Elevated CSF outflow resistance associated with impaired lymphatic CSF absorption in a rat model of kaolin-induced communicating hydrocephalus.

BACKGROUND: We recently reported a lymphatic cerebrospinal fluid (CSF) absorption deficit in a kaolin model of communicating hydrocephalus in rats with ventricular expansion correlating negatively with the magnitude of the impediment to lymphatic function. However, it is possible that CSF drainage was not significantly altered if absorption at other sites compensated for the lymphatic defect. The purpose of this study was to investigate the impact of the lymphatic absorption deficit on global CSF absorption (CSF outflow resistance). METHODS: Kaolin was injected into the basal cisterns of Sprague Dawley rats. The development of hydrocephalus was assessed using magnetic resonance imaging (MRI). In one group of animals at about 3 weeks after injection, the movement of intraventricularly injected iodinated human serum albumin (125I-HSA) into the olfactory turbinates provided an estimate of CSF transport through the cribriform plate into nasal lymphatics (n = 18). Control animals received saline in place of kaolin (n = 10). In a second group at about 3.5 weeks after kaolin injection, intraventricular pressure was measured continuously during infusion of saline into the spinal subarachnoid space at various flow rates (n = 9). CSF outflow resistance was calculated as the slope of the steady-state pressure versus flow rate. Control animals for this group either received no injections (intact: n = 11) or received saline in place of kaolin (n = 8). RESULTS: Compared to saline injected controls, lateral ventricular volume in the kaolin group was significantly greater (0.087 +/- 0.013 ml, n = 27 versus 0.015 +/- 0.001 ml, n = 17) and lymphatic function was significantly less (2.14 +/- 0.72% injected/g, n = 18 versus 6.38 +/- 0.60% injected/g, n = 10). Additionally, the CSF outflow resistance was significantly greater in the kaolin group (0.46 +/- 0.04 cm H2O microL(-1) min, n = 9) than in saline injected (0.28 +/- 0.03 cm H2O microL(-1) min, n = 8) or intact animals (0.18 +/- 0.03 cm H2O microL(-1) min, n = 11). There was a significant positive correlation between CSF outflow resistance and ventricular volume. CONCLUSIONS: The data suggest that the impediment to lymphatic CSF absorption in a kaolin-induced model of communicating hydrocephalus has a significant impact on global CSF absorption. A lymphatic CSF absorption deficit would appear to play some role (either direct or indirect) in the pathogenesis of ventriculomegaly.

Intraventricular injection of antibodies to beta1-integrins generates pressure gradients in the brain favoring hydrocephalus development in rats.

In some tissues, the injection of antibodies to the beta(1)-integrins leads to a reduction in interstitial fluid pressure, indicating an active role for the extracellular matrix in tissue pressure regulation. If perturbations of the matrix occur in the periventricular area of the brain, a comparable lowering of interstitial pressures may induce transparenchymal pressure gradients favoring ventricular expansion. To examine this concept, we measured periventricular (parenchymal) and ventricular pressures with a servo-null micropipette system (2-microm tip) in adult Wistar rats before and after anti-integrin antibodies or IgG/IgM isotype controls were injected into a lateral ventricle. In a second group, the animals were kept for 2 wk after similar injections and after euthanization, the brains were removed and assessed for hydrocephalus. In experiments in which antibodies to beta(1)-integrins (n = 10) but not isotype control IgG/IgM (n = 7) were injected, we observed a decline in periventricular pressures relative to the preinjection values. Under similar circumstances, ventricular pressures were elevated (n = 10) and were significantly greater than those in the periventricular interstitium. We estimated ventricular to periventricular pressure gradients of up to 4.3 cmH(2)O. In the chronic preparations, we observed enlarged ventricles in many of the animals that received injections of anti-integrin antibodies (21 of 29 animals; 72%) but not in any animal receiving the isotype controls. We conclude that modulation/disruption of beta(1)-integrin-matrix interactions in the brain generates pressure gradients favoring ventricular expansion, suggesting a novel mechanism for hydrocephalus development.

Memantine treatment of juvenile rats with kaolin-induced hydrocephalus.

Memantine is a selective, non-competitive N-methyl-D-aspartate (NMDA) receptor antagonist that has previously been shown to have neuroprotective qualities in some animal models of neurologic disease. We hypothesized that memantine therapy would improve behavioral, neuropathological, and/or biochemical outcomes in juvenile rats with kaolin-induced hydrocephalus. Three-week old rats received an injection of kaolin (aluminum silicate) into the cisterna magna. Magnetic resonance imaging was performed one week later to assess ventricle size and stratify rats to three treatment groups. Rats were blindly treated daily for three weeks with saline or 10 or 30 mg/kg/day memantine. Behavior measures were performed weekly. Histologic and biochemical evaluations were performed at termination. Hydrocephalic rats showed no differences in weight among treatment groups. Memantine treatment stabilized ventricular enlargement in both low and high dose groups. The high dose group exhibited increased motor activity in open field chambers compared to the vehicle-treated group. However, there were no significant differences between the three hydrocephalic treatment groups for other behavioral tasks. Ventriculomegaly was associated with periventricular white matter damage. Glial fibrillary acidic protein (GFAP) content was higher in the low dose memantine group compared to vehicle-treated group, but there were no differences in GFAP-immunoreactive astrocytes or Iba-1- immunoreactive microglia between groups. Memantine therapy stabilized ventricular expansion and improved some behavioral measures but did not reduce brain tissue changes in juvenile rats with kaolin-induced hydrocephalus.

Properties of the lymphatic cerebrospinal fluid transport system in the rat: impact of elevated intracranial pressure.

