Pharmacological Modulation of Glutamatergic and Neuroinflammatory Pathways in a Lafora Disease Mouse Model

Abstract

Lafora disease (LD) is a fatal rare neurodegenerative disorder that affects young adolescents and has no treatment yet. The hallmark of LD is the presence of polyglucosan inclusions (PGs), called Lafora bodies (LBs), in the brain and peripheral tissues. LD is caused by mutations in either EPM2A or EPM2B genes, which, respectively, encode laforin, a glucan phosphatase, and malin, an E3-ubiquitin ligase, with identical clinical features. LD knockout mouse models (Epm2a − / − and Epm2b − / −) recapitulate PG body accumulation, as in the human pathology, and display alterations in glutamatergic transmission and neuroinflammatory pathways in the brain. In this work, we show the results of four pre-clinical trials based on the modulation of glutamatergic transmission (riluzole and memantine) and anti-neuroinflammatory interventions (resveratrol and minocycline) as therapeutical strategies in an Epm2b − / − mouse model. Drugs were administered in mice from 3 to 5 months of age, corresponding to early stage of the disease, and we evaluated the beneficial effect of the drugs by in vivo behavioral phenotyping and ex vivo histopathological brain analyses. The behavioral assessment was based on a battery of anxiety, cognitive, and neurodegenerative tests and the histopathological analyses included a panel of markers regarding PG accumulation, astrogliosis, and microgliosis. Overall, the outcome of ameliorating the excessive glutamatergic neurotransmission present in Epm2b − / − mice by memantine displayed therapeutic effectiveness at the behavioral levels. Modulation of neuroinflammation by resveratrol and minocycline also showed beneficial effects at the behavioral level. Therefore, our study suggests that both therapeutical strategies could be beneficial for the treatment of LD patients. Graphical abstract A mouse model of Lafora disease (Epm2b-/-) was used to check the putative beneficial effect of different drugs aimed to ameliorate the alterations in glutamatergic transmission and/or neuroinflammation present in the model. Drugs in blue gave a more positive outcome than the rest.

Beneficial effect of fingolimod in a Lafora disease mouse model by preventing reactive astrogliosis-derived neuroinflammation and brain infiltration of T-lymphocytes.

“…To provide an alternative to the use of metformin, our group has checked the possible bene cial effect of alternative repurposing drugs. We recently reported the bene cial effect of propranolol [20] and memantine and minocycline [21].…”

“…The order of the behavioral tests and resting time between them were the same for each mouse. The selected battery of behavioral tests was based on the results obtained in previous reports for the use of Epm2b-/-mice [20], [21]. It consisted of three tests performed in the following order: hindlimb clasping, open eld, and object location memory (OLM).…”

“…The samples in para n were sagittal sectioned at 4 μm using a microtome. The presence of polyglucosans (PGs) inclusions (the hallmark of Lafora disease) in Epm2b-/mice, was assessed by periodic acid Schiff (PAS) staining as detailed in [20], [21]. Six mice (3 females and 3 males) were analyzed in total per each group (WT or Epm2b-/untreated and Epm2b-/treated with ngolimod or dimetylfumarate).…”

“…Brain samples derived from six individual males and females from each group of mice were analyzed by immuno uorescence to asses reactive astrocytes and microglia, as previously described in [20], [21]. In parallel ten individual mice per group were analyzed for T-lymphocytes in ltration as described in [15].…”

Beneficial Effect of Fingolimod in a Lafora Disease Mouse Model by Preventing Reactive Astrogliosis-Derived Neuroinflammation and Brain Infiltration of T-lymphocytes

“…Most of the results described above have been obtained by using LD knockout (KO) mouse models of LD with a complete loss of function of laforin ( Epm2a-/- ) [ 16 ] or malin ( Epm2b-/- ) [ 17 ]. These animals partially mimic human symptoms such as early accumulation of PGs in brain and peripheral tissues, abnormal postures of the hindlimb, memory defects, and hyperactivity with disrupted attention [ 16 – 21 ].…”