AQP4
Aquaporin-4 (AQP-4) (Mercurial-insensitive water channel) (MIWC) (WCH4)
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Aging is associated with impairment of the paravascular pathway caused by the activation of astrocytes and depolarization of protein aquaporin‑4 (AQP4) water channels, resulting in the accumulation of protein waste, including amyloid β (Aβ), in the brain parenchyma. The secreted glycoprotein slit guidance ligand 2 (Slit2) is important in regulating the function of the central nervous system and inflammatory response process. In the present study, 15‑month‑old Slit2 overexpression transgenic mice (Slit2‑Tg mice) and two‑photon fluorescence microscopy were used to evaluate the dynamic clearance of the paravascular pathway and the integrity of the blood‑brain barrier (BBB). The reactivity of astrocytes, polarity of AQP4 and deposition of Aβ in the brain parenchyma were analyzed by immunofluorescence. A Morris water maze test was used to examine the effect of Slit2 on spatial memory cognition in aging mice. It was found that the overexpression of Slit2 improved the clearance of the paravascular pathway by inhibiting astrocyte activation and maintaining AQP4 polarity on the astrocytic endfeet in Slit2‑Tg mice. In addition, Slit2 restored the disruption of the BBB caused by aging. The accumulation of Aβ was significantly reduced in the brain of Slit2‑Tg mice. Furthermore, the water maze experiment showed that Slit2 improved spatial memory cognition in the aging mice. These results indicated that Slit2 may have the potential to be used in the prevention and treatment of neurodegenerative diseases in the elderly.
MeSH Terms
- Aging
- Animals
- Aquaporin 4
- Astrocytes
- Blood-Brain Barrier
- Brain
- Female
- Fluorescent Antibody Technique
- Humans
- Intercellular Signaling Peptides and Proteins
- Male
- Maze Learning
- Mice
- Mice, Inbred C57BL
- Mice, Transgenic
- Nerve Tissue Proteins
- Spatial Memory
Ageing-related tau astrogliopathy (ARTAG) appears in subependymal, subpial, perivascular, white matter (WM) and grey matter (GM) locations. Physical effects, blood-brain barrier dysfunction and blood- or vessel-related factors have been considered as aetiology. As connexin-43 (Cx43) and aquaporin-4 (AQP4) are related to these, we hypothesized that their immunoreactivity (IR) varies with ARTAG in a location-specific manner. We performed a morphometric immunohistochemical study measuring the densities of IR of Cx43, AQP4, AT8 (phospho-tau) and glial fibrillar acidic protein (GFAP). We analysed the amygdala and hippocampus in age-matched cases with (n = 19) and without (n = 20) ARTAG in each of the locations it aggregates. We show a dramatic increase (>6-fold; P < 0.01) of Cx43 density of IR in ARTAG cases correlating strongly with AT8 density of IR, irrespective of the presence of neuronal tau pathology or reactive gliosis measured by GFAP density of IR, in the GM. In contrast, AQP4 density of IR was increased only in the WM and GM, and was associated with increased AT8 density of IR only in WM and perivascular areas. Our study reveals distinctive astroglial responses in each of the locations associated with ARTAG. Our observations support the concept that factors related to brain-fluid interfaces and water-ion imbalances most likely play a role in the generation of ARTAG. As Cx43 is crucial for maintaining neuronal homeostasis, the ARTAG-dependent increase of Cx43 density of IR suggests that the development of ARTAG in the GM most likely indicates an early response to the degeneration of neurons.
MeSH Terms
- Aged
- Aged, 80 and over
- Aging
- Aquaporin 4
- Astrocytes
- Biomarkers
- Brain
- Connexin 43
- Female
- Humans
- Male
- Tauopathies
Keywords
- Tau
- ageing-related tau astrogliopathy
- aquaporin-4
- blood-brain barrier
- connexin 43
- glial fibrillar acidic protein
Age is characterized by chronic inflammation, leading to synaptic dysfunction and dementia because the clearance of protein waste is reduced. The clearance of proteins depends partly on the permeation of the blood-brain barrier (BBB) or on the exchange of water and soluble contents between the cerebrospinal fluid (CSF) and the interstitial fluid (ISF). A wealth of evidence indicates that physical exercise improves memory and cognition in neurodegenerative diseases during aging, such as Alzheimer's disease (AD), but the influence of physical training on glymphatic clearance, BBB permeability and neuroinflammation remains unclear. In this study, glymphatic clearance and BBB permeability were evaluated in aged mice using [i]in vivo[/i] two-photon imaging. The mice performed voluntary wheel running exercise and their water-maze cognition was assessed; the expression of the astrocytic water channel aquaporin 4 (AQP4), astrocyte and microglial activation, and the accumulation of amyloid beta (Aβ) were evaluated with immunofluorescence or an enzyme-linked immunosorbent assay (ELISA); synaptic function was investigated with [i]Thy1[/i]-green fluorescent protein (GFP) transgenic mice and immunofluorescent staining. Voluntary wheel running significantly improved water-maze cognition in the aged mice, accelerated the efficiency of glymphatic clearance, but which did not affect BBB permeability. The numbers of activated astrocytes and microglia decreased, AQP4 expression increased, and the distribution of astrocytic AQP4 was rearranged. Aβ accumulation decreased, whereas dendrites, dendritic spines and postsynaptic density protein (PSD95) increased. Our study suggests that voluntary wheel running accelerated glymphatic clearance but not BBB permeation, improved astrocytic AQP4 expression and polarization, attenuated the accumulation of amyloid plaques and neuroinflammation, and ultimately protected mice against synaptic dysfunction and a decline in spatial cognition. These data suggest possible mechanisms for exercise-induced neuroprotection in the aging brain.
