Abstract:
Over the last two decades, building evidence has shown that antibodies in the peripheral circulatory system are able to cross the blood-brain-barrier (BBB) and enter the central nervous system (CNS) to modulate brain function, as supported by findings in vaccination strategies for Alzheimer’s disease (AD) in mouse models and autoimmune disease models. Our laboratory has previously shown that naïve rats vaccinated against the GluN1 subunit of the N-methyl-D-aspartate (NMDA) receptor developed high anti-GluN1 antibody titres, with GluN1 antibodies found at low levels in the cerebrospinal fluids. Moreover, GluN1-vaccinated rats showed enhanced learning and memory function. These exciting observations are crucial for developing an immunotherapeutic strategy for neurodegenerative disease treatments as the NMDA receptor is not only essential for brain function, but is also involved in the pathology of a number of brain diseases. This aim of the thesis was to characterise the mechanism(s) by which anti-GluN1 antibodies mediate improved learning and memory in rats. Recombinant GluN1 (recGluN1) vaccination was not found to be associated with any neurotoxicity in the rat brain and NMDA receptor expression levels in recGluN1-vaccinated rat hippocampi were not different compared to control-vaccinated animals. Interestingly, passive transfer of rat anti-GluN1 antibodies to naïve mice conferred improved performance in tests of object recognition and fear-associative learning and memory, providing more evidence that rat anti-GluN1 antibodies mediate improved learning and memory. NMDA receptor-linked protein kinase B (PKB) and extracellular signal-regulated kinase 2 (ERK2) activities were upregulated, but Ca2+/calmodulin-dependent protein kinase II (CaMKII) activity was downregulated, as a result of binding and modulation of NMDA receptor function by anti-GluN1 antibodies. Rat anti-GluN1 antibodies bound with high affinity to the GluN1 glycine-binding site. Autoantibodies to brain antigens including GluN1 are also commonly found in humans with neurological disorders and whether these antibodies could affect neuronal function is unclear. We generated epitope binding profiles for 14 human stroke sera that also showed reactivity to the GluN1 subunit and discovered they had different epitope binding profiles. Neuronal cell death is a common clinical symptom of stroke; therefore, as a proof of concept, purified antibodies from four stroke sera with distinctively different anti-GluN1 epitope binding profiles were applied to rat primary hippocampal neuronal cultures, with neurotoxicity being associated with two stroke samples. This thesis demonstrates that the anti-GluN1 antibody generated by recGluN1 vaccination (but not human stroke-derived) is beneficial to neuronal function in the CNS, and the hope is that the characterisation of the effects of anti-GluN1 antibodies on neuronal function will further guide the development of antibody-based immunotherapeutic strategies for stroke treatment and rehabilitation.