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. 2015 Jul;40(8):1969-78.
doi: 10.1038/npp.2015.47. Epub 2015 Feb 11.

Septal Glucagon-Like Peptide 1 Receptor Expression Determines Suppression of Cocaine-Induced Behavior

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Free PMC article

Septal Glucagon-Like Peptide 1 Receptor Expression Determines Suppression of Cocaine-Induced Behavior

Anne E Harasta et al. Neuropsychopharmacology. .
Free PMC article

Abstract

Glucagon-like peptide 1 (GLP-1) and its receptor GLP-1R are a key component of the satiety signaling system, and long-acting GLP-1 analogs have been approved for the treatment of type-2 diabetes mellitus. Previous reports demonstrate that GLP-1 regulates glucose homeostasis alongside the rewarding effects of food. Both palatable food and illicit drugs activate brain reward circuitries, and pharmacological studies suggest that central nervous system GLP-1 signaling holds potential for the treatment of addiction. However, the role of endogenous GLP-1 in the attenuation of reward-oriented behavior, and the essential domains of the mesolimbic system mediating these beneficial effects, are largely unknown. We hypothesized that the central regions of highest Glp-1r gene activity are essential in mediating responses to drugs of abuse. Here, we show that Glp-1r-deficient (Glp-1r(-/-)) mice have greatly augmented cocaine-induced locomotor responses and enhanced conditional place preference compared with wild-type (Glp-1r(+/+)) controls. Employing mRNA in situ hybridization we located peak Glp-1r mRNA expression in GABAergic neurons of the dorsal lateral septum, an anatomical site with a crucial function in reward perception. Whole-cell patch-clamp recordings of dorsal lateral septum neurons revealed that genetic Glp-1r ablation leads to increased excitability of these cells. Viral vector-mediated Glp-1r gene delivery to the dorsal lateral septum of Glp-1r(-/-) animals reduced cocaine-induced locomotion and conditional place preference to wild-type levels. This site-specific genetic complementation did not affect the anxiogenic phenotype observed in Glp-1r(-/-) controls. These data reveal a novel role of GLP-1R in dorsal lateral septum function driving behavioral responses to cocaine.

