AQP4-null Mice Research Articles

Aquaporin-4 deletion ameliorates hypoglycemia-induced BBB permeability by inhibiting inflammatory responses

BackgroundSevere hypoglycemia induces brain edema by upregulating aquaporin-4 (AQP4) expression and by degrading tight junctions. Acute severe hypoglycemia induces a proinflammatory environment that may contribute to a disruption in the epithelial barrier by decreasing tight junction protein expression. Interestingly, the altered AQP4 expression has been considered to play a critical role in neuroinflammation during acute brain injury. It has been shown that AQP4 deletion reduces brain inflammation in AQP4-null mice after intracerebral LPS injection. However, the effect of AQP4 deletion regarding protection against hypoglycemia-induced blood-brain barrier (BBB) breakdown is unknown.MethodsAn acute severe hypoglycemic stress model was established via injection of 4 unit/kg body weight of insulin. Evans blue (EB) staining and water measurement were used to assess BBB permeability. Western blot, reverse transcription polymerase chain reaction, and immunofluorescence were used to detect the expression of related proteins. The production of cytokines was assessed via enzyme-linked immunosorbent assay.ResultsHypoglycemia-induced brain edema and BBB leakage were reduced in AQP4−/− mice. AQP4 deletion upregulated PPAR-γ and inhibited proinflammatory responses. Moreover, knockdown of aquaporin-4 by small interfering RNA in astrocytes co-cultured with endothelial cells effectively reduced transendothelial permeability and degradation of tight junctions. Treatment with PPAR-γ inhibitors showed that upregulation of PPAR-γ was responsible for the protective effect of AQP4 deletion under hypoglycemic conditions.ConclusionsOur data suggest that AQP4 deletion protects BBB integrity by reducing inflammatory responses due to the upregulation of PPAR-γ expression and attenuation of proinflammatory cytokine release. Reduction in AQP4 may be protective in acute severe hypoglycemia.

P-G4 Molecular characterization of hippocampal changes in an animal model of HIV associated primary central nervous system lymphoma

Primary Central Nervous System Lymphoma (PCNSL) is a rare form of extra nodal non-Hodgkin's lymphoma. HIV patients are more prone to PCNSL. It may arise from a systemic lymphoma that seeds multiple organs, including the brain. The predominant histology of PCNSL is large B-cell lymphoma. Molecular studies on systemic AIDS/NHL is well characterized, however, information on molecular studies of PCNSL is limited due in part to accessibility of tissue and lack of animal model. To date, no animal model has been developed, which recapitulates both histopathologic and molecular features of this disease, including the immune phenotypic state. We recently developed an animal model of PCNSL. The observation of a phenotype of a HIV -1 transgenic mouse model that develops a B- cell lymphoma that mimics the disease is seen in AIDS patients with NHL Brain lymphoma patients. HIV associated B-cell lymphoma patients as well as HIV infected patients suffer from cognatic impairment. Brain hippocampus region regulates cognatic behavior. Therefore, we were interested in finding the cellular and molecular changes in the hippocampus of these animals. Interestingly, we identified a significant increase of infiltrating leucocytes, T cells, B cells, and macrophages/microglial cells in the hippocampal region of those mice. Astrogliosis is also observed. Aquaporin-4 (AQP4) is the predominant water channel expressed by astrocytes. The regulation of AQP4 has been extensively investigated in various neuropathological conditions; however, the functional role of AQP4 in synaptic plasticity, learning, and memory is only beginning to be elucidated. We have explored the role of AQP4 and its influence on hippocampus and its potential relationship with synaptic plasticity of these mice. Recent in vitro and in vivo studies using AQP4-null and wild-type mice, in particular, the impairment of cognatic function observed in the hippocampus. (Szu JI and Binder DK. Front Integr Neurosci. 2016 Feb 24; 10:8). In this context we found increased AQP4 and synaptic plasticity (by detecting synaptophysin and synapsine-1) in the hippocampus of these animals. This suggests that AQP4 plays an important role in regulating synaptic plasticity in the hippocampus of these mice. Astrocytes play a role in synaptic plasticity. However, there are only a few studies that implicate a direct relationship of AQP4 in synaptic plasticity. All together, these studies highlight the potential influence of AQP4 in synaptic plasticity and memory of these animals probably due to compensatory mechanism.

