Adult hippocampal neurogenesis is impaired by transient and moderate developmental thyroid hormone disruption.

The hippocampus maintains a capacity for neurogenesis throughout life, a capacity that is reduced in models of adult onset hypothyroidism. The effects of developmental thyroid hormone (TH) insufficiency on neurogenesis in the adult hippocampus, however, has not been examined. Graded degrees of TH insufficiency were induced in pregnant rat dams by administration of 0, 3 or 10ppm of 6-propylthiouracil (PTU) in drinking water from gestational day (GD) 6 until weaning. Body, brain, and hippocampal weight were reduced on postnatal day (PN) 14, 21, 78 and hippocampal volume was smaller at the 10 but not 3ppm dose level. A second experiment examined adult hippocampal neurogenesis following developmental or adult onset hypothyroidism. Two male offspring from 0 and 3ppm exposed dams were either maintained on control water or exposed to 3ppm PTU to create 4 distinct treatment conditions (Control-Control; Control-PTU, PTU-Control, PTU-PTU) based on developmental and adult exposures. Beginning on the 28th day of adult exposure to 0 or 3ppm PTU, bromodeoxyuridine (BrdU, 50mg/kg, ip) was administered twice daily for 5days, and one male from each treatment was sacrificed 24h and 28days after the last BrdU dose and brains processed for immunohistochemistry. Although no volume changes were seen in the hippocampus of the neonate at 3ppm, thinning of the granule cell layer emerged in adulthood. Developmental TH insufficiency produced a reduction in newly born cells, reducing BrdU+ve cells at 1 with no further reduction at 28-days post-BrdU. Similar findings were obtained using the proliferative cell marker Ki67. Neuronal differentiations was also altered with fewer doublecortin (Dcx) expressing cells and a higher proportion of immature Dcx phenotypes seen after developmental but not adult TH insufficiency. An impaired capacity for neurogenesis may contribute to impairments in synaptic plasticity and cognitive deficits previously reported by our laboratory and others following moderate degrees of developmental TH insufficiency induced by this PTU model.

Subcortical band heterotopia in rat offspring following maternal hypothyroxinaemia: structural and functional characteristics.

Thyroid hormones (TH) play crucial roles in brain maturation and are important for neuronal migration and neocortical lamination. Subcortical band heterotopia (SBH) represent a class of neuronal migration errors in humans that are often associated with childhood epilepsy. We have previously reported the presence of SBH in a rodent model of low level hypothyroidism induced by maternal exposure to the goitrogen, propylthiouracil (PTU). In the present study, we report the dose-response characteristics of this developmental malformation and the connectivity of heterotopic neurones with other brain regions, as well as their functionality. Pregnant rats were exposed to varying concentrations of PTU through the drinking water (0-10 p.p.m.) beginning on gestational day 6 to produce graded levels of TH insufficiency. Dose-dependent increases in the volume of the SBH present in the corpus callosum were documented in the adult offspring, with a clear presence at concentrations of PTU that resulted in minor (< 15%) reductions in maternal serum thyroxine as measured when pups were weaned. SBH contain neurones, oligodendrocytes, astrocytes and microglia. Monoaminergic and cholinergic processes were prevalent and many of the axons were myelinated. Anatomical connectivity of SBH neurones to cortical neurones and the synaptic functionality of these anatomical connections was verified by ex vivo field potential recordings. SBH persisted in adult offspring despite a return to euthyroid status on termination of exposure and these offspring displayed an increased sensitivity to seizures. Features of this model are attractive with respect to the investigation of the molecular mechanisms of cortical development, the effectiveness of therapeutic intervention in hypothyroxinaemia during pregnancy and the impact of the very modest TH imbalance that accompanies exposure to environmental contaminants.

Vascular endothelial growth factor is up-regulated after status epilepticus and protects against seizure-induced neuronal loss in hippocampus.

Vascular endothelial growth factor (VEGF) is a protein factor which has been found to play a significant role in both normal and pathological states. Its role as an angiogenic factor is well-established. More recently, VEGF has been shown to protect neurons from cell death both in vivo and in vitro. While VEGF's potential as a protective factor has been demonstrated in hypoxia-ischemia, in vitro excitotoxicity, and motor neuron degeneration, its role in seizure-induced cell loss has received little attention. A potential role in seizures is suggested by Newton et al.'s [Newton SS, Collier EF, Hunsberger J, Adams D, Terwilliger R, Selvanayagam E, Duman RS (2003) Gene profile of electroconvulsive seizures: Induction of neurotrophic and angiogenic factors. J Neurosci 23:10841-10851] finding that VEGF mRNA increases in areas of the brain that are susceptible to cell loss after electroconvulsive-shock induced seizures. Because a linear relationship does not always exist between expression of mRNA and protein, we investigated whether VEGF protein expression increased after pilocarpine-induced status epilepticus. In addition, we administered exogenous VEGF in one experiment and blocked endogenous VEGF in another to determine whether VEGF exerts a neuroprotective effect against status epilepticus-induced cell loss in one vulnerable brain region, the rat hippocampus. Our data revealed that VEGF is dramatically up-regulated in neurons and glia in hippocampus, thalamus, amygdala, and neocortex 24 h after status epilepticus. VEGF induced significant preservation of hippocampal neurons, suggesting that VEGF may play a neuroprotective role following status epilepticus.