Figures

Figure 1
Figure 1
Glp-1r mRNA is enriched in the lateral septum. (a) Illustration of a coronal mouse brain section. (a1, a2) Glp-1r mRNA-expressing cells (purple) in the medial amygdaloid nucleus (red box in (a)) or the hypothalamus (blue box in (a)) labeled by arrowheads. (a3, a4) High magnification of individual Glp-1r mRNA-positive cells. (b) Overview of Glp-1r mRNA expression (black staining) domains in the sagittal plane (Allen Brain Atlas ID 73606497_125). (b1, b2) Close-ups of the NAc and the VTA showing little Glp-1r expression. (c) In the coronal plane, highest Glp-1r expression (silver impregnation) was identified in the LS (Allen Brain Atlas ID 74511737_345). (d) The Glp-1r sense probe produced no signal. (e) Abundant Glp-1r mRNA expression (pink) was confirmed in the dLS using a specific DIG-labeled riboprobe. (e1) Higher magnification of the dLS indicated by the box in (e) depicting strong cellular signal detection of Glp-1r mRNA. (f) Fluorescent detection of Glp-1r (red) and (g) Gad65 (green) mRNA in the dLS. (h) Nuclear counterstaining with DAPI (blue). (i) Merged picture reveals that almost all Glp-1r-positive septal neurons are GABAergic. Bars: (a1, a2, d, e) 75 μm; (a3, a4, e, f) 10 μm; (b) 1 mm; (b1, b2) 100 μm; (c) 200 μm. dLS, dorsal lateral septum; DMH, dorsomedial hypothalamic nucleus; Hippo, hippocampus; LV, lateral ventricle; MeA, medial amygdaloid nucleus; NAc, nucleus accumbens; opt, optic tract; PG, pontine gray; VTA, ventral tegmental area.
Figure 2
Figure 2
Increased firing of Glp-1r−/− septal neurons. (a) The time to fire an AP is delayed in Glp-1r−/− (total of 23 cells from 10 animals) compared with Glp-1r+/+ (total of 21 cells from 11 animals) neurons (P=0.02). (b) The number of APs at twice threshold was elevated in Glp-1r−/− compared with Glp-1r+/+ controls (P=0.04). (c) Representative responses of Glp-1r+/+ (left) and Glp-1r−/− (right) neuron, respectively, to 600 ms depolarizing current steps at 100 and 150 pA. (d) Responses (number of APs evoked by 600 ms stimulus) of Glp-1r+/+ and Glp-1r−/− cells across a range of step current injections from 10 to 150 pA revealed an increased activity of Glp-1r−/− neurons (two-way repeated measures ANOVA × genotype effect: F(1, 43)=6.48, P=0.01). *P<0.05, **P<0.01.
Figure 3
Figure 3
AAV-mediated Glp-1r gene delivery to the dLS. (a) AAV expression cassettes. ITR, inverted terminal repeat; CBA, chicken β-actin promoter; pA, polyadenylation signal; WPRE, woodchuck hepatitis virus posttranscriptional regulatory element. (b) Biological activity of GLP-1R-GFP was confirmed by expression in heterologous HEK293 cells and exposure to Ex4 (10 nM) for the indicated durations (0–50 min). The dynamics of ERK phosphorylation were similar following activation of the tagged GLP-1R or the native receptor. Total ERK served as loading control. Note that the GFP antibody exclusively recognized the tagged recombinant receptor. (c) Recombinant GLP-1R-GFP traffics to the cell surface in HEK293 cells and in the mouse LS (top row), whereas GFP controls show cytosolic expression (bottom row). Anti-GFP immunofluorescence (green); DAPI (blue) nuclear counterstain. Bars: 10 μm. (d) Schematic illustration of vector spread assessed by AAV-transduced neuronal somata; cc, corpus callosum; dLS, dorsal lateral septum; LV, lateral ventricle.
Figure 4
Figure 4
Genetic complementation of GLP-1R expression in the dLS rescues cocaine-induced, but not anxiety-related, behaviors in Glp-1r-deficient mice. (a) Glp-1r−/− mice and Glp-1r−/− mice injected with AAV-GLP-1R-GFP spent less time in the center of the EPM (left panel, one-way ANOVA F=8.049; P=0.0012); Glp-1r+/+ controls (n=17) compared with Glp-1r−/− mice (n=15; P<0.05), or Glp-1r+/+ controls compared with Glp-1r−/−+AAV-GLP-1R-GFP (n=9; P<0.01). Middle panel: Glp-1r−/− mice and Glp-1r−/− mice injected with AAV-GLP-1R-GFP spent less time in the open arms (one-way ANOVA F=14.34; P<0.0001) compared with Glp-1r+/+ controls (P<0.0001) and entered the open arms less frequently (right panel; one-way ANOVA F=9.65; P<0.001; post hoc test P<0.01). (b) Altered locomotor response to cocaine for all groups over the course of the paradigm (two-way repeated measures ANOVA genotype effect F(2, 41)=9.771, P=0.0003). There was no difference between genetically complemented Glp-1r−/− mice (n=9) and Glp-1r+/+ controls (n=17) following acute cocaine exposure (day 1; P>0.9999). Intact development and expression of cocaine sensitization (two-way repeated measures ANOVA cocaine effect F(2, 82)=21.00, P<0.0001) was present in Glp-1r+/+ and Glp-1r−/− mice (n=15) but absent in Glp-1r−/−+AAV-GLP-1R-GFP mice (day 1–day 5, P=0.5141 and day 1–day 12, P=0.2919). Note that there was no difference between Glp-1r+/+ controls and Glp-1r−/−+AAV-GLP-1R-GFP on any given day. #P compared with day 1; *P compared with all groups on a given day. (c) Genetically complemented Glp-1r−/− mice displayed a normal CPP phenotype (one-way ANOVA F=7.037, P=0.0023. The CPP score represents the difference between the time spent in the cocaine compartment before and following conditioning. * or #P<0.05, ** or ##P<0.01, ***P<0.001.
Figure 5
Figure 5
Septal GLP-1R regulates motivated behavior. GLP-1R-expressing GABAergic septal neurons modulate cellular activity following stimulation. GLP-1Rs are present in both somata and terminals of these neurons (Göke et al, 1995; Shirazi et al, 2013) that form inhibitory synapses with GABAergic interneurons in the VTA (Luo et al, 2011). Their inhibition, in turn, leads to activation of dopaminergic VTA neurons via disinhibition. Hence, this dLS–VTA pathway controls drug-induced behaviors such as locomotor sensitization and drug seeking. DA, dopamine. Adapted from Luo et al (2011).

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