamine (DA)-induced inhibition of NAc-projecting DA neuron firing. (A) Single-cell polymerase chain reaction (PCR) analysis in retrogradely labeled ventral tegmental area (VTA) DA neurons from different projections. (B–D) VTA DA neuron firing recorded with loose cell-attached patch recordings. XE991 (3 μM), a Kv7 blocker, reversed the DA-induced inhibition of neuron firing in WT (B) and Kir3.2−/− (D) mice, but not in Kv7.4−/− mice (C). (E) The effect of DA on spontaneous firing in DA neurons from Kv7.4−/−/Kir3.2−/− mice. (F) The percentage of VTA-NAc DA neurons with firing rate being inhibited to less than 20% (complete inhibition) in WT, Kv7.4−/−, Kir3.2−/− and Kv7.4−/−/Kir3.2−/− mice. (G) Average DA inhibition on firing rate in all recorded VTA-NAc DA neurons from WT, Kv7.4−/−, Kir3.2−/− and Kv7.4−/−/Kir3.2−/− mice. One-way repeated-measures ANOVA with Bonferroni post hoc test. ***p < 0.001, **p < 0.01, *p < 0.05; N.S., not significant. One-way repeated-measures ANOVA with Bonferroni post hoc test. N, number of animals; n, number of recordings.

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  Figure 2. Dopamine activates Kv7/M current in NAc-projecting VTA DA neurons. (A) Kv7/M currents recorded from DA neurons of WT (i,ii) and Kv7.4−/− (iii,iv) mice. Shown are time-courses of M current amplitudes recorded using whole-cell patch-clamp from VTA slices (for details, see “Materials and Methods” section). Summarized data are presented in (B). Error bars represent SEM. ***p < 0.001, *p < 0.05; N.S., not significant. One-way repeated-measures ANOVA with Bonferroni test for WT mice, paired t-test for Kv7.4−/− mice. N = number of animals; n, number of recordings.

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  Figure 3. Dopamine activates Kv7/M in VTA DA neurons through dopamine D2 receptor and related cell signaling. (Ai,ii). Example of time-course of Kv7/M currents activated by DA in the presence of D2 receptor inhibitor sulpiride (300 nM) and D1 receptor inhibitor SCH23390 (1 μM). (Aiii) Summarized effects of D2 receptor agonist quinpirole (300 nM), sulpiride (300 nM), SCH23390 (1 μM) and Kv7/M channel inhibitor XE991 (3 μM) on DA-induced activation of Kv7/M currents in VTA DA neurons. ***p < 0.001, **p < 0.01, *p < 0.05; N.S., not significant, paired t-test. N = number of animals; n = number of recordings. (B) Effect of sulpiride (300 nM) and SCH23390 (1 μM; i,ii) on the resting membrane potential of VTA DA neurons. Summarized effects of sulpiride and SCH23390 on DA-induced hyperpolarization of VTA DA neurons (iii). *p < 0.05; N.S., not significant, One-way repeated-measures ANOVA with Bonferroni test. (C) Example traces from cell-attached recordings of VTA DA neurons activated by DA in the presence of sulpiride and SCH23390 are shown in (Ci). (Cii) Summarized effects of DA, sulpiride and SCH23390 on DA-induced inhibition of the spontaneous firing in VTA DA neurons. *p < 0.05; N.S., not…

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  Figure 4. DA-induced inhibition on the firing of NAc-projecting VTA DA neurons is reduced in the social defeat model of mice. (A,B) Example traces from cell-attached recordings of NAc-projecting VTA DA neurons are shown before and after administration of fasudil (a selective Kv7.4 opener; 10 μM) and DA (20 μM) in the control and the social defeat model mice. (C) Summarized the effect of fasudil and DA on the spontaneous firing frequency of NAc-projecting VTA DA neurons. **p < 0.01, *p < 0.05; N.S., not significant, paired t-test. N = number of animals; n = number of recordings. (D) Normalized firing frequency based on the data shown in C. Error bars represent SEM. **p < 0.01, *p < 0.05; N.S., not significant, one-sample Wilcoxon signed ranks test.