Hierarchical glucocorticoid-endocannabinoid interplay regulates the activation of the nucleus accumbens by insulin
Graphical abstract
Introduction
Insulin signaling in the brain is crucial for both central and systemic metabolic homeostasis (Kleinridders et al., 2014, Kullmann et al., 2015). Research in humans and animals concluded that insulin influences glucose regulation in the forebrain (Duarte et al., 2012). For instance, [18F]FDG-PET studies documented that global cerebral glucose uptake rates can be stimulated by acute hyperinsulinemia only in patients with impaired glucose tolerance (Hirvonen et al., 2011), unless endogenous insulin production is suppressed in the healthy control group (Bingham et al., 2002). These findings suggest that even under normoinsulinemia, brain insulin levels are high enough to mask certain central effects of exogenous insulin. Notably though, mapping local metabolic rates in different brain areas, both increased and decreased glucose uptake have been reported in response to an acute bolus of insulin (Anthony et al., 2006). Local metabolic rates do not necessarily match changes in global metabolic rates: insulin for instance stimulates glucose uptake on average by 27% in the ventral striatum of healthy humans, and only by 13% in insulin resistant patients (Anthony et al., 2006).
The ventral striatum is called the nucleus accumbens in non-primate mammals, and is a major relay nucleus of the reward circuitry (Kenny, 2011). Insulin signals converge here that underlie palatable food preference and food reward (Volkow et al., 2008, Kenny, 2011, Kullmann et al., 2015). The modulation of mesoaccumbal transmitter release and synaptic plasticity is also crucial in drug-induced reward and reinforcement of drug abuse − including that of cannabis—which are highly dependent on frontal cortical-mesoaccumbal endocannabinoid signaling (Robbe et al., 2003, Maldonado et al., 2013, Covey et al., 2014). Consequently, food and drug addiction share common mechanisms (Volkow et al., 2008, Kenny, 2011), and insulin resistance is associated with the physical and functional deterioration of the nucleus accumbens, which exacerbates addictive behaviour including the increased intake of caloric and palatable food (Isganaitis and Lustig, 2005, Chen et al., 2013, O’Dell et al., 2014).
Various molecular mechanisms can hinder insulin signaling at insulin receptors (InsRs). For example, glucocorticoids are well known to cause insulin resistance and obesity (Andrews and Walker, 1999). Strikingly, a recent study revealed that glucocorticoids can trigger the metabolic syndrome by engaging endocannabinoid signaling in the periphery (Bowles et al., 2015). Glucocorticoid receptor (GcR) activation has also been reported to stimulate endocannabinoid synthesis at central synapses (Di et al., 2003, Hill and McEwen, 2009, Hill et al., 2011). The metabotropic CB1 cannabinoid receptor (CB1R) is the principal cognate receptor in the brain of the two most-studied endocannabinoids, anandamide and 2-AG (Katona and Freund, 2012, Murataeva et al., 2014). The CB1R is present in the nucleus accumbens (Mátyás et al., 2007, Pickel et al., 2006, Winters et al., 2012). Remarkably, CB1Rs form functional and physical heteromers with the β-chain of the InsRs in both pancreatic islets and neuronal cell lines (Dalton and Howlett, 2012, Kim et al., 2012), and cannabimimetics prevent insulin-induced autophosphorylation of the InsR β-chain and the consequent activation of the PI3K-Akt pathway.
Here we sought answers to two main questions, namely whether insulin also stimulates glucose uptake in the accumbal slice as it does in vivo (Anthony et al., 2006), and if the effect of insulin is influenced by glucocorticoid and endocannabinoid signaling. This would also support the use of in vitro [3H]DG assays to study basic mechanisms of food intake regulation, providing a viable alternative to in vivo [18F]FDG-PET studies.
Section snippets
Ethics statement and animals
All studies were conducted in accordance with the principles and procedures outlined as “3Rs” in the guidelines of EU (86/609/EEC), FELASA, and the National Centre for the 3Rs (the ARRIVE; Kilkenny et al., 2010). Studies were approved by the Animal Care Committee of the Center for Neuroscience and Cell Biology of the University of Coimbra, Portugal. We also applied the ARRIVE guideline for the design and execution of in vitro pharmacological experiments (see below), as well as for data
Insulin receptor (InsR) activation stimulates glucose uptake in the accumbal slices
The first 30 min of deoxyglucose uptake in the brain displays linear kinetics (Schmidt et al., 1989, Lemos et al., 2012). The mean glucose uptake value, as calculated from specific [3H]DG uptake, amounted to 68.4 ± 1.9 nmol/mg protein in the 30 min uptake period in the accumbal slices (n = 51 rats, i.e. pooled from all control animals throughout the study). One-min pretreatment with insulin at 3 nM (n = 7) and at 300 nM (n = 20), but not at 0.3 nM (n = 10) stimulated [3H]DG uptake in the range of 20–24% (P <
Discussion
The novel findings in this study indicate that insulin stimulates glucose uptake in the rat accumbal slice in vitro via insulin receptor (InsR) activation. Certainly, this is consistent with an earlier in vivo [18F]FDG-PET study in humans (Anthony et al., 2006), thus documenting that studying insulin action in rat accumbal slices is a viable alternative approach that may be useful for dissecting mechanisms. Furthermore, glucocorticoid receptor (GcR) activation triggers the release of the
Conflict of interest
None.
List of author contribution
Study design: AK. Neurochemical assays: BSP, CL, FNK, CSS, AK, EC. Immunoprecipitation: JMM, RJR, KM, BSP, CSS. Financing: AK, RAC. First draft: AK. Final form of the manuscript: all authors.
Acknowledgments
This work was supported by grants from the Fundação para a Ciência e a Tecnologia to A.K. (PTDC/SAU-OSM/105663/2008 and PTDC/SAU-NEU/100729/2008), to R.A.C. (PTDC/SAU-NSC/122254/2010), to E.C. (PTDC/SAU-OSM/104124/2008) and to R.J.R. (EXPL/NEU-NMC/0671/2012); as well as from DARPA to R.A.C. (W911NF-10-1-0059). This work was also supported by Programa Mais Centro under project CENTRO-07-ST24-FEDER-002006, COMPETE, PEst-C/SAU/LA0001/2013-2014, and the strategic project UID/NEU/04539/2013 as well
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2018, Brain Research BulletinCitation Excerpt :The authors suggest that 2-AG binds to the CB1R-InsR (insulin receptor) heteromer and inhibits the glucose uptake induced by insulin. These findings also indicate the existence of a possible interaction between CB1R and GcR, i.e. a glucocorticoid receptor (Pinheiro et al., 2016). Additionally, the levels of both endocannabinoid 2-AG and glucocorticoid cortisol fluctuate according to circadian rhythms, showing the highest serum concentrations in the morning (Hanlon et al., 2015; Chung et al., 2011).
Endocannabinoids: Effectors of glucocorticoid signaling
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- 1
Present address: Experimental Psychiatry Unit, Center for Psychiatry and Psychotherapy, Medical University of Innsbruck, Austria.
- 2
These authors contributed equally to the work.