Keywords
- aging
- inflammation
- interstitial fluid
- paravascular space
- spatial memory
- wheel running
Amyloid-β deposition in senile plaques is one of the main pathological changes in Alzheimer's disease (AD). We previously reported that aquaporin-4 (AQP4) is redistributed within the astrocytes in cerebral amyloid angiopathy in the tg-ArcSwe mouse model of AD, suggesting that AQP4 may participate in amyloid-β deposition. However, the role of AQP4 in plaque formation is not currently clear. The objective of the current study was to explore the AQP4 distribution within plaques in the tg-ArcSwe mice in more depth by the combined application of immunofluorescence cytochemistry and immunogold electron microscopy. In addition, the astrocyte marker, glial fibrillary acidic protein (GFAP), was studied in association with AQP4. We demonstrated a robust upregulation of AQP4 expression in areas of plaques. Compared to GFAP, AQP4 appeared predominantly at later stages of plaque formation, in older mice, and within the processes of astrocytes. In combination with GFAP, AQP4 differentiated plaques into three progression stages under light microscopy. This suggests that AQP4 expression was associated with amyloid deposition and astrocyte pathology in the Tg-ArcSwe mouse model of AD. This provides novel proof for the involvement of AQP4 in the process of amyloid deposition in AD.
MeSH Terms
- Aging
- Alzheimer Disease
- Amyloid beta-Peptides
- Amyloid beta-Protein Precursor
- Animals
- Aquaporin 4
- Astrocytes
- Brain
- Disease Models, Animal
- Disease Progression
- Glial Fibrillary Acidic Protein
- Humans
- Mice, Inbred C57BL
- Mice, Transgenic
- Peptide Fragments
- Plaque, Amyloid
Keywords
- Alzheimer’s disease
- aquaporin 4
- astrocyte
- glial fibrillary acidic protein
- senile plaques
Hydrocephalus is a complex neurological disorder with a pervasive impact on the central nervous system. Previous work has demonstrated derangements in the biochemical profile of cerebrospinal fluid (CSF) in hydrocephalus, particularly in infants and children, in whom neurodevelopment is progressing in parallel with concomitant neurological injury. The objective of this study was to examine the CSF of children with congenital hydrocephalus (CHC) to gain insight into the pathophysiology of hydrocephalus and identify candidate biomarkers of CHC with potential diagnostic and therapeutic value. CSF levels of amyloid precursor protein (APP) and derivative isoforms (sAPPα, sAPPβ, Aβ42), tau, phosphorylated tau (pTau), L1CAM, NCAM-1, aquaporin 4 (AQP4), and total protein (TP) were measured by ELISA in 20 children with CHC. Two comparative groups were included: age-matched controls and children with other neurological diseases. Demographic parameters, ventricular frontal-occipital horn ratio, associated brain malformations, genetic alterations, and surgical treatments were recorded. Logistic regression analysis and receiver operating characteristic curves were used to examine the association of each CSF protein with CHC. CSF levels of APP, sAPPα, sAPPβ, Aβ42, tau, pTau, L1CAM, and NCAM-1 but not AQP4 or TP were increased in untreated CHC. CSF TP and normalized L1CAM levels were associated with FOR in CHC subjects, while normalized CSF tau levels were associated with FOR in control subjects. Predictive ability for CHC was strongest for sAPPα, especially in subjects ≤12 months of age (p<0.0001 and AUC = 0.99), followed by normalized sAPPβ (p = 0.0001, AUC = 0.95), tau, APP, and L1CAM. Among subjects ≤12 months, a normalized CSF sAPPα cut-point of 0.41 provided the best prediction of CHC (odds ratio = 528, sensitivity = 0.94, specificity = 0.97); these infants were 32 times more likely to have CHC. CSF proteins such as sAPPα and related proteins hold promise as biomarkers of CHC in infants and young children, and provide insight into the pathophysiology of CHC during this critical period in neurodevelopment.
MeSH Terms
- Aging
- Amyloid beta-Protein Precursor
- Biomarkers
- Child
- Female
- Humans
- Hydrocephalus
- Infant
- Infant, Newborn
- Male
Cognitive impairment and dementia, including Alzheimer disease (AD), are common within the aging population, yet the factors that render the aging brain vulnerable to these processes are unknown. Perivascular localization of aquaporin-4 (AQP4) facilitates the clearance of interstitial solutes, including amyloid-β, through the brainwide network of perivascular pathways termed the glymphatic system, which may be compromised in the aging brain. To determine whether alterations in AQP4 expression or loss of perivascular AQP4 localization are features of the aging human brain and to define their association with AD pathology. Expression of AQP4 was analyzed in postmortem frontal cortex of cognitively healthy and histopathologically confirmed individuals with AD by Western blot or immunofluorescence for AQP4, amyloid-β 1-42, and glial fibrillary acidic protein. Postmortem tissue and clinical data were provided by the Oregon Health and Science University Layton Aging and Alzheimer Disease Center and Oregon Brain Bank. Postmortem tissue from 79 individuals was evaluated, including cognitively intact "young" individuals aged younger than 60 years (range, 33-57 years), cognitively intact "aged" individuals aged older than 60 years (range, 61-96 years) with no known neurological disease, and individuals older than 60 years (range, 61-105 years) of age with a clinical history of AD confirmed by histopathological evaluation. Forty-eight patient samples (10 young, 20 aged, and 18 with AD) underwent histological analysis. Sixty patient samples underwent Western blot analysis (15 young, 24 aged, and 21 with AD). Expression of AQP4 protein, AQP4 immunoreactivity, and perivascular AQP4 localization in the frontal cortex were evaluated. Expression of AQP4 was associated with advancing age among all individuals (R2 = 0.17; P = .003). Perivascular AQP4 localization was significantly associated with AD status independent of age (OR, 11.7 per 10% increase in localization; z = -2.89; P = .004) and was preserved among eldest individuals older than 85 years of age who remained cognitively intact. When controlling for age, loss of perivascular AQP4 localization was associated with increased amyloid-β burden (R2 = 0.15; P = .003) and increasing Braak stage (R2 = 0.14; P = .006). In this study, altered AQP4 expression was associated with aging brains. Loss of perivascular AQP4 localization may be a factor that renders the aging brain vulnerable to the misaggregation of proteins, such as amyloid-β, in neurodegenerative conditions such as AD.