Previous studies suggested that a major portion of cerebrospinal fluid (CSF) is absorbed by extracranial lymphatics located in the olfactory turbinates. The objective of this study was to determine the impact of elevated intracranial pressure (ICP) on downstream cervical lymphatic pressures in the rat. Pressures were measured in the deep cervical lymph nodes using a servo-null micropressure system. A catheter was placed in a lateral ventricle and fluid was infused from a reservoir at defined ICPs. When Ringer's solution was infused, elevations of ICP from 10 to 50 cm H2O resulted on average in a reduction of diastolic cervical node pressures. In contrast, when a diluted plasma solution (80% plasma in Ringer's) was infused, downstream diastolic lymphatic pressures increased as ICP was elevated to 50 cm H2O. These data are consistent with the view that much of the CSF-derived water that convects into the lymphatics is absorbed into the ethmoidal or nodal blood vessels. This study supports the concept of fluid continuity between the subarachnoid space and extracranial lymphatics and suggests that this loss of CSF-derived water may act as a safety mechanism to reduce the volume load to the downstream lymphatic vessels.

Development of cerebrospinal fluid absorption sites in the pig and rat: connections between the subarachnoid space and lymphatic vessels in the olfactory turbinates.

The textbook view that cerebrospinal fluid (CSF) absorption occurs mainly through the arachnoid granulations and villi is being challenged by quantitative and qualitative studies that support a major role for the lymphatic circulation in CSF transport. There are many potential sites at which lymphatics may gain access to CSF but the primary pathway involves the movement of CSF through the cribriform plate foramina in association with the olfactory nerves. Lymphatics encircle the nerve trunks on the extracranial surface of the cribriform plate and absorb CSF. However, the time during development in which the CSF compartment and extracranial lymphatic vessels connect anatomically is unclear. In this report, CSF-lymphatic connections were investigated using the silastic material Microfil and a soluble Evan's blue-protein complex in two species; one in which significant CSF synthesis by the choroid plexus begins before birth (pigs) and one in which CSF secretion is markedly up regulated within the first weeks after birth (rats). We examined a total of 46 pig fetuses at embryonic (E) day E80-81, E92, E101, E110 (birth at 114 days). In rats, we investigated a total of 115 animals at E21 (birth at 21 days), postnatal (P) day P1-P9, P12, P13, P15, P22, and adults. In pigs, CSF-lymphatic connections were observed in the prenatal period as early as E92. Before this time (E80-81 fetuses) CSF-lymphatic connections did not appear to exist. In rats, these associations were not obvious until about a week after birth. These data suggest that the ability of extracranial lymphatic vessels to absorb CSF develops around the time that significant volumes of CSF are being produced by the choroid plexus and further support an important role for lymphatic vessels in CSF transport.

A MATHEMATICAL ANALYSIS OF PHYSIOLOGICAL AND MOLECULAR MECHANISMS THAT MODULATE PRESSURE GRADIENTS AND FACILITATE VENTRICULAR EXPANSION IN HYDROCEPHALUS.

Perhaps the greatest paradox in the hydrocephalus field is the failure of researchers to consistently measure transmantle pressure gradients (ventricle to subarachnoid space) in either human or animal models of the communicating form of the disorder. Without such a gradient, conceptualization of how ventricular distention occurs is difficult. Based on evidence from both a mathematical model [35] and experiments in skin [51], we observed that the intraventricular injection of anti-ß1 integrin antibodies in rat brains results in a reduction of periventricular pressures to values below those monitored in the ventricles. In addition, many of these animals developed hydrocephalus [30]. We conclude that the dissociation of ß1 integrins from the surrounding matrix fibers generates pressure gradients favouring ventricular expansion suggesting a novel mechanism for hydrocephalus development. Several issues, however, need further clarification. If hydrostatic pressure declines in the periventricular tissues then fluid absorption must occur. Aquaporin-4 (AQP4) is a likely candidate for this absorption as it is the predominant water channel in the brain. Indeed, when capillary function is negated, periventricular interstitial fluid pressures increase after anti-ß1 integrin antibody administration. This suggests that capillary absorption of parenchymal water may play a pivotal role in the generation of pressure gradients in our hydrocephalus model. Focusing on these issues, we present two poroelastic models to investigate the role of intramantle pressure gradients in ventriculomegaly and to determine if integrin-matrix disassociation represents a complete causative mechanism for hydrocephalus development.

Quantification of cerebrospinal fluid transport across the cribriform plate into lymphatics in rats.

A major pathway by which cerebrospinal fluid (CSF) is removed from the cranium is transport through the cribriform plate in association with the olfactory nerves. CSF is then absorbed into lymphatics located in the submucosa of the olfactory epithelium (olfactory turbinates). In an attempt to provide a quantitative measure of this transport, 125I-human serum albumin (HSA) was injected into the lateral ventricles of adult Fisher 344 rats. The animals were killed at 10, 20, 30, 40, and 60 min after injection, and tissue samples, including blood (from heart puncture), skeletal muscle, spleen, liver, kidney, and tail were excised for radioactive assessment. The remains were frozen. To sample the olfactory turbinates, angled coronal tissue sections anterior to the cribriform plate were prepared from the frozen heads. The average concentration of 125I-HSA was higher in the middle olfactory turbinates than in any other tissue with peak concentrations achieved 30 min after injection. At this point, the recoveries of injected tracer (percent injected dose/g tissue) were 9.4% middle turbinates, 1.6% blood, 0.04% skeletal muscle, 0.2% spleen, 0.3% liver, 0.3% kidney, and 0.09% tail. The current belief that arachnoid projections are responsible for CSF drainage fails to explain some important issues related to the pathogenesis of CSF disorders. The rapid movement of the CSF tracer into the olfactory turbinates further supports a role for lymphatics in CSF absorption and provides the basis of a method to investigate the novel concept that diseases associated with the CSF system may involve impaired lymphatic CSF transport.