Zsolt Zador

Increased seizure duration and slowed potassium kinetics in mice lacking aquaporin-4 water channels

The glial water channel aquaporin‐4 (AQP4) has been hypothesized to modulate water and potassium fluxes associated with neuronal activity. In this study, we examined the seizure phenotype of AQP4 −/− mice using in vivo electrical stimulation and electroencephalographic (EEG) recording. AQP4 −/− mice were found to have dramatically prolonged stimulation‐evoked seizures after hippocampal stimulation compared to wild‐type controls (33 ± 2 s vs. 13 ± 2 s). In addition, AQP4 −/− mice were found to have a higher seizure threshold (167 ± 17 μA vs. 114 ± 10 μA). To assess a potential effect of AQP4 on potassium kinetics, we used in vivo recording with potassium‐sensitive microelectrodes after direct cortical stimulation. Although there was no significant difference in baseline or peak [K+]o, the rise time to peak [K+]o (t1/2, 2.3 ± 0.5 s) as well as the recovery to baseline [K+]o (t1/2, 15.6 ± 1.5 s) were slowed in AQP4 −/− mice compared to WT mice (t1/2, 0.5 ± 0.1 and 6.6 ± 0.7 s, respectively). These results implicate AQP4 in the expression and termination of seizure activity and support the hypothesis that AQP4 is coupled to potassium homeostasis in vivo.

Bypass surgery for complex brain aneurysms: an assessment of intracranial-intracranial bypass.

OBJECTIVEBypass surgery for brain aneurysms is evolving from extracranial-intracranial (EC-IC) to intracranial-intracranial (IC-IC) bypasses that reanastomose parent arteries, revascularize efferent branches with in situ donor arteries or reimplantation, and reconstruct bifurcated anatomy with grafts that are entirely intracranial. We compared results with these newer IC-IC bypasses to conventional EC-IC bypasses. METHODSDuring a 10-year period, 82 patients underwent bypass surgery as part of their aneurysm management. A quarter of the patients presented with ruptured aneurysms and two-thirds presented with compressive symptoms from unruptured aneurysms. Most aneurysms (82%) had non-saccular morphology and 56% were giant sized. Common locations included the cavernous internal carotid artery (23%), middle cerebral artery (20%), and posteroinferior cerebellar artery (12%). RESULTSForty-seven patients (57%) received EC-IC bypasses and 35 patients (43%) received IC-IC bypasses, including 9 in situ bypasses, 6 reimplantations, 11 reanastomoses, and 9 intracranial grafts. Aneurysm obliteration rates were comparable in EC-IC and IC-IC bypass groups (97.9% and 97.1%, respectively), as were bypass patency rates (94% and 89%, respectively). Three patients died (surgical mortality, 3.7%), and 4 patients were permanently worse as a result of bypass occlusions (neurological morbidity, 4.9%). At late follow-up (mean duration, 41 months), good outcomes (Glasgow Outcome Scale score 5 or 4) were measured in 68 patients (90%) overall, and were similar in EC-IC and IC-IC bypass groups (91% and 89%, respectively). Changes in Glasgow Outcome Scale score were slightly more favorable with IC-IC bypass (6% worse or dead after IC-IC bypass versus 14% with EC-IC bypass). CONCLUSIONIC-IC bypasses compare favorably to EC-IC bypasses in terms of aneurysm obliteration rates, bypass patency rates, and neurological outcomes. IC-IC bypasses can be more technically challenging to perform, but they do not require harvest of extracranial donor arteries, spare patients a neck incision, shorten interposition grafts, are protected inside the cranium, use caliber-matched donor and recipient arteries, and are not associated with ischemic complications during temporary arterial occlusions. IC-IC bypass can replace conventional EC-IC bypass with more anatomic reconstructions for selected aneurysms involving the middle cerebral artery, posteroinferior cerebellar artery, anterior cerebral artery, and basilar apex.

Glial Cell Aquaporin-4 Overexpression in Transgenic Mice Accelerates Cytotoxic Brain Swelling

Aquaporin-4 (AQP4) is a water transport protein expressed in glial cell plasma membranes, including glial cell foot processes lining the blood-brain barrier. AQP4 deletion in mice reduces cytotoxic brain edema produced by different pathologies. To determine whether AQP4 is rate-limiting for brain water accumulation and whether altered AQP4 expression, as occurs in various pathologies, could have functional importance, we generated mice that overexpressed AQP4 in brain glial cells by a transgenic approach using the glial fibrillary acid protein promoter. Overexpression of AQP4 protein in brain by ∼2.3-fold did not affect mouse survival, appearance, or behavior, nor did it affect brain anatomy or intracranial pressure (ICP). However, following acute water intoxication produced by intraperitoneal water injection, AQP4-overexpressing mice had an accelerated progression of cytotoxic brain swelling, with ICP elevation of 20 ± 2 mmHg at 10 min, often producing brain herniation and death. In contrast, ICP elevation was 14 ± 2 mmHg at 10 min in control mice and 9.8 ± 2 mmHg in AQP4 knock-out mice. The deduced increase in brain water content correlated linearly with brain AQP4 protein expression. We conclude that AQP4 expression is rate-limiting for brain water accumulation, and thus, that altered AQP4 expression can be functionally significant.