The Role of Astrocytic Aquaporin-4 in Synaptic Plasticity and Learning and Memory.

Aquaporin-4 (AQP4) is the predominant water channel expressed by astrocytes in the central nervous system (CNS). AQP4 is widely expressed throughout the brain, especially at the blood-brain barrier where AQP4 is highly polarized to astrocytic foot processes in contact with blood vessels. The bidirectional water transport function of AQP4 suggests its role in cerebral water balance in the CNS. The regulation of AQP4 has been extensively investigated in various neuropathological conditions such as cerebral edema, epilepsy, and ischemia, however, the functional role of AQP4 in synaptic plasticity, learning, and memory is only beginning to be elucidated. In this review, we explore the current literature on AQP4 and its influence on long term potentiation (LTP) and long term depression (LTD) in the hippocampus as well as the potential relationship between AQP4 and in learning and memory. We begin by discussing recent in vitro and in vivo studies using AQP4-null and wild-type mice, in particular, the impairment of LTP and LTD observed in the hippocampus. Early evidence using AQP4-null mice have suggested that impaired LTP and LTD is brain-derived neurotrophic factor dependent. Others have indicated a possible link between defective LTP and the downregulation of glutamate transporter-1 which is rescued by chronic treatment of β-lactam antibiotic ceftriaxone. Furthermore, behavioral studies may shed some light into the functional role of AQP4 in learning and memory. AQP4-null mice performances utilizing Morris water maze, object placement tests, and contextual fear conditioning proposed a specific role of AQP4 in memory consolidation. All together, these studies highlight the potential influence AQP4 may have on long term synaptic plasticity and memory.

The central role of aquaporins in the pathophysiology of ischemic stroke

Stroke is a complex and devastating neurological condition with limited treatment options. Brain edema is a serious complication of stroke. Early edema formation can significantly contribute to infarct formation and thus represents a promising target. Aquaporin (AQP) water channels contribute to water homeostasis by regulating water transport and are implicated in several disease pathways. At least 7 AQP subtypes have been identified in the rodent brain and the use of transgenic mice has greatly aided our understanding of their functions. AQP4, the most abundant channel in the brain, is up-regulated around the peri-infarct border in transient cerebral ischemia and AQP4 knockout mice demonstrate significantly reduced cerebral edema and improved neurological outcome. In models of vasogenic edema, brain swelling is more pronounced in AQP4-null mice than wild-type providing strong evidence of the dual role of AQP4 in the formation and resolution of both vasogenic and cytotoxic edema. AQP4 is co-localized with inwardly rectifying K+-channels (Kir4.1) and glial K+ uptake is attenuated in AQP4 knockout mice compared to wild-type, indicating some form of functional interaction. AQP4-null mice also exhibit a reduction in calcium signaling, suggesting that this channel may also be involved in triggering pathological downstream signaling events. Associations with the gap junction protein Cx43 possibly recapitulate its role in edema dissipation within the astroglial syncytium. Other roles ascribed to AQP4 include facilitation of astrocyte migration, glial scar formation, modulation of inflammation and signaling functions. Treatment of ischemic cerebral edema is based on the various mechanisms in which fluid content in different brain compartments can be modified. The identification of modulators and inhibitors of AQP4 offer new therapeutic avenues in the hope of reducing the extent of morbidity and mortality in stroke.

