Examining the Duration of Carryover Effect in Patients With Chronic Pain Treated With Spinal Cord Stimulation (EChO Study): An Open, Interventional, Investigator-Initiated, International Multicenter Study.

OBJECTIVES: Spinal cord stimulation (SCS) is a surgical treatment for severe, chronic, neuropathic pain. It is based on one to two lead(s) implanted in the epidural space, stimulating the dorsal column. It has long been assumed that when deactivating SCS, there is a variable interval before the patient perceives the return of the pain, a phenomenon often termed echo or carryover effect. Although the carryover effect has been problematized as a source of error in crossover studies, no experimental investigation of the effect has been published. This open, prospective, international multicenter study aimed to systematically document, quantify, and investigate the carryover effect in SCS. MATERIALS AND METHODS: Eligible patients with a beneficial effect from their SCS treatment were instructed to deactivate their SCS device in a home setting and to reactivate it when their pain returned. The primary outcome was duration of carryover time defined as the time interval from deactivation to reactivation. Central clinical parameters (age, sex, indication for SCS, SCS treatment details, pain score) were registered and correlated with carryover time using nonparametric tests (Mann-Whitney/Kruskal-Wallis) for categorical data and linear regression for continuous data. RESULTS: In total, 158 patients were included in the analyses. A median carryover time of five hours was found (interquartile range 2.5;21 hours). Back pain as primary indication for SCS, high-frequency stimulation, and higher pain score at the time of deactivation were correlated with longer carryover time. CONCLUSIONS: This study confirms the existence of the carryover effect and indicates a remarkably high degree of interindividual variation. The results suggest that the magnitude of carryover may be correlated to the nature of the pain condition and possibly stimulation paradigms. CLINICAL TRIAL REGISTRATION: The Clinicaltrials.gov registration number for the study is NCT03386058.

Distinct Cholesterol Localization in Glioblastoma Multiforme Revealed by Mass Spectrometry Imaging.

Glioblastoma multiforme (GBM) is the most common and aggressive brain tumor in adults and is highly resistant to chemo- and radiotherapies. GBM has been associated with alterations in lipid contents, but lipid metabolism reprogramming in tumor cells is not fully elucidated. One of the key hurdles is to localize the lipid species that are correlated with tumor growth and invasion. A better understanding of the localization of abnormal lipid metabolism and its vulnerabilities may open up to novel therapeutic approaches. Here, we use time-of-flight secondary ion mass spectrometry (ToF-SIMS) to spatially probe the lipid composition in a GBM biopsy from two regions with different histopathologies: one region with most cells of uniform size and shape, the homogeneous part, and the other with cells showing a great variation in size and shape, the heterogeneous part. Our results reveal elevated levels of cholesterol, diacylglycerols, and some phosphatidylethanolamine in the homogeneous part, while the heterogeneous part was dominated by a variety of fatty acids, phosphatidylcholine, and phosphatidylinositol species. We also observed a high expression of cholesterol in the homogeneous tumor region to be associated with large cells but not with macrophages. Our findings suggest that ToF-SIMS can distinguish in lipid distribution between parts within a human GBM tumor, which can be linked to different molecular mechanisms.

Applicability and Validity of an e-Health Tool for the Appropriate Referral and Selection of Patients With Chronic Pain for Spinal Cord Stimulation: Results From a European Retrospective Study.

OBJECTIVES: To support rational decision-making on spinal cord stimulation (SCS), a European expert panel developed an educational e-health tool using the RAND/University of California at Los Angeles Appropriateness Method. This retrospective study aimed to determine the applicability and validity of the tool using data from patients for whom SCS had been considered. MATERIALS AND METHODS: A total of 12 European implant centers retrieved data from 25 to 50 consecutive patients for whom SCS was considered in 2018-2019. For each patient, data were captured on the clinical and psychosocial variables included in the e-health tool, center decisions on SCS, and patient outcomes. Patient outcomes included global perception of effect by the patient and observer, and pain reduction (numeric pain rating scale) at six-month follow-up. RESULTS: In total, 483 patients were included, of whom 133 received a direct implant, 258 received an implant after a positive trial, 32 had a negative trial, and 60 did not receive SCS for reasons other than a negative trial. The most frequent indication was persistent spinal pain syndrome type 1 and type 2 (74%), followed by neuropathic pain syndromes (13%), complex regional pain syndrome (12%), and ischemic pain syndromes (0.8%). Data on the clinical and psychosocial variables were complete for 95% and 93% of patients, respectively, and missing data did not have a significant impact on the study outcomes. In patients who had received SCS, panel recommendations were significantly associated with patient outcomes (p < 0.001 for all measures). Substantial improvement ranged from 25% if the e-health tool outcome was "not recommended" to 83% if SCS was "strongly recommended". In patients who underwent a trial (N = 290), there was 3% of trial failure when SCS was "strongly recommended" vs 46% when SCS was "not recommended". CONCLUSIONS: Retrospective application of the e-health tool on patient data showed a strong relationship between the panel recommendations and both SCS trial results and treatment outcomes.

