Study protocol for OptimalTTF-2: enhancing Tumor Treating Fields with skull remodeling surgery for first recurrence glioblastoma: a phase 2, multi-center, randomized, prospective, interventional trial.

BACKGROUND: OptimalTTF-2 is a randomized, comparative, multi-center, investigator-initiated, interventional study aiming to test skull remodeling surgery in combination with Tumor Treating Fields therapy (TTFields) and best physicians choice medical oncological therapy for first recurrence in glioblastoma patients. OptimalTTF-2 is a phase 2 trial initiated in November 2020. Skull remodeling surgery consists of five burrholes, each 15 mm in diameter, directly over the tumor resection cavity. Preclinical research indicates that this procedure enhances the effect of Tumor Treating Fields considerably. We recently concluded a phase 1 safety/feasibility trial that indicated improved overall survival and no additional toxicity. This phase 2 trial aims to validate the efficacy of the proposed intervention. METHODS: The trial is designed as a comparative, 1:1 randomized, minimax two-stage phase 2 with an expected 70 patients to a maximum sample size of 84 patients. After 12-months follow-up of the first 52 patients, an interim futility analysis will be performed. The two trial arms will consist of either a) TTFields therapy combined with best physicians choice oncological treatment (control arm) or b) skull remodeling surgery, TTFields therapy and best practice oncology (interventional arm). Major eligibility criteria include age ≥ 18 years, 1st recurrence of supratentorial glioblastoma, Karnofsky performance score ≥ 70, focal tumor, and lack of significant co-morbidity. Study design aims to detect a 20% increase in overall survival after 12 months (OS12), assuming OS12 = 40% in the control group and OS12 = 60% in the intervention group. Secondary endpoints include hazard rate ratio of overall survival and progression-free survival, objective tumor response rate, quality of life, KPS, steroid dose, and toxicity. Toxicity, objective tumor response rate, and QoL will be assessed every 3rd month. Endpoint data will be collected at the end of the trial, including the occurrence of suspected unexpected serious adverse reactions (SUSARs), unacceptable serious adverse events (SAEs), withdrawal of consent, or loss-to-follow-up. DISCUSSION: New treatment modalities are highly needed for first recurrence glioblastoma. Our proposed treatment modality of skull remodeling surgery, Tumor Treating Fields, and best practice medical oncological therapy may increase overall survival significantly. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT0422399 , registered 13. January 2020.

Cryoneurolysis' outcome on pain experience (COPE) in patients with low-back pain: study protocol for a single-blinded randomized controlled trial.

BACKGROUND: Low-back pain, including facet joint pain, accounts for up to 20 % of all sick leaves in DenmarkA proposed treatment option is cryoneurolysis. This study aims to investigate the effect of cryoneurolysis in lumbar facet joint pain syndrome. METHODS: A single-center randomized controlled trial (RCT) is performed including 120 participants with chronic facet joint pain syndrome, referred to the Department of Neurosurgery, Aarhus University Hospital. Eligible patients receive a diagnostic anesthetic block, where a reduction of pain intensity ≥ 50 % on a numerical rating scale (NRS) is required to be enrolled. Participants are randomized into three groups to undergo either one treatment of cryoneurolysis, radiofrequency ablation or placebo. Fluoroscopy and sensory stimulation is used to identify the intended target nerve prior to administrating the above-mentioned treatments. All groups receive physiotherapy for 6 weeks, starting 4 weeks after treatment. The primary outcome is the patients' impression of change in pain after intervention (Patient Global Impression of Change (PGIC)) at 4 weeks follow-up, prior to physiotherapy. Secondary outcomes are a reduction in low-back pain intensity (numeric rating scale) and quality of life (EQ-5D, SF-36) and level of function (Oswestry Disability Index), psychological perception of pain (Pain Catastrophizing Scale) and depression status (Major Depression Inventory). Data will be assessed at baseline (T0), randomization (T1), day one (T2), 4 weeks (T3), 3 (T4), 6 (T5) and 12 months (T6). DISCUSSION: This study will provide information on the effectiveness of cryoneurolysis vs. the effectiveness of radiofrequency ablation or placebo for patients with facet joint pain, and help to establish whether cryoneurolysis should be implemented in clinical practice for this patient population. TRIAL REGISTRATION: The trial is approved by the ethical committee of Central Jutland Denmark with registration number 1-10-72-27-19 and the Danish Data Protection Agency with registration number 666,852. The study is registered at Clinicaltrial.gov with the ID number NCT04786145 .