Aquaporin-4 deletion in mice reduces encephalopathy and brain edema in experimental acute liver failure

Brain edema and associated astrocyte swelling leading to increased intracranial pressure are hallmarks of acute liver failure (ALF). Elevated blood and brain levels of ammonia have been implicated in the development of brain edema in ALF. Cultured astrocytes treated with ammonia have been shown to undergo cell swelling and such swelling was associated with an increase in the plasma membrane expression of aquaporin-4 (AQP4) protein. Further, silencing the AQP4 gene in cultured astrocytes was shown to prevent the ammonia-induced cell swelling. Here, we examined the evolution of brain edema in AQP4-null mice and their wild type counterparts (WT-mice) in different models of ALF induced by thioacetamide (TAA) or acetaminophen (APAP). Induction of ALF with TAA or APAP significantly increased brain water content in WT mice (by 1.6%±0.3 and 2.3±0.4%, respectively). AQP4 protein was significantly increased in brain plasma membranes of WT mice with ALF induced by either TAA or APAP. In contrast to WT-mice, brain water content did not increase in AQP4-null mice. Additionally, AQP4-null mice treated with either TAA or APAP showed a remarkably lesser degree of neurological deficits as compared to WT mice; the latter displayed an inability to maintain proper gait, and demonstrated a markedly reduced exploratory behavior, with the mice remaining in one corner of the cage with its head tilted downwards. These results support a central role of AQP4 in the brain edema associated with ALF.

Aquaporin-4 antibodies, CNS acidosis and neuromyelitis optica: A potential link

BackgroundNeuromyelitis optica (NMO, Devic’s syndrome) is a severely disabling disorder of the central nervous system characterized by optic neuritis and longitudinally extensive myelitis. In around 80% of cases, NMO is caused by autoantibodies to astrocytic aquaporin-4 (AQP4), the most abundant water channel in the CNS. Acute NMO attacks are frequently accompanied by elevated levels of lactate in the cerebrospinal fluid (CSF). As a strongly dissociated anion (pK′=3.7) directly changing the strong ion difference, lactate causes a reduction in the dependent anion [HCO3-] and a rise in [H+], resulting in “metabolic” acidosis in the CSF. CSF acidosis also develops during respiratory failure due to brainstem or high cervical spinal cord lesions, the most common cause of death in NMO. However, lactic acid and more generally, a decrease in pH, has been shown to increase the membrane expression of AQP4 in astrocytes. An increase in AQP4 membrane expression during acute NMO attacks could potentially enhance the complement-mediated humoral immune reaction against AQP4-expressing astrocytes characteristic for NMO and, thus, result in more severe astrocytic damage. Moreover, lactate and acidosis have been shown to cause astrocytic swelling and to affect astrocytic viability, potentially rendering astrocytes more susceptible to AQP4-Ab-mediated damage. Finally, increased AQP4 expression could be an independent risk factor in NMO and other forms of CNS inflammation, as indicated by the finding of grossly attenuated experimental autoimmune encephalomyelitis in AQP4-null mice. Therefore, we hypothesize that CSF acidosis might play a role in the pathophysiology of AQP4-Ab-positive NMO and that alterations in CSF pH might possibly influence the outcome of acute attacks in this condition. In addition, we discuss potential clinical implications and make proposals on how to test the hypothesis. Finally, other factors that influence astrocytic AQP4 membrane expression and might play a role in NMO are discussed.

Protective role of brain water channel AQP4 in murine cerebral malaria

Tragically common among children in sub-Saharan Africa, cerebral malaria is characterized by rapid progression to coma and death. In this study, we used a model of cerebral malaria appearing in C57BL/6 WT mice after infection with the rodent malaria parasite Plasmodium berghei ANKA. Expression and cellular localization of the brain water channel aquaporin-4 (AQP4) was investigated during the neurological syndrome. Semiquantitative real-time PCR comparing uninfected and infected mice showed a reduction of brain AQP4 transcript in cerebral malaria, and immunoblots revealed reduction of brain AQP4 protein. Reduction of brain AQP4 protein was confirmed in cerebral malaria by quantitative immunogold EM; however, polarized distribution of AQP4 at the perivascular and subpial astrocyte membranes was not altered. To further examine the role of AQP4 in cerebral malaria, WT mice and littermates genetically deficient in AQP4 were infected with P. berghei. Upon development of cerebral malaria, WT and AQP4-null mice exhibited similar increases in width of perivascular astroglial end-feet in brain. Nevertheless, the AQP4-null mice exhibited more severe signs of cerebral malaria with greater brain edema, although disruption of the blood-brain barrier was similar in both groups. In longitudinal studies, cerebral malaria appeared nearly 1 d earlier in the AQP4-null mice, and reduced survival was noted when chloroquine rescue was attempted. We conclude that the water channel AQP4 confers partial protection against cerebral malaria.