Patient selection for spinal cord stimulation: The importance of an integrated assessment of clinical and psychosocial factors.

BACKGROUND: A previously developed educational e-health tool considers both clinical and psychosocial factors when selecting patients with chronic pain for spinal cord stimulation (SCS). The validity of the composite recommendations was evaluated in a retrospective study, demonstrating a strong relationship with patient outcomes after SCS. METHODS: An additional retrospective analysis was performed to determine the added value of a psychosocial evaluation as part of the decision-making process on SCS. Data concerned 482 patients who were considered for SCS in 2018-2019. The analysis focused on the relationship between the different layers of the tool recommendations (clinical, psychosocial, composite) with trial results and patient outcomes at 6 months after SCS. Of the initial study population, 381 patients underwent SCS and had follow-up data on at least one of three pain-related outcome measures. RESULTS: Pain improvement was observed in 76% of the patients for whom SCS was strongly recommended based on merely the clinical aspects. This percentage varied by the level of psychosocial problems and ranged from 86% in patients without any compromising psychosocial factors to 60% in those with severe problems. Similarly, the severity of psychosocial problems affected trial results in patients for whom SCS was either recommended or strongly recommended. CONCLUSIONS: The strong relationship between psychosocial factors embedded in the SCS e-health tool and patient outcomes supports an integrated and multidisciplinary approach in the selection of patients for SCS. The educational e-health tool, combining both clinical and psychosocial aspects, is believed to be helpful for further education and implementation of this approach. SIGNIFICANCE STATEMENT: This study confirms the relevance of the psychosocial factors embedded in the educational SCS e-health tool (https://scstool.org/). The strong relationship between the severity of psychosocial factors with patient outcomes supports conducting a comprehensive psychological and behavioural assessment when determining the eligibility of patients for SCS.

The Neurostimulation Appropriateness Consensus Committee (NACC): Recommendations on Best Practices for Cervical Neurostimulation.

INTRODUCTION: The International Neuromodulation Society convened a multispecialty group of physicians based on expertise with international representation to establish evidence-based guidance on the use of neurostimulation in the cervical region to improve outcomes. This Neurostimulation Appropriateness Consensus Committee (NACC) project intends to provide evidence-based guidance for an often-overlooked area of neurostimulation practice. MATERIALS AND METHODS: Authors were chosen based upon their clinical expertise, familiarity with the peer-reviewed literature, research productivity, and contributions to the neuromodulation literature. Section leaders supervised literature searches of MEDLINE, BioMed Central, Current Contents Connect, Embase, International Pharmaceutical Abstracts, Web of Science, Google Scholar, and PubMed from 2017 (when NACC last published guidelines) to the present. Identified studies were graded using the US Preventive Services Task Force criteria for evidence and certainty of net benefit. Recommendations are based on the strength of evidence or consensus when evidence was scant. RESULTS: The NACC examined the published literature and established evidence- and consensus-based recommendations to guide best practices. Additional guidance will occur as new evidence is developed in future iterations of this process. CONCLUSIONS: The NACC recommends best practices regarding the use of cervical neuromodulation to improve safety and efficacy. The evidence- and consensus-based recommendations should be utilized as a guide to assist decision making when clinically appropriate.

Repetitive transcranial magnetic stimulation of the primary motor cortex in management of chronic neuropathic pain: a systematic review.

OBJECTIVES: Repetitive transcranial magnetic stimulation (rTMS) of the primary motor cortex (M1) with frequencies 5-20 Hz is an expanding non-invasive treatment for chronic neuropathic pain (NP). Outcome data, however, show considerable inhomogeneity with concern to the levels of effect due to the great diversity of treated conditions. The aim of this review was to survey the literature regarding the efficacy and safety of M1 rTMS, and the accuracy to predict a positive response to epidural motor cortex stimulation (MCS) which is supposed to give a more longstanding pain relief. METHODS: A systematic literature search was conducted up to June 2019 in accordance with the PRISMA guidelines. We used the PICO Model to define two specific clinical questions: (1) Does rTMS of M1 relieve NP better than sham treatment? (2) Can the response to rTMS be used to predict the effect of epidural MCS? After article selection, data extraction, and study quality assessment, the certainty of evidence of treatment effect was defined using the GRADE system. RESULTS: Data on 5-20 Hz (high-frequency) rTMS vs. sham was extracted from 24 blinded randomised controlled trials which were of varying quality, investigated highly heterogeneous pain conditions, and used excessively variable stimulation parameters. The difference in pain relief between active and sham stimulation was statistically significant in 9 of 11 studies using single-session rTMS, and in 9 of 13 studies using multiple sessions. Baseline data could be extracted from 6 single and 12 multiple session trials with a weighted mean pain reduction induced by active rTMS, compared to baseline, of -19% for single sessions, -32% for multiple sessions with follow-up <30 days, and -24% for multiple sessions with follow-up ≥30 days after the last stimulation session. For single sessions the weighted mean difference in pain reduction between active rTMS and sham was 15 percentage points, for multiple sessions the difference was 22 percentage points for follow-ups <30 days, and 15 percentage points for follow-ups ≥30 days. Four studies reported data that could be used to evaluate the accuracy of rTMS to predict response to MCS, showing a specificity of 60-100%, and a positive predictive value of 75-100%. No serious adverse events were reported. CONCLUSIONS: rTMS targeting M1 can result in significant reduction of chronic NP which, however, is transient and shows a great heterogeneity between studies; very low certainty of evidence for single sessions and low for multiple sessions. Multiple sessions of rTMS can maintain a more longstanding effect. rTMS seems to be a fairly good predictor of a positive response to epidural MCS and may be used to select patients for implantation of permanent epidural electrodes. More studies are needed to manifest the use of rTMS for this purpose. Pain relief outcomes in a longer perspective, and outcome variables other than pain reduction need to be addressed more consistently in future studies to consolidate the applicability of rTMS in routine clinical practice.