Nasal application of HSV encoding human preproenkephalin blocks craniofacial pain in a rat model of traumatic brain injury.

According to Centers for Disease Control and Prevention, each year, an estimated 1.7 million Americans sustain a traumatic brain injury (TBI), which frequently leads to chronic craniofacial pain. In this study we examine a gene therapy approach to the treatment of post-TBI craniofacial neuropathic pain using nasal application of a herpes simplex virus (HSV)-based vector expressing human proenkephalin (SHPE) to target the trigeminal ganglia. Mild TBI was induced in rats by the use of a modified fluid percussion model. Two days after mild TBI, following the development of facial mechanical allodynia, animals received either an intranasal application of vehicle or recombinant HSV encoding human preproenkephalin or lacZ reporter gene encoding control vector (SHZ.1). Compared with baseline response thresholds, mild TBI in SHZ.1 or vehicle-treated animals induced a robust craniofacial allodynia lasting at least 45 days. On the other hand, nasal SHPE application 2 days post-TBI attenuated facial allodynia, reaching significance by day 4-7 and maintaining this effect throughout the duration of the experiment. Immunohistochemical examination revealed strong expression of human proenkephalin in trigeminal ganglia of SHPE, but not SHZ.1-treated rats. This study demonstrates that intranasal administration of HSV-based gene vectors may be a viable, non-invasive means of treating chronic craniofacial pain, including post-TBI pain.

Hypothalamic deep brain stimulation influences autonomic and limbic circuitry involved in the regulation of aggression and cardiocerebrovascular control in the Göttingen minipig.

BACKGROUND: Deep brain stimulation (DBS) in the ventral tuberal hypothalamus (VTH) is currently under investigation for the treatment of severe obesity. Stimulation impact on a number of closely related hypothalamic neural systems could potentially influence normal hypothalamic function and thereby generate adverse side effects. OBJECTIVE: To assess the feasibility and safety of VTH DBS in a non-primate large animal model. METHODS: In the VTH of 6 Göttingen minipigs, quadropolar leads were implanted bilaterally (n = 2) or unilaterally (n = 4), using optimized MRI sequences allowing identification of major diencephalic landmarks. Heart rate, weight, behavior and nighttime locomotor activity were recorded throughout the study period. Two of the unilaterally implanted minipigs were examined with [15O]H2O positron emission tomography (PET) scans performed in DBS-off and DBS-on mode. RESULTS: VTH DBS elicited an amplitude-dependent increase in heart rate and transient aggressive behavior. PET demonstrated that VTH DBS caused a global increase in cerebral blood flow velocities and decreased mean transit time. CONCLUSIONS: VTH DBS results in behavioral and physiological changes, which may derive from activation of closely related limbic and autonomic networks. Caution and further studies of longer length should be requested before this procedure is used more widely in humans.

Brain death induced by cerebral haemorrhage - a new porcine model evaluated by CT angiography.

BACKGROUND: Brain death and complications to brain death affects the function of organs in the potential donor. Previous animal models of brain death have not been able to fully elucidate the mechanisms behind this organ dysfunction, and none of the available animal models mimic the most common insult prior to brain death: intracerebral haemorrhage. The objective of this study was to develop a large animal model of brain death based on a controlled intracerebral haemorrhage and verified by computerised tomographic angiography (CTA). METHODS: Twenty pigs (range: 26.6-31.2 kg) were randomised to brain death or control. Brain death was induced by infusion of blood through a stereotaxically placed needle in the internal capsule. Brain death was confirmed by the measured intracranial pressure (ICP), lack of corneal and pupillary light reflexes, and atropine test. CTA was performed 120-180 min after brain death. The pigs were observed for 8 h after brain death. RESULTS: Brain death was declared when the ICP exceeded mean arterial pressure after a median of 36 min (range: 28-51 min). Significant increases in heart rate, and mean arterial pressure (MAP) were followed by a steep decrease. With fluid therapy, the animals demonstrated haemodynamic stability. Reflexes disappeared, and atropine did not induce an increase in heart rate in the brain dead animals. CTA confirmed loss of cerebral circulation. CONCLUSION: This study offers a standardised, clinically relevant porcine model of brain death induced by a haemorrhagic attack. Brain death was verified by the disappearance of corneal and pupil reflex, atropine test, and CTA.

