Aquaporin‐4 (AQP4) is the main water channel in brain and is enriched in perivascular astrocyte processes abutting brain microvessels. There is a rich literature on the role of AQP4 in experimental ...stroke. While its role in oedema formation following middle cerebral artery occlusion (MCAO) has been studied extensively, its specific impact on infarct volume remains unclear. This study investigated the effects of total and partial AQP4 deletion on infarct volume in mice subjected to distal medial cerebral artery (dMCAO) occlusion. Compared to MCAO, this model induces smaller infarcts confined to neocortex, and less oedema. We show that AQP4 deletion significantly reduced infarct volume as assessed 1 week after dMCAO, suggesting that the role of AQP4 in stroke goes beyond its effect on oedema formation and dissolution. The reduction in infarct volume was associated with increased astrocyte reactivity in the peri‐infarct areas. No significant differences were observed in the number of microglia among the genotypes. These findings provide new insights in the role of AQP4 in ischaemic injury indicating that AQP4 affects both infarct volume and astrocyte reactivity in the peri‐infarct zone.
Key points
Aquaporin‐4 (AQP4) is the main water channel in brain and is enriched in perivascular astrocyte processes abutting microvessels.
A rich literature exists on the role of AQP4 in oedema formation following middle cerebral artery occlusion (MCAO).
We investigated the effects of total and partial AQP4 deletion on infarct volume in mice subjected to distal medial cerebral artery occlusion (dMCAO), a model inducing smaller infarcts confined to neocortex and less oedema compared to MCAO.
AQP4 deletion significantly reduced infarct volume 1 week after dMCAO, suggesting a broader role for AQP4 in stroke beyond oedema formation.
The reduction in infarct volume was associated with increased astrocyte reactivity in the peri‐infarct areas, while no significant differences were observed in the number of microglia among the genotypes.
These findings provide new insights into the role of AQP4 in stroke, indicating that AQP4 affects both infarct volume and astrocyte reactivity in the peri‐infarct zone.
figure legend Infarct size and astrocyte reactivity in wild‐type (WT) mice compared with AQP4 knockout (AQP4−/−) littermates, following distal medial cerebral artery occlusion. Genetic deletion of Aqp4 (AQP4−/−) results in reduced infarct size and heightened astrocyte reactivity, as evidenced by increased glial fibrillary acidic protein (GFAP) immunostaining in the infarct border zone.
Proper function of the retina depends heavily on a specialized form of retinal glia called Müller cells. These cells carry out important homeostatic functions that are contingent on their polarized ...nature. Specifically, the Müller cell endfeet that contact retinal microvessels and the corpus vitreum show a tenfold higher concentration of the inwardly rectifying potassium channel K
4.1 than other Müller cell plasma membrane domains. This highly selective enrichment of K
4.1 allows K+ to be siphoned through endfoot membranes in a special form of spatial buffering. Here, we show that K
4.1 is enriched in endfoot membranes through an interaction with β1-syntrophin. Targeted disruption of this syntrophin caused a loss of K
4.1 from Müller cell endfeet without affecting the total level of K
4.1 expression in the retina. Targeted disruption of α1-syntrophin had no effect on K
4.1 localization. Our findings show that the K
4.1 aggregation that forms the basis for K+ siphoning depends on a specific syntrophin isoform that colocalizes with K
4.1 in Müller endfoot membranes.
