Distinct roles of ascorbic acid in extracellular vesicles and free form: Implications for metabolism and oxidative stress in presymptomatic Huntington's disease

Huntington's disease (HD) is a neurodegenerative disorder caused by a CAG trinucleotide repeat expansion in the first exon of the huntingtin gene. The huntingtin protein (Htt) is ubiquitously expressed and localized in several organelles, including endosomes, where it plays an essential role in intracellular trafficking. Presymptomatic HD is associated with a failure in energy metabolism and oxidative stress. Ascorbic acid is a potent antioxidant that plays a key role in modulating neuronal metabolism and is highly concentrated in the brain. During synaptic activity, neurons take up ascorbic acid released by glial cells; however, this process is disrupted in HD. In this study, we aim to elucidate the molecular and cellular mechanisms underlying this dysfunction. Using an electrophysiological approach in presymptomatic YAC128 HD slices, we observed decreased ascorbic acid flux from astrocytes to neurons, which altered neuronal metabolic substrate preferences. Ascorbic acid efflux and recycling were also decreased in cultured astrocytes from YAC128 HD mice. We confirmed our findings using GFAP-HD160Q, an HD mice model expressing mutant N-terminal Htt mainly in astrocytes. For the first time, we demonstrated that ascorbic acid is released from astrocytes via extracellular vesicles (EVs). Decreased number of particles and exosomal markers were observed in EV fractions from cultured YAC128 HD astrocytes and Htt-KD cells. We observed reduced number of multivesicular bodies (MVBs) in YAC128 HD striatum via electron microscopy, suggesting mutant Htt alters MVB biogenesis. EVs containing ascorbic acid effectively reduced reactive oxygen species, whereas “free” ascorbic acid played a role in modulating neuronal metabolic substrate preferences. These findings suggest that the early redox imbalance observed in HD arises from a reduced release of ascorbic acid-containing EVs by astrocytes. Meanwhile, a decrease in "free" ascorbic acid likely contributes to presymptomatic metabolic impairment. • This study describes the cellular and molecular mechanisms underlying the presymptomatic decrease in ascorbic acid transfer from astrocytes to neurons observed in Huntington’s disease (HD), which disrupts neuronal metabolic preferences and contributes to early redox dysfunction. • This research demonstrated for the first time that ascorbic acid is released from astrocytes via extracellular vesicles, and there is a notable reduction in the number of these vesicles and exosomal markers in HD models. • EVs containing ascorbic acid reduced reactive oxygen species, while "free" ascorbic acid influenced neuronal metabolic substrate preferences. These findings suggest that early redox imbalance in HD stems from reduced astrocytic release of ascorbic acid-containing EVs, with decreased "free" ascorbic acid contributing to presymptomatic metabolic impairment Huntington's disease affects brain function through altered metabolism & oxidative stress, linked to reduced astrocytic ascorbic acid release.