Thyroid hormone insufficiency during brain development reduces parvalbumin immunoreactivity and inhibitory function in the hippocampus.

Thyroid hormones are necessary for brain development. gamma-Amino-butyric acid (GABA)ergic interneurons comprise the bulk of local inhibitory circuitry in brain, many of which contain the calcium binding protein, parvalbumin (PV). A previous report indicated that severe postnatal hypothyroidism reduces PV immunoreactivity (IR) in rat neocortex. We examined PV-IR and GABA-mediated synaptic inhibition in the hippocampus of rats deprived of thyroid hormone from gestational d 6 until weaning on postnatal d 30. Pregnant dams were exposed to propylthiouracil (0, 3, 10 ppm) via the drinking water, which decreased maternal serum T(4) by approximately 50-75% and increased TSH. At weaning, T(4) was reduced by approximately 70% in offspring in the low-dose group and fell below detectable levels in high-dose animals. PV-IR was diminished in the hippocampus and neocortex of offspring killed on postnatal d 21, an effect that could be reversed by postnatal administration of T(4). Dose-dependent decreases in the density of PV-IR neurons were observed in neocortex and hippocampus, with the dentate gyrus showing the most severe reductions (50-75% below control counts). Altered staining persisted to adulthood despite the return of thyroid hormones to control levels. Developmental cross-fostering and adult-onset deprivation studies revealed that early postnatal hormone insufficiency was required for an alteration in PV-IR. Synaptic inhibition of the perforant path-dentate gyrus synapse evaluated in adult offspring, in vivo, revealed dose-dependent reductions in paired pulse depression indicative of a suppression of GABA-mediated inhibition. These data demonstrate that moderate degrees of thyroid hormone insufficiency during the early postnatal period permanently alters interneuron expression of PV and compromises inhibitory function in the hippocampus.

Chronic developmental lead exposure reduces neurogenesis in adult rat hippocampus but does not impair spatial learning.

The dentate granule cell (DG) layer of the hippocampal formation has the distinctive property of ongoing neurogenesis that continues throughout adult life. Although the function of these newly generated neurons and the mechanisms that control their birth are unknown, age, activity, diet and psychosocial stress have all been demonstrated to regulate this type of neurogenesis. Little information on the impact of environmental insults on this process has appeared to date. Developmental lead (Pb) exposure has been well documented to impair cognitive function in children and animals and reduce activity-dependent synaptic plasticity in the hippocampus of rodents. Therefore, we examined the effects of this classic environmental neurotoxicant on hippocampal-dependent learning and adult neurogenesis in the hippocampus. Pregnant rats were exposed to a low level of Pb-acetate (0.2%) via the drinking water from late gestation (GD 16) until weaning on postnatal day 21 (PN 21). At weaning, half of the Pb-exposed animals were weaned to control drinking water and the remainder were maintained on Pb water until termination of the study. Animals were paired- housed and on PN 75 were administered a series of injections of a thymidine analog bromodeoxyuridine (BrdU), a marker of DNA synthesis that labels proliferating cells and their progeny. At 12-h intervals for 12 days, rats received an ip injection of BrdU (50 mg/kg). Subjects were sacrificed and perfused 24 h and 28 days after the last injection. Spatial learning was assessed in an independent group of animals beginning on PN 110 using a Morris water maze. No Pb-induced impairments were evident in water maze learning. Immunohistochemistry for the detection of BrdU-labeled cells was performed on 40-microm coronal sections throughout the hippocampus. Continuous exposure to Pb (Life) reduced the total number of BrdU-positive cells at 28 days without affecting the total number of labeled cells evident 24 h after the last injection. No differences in the number of progenitor cells labeled or surviving were seen between control and treated animals whose Pb exposure was terminated at weaning. Double labeling with BrdU and the glial specific marker, glial acidic fibrillary protein (GFAP) indicated that the bulk of the surviving cells were of a neuronal rather than a glial phenotype. These data reveal that chronic low-level Pb exposure reduces the capacity for neurogenesis in the adult hippocampus. Despite deficits in synaptic plasticity previously reported from our laboratory, and now structural plasticity, no significant impact on spatial learning was detected.

Perforant path activation of ectopic granule cells that are born after pilocarpine-induced seizures.

Granule cells in the dentate gyrus are born throughout life, and various stimuli can affect their development in the adult brain. Following seizures, for instance, neurogenesis increases greatly, and some new cells migrate to abnormal (ectopic) locations, such as the hilus. Previous electrophysiological studies of this population have shown that they have intrinsic properties that are similar to normal granule cells, but differ in other characteristics, consistent with abnormal integration into host circuitry. To characterize the response of ectopic hilar granule cells to perforant path stimulation, intracellular recordings were made in hippocampal slices from rats that had pilocarpine-induced status epilepticus and subsequent spontaneous recurrent seizures. Comparisons were made with granule cells located in the granule cell layer of both pilocarpine- and saline-treated animals. In addition, a few ectopic hilar granule cells were sampled from saline-treated rats. Remarkably, hilar granule cells displayed robust responses, even when their dendrites were not present within the molecular layer, where perforant path axons normally terminate. The evoked responses of hilar granule cells were similar in several ways to those of normally positioned granule cells, but there were some differences. For example, there was an unusually long latency to onset of responses evoked in many hilar granule cells, especially those without molecular layer dendrites. Presumably this is due to polysynaptic activation by the perforant path. These results indicate that synaptic reorganization after seizures can lead to robust activation of newly born hilar granule cells by the perforant path, even when their dendrites are not in the terminal field of the perforant path. Additionally, the fact that these cells can be found in normal tissue and develop similar synaptic responses, suggests that seizures, while not necessary for their formation, strongly promote their generation and the development of associated circuits, potentially contributing to a lowered seizure threshold.

Spontaneous recurrent seizures after pilocarpine-induced status epilepticus activate calbindin-immunoreactive hilar cells of the rat dentate gyrus.

Although it is now established that neurogenesis of dentate gyrus granule cells increases after experimental seizures, little is currently known about the function of the new granule cells. One question is whether they become integrated into the network around them. Recent experiments that focused on the newly born granule cells in the hilus showed that indeed the new cells appear to become synchronized with host hippocampal neurons [Scharfman et al. (2000) J. Neurosci. 20, 6144-6158]. To address this issue further, we asked whether the new hilar granule cells were active during spontaneous limbic seizures that follow status epilepticus induced by pilocarpine injection. Thus, we perfused rats after spontaneous seizures and stained sections using antibodies to c-fos, a marker of neural activity, and calbindin, a marker of the newly born hilar granule cells [Scharfman et al. (2000) J. Neurosci. 20, 6144-6158]. We asked whether calbindin-immunoreactive hilar neurons were also c-fos-immunoreactive.C-fos was highly expressed in calbindin-immunoreactive hilar neurons. Approximately 23% of hilar cells that expressed c-fos were double-labeled for calbindin. In addition, other types of hilar neurons, i.e. those expressing parvalbumin or neuropeptide Y, also expressed c-fos. Yet other hippocampal neurons, including granule cells and pyramidal cells, had weak expression of c-fos at the latency after the seizure that hilar neuron expression occurred. In controls, there was very little c-fos or calbindin expression in the hilus.These results indicate that calbindin-immunoreactive hilar cells are activated by spontaneous seizures. Based on the evidence that many of these cells are likely to be newly born, the data indicate that new cells can become functionally integrated into limbic circuits involved in recurrent seizure generation. Furthermore, they appear to do so in a manner similar to many neighboring hilar neurons, apparently assimilating into the local environment. Finally, the results show that a number of hilar cell types are activated during chronic recurrent seizures in the pilocarpine model, a surprising result given that many hilar neurons are thought to be damaged soon after pilocarpine-induced status epilepticus.

Survival of dentate hilar mossy cells after pilocarpine-induced seizures and their synchronized burst discharges with area CA3 pyramidal cells.

The clinical and basic literature suggest that hilar cells of the dentate gyrus are damaged after seizures, particularly prolonged and repetitive seizures. Of the cell types within the hilus, it appears that the mossy cell is one of the most vulnerable. Nevertheless, hilar neurons which resemble mossy cells appear in some published reports of animal models of epilepsy, and in some cases of human temporal lobe epilepsy. Therefore, mossy cells may not always be killed after severe, repeated seizures. However, mossy cell survival in these studies was not completely clear because the methods did allow discrimination between mossy cells and other hilar cell types. Furthermore, whether surviving mossy cells might have altered physiology after seizures was not examined. Therefore, intracellular recording and intracellular dye injection were used to characterize hilar cells in hippocampal slices from pilocarpine-treated rats that had status epilepticus and recurrent seizures ('epileptic' rats). For comparison, mossy cells were also recorded from age-matched, saline-injected controls, and pilocarpine-treated rats that failed to develop status epilepticus. Numerous hilar cells with the morphology, axon projection, and membrane properties of mossy cells were recorded in all three experimental groups. Thus, mossy cells can survive severe seizures, and those that survive retain many of their normal characteristics. However, mossy cells from epileptic tissue were distinct from mossy cells of control rats in that they generated spontaneous and evoked epileptiform burst discharges. Area CA3 pyramidal cells also exhibited spontaneous and evoked bursts. Simultaneous intracellular recordings from mossy cells and pyramidal cells demonstrated that their burst discharges were synchronized, with pyramidal cell discharges typically beginning first. From these data we suggest that hilar mossy cells can survive status epilepticus and chronic seizures. The fact that mossy cells have epileptiform bursts, and that they are synchronized with area CA3, suggest a previously unappreciated substrate for hyperexcitability in this animal model.

