Increased blood–brain barrier permeability and alterations in perivascular astrocytes and pericytes induced by intracisternal glutaric acid

Abstract

BackgroundGlutaric acid (GA) is a dicarboxylic acid that accumulates in millimolar concentrations in glutaric acidemia I (GA-I), an inherited neurometabolic childhood disease characterized by extensive neurodegeneration. Vascular dysfunction is a common and early pathological feature in GA-I, although the underlying mechanisms remain unknown. In the present study, we have used a previously-validated rat model of GA-I to determine the effect of GA on the blood- brain barrier (BBB) and the neurovascular unit.MethodsNewborn rat pups received a single injection of GA (1 μmol/g) or vehicle into the cisterna magna. BBB permeability was analyzed at 14 and 30 days post injection (DPI) by assessing Evans blue (EB) and immunoglobulin G (IgG) extravasation. Blood vessels and microglia were labeled with tomato lectin. Characterization of EB positive cells was made by double labeling with antibodies to astrocyte and neuronal markers. Immunohistochemistry against aquaporin 4 (AQP4), β receptor of the platelet derived growth factor (PDGFRβ) and laminin was used to recognize astrocyte endfeet, pericytes and basal lamina. Zonula occludens 1 (ZO-1) and occludin striatal expression was assessed by Western blotting.ResultsPerinatal intracisternal GA administration caused an increased extravasation of free EB, but not of IgG, into the striatal parenchyma at 14 and 30 DPI. EB extravasated through the BBB was internalized exclusively into neurons. GA-injected animals did not show significant changes in the area of small blood vessels in the striatum, but at 30 DPI there was a significant decrease in AQP4, PDGFRβ and laminin positive areas associated with small blood vessels. Occludin and ZO-1 expression in the striatal tissue was unchanged in all conditions analyzed.ConclusionsThe present study shows a previously-unknown effect of a perinatal administration of a single intracisternal GA injection on BBB permeability and on key components of the neurovascular unit. The results suggest BBB leakage is a pathogenic mechanism and a potential therapeutic target for patients with GA-I.

A role of astrocytes in mediating postnatal neurodegeneration in Glutaric acidemia‐type 1

“…Transmission electron microscopy showed that striatal OLs from rats injected with GA had endoplasmic reticulum (ER) with significant swollen vesicles indicating significant ER stress. Abnormal myelin sheaths and thinner axons were also consistently found in GAI injected animals [35], all resembling the progressive myelin alterations reported in GAI patients [23,51]. Interestingly, GAI induced myelination failure was restricted to the basal ganglia, the most vulnerable brain area in GAI patients [19,20,23].…”

“…Interestingly, EB extravasation through the BBB was internalized in striatal neurons but not in astrocytes or microglial cells. This suggests concurrent membrane anomalies in damaged striatal neurons allowing passive entrance of the dye [35].…”

“…We found that perinatal icv administration of GA to newborn rats provoked serious damage in postnatal myelination that was typically delayed by 2 weeks after the treatment. Such myelination failure was characterized by decreased expression of myelin associated glycoprotein and myelin binding protein in the striatal bundles [35]. Compared with controls, myelination in GA‐animals was weaker and less developed, suggesting a direct correlation with delayed OL differentiation.…”

“…An alternative hypothesis proposes that neurodegeneration in GAI is caused by non‐cell autonomous mechanisms in which interactions between glial or neuronal cells contribute to neurodegenerative processes [14,32–35]. In particular, this hypothesis is based on the observation that increased GA levels in the CNS trigger long lasting astrocyte dysfunction that in turn, subsequently elicits secondary striatal neuronal loss [32], delayed myelination [34], altered BBB permeability and neurovascular unit failure [35]. Taken together, this new proposed mechanism for GAI pathogenesis could lead to new preventive and/or therapeutic approaches for patients.…”

Experimental evidence that bioenergetics disruption is not mainly involved in the brain injury of glutaryl-CoA dehydrogenase deficient mice submitted to lysine overload

“…However, it should be stressed that bioenergetics disruption is capable of inducing secondary excitotoxicity (Danbolt, 2001;Novelli et al, 1988). Therefore, it is conceivable that moderate disturbance of bioenergetics, associated with excitotoxicity (Kolker et al, 2002b;Lagranha et al, 2014;Wajner et al, 2004), oxidative stress Seminotti et al, 2012Seminotti et al, , 2013 and increased blood-brain barrier permeability (Isasi et al, 2014;Zinnanti et al, 2014) may contribute to the brain injury in GA I.…”