Achucarro Seminars

14 May [2021]

at 13.45 CET

High resolution imaging of cannabinoid receptors in glial cells in a mouse model of Alzheimer's disease

Itziar Terradillos

Laboratory of Ultrastructural and Functional Neuroanatomy of the Synapse (Achucarro)


Abstract

Alzheimer's disease (AD) is the most common form of dementia characterized by neuritic plaques formation and accumulation of neurofibrillary tangles. Effective therapies are not yet available, thus new targets are actively searched for tackling the disease. The potential of cannabinoids to act on several processes involved in AD pathogenesis (oxidative stress, Aβ and τ protein metabolism, inflammation, mitochondrial dysfunction, excitotoxicity),  prompted us to carry out investigations on the nexus between inflammation and the endocannabinoid system (ECS) in animal models of AD. The ECS is made up of cannabinoid receptors (mostly CB1, CB2) and other components. The CB1 is one of the most abundant G-protein coupled receptors in the brain. Our laboratory has recently revealed in the hippocampus that about 56% of the total CB1 labeling is in GABAergic terminals, ~12% in glutamatergic terminals, ~6% in astrocytes, ~15% in mitochondria and ~11% in unidentified compartments including microglia. Although microglia barely contains (if any) CB1 at resting conditions, it seems to be expressed in this glial cell type depending on the conditions. Likewise, CB2 increases in microglia in inflammatory environments and in activated microglia in the brain of patients with Alzheimer´s disease. In animal models, the EGFP expression in mice CB2EGFP/f/f crossed with mice expressing 5 familial mutations of the Alzheimer´s disease (5xFAD) (CB2EGFP/f/f/5xFAD) coincides with the presence of neuritic plaques in several brain regions and is restricted to microglial cells in the vicinity of the neuritic plaques (López et al, 2018). Astrocytes also participate in inflammatory responses through their capacity to release pro-inflammatory molecules, diminished by anti-inflammatory reactions mediated by endocannabinoids acting on astroglial CB1 receptors. Therefore, given the relevant role of both the ECS and neuroglia in inflammatory mechanisms and the implication of the ECS in AD, it is timely to investigate the subcellular localization of CB1 receptors in astrocytes and microglia in animal models of AD.

I will present in my talk the recent findings using a transgenic mouse model expressing EGFP under the control of the gen cnr2 promoter preceded by the insertion of an IRES (internal ribosomal entry site) sequence in the 3´untranslated (UTR) region of the cnr2 gen. These mice were crossed with mice expressing 5 familial Azheimer´s disease mutations (5xFAD) (López et al, 2018). I applied to the subiculum (where the plaques were most abundant) of 6.5-month-old wild type (WT) and CB2EGFP/f/f/5xFAD mutant mice, antibodies against GLAST (for astroglia) and Iba1 (for microglia) in combination with specific CB1 antibodies and high-resolution immunoelectron microscopy. CB1 receptors were localized in 17.75 ± 1.21% of the astroglial processes in WT and in 21.24 ± 2.37% in CB2EGFP/f/f/5xFAD mice (ns; P: 0.6421). Furthermore, 3.79 ± 2.10% of the microglial processes in WT contained CB1, while 6.27 ± 1.15% of them had the receptor in CB2EGFP/f/f/5xFAD mice (**; P: 0.0033). Furthermore, the number of CB1 particles in microglia (per 100µm2) was higher in the mutants (1.31 ± 0.18) than in WT (0.48 ± 0.13)(***; P: 0.0009). 

These changes in CB1 receptor localization in microglial cells that were also associated with changes in cell morphology in the mouse model of AD, will be further investigated to elucidate their impact on the (anti)inflammatory response, tripartite synapse homeostasis and synaptic plasticity, whose disruption underlies brain dysfunction.

Currently we are hosting our seminar in Zoom. The link is available to external people on request.