Neuron-astroglial communications : NMDA-induced proheparanase modulation of synaptic strength

Abstract

Recent findings of neuronal expression of heparanase and the possible role of perisynaptic proheparanase in triggering decrease in both basal and long-term potentiation of synaptic strength set the stage for this work to find sources of proheparanase in the synaptic environment in the context of the tripartite synapse, comprising pre- and post-synaptic neurons and peri-synaptic astrocytes. With highly purified hippocampal neurons forming networks at around 21 DIV and low-dose NMDA (glutamate analogue) to chemically induce long-term depression of synaptic strength, proheparanase secreted into the serum-free medium was found to be twice that of controls. Similar treatment of purified astrocytes in culture showed medium proheparanase 5-fold that of controls, suggesting astrocytic capacity for response to low-dose NMDA exceeding that of neurons. Taking the cue that exosomes carry membrane-clustered proheparanase for release into the extracellular environment, the exosome fraction was recovered from the conditioned medium of astrocyte cultures following differential centrifugation and then ultracentrifugation. Indeed, proheparanase was detectable in the exosome fraction as marked by immunopositivities for Lamp1, TSG101 and Alix. Phorbol ester (activator of protein kinase C pathway) and low-dose NMDA treatments of hippocampal neurons triggered additive effects on release of proheparanase into the extracellular environment, via exocytosis and exosomes respectively. Exosomal proheparanse affected a site-specific phosphorylation of GluA2-containing AMPA receptors. By contrast, long-term deficiency of proheparanase in hippocampal neurons decreased the GluA2 site-specific phosphorylation and increased total GluA2-containing AMPA receptor level. The study presented here provides evidence that astrocytes can respond to the spill-over of glutamate during neuronal transmission and react via proheparanase to modulate synaptic transmission. Hence, the current study supports and delineates a new means by which astrocytes can modulate synaptic strength. The finding provides insights into astrocytes as a potential target for treating cognitive impairments involving the hippocampus.