Elevated intracranial pressure after head trauma can be suppressed by antisecretory factor-a pilot study.

BACKGROUND: Control of intracranial pressure (ICP) is a key element in neurointensive care for directing treatment decisions in patients with severe traumatic brain injury (TBI). The anti-inflammatory protein antisecretory factor (AF) has been demonstrated to reduce experimentally induced high ICP in animal models. This report describes the first steps to investigate the uptake, safety, and influence of AF for reduction of elevated ICP in patients with TBI in a clinical setting. METHOD: Four patients with severe TBI (Glasgow Coma Scale < 9) that required neurointensive care with ICP monitoring due to signs of refractory intracranial hypertension were investigated. One hundred milliliters of Salovum®, a commercially available egg yolk powder with high contents of AF peptides, was administrated either via nasogastric (patients 1 and 2) or rectal tube (patients 2, 3, and 4) every 8 h for 2 to 3 days as a supplement to the conventional neurointensive care. ICP was registered continuously. Plasma levels of AF were measured by enzyme-linked immunosorbent assay (ELISA) to confirm that Salovum® was absorbed appropriately into the bloodstream. RESULTS: In the first two patients, we observed that when delivered by the nasogastric route, there was an accumulation of the Salovum® solution in the stomach with difficulties to control ICP due to impaired gastric emptying. Therefore, we tested to administer Salovum® rectally. In the third and fourth patients, who both showed radiological signs of extensive brain edema, ICP could be controlled during the course of rectal administration of Salovum®. The ICP reduction was statistically significant and was accompanied by an increase in blood levels of AF. No adverse events that could be attributed to AF treatment or the rectal approach for Salovum® administration were observed. CONCLUSIONS: The outcomes suggest that AF can act as a suppressor of high ICP induced by traumatic brain edema. Use of AF may offer a new therapeutic option for targeting cerebral edema in clinical practice.

Appropriate referral and selection of patients with chronic pain for spinal cord stimulation: European consensus recommendations and e-health tool.

BACKGROUND: Spinal cord stimulation (SCS) is an established treatment for chronic neuropathic, neuropathic-like and ischaemic pain. However, the heterogeneity of patients in daily clinical practice makes it often challenging to determine which patients are eligible for this treatment, resulting in undesirable practice variations. This study aimed to establish patient-specific recommendations for referral and selection of SCS in chronic pain. METHODS: A multidisciplinary European panel used the RAND/UCLA Appropriateness Method (RUAM) to assess the appropriateness of (referral for) SCS for 386 clinical scenarios in four pain areas: chronic low back pain and/or leg pain, complex regional pain syndrome, neuropathic pain syndromes and ischaemic pain syndromes. In addition, the panel identified a set of psychosocial factors that are relevant to the decision for SCS treatment. RESULTS: Appropriateness of SCS was strongly determined by the neuropathic or neuropathic-like pain component, location and spread of pain, anatomic abnormalities and previous response to therapies targeting pain processing (e.g. nerve block). Psychosocial factors considered relevant for SCS selection were as follows: lack of engagement, dysfunctional coping, unrealistic expectations, inadequate daily activity level, problematic social support, secondary gain, psychological distress and unwillingness to reduce high-dose opioids. An educational e-health tool was developed that combines clinical and psychosocial factors into an advice on referral/selection for SCS. CONCLUSIONS: The RUAM was useful to establish a consensus on patient-specific criteria for referral/selection for SCS in chronic pain. The e-health tool may help physicians learn to apply an integrated approach of clinical and psychosocial factors. SIGNIFICANCE: Determining the eligibility of SCS in patients with chronic pain requires careful consideration of a variety of clinical and psychosocial factors. Using a systematic approach to combine evidence from clinical studies and expert opinion, a multidisciplinary European expert panel developed detailed recommendations to support appropriate referral and selection for SCS in chronic pain. These recommendations are available as an educational e-health tool (https://www.scstool.org/).

Optimizing the Management and Outcomes of Failed Back Surgery Syndrome: A Proposal of a Standardized Multidisciplinