Age determination of subdural hematomas: survey among radiologists.

Abusive head trauma is a severe form of child abuse. One important diagnostic finding is the presence of a subdural hematoma. Age determination of subdural hematomas is important to relate radiological findings to the clinical history presented by the caregivers. In court this topic is relevant as dating subdural hematomas can lead to identification of a suspect. The aim of our study is to describe the current practice among radiologists in the Netherlands regarding the age determination of subdural hematomas in children. This is a cross-sectional study, describing the results of an online questionnaire regarding dating subdural hematomas among pediatric and neuro-radiologists in the Netherlands. The questionnaire consisted of sociodemographic questions, theoretical questions and eight pediatric cases in which the participants were asked to date subdural hematomas based on imaging findings. Fifty-one out of 172 radiologists (30 %) filled out the questionnaire. The percentage of participants that reported it was possible to date the subdural hematoma varied between 58 and 90 % for the eight different cases. In four of eight cases (50 %), the age of the subdural hematoma as known from clinical history fell within the range reported by the participants. None of the participants was "very certain" of their age determination. The results demonstrate that there is a considerable practice variation among Dutch radiologists regarding the age determination of subdural hematomas. This implicates that dating of subdural hematomas is not suitable to use in court, as no uniformity among experts exists.

Chronic but not acute foot-shock stress leads to temporary suppression of cell proliferation in rat hippocampus.

Stressful experiences, especially when prolonged and severe are associated with psychopathology and impaired neuronal plasticity. Among other effects on the brain, stress has been shown to negatively regulate hippocampal neurogenesis, and this effect is considered to be exerted via glucocorticoids. Here, we sought to determine the temporal dynamics of changes in hippocampal neurogenesis after acute and chronic exposure to foot-shock stress. Rats subjected to a foot-shock procedure showed strong activation of the hypothalamic-pituitary-adrenal (HPA) axis, even after exposure to daily stress for 3 weeks. Despite a robust release of corticosterone, acute foot-shock stress did not affect the rate of hippocampal cell proliferation. In contrast, exposure to foot-shock stress daily for 3 weeks led to reduced cell proliferation 2 hours after the stress procedure. Interestingly, this stress-induced effect did not persist and was no longer detected 24 hours later. Also, while chronic foot-shock stress had no impact on survival of hippocampal cells that were born before the stress procedure, it led to a decreased number of doublecortin-positive granule neurons that were born during the chronic stress period. Thus, whereas a strong activation of the HPA axis during acute foot-shock stress is not sufficient to reduce hippocampal cell proliferation, repeated exposure to stressful stimuli for prolonged period of time ultimately results in dysregulated neurogenesis. In sum, this study supports the notion that chronic stress may lead to cumulative changes in the brain that are not seen after acute stress. Such changes may indicate compromised brain plasticity and increased vulnerability to neuropathology.

Fear conditioning and shock intensity: the choice between minimizing the stress induced and reducing the number of animals used.

Many fear conditioning studies use electric shock as the aversive stimulus. The intensity of shocks varies throughout the literature. In this study, shock intensities ranging from 0 to 1.5 mA were used, and the effects on the rats assessed by both behavioural and biochemical stress parameters. Results indicated a significant difference with respect to defaecation and freezing behaviour between controls and those animals that received a shock. Significant differences in corticosterone levels were also noted between controls and those groups that received a shock. No significant differences were found between the shock groups with regards to the stress parameters measured in our fear conditioning paradigm, indicating that the two shock groups were similarly stressed. Increased significance levels were noted for freezing behaviour as well as a lower standard error of means was found in the highest shock intensity group. We therefore recommend using the higher shock intensity (1.5 mA) as the rats in the higher shock intensity group were more homogeneously fear-conditioned and therefore the results should be more reproducible and robust than in the lower shock intensity group. This would allow for fewer rats to be used in order to gain an accurate impression of the conditioning paradigm employed.

Monitoring arterio-venous differences of glucose and lactate in the anesthetized rat with or without brain damage with ultrafiltration and biosensor technology.

Continuous monitoring of arterio-venous glucose and lactate differences may serve as a diagnostic tool to assess normal brain function and brain pathology. We describe a method and some results obtained with arterio-venous measurements of glucose and lactate in the blood of the halothane-anesthetized rat and after brain injury. The method is based on low flow rate ultrafiltration for continuous collection of blood filtrate combined with flow injection analysis and biosensors for the detection of glucose and lactate. We measured the glucose and lactate concentration every minute in the jugular vein and the aorta at control conditions and during and after inflation of an embolectomy-balloon for 2 min. Net cerebral lactate efflux and glucose uptake was seen under control conditions and at low blood lactate levels. During brain injury both lactate release and glucose uptake were reduced and there was a net lactate influx at high arterial lactate levels. These results indicate that the flux of lactate in and out of the brain is not only dependent on the lactate concentration in the brain, but on blood levels as well, possibly because of bi-directional flux through the monocarboxylate transporter type 1.