Aquaporin-4 Reduces Post-Traumatic Seizure Susceptibility by Promoting Astrocytic Glial Scar Formation in Mice

Seizures are important neurological complications after traumatic brain injury (TBI) and are reported for up to 50% of patients with TBI. Despite several studies, no drug strategy has been able to alter the biological events leading to epileptogenesis. The glial water channel, aquaporin-4 (AQP4), was shown to facilitate cytotoxic cell swelling in ischemia and glial scar formation after stab wound injury. In this study, we examined post-traumatic seizure susceptibility of AQP4-deficient mice (AQP4-/-) after injection of pentylenetetrazole (PTZ) 1 month after controlled cortical impact (CCI) and compared them to wild-type sham injury controls. After PTZ injection, AQP4-/- mice demonstrated dramatically shortened seizure latency (120 ± 40 vs. 300 ± 70 sec; p < 0.001) and increased seizure severity (grade 7.5 ± 0.4 vs. 5.8 ± 0.4; p < 0.001) compared to their wild-type counterparts. Morphometric analysis demonstrated a significant 2-fold reduction in astrocytosis, with a concomitant increase in microgliosis in injured AQP4-null mice compared to their injured wild-type counterparts (44 ± 2 vs. 24 ± 3 cells per high power field [cells/hpf], respectively; p < 0.0001). Minocycline, an inhibitor of microglia, reversed the post-TBI epilepsy phenotype of AQP4-null mice. After minocycline treatment, AQP4-/- mice demonstrated similar latency of seizures evoked by PTZ (723 ± 35 vs. 696 ± 38 sec; p > 0.05) and severity of seizures evoked by PTZ (grade 4.0 ± 0.5 vs. 3.81 ± 0.30; p > 0.05) compared to wild-type counterparts. Immunohistochemical analysis demonstrated decreased immunostaining of microglia to levels comparable to wild-type (12 ± 2 vs. 11 ± 4 cells/hpf, respectively; p > 0.05). Taken together, these results suggest a protective role of AQP4 in post-traumatic seizure susceptibility by promoting astrogliosis, formation of a glial scar, and preventing microgliosis.

Aquaporins at a glance

With a concentration of 55,000 mM, water is by far the most prevalent molecule in biological systems. For many years it was assumed that biological membranes are freely water permeable, so that the existence of a family of water channels would not have been predicted. However, biophysical studies in

Expression and function of aquaporin-4 in olfactory system of mice

Objective To investigate the expression and function of aquaporin-4(AQP4) in the olfactory system of mice.Methods The differences of AQP4 expression in olfaction system between wild-type and AQP4-null mice were studied by immunoblotting and immunofluorescence methods. The differences of mouse olfactory functions in the two groups were examined by two olfactory behavioral assays: the buried food pellet test and olfaction maze test,and the odorant-stimulated electroolfactogram(EOG) recording.Results The results of immunoblotting and immunofluorescence showed no AQP4 expression in the olfactory system in AQP4-null mice.Immunofluorescence result also indicated that AQP4 was mainly distributed in the membrane of support cells,duct cells of Bowmann's gland,basal cells of olfactory epithelium,membrane of Bowmann's gland epithelial cells,olfactory sheath cells surrounding the olfactory bundles,and the membrane of cells in the olfactory bundle layer and glomeruler layer.The results of olfactory behavioral assay were significantly different between the two groups at all time points tested in both the olfaction maze test and the buried food pellet test(P0.05).It was showed that the EOGs under different pressures of saturated trimethylamine had a similar shape in both groups,and the amplitude of EOGs increased with the increase of pressure.While under the same pressure,the EOG amplitude of AQP4-null mice was significantly lower than that of wild-type mice(P0.05).Conclusion AQP4 is widely distributed in the olfactory system of mice,including the olfactory mucosa,olfactory nerve,and olfactory bulb,which can protect the olfactory neural bundle and facilitate neural signal transfer.