Development of neuromodulation treatments in a large animal model--do neurosurgeons dream of electric pigs?

The Göttingen minipig has been established as a translational research animal for neurological and neurosurgical disorders. This animal has a large gyrencephalic brain suited for examination at sufficient resolution with conventional clinical scanning modalities. The large brain, further, allows use of standard neurosurgical techniques and can accommodate clinical neuromodulatory devises such as deep brain stimulation (DBS) electrodes and encapsulated cell biodelivery devices making the animal ideal for basic scientific studies on neuromodulation mechanisms and preclinical tests of new neuromodulation technology for human use. The use of the Göttingen minipig is economical and does not have the concerns of the public associated with the experimental use of primates, cats, and dogs, thus providing a cost-effective research model for translation of rodent data before clinical trials are initiated.

Neuromodulation in a minipig MPTP model of Parkinson disease.

Large animal neuroscience enables the use of conventional clinical brain imagers and the direct use and testing of surgical procedures and equipment from the human clinic. The greater complexity of the large animal brain additionally enables a more direct translation to human brain function in health and disease. Economical, ethical, scientific and practical issues may on the other hand hamper large animal neuroscience. Large animal neuroscience should therefore either be performed in order to examine large animal species dependent problems or to complement promising small animal basic studies by constituting an intermediate research system, bridging small animal CNS research to the human CNS. We have, accordingly, during the last ten years used the Gottingen minipig to examine neuromodulatory treatment modalities such as stem cell transplantation and deep brain stimulation directed towards Parkinson disease. This has been accomplished by the development of a MPTP-based large animal model of Parkinson disease in the Gottingen minipig and the development of stereotaxic and surgical approaches needed to manipulate the Gottingen minipig CNS. The instituted changes in the CNS can be evaluated in the live animal by brain imaging (PET and MR), cystometry, gait analysis, neurological evaluation and by post mortem examination based on histology and stereological analysis.

Chronic subthalamic high-frequency deep brain stimulation in Parkinson's disease--a histopathological study.

This study describes the pathological findings in the brain of a patient with Parkinson's disease (PD) treated with bilateral subthalamic high-frequency deep brain stimulation (STN DBS) for 29 months prior to death. After routine neuropathological examination, tissue blocks containing the electrode tracts, the subthalamic nucleus (STN), the substantia nigra and the pre-frontal cortex were paraffin embedded and cut into 5-microm-thick serial sections and stained with several conventional staining methods and immunohistochemistry. Bilateral nigral depigmentation, cell loss and Lewy body formation confirmed the diagnosis of PD. Microscopic evaluation furthermore confirmed the location of the electrodes in the STN. The electrode tracts were surrounded by a 150-microm-wide glial fibrillary acidic protein (GFAP)-positive capsule consisting of a thin collagen layer lining the lumen of the tract, whilst an area with few cells and axons constituted the capsule wall towards the surrounding normal brain tissue. The brain tissue appeared normal outside the capsule boundaries with no difference in areas of stimulation compared with areas of no stimulation. Our results correspond with previous studies performed after fewer months of STN DBS and indicate mild histopathological changes in the vicinity of the electrode tract, appearing to result from the electrode placement and not from the electrical stimulation.

Does chronic low-dose treatment with cyclosporine influence the brain? A histopathological study in pigs.

Twenty percent of the patients immunosuppressed with cyclosporine A (CsA) develop neurological side effects such as tremor, paresthesias, headache, seizures, visual disorders, paresis, and coma-CsA encephalopathy. The encephalopathy resolves on CsA discharge; autopsies of recovered patients are normal. Characteristic lesions are seen on magnetic resonance imaging (MRI) during the period of encephalopathy. MRI of asymptomatic patients receiving CsA as well as most recovered patients are normal. Several theories of pathogenesis have been proposed, but none has been firmly established. The current placebo-controlled study, blinded to the investigator, was accordingly initiated to elucidate histopathological changes in the brain. Twelve adult Göttingen minipigs were randomized into two groups treated with either low-dose CsA (10 mg/kg/d) or no treatment for 6 months. Behavior, blood pressure, and blood parameters were measured throughout the study. All animals had a cerebral MRI before sacrifice. Three control pigs and one CsA-treated pig died during observation and were excluded from the study. None of the remaining eight pigs displayed behavioral signs or MRI-visible lesions characteristic of CsA encephalopathy. The brains appeared all normal on the gross pathological examination, but microscopy revealed perivascular, meningeal, and neuronal tissue infiltration with granulocytes and mononuclear cells in one CsA-treated pig, while the remaining pigs were without histopathological lesions. Pathological changes were noticed in one out of five CsA-treated animals, corresponding to the percentage of patients treated with CsA who develop CsA encephalopathy. To pursue this finding, two studies, one using CsA 20 mg/kg/d for 6 months and one using CsA 10 mg/kg/d for 12 months, have been initiated.