Effects of brain death and hemodynamic status on function and immunologic activation of the potential donor liver in the rat.

OBJECTIVE: To assess the effect on the function and immunologic status of potential donor livers of the duration of brain death combined with the presence and absence of hemodynamic instability in the donor. SUMMARY BACKGROUND DATA: Brain death, regarded as a given condition in organ transplantation, could have significant effects on the donor organ quality. METHODS: Brain death was induced in Wistar rats. Short or long periods of brain death in the presence or absence of hemodynamic instability were applied. Sham-operated rats served as controls. Organ function was studied by monitoring standard serum parameters. The inflammatory status of the liver was assessed by determining the immediate early gene products, the expression of cell adhesion molecules, and the influx of leukocytes in the liver. RESULTS: Progressive organ dysfunction was most pronounced in hemodynamically unstable brain-dead donors. Irrespective of hemodynamic status, a progressive inflammatory activation could be observed in brain-dead rats compared with controls. CONCLUSIONS: Brain death causes progressive liver dysfunction, which is made worse by the coexistence of hemodynamic instability. Further, brain death activates the inflammatory status of the potential donor liver, irrespective of the presence of hypotension. The changes observed may predispose the graft to additional damage from ischemia and reperfusion in the transplant procedure.

Anatomical demonstration of the suprachiasmatic nucleus-pineal pathway.

A polysynaptic pathway is proposed to transmit light information from the retina through the suprachiasmatic nucleus of the hypothalamus (SCN) to the pineal. In the present study, the powerful transneuronal tracer, pseudorabies virus (PRV), was used to provide a detailed description of this pathway. PRV injected into the pineal subsequently labeled the superior cervical ganglion, the intermediolateral column of the upper thoracic cord, the autonomic division of the paraventricular nucleus of the hypothalamus (PVN), and the SCN. Neurons in the autonomic division of the PVN were the only PRV-labeled neurons in the hypothalamus shown to receive input from the SCN as demonstrated by the presence of vasoactive intestinal polypeptide axonal contacts. This observation concurred with the presence of ventrally placed neurons in the SCN that could only be observed a day after the appearance of PVN-labeled neurons. Nevertheless the majority of the neurons were found in the dorsomedial position of the SCN, associated with the vasopressin-containing population of SCN neurons. Confocal laser scanning microscopy showed double-labeled neurons containing PRV and vasopressin or PRV and vasoactive intestinal polypeptide. Specificity of tracing was also established by prior removal of the superior cervical ganglion, resulting in a complete absence of the tracer but in the pineal. Thus, the present study provides the anatomical basis for circadian control of melatonin secretion.

Induction of organ dysfunction and up-regulation of inflammatory markers in the liver and kidneys of hypotensive brain dead rats: a model to study marginal organ donors.

BACKGROUND: Marginal donors exposed to the full array of effects induced by brain death are characterized by low success rates after transplantation. This study examined whether organs from marginal brain dead animals show any change in organ function or tissue activation making them eventually more susceptible for additional damage during preservation and transplantation. METHODS: To study this hypothesis we first focused on effects of brain death on donor organ quality by using a brain death model in the rat. After induction of brain death, Wistar rats were ventilated for 1 and 6 hr and then killed. Sham-operated rats served as controls. Organ function was studied using standard serum parameters. Tissue activation of liver and kidney was assessed by staining of immediate early gene products (IEG: FOS, JUN), and inflammatory markers; cell adhesion molecules (Intercellular adhesion molecule-1, vascular cell adhesion molecule-1), leukocyte infiltrates (CD45, T cell receptor, CD8, CD4), and MHC class II. RESULTS: During brain death progressive organ dysfunction was observed that coincided with a significant increase in activation of immediate early genes, intercellular adhesion molecule-1, vascular cell adhesion molecule-1, CD45, and MHC class II versus nonbrain dead controls. In liver tissue also the markers for T cell receptor and CD8 significantly increased. CONCLUSIONS: These findings suggest that an immune activation with increased endothelial cell activation and immediate early gene expression occurs in marginal donors after brain death induction. We suggest that brain death should not longer be regarded as a given nondeleterious condition but as a dynamic process with potential detrimental effects on donor organs that could predispose grafts for increased alloreactivity after transplantation.