Aquaporin-4: orthogonal array assembly, CNS functions, and role in neuromyelitis optica

Aquaporin-4 (AQP4) is a water-selective transporter expressed in astrocytes throughout the central nervous system, as well as in kidney, lung, stomach and skeletal muscle. The two AQP4 isoforms produced by alternative spicing, M1 and M23 AQP4, form heterotetramers that assemble in cell plasma membranes in supramolecular structures called orthogonal arrays of particles (OAPs). Phenotype analysis of AQP4-null mice indicates the involvement of AQP4 in brain and spinal cord water balance, astrocyte migration, neural signal transduction and neuroinflammation. AQP4-null mice manifest reduced brain swelling in cytotoxic cerebral edema, but increased brain swelling in vasogenic edema and hydrocephalus. AQP4 deficiency also increases seizure duration, impairs glial scarring, and reduces the severity of autoimmune neuroinflammation. Each of these phenotypes is likely explicable on the basis of reduced astrocyte water permeability in AQP4 deficiency. AQP4 is also involved in the neuroinflammatory demyelinating disease neuromyelitis optica (NMO), where autoantibodies (NMO-IgG) targeting AQP4 produce astrocyte damage and inflammation. Mice administered NMO-IgG and human complement by intracerebral injection develop characteristic NMO lesions with neuroinflammation, demyelination, perivascular complement deposition and loss of glial fibrillary acidic protein and AQP4 immunoreactivity. Our findings suggest the potential utility of AQP4-based therapeutics, including small-molecule modulators of AQP4 water transport function for therapy of brain swelling, injury and epilepsy, as well as small-molecule or monoclonal antibody blockers of NMO-IgG binding to AQP4 for therapy of NMO.

Absence of Aquaporin-4 in Skeletal Muscle Alters Proteins Involved in Bioenergetic Pathways and Calcium Handling

Aquaporin-4 (AQP4) is a water channel expressed at the sarcolemma of fast-twitch skeletal muscle fibers, whose expression is altered in several forms of muscular dystrophies. However, little is known concerning the physiological role of AQP4 in skeletal muscle and its functional and structural interaction with skeletal muscle proteome. Using AQP4-null mice, we analyzed the effect of the absence of AQP4 on the morphology and protein composition of sarcolemma as well as on the whole skeletal muscle proteome. Immunofluorescence analysis showed that the absence of AQP4 did not perturb the expression and cellular localization of the dystrophin-glycoprotein complex proteins, aside from those belonging to the extracellular matrix, and no alteration was found in sarcolemma integrity by dye extravasation assay. With the use of a 2DE-approach (BN/SDS-PAGE), protein maps revealed that in quadriceps, out of 300 Coomassie-blue detected and matched spots, 19 proteins exhibited changed expression in AQP4−/− compared to WT mice. In particular, comparison of the protein profiles revealed 12 up- and 7 down-regulated protein spots in AQP4−/− muscle. Protein identification by MS revealed that the perturbed expression pattern belongs to proteins involved in energy metabolism (i.e. GAPDH, creatine kinase), as well as in Ca2+ handling (i.e. parvalbumin, SERCA1). Western blot analysis, performed on some significantly changed proteins, validated the 2D results. Together these findings suggest AQP4 as a novel determinant in the regulation of skeletal muscle metabolism and better define the role of this water channel in skeletal muscle physiology.