A MRI-compatible stereotaxic localizer box enables high-precision stereotaxic procedures in pigs.

We present a nonmagnetic Plexiglas stereotaxic localizer box that can be fitted directly to the pig skull by aluminum screws, allowing stereotaxic MRI or ventriculography and subsequent high-precision stereotaxic procedures. The localizer box was used to target the subthalamic nucleus (STN) bilaterally in five female Göttingen minipigs. Stereotaxic markers were inserted in the pig skull, the head fixated in the localizer box by aluminum screws inserted bilaterally in the zygoma bone with the hard palate locked on a horizontal palate holder. MRI was obtained on a 3T-MR-imager revealing the relation between the inserted markers and the estimated STN-position, and thus the target coordinates. After the MRI, a stereotaxic frame with attached micromanipulator was locked on to the localizer box converting it into a stereotaxic device. The stereotaxic markers were exposed and used as starting point for the stereotaxic procedure, whereby a microelectrode for electrolytic lesioning was inserted in the STN. Postmortem histological analysis revealed 70% correct STN-targeting. The average distance from the lesion center to the STN center was 1.2 mm with a S.D. of 1.1 mm. The most displaced lesion being 3.6 mm from the STN center. We conclude that the described localizer box secure firm head fixation, allowing stereotaxic MRI and subsequent conversion into a stereotaxic device for high-precision stereotaxic procedures.

Effects of high frequency deep brain stimulation on urine storage and voiding function in conscious minipigs.

AIMS: Suprapontine neural integration during the storage phase is decisive for the timing of voiding. Neurological disorders like Parkinson's disease are thus frequently complicated by bladder dysfunction. The aim of the present study was to investigate the effect of high frequency deep brain stimulation on the urine storage and voiding function in conscious Parkinsonian minipigs. MATERIALS AND METHODS: Five Goettingen minipigs had a Parkinsonism-like state induced by intoxication with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). An electrode for chronic stimulation was placed unilaterally in the subthalamic nucleus. The effect of stimulation was determined by the outcome of transurethral cystometries performed with the stimulation in on- and off-mode. RESULTS: Of 20 planned cystometry-sessions 18 were completed. Six incomplete voidings occurred in stimulation on-mode and five in off-mode. Interruption of the stimulation for 2 days was followed by a significant increase in pressure rise on filling to cystometric capacity, from 7 to 21 cmH(2)O (P = 0.005), and an insignificant reduction in cystometric capacity from 30 to 26 ml/kg bodywt. (P = 0.370), leading to a significant decrease in bladder compliance from 124 to 34 ml/cmH(2)O (P = 0.013). CONCLUSIONS: Transurethral cystometry was a feasible examination technique in pigs. The findings demonstrate that high frequency deep brain stimulation changes the bladder characteristics in the storage phase. Since bladder pressure and capacity responded differently to interruption of stimulation distinct neural mechanisms must be involved in the modulation of sensory information on bladder tension and stretch.

An autometallographic technique for myelin staining in formaldehyde-fixed tissue.

A new autometallographic (AMG) technique for staining myelin in formaldehyde- or paraformaldehyde- (PFA) fixed tissue is presented. The tissue sections were exposed to AMG development without prior treatment with silver salts. The method was examined on PFA-fixed tissue from mouse, rat, pig, and formaldehyde-fixed human autopsy material. Samples from brain, spinal cord, cranial, and spinal nerves were either cut on a vibratome, frozen and cryostat sectioned, or embedded and microtome sectioned, before AMG development and counterstaining. The AMG-myelin technique results in a specific black/dark-brown staining of myelin in all parts of the CNS and PNS. It works on all species examined, independent of the histological preparation techniques applied. The AMG staining is stable, stays unchanged through decades, allows counterstaining, and has previously been used with immunohistochemical techniques. On perfusion-fixed tissue the technique works without further fixation, but the intensity of the AMG-myelin staining is increased by increased postfixation time. Additionally, immersion fixation has to last for days depending on the size of the tissue block in order to obtain proper myelin staining. The most feasible explanation of the chemical events underlying the AMG-myelin technique is that nano-sized clusters of metallic silver are formed in the myelin as a result of chemical bounds with reducing capacity, exposed or created by the formaldehyde molecule. The AMG method is simple to perform and as specific as the conventional osmium and luxol fast blue stainings. The present technique is thus an effective, simple, inexpensive, and quick myelin staining method of formaldehyde- or PFA-fixed tissue.

