Natural Cannabinoids Improve Dopamine Neurotransmission and Tau and Amyloid Pathology in a Mouse Model of Tauopathy

Abstract. Cannabinoids are neuroprotective in models of neurodegenerative dementias. Their effects are mostly mediated through CB1 and CB2 receptor-dependent modulation of excitotoxicity, inflammation, oxidative stress, and other processes. We tested the effects of Sativex ® , a mixture of 9 -tetrahydrocannabinol and cannabidiol, acting on both CB1 and CB2 receptors, in parkin-null, human tau overexpressing (PK −/− /Tau VLW ) mice, a model of complex frontotemporal dementia, parkinsonism, and lower motor neuron disease. The animals received Sativex ® , 4.63 mg/kg, ip, daily, for one month, at six months of age, at the onset of the clinical symptoms. We evaluated the effects of Sativex ® on behavior, dopamine neurotransmission, glial activation, redox state, mitochondrial activity, and deposition of abnormal proteins. PK −/− /Tau VLW mice developed the neurological deficits, but those treated with Sativex ® showed less abnormal behaviors related to stress, less auto and hetero-aggression, and less stereotypy. Sativex ® significantly reduced the intraneuronal, MAO-related free radicals produced during dopamine metabolism in the limbic system. Sativex ® also decreased gliosis in cortex and hippocampus, increased the ratio reduced/oxidized glutathione in the limbic system, reduced the levels of iNOS, and increased those of complex IV in the cerebral cortex. With regard to tau and amyloid pathology, Sativex ® reduced the deposition of both in the hippocampus and cerebral cortex of PK −/− /Tau VLW mice and increased autophagy. Sativex ® , even after a short administration in animals with present behavioral and pathological abnormalities, improves the phenotype, the oxidative stress, and the deposition of proteins in PK −/− /Tau VLW mice, a model of complex neurodegenerative disorders

NMDA receptor gene variations as modifiers in Huntington disease: A replication study

Several candidate modifier genes which, in addition to the pathogenic CAG repeat expansion, influence the age at onset (AO) in Huntington disease (HD) have already been described. The aim of this study was to replicate association of variations in the N-methyl D-aspartate receptor subtype genes GRIN2A and GRIN2B in the "REGISTRY" cohort from the European Huntington Disease Network (EHDN). The analyses did replicate the association reported between the GRIN2A rs2650427 variation and AO in the entire cohort. Yet, when subjects were stratified by AO subtypes, we found nominally significant evidence for an association of the GRIN2A rs1969060 variation and the GRIN2B rs1806201 variation. These findings further implicate the N-methyl D-aspartate receptor subtype genes as loci containing variation associated with AO in HD

NMDA receptor gene variations as modifiers in Huntington disease: a replication study.

none12siSeveral candidate modifier genes which, in addition to the pathogenic CAG repeat expansion, influence the age at onset (AO) in Huntington disease (HD) have already been described. The aim of this study was to replicate association of variations in the N-methyl D-aspartate receptor subtype genes GRIN2A and GRIN2B in the "REGISTRY" cohort from the European Huntington Disease Network (EHDN). The analyses did replicate the association reported between the GRIN2A rs2650427 variation and AO in the entire cohort. Yet, when subjects were stratified by AO subtypes, we found nominally significant evidence for an association of the GRIN2A rs1969060 variation and the GRIN2B rs1806201 variation. These findings further implicate the N-methyl D-aspartate receptor subtype genes as loci containing variation associated with AO in HD.openC. Saft; J. T. Epplen; S. Wieczorek; G. B. Landwehrmeyer; R. A. C;J. G. de;M. Dose;S. J. Tabrizi;D. Craufurd;R. E. G.;Investigators of the European Huntington's Disease Network [E. Di Maria];L. ArningC., Saft; J. T., Epplen; S., Wieczorek; G. B., Landwehrmeyer; R. A., C; J. G., De; M., Dose; S. J., Tabrizi; D., Craufurd; R. E., G.; DI MARIA, Emilio; L., Arnin

Effects of epoxomicin and trehalose on ERK-1/2 and HSP70 chaperone protein activation in HD fibroblasts.

<p>(A) Western blot of p-ERK-1/2 expression with regard to total ERK and its corresponding densitometric analysis in control and HD fibroblasts. (B) Western blot of HSP70 expression and its corresponding densitometric analysis. Values are expressed as the mean ± SD, <i>n</i> = 4 patients. The data of each patient was obtained using 4 replicates. Statistical analysis was performed by one-way ANOVA with repeated measures followed by Bonferroni multiple comparison test: *p<0.05, ***p<0.001 <i>vs</i> Solvent; +++p<0.001 HD <i>vs</i> controls, ΔΔΔp<0.001 trehalose + epoxomicin <i>vs</i> epoxomicin. There is an interaction between epoxomicin effect and genotype in ERK activation (F = 71.13 with a p value  = <0.0001). In HD, there is an interaction between the epoxomicin and trehalose effects in ERK activation (F = 12.67 with a p value  = 0.0013).</p

Epoxomicin and trehalose differential effects in cellular viability on control and HD human skin fibroblasts.

