Fig. 4.

EtOH markedly enhanced CIN firing rate with block by α-conotoxins or knockdown of α6*-nAChRs. A Illustration showing the experimental framework wherein AAV-flex-mRuby2-α6-miRNA was injected into the VTA 3 weeks prior and NAc CINs were recorded in cell-attached, voltage-clamp mode in VGAT-Cre/GAD67-GFP mice. B Immunohistochemical panel showing that CINs in VGAT-Cre/GAD67-GFP mice are innervated by VTA-NAc GABAergic projections expressing α6*-nAChRs. Imaged using oil immersion 40 × objective (Olympus, UPlanFLN 1.30 numerical aperture). C CINs visualized in ChAT mice were characterized by autoreceptor inhibition by the M2 agonist muscarine. Insets are 5-s representative spike recordings before (a), during (b), and after (c) muscarine (wash). Times for (a, b, c) are indicated on the representative ratemeter below. D In putative CINs inhibited by muscarine in VGAT-Cre mice, firing rate is enhanced by increasing doses of EtOH. E Block of EtOH enhancement of CIN firing rate at high doses by MII. F Block of EtOH enhancement of CIN firing rate in mice injected with AAV-flex-mRuby2-α6-miRNA into the VTA. G Summary of effects of α6-miRNA or MII on CIN baseline firing rate. H Summary of EtOH effects in mice treated with MII or α6-miRNA. Values in parentheses are n values. Asterisks *, **, and *** indicate significance levels p < 0.05, p < 0.01, and p < 0.001, respectively