Impedance recordings to determine change in extracellular volume in the brain following cardiac arrest in broiler chickens.

The present study describes a method to determine the onset and development of brain damage in broiler chickens. Exsanguination disrupts the brain metabolism and causes the brain to become ischemic. Energy-requiring systems in the cell membrane fail, which results in an ionic shift over the membrane, accompanied by a water influx into the cell. This cellular edema decreases the extracellular volume of brain tissue. In mammals, this brain damage has been measured by recording brain impedance. We adapted this approach for use with poultry. Five to six-week-old commercial broilers were equipped with impedance recording electrodes in the striatum area of the brain. Cardiac arrest was induced by means of an intravenous injection of MgCl2 and brain impedance was recorded for 30 min. The resulting curves showed a high similarity to those obtained in rats. No effects of 12 h antemortem feed deprivation on the size and rate of change in brain impedance could be found. Both in anesthetized and conscious birds, a change in brain impedance was found. We conclude that brain impedance can be used to determine the development of ischemic brain damage in broiler chickens.

Forebrain parasympathetic control of heart activity: retrograde transneuronal viral labeling in rats.

Dysfunction of parasympathetic command neurons may be a cause of cardiac autonomic imbalance, which has been implicated as a pathogenic mechanism of lethal arrhythmias. The locations in the brain of these command neurons are not known. The aim of this investigation is to identify selectively the parasympathetic command neurons in the forebrain. Male Wistar rats were inoculated in the left ventricular myocardium with 2 ml of a 3 x 10(6) plaque-forming units/ml of a pseudorabies virus (PRV)-Bartha solution. Eighteen hours after the infection, the spinal cord was transected at T1. Six of fourteen rats showed PRV-immunoreactive cells in the forebrain after 6 postoperative survival days. Bilaterally, the infections were located in the prelimbic, anterior cingulate, frontal, and insular cortexes, various hypothalamic and midbrain nuclei, the nucleus of the solitary tract, the dorsal motor vagus, and periambiguus nuclei. Control animals receiving intravenous PRV-Bartha injections were not infected. Using transneuronal retrograde viral labeling and spinal cord transection, we were able to localize the forebrain parasympathetic command neurons that maintain cardiac autonomic balance. The virus-infected cells were localized in regions that previously showed susceptibility for immune activation-mediated selective cerebral endothelial leakage. We hypothesize that such selective endothelial leakage could induce autonomic imbalance after myocardial infarction.

Evidence that stress activates glial lactate formation in vivo assessed with rat hippocampus lactography.

Extracellular lactate of the rat hippocampus is inter alia increased by immobilization stress. The origin of lactate is, however, not well established, so it is not known whether it is mainly derived form neurons or glial cells. Dialysates were collected shortly (1 or 2 days) or with a delay (14 or 15 days) after implantation of the probe. In the short-term experiment lactate increased after stress, both with or without glucose added to the perfusate. In the long-term experiment there was marked gliosis around the dialysis probe and the stress effects were seen only in the presence of 5 mM glucose. The results are consistent with the idea that stress induces glycogenolysis and lactate export from astroglial cells via neurotransmitter or hormonal related processes.

Possible glial contribution of rat hippocampus lactate as assessed with microdialysis and stress.

Microdialysis for the continuous monitoring of lactate ("lactography") was applied in rat brain hippocampus in an attempt to establish whether lactate is of neuronal or glial origin. Lactate was analyzed with an electrochemical assay after enzymatic oxidation in dialysates derived from a short-term (1 or 2 days after implantation of the probe) and a long-term (14 or 15 days) preparation. In the short-term experiment the lactate levels in the dialysate were higher without glucose in the perfusate, whereas in the long-term experiment a several fold increase in lactate was observed in the presence of 5 mM glucose. During stress, increases in lactate were virtually similar both in the acute (with or without glucose in the perfusate) and in the chronic preparation (response in the presence of glucose only). In the long-term preparation presence of reactive astroglia cells was visualized immunohistochemically with antibodies against glial fibrillary acidic protein. Damage of the hippocampus and the corpus callosum was seen in the chronic preparation with silver impregnation staining. These results emphasize the importance of the presence of glucose in the perfusate and they are consistent with the idea that glial cells contribute to extracellular lactate in the rat hippocampus in vivo and that stress activates the astroglial glycogen pool.

Region-specific alterations of calbindin-D28k immunoreactivity in the rat hippocampus following adrenalectomy and corticosterone treatment.