<p>(A) Dose-dependent effects of epoxomicin in caspase-3 activation, an indicator of apoptosis. (B) Photomicrographs of activated caspase-3⁺ cells (green) and total nuclei stained with bis-benzimide (blue) after epoxomicin and trehalose treatments. (Scale bar  = 20 µm). (C) Percent of activated caspase-3⁺ cells in control and HD fibroblasts after epoxomicin and trehalose treatments. Values are expressed as the mean ± SD, <i>n</i> = 4 patients. Control cell number (mean per field) 31.87±1.124, <i>n</i> = 4. HD cell number (mean per field) 44.75±2.456, <i>n</i> = 4. The data of each patient was obtained using 4 replicates. Statistical analysis was performed by one-way ANOVA with repeated measures followed by Bonferroni multiple comparison test: *p<0.05, **p<0.01, ***p<0.001 <i>vs</i> Solvent; +p<0.05, ++p<0.01, +++p<0.001 HD <i>vs</i> controls; ΔΔp<0.01, ΔΔΔp<0.001 trehalose + epoxomicin <i>vs</i> epoxomicin; δδδp<0.001 3-methyladenine + trehalose + epoxomicin <i>vs</i> trehalose + epoxomicin.</p

Trehalose protects against accumulation of Huntingtin and poly-ubiquitinated protein induced by epoxomicin and increases UPS activity.

<p>(A) Huntingtin immunocytochemistry (green) and total nuclei stained with bis-benzimide (blue). The histogram shows the ratio of Huntingtin (IOD) with respect to the total cell number. (Scale bar  = 20 µm) (B) Ubiquitinated protein accumulation and its corresponding densitometric analysis. (C) Chymotrypsin-like proteasome activity. Values are expressed as the mean ± SD, <i>n</i> = 4 patients. Control cell number (mean per field) 32,66±1.472, <i>n</i> = 4. HD cell number (mean per field) 44,67±0.3405, <i>n</i> = 4. The data of each patient was obtained using 4 replicates. Statistical analysis was performed by one-way ANOVA with repeated measures followed by Bonferroni multiple comparison test: *p<0.05, ***p<0.001 <i>vs</i> solvent; +p<0.05, +++p<0.001 HD <i>vs</i> controls; ΔΔΔp<0.001 trehalose + epoxomicin <i>vs</i> epoxomicin.</p

Macroautophagyc pathway and chaperone-mediated autophagy in control and HD human skin fibroblasts.

<p>(A) LC3 immunocytochemistry (green) and total nuclei stained with bis-benzimide (blue) and the percentage of LC3 positive cells with respect to the total cell number. (B) LAMP2A immunocytochemistry (green) and total nuclei stained with bis-benzimide (blue) and percentage of LAMP2A positive cells. (C) LAMP2A (green) and HSC70 (red) colocalization (yellow) and percentage of LAMP2A and HSC70 colocalization respect to the total cell number. (Scale bar  = 20 µm). Values are expressed as the mean ± SD, <i>n</i> = 4 patients. Control cell number (mean per field) 30.94±2.012, <i>n</i> = 4. HD cell number (mean per field) 46.89±4.587, <i>n</i> = 4. The data of each patient was obtained using 4 replicates. Statistical analysis was performed by one-way ANOVA with repeated measures followed by Bonferroni multiple comparison test: **p<0.01, ***p<0.001 <i>vs</i> Solvent; +<0.05, +p<0.05, ++p<0.01, +++p<0.001 HD <i>vs</i> controls; ΔΔΔp<0.001 trehalose + epoxomicin <i>vs</i> epoxomicin.</p

Epoxomicin increases ROS levels, which are reduced by trehalose in HD firboblasts.

<p>(A) 2′, 7′–dichlorofluorescin (DCF) immunocytochemistry (green) and total nuclei stained with bis-benzimide (blue) in control and HD fibroblasts and (B) percentage of DCF positive cells respect to the total number. (Scale bar  = 20 µm). Values are expressed as the mean ± SD, <i>n</i> = 4 patients. Control cell number (mean per field) 28.83±0.9280, <i>n</i> = 4. HD cell number (mean per field) 46.40±5.247, <i>n</i> = 4. The data of each patient was obtained using 4 replicates. Statistical analysis was performed by one-way ANOVA with repeated measures followed by Bonferroni multiple comparison test: ***p<0.001 <i>vs</i> solvent; +p<0.05, +++p<0.001 HD <i>vs</i> controls; ΔΔΔp<0.001 trehalose + epoxomicin <i>vs</i> epoxomicin.</p