Aquaporins: role in cerebral edema and brain water balance.

The regulation of water balance in the brain is crucial. A disruption in this equilibrium causes an increase in brain water content that significantly contributes to the pathophysiology of traumatic brain injury, hydrocephalus, and a variety of neurological disorders. The discovery of the aquaporin (AQP) family of membrane water channels has provided important new insights into the physiology and pathology of brain water homeostasis. A number of recent studies are described in the review that demonstrated the important role of AQP1 and AQP4 in brain water balance and cerebral edema. Phenotypic analyses of AQP deficient mice have allowed us to explore the role of these membrane water channels in the mechanisms of cytotoxic edema, vasogenic edema, and CSF production. These studies indicate that AQP4 plays significant role in the development of cytotoxic edema and the absorption of excess brain water resulting from vasogenic edema. They also have demonstrated the role of AQP1 in CSF production and maintenance of steady-state ICP. The ability to modulate water flux through AQP deletion has provided new insights into brain water homeostasis and suggested a number of new research directions. However, these efforts have not yet translated to the treatment human clinical diseases. These advances will require the development of AQP inhibitors and activators to establish the benefit modulating the function of these water channels.

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 ~ 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/b16) 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.—Lu, D. C., Zhang, H., Zador, Z., Verkman, A. S. Impaired olfaction in mice lacking aquaporin‐4 water channels. FASEB J. 22, 3216–3223 (2008)

Novel variants in human Aquaporin-4 reduce cellular water permeability

Cerebral edema contributes significantly to morbidity and mortality after brain injury and stroke. Aquaporin-4 (AQP4), a water channel expressed in astrocytes, plays a key role in brain water homeostasis. Genetic variants in other aquaporin family members have been associated with disease phenotypes. However, in human AQP4, only one non-synonymous single-nucleotide polymorphism (nsSNP) has been reported, with no characterization of protein function or disease phenotype. We analyzed DNA from an ethnically diverse cohort of 188 individuals to identify novel AQP4 variants. AQP4 variants were constructed by site-directed mutagenesis and expressed in cells. Water permeability assays in the cells were used to measure protein function. We identified 24 variants in AQP4 including four novel nsSNPs (I128T, D184E, I205L and M224T). We did not observe the previously documented M278T in our sample. The nsSNPs found were rare ( approximately 1-2% allele frequency) and heterozygous. Computational analysis predicted reduced function mutations. Protein expression and membrane localization were similar for reference AQP4 and the five AQP4 mutants. Cellular assays confirmed that four variant AQP4 channels reduced normalized water permeability to between 26 and 48% of the reference (P < 0.001), while the M278T mutation increased normalized water permeability (P < 0.001). We identified multiple novel AQP4 SNPs and showed that four nsSNPs reduced water permeability. The previously reported M278T mutation resulted in gain of function. Our experiments provide insight into the function of the AQP4 protein. These nsSNPs may have clinical implications for patients with cerebral edema and related disorders.

Microfiberoptic fluorescence photobleaching reveals size-dependent macromolecule diffusion in extracellular space deep in brain

Diffusion in brain extracellular space (ECS) is important for nonsynaptic intercellular com‐munication, extracellular ionic buffering, and delivery of drugs and metabolites. We measured macromolecu‐lar diffusion in normally light‐inaccessible regions of mouse brain by microfiberoptic epifluorescence photo‐bleaching, in which a fiberoptic with a micron‐size tip is introduced deep in brain tissue. In brain cortex, the diffusion of a noninteracting molecule [fluorescein isothiocyanate (FITC)‐dextran, 70 kDa] was slowed 4.5 ± 0.5‐fold compared with its diffusion in water (Do/D), and was depth‐independent down to 800 μm from the brain surface. Diffusion was significantly accelerated (Do/D of 2.9±0.3) in mice lacking the glial water channel aquaporin‐4. FITC‐dextran diffusion varied greatly in different regions of brain, with Do/D of 3.5 ± 0.3 in hippocampus and 7.4 ± 0.3 in thalamus. Remarkably, Do/D in deep brain was strongly dependent on solute size, whereas diffusion in cortex changed little with solute size. Mathematical modeling of ECS diffusion required nonuniform ECS dimensions in deep brain, which we call “heterometricity,” to account for the size‐dependent diffusion. Our results provide the first data on molecular diffusion in ECS deep in brain in vivo and demonstrate previously unrecognized hindrance and heterometricity for diffusion of large macromolecules in deep brain.—Zador Z., Magzoub, M., Jin, S., Manley, G. T., Papadopoulos, M. C., Verkman A. S. Microfiberoptic fluorescence photobleaching reveals size‐dependent macromolecule diffusion in extracellular space deep in brain. FASEB J. 22, 870–879 (2008)

Distal Aneurysms of Intracranial Arteries: Application of Numerical Nomenclature, Predilection for Cerebellar Arteries, and Results of Surgical Management

