Endoscope-assisted posterior approach for the resection of ventral intradural spinal cord tumors: report of two cases.

We report our experience with the removal of ventral intradural spinal tumors through a posterior approach with the assistance of an endoscope. The endoscope allowed us to remove the tumor with minimal retraction of the spinal cord and to inspect for involvement of structures that were difficult to visualize with the microscope. Moreover, it obviated the use of a more involved anterior approach. This is the second report of the utilization of the endoscope for removal of intradural spinal cord tumors. Our data are discussed in the context of prior reported morphometric studies involving the spinal cord and expand the indications for the endoscope.

An efficient method for the culturing and generation of neurons and astrocytes from second trimester human central nervous system tissue.

The isolation, culturing and expansion of human neural progenitors cells has important potential clinical applications in cellular transplantation strategies as well as in developmental studies involving the central nervous system (CNS). This study describes an efficient method to culture neurons and astrocytes as primary cultures, as well as from proliferative progenitor cells derived from second trimester fetal CNS tissue. Second trimester fetal human tissue was mechanically dissociated and subjected to trypsin-dissociation and trituration. The resulting suspension was passed over a Percoll density gradient. The middle (second) fraction of cells was centrifuged to yield a homogenous population of cells with 80%-90% viability. These cells were either cultured directly on laminin coated dishes with defined medium supplemented with fetal bovine serum or in defined medium supplemented with growth factors including epidermal growth factor, basic fibroblast growth factor and leukemia inhibitory factor. The primary cell cultures yielded neurons and astrocytes after 3-5 days in vitro verified by immunostaining with MAP2ab and GFAP. Cells exposed to growth factor supplemented medium formed free-floating spheres within one week. Upon growth factor removal and plating on laminin-coated dishes, brain derived spheres gave rise to neurons, astrocytes and oligodendrocytes; spinal cord derived spheres generated only astrocytes. This protocol describes an efficient method to generate and culture neurons and astrocytes from second trimester human CNS tissue that may be useful in transplantation and developmental studies.

Comparison of neural precursor cell fate in second trimester human brain and spinal cord.

Neural transplantation holds promise for the treatment of traumatic brain and spinal cord injury by replacing lost cellular elements as well as repairing neural damage. Fetal human stem cells derived from central nervous system (CNS) tissue are potential transplantable sources for all cell types found in the mature human nervous system including neurons, astrocytes and oligodendroglia. Although nearly all areas of the fetal human neuraxis contain undifferentiated neural precursor cells, the phenotypic fate of the daughter cells might vary from one region to another during a specific developmental period. The purpose of this study was to compare the various cell types derived from neural precursors cultured from second trimester fetal human brain and spinal cord. To this end, brains (n = 8) and spinal cords (n = 8) of 15-24 week fetuses were dissociated and grown in culture medium supplemented with epidermal growth factor (EGF), basic fibroblast growth factor (FGF) and leukemia inhibitory factor (LIF). The proliferating precursor cells from both brain and spinal cord grew as spherical masses that were plated on laminin-coated dishes after seven days in culture. During the next 5-7 days, the cells that emerged from these spheres were fixed and processed for immunocytochemistry. Brain derived spheres gave rise to cells expressing antigens specific for neurons (MAP-2ab and neuron specific-intermediate filaments), astrocytes (GFAP) and oligodendrocytes (A007). In contrast, cells that emerged from spinal cord derived spheres were only immunoreactive for GFAP. These data suggest that neuroepithelial precursor cells from different CNS regions, although similar in their responsiveness to proliferative growth factors, might differ in their ability to generate different cell types in the adult CNS.

Neurotransmitter distribution in the second trimester fetal human corpus striatum.