MeSH Terms
- Adult
- Age Factors
- Aged
- Aged, 80 and over
- Aging
- Alzheimer Disease
- Amyloid beta-Peptides
- Analysis of Variance
- Aquaporin 4
- Cognition
- Diagnosis
- Female
- Frontal Lobe
- Gene Expression Regulation
- Glial Fibrillary Acidic Protein
- Humans
- Male
- Middle Aged
- Peptide Fragments
- Plaque, Amyloid
Aquaporin (AQP) 1 and AQP 4 are expressed in human heart and several studies have been focused on these two aquaporins. For this purpose, the present study is aimed to research the effects of aging on AQP 1 and AQP 4 in heart tissue. In this study, 14 Balb/C type white mice were used. Animals were divided into two equal groups. Group I consisted of 2-month-old young animals (n=7), and group II consisted of 18-month-old animals (n=7). To determine the AQP1 and AQP4 expression in the myocardium, the heart tissue was removed to perform western blotting and immunohistochemical and histopathological evaluations. Muscle fibers of the heart in aged animals were more irregular and loosely organized in hematoxylin-eosin (H-E) stained sections. Hscore analysis revealed that AQP1 and AQP4 immunoreactivity significantly increased in heart tissues of old mice compared with those of young mice (p<0.001). In addition, AQP1 and AQP4 protein expressions in the tissues of old animals were increased significantly according to western blot analysis (p=0.018 and p<0.001 for AQP1 and AQP4, respectively). Increased AQP1 and AQP4 levels in the heart tissue may be correlated with the maintenance of water and electrolytes balance, which decreases with aging. In this context, it might be the result of a compensatory response to decreased AQP4 functions. In addition, this increase with aging as demonstrated in our study might be one of the factors that increases the tendency of ischemia in elder people.
MeSH Terms
- Aging
- Animals
- Aquaporin 1
- Aquaporin 4
- Female
- Mice
- Mice, Inbred BALB C
- Myocardial Ischemia
- Myocardium
White matter hyperintensities as seen on brain T2-weighted magnetic resonance imaging are associated with varying degrees of cognitive dysfunction in stroke, cerebral small vessel disease and dementia. The pathophysiological mechanisms within the white matter accounting for cognitive dysfunction remain unclear. With the hypothesis that gliovascular interactions are impaired in subjects with high burdens of white matter hyperintensities, we performed clinicopathological studies in post-stroke survivors, who had exhibited greater frontal white matter hyperintensities volumes that predicted shorter time to dementia onset. Histopathological methods were used to identify substrates in the white matter that would distinguish post-stroke demented from post-stroke non-demented subjects. We focused on the reactive cell marker glial fibrillary acidic protein (GFAP) to study the incidence and location of clasmatodendrosis, a morphological attribute of irreversibly injured astrocytes. In contrast to normal appearing GFAP astrocytes, clasmatodendrocytes were swollen and had vacuolated cell bodies. Other markers such as aldehyde dehydrogenase 1 family, member L1 (ALDH1L1) showed cytoplasmic disintegration of the astrocytes. Total GFAP cells in both the frontal and temporal white matter were not greater in post-stroke demented versus post-stroke non-demented subjects. However, the percentage of clasmatodendrocytes was increased by >2-fold in subjects with post-stroke demented compared to post-stroke non-demented subjects (P = 0.026) and by 11-fold in older controls versus young controls (P < 0.023) in the frontal white matter. High ratios of clasmotodendrocytes to total astrocytes in the frontal white matter were consistent with lower Mini-Mental State Examination and the revised Cambridge Cognition Examination scores in post-stroke demented subjects. Double immunofluorescent staining showed aberrant co-localization of aquaporin 4 (AQP4) in retracted GFAP astrocytes with disrupted end-feet juxtaposed to microvessels. To explore whether this was associated with the disrupted gliovascular interactions or blood-brain barrier damage, we assessed the co-localization of GFAP and AQP4 immunoreactivities in post-mortem brains from adult baboons with cerebral hypoperfusive injury, induced by occlusion of three major vessels supplying blood to the brain. Analysis of the frontal white matter in perfused brains from the animals surviving 1-28 days after occlusion revealed that the highest intensity of fibrinogen immunoreactivity was at 14 days. At this survival time point, we also noted strikingly similar redistribution of AQP4 and GFAP astrocytes transformed into clasmatodendrocytes. Our findings suggest novel associations between irreversible astrocyte injury and disruption of gliovascular interactions at the blood-brain barrier in the frontal white matter and cognitive impairment in elderly post-stroke survivors. We propose that clasmatodendrosis is another pathological substrate, linked to white matter hyperintensities and frontal white matter changes, which may contribute to post-stroke or small vessel disease dementia.
MeSH Terms
- Aged
- Aged, 80 and over
- Aging
- Aldehyde Dehydrogenase
- Animals
- Aquaporin 4
- Astrocytes
- Blood-Brain Barrier
- Case-Control Studies
- Cognition Disorders
- Dementia
- Female
- Frontal Lobe
- Glial Fibrillary Acidic Protein
- Humans
- Magnetic Resonance Imaging
- Male
- Middle Aged
- Neuroimaging
- Neuropsychological Tests
- Oxidoreductases Acting on CH-NH Group Donors
- Papio anubis
- Stroke
- White Matter
Keywords
- ageing
- blood–brain barrier
- clasmatodendrocyte
- post-stroke dementia
- white matter
Traumatic brain injury (TBI) is one of the major risk factors of dementia, aging, and cognitive impairments, etc. We have previously reported that expression of the astrocytic glutamate transporter GLT-1/EAAT2 is downregulated in the pericontusional cortex of adult and old mice in post-TBI time-dependent manner, and the process of decline starts before in old than in adult TBI mice. However, relationship between age- and TBI-dependent alterations in GLT-1/EAAT2 expression and interactions of transcription factors NF-κB and N-myc with their cognate GLT-1/EAAT2 promoter sequences, an important step of its transcriptional control, is not known. To understand this, we developed TBI mouse model by modified chronic head injury (CHI) method, analyzed expression of GFAP, TNF-α, and AQP4 by RT-PCR for its validation, and analyzed interactions of NF-κB and N-myc with GLT-1/EAAT2 promoter sequences by electrophoretic mobility shift assay (EMSA). Our EMSA data revealed that interactions of NF-κB and N-myc with GLT-1/EAAT2 promoter sequences was significantly elevated in the ipsi-lateral cortex of both adult and old TBI mice in post-TBI time-dependent manner; however, these interactions started immediately in the old compared to that in adult TBI mice, which could be attributed to our previously reported age- and post-TBI time-dependent differential expression of GLT-1/EAAT2 in the pericontusional cortex.