Electrophysiological effects of exogenous and endogenous kynurenic acid in the rat brain: studies in vivo and in vitro.

In this review, recent studies on the electrophysiological effects of de novo synthesized ("endogenous") kynurenic acid (KYNA) are discussed. Endogenous KYNA is normally formed as a byproduct of tryptophan metabolism. Evidence for a physiological role in neuronal excitability has not been strong, in part because brain levels are much lower than the KD of KYNA at the glycine site of the NMDA receptor, where KYNA is thought to exert its most potent effect. The results suggest that, unexpectedly, even low concentrations of endogenous KYNA have physiological consequences. These levels of KYNA reduced the number of hippocampal slices with spontaneous epileptiform discharges after exposure to buffer lacking magnesium. However, effects on evoked responses to single afferent stimuli were not detected. Taken together, the data argue for a potentially important role of endogenous KYNA in suppression of seizure-like activity, and suggest a novel approach to anticonvulsant drug development that could have few side effects.

Boron neutron capture therapy of brain tumors: biodistribution, pharmacokinetics, and radiation dosimetry sodium borocaptate in patients with gliomas.

OBJECTIVE: The purpose of this study was to obtain tumor and normal brain tissue biodistribution data and pharmacokinetic profiles for sodium borocaptate (Na2B12H11SH) (BSH), a drug that has been used clinically in Europe and Japan for boron neutron capture therapy of brain tumors. The study was performed with a group of 25 patients who had preoperative diagnoses of either glioblastoma multiforme (GBM) or anaplastic astrocytoma (AA) and were candidates for debulking surgery. Nineteen of these patients were subsequently shown to have histopathologically confirmed diagnoses of GBM or AA, and they constituted the study population. METHODS: BSH (non-10B-enriched) was infused intravenously, in a 1-hour period, at doses of 15, 25, and 50 mg boron/kg body weight (corresponding to 26.5, 44.1, and 88.2 mg BSH/kg body weight, respectively) to groups of 3, 3, and 13 patients, respectively. Multiple samples of tumor tissue, brain tissue around the tumors, and normal brain tissue were obtained at either 3 to 7 or 13 to 15 hours after infusion. Blood samples for pharmacokinetic studies were obtained at times up to 120 hours after termination of the infusion. Sixteen of the patients underwent surgery at the Beijing Neurosurgical Institute and three at The Ohio State University, where all tissue samples were subsequently analyzed for boron content by direct current plasma-atomic emission spectroscopy. RESULTS: Blood boron values peaked at the end of the infusion and then decreased triexponentially during the 120-hour sampling period. At 6 hours after termination of the infusion, these values had decreased to 20.8, 29.1, and 62.6 microg/ml for boron doses of 15, 25, and 50 mg/kg body weight, respectively. For a boron dose of 50 mg/kg body weight, the maximum (mean +/- standard deviation) solid tumor boron values at 3 to 7 hours after infusion were 17.1+/-5.8 and 17.3+/-10.1 microg/g for GBMs and AAs, respectively, and the mean tumor value averaged across all samples was 11.9 microg/g for both GBMs and AAs. In contrast, the mean normal brain tissue values, averaged across all samples, were 4.6+/-5.1 and 5.5+/-3.9 microg/g and the tumor/normal brain tissue ratios were3.8 and 3.2 for patients with GBMs and AAs, respectively. The large standard deviations indicated significant heterogeneity in uptake in both tumor and normal brain tissue. Regions histopathologically classified either as a mixture of tumor and normal brain tissue or as infiltrating tumor exhibited slightly lower boron concentrations than those designated as solid tumor. After a dose of 50 mg/kg body weight, boron concentrations in blood decreased from 104 microg/ml at 2 hours to 63 microg/ml at 6 hours and concentrations in skin and muscle were 43.1 and 39.2 microg/g, respectively, during the 3- to 7-hour sampling period. CONCLUSION: When tumor, blood, and normal tissue boron concentrations were taken into account, the most favorable tumor uptake data were obtained with a boron dose of 25 mg/kg body weight, 3 to 7 hours after termination of the infusion. Although blood boron levels were high, normal brain tissue boron levels were almost always lower than tumor levels. However, tumor boron concentrations were less than those necessary for boron neutron capture therapy, and there was significant intratumoral and interpatient variability in the uptake of BSH, which would make estimation of the radiation dose delivered to the tumor very difficult. It is unlikely that intravenous administration of a single dose of BSH would result in therapeutically useful levels of boron. However, combining BSH with boronophenylalanine, the other compound that has been used clinically, and optimizing their delivery could increase tumor boron uptake and potentially improve the efficacy of boron neutron capture therapy.

Granule-like neurons at the hilar/CA3 border after status epilepticus and their synchrony with area CA3 pyramidal cells: functional implications of seizure-induced neurogenesis.

