Aquaporin-4-Deficient Mice Have Increased Extracellular Space without Tortuosity Change

Department of Neurosurgery and Brain and Spinal Injury Center, University of California, San Francisco, San Francisco, California 94110, USA.
The Journal of Neuroscience : The Official Journal of the Society for Neuroscience (Impact Factor: 6.34). 06/2008; 28(21):5460-4. DOI: 10.1523/JNEUROSCI.0257-08.2008
Source: PubMed


Aquaporin-4 (AQP4) is the major water channel expressed at fluid-tissue barriers throughout the brain and plays a crucial role in cerebral water balance. To assess whether these channels influence brain extracellular space (ECS) under resting physiological conditions, we used the established real-time iontophoresis method with tetramethylammonium (TMA(+)) to measure three diffusion parameters: ECS volume fraction (alpha), tortuosity (lambda), and TMA(+) loss (k'). In vivo measurements were performed in the somatosensory cortex of AQP4-deficient (AQP4(-/-)) mice and wild-type controls with matched age. Mice lacking AQP4 showed a 28% increase in alpha (0.23 +/- 0.007 vs 0.18 +/- 0.003) with no differences in lambda (1.62 +/- 0.04 vs 1.61 +/- 0.02) and k' (0.0045 +/- 0.0001 vs 0.0031 +/- 0.0009 s(-1)). Additional recordings in brain slices showed similarly elevated alpha in AQP4(-/-) mice, and no differences in lambda and k' between the two genotypes. This is the first direct comparison of ECS properties in adult mice lacking AQP4 water channels with wild-type animals and demonstrates a significant enlargement of the volume fraction but no difference in hindrance to TMA(+) diffusion, expressed as tortuosity. These findings provide direct evidence for involvement of AQP4 in modulation of the ECS volume fraction and provide a basis for future modeling of water and ion transport in the CNS.

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Available from: Xiaoming Yao, Jan 14, 2015
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    • "In accord with our finding of increased seizure susceptibility and reduced ECS volume in Has3 / mice, decreased seizure susceptibility and enlarged ECS volume were previously reported in aquaporin-4-deficient mice (Binder et al., 2004; Yao et al., 2008). We also note that other studies have demonstrated modulation of ongoing epileptiform activity by osmotically induced ECS volume changes (Andrew et al., 1989; Traynelis and Dingledine, 1989; Dudek et al., 1998). "
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    ABSTRACT: Hyaluronan (HA), a large anionic polysaccharide (glycosaminoglycan), is a major constituent of the extracellular matrix of the adult brain. To address its function, we examined the neurophysiology of knock-out mice deficient in hyaluronan synthase (Has) genes. Here we report that these Has mutant mice are prone to epileptic seizures, and that in Has3(-/-) mice, this phenotype is likely derived from a reduction in the size of the brain extracellular space (ECS). Among the three Has knock-out models, namely Has3(-/-), Has1(-/-), and Has2(CKO), the seizures were most prevalent in Has3(-/-) mice, which also showed the greatest HA reduction in the hippocampus. Electrophysiology in Has3(-/-) brain slices demonstrated spontaneous epileptiform activity in CA1 pyramidal neurons, while histological analysis revealed an increase in cell packing in the CA1 stratum pyramidale. Imaging of the diffusion of a fluorescent marker revealed that the transit of molecules through the ECS of this layer was reduced. Quantitative analysis of ECS by the real-time iontophoretic method demonstrated that ECS volume was selectively reduced in the stratum pyramidale by ∼40% in Has3(-/-) mice. Finally, osmotic manipulation experiments in brain slices from Has3(-/-) and wild-type mice provided evidence for a causal link between ECS volume and epileptiform activity. Our results provide the first direct evidence for the physiological role of HA in the regulation of ECS volume, and suggest that HA-based preservation of ECS volume may offer a novel avenue for development of antiepileptogenic treatments.
    Preview · Article · Apr 2014 · The Journal of Neuroscience : The Official Journal of the Society for Neuroscience
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    • "There are controversial reports in the literature about the tortuosity changes in AQP4-deficient mice observed using different methods. While the study of Yao and colleagues [26] is in agreement with our findings, diffusion studies with dextran polymers using fluorescence recovery after photobleaching (FRAP) showed, in contrast, a decrease of about 10–20% in tortuosity in the neocortex of AQP4−/− mice [30], [31]. Since the recording intervals are significantly shorter and the molecular weight of the used extracellular marker is higher in the FRAP method compared to RTI-TMA method, the diffusion parameters measured by these methods may differ. "
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    ABSTRACT: Aquaporin-4 (AQP4) is the primary cellular water channel in the brain and is abundantly expressed by astrocytes along the blood-brain barrier and brain-cerebrospinal fluid interfaces. Water transport via AQP4 contributes to the activity-dependent volume changes of the extracellular space (ECS), which affect extracellular solute concentrations and neuronal excitability. AQP4 is anchored by α-syntrophin (α-syn), the deletion of which leads to reduced AQP4 levels in perivascular and subpial membranes. We used the real-time iontophoretic method and/or diffusion-weighted magnetic resonance imaging to clarify the impact of α-syn deletion on astrocyte morphology and changes in extracellular diffusion associated with cell swelling in vitro and in vivo. In mice lacking α-syn, we found higher resting values of the apparent diffusion coefficient of water (ADCW) and the extracellular volume fraction (α). No significant differences in tortuosity (λ) or non-specific uptake (k'), were found between α-syn-negative (α-syn -/-) and α-syn-positive (α-syn +/+) mice. The deletion of α-syn resulted in a significantly smaller relative decrease in α observed during elevated K(+) (10 mM) and severe hypotonic stress (-100 mOsmol/l), but not during mild hypotonic stress (-50 mOsmol/l). After the induction of terminal ischemia/anoxia, the final values of ADCW as well as of the ECS volume fraction α indicate milder cell swelling in α-syn -/- in comparison with α-syn +/+ mice. Shortly after terminal ischemia/anoxia induction, the onset of a steep rise in the extracellular potassium concentration and an increase in λ was faster in α-syn -/- mice, but the final values did not differ between α-syn -/- and α-syn +/+ mice. This study reveals that water transport through AQP4 channels enhances and accelerates astrocyte swelling. The substantially altered ECS diffusion parameters will likely affect the movement of neuroactive substances and/or trophic factors, which in turn may modulate the extent of tissue damage and/or drug distribution.
    Full-text · Article · Jul 2013 · PLoS ONE
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    • "In contrast to the slowed diffusion produced by brain edema and seizure activity, ECS diffusion was faster in Aqp4 2/2 mice, indicating ECS expansion in AQP4 deficiency (Binder et al., 2004b). Similar results were obtained with follow-up studies using the TMA 1 method (Yao et al., 2008). "
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    ABSTRACT: Recent studies have implicated glial cells in modulation of synaptic transmission, so it is plausible that glial cells may have a functional role in the hyperexcitability characteristic of epilepsy. Indeed, alterations in distinct astrocyte membrane channels, receptors, and transporters have all been associated with the epileptic state. This review focuses on the potential roles of the glial water channel aquaporin-4 (AQP4) in modulation of brain excitability and in epilepsy. We will review studies of mice lacking AQP4 (Aqp4(-/-) mice) or α-syntrophin (an AQP4 anchoring protein) and discuss the available human studies demonstrating alterations of AQP4 in human epilepsy tissue specimens. We will conclude with new studies of AQP4 regulation and discuss the potential role of AQP4 in the development of epilepsy (epileptogenesis). While many questions remain unanswered, the available data indicate that AQP4 and its molecular partners may represent important new therapeutic targets.
    Full-text · Article · Aug 2012 · Glia
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