Increased brain edema in aqp4-null mice in an experimental model of subarachnoid hemorrhage

We investigated the role of the glial water channel protein aquaporin-4 in brain edema in a mouse model of subarachnoid hemorrhage in which 30 μl of blood was injected into the basal cisterns. Brain water content, intracranial pressure and neurological score were compared in wildtype and aquaporin-4 null mice. We also measured blood-brain barrier permeability, and the osmotic permeability of the glia limitans, one of the routes of edema elimination. Wildtype and aquaporin-4 null mice had comparable baseline brain water content, intracranial pressure and neurological score. At 6 h after blood injection, aquaporin-4 null mice developed more brain swelling than wildtype mice. Brain water content increased by 1.5±0.1% vs. 0.5±0.2% (Mean±Standard Error, P<0.0005) and intracranial pressure by 36±5 vs. 21±3 mm Hg ( P<0.05) above pre-injection baseline, and neurological score was worse at 18.0 vs. 24.5 (median, P<0.05), respectively. Although subarachnoid hemorrhage produced comparable increases in blood–brain barrier permeability in wildtype and aquaporin-4 null mice, aquaporin-4 null mice had a twofold reduction in glia limitans osmotic permeability. We conclude that aquaporin-4 null mice manifest increased brain edema following subarachnoid hemorrhage as a consequence of reduced elimination of excess brain water.

Aquaporin-4 deficiency induces subfertility in female mice

To investigate the effects of aquaporin-4 (AQP4) on female fertility. Prospective, randomized animal study. Medical university. AQP4(+/+) and AQP4(-/-) female mice. Eight- to 10-week-old virgin female AQP4(+/+) and AQP4(-/-) mice were subjected to a mating study. Superovulation was carried out on 4-week-old AQP4(+/+) and AQP4(-/-) mice. The protocol consisted of a single intraperitoneal (IP) injection with 5 units of pregnant mare serum gonadotropin (PMSG) for 48 hours, followed by 5 units of human chorionic gonadotropin (hCG) for an additional 14 or 48 hours. Pregnancy success, oocyte count, morphological changes in ovaries and uteri, and alterations of estrous cycle. AQP4-null mice displayed subfertility as evidenced by a lower rate of pregnancy and decreased litter size. Further studies revealed that in AQP4-null mice, there were fewer numbers of antral follicles and corpora lutea in the ovaries and that uteri presented as hyporeactive to gonadotropins. In addition, in AQP4-null mice, superovulation induced by hCG and PMSG yielded fewer numbers of antral follicles, atretic follicles, corpora lutea, and oocytes in ovaries, and uteri exhibited a decreased response to exogenous gonadotropins. AQP4 deficiency induces subfertility in female mice.

Sporadic obstructive hydrocephalus in Aqp4 null mice

Aquaporin-4 (Aqp4) is a water transport protein expressed in glia and ependymocytes in brain. We report here the unexpected occurrence of severe obstructive hydrocephalus in a random subset of Aqp4 knockout mice. Of 612 Aqp4 knockout mice produced by heterozygote-heterozygote or knockout-knockout breedings, 9.6% of offspring manifested progressive encephalomegaly. Encephalomegaly was never seen in wild-type or Aqp4 heterozygous mice. Examination of the subset encephalomegalic mice revealed marked triventricular hydrocephalus (lateral ventricle size approximately 500 mm(3)), elevated intracranial pressure (19 +/- 3 vs. 6.1 +/- 0.6 mm Hg), and death by age 6 weeks, with a median survival of 28 days. Intraventricular dye injection studies revealed total obstruction of the cerebral aqueduct. Evans blue extravasation studies indicated an intact blood-brain barrier in the hydrocephalic mice. Brain histology revealed reduced ventricular size and ependymocyte disorganization in some nonhydrocephalic Aqp4 null mice. Our studies establish Aqp4 deletion as a predisposing factor for the development of congenital obstructive hydrocephalus in mice. We suggest that AQP4 polymorphisms might also contribute to the development of aqueduct stenosis in humans.