Effects of intact glucosinolates and products produced from glucosinolates in myrosinase-catalyzed hydrolysis on the potato cyst nematode (Globodera rostochiensis Cv. Woll).

The potato cyst nematode (Globodera rostochiensis cv. Woll) is responsible for large yield losses in the potato crop, and opportunities for reducing the attack of these plant nematode species are, therefore, important. This study has been devoted to the testing of the in vitro effects on the potato cyst nematode of eight glucosinolates [prop-2-enyl-, but-3-enyl-, (R)-4-methylsulfinylbut-3-enyl-, benzyl-, phenethyl-, 4-hydroxybenzyl-, (2S)-2-hydroxybut-3-enyl-, and (2R)-2-hydroxy-2-phenylethylglucosinolate] as well as the effects of the products of this myrosinase-catalyzed hydrolysis. The glucosinolates were used at three concentrations, 0.05, 0.3, and 1.0 mg/mL, in the presence or absence of the enzyme myrosinase. The effects of the compounds on the mortality were monitored every 8 h for a 72 h period. No effects were found for any of the intact glucosinolates. However, when active myrosinase was included with 1 mg/mL phenethylglucosinolate at pH 6.5, 100% mortality was observed within just 16 h. A similar effect was achieved at the same concentration of benzyl- and prop-2-enylglucosinolates in the myrosinase-containing solutions, although longer exposures were required (24 and 40 h, respectively). The main aglucone products released from the glucosinolates with pronounced effects on the nematodes were shown to be the corresponding isothiocyanates. The results suggest that mixtures of these specific glucosinolates and active myrosinase or autolysis of plant materials containing these enzymes and glucosinolates might be used to control the potato cyst nematode in the soil.

New strategies for the treatment of Parkinson's disease hold considerable promise for the future management of neurodegenerative disorders.

Neurodegenerative diseases are often considered incurable with no efficient therapies to modify or halt the progress of disease, and ultimately lead to reduced quality of life and to death. Our knowledge of the nervous system in health and disease has, however, increased considerably during the last fifty years and today, neuroscience reveals promising new strategies to deal with disorders of the nervous system. Some of these results have been implemented with success in the treatment of Parkinson's disease, a common neurodegenerative illness affecting approximately 1% of the population aged seventy or more. Parkinson's disease is characterized by a massive loss of dopaminergic neurons in the substantia nigra, leading to severe functional disturbance of the neuronal circuitry in the basal ganglia. A thorough description of basal ganglia circuitry in health and disease is presented. We describe how the functional disturbances seen in Parkinson's disease may be corrected at specific sites in this circuitry by medical treatment or, in advanced stages of Parkinson's disease, by neurosurgical methods. The latter include lesional surgery, neural transplantation and deep brain stimulation, together with future treatment strategies using direct or indirect implantation of genetically modified cell-lines capable of secreting neurotrophic factors or neurotransmitters. Advantages and disadvantages are briefly mentioned for each strategy and the implications for the future and the possible use of these interventions in other neurodegenerative diseases are discussed, with special emphasis on deep brain stimulation.

New strategies for embedding, orientation and sectioning of small brain specimens enable direct correlation to MR-images, brain atlases, or use of unbiased stereology.

