Effects of acute versus repeated cocaine exposure on the expression of endocannabinoid signaling-related proteins in the mouse cerebellum

Abstract

Growing awareness of cerebellar involvement in addiction is based on the cerebellum’s intermediary position between motor and reward, potentially acting as an interface between motivational and cognitive functions. Here, we examined the impact of acute and repeated cocaine exposure on the two main signaling systems in the mouse cerebellum: the endocannabinoid (eCB) and glutamate systems. To this end, we investigated whether eCB signaling-related gene and protein expression {cannabinoid receptor type 1 receptors and enzymes that produce [diacylglycerol lipase alpha/beta (DAGLα/β) and N-acyl phosphatidylethanolamine phospholipase D (NAPE-PLD)] and degrade [monoacylglycerol lipase (MAGL) and fatty acid amino hydrolase (FAAH)] eCB} were altered. In addition, we analyzed the gene expression of relevant components of the glutamate signaling system [glutamate synthesizing enzymes liver-type glutaminase isoform (LGA) and kidney-type glutaminase isoform (KGA), metabotropic glutamatergic receptor (mGluR3/5), NMDA-ionotropic glutamatergic receptor (NR1/2A/2B/2C) and AMPA-ionotropic receptor subunits (GluR1/2/3/4)] and the gene expression of tyrosine hydroxylase (TH), the rate-limiting enzyme in catecholamine biosynthesis, because noradrenergic terminals innervate the cerebellar cortex. Results indicated that acute cocaine exposure decreased DAGLα expression, suggesting a down-regulation of 2-arachidonylglycerol (2-AG) production, as well as gene expression of TH, KGA, mGluR3 and all ionotropic receptor subunits analyzed in the cerebellum. The acquisition of conditioned locomotion and sensitization after repeated cocaine exposure were associated with an increased NAPE-PLD/FAAH ratio, suggesting enhanced anandamide production, and a decreased DAGLβ/MAGL ratio, suggesting decreased 2-AG generation. Repeated cocaine also increased LGA gene expression but had no effect on glutamate receptors. These findings indicate that acute cocaine modulates the expression of the eCB and glutamate systems. Repeated cocaine results in normalization of glutamate receptor expression, although sustained changes in eCB is observed. We suggest that cocaine-induced alterations to cerebellar eCB should be considered when analyzing the adaptations imposed by psychostimulants that lead to addiction.

Endocannabinoid signalling in reward and addiction

“…Psychostimulants generally produce modest disruptions in brain EC content with subtle increases and decreases in 2-AG concentration in forebrain following non-contingent acute and chronic cocaine exposure, respectively (no other alterations evident regardless of region analyzed) 48, 52 . Moreover, voluntary cocaine self-administration does not alter rat extracellular NAc EC levels 30 but decreases 2-AG content in frontal cortex and hippocampal tissue 53–55 .…”

Glutamate and Brain Glutaminases in Drug Addiction

“…In the PFC, GLS2 mRNA increases in acute cocaine priming, as well as in acute cocaine/sensitized mice (Fig. 3) [63,65], while its expression levels remain essentially unchanged in hippocampus and cerebellum [63,64] (although in this last region, a significant increase was detected for the C-C group vs the V-V group [66]). It is noteworthy that both GA isoforms showed a strikingly different pattern in the cocaine addiction paradigm described above.…”

Have we been ignoring the elephant in the room? Seven arguments for considering the cerebellum as part of addiction circuitry

“…This effect is mediated by D1, D2, GABA B and NMDA 10 cerebellar receptors (Couceyro et al, 1994;Klitenick et al, 1995). Interestingly, glutamateendocannabinoid synaptic interactions within the cerebellum have been also associated with cocaine-induced sensitisation (Palomino et al, 2014). Very recently, we have investigated molecular and structural plasticity in the cerebellum of sensitized mice (Vazquez-Sanroman et al, 2015a;Vazquez-Sanroman et al, 2015b).…”

“…Overall, addictive drugs modify cerebellar glutamate and endocannabinoid interactions (Netzeband et al, 1999;Rubino et al, 2004;Palomino et al, 2014), neuromodulators like norepinephrine and dopamine (Ferrucci et al, 2007;Yin et al, 2010;Bekheet et al, 2010), intracellular signalling transduction pathways (Sanna et al, 2002;Alfonso-Loeches and Guerri, 2011), as well as gene expression (Enoch et al, 2014;Palomino et al, 2014). Several of these drug-induced molecular changes have been correlated with drug tolerance, drug dependence and the withdrawal syndrome (Hutcheson et al, 1998;Sanna et al, 2002;Rubino et al, 2004;Vinod et al, 2006).…”