Impaired olfaction in mice lacking aquaporin‐4 water channels

Aquaporin-4 (AQP4) is a water-selective transport protein expressed in glial cells throughout the central nervous system. AQP4 deletion in mice produces alterations in several neuroexcitation phenomena, including hearing, vision, epilepsy, and cortical spreading depression. Here, we report defective olfaction and electroolfactogram responses in AQP4-null mice. Immunofluorescence indicated strong AQP4 expression in supportive cells of the nasal olfactory epithelium. The olfactory epithelium in AQP4-null mice had identical appearance, but did not express AQP4, and had approximately 12-fold reduced osmotic water permeability. Behavioral analysis showed greatly impaired olfaction in AQP4-null mice, with latency times of 17 +/- 0.7 vs. 55 +/- 5 s in wild-type vs. AQP4-null mice in a buried food pellet test, which was confirmed using an olfactory maze test. Electroolfactogram voltage responses to multiple odorants were reduced in AQP4-null mice, with maximal responses to triethylamine of 0.80 +/- 0.07 vs. 0.28 +/- 0.03 mV. Similar olfaction and electroolfactogram defects were found in outbred (CD1) and inbred (C57/bl6) mouse genetic backgrounds. Our results establish AQP4 as a novel determinant of olfaction, the deficiency of which probably impairs extracellular space K(+) buffering in the olfactory epithelium.

Greatly improved neurological outcome after spinal cord compression injury in AQP4-deficient mice

Aquaporin-4 (AQP4) is a water channel protein expressed in astrocytes throughout the CNS. In brain, AQP4 facilitates water balance and glial scar formation, which are important determinants of outcome after injury. Here, we provide evidence for AQP4-dependent spinal cord swelling following compression injury, resulting in remarkably improved outcome in AQP4-null mice. Two days after transient T6 spinal cord compression injury, wild-type mice developed more severe hindlimb weakness than AQP4-null mice, as assayed by the Basso open-field motor score, inclined plane method and footprint analysis. Basso motor scores were 1.3 +/- 0.5 (wild-type) versus 4.9 +/- 0.6 (AQP4-null) (SE, P < 0.001). Improved motor outcome in AQP4-null mice was independent of mouse strain and persisted at least 4 weeks. AQP4-null mice also had improved sensory outcome at 2 days, as assessed by spinal somatosensory evoked responses, with signal amplitudes approximately 10 microV (uninjured), 1.7 +/- 0.7 microV (wild-type) and 6.4 +/- 1.3 microV (AQP4-null) (P < 0.01). The improved motor and sensory indices in AQP4-null mice corresponded to remarkably less neuronal death and myelin vacuolation, as well as reduced spinal cord swelling and intraparenchymal spinal cord pressure measured at T6 at 2 days after injury. AQP4 immunoreactivity at the injury site was increased in grey and white matter at 48 h. Taken together, our findings indicate that AQP4 provides a major route for excess water entry into the injured spinal cord, which in turn causes spinal cord swelling and elevated spinal cord pressure. Our data suggest AQP4 inhibition or downregulation as novel early neuroprotective manoeuvres in spinal cord injury.

