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. Author manuscript; available in PMC: 2017 Dec 19.
Published in final edited form as: Nat Neurosci. 2017 Jun 19;20(8):1096–1103. doi: 10.1038/nn.4590

Figure 5.

Figure 5

Targeting Cadm1 expression in the hippocampus and hypothalamus reduces body weight. (a) Representative coronal brain section of Cadm1 floxed mice after stereotaxic rAAV8-CaMKIIa-mCherry-Cre (AAV-Cre) injection in the dentate gyrus (DG) and CA3 regions to target Cadm1 expression in hippocampal neurons. (b) Co-localization of mCherry with NeuN and Vglut2 after delivery of AAV- CaMKIIa-Cre to the hippocampus. (c) Body weight curves of floxed Cadm1 mice (flox/flox) (n=6) and control littermates (+/?) (n=6) after delivery of AAV-Cre to the hippocampus. (d) Energy expenditure from individual animals plotted against lean body mass in 9-week old floxed Cadm1 mice (flox/flox) (n=7) and control littermates (+/?) (n=6) after delivery of AAV-Cre to the hippocampus. (e) Energy expenditure from individual animals plotted against activity measured in 9-week old floxed Cadm1 mice (flox/flox) (n=7) and control littermates (+/?) (n=6) after delivery of AAV-Cre to the hippocampus. (f) Quantification of food intake measured in floxed Cadm1 mice (flox/flox) (n=7) and control littermates (+/?) (n=6) after delivery of AAV-Cre to the hippocampus. (g) Stereotactic delivery of a mCherry-expressing AAV to paraventricular nucleus (PVH) (red) reveals PVH efferent projections to eGFP-positive POMC neurons (green) within the Arc region of the hypothalamus. Dotted white circle outlines the cell body of an eGFP-POMC-positive cell, used for 3D reconstruction. (h) Surface rendering of Amira 3D reconstruction of an eGFP-POMC-positive cell body receiving afferent input from the PVH. Cell is represented in red, while the synaptic input is color-coded, with the cold to warm colors spreading from 0 to 250nm distance between axonal varicosities and the soma (see color-coded horizontal bar for the distance definition). (i) Histograms show the number of anterograde AAV-mCherry-labelled PVH axonal varicosities found within the distance of 250nm from the POMC-positive cell body. (j) Double immunostaining of Cadm1 and mCherry or synaptophysin (Syp) indicates that PVH regions provide Cadm1-positive efferent projections to the ARC region of hypothalamus. POMC-eGFP transgenic mice received an AAV-mCherry injection to the PVH, and were subsequently immunostained for Cadm1 (green) and mCherry (red) or synaptophysin (magenta). (k) Representative coronal brain section of Cadm1 floxed mice after stereotaxic injection of AAV8-CaMKIIa-mCherry-Cre to hypothalamic neurons in the PVH, and ventromedial hypothalamus (VMH) regions to drive Cre expression to excitatory neurons in these regions. Section is showing mCherry expression and staining for vasopressin (green) and nuclei counterstained with DAPI (blue). (l) Confocal imaging of mCherry (red) and vasopressin (green). (m) Body weight curves of floxed Cadm1 mice (flox/flox) (n=9) and control littermates (+/?) (n=11) after delivery of AAV-Cre to the hypothalamus. (n) Quantification of energy expenditure (EE) adjusted to lean body mass (LBM) in 10-week old floxed Cadm1 mice (flox/flox) (n=10) and control littermates (+/?) (n=9) after delivery of AAV-Cre to the hypothalamus. (o) Quantification of energy expenditure (EE) adjusted to locomotor activity in 10-week old floxed Cadm1 mice (flox/flox) (n=10) and control littermates (+/?) (n=9) after delivery of AAV-Cre to the hypothalamus. (p) Quantification of food intake measured in floxed Cadm1 mice (flox/flox) (n=10) and control littermates (+/?) (n=9) after delivery of AAV-Cre to the hypothalamus. All results are presented as mean ± s.e.m. *P<0.05 and **P<0.01. Boxplots show median, lower and upper quartiles (box), and minimum and maximum (whiskers). Statistical analyses are described in the Methods and Supplementary Table 3.