The aim of this study was (i) to compare the immunocytochemical distribution of the calcium-binding protein calbindin-D28k (CB) in the hippocampus of rats with the pattern of neurodegeneration following adrenalectomy (ADX) using silver impregnation, and (ii) to investigate the CB-immunoreactivity in the hippocampus following 3 weeks corticosterone treatment. 24 h following ADX no degenerative changes, nor alterations in CB-immunoreactivity were found in the hippocampus. Both 3 and 21 days following ADX neurodegeneration in the dentate gyrus could be observed which was accompanied with a loss of CB-immunoreactive (CB-ir) cells in that parts of the dentate gyrus suffering neuronal degeneration. Additionally we observed a marked loss of CB-ir in the CA1 area both 3 and 21 days following ADX. Three weeks daily corticosterone treatment (10 mg/day) induced a marked increase of CB-ir exclusively in the CA1 pyramidal cell layer. We conclude that (i) there is a close relationship between the loss of CB-immunoreactive cells in the DG and the neuronal degeneration in the dentate gyrus following ADX, and (ii) corticosterone appears to be involved in the regulation of calbindin-D28k in the CA1 pyramidal cell layer.

Macular grid laser photocoagulation in uveitis.

AIMS/BACKGROUND: The aim of this study was to evaluate whether grid laser photocoagulation of the macula is beneficial in the treatment of cystoid macular oedema in patients with uveitis. METHODS: Six eyes of five patients with long standing cystoid macular oedema due to chronic uveitis were treated by grid laser photocoagulation of the macula. RESULTS: In the first weeks after treatment a temporary increase of oedema and paracentral scotomas were observed. At the long term follow up of more than 18 months in all patients, macular oedema had been reduced significantly or disappeared in all eyes treated. One eye had a significant increase in Snellen acuity, three eyes more or less stabilised, and two eyes deteriorated. CONCLUSION: The beneficial effect of laser treatment on visual acuity in patients with uveitis might be more favourable if performed at an earlier stage of the disease.

Induction of glial fibrillary acidic protein immunoreactivity in the rat dentate gyrus after adrenalectomy: comparison with neurodegenerative changes using silver impregnation.

In the present study we performed a light microscopic anatomical comparison of adrenalectomy (ADX)-induced neurodegeneration using silver impregnation and reaction of astroglial cells using GFAP immunocytochemistry in the hippocampus of the rat. Three survival times following ADX were studied: 24 hours, 3 days, and 3 weeks. Twenty-four hours following ADX we found no degenerative changes or altered GFAP immunostaining. Three days after adrenalectomy, argyrophilic somata appeared in the granular cell layer of the dentate gyrus. Argyrophilic dendrites were seen in the molecular layer of the dentate gyrus and neuritic argyrophilia were seen in the mossy fiber layer. Induction of GFAP immunoreactivity occurred simultaneously with degeneration. Increased GFAP immunoreactivity could be observed 3 days after adrenalectomy in the molecular layer of the dentate gyrus, granular cell layer, sub and supragranular cell layer, and mossy fiber layer. Size and shape of astroglial cells were changed, and their processes in the molecular layer changed from unidirectional to randomly organized. Degeneration and astroglial reaction were more pronounced 3 weeks after adrenalectomy and both were prevented by adding corticosterone to the drinking solution. Animals that did not show degenerative changes showed no increased GFAP immunoreactivity, while both effects were confined to the dentate gyrus and mossy fiber zone. These results show that there is a close relationship between the induction of GFAP immunoreactivity in the hippocampus of the rat and neuronal degeneration in the dentate gyrus following ADX, both in time and space.

Time course and distribution of neuronal degeneration in the dentate gyrus of rat after adrenalectomy: a silver impregnation study.

Recently, Sloviter et al. reported that adrenalectomy (ADX) of young adult rats after 3 months led to a selective loss of granule neurons in the dentate gyrus (DG) and that this loss could be prevented by low doses of corticosterone. In the present study, the ADX-induced neuronal degeneration was investigated in Wistar rats, using a silver impregnation method for degenerating neurons. To examine the time course and distribution of the ADX-induced degeneration, young adult male rats were allowed to survive 2, 3, and 5 days and 1, 2, and 3 weeks after ADX. Argyrophilic neurons were present in the dentate granule cell layer on the second day following ADX. Three days after ADX, the number of argyrophilic granule neurons was much more abundant, and it increased gradually with longer post-ADX survival times. Argyrophilia was specifically confined to dentate granule cells and was accompanied by the occurrence of pyknotic nuclei as observed in adjacent cresyl violet-stained sections. There were significant differences between individual rats in quantity of argyrophilia. About one fifth of the ADX rats showed sporadic or no argyrophilia, in spite of plasma corticosterone levels below the detection limit (10 ng/mL). Sham-operated rats and ADX rats receiving corticosterone (10 mg/L) or dexamethasone (15 mg/L) in their drinking water did not display any argyrophilic neurons in the dentate gyrus. The distribution of the argyrophilia within the DG was highly characteristic with the highest number of degenerating cells in the hidden blade of the middle and the temporal thirds of the DG.(ABSTRACT TRUNCATED AT 250 WORDS)

Analysis of IL-6 levels in human vitreous fluid obtained from uveitis patients, patients with proliferative intraocular disorders and eye bank eyes.