Aquaporin-4 (AQP4) is the predominant water channel in the brain and is expressed in high density in astrocytes. By fluxing water along osmotic gradients, AQP4 contributes to brain volume and ion ...homeostasis. Here we ask whether deletion of
Aqp4
leads to upregulation of the gap junctional proteins connexin-43 (Cx43) and connexin-30 (Cx30). These molecules couple adjacent astrocytes to each other and allow water and ions to redistribute within the astrocyte syncytium. Immunogold analysis of parietal cortex and hippocampus showed that the number of gap junctions per capillary profile is increased in AQP4 knockout (AQP4 KO) mice. The most pronounced changes were observed for Cx43 in hippocampus where the number of connexin labeled gap junctions increased by 100% following AQP4 KO. Western blot analysis of whole tissue homogenates showed no change in the amount of Cx43 or Cx30 protein after AQP4 KO. However, AQP4 KO led to a significant increase in the amount of Cx43 in a Triton X-100 insoluble fraction. This fraction is associated with connexin assembly into gap junctional plaques in the plasma membrane. In line with our immunoblot data, RT-qPCR showed no significant increase in Cx43 and Cx30 mRNA levels after AQP4 KO. Our findings suggest that AQP4 KO leads to increased aggregation of Cx43 into gap junctions and provide a putative mechanistic basis for the enhanced tracer coupling in hippocampi of AQP4 KO mice. The increased number of gap junctions in AQP4 deficient mice may explain why
Aqp4
deletion has rather modest effects on brain volume and K
+
homeostasis.
Proper function of the retina depends heavily on a specialized form of retinal glia called Müller cells. These cells carry out important homeostatic functions that are contingent on their polarized ...nature. Specifically, the Müller cell endfeet that contact retinal microvessels and the corpus vitreum show a tenfold higher concentration of the inwardly rectifying potassium channel Kir4.1 than other Müller cell plasma membrane domains. This highly selective enrichment of Kir4.1 allows K+ to be siphoned through endfoot membranes in a special form of spatial buffering. Here, we show that Kir4.1 is enriched in endfoot membranes through an interaction with β1‐syntrophin. Targeted disruption of this syntrophin caused a loss of Kir4.1 from Müller cell endfeet without affecting the total level of Kir4.1 expression in the retina. Targeted disruption of α1‐syntrophin had no effect on Kir4.1 localization. Our findings show that the Kir4.1 aggregation that forms the basis for K+ siphoning depends on a specific syntrophin isoform that colocalizes with Kir4.1 in Müller endfoot membranes.
Proper function of the retina depends heavily on a specialized form of retinal glia called Müller cells. These cells carry out important homeostatic functions that are contingent on their polarized ...nature. Specifically, the Müller cell endfeet that contact retinal microvessels and the corpus vitreum show a tenfold higher concentration of the inwardly rectifying potassium channel Kir4.1 than other Müller cell plasma membrane domains. This highly selective enrichment of Kir4.1 allows K+ to be siphoned through endfoot membranes in a special form of spatial buffering. Here, we show that Kir4.1 is enriched in endfoot membranes through an interaction with β1‐syntrophin. Targeted disruption of this syntrophin caused a loss of Kir4.1 from Müller cell endfeet without affecting the total level of Kir4.1 expression in the retina. Targeted disruption of α1‐syntrophin had no effect on Kir4.1 localization. Our findings show that the Kir4.1 aggregation that forms the basis for K+ siphoning depends on a specific syntrophin isoform that colocalizes with Kir4.1 in Müller endfoot membranes.
Main Points
Inwardly rectifying K+ channel Kir4.1 is concentrated at the perivascular and subvitreal Müller cell endfeet.
Targeted deletion of β1‐syntrophin leads to significant loss of Kir4.1 at the Müller cell endfeet without changing its expression levels.
Biallelic PIGT variants were previously reported in seven patients from three families with Multiple Congenital Anomalies-Hypotonia Seizures Syndrome 3 (MCAHS3), characterized by epileptic ...encephalopathy, hypotonia, global developmental delay/intellectual disability, cerebral and cerebellar atrophy, craniofacial dysmorphisms, and skeletal, ophthalmological, cardiac, and genitourinary abnormalities. We report a novel homozygous PIGT missense variant c.1079G>T (p.Gly360Val) in two brothers with several of the typical features of MCAHS3, but in addition, pyramidal tract neurological signs. Notably, they are the first patients with MCAHS3 without skeletal, cardiac, or genitourinary anomalies. PIGT encodes a crucial subunit of the glycosylphosphatidylinositol (GPI) transamidase complex, which catalyzes the attachment of proteins to GPI-anchors, attaching the proteins to the cell membrane. In vitro studies in cells from the two brothers showed reduced levels of GPI-anchors and GPI-anchored proteins on the cell surface, supporting the pathogenicity of the novel PIGT variant.
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