Hyaluronidase inhibition accelerates functional recovery from stroke in the mouse brain

Perineuronal nets (PNNs) are presumed to limit plasticity in adult animals. Ischaemic stroke results in the massive breakdown of PNNs resulting in rejuvenating states of neuronal plasticity, but the mechanisms of this phenomenon are largely unknown. As hyaluronic acid (HA) is the structural backbone of PNNs, we hypothesized that these changes are a consequence of the altered expression of HA metabolism enzymes. Additionally, we investigated whether early hyaluronidase inhibition interferes with post-stroke PNN reduction and behavioural recovery. We investigated the mRNA/protein expression of these enzymes in the perilesional, remote and contralateral cortical regions in mice at different time points after photothrombosis, using quantitative real-time polymerase chain reaction and immunofluorescence. A skilled reaching test was employed to test hyaluronidase inhibitor L-ascorbic acid 6-hexadecanoate influence on post-stroke recovery. We found the simultaneous up-regulation of mRNA of HA synthesizing and degrading enzymes in the perilesional area early after stroke, suggesting an acceleration of HA turnover in ischaemic animals. Immunostaining revealed differential cellular localization of enzymes, with hyaluronidase 1 in astrocytes and hyaluronan synthase 2 in astrocytes and neurons, and post-stroke up-regulation of both of them in astrocytes. β-glucuronidase was observed in neurons but post-stroke up-regulation occurred in microglia. Inhibition of hyaluronidase activity early after stroke resulted in improved performance in skilled reaching test, without affecting the numbers of PNNs. These results suggest that after stroke, a substantial reorganization of polysaccharide content occurs, and interfering with this process at early time has a beneficial effect on recovery.