One experimental strategy that may offer hope in the neurodegenerative disorder Huntington's disease (HD) has been neural transplantation. In HD, most of the pathological changes occur in the corpus striatum. Fetal human striatal implants will most likely be the first transplant strategy attempted in clinical trials to replace lost neurons and/or prevent the degeneration of neurons destined to die. The temporal expression of neurotransmitters in the developing human corpus striatum is a key factor in determining the optimum age of transplantable tissue. To this end, an immunocytochemical analysis of various neurotransmitters was performed on second trimester human brains. Antibodies against acetylcholine, gamma-aminobutyric acid, enkephalin, neuropeptide-Y and substance P were used in ten human fetal brains ranging from 13 to 21 weeks gestation. The presence and pattern of distribution for these neurotransmitters varied in the different parts of the corpus striatum (globus pallidus, putamen, caudate nucleus). These results are compared to the already existing data for the adult human corpus striatum.

Transplantation of human fetal brain cells into ischemic lesions of adult gerbil hippocampus.

OBJECT: The goal of this study was to establish whether transplanted cells derived from fetal human brain can survive in an ischemic lesion. METHODS: Sixteen adult male Mongolian gerbils underwent transient bilateral common carotid artery occlusion. One week later, cell suspensions prepared from fetal human brain were injected using stereotactic guidance into the CA1 region of the hippocampus on one side. On the contralateral side injection of the cell suspension medium only was performed. One week after transplantation, the animals were perfusion fixed and their brains were processed for histological studies as well as expression of neuron and glia-specific antigens. Data from ischemic animals were compared with eight nonischemic gerbils that served as sham-operated controls. Last, the in vivo data were correlated with observations made from matching in vitro cultures of the fetal brain cell suspension. The in vivo data indicated that transplanted human fetus-derived brain cells survived in ischemic lesions of gerbil hippocampus after 1 week, provided that the host animal underwent adequate immunosuppression and the transplanted cells were not incorporated into the scar caused by the transplantation procedure. Unlike their in vivo counterparts, after 1 week, most cultured fetal brain cells expressed either neuron- or astrocyte-specific antigens. CONCLUSIONS: This work demonstrates that xenotransplanted fetal human brain cells are able to survive in an ischemic lesion in a rodent model. These data might be useful for future neural transplantation studies of treatments for cerebrovascular ischemia in humans.

Postural changes in intracranial pressure in chronically shunted patients.

A subset of hydrocephalic patients with indwelling shunts become symptomatic when they are upright and active. Intracranial pressure (ICP) measurements in these patients have shown a significant drop in pressure when the patient is upright with return to normal levels when the patient is supine. In 20 chronically shunted hydrocephalic patients who previously had no siphon protection devices, ICP changes in supine and upright position were studied at the time when the patient had external ventriculostomy for treatment of shunt infection. Our hypothesis was that these patients might display rapid changes in ICP from fluid shifts occurring in non-CSF compartments. To minimize the effects of hysteresis, drift and zero-point error, measurements were made using a fluid manometer rather than a strain gauge pressure transducer. The pressure-volume index was estimated using the standard technique of bolus injection. Intracranial CSF volume was estimated on CT scans. The fluid shift was calculated using a mathematical model of the CSF compartment that incorporates negative pressure and volume components that permits simulation of siphoning. Sixteen patients had small, slit ventricles; 3 patients had moderate-sized ventricles and in 1 patient the ventricular size was normal. The average intracranial CSF volume estimated on CT scan was 12 cm(3). There was a mean drop in ICP in the upright position of 159 mm H(2)O. The mean PVI of 42 ml suggested a volume displacement out of proportion to the available intracranial CSF volume. Based on these findings, we conclude that even in the absence of drainage through the shunt, chronically shunted patients still display a fall in ICP when assuming the upright position. This raises the possibility of fluid shifts other than of CSF through nonshunt pathways. Possible mechanisms involving altered CSF-venous system interaction are discussed.

Cellular transplantation and spinal cord injury.