MeSH Terms
- Aging
- Animals
- Aquaporin 4
- Brain Injuries, Traumatic
- Cerebral Cortex
- Electrophoretic Mobility Shift Assay
- Excitatory Amino Acid Transporter 2
- Glial Fibrillary Acidic Protein
- Male
- Mice
- N-Myc Proto-Oncogene Protein
- NF-kappa B
- Promoter Regions, Genetic
- Protein Binding
- RNA, Messenger
- Tumor Necrosis Factor-alpha
Keywords
- Aging
- GLT-1/EAAT2
- N-myc
- NF-κB
- Traumatic brain injury
In the brain, protein waste removal is partly performed by paravascular pathways that facilitate convective exchange of water and soluble contents between cerebrospinal fluid (CSF) and interstitial fluid (ISF). Several lines of evidence suggest that bulk flow drainage via the glymphatic system is driven by cerebrovascular pulsation, and is dependent on astroglial water channels that line paravascular CSF pathways. The objective of this study was to evaluate whether the efficiency of CSF-ISF exchange and interstitial solute clearance is impaired in the aging brain. CSF-ISF exchange was evaluated by in vivo and ex vivo fluorescence microscopy and interstitial solute clearance was evaluated by radiotracer clearance assays in young (2-3 months), middle-aged (10-12 months), and old (18-20 months) wild-type mice. The relationship between age-related changes in the expression of the astrocytic water channel aquaporin-4 (AQP4) and changes in glymphatic pathway function was evaluated by immunofluorescence. Advancing age was associated with a dramatic decline in the efficiency of exchange between the subarachnoid CSF and the brain parenchyma. Relative to the young, clearance of intraparenchymally injected amyloid-β was impaired by 40% in the old mice. A 27% reduction in the vessel wall pulsatility of intracortical arterioles and widespread loss of perivascular AQP4 polarization along the penetrating arteries accompanied the decline in CSF-ISF exchange. We propose that impaired glymphatic clearance contributes to cognitive decline among the elderly and may represent a novel therapeutic target for the treatment of neurodegenerative diseases associated with accumulation of misfolded protein aggregates.
MeSH Terms
- Aging
- Animals
- Aquaporin 4
- Brain
- Cerebrovascular Circulation
- Female
- Male
- Metabolic Clearance Rate
- Mice
- Mice, Inbred C57BL
- Microscopy, Fluorescence, Multiphoton
- Neuroglia
Few data are available for patients with a late onset (≥ 50 years) of neuromyelitis optica (LONMO) or neuromyelitis optica spectrum disease (LONMOSD), defined by an optic neuritis/longitudinally extensive transverse myelitis with aquaporin-4 antibodies (AQP4-Ab). To characterize LONMO and LONMOSD, and to analyze their predictive factors of disability and death. We identified 430 patients from four cohorts of NMO/NMOSD in France, Germany, Turkey and UK. We extracted the late onset patients and analyzed them for predictive factors of disability and death, using the Cox proportional model. We followed up on 63 patients with LONMO and 45 with LONMOSD during a mean of 4.6 years. This LONMO/LONMOSD cohort was mainly of Caucasian origin (93%), women (80%), seropositive for AQP4-Ab (85%) and from 50 to 82.5 years of age at onset. No progressive course was noted. At last follow-up, the median Expanded Disability Status Scale (EDSS) scores were 5.5 and 6 in the LONMO and LONMOSD groups, respectively. Outcome was mainly characterized by motor disability and relatively good visual function. At last follow-up, 14 patients had died, including seven (50%) due to acute myelitis and six (43%) because of opportunistic infections. The EDSS 4 score was independently predicted by an older age at onset, as a continuous variable after 50 years of age. Death was predicted by two independent factors: an older age at onset and a high annualized relapse rate. LONMO/LONMOSD is particularly severe, with a high rate of motor impairment and death.
MeSH Terms
- Age of Onset
- Aged
- Aged, 80 and over
- Aquaporin 4
- Autoantibodies
- Biomarkers
- Cause of Death
- Chi-Square Distribution
- Disability Evaluation
- Disease Progression
- Europe
- Female
- Humans
- Kaplan-Meier Estimate
- Male
- Middle Aged
- Motor Activity
- Multivariate Analysis
- Neuromyelitis Optica
- Predictive Value of Tests
- Prognosis
- Proportional Hazards Models
- Retrospective Studies
- Risk Factors
- Severity of Illness Index
- Time Factors
Keywords
- Aging
- aquaporin antibody
- aquaporin-4
- late onset
- morbidity
- mortality
- neuromyelitis optica
- prognosis
- spectrum disorders
To investigate the effects of aquaporin-4 (AQP4) gene knockout on the behavior changes and cerebral morphology during aging in mice,and to compare that of young and aged mice between AQP4 knockout mice (AQP4(-/-)) and wild type mice (AQP4( / )). Fifty-eight CD-1 mice were divided into four groups: young (2-3 months old) AQP4(-/-), aged (17-19 months old) AQP4(-/-), young AQP4( / ) and aged AQP4( / ). The activity levels and exploring behavior of mice were tested in open field. The neurons were stained with toluidine blue and NeuN, the astrocytes and microglia were stained with GFAP and Iba-1, respectively. The morphological changes of neuron, astrocyte and microglia were then analyzed. Compared with young mice, the total walking distance in open field of aged AQP4( / ) mice and aged AQP4(-/-) mice decreased 41.2% and 44.1%, respectively (P<0.05); while there was no difference in the ratio of distance and retention time in the central area of open field. The density of neuron in cortex of aged AQP4( / ) mice and aged AQP4(-/-) mice decreased 19.6% and 15.8%, respectively (P<0.05), while there was no difference in the thickness of neuron cell body in hippocampus CA1 region. The density of astrocyte in hippocampus CA3 region of aged AQP4( / ) mice and aged AQP4(-/-) mice increased 57.7% and 64.3%, respectively (P<0.001), while there was no difference in the area of astrocyte. The area of microglia in hippocampus CA3 region of aged AQP4( / ) mice and aged AQP4(-/-) mice increased 46.9% and 52.0%, respectively (P<0.01), while there was no difference in the density of microglia. Compared with AQP4( / ) mice, the young and aged AQP4(-/-) mice showed smaller area of astrocyte in hippocampus CA3 region, reduced 18.0% in young mice and 23.6% in aged mice. There was no difference between AQP4( / ) mice and AQP4(-/-) mice for other observed indexes. AQP4 may be involved in change of astrocyte and astrocyte-related behaviors during aging. AQP4 gene knockout may have limited effects on the change of neuron, microglia and most neuronal behaviors in aging process.