A group of neurons with the characteristics of dentate gyrus granule cells was found at the hilar/CA3 border several weeks after pilocarpine- or kainic acid-induced status epilepticus. Intracellular recordings from pilocarpine-treated rats showed that these "granule-like" neurons were similar to normal granule cells (i. e., those in the granule cell layer) in membrane properties, firing behavior, morphology, and their mossy fiber axon. However, in contrast to normal granule cells, they were synchronized with spontaneous, rhythmic bursts of area CA3 pyramidal cells that survived status epilepticus. Saline-treated controls lacked the population of granule-like cells at the hilar/CA3 border and CA3 bursts. In rats that were injected after status epilepticus with bromodeoxyuridine (BrdU) to label newly born cells, and also labeled for calbindin D(28K) (because it normally stains granule cells), many double-labeled neurons were located at the hilar/CA3 border. Many BrdU-labeled cells at the hilar/CA3 border also were double-labeled with a neuronal marker (NeuN). Taken together with the recent evidence that granule cells that are born after seizures can migrate into the hilus, the results suggest that some newly born granule cells migrate as far as the CA3 cell layer, where they become integrated abnormally into the CA3 network, yet they retain granule cell intrinsic properties. The results provide insight into the physiological properties of newly born granule cells in the adult brain and suggest that relatively rigid developmental programs set the membrane properties of newly born cells, but substantial plasticity is present to influence their place in pre-existing circuitry.

Improved survival after boron neutron capture therapy of brain tumors by Cereport-mediated blood-brain barrier modulation to enhance delivery of boronophenylalanine.

OBJECTIVE: Cereport (Alkermes, Inc., Cambridge, MA), or, as it has been previously called, RMP-7 (receptor-mediated permeabilizer-7), is a bradykinin analog that has been shown to produce a transient, pharmacologically mediated opening of the blood-brain barrier. The purpose of the present study was to determine whether the efficacy of boron neutron capture therapy (BNCT) could be enhanced by means of intracarotid (i.c.) infusion of Cereport, in combination with intravenous (i.v.) injection or i.c. infusion of boronophenylalanine (BPA) in the F98 rat glioma model. METHODS: For biodistribution studies, Fischer rats bearing intracerebral implants of the F98 glioma received i.v. or i.c. injections of 300 or 500 mg/kg body weight (b.w.) of BPA with or without i.c. infusion of 1.5 microg/kg b.w. of Cereport. For therapy studies, BNCT was initiated 14 days after intracerebral implantation of 10(3) F98 cells. The i.v. or i.c. injection of BPA (500 mg/kg b.w.) was given with or without Cereport, and the animals were irradiated 2.5 hours later at the Brookhaven Medical Research Reactor with a collimated beam of thermal neutrons delivered to the head. RESULTS: At a BPA dose of 500 mg/kg b.w., tumor boron concentrations (mean +/- standard deviation) were 55.7 +/- 9.6 microg/g with Cereport versus 33.6 +/- 3.9 microg/g without Cereport at 2.5 hours after i.c. infusion of BPA, and concentrations were 29.4 +/- 9.9 microg/g with Cereport versus 15.4 +/- 3.5 microg/g without Cereport (P < 0.05) after i.v. injection of BPA. After i.c. administration of BPA and Cereport, the tumor-to-blood ratio was 5.4 +/- 0.6, and the tumor-to-brain ratio was 5.2 +/- 2.4. After BNCT with BPA at a dose of 500 mg/kg, the survival time was 50 +/- 16 days for i.c. administration of BPA with Cereport versus 40 +/- 6 days without Cereport (P = 0.05), 38 +/- 4 days for i.v. administration of BPA with Cereport versus 34 +/- 3 days without Cereport (P = 0.02), 28 +/- 5 days for irradiated controls, and 23 +/- 3 days for untreated controls. Compared with untreated controls, there was a 117% increase in lifespan in rats that received an i.c. infusion of Cereport and then BPA, and an 86% increase in lifespan in rats that received i.c. administration of BPA without Cereport. CONCLUSION: These studies have established that i.c. administration of Cereport can not only increase tumor uptake of BPA, but also enhance the efficacy of BNCT.

Kynurenergic manipulations influence excitatory synaptic function and excitotoxic vulnerability in the rat hippocampus in vivo.

Competing enzymatic mechanisms degrade the tryptophan metabolite L-kynurenine to kynurenate, an inhibitory and neuroprotective compound, and to the neurotoxins 3-hydroxykynurenine and quinolinate. Kynurenine 3-hydroxylase inhibitors such as PNU 156561 shift metabolism towards enhanced kynurenate production, and this effect may underlie the recently discovered anticonvulsant and neuroprotective efficacy of these drugs. Using electrophysiological and neurotoxicological endpoints, we now used PNU 156561 as a tool to examine the functional interplay of kynurenate, 3-hydroxykynurenine and quinolinate in the rat hippocampus in vivo. First, population spike amplitude in area CA1 and the extent of quinolinate-induced excitotoxic neurodegeneration were studied in animals receiving acute or prolonged intravenous infusions of L-kynurenine, PNU 156561, (L-kynurenine+PNU 156561) or kynurenate. Only the latter two treatments, but not L-kynurenine or PNU 156561 alone, caused substantial inhibition of evoked responses in area CA1, and only prolonged (3h) infusion of (L-kynurenine+PNU 156561) or kynurenate was neuroprotective. Biochemical analyses in separate animals revealed that the levels of kynurenate attained in both blood and brain (hippocampus) were essentially identical in rats receiving extended infusions of L-kynurenine alone or (L-kynurenine+PNU 156561) (4 and 7microM, respectively, after an infusion of 90 or 180min). However, addition of the kynurenine 3-hydroxylase inhibitor resulted in a significant decrement in the formation of 3-hydroxykynurenine and quinolinate in both blood and brain. These data suggest that the ratio between kynurenate and 3-hydroxykynurenine and/or quinolinate in the brain is a critical determinant of neuronal excitability and viability. The anticonvulsant and neuroprotective potency of kynurenine 3-hydroxylase inhibitors may therefore be due to the drugs' dual action on both branches of the kynurenine pathway of tryptophan degradation.