We present a newly developed brain slicing machine and technique for tissue embedding, which enable orientation of fresh or fixed brain tissue from small laboratory animals, in any given position, and subsequent tissue sectioning into slabs with an optional thickness between 0.5 and 20 mm. The oriented tissue slabs may be analysed directly, or processed further on a cryostat or vibratome, into thin stainable histological sections, and aligned to MR-images or brain atlases, depending on the reference used for the initial orientation. Additionally, we describe a new embedding medium (HistOmer) which is an alginate cold polymer ready for instant use after mixing with water. HistOmer allows accurate positioning of the tissue during embedding, and at the same time supports and protects the embedded tissue during sectioning. HistOmer is, therefore, described comprehensively and compared with other commonly used embedding media. This novel slicing technique may also, as illustrated, be used to perform isotropic random orientation of the embedded tissue, before sectioning into tissue slabs of the same thickness. The technique thereby fulfills the requirements for optimal tissue sampling and preparation needed for an unbiased stereologic analysis.

Normalization of markers for dopamine innervation in striatum of MPTP-lesioned miniature pigs with intrastriatal grafts.

As part of the DaNeX study, the uptake and binding of several positron emitting tracers was recorded in brain of healthy Göttingen minipigs, in minipigs with a syndrome of parkinsonism due to MPTP intoxication, and in parkinsonian minipigs which had received intrastriatal grafts of mesencephalic neurons from fetal pigs. The specific binding of [11C]NS 2214 to catecholamine uptake sites was reduced by two thirds in striatum of the intoxicated animals, while the rate constant for the decarboxylation of [18F]fluorodopa was reduced by 50% in the intoxicated animals. Several months after grafting, both pre-synaptic markers of dopamine fibres were normal in striatum. Dopamine depletion or grafting were without effect on the cerebral perfusion rate, measured with [15O]-water, did not alter the rate of oxygen metabolism (CMRO2) in brain, and did not alter the binding potential of tracers for dopamine D1 or D2 receptors in pig striatum. However, the grafting was associated with a local increase in the binding of [11C]PK 11195, a tracer for reactive gliosis, suggesting that an immunological reaction occurs at the site of graft, which might potentially have reduced the graft patency. However, this apparent immunological response did not preclude the re-establishment of normal [18F]fluorodopa and [11C]NS 2214 uptake in the allografted striatum.

[Treatment of severe Parkinson disease with electric stimulation of the subthalamic nuclei]. / Behandling af svaer Parkinsons sygdom med elektrisk stimulation i nucleus subthalamicus.

INTRODUCTION: Patients with Parkinson's disease suffering from severe fluctuations, i.e. dyskinesias and on-off periods and/or severe tremor, who cannot be improved by adjustment of the medical treatment, can be treated with deep brain stimulation via leads implanted bilaterally into the subthalamic nuclei. METHODS: Nine patients with advanced idiopathic Parkinson's disease were treated with stereotactic and bilateral implantation of leads into the subthalamic nuclei. All patients had levodopa responsive Parkinson's disease and levodopa induced fluctuations in the form of on-off periods and dyskinesias. The mean age was 61 years and the mean duration of disease 15 years. The patients were followed for 12 months and evaluated with the Unified Parkinson's Disease Rating Scale off and on medications before and after lead implantation. RESULTS: The Hoehn & Yahr stage fell from 3.9/2.8 before lead implantation and off/on medication to 3.0/2.3 with lead stimulation turned on and off/on medication. The activity of daily living (ADL) index was significantly improved and reduced from 27/13 before lead implantation and off/on medication to 12/7 with the stimulation turned on and off/on medication i.e. with 55%/46% on stimulation. The motor score was likewise significantly improved and reduced from 55/28 to 24/17 i.e. with 56%/39% on stimulation. The most important results of deep brain stimulation in the subthalamic nuclei is the significant reduction of the motor fluctuations. On stimulation off periods were reduced from 30% to 6% and the dyskinesias were significantly reduced from 47% to 14% of the daytime. The mean dose of levodopa equivalent medication was reduced with 26%. DISCUSSION: In conclusion, patients with advanced Parkinson's disease and levodopa induced fluctuations can be successfully treated with bilateral high frequency electric stimulation of leads implanted into the subthalamic nuclei.

Relationship between residual cerebral blood flow and oxygen metabolism as predictive of ischemic tissue viability: sequential multitracer positron emission tomography scanning of middle cerebral artery occlusion during the critical first 6 hours after stroke in pigs.