MeSH Terms
- Aging
- Animals
- Aquaporin 4
- Astrocytes
- Behavior, Animal
- Brain
- Female
- Male
- Mice
- Mice, Knockout
- Microglia
- Neurons
Collaborating on studies of subchronic daily intoxication in juvenile and adult rats, we examined whether the repetitive ethanol treatments at these two life stages altered levels of key neuroinflammation-associated proteins-aquaporin-4 (AQP4), certain phospholipase A2 (PLA2) enzymes, PARP-1 and caspase-3-in hippocampus (HC) and entorhinal cortex (EC). Significant changes in the proteins could implicate activation of specific neuroinflammatory signaling pathways in these rats as well as in severely binge-intoxicated adult animals that are reported to incur degeneration of vulnerable neurons in HC and EC. Male Wistar rats, ethanol-intoxicated (3 g/kg i.p.) once daily for 6 days over an 8-day interval beginning at 37 days old and repeated at age 68-75 days, were sacrificed 1 h after the day 75 dose (blood ethanol, 200- 230 mg/dl). Analysis of HC with an immunoblot technique showed that AQP4, Ca( 2)-dependent PLA2 (cPLA2 IVA), phosphorylated (activated) p-cPLA2, cleaved (89 kD) PARP (c-PARP), and caspase-3 levels were significantly elevated over controls, whereas Ca( 2)-independent PLA2 (iPLA2 VIA) was reduced ∼70%; however, cleaved caspase-3 was undetectable. In the EC, AQP4 was unchanged, but cPLA2 and p-cPLA2 were significantly increased while iPLA2 levels were diminished (∼40%) similar to HC, although just outside statistical significance (p = 0.06). In addition, EC levels of PARP-1 and c-PARP were significantly increased. The ethanol-induced activation of cPLA2 in association with reduced iPLA2 mirrors PLA2 changes in reports of neurotrauma and also of dietary omega-3 fatty acid depletion. Furthermore, the robust PARP-1 elevations accompanied by negligible caspase-3 activation indicate that repetitive ethanol intoxication may be potentiating non-apoptotic neurodegenerative processes such as parthanatos. Overall, the repetitive ethanol treatments appeared to instigate previously unappreciated neuroinflammatory pathways in vivo. The data provide insights into mechanisms of binge ethanol abuse that might suggest new therapeutic approaches to counter neurodegeneration and dementia.
MeSH Terms
- Aging
- Alcoholic Intoxication
- Animals
- Aquaporin 4
- Caspase 3
- Entorhinal Cortex
- Group VI Phospholipases A2
- Hippocampus
- Male
- Phospholipases A2, Cytosolic
- Poly (ADP-Ribose) Polymerase-1
- Poly(ADP-ribose) Polymerases
- Rats
- Rats, Wistar
- Signal Transduction
Glial cells, besides participating as passive supporting matrix, are also proposed to be involved in the optimization of the interstitial space for synaptic transmission by tight control of ionic and water homeostasis. In adult mouse brain, inwardly rectifying K (Kir4.1) and aquaporin-4 (AQP4) channels localize to astroglial endfeets in contact with brain microvessels and glutamate synapses, optimizing clearance of extracellular K( ) and water from the synaptic layers. However, it is still unclear whether there is an age-dependent difference in the expressions of Kir4.1 and AQP4 channels specifically during postnatal development and aging when various marked changes occur in brain and if these changes region specific. RT-PCR and immunoblotting was conducted to compare the relative expression of Kir4.1 and AQP4 mRNA and protein in the early and mature postnatal (0-, 15-, 45-day), adult (20-week), and old age (70-week) mice cerebral and cerebellar cortices. Expressions of Kir4.1 and AQP4 mRNA and protein are very low at 0-day. A pronounced and continuous increase was observed by mature postnatal ages (15-, 45-days). However, in the 70-week-old mice, expressions are significantly up-regulated as compared to 20-week-old mice. Both genes follow the same age-related pattern in both cerebral and cerebellar cortices. The time course and expression pattern suggests that Kir4.1 and AQP4 channels may play an important role in brain K( ) and water homeostasis in early postnatal weeks after birth and during aging.