Boron neutron capture therapy of brain tumors: enhanced survival and cure following blood-brain barrier disruption and intracarotid injection of sodium borocaptate and boronophenylalanine.

PURPOSE: Boronophenylalanine (BPA) and sodium borocaptate (Na(2)B(12)H(11)SH or BSH) have been used clinically for boron neutron capture therapy (BNCT) of high-grade gliomas. These drugs appear to concentrate in tumors by different mechanisms and may target different subpopulations of glioma cells. The purpose of the present study was to determine if the efficacy of BNCT could be further improved in F98-glioma-bearing rats by administering both boron compounds together and by improving their delivery by means of intracarotid (i.c.) injection with or without blood-brain barrier disruption (BBB-D). METHODS AND MATERIALS: For biodistribution studies, 10(5) F98 glioma cells were implanted stereotactically into the brains of syngeneic Fischer rats. Eleven to 13 days later animals were injected intravenously (i.v.) with BPA at doses of either 250 or 500 mg/kg body weight (b.w.) in combination with BSH at doses of either 30 or 60 mg/kg b.w. or i.c. with or without BBB-D, which was accomplished by i.c. infusion of a hyperosmotic (25%) solution of mannitol. For BNCT studies, 10(3) F98 glioma cells were implanted intracerebrally, and 14 days later animals were transported to the Brookhaven National Laboratory (BNL). They received BPA (250 mg/kg b.w.) in combination with BSH (30 mg/kg b.w. ) by i.v. or i.c. injection with or without BBB-D, and 2.5 hours later they were irradiated with a collimated beam of thermal neutrons at the BNL Medical Research Reactor. RESULTS: The mean tumor boron concentration +/- standard deviation (SD) at 2.5 hours after i. c. injection of BPA (250 mg/kg b.w.) and BSH (30 mg/kg b.w.) was 56. 3 +/- 37.8 microgram/g with BBB-D compared to 20.8 +/- 3.9 microgram/g without BBB-D and 11.2 +/- 1.8 microgram/g after i.v. injection. Doubling the dose of BPA and BSH produced a twofold increase in tumor boron concentrations, but also concomitant increases in normal brain and blood levels, which could have adverse effects. For this reason, the lower boron dose was selected for BNCT studies. The median survival time was 25 days for untreated control rats, 29 days for irradiated controls, 42 days for rats that received BPA and BSH i.v., 53 days following i.c. injection, and 72 days following i.c. injection + BBB-D with subsets of long-term survivors and/or cured animals in the latter two groups. No histopathologic evidence of residual tumor was seen in the brains of cured animals. CONCLUSIONS: The combination of BPA and BSH, administered i.c. with BBB-D, yielded a 25% cure rate for the heretofore incurable F98 rat glioma with minimal late radiation-induced brain damage. These results demonstrate that using a combination of boron agents and optimizing their delivery can dramatically improve the efficacy of BNCT in glioma-bearing rats.

Preoperative antibiotics and steroids in vestibular schwannoma excision.

OBJECTIVE: To examine the benefits of preoperative admission for intravenous steroids and antibiotics for patients undergoing vestibular schwannoma excision. STUDY DESIGN: Retrospective cohort study. METHODS: One hundred twenty patients with pathologically confirmed vestibular schwannoma followed for at least 1 year after surgery were included. Sixty patients were assigned to the preoperative admission group and 60 patients to the same-day-admission surgery group. The preoperative admission group was given intravenous dexamethasone (0.1 mg/kg) and intravenous cefazolin (1 g) beginning 12 hours before surgery. The same-day-surgery group received the same dosage of medication beginning at induction of anesthesia. OUTCOMES: Facial nerve function, meningitis, and wound infection rates, duration of hospital stay, and readmission rates were examined. RESULTS: There was no statistical difference in facial nerve function between the groups when controlling for tumor size. Likewise, there was no difference in meningitis or wound infection rates in the groups. As expected, hospital stay was significantly reduced but readmission rates were not affected. CONCLUSIONS: There are no apparent facial nerve function or infection control benefits to 1-day preoperative admission for intravenous steroids and antibiotics for patients undergoing vestibular schwannoma excision.

Actions of brain-derived neurotrophic factor in slices from rats with spontaneous seizures and mossy fiber sprouting in the dentate gyrus.

This study examined the acute actions of brain-derived neurotrophic factor (BDNF) in the rat dentate gyrus after seizures, because previous studies have shown that BDNF has acute effects on dentate granule cell synaptic transmission, and other studies have demonstrated that BDNF expression increases in granule cells after seizures. Pilocarpine-treated rats were studied because they not only have seizures and increased BDNF expression in granule cells, but they also have reorganization of granule cell "mossy fiber" axons. This reorganization, referred to as "sprouting," involves collaterals that grow into novel areas, i.e., the inner molecular layer, where granule cell and interneuron dendrites are located. Thus, this animal model allowed us to address the effects of BDNF in the dentate gyrus after seizures, as well as the actions of BDNF on mossy fiber transmission after reorganization. In slices with sprouting, BDNF bath application enhanced responses recorded in the inner molecular layer to mossy fiber stimulation. Spontaneous bursts of granule cells occurred, and these were apparently generated at the site of the sprouted axon plexus. These effects were not accompanied by major changes in perforant path-evoked responses or paired-pulse inhibition, occurred only after prolonged (30-60 min) exposure to BDNF, and were blocked by K252a. The results suggest a preferential action of BDNF at mossy fiber synapses, even after substantial changes in the dentate gyrus network. Moreover, the results suggest that activation of trkB receptors could contribute to the hyperexcitability observed in animals with sprouting. Because human granule cells also express increased BDNF mRNA after seizures, and sprouting can occur in temporal lobe epileptics, the results may have implications for understanding temporal lobe epilepsy.

Kindled seizures activate both branches of the autonomic nervous system.

Amygdaloid kindled seizures in the rat induce an abrupt elevation of blood pressure accompanied by a significant decrease in heart rate. The autonomic pharmacology of this response was examined in unanesthetized kindled rats. Muscarinic receptor blockade with atropine (1 mg/kg, intravenous (i.v.)) abolished the seizure-induced bradycardia. The seizure-induced hypertension was unaffected by beta-adrenergic blockade with timolol (1 mg/kg, i.v.), but was reduced by phentolamine (5 mg/kg, subcutaneous (s.c.)), an alpha-adrenergic receptor antagonist. A chemical sympathectomy was induced with 6-hydroxydopamine (100 mg/kg, i.v.), an agent that does not cross the blood-brain barrier. This eliminated the pressor response but did not completely block the seizure-induced bradycardia. The effectiveness of 6-hydroxydopamine was tested with tyramine (0.5 mg/kg, i.v.) an agent that releases endogenous catecholamines. These results indicate amygdaloid kindled seizures activate both branches of the autonomic nervous system. The bradycardia was mediated by the parasympathetic system; the pressor response was caused by an increase in peripheral resistance due to alpha-adrenergic receptor activation. More important, these findings show that kindling is a useful seizure model for future studies on the effect of seizures on cardiovascular function and possible mechanisms of seizure-related sudden unexplained death.

Boron neutron capture therapy of brain tumors: an emerging therapeutic modality.

Boron neutron capture therapy (BNCT) is based on the nuclear reaction that occurs when boron-10, a stable isotope, is irradiated with low-energy thermal neutrons to yield alpha particles and recoiling lithium-7 nuclei. For BNCT to be successful, a large number of 10B atoms must be localized on or preferably within neoplastic cells, and a sufficient number of thermal neutrons must be absorbed by the 10B atoms to sustain a lethal 10B (n, alpha) lithium-7 reaction. There is a growing interest in using BNCT in combination with surgery to treat patients with high-grade gliomas and possibly metastatic brain tumors. The present review covers the biological and radiobiological considerations on which BNCT is based, boron-containing low- and high-molecular weight delivery agents, neutron sources, clinical studies, and future areas of research. Two boron compounds currently are being used clinically, sodium borocaptate and boronophenylalanine, and a number of new delivery agents are under investigation, including boronated porphyrins, nucleosides, amino acids, polyamines, monoclonal and bispecific antibodies, liposomes, and epidermal growth factor. These are discussed, as is optimization of their delivery. Nuclear reactors currently are the only source of neutrons for BNCT, and the fission reaction within the core produces a mixture of lower energy thermal and epithermal neutrons, fast or high-energy neutrons, and gamma-rays. Although thermal neutron beams have been used clinically in Japan to treat patients with brain tumors and cutaneous melanomas, epithermal neutron beams now are being used in the United States and Europe because of their superior tissue-penetrating properties. Currently, there are clinical trials in progress in the United States, Europe, and Japan using a combination of debulking surgery and then BNCT to treat patients with glioblastomas. The American and European studies are Phase I trials using boronophenylalanine and sodium borocaptate, respectively, as capture agents, and the Japanese trial is a Phase II study. Boron compound and neutron dose escalation studies are planned, and these could lead to Phase II and possibly to randomized Phase III clinical trials that should provide data regarding therapeutic efficacy.

