Damaged Neocortical Perineuronal Nets Due to Experimental Focal Cerebral Ischemia in Mice, Rats and Sheep

Abstract

As part of the extracellular matrix (ECM), perineuronal nets (PNs) are polyanionic, chondroitin sulfate proteoglycan (CSPG)-rich coatings of certain neurons, known to be affected in various neural diseases. Although these structures are considered as important parts of the neurovascular unit (NVU), their role during evolution of acute ischemic stroke and subsequent tissue damage is poorly understood and only a few preclinical studies analyzed PNs after acute ischemic stroke. By employing three models of experimental focal cerebral ischemia, this study was focused on histopathological alterations of PNs and concomitant vascular, glial and neuronal changes according to the NVU concept. We analyzed brain tissues obtained 1 day after ischemia onset from: (a) mice after filament-based permanent middle cerebral artery occlusion (pMCAO); (b) rats subjected to thromboembolic MACO; and (c) sheep at 14 days after electrosurgically induced focal cerebral ischemia. Multiple fluorescence labeling was applied to explore simultaneous alterations of NVU and ECM. Serial mouse sections labeled with the net marker Wisteria floribunda agglutinin (WFA) displayed largely decomposed and nearly erased PNs in infarcted neocortical areas that were demarcated by up-regulated immunoreactivity for vascular collagen IV (Coll IV). Subsequent semi-quantitative analyses in mice confirmed significantly decreased WFA-staining along the ischemic border zone and a relative decrease in the directly ischemia-affected neocortex. Triple fluorescence labeling throughout the three animal models revealed up-regulated Coll IV and decomposed PNs accompanied by activated astroglia and altered immunoreactivity for parvalbumin, a calcium-binding protein in fast-firing GABAergic neurons which are predominantly surrounded by neocortical PNs. Furthermore, ischemic neocortical areas in rodents simultaneously displayed less intense staining of WFA, aggrecan, the net components neurocan, versican and the cartilage link protein (CRTL) as well as markers in net-bearing neurons such as the potassium channel subunit Kv3.1b and neuronal nuclei (NeuN). In summary, theconsistent observations based on three different stroke models confirmed that PNs are highly sensitive constituents of the NVU along with impaired associated GABAergic neurons. These results suggest that PNs could be promising targets of future stroke treatment, and further studies should address their reorganization and plasticity in both stabilizing the acute stroke as well as supportive effects during the chronic phase of stroke.

The Perineuronal ‘Safety’ Net? Perineuronal Net Abnormalities in Neurological Disorders

“…In contrast, in cortical areas surrounding the infarct and outside of the glial scar, WFA-expressing PNNs are completely degraded by day 7, and CSPG immunoreactivity is down regulated although they remain detectable up to 14 days post-injury ( Hobohm et al, 2005 ). Similar degradation of WFA-expressing PNNs in the neocortex has also been observed following thromboembolic middle cerebral artery occlusion in rats, permanent middle artery occlusion in mice, and focal cerebral ischemia in sheep ( Härtig et al, 2017 ). Interestingly, reduced PV-immunoreactivity and PV cell function have also been observed following stroke and this decrease in inhibitory cell function may underlie epilepsy-like activity following stroke ( Hobohm et al, 2005 ; Xie et al, 2014 ).…”

Chondroitin sulfate metabolism in the brain

“…These data support the notion that removal of CS chains from CSPGs molecules allows alterations in the neural circuitry and results in increased plasticity (Fox and Caterson, 2002). Moreover, it appears that the numbers of PNNs, while generally stable in adult brain, can be dramatically decreased upon ischemia, seizures and traumatic brain injury (Hobohm et al, 2005;Harris et al, 2010;Karetko-Sysa et al, 2011;McRae et al, 2012;Yi et al, 2012;Härtig et al, 2017). In view of the above mentioned data, this decrease can be considered as an attempt of the impaired brain to reinstate plasticity in order to allow axonal sprouting and compensation of damaged function (Carmichael et al, 2017).…”

Structural Variation of Chondroitin Sulfate Chains Contributes to the Molecular Heterogeneity of Perineuronal Nets

“…It was reported that the glycan structures of CSPGs were affected in schizophrenia and epilepsy (Pantazopoulos et al, 2010 , 2015 ; Yutsudo and Kitagawa, 2015 ; Miyata and Kitagawa, 2016b ). PNNs are markedly reduced in experimental ischemic stroke models (Härtig et al, 2016 , 2017 ). Thus, it will be fascinating to assess the contribution of the molecular heterogeneity of PNN in models of these diseases.…”