Spinal cord injury is often characterized by immediate and irreversible loss of sensory and motor functions below the level of injury. Cellular transplantation in various experimental models of spinal cord injury has been used as a strategy for reducing deficits and improving functional recovery. The general strategy has been aimed at promoting regeneration of intrinsic injured axons with the development of alternative pathways that facilitate a partial functional connection. Other objectives of cellular transplantation studies have included replacement of lost cellular elements, alleviation of chronic pain, and modulation of the inflammatory response after injury. This review focuses on the cell types that have been used in spinal cord transplantation studies in the context of evolving biological perspectives, technological advances, and new therapeutic strategies and serves as a point of reference for future studies.

Chemical meningitis from bile reflux in a lumbar-gallbladder shunt.

OBJECTIVE AND IMPORTANCE: The gallbladder is used to divert cerebrospinal fluid (CSF) in patients with hydrocephalus when all other sites have been exhausted. This is seen in hydrocephalic patients who have reached teenage years but have undergone multiple shunt revisions, abdominal operations and repeated neck vein cannulations during childhood. One complication of the lumbar-gallbladder shunt is discussed as well as its pathophysiologic theory and management. CLINICAL PRESENTATION: A case of a patient with a lumbar-gallbladder shunt who developed chemical meningitis from reflux of bile into the CSF space is presented. The patient presented with generalized seizures. INTERVENTION: Included: ventilatory support, externalization of the shunt, correction of the metabolic abnormalities and administration of anticonvulsants and steroids. CONCLUSION: This case illustrates an unusual occurrence of reflux of bile into the CNS through a lumbar-gallbladder shunt in a patient with long-term complex communicating hydrocephalus. It also demonstrates its mode of presentation and successful management. To our knowledge, this is the first case report of its kind.

Anterolateral lumbar lipomyelomeningocele: case report and review of the literature.

OBJECTIVE AND IMPORTANCE: Meningoceles associated with defects of the abdominal wall are exceedingly rare. One such complex case is presented along with a review of the relevant literature. The current pathophysiological theories and surgical management are discussed. CLINICAL PRESENTATION: A case of a patient with an anterolateral lumbar lipomyelomeningocele associated with multiple congenital anomalies, including defects in the abdominal wall and urogenital system, is presented. The lipomyelomeningocele presented as an expanding abdominal mass. INTERVENTION: Ventriculoatrial shunting and two operations to repair the myelomeningocele were performed to control the expanding abdominal mass. CONCLUSION: This report illustrates that the surgical management of complex lipomyelomeningoceles is similar to the more common types. It also demonstrates that in the infant in whom the intra-abdominal cerebrospinal fluid collection is the primary cause of the symptoms, cerebrospinal fluid shunting may be used to delay the definitive repair until the dura strengthens, thus avoiding the complications of complex repairs of dura with low tensile strength.

Ependymal/subependymal zone cells of postnatal and adult songbird brain generate both neurons and nonneuronal siblings in vitro and in vivo.

The songbird forebrain continues to generate neurons in adulthood, from precursor cells located in the ependymal /subependymal zone (SZ) over the mediocaudal neostriatum. Precursor mitosis is followed by migration of neuronal daughter cells into the underlying forebrain, along radial fibers derived from the SZ. To define the ontogeny of both the new neurons and their radial guide cells, we employed retroviral insertion of the lacZ gene into neostriatal SZ precursor cells derived from postnatal and adult songbirds. We found that single SZ cells generate both neurons and substrate glia in vitro, and in an analogous fashion, both neurons and radial cells in vivo. This suggests that newly generated neurons and radial cells of the adult avian brain derive from a common pluripotential progenitor.

Transient coupling of Ng-CAM expression to NgCAM-dependent calcium signaling during migration of new neurons in the adult songbird brain.