Aquaporin-4 and brain edema

Aquaporin-4 (AQP4) is a water-channel protein expressed strongly in the brain, predominantly in astrocyte foot processes at the borders between the brain parenchyma and major fluid compartments, including cerebrospinal fluid (CSF) and blood. This distribution suggests that AQP4 controls water fluxes into and out of the brain parenchyma. Experiments using AQP4-null mice provide strong evidence for AQP4 involvement in cerebral water balance. AQP4-null mice are protected from cellular (cytotoxic) brain edema produced by water intoxication, brain ischemia, or meningitis. However, AQP4 deletion aggravates vasogenic (fluid leak) brain edema produced by tumor, cortical freeze, intraparenchymal fluid infusion, or brain abscess. In cytotoxic edema, AQP4 deletion slows the rate of water entry into brain, whereas in vasogenic edema, AQP4 deletion reduces the rate of water outflow from brain parenchyma. AQP4 deletion also worsens obstructive hydrocephalus. Recently, AQP4 was also found to play a major role in processes unrelated to brain edema, including astrocyte migration and neuronal excitability. These findings suggest that modulation of AQP4 expression or function may be beneficial in several cerebral disorders, including hyponatremic brain edema, hydrocephalus, stroke, tumor, infection, epilepsy, and traumatic brain injury.

Noninvasive early detection of brain edema in mice by near-infrared light scattering

Brain edema accounts for significant morbidity and mortality in many neurologic conditions such as head trauma, stroke, meningitis, and brain tumor. The water channel aquaporin-4 (AQP4) has been found to be an important determinant of brain water accumulation and clearance of excess brain water. We report the development of a noninvasive near-infrared (NIR) light-scattering method to compare the early kinetics of brain swelling in normal and AQP4-deficient mice. Brain tissue was illuminated through the intact skull with NIR light at 850 nm, and steady-state scattered light intensity was monitored at an angle of 90 degrees at a position on the skull approximately 10 mm from the illuminated site. NIR light scattering reversibly increased with brain swelling (DeltaI/Io approximately 25% per 1% increase in brain water content), but was insensitive to changes in cerebral blood flow, blood oxygenation, or blood flow-related changes in intracranial pressure (ICP). DeltaI/Io increased approximately linearly with brain water content as measured by wet-to-dry weight ratios. Acute water intoxication (intraperitoneal water, 20% body weight) produced a gradual increase in DeltaI/Io of 12 +/- 4% in wild-type mice at 5 min, much greater than that of 2 +/- 1% in AQP4-null mice. Correlation of the NIR signal with ICP showed that increased DeltaI/Io preceded measurable increases in ICP, indicating the ability of the NIR method to detect early brain edema before ICP elevation. NIR light scattering provides a simple noninvasive method to monitor brain edema in mice, with potential clinical applications.

Aquaporin-4 Gene Disruption in Mice Protects against Impaired Retinal Function and Cell Death after Ischemia

Water channel aquaporin (AQP)-4 is expressed in Muller cells in retina, which are similar to astroglial cells in the central nervous system, where AQP4 deletion protects against cytotoxic brain edema after cerebral ischemia. A transient ischemia-reperfusion model was used to determine whether AQP4 deletion in mice protects the retina. Retinal function and morphology were assessed in wild-type versus AQP4-deficient mice after ischemic damage produced by a 45- to 60-minute elevation of intraocular pressure to 120 mm Hg. Retinal function was assessed by electroretinography, and retinal structure by light microscopy. Extracellular space (ECS) size in fluorescently stained retinal slices was assessed by fluorescence recovery after photobleaching. Retinal function and cell survival were significantly improved in AQP4-deficient mice in both inbred (C57/bl6) and outbred (CD1) genetic backgrounds. By electroretinography, b-wave amplitude was reduced by 75% to 83% at 1 to 4 days after ischemia in wild-type mice versus 48% to 51% in AQP4-null CD1 mice. Reductions were 53% to 72% versus <34% in C57/bl6 mice. Retinal structure and cell count were preserved in AQP4-null mice, particularly in the inner nuclear and plexiform layers of the retina, where Müller cells are concentrated. At 4 days after ischemia, inner retinal thickness was thinned by 43% in wild-type mice versus 11% in AQP4-null mice. Several mechanisms for retinal protection were investigated, including ECS expansion, reduced early swelling, and altered Kir4.1 K(+) channel expression. AQP4 deletion in mice is neuroprotective in a transient ischemia model of retinal injury, suggesting the possible use of AQP4 inhibitors in retinal vascular occlusive and ischemic diseases.