(a) Left, targeting scheme for hM4Di-mCherry. Right, localization of hM4Di-mCherry. 3V, third ventricle; Arc, arcuate nucleus; D, dorsal; L, lateral; LH, lateral hypothalamus; M, medial; V: ventral. Inset, hM4Di-mCherry in Agrp cells at higher zoom. Scale bar, 500 μm. Representative example from four brains. (b) Effect of 5μM CNO on Agrp-hM4Di cell firing. Representative example of five cells. (c) Left, group data of membrane potential effect in experiment shown in b (n=5 cells). P value is from a paired t-test, t=6.689, d.f.=4. Right, Spike firing responses in experiment shown in b for all cells. (d) Left, effects of EtOH (2 g kg−1 i.p.) and CNO (5 mg kg−1 i.p.) on food intake of Agrp-Cre mice expressing hM4Di or ChR2 in Agrp cells. Food intake under each drug condition represents a mean of 3 days of drug treatment (same design as in Fig. 1a). Four male mice per group (age/gender-matched littermate pairs). Two-way repeated measures analysis of variance: treatment F(3, 18)=129.9, P<0.0001; genotype F(1, 6)=45.44, P=0.0005; interaction F(3, 18)=53.28, P<0.0001; numbers between bars are P values from Tuckey’s multiple comparison corrections. Right, Agrp cell-driven food intake (defined as reduction in food intake evoked by CNO in Agrp-hM4Di mice) in the presence and absence of EtOH (n=8 mice). Number above bars is P value from a two-tailed paired t-test, t=5.055, d.f.=7. Values shown are individual points and/or means±s.e.m.