Storage of aversive memories is of utmost importance for survival, allowing animals to avoid upcoming similar stimuli. However, without reinforcement, the learned avoidance response gradually decreases over time. Although the molecular mechanisms controlling this extinction process are not well known, there is evidence that the endocannabinoid system plays a key role through CB1 receptor-mediated modulation of cholinergic signaling. In this study, we measured fear extinction throughout 7 months using naïve rats, assessed in passive avoidance (PA) test in a non-reinforced manner. Then, we evaluated the effect of gentle handling and non-aversive novel object recognition test (NORT) on the extinction and expression of fear memories by measuring passive avoidance responses. Neurochemical correlates were analyzed by functional autoradiography for cannabinoid, cholinergic, and dopaminergic receptors. Despite results showing a gradual decrease of passive avoidance response, it did not fully disappear even after 7 months, indicating the robustness of this process. Meanwhile, in rats that received gentle handling or performed NORT after receiving the PA aversive stimulus, extinction occurred within a week. In contrast, gentle handling performed before receiving the aversive stimulus exacerbated fear expression and triggered escape response in PA. The neurochemical analysis showed increased cannabinoid and cholinergic activity in the nucleus basalis magnocellularis (NBM) in rats that had performed only PA, as opposed to rats that received gentle handling before PA. Additionally, a correlation between CB1 mediated-signaling in the NBM and freezing in PA was found, suggesting that the endocannabinoid system might be responsible for modulating fear response induced by aversive memories.

Kv3.3 proteins are pore-forming subunits of voltage-dependent potassium channels, and mutations in the gene encoding for Kv3.3 have recently been linked to human disease, spinocerebellar ataxia 13, with cerebellar and extracerebellar symptoms. To understand better the functions of Kv3.3 subunits in brain, we developed highly specific antibodies to Kv3.3 and analyzed immunoreactivity throughout mouse brain. We found that Kv3.3 subunits are widely expressed, present in important forebrain structures but particularly prominent in brainstem and cerebellum. In forebrain and midbrain, Kv3.3 expression was often found colocalized with parvalbumin and other Kv3 subunits in inhibitory neurons. In brainstem, Kv3.3 was strongly expressed in auditory and other sensory nuclei. In cerebellar cortex, Kv3.3 expression was found in Purkinje and granule cells. Kv3.3 proteins were observed in axons, terminals, somas, and, unlike other Kv3 proteins, also in distal dendrites, although precise subcellular localization depended on cell type. For example, hippocampal dentate granule cells expressed Kv3.3 subunits specifically in their mossy fiber axons, whereas Purkinje cells of the cerebellar cortex strongly expressed Kv3.3 subunits in axons, somas, and proximal and distal, but not second- and third-order, dendrites. Expression in Purkinje cell dendrites was confirmed by immunoelectron microscopy. Kv3 channels have been demonstrated to rapidly repolarize action potentials and support high-frequency firing in various neuronal populations. In this study, we identified additional populations and subcellular compartments that are likely to sustain high-frequency firing because of the expression of Kv3.3 and other Kv3 subunits.

Williams-Beuren syndrome (WBS) is a rare genetic multisystemic disorder characterized by mild-to-moderate intellectual disability and hypersocial phenotype, while the most life-threatening features are cardiovascular abnormalities. Nowadays, there are no pharmacological treatments to directly ameliorate the main traits of WBS. The endocannabinoid system (ECS), given its relevance for both cognitive and cardiovascular function, could be a potential druggable target in this syndrome. We analyzed the components of the ECS in the complete deletion (CD) mouse model of WBS and assessed the impact of its pharmacological modulation in key phenotypes relevant for WBS. CD mice showed the characteristic hypersociable phenotype with no preference for social novelty and poor short-term object-recognition performance. Brain cannabinoid type-1 receptor (CB1R) in CD male mice showed alterations in density and coupling with no detectable change in main endocannabinoids. Endocannabinoid signaling modulation with subchronic (10 days) JZL184, a selective inhibitor of monoacylglycerol lipase, specifically normalized the social and cognitive phenotype of CD mice. Notably, JZL184 treatment improved cardiovascular function and restored gene expression patterns in cardiac tissue. These results reveal the modulation of the ECS as a promising novel therapeutic approach to improve key phenotypic alterations in WBS.