Several studies suggest a role for IL-6 in the pathogenesis of uveitis. Earlier we have shown that aqueous humour obtained from patients with uveitis contained raised levels of IL-6. In the study described here we investigated the IL-6 levels in vitreous fluid samples obtained from 75 uveitis patients with different uveitis entities. Vitreous samples from 14 patients with proliferative intraocular disorders (PID) and 29 eye bank eyes were used as controls. All the samples were tested in the IL-6 B9 bioassay as well as in a sensitive ELISA for IL-6. Raised IL-6 levels were detected in the vitreous fluid of uveitis patients as well as patients with PID, implicating IL-6 as a common inflammatory mediator. The highest mean level of IL-6 was found in the vitreous fluid of patients with acute retinal necrosis. The mean IL-6 levels measured by the ELISA were higher compared to the levels measured by the B9 bioassay. This may be caused by the presence of B9 bioassay inhibitory factors in the vitreous fluid of these patients.

Continuous monitoring of extracellular lactate concentration by microdialysis lactography for the study of rat muscle metabolism in vivo.

A method is described for the measurement and on-line monitoring of muscular extracellular lactate concentration in both anaesthetized and freely moving rats. This method is based on microdialysis sampling and lactic dehydrogenase-catalysed nicotinamide adenine dinucleotide, reduced (NADH)-fluorescence detection techniques. In vivo calibration revealed a resting extracellular lactate concentration of 1.92 +/- 0.13 mmol/l (+/- SEM) in the gastrocnemius muscle of adult male Wistar rats (n = 6), while the average whole-blood lactate level was 0.76 +/- 0.12 mmol/l (+/- SEM). This measured extracellular lactate concentration was 1.73-times higher than that deduced from the arterial lactate concentration. Blocking glycolysis with iodoacetate reduced the extracellular lactate concentration to 52 +/- 6% (+/- SEM, n = 4) of the resting level. The extracellular lactate concentration in rat gastrocnemius muscle had increased to significantly (P less than or equal to 0.05) different levels, 2.4 +/- 0.03 (+/- SEM) or 4.0 +/- 0.55 (+/- SEM) times the control value, 1 h after aortic clamping (n = 3) or cardiac arrest (n = 3), respectively. Stimulation of the sciatic nerve induced elevations of the extracellular lactate concentration in the tibialis anterior muscle which were linearly related to the recorded isometric force-time integral. We also monitored on-line the changes in extracellular lactate concentration in the tibialis anterior muscle of a swimming rat. Our results indicate that microdialysis lactate reflects also intracellular metabolism. Lactography may be a useful alternative to biopsies and nuclear magnetic resonance spectroscopy in clinical medicine and physiology for the monitoring of metabolism in vivo.

Rat striatal cation shifts reflecting hypoxic-ischemic damage can be predicted by on-line impedance measurements.

We investigated the earliest time at which irreversible damage takes place after hypoxia-ischemia in the Levine preparation of rats. In 60 rats anesthetized with chloral hydrate and maintained at one of three body temperatures, we unilaterally ligated the left common carotid artery and placed electrodes in the striatum to measure impedance (reflecting the extracellular space) during hypoxia, recovery, and/or cardiac arrest. We measured blood gases and pH at regular intervals during hypoxia in 47 rats and assessed blood-brain barrier function with Evans blue and tissue damage using Na+:K+ ratios. Shortly after hypoxia, impedance normalized in 24 rats without brain damage (normal Na+:K+ ratios, 4 hours of recovery). Sustained elevation of striatal impedance during recovery in six rats was related to an elevated Na+:K+ ratio and a disrupted blood-brain barrier. Damage was not obviously related to blood gases, pH, or the net reduction of the extracellular space during hypoxia. Hypothermia in 17 rats prevented impedance changes, and no striatal damage was found. Thus, irreversible brain damage very likely occurs during or very shortly after hypoxia. Persistent reduction of the extracellular space indicates tissue damage and can be used to monitor potential in vivo therapeutic measures.