Rab5-mediated endo-lysosomal mistrafficking associated with impaired oxytocin receptor signalling in early Alzheimer’s disease

Endosomal dysfunction is increasingly recognised as an early event in Alzheimer’s disease. Hyperactivation of the early endosomal regulator Rab5 has been implicated in early AD-associated pathology, but its downstream signalling consequences and therapeutic relevance remain incompletely defined. We investigated whether Rab5-associated endo-lysosomal abnormalities are linked to oxytocin receptor trafficking defects and impaired receptor-associated signalling in early Alzheimer’s disease-relevant settings. Young APP/PS1 mice were used to assess endogenous endo-lysosomal abnormalities in the hippocampus. A hippocampus-targeted Rab5 overactivation model was established by stereotaxic delivery of AAV-hSyn-EGFP-Rab5Q79L to test whether sustained Rab5 activation was sufficient to induce similar alterations. Oxytocin receptor localisation, endosomal marker colocalisation, and OTR-associated downstream signalling readouts were evaluated by confocal immunofluorescence and western blotting. Receptor internalisation/recycling assays, membrane protein western blotting, OT rechallenge, and OTR antagonist experiments were performed in HEK293T cells to assess plasma-membrane OTR recovery and OT-responsive ERK/CREB phosphorylation. RNA sequencing, synaptic marker analysis, resting-state functional magnetic resonance imaging, structural magnetic resonance imaging, and behavioural testing were used to assess downstream molecular, circuit-level, and functional consequences. Group comparisons were analysed using one-way analysis of variance followed by Tukey’s post hoc test. Young APP/PS1 mice exhibited enlarged Rab5-positive early endosomal structures, increased overlap of Rab5 with Rab7 and LAMP1, abnormal intracellular redistribution of the oxytocin receptor, and reduced phosphorylation of ERK and CREB in the hippocampus. Rab5 overactivation reproduced Rab5-associated endosomal abnormalities and was accompanied by altered OTR endosomal association, impaired plasma-membrane OTR recovery, and reduced OT-evoked ERK/CREB phosphorylation. These receptor-level and signalling changes were accompanied by disruption of neuronal signalling-related transcriptional programmes, loss of excitatory and inhibitory synapses, reduced spontaneous brain activity, weakened hippocampal-medial prefrontal cortex functional connectivity, hippocampal atrophy, and cognitive impairment. Across the two models, MW150 treatment was associated with partial improvement in endosomal abnormalities, OTR-associated signalling readouts, and behavioural measures; additional synaptic and imaging improvements were observed in the Rab5-overactivation model. Rab5-associated endo-lysosomal trafficking abnormalities are accompanied by altered oxytocin receptor localisation, reduced OTR-associated signalling readouts, and multi-level early AD-relevant phenotypes. The partial improvement in molecular, synaptic, network, and behavioral abnormalities by MW150 supports the modifiability of the p38α–Rab5/endosomal axis. At the same time, further work is needed to establish whether OTR trafficking directly mediates these downstream phenotypes.