Induction of aquaporin 1 but not aquaporin 4 messenger RNA in rat primary brain microvessel endothelial cells in culture

Abstract

Aquaporins (AQPs) are a family of proteins that mediate water transport across cells, but the extent to which they are involved in water transport across endothelial cells of the blood-brain barrier is not clear. Expression of AQP1 and AQP4 in rat brain microvessel endothelial cells was investigated in order to determine whether these isoforms were present and, in particular, to examine the hypothesis that brain endothelial expression of AQPs is dynamic and regulated by astrocytic influences. Reverse-transcriptase-polymerase chain reaction (RT-PCR) and immunocytochemistry showed that AQP1 mRNA and protein are present at very low levels in primary rat brain microvessel endothelial cells, and are up-regulated in passaged cells. Upon passage, endothelial cell expression of mdr1a mRNA is decreased, indicating loss of blood-brain barrier phenotype. In passage 4 endothelial cells, AQP1 mRNA levels are reduced by coculture above rat astrocytes, demonstrating that astrocytic influences are important in maintaining the low levels of AQP1 characteristic of the bloodbrain barrier endothelium. Reverse-transcriptase-PCR revealed very low levels of AQP1 mRNA present in the RBE4 rat brain microvessel endothelial cell line, with no expression detected in primary cultures of rat astrocytes or in the C6 rat glioma cell line. In contrast, AQP4 mRNA is strongly expressed in astrocytes, but no expression is found in primary or passaged brain microvessel endothelial cells, or in RBE4 or C6 cells. Our results support the concept that expression of AQP1, which is seen in many non-brain endothelia, is suppressed in the specialized endothelium of the blood-brain barrier.

Glial-conditional deletion of aquaporin-4 ( Aqp4 ) reduces blood–brain water uptake and confers barrier function on perivascular astrocyte endfeet

“…The immunoblots were in agreement, as no detectable signal remained in brain extracts derived from glialconditional knockout animals. In line with our data, cultured brain microvessel endothelial cells failed to express AQP4 mRNA (15).…”

The role of brain barriers in fluid movement in the CNS: is there a ‘glymphatic’ system?

“…As the flow rate per unit area is low compared with choroid plexus secretion of CSF, it is not possible to demonstrate fluid production from segments of isolated brain capillary, but the net result is to supply local regions of the neuropil with fresh fluid of optimal composition for neural function. Interestingly, the brain endothelium shows down-regulation of the AQP1 highly expressed on non-brain endothelial cells [51], consistent with the need to limit the rate of fluid production to maximize local ionic homeostasis. If a large percentage of the ISF were provided from recirculated CSF as might be implied by the 'glymphatic' hypothesis, this would contain waste products from all the upstream sites exposed to that CSF, a much less suitable medium for optimal neural function.…”

Agmatine Attenuates Brain Edema through Reducing the Expression of Aquaporin-1 after Cerebral Ischemia

“…The disruption of BBB was observed 22 h after 2 h MCAO in this study (Table 1). AQP-1 and AQP-4 are the two members of AQP family known to be expressed in the central nervous system, and it is possible that these proteins contribute to water transport across the BBB (Dolman et al, 2005). In a study of AQP-1 expression in human brain, a small number of microvessels were positively stained, but they were markedly upregulated in endothelium in astrocytomas and metastatic carcinomas (Verkman, 2002).…”