The adult avian forebrain continues to generate neurons from subependymal zone (SZ) precursor cells, whose neuronal progeny migrate into the brain upon radial guide fibers. These neurons express the immunoglobulin-family adhesion molecule NgCAM, and their migration in culture is disrupted by anti-NgCAM Fab. Confocal imaging of adult zebra finch SZ loaded with the calcium indicator fluo-3, as well as ratio imaging with the indicator fura-2, revealed that migrating new neurons responded to microgram amounts of NgCAM with reversible increments in cytosolic calcium. The calcium response to NgCAM antigen was developmentally restricted, in that it was only manifested by neurons for roughly the 3- to 4-day period between 6 and 9 DIV, even though NgCAM expression persisted tonically thereafter. The period during which NgCAM elicited a calcium signal corresponded to the postmitotic age at which new, bipolar neurons leave the adult SZ to enter the brain parenchyma in vivo. Accordingly, the calcium response to NgCAM was largely limited to morphologically bipolar cells. Anti-NgCAM IgG also evoked a neuronal calcium signal over the same restricted period that NgCAM protein exerted its effect. These findings suggest a dynamic coupling and uncoupling of calcium-dependent signal transduction pathways to a stably expressed surface adhesion molecule, whose function in a given neuron may therefore evolve with cellular maturation.

Hu protein as an early marker of neuronal phenotypic differentiation by subependymal zone cells of the adult songbird forebrain.

The avian forebrain exhibits neurogenesis in adulthood, with neuronal production from ependymal/subependymal zone (SZ) precursor cells. To follow the commitment of newborn cells to neuronal lineage, we used their expression of the Hu family of neuronal RNA-binding proteins to identify them before their migration from the SZ. Adult canaries were injected with [3H]thymidine as a marker of DNA replication, sacrificed after varying intervals, stained for Hu, and autoradiographed. We found that Hu was not expressed by premitotic precursor cells, but rather appeared within hours in their neuronal progeny, which did not embark on parenchymal migration until 4 to 7 days later. Hu was expressed by all neurons, but not glia, both in vivo and in vitro, as determined by ultrastructural analysis as well as co-localization of Hu and cell-type selective antigens. In addition, co-staining for Hu and N-cadherin, whose expression is down-regulated on neuronal emigration from the SZ, revealed their initial co-expression by neuronal daughter cells still within the SZ. These results suggest that Hu expression may be used as a very early indicator of neuronal differentiation by SZ cells. Furthermore, the data indicate that in the adult avian brain, neuronal phenotype is established within hours of precursor mitosis, even though the neuronal daughter cells do not initiate parenchymal migration for at least 4 days thereafter, following their down-regulation of N-cadherin.

Estrogens and non-estrogenic ovarian influences combine to promote the recruitment and decrease the turnover of new neurons in the adult female canary brain.

The higher vocal center (HVC) of the songbird forebrain exhibits persistent neurogenesis in adulthood, particularly in a region of the mediocaudal neostriatum that is associated with a subventricular layer of estrogen receptive cells. We asked whether estrogens might influence adult neurogenesis, by assessing the effect of ovariectomy on HVC neuronal production in the adult female canary. Fifteen 1-year-old females were separated into groups of ovariectomized, estradiol-replaced ovariectomized, and gonadally intact birds. To label dividing cells and their progeny, the birds were given [3H]thymidine for 8 days, killed 32 days later, and their brains autoradiographed. A significant rise was noted in the number of HVC neurons per section in estradiol-treated birds relative to the untreated control birds. The number of [3H]thymidine-labeled HVC neurons was also higher in the estrogen-treated birds; however, the neuronal labeling index (LI) did not vary as a function of estradiol replacement, as the total number of HVC neurons rose in parallel with the added new neurons. In contrast, the neuronal LI did rise as a result of ovariectomy, and this ovariectomy-associated increase in the LI was not reversed by estradiol. Among non-neuronal cell types, the endothelial LI was higher in estrogen-treated birds than in their untreated counterparts, suggesting estrogen-associated angiogenesis. Radioimmunoassay confirmed that serum estradiol was reduced in the castrated birds. Since estrogen appeared to promote the survival of [3H]thymidine+ neurons, we next sought to determine whether estrogen acted directly on the newly generated neurons, or rather indirectly through an intermediary cell population. To this end, we asked whether the new neurons or their precursors expressed estrogen receptor immunoreactivity (ER-IR). Five adult male canaries were given [3H]thymidine for periods ranging from 2 to 28 days, killed at varying times up to 3 weeks thereafter, then probed for ER-IR and autoradiographed. [3H]thymidine+ cells displayed no detectable ER-IR within their first 4 weeks of postmitotic life. Rather, during migration from the ventricular zone (VZ), the new neurons traversed a layer of mitotically quiescent, ER+ subventricular cells. Double labeling for ER-IR and cell-type selective antigens confirmed that these ER+ cells were neurons. These results indicate that the early survival of new neurons in the adult songbird HVC is promoted by estrogen, and may be mediated by the estrogen-stimulated paracrine release of neurotrophic agents by ER-IR subventricular neurons. Our data suggest that estrogen's promotion of neuronal survival may operate concurrently with an estrogen-independent ovarian suppression of neuronal mitogenesis.