BACKGROUND Distal intracranial aneurysms are rare, have unclear origins, and are frequently nonsaccular. Published clinical experience with these aneurysms is limited. OBJECTIVE To examine differences between distal aneurysms of cerebral and cerebellar arteries and to examine results associated with surgical therapy in 140 patients. METHODS Distal aneurysms in the cerebral arteries were defined as outside the circle of Willis, on or beyond the A2 anterior cerebral artery, M2 middle cerebral artery, or P2 posterior cerebral segments. Distal aneurysms in the cerebellar arteries were on or beyond the s2 superior cerebellar artery, a2 anterior inferior cerebellar artery, or p2 posterior inferior cerebellar artery segments. Clinical data, microsurgical technique, and patient outcomes were reviewed. RESULTS The incidence of distal cerebellar artery aneurysms was 4.3 times greater than distal cerebral artery aneurysms (6.5% vs. 28.6%; P< 0.01). The A3 anterior cerebral artery segment and the p2 and p3 posterior inferior cerebellar artery segments were the most common sites. Presentation with aneurysm rupture was more frequent with cerebellar aneurysms (65% vs. 40%; P< 0.05). Distal cerebellar artery aneurysms were less likely than distal cerebral artery aneurysms to be clipable (40% vs. 72%; P< 0.01), with 42% treated with trapping alone. Overall, 14% required a bypass. CONCLUSIONS Distal intracranial aneurysms have a predilection for cerebellar arteries and are not as rare as the literature suggests. Application of standardized nomenclature for segmental anatomy to these lesions will increase the precision of anatomic description and clarity of clinical discourse. Although technically difficult, good clinical results can be expected with surgical management.

Rotational vertebral artery occlusion-series of 9 cases.

BACKGROUND:Rotational vertebral artery syndrome (RVAS) is a rare entity about which previously published studies are mostly limited to individual case reports. OBJECTIVE:To report our decade-long experience with this syndrome in 9 patients with compression ranging from the occiput to C6. METHODS:We utilized a posterior approach for lesions rostral to C4 and an anterior approach for lesions at or caudal to C4. Furthermore, we demonstrated the feasibility and efficacy of a minimally invasive posterior cervical approach. Patient profile, operative indications, surgical approach, operative findings, complications, and long-term follow-up were reviewed and discussed. RESULTS:Average follow-up was 47 months. All procedures provided excellent outcomes by Glasgow Outcome Scale scores. The anterior approach had significantly less blood loss (187.5 mL vs 450 mL, P = .00016) and shorter hospitalization length (2 days vs 4.5 days; P = .0001) compared with the far-lateral approach. There was one complication of cervical instability in the far-lateral approach cohort. As an alternative to the far-lateral surgery, a minimally invasive approach resulted in shorter hospitalization (2 days) and less blood loss (10 mL) while avoiding the complication of cervical instability. CONCLUSION:We demonstrated the safety, efficacy, and durability of 3 surgical approaches for RVAS. Proper examination, preoperative imaging, and surgical planning were necessary for a satisfactory outcome.

The new licox combined brain tissue oxygen and brain temperature monitor: Assessment of in vitro accuracy and clinical experience in severe traumatic brain injury

OBJECTIVE Monitoring of brain tissue oxygen tension is increasingly being used to monitor patients after severe traumatic brain injury and to guide therapies aimed at maintaining brain tissue oxygen tension above threshold levels. The new Licox PMO combined oxygen and temperature catheter (Integra LifeSciences, Plainsboro, NJ) combines measurements of oxygen tension and temperature in a single probe inserted through a bolt mechanism. In this study, we sought to evaluate the accuracy of the new Licox PMO probe under controlled laboratory conditions and to assess the accuracy of oxygen tension and temperature measurements and the new automated card calibration system. We also describe our clinical experience with the Licox PMO probe. METHODS Oxygen tension was measured in a 2-chambered apparatus at different oxygen tensions and temperatures. The new card calibration system was compared with a manually calibrated system. Rates of hematoma, infection, and dislodgement in our clinical experience were recorded. RESULTS The new Licox PMO probe accurately measures oxygen tension over a wide range of oxygen concentrations and physiological temperatures, but it does have a small tendency to underestimate oxygen tension (mean error, −3.8 ± 3.5%) that is more pronounced between the temperatures of 33 and 39°C. The thermistor of the PMO probe also has a tendency to underestimate temperature when compared with a resistance thermometer (mean error, −0.67 ± 0.22°C). The card calibration system was also found to introduce some variability in measurements of oxygen tension when compared with a manually calibrated system. Clinical experience with the new probe indicates good placement within the white matter using the improved bolt system and low rates of hematoma (2.9%), infection (0%), and dislodgement (5.9%). conclusion The new Licox PMO probe is accurate but has a small, consistent tendency to under-read oxygen tension that is more pronounced at higher temperatures. The probe tends to under-read temperature by 0.5 to 0.8°C across temperatures, suggesting that caution should be used when brain temperature is measured with the Licox PMO probe and used to guide temperature-directed treatment strategies. The Licox PMO probe improves upon previous models in allowing consistent and accurate placement in the white matter and obviating the need for placement of 2 separate probes to measure oxygen tension and temperature.