OBJECT: The authors tested the hypothesis that oxygen metabolism is the key factor linking the long-term viability of ischemic brain tissue to the magnitude of residual blood flow during the first 6 hours following a stroke. METHODS: Eleven anesthetized pigs underwent a series of positron emission tomography studies to measure cerebral blood flow (CBF) and metabolism before and for 7 hours after the animals were subjected to permanent middle cerebral artery (MCA) occlusion. The extent of collateral blood supply was assessed using angiography. Abnormal metabolism of the ischemic tissue progressed as a function of time in inverse proportion to the magnitude of residual CBF, and the volume of the infarct grew in inverse proportion to the residual blood supply. Ten hours after occlusion of the MCA, the infarct topographically matched the tissue with a cerebral metabolic rate of oxygen consumption below 50% of values measured on the contralateral side. This was also the threshold for the decline of the oxygen extraction fraction below normal, which was critical for the prediction of nonviable ischemic tissue. Mildly ischemic tissue (CBF > 30 ml/100 g/min) did not reach the cerebral metabolic rate of oxygen threshold of viability during the first 6 hours after MCA occlusion; moderately ischemic tissue (CBF 12-30 m1/100 g/ min) reached the threshold of viability in 3 hours; and severely ischemic tissue (CBF < 12 ml/100 g/min) remained viable for less than 1 hour. CONCLUSIONS: The relationship between the residual CBF and both oxygen metabolism and extraction is critical to the evolution of metabolic deficiency and lesion size after stroke.

Expression of zinc-positive cells and terminals in fetal neocortical homografts to adult rat depends on lesion type and rearing conditions.

Zinc-positive neurons and terminals, known to be associated with the glutamatergic projections in the brain, can be demonstrated by the histochemical Timm method and later modifications thereof. The adult rat neocortex contain a uniform lamination of zinc-positive cells with specific projections to, e.g., the striatum. We have previously reported that fetal neocortical grafts implanted in the adult rat neocortex combined with rearing in an enriched environment can improve behavioral functions and reduce the secondary atrophy of thalamus after cortex infarction in adult rats. In order to examine whether the expression of zinc positivity is ontogenetically inherent to neocortical neurons we grafted fetal neocortical tissue to aspiration or ischemic lesions of the frontoparietal neocortex of adult rats, followed by histochemical visualization of the vesicular zinc pool by selenite or sulfide. One further aim of the study was to elucidate to what extent the distribution of zinc-containing neurons and terminals in the grafts depended on rearing under different environmental conditions. The foremost finding of the present study was that the overall density of zinc-containing terminals in fetal cortical transplants placed in brain infarcts of adult spontaneously hypertensive rats is higher when the rats are reared in an enriched environment. Moreover, the presence and expression of zinc-positive neurons and terminals do not seem to be ontogenetically inherent to the cortical neurons as the fetal neocortical grafts placed in aspiration lesions contained no zinc-selenide-positive neurons and few or no zinc-selenide-positive terminals. The presence or expression of zinc-positive cells may thus be induced by ingrowth of fibers and terminals from the host brain as transplants placed in the ischemic lesions expressed both zinc-positive neurons and terminals.

Determination of ascorbigens in autolysates of various Brassica species using supercritical fluid chromatography.

A new method of analysis based on normal phase supercritical fluid chromatography (SFC) has been developed for investigation of ascorbigens [2-C-(indol-3-ylmethyl)-beta-L-xylo-3-hexulofuranosonic acid gamma-lactone derivatives]. This method has been adapted to preparative isolation and quantitative determinations of individual ascorbigens comprising ascorbigen, neoascorbigen, and 4-methoxyascorbigen. The structures of these compounds have been revealed from 1D ((1)H, (13)C) and 2D (COSY, HMQC, HMBC) NMR experiments. The developed SFC method had an acceptable linearity for the ascorbigens with correlation coefficients (R(2)) > 0.9995 (n = 10) in the range of 0.13-4.9 nmol injected, detection limits were below 13 pmol, retention time stabilities were excellent, and relative response factors have been determined. The SFC method has been used for determination of ascorbigens produced during autolysis of indol-3-ylmethylglucosinolates in various Brassica vegetables and rapeseed seedlings. Generally, 30-60% of the indol-3-ylmethylglucosinolates in the plants were transformed into ascorbigens, with the concentration in autolysates varying from 0.51 +/- 0.002 to 3.72 +/- 0.21 micromol/g of dry weight (DW) for ascorbigen, from 0.05 +/- 0.01 to 2.42 +/- 0.23 micromol/g of DW for neoascorbigen, and from 0.03 +/- 0.002 to 0.84 +/- 0.07 micromol/g of DW for 4-methoxyascorbigen.