In vitro neuronal production and differentiation by precursor cells derived from the adult human forebrain.

It has traditionally been held that the adult brain is incapable of significant self-repair, due in part to its inability to generate new neurons. Nevertheless, rodents and birds have been found to harbor neural precursor cells in adulthood. We asked whether the adult human brain might retain such precursors, by culturing samples of temporal lobe under conditions permissive for neuronal differentiation, while exposed to 3H-thymidine. Adult human temporal lobe cultures, derived from cortex, subcortex, and periventricular subependymal zone (SZ), were incubated for 7-28 d, stained for neuronal and glial antigens, and autoradiographed. Neuron-like cells were found in explant outgrowths and monolayer dissociates of SZ and periventricular white matter, but not cortex; they expressed neuronal antigens including MAP-2, MAP-5, NF, and N-CAM, and were GFAP-. Neurons responded to K+ depolarization with rapid and reversible increases in intracellular Ca2+, with much greater increments than those noted in glia. Although most neurons were not 3H-thymidine labeled, a small number of MAP-2+ and MAP-5+/GFAP- cells did incorporate 3H-thymidine, suggesting neuronal production from precursor mitosis. Rare 3H-thymidine+ neurons were also found in cultures of subventricular white matter; in these, GFAP+ astrocytic mitogenesis was common, while O4+ oligodendrocytes, although the predominant cell type, were largely postmitotic. Thus, the adult human forebrain harbors precursor cells that retain the potential for neuronal production and differentiation in vitro.

N-cadherin and Ng-CAM/8D9 are involved serially in the migration of newly generated neurons into the adult songbird brain.

In the adult avian forebrain, neurons continue to be produced in the subependymal zone (SZ), from which they migrate upon radial fibers. To identify ligands regulating this process, we studied N-cadherin and Ng-CAM/8D9 expression in HVC, a neurogenic region of the canary neostriatum. N-cadherin was relatively restricted to the SZ and was expressed by dividing, [3H]thymidine-labeled precursor cells. However, cellular N-cadherin was down-regulated prior to neuronal migration from the SZ. Addition of anti-N-cadherin Fab hastened neuronal migration from adult SZ explants, without influencing neuronal number. Unlike N-cadherin, Ng-CAM/8D9 was expressed by migrating neurons. Anti-8D9 Fab inhibited neuronal migration upon cultured ependymoglia, which did not express Ng-CAM/8D9. Thus, the departure of new neurons from the adult SZ may require their suppression of N-cadherin, whereas their subsequent migration and survival may depend upon neuronal expression of Ng-CAM/8D9 and its interaction with a heterophilic radial cell receptor.

Ruptured mycotic aneurysm presenting as an intraparenchymal hemorrhage and nonadjacent acute subdural hematoma: case report and review of the literature.

The case of a mycotic aneurysm presenting as an intraparenchymal hemorrhage and acute subdural hematoma is described. A 36-year-old man with a history of headaches and fever presented in coma. Brain computed tomography scan revealed a left occipital intraparenchymal hemorrhage and a nonadjacent acute subdural hematoma that was evacuated. Angiography demonstrated an aneurysm on the distal branch of the left posterior cerebral artery consistent with a mycotic aneurysm. This case and a review of the literature are discussed.