MeSH Terms
- Aging
- Animals
- Brain
- Electrophoresis, Polyacrylamide Gel
- Male
- Mice
- Mice, Inbred AKR
- Neuroglia
- Potassium Channels, Inwardly Rectifying
- RNA, Messenger
- Reverse Transcriptase Polymerase Chain Reaction
- Up-Regulation
Sodium-sensitive hypertension is caused by renal tubular dysfunction, leading to increased retention of sodium and water. Previous findings have suggested that single-nucleotide polymorphisms of the cytoskeletal protein, α-adducin, are associated with increased membrane expression of the Na/K pump and abnormal renal sodium transport in Milan hypertensive strain (MHS) rats and in humans. However, the possible contribution of renal aquaporins (AQPs) to water retention remains undefined in MHS rats. Kidneys from MHS rats were analyzed and compared with those from age-matched Milan normotensive strain (MNS) animals by quantitative-PCR, immunoblotting, and immunoperoxidase. Endocytosis assay was performed on renal cells stably expressing AQP4 and co-transfected either with wild-type normotensive (NT) or with mutated hypertensive (HT) α-adducin. Semiquantitative immunoblotting revealed that AQP1 abundance was significantly decreased only in HT MHS whereas AQP2 was reduced in both young pre-HT and adult-HT animals. On the other hand, AQP4 was dramatically upregulated in MHS regardless of the age. These results were confirmed by immunoperoxidase microscopy. Endocytosis assays clearly showed that the expression of mutated adducin strongly reduced the rate of constitutive AQP4 endocytosis, thereby increasing its abundance at the plasma membrane. We provide here the first evidence that AQP1, AQP2, and AQP4 are dysregulated in the kidneys of MHS animals. In particular, we provide evidence that α-adducin mutations may be responsible for AQP4 upregulation. The downregulation of AQP1 and AQP2 and the upregulation of AQP4 may be relevant for the onset and maintenance of salt-sensitive hypertension.
MeSH Terms
- Absorption
- Aging
- Animals
- Aquaporin 4
- Aquaporins
- Blood Pressure
- Calmodulin-Binding Proteins
- Disease Models, Animal
- Endocytosis
- Hypertension
- Kidney
- Male
- Polymorphism, Genetic
- Rats
- Rats, Mutant Strains
- Salt Tolerance
- Water
Aquaporins (AQP) 1 and 4 are water channel proteins localized respectively at the level of the blood-cerebrospinal fluids (CSF) and blood brain (BBB) barriers. These barriers represent the sites of exchange between blood and nervous tissue and between blood, choroid plexus and CSF in brain ventricles respectively. Damage of these barriers may alter transfer of substances between blood and nervous tissue. In spontaneously hypertensive rats (SHR) chronic hypertension may induce BBB dysfunction and pronounced defects in the integrity of the blood-CSF barrier. AQP1 is expressed in the apical membrane of choroid plexus epithelium. AQP4 is expressed by astrocyte foot processes near blood vessels. The present study has assessed the expression of AQP1 and AQP4 in the brain of SHR in pre-hypertensive (2 months of age), developing hypertension (4 months of age) and established hypertension (6 months of age) stages. Age-matched Wistar-Kyoto (WKY) rats were used as normotensive reference group. AQP1 expression is increased in choroid plexus epithelium of 6-month-old SHR. An increased expression of AQP4 was found in frontal cortex, striatum, and hippocampus of 4- and 6-month-old SHR compared to younger cohorts and age-matched WKY rats. These findings suggest that the increase in AQP expression may alter fluid exchange in BBB and/or in blood-CSF barrier. This situation in case of an acute or excessively elevated rise of blood pressure can promote BBB changes causing the brain damage occurring in this animal model of hypertension.
MeSH Terms
- Aging
- Animals
- Aquaporin 1
- Aquaporin 4
- Astrocytes
- Brain
- Choroid Plexus
- Chronic Disease
- Corpus Striatum
- Disease Models, Animal
- Disease Progression
- Endothelium, Vascular
- Frontal Lobe
- Hippocampus
- Intracranial Hypertension
- Male
- Rats
- Rats, Inbred SHR
- Rats, Inbred WKY
Salivary glands are involved in secretion of saliva, which is known to participate in the protection and hydratation of mucosal structures within the oral cavity, oropharynx and oesophagus, the initiation of digestion, some antimicrobial defence, and the protection from chemical and mechanical stress. Saliva secretion is a watery fluid containing electrolytes and a mixture of proteins and can be stimulated by muscarinic and adrenergic agonists. Since water movement is involved in saliva secretion, the expression, localization and function of aquaporins (AQPs) have been studied in salivary glands. This review will focus on the expression, localization and functional roles of the AQPs identified in salivary glands. The presence of AQP1, AQP5 and AQP8 has been generally accepted by many, while the presence of AQP3, AQP4, AQP6 and AQP7 still remains controversial. Functionally, AQP5 seems to be the only AQP thus far to be clearly playing a major role in the salivary secretion process. Modifications in AQPs expression and/or distribution have been reported in xerostomic conditions.
MeSH Terms
- Aging
- Animals
- Aquaporin 5
- Aquaporins
- Biological Transport, Active
- Cell Line
- Diabetes Mellitus
- Head and Neck Neoplasms
- Humans
- Protein Transport
- Salivary Glands
- Sjogren's Syndrome
- Water
- Xerostomia
The precise localization of aquaporin (AQP)1 and AQP4 was studied in iris and ciliary epithelial cells, in both mature and developing rats, to elucidate the molecular mechanisms underlying aqueous humor balance. Anterior segments of eyes dissected from embryonic day (E)13, E15, E18, and E20, postnatal day (P)0, P7, and P14, and postnatal week 8 rats were subjected to immunofluorescence analysis with AQP isoform-specific antibodies. In adult rat eye, AQP1 was localized to the apical and basolateral plasma membranes of iris epithelial cell layers and of anterior ciliary non-pigmented epithelial (NPE) cells. Conversely, AQP4 was localized to the basolateral plasma membrane of NPE cells in ciliary epithelium and the posterior iris. Developmentally, AQP1 was detected as early as E15 in immature iris and ciliary epithelial cells, and expression persisted throughout development up to adulthood. In contrast, AQP4 was first observed at P7 in the developing pars plicata, and the AQP4-positive area gradually spread to cover the entire pars plicata as development proceeded. These findings indicate that both AQP1 and AQP4 contribute to aqueous humor secretion in the rat eye, thereby maintaining proper intraocular pressure. Moreover, AQP appears to play a major role in aqueous humor secretion in early eye development. This study thus provides a basis for understanding the molecular mechanisms of aqueous humor secretion in pathological and physiological conditions.