Enhanced delivery of boronophenylalanine for neutron capture therapy of brain tumors using the bradykinin analog Cereport (Receptor-Mediated Permeabilizer-7).

OBJECTIVE: Using the well-characterized F98 rat glioma model, the purpose of the present study was to determine whether the delivery of boronophenylalanine (BPA) could be enhanced by prior administration of the bradykinin analog Cereport (Alkermes, Inc., Cambridge, MA) (previously known as Receptor-Mediated Permeabilizer-7), which produces a transient, pharmacologically mediated opening of the blood-brain barrier. METHODS: Two series of experiments were performed in F98 glioma-bearing rats that had received either intracarotid (i.c.) or intravenous infusions of Cereport (at doses ranging from 1.5 to 7.5 microg/kg of body weight), followed by i.c. (or intravenous) injection of BPA (300 mg/kg of body weight). Animals were killed 0.5, 2.5, or 4 hours later, samples of blood, skin, muscle, and eye were obtained, brains were removed, and tumors were excised for boron determination by direct current plasma-atomic emission spectroscopy. RESULTS: Averaged over all time points, i.c. infusion of Cereport significantly enhanced tumor boron uptake (P = 0.0001), compared with the excipient (saline) control values. Tumor boron values were equivalent at 0.5 (36.0 microg/g) and 2.5 hours (38.5 microg/g) after i.c. administration of Cereport and BPA and then decreased by 33% (to 25.7 microg/g) at 4 hours. These tumor boron uptake values were significantly different (alpha = 0.05), compared with values measured at the corresponding times after i.c. administration of BPA without Cereport (22.6, 21.8, and 15.3 microg/g, respectively). Although no time-related effects were observed, i.c. administration of Cereport followed by intravenous administration of BPA also significantly enhanced (alpha = 0.05) tumor boron uptake at 0.5, 2.5, and 4 hours (27.4, 30.3, and 28.0 microg/g, respectively), compared with values obtained without Cereport (11.3, 13.4, and 15.2 microg/g, respectively). Boron levels in normal brain tissue from tumor-bearing and non-tumor-bearing cerebral hemispheres and in blood were not significantly different from those measured in saline-treated control animals. CONCLUSION: This study established that i.c. infusion of Cereport significantly increased delivery of BPA to F98 rat gliomas, and this could enhance the efficacy of boron neutron capture therapy of this tumor.

Chronic changes in synaptic responses of entorhinal and hippocampal neurons after amino-oxyacetic acid (AOAA)-induced entorhinal cortical neuron loss.

Chronic changes in synaptic responses of entorhinal and hippocampal neurons after amino-oxyacetic acid (AOAA)-induced entorhinal neuron loss. J. Neurophysiol. 80: 3031-3046, 1998. Synaptic responses of entorhinal cortical and hippocampal neurons were examined in vivo and in vitro, 1 mo to 1.5 yr after a unilateral entorhinal lesion caused by a focal injection of amino-oxyacetic acid (AOAA). It has been shown previously that injection of AOAA into the medial entorhinal cortex produces cell loss in layer III preferentially. Although behavioral seizures stopped approximately 2 h after AOAA treatment, abnormal evoked responses were recorded as long as 1.5 yr later in the entorhinal cortex and hippocampus. In the majority of slices from AOAA-treated rats, responses recorded in the superficial layers of the medial entorhinal cortex to white matter, presubiculum, or parasubiculum stimulation were abnormal. Extracellularly recorded responses to white matter stimulation were prolonged and repetitive in the superficial layers. Intracellular recordings showed that residual principal cells in superficial layers produced prolonged, repetitive excitatory postsynaptic potentials (EPSPs) and discharges in response to white matter stimulation compared with brief EPSPs and a single discharge in controls. Responses of deep layer neurons of AOAA-treated rats did not differ from controls in their initial synaptic response. However, in a some of these neurons, additional periods of excitatory activity occurred after a delay. Abnormal responses were recorded from slices ipsilateral as well as contralateral to the lesioned hemisphere. Recordings from the entorhinal cortex in vivo were abnormal also, as demonstrated by prolonged and repetitive responses to stimulation of the area CA1/subiculum border. Evoked responses of hippocampal neurons, recorded in vitro or in vivo, demonstrated abnormalities in selected pathways, such as responses of CA3 neurons to hilar stimulation in vitro. There was a deficit in the duration of potentiation of CA1 population spikes in response to repetitive CA3 stimulation in AOAA-treated rats. Theta activity was reduced in amplitude in area CA1 and the dentate gyrus of AOAA-treated rats, although evoked responses to angular bundle stimulation could not be distinguished from controls. The results demonstrate that a preferential lesion of layer III of the entorhinal cortex produces a long-lasting change in evoked and spontaneous activity in parts of the entorhinal cortex and hippocampus. Given the similarity of the lesion produced by AOAA and entorhinal lesions in temporal lobe epileptics, these data support the hypothesis that preferential damage to the entorhinal cortex contributes to long-lasting changes in excitability, which could be relevant to the etiology of temporal lobe epilepsy.