MeSH Terms
- Aging
- Animals
- Aquaporin 1
- Aquaporin 4
- Ciliary Body
- Epithelial Cells
- Female
- Fluorescein-5-isothiocyanate
- Fluorescent Antibody Technique, Indirect
- Fluorescent Dyes
- Immunohistochemistry
- Iris
- Pregnancy
- Rats
- Rats, Sprague-Dawley
Whether the water channel protein AQP4 is involved in the very early cell swelling and brain oedema observed with cerebral hypoxia-ischaemia (HI) and whether this response depends on the maturity of brain were investigated by comparing regional changes in AQP4 protein expression and signal intensity on magnetic resonance (MR) images in immature and juvenile brains. Maps of T2 and the apparent diffusion coefficient (ADC) of water were acquired in 1- and 4-week-old rats at times prior to HI, within the last 5 min of HI and 1 h or 24 h afterwards. AQP4 expression assessed with Western blotting was not significantly reduced until 24 h post-HI irrespective of age. However, AQP4 immunostaining was decreased at the end of HI and at 1 h or 24 h after HI in the hemisphere ipsilateral to the occlusion with changes being similar in both age groups and coinciding well with regional reductions in ADC. IgG immunostaining to assess blood-brain barrier integrity and T2 were unchanged at early time points in 4-week old rats despite decreases in AQP4 immunostaining. Thus, at early time points there were decreases in AQP4 detected with immunostaining but not Western blotting methods. However, the good correlation between alterations in ADC and AQP4 immunostaining suggests that changes in the AQP4 are involved in some of the early changes in brain water distribution observed in hypoxia-ischemia, and supports the speculation that AQP4 is involved in the transport of water across the perivascular membranes into the vascular lumen.
MeSH Terms
- Aging
- Animals
- Animals, Newborn
- Aquaporin 4
- Aquaporins
- Brain Edema
- Female
- Gene Expression Regulation
- Hypoxia-Ischemia, Brain
- Magnetic Resonance Imaging
- Pregnancy
- Rats
Kidneys of new-born animals are resistant to arginine vasopressin (AVP). The ability of the hormone to regulate water permeability of the collecting duct can be seen from weaning period, probably due to the maturation of the intracellular signaling pathway. The purpose of the present work was to investigate the effect of V2 receptor agonist dDAVP on the water permeability of OMCD basolateral membrane in 10-, 22- and 60-day old Wistar rats. We also estimated ontogenetic gene expression of AQP2, AQP3, AQP4 and V2 receptor. Osmotic water permeability (Pf) of the basolateral membrane of microdissected OMCD was measured under control conditions and after incubation with the agonist V2 receptor desmopressin (dDAVP; 10(-7) M). Water permeability in 10- and 22-day old rats under control conditions were significantly higher than in adults. Desmopressin stimulated significant increase of this parameter in 22-day old pups (Pf = = 125 /- 4.85; Pf = 174 /- 8.2 microns/s, p < 0.001) and adult rats (Pf = 100.5 /- 7.38; Pf = 178.8 /- 9.54 microns/s, p < 0.001). Osmotic water permeability of the OMCD basolateral membrane in 10-day old rats does not depend on dDAVP (Pf = 172.5 /- 23.8; Pf = 164.8 /- 34 microns/s). With the RT-PCR, we observed a gradual increase of AQP2 and V2 receptor genes expression during postnatal ontogenesis. The gene expression of AQP3 and AQP4 remained unchanged during postnatal ontogenesis. In general, the water permeability of the OMCD basolateral membrane of rats can be stimulated by AVP since the 22nd day of postnatal life. The water permeability of the OMCD basolateral membrane under control conditions gradually decreased during postnatal development, while gene expression of AQP3 and AQP4 was unchanged. The mechanism of this decrease remains to be established.
MeSH Terms
- Aging
- Animals
- Aquaporin 2
- Aquaporin 6
- Aquaporins
- Cell Membrane Permeability
- Deamino Arginine Vasopressin
- Female
- Kidney Medulla
- Kidney Tubules, Collecting
- Male
- Osmosis
- Rats
- Rats, Wistar
- Receptors, Vasopressin
- Renal Agents
- Water
The expression and localization of aquaporins (AQP1-AQP5), members of the water channel family, in the developing rat submandibular gland were analysed using RT-PCR, Northern blotting and immunohistochemistry to explore their relation to the development of this salivary gland. RT-PCR analysis revealed unique expression patterns of each AQP. AQP1 was expressed constitutively during prenatal development, whereas the expression of AQP5 became more intense in the course of development from embryonic day 16.5 (E16) to E20. These expression patterns concurred with the results of Northern blot analysis. AQP3 and AQP4 mRNAs in the prenatal development were not detected in Northern blots, although they were detected by RT-PCR. During postnatal development, AQP5 and AQP1 mRNAs were expressed continuously, but no message for AQP3 or AQP4 was detected. AQP2 mRNA was not detected during either prenatal or postnatal development in this tissue. Immunohistochemical studies revealed that AQP5 was first localized at the apical membrane of proacinar cells at E18, and then became clearly distributed at the apical membrane of acinar cells in accordance with the differentiation and establishment of the mature acini. In addition, some vasculature also showed immunoreactivity for AQP5. AQP1 was immunolocalized in the blood vessels, including capillaries, of the gland throughout development. These observations suggest the existence of transcriptional regulation of rat AQP5, which is one of the most probable regulators of saliva production and secretion, during the establishment of the functional submandibular salivary gland.
MeSH Terms
- Aging
- Amino Acid Sequence
- Animals
- Aquaporins
- Biological Transport, Active
- Blotting, Northern
- Body Water
- Female
- Immunohistochemistry
- Molecular Sequence Data
- Pregnancy
- RNA
- RNA, Antisense
- Rats
- Rats, Sprague-Dawley
- Reverse Transcriptase Polymerase Chain Reaction
- Sodium-Potassium-Exchanging ATPase
- Submandibular Gland
Water transport between the perilymph and endolymph is important in regulations of volume and osmotic pressure of the inner ear labyrinth. It is now known that expression of water channels (aquaporins or AQPs) in the cell membrane dramatically increases the ability of water to cross epithelial cells. The aims of the current study were to investigate the cellular localization of AQPs by immunolabeling, and to study the developmental expression and relative abundance of various subtypes of AQPs. We report here that AQP3, AQP7 and AQP9 were expressed in the inner ear. Specific subtypes of AQPs were found in discrete regions expressed by both epithelial cells and fibrocytes in cochlear and vestibular organs. Semi-quantitative measurements showed that AQP4 and AQP1 were the two most abundantly expressed AQP subtypes in the inner ear, and their expressions were dramatically upregulated during development. These data showed a highly localized and largely non-overlapping distribution pattern for different subtypes of AQPs in the inner ear, suggesting the existence of regional subtype-specific water transport pathways, and global regulation of water transport in the inner ear may require concerted actions of multiple types of AQPs.
MeSH Terms
- Aging
- Animals
- Aquaporins
- Biological Transport
- Cochlea
- Ear, Inner
- Endolymphatic Sac
- Immunohistochemistry
- Mice
- Protein Isoforms
- Vestibule, Labyrinth
- Water
Aquaporin-4 (AQP4) is a member of the aquaporin water-channel family. AQP4 is expressed primarily in the brain, but it is also present in the collecting duct of the kidney, where it is located in the basolateral plasma membrane of principal cells and inner medullary collecting duct (IMCD) cells. Recent studies in the mouse also have reported the presence of AQP4 in the basolateral membrane of the proximal tubule. The purpose of this study was to establish the pattern of AQP4 expression during kidney development and in the adult kidney of both the mouse and the rat. Kidneys of adult and 3-, 7-, and 15-d-old mice and rats were preserved for immunohistochemistry and processed using a peroxidase pre-embedding technique. In both the mouse and the rat, strong basolateral immunostaining was observed in IMCD cells and principal cells in the medullary collecting duct at all ages examined. Labeling was weaker in the cortical collecting duct and the connecting tubule, and there was no labeling of connecting tubule cells in the mouse. In adult mouse kidney, strong AQP4 immunoreactivity was observed in the S3 segment of the proximal tubule. However, there was little or no labeling in the cortex or around the corticomedullary junction in 3- and 7-d-old mice. Between 7 and 15 d of age, distinct AQP4 immunoreactivity appeared in the S3 segment of the mouse proximal tubule concomitant with the differentiation of this segment of the nephron. Labeling of proximal tubules was never observed in the rat kidney. These results suggest that there are differences in transepithelial water transport between mouse and rat or that additional, not yet identified water channels exist in the rat proximal tubule.
MeSH Terms
- Aging
- Animals
- Animals, Newborn
- Aquaporin 4
- Aquaporins
- Immunohistochemistry
- Kidney
- Mice
- Rats
- Tissue Distribution
Developmental expression of aquaporin water transport proteins is not well understood in respiratory tract or secretory glands; here we define aquaporin protein ontogeny in rat. Expression of aquaporin-3 (AQP3), AQP4, and AQP5 proteins occurs within 2 wk after birth, whereas AQP1 first appears before birth. In most tissues, aquaporin protein expression increases progressively, although transient high-level expression is noted in distal lung (AQP4 at postnatal day 2) and trachea (AQP5 at postnatal day 21 and AQP3 at postnatal day 42). In mature animals, AQP5 is abundant in distal lung and salivary glands, AQP3 and AQP4 are present in trachea, and AQP1 is present in all of these tissues except salivary glands. Surprisingly, all four aquaporin proteins are highly abundant in nasopharynx. Unlike AQP1, corticosteroids did not induce expression of AQP3, AQP4, or AQP5 in lung. Our results seemingly implicate aquaporins in proximal airway humidification, glandular secretion, and perinatal clearance of fluid from distal airways. However, the studies underscore a need for detailed immunohistochemical characterizations and definitive functional studies.
MeSH Terms
- Adrenal Cortex Hormones
- Aging
- Amino Acid Sequence
- Animals
- Antibodies
- Aquaporin 1
- Aquaporin 3
- Aquaporin 4
- Aquaporin 5
- Aquaporins
- Betamethasone
- Embryonic and Fetal Development
- Eye
- Gene Expression Regulation, Developmental
- Ion Channels
- Lung
- Membrane Proteins
- Molecular Sequence Data
- Organ Specificity
- Peptide Fragments
- Rats
- Respiratory Physiological Phenomena
- Respiratory System
- Salivary Glands
The mRNA expression and localization of the aquaporin (AQP) family in rat kidney were examined by ribonuclease protection assay and immunohistochemistry. AQP1, AQP2, AQP3, and AQP4 mRNA were hardly detectable in 16-day gestation fetuses. AQP1 mRNA was explosively expressed at 1 wk, keeping the level throughout life. AQP2 mRNA expression was apparently noticed in 18-day fetuses and was enhanced gradually with age to reach a plateau at 4 wk. AQP3 and AQP4 mRNA expression was significantly found at birth but was not changed remarkably thereafter. AQP2 protein appeared first at the apical side of collecting duct cells in 18-day fetuses. The staining intensity at the site increased with age, and basolateral staining was added in adult rats. AQP3 was distinctly demonstrated at the basolateral side of collecting duct cells after birth, and the staining intensity was almost stable throughout life. The progressive induction of AQP2 expression in the first 4 wk after birth is presumed to contribute to the maturation of urinary concentrating capacity during the kidney development.
MeSH Terms
- Aging
- Animals
- Animals, Newborn
- Embryo, Mammalian
- Embryonic and Fetal Development
- Immunohistochemistry
- Ion Channels
- Kidney
- RNA, Messenger
- Rats
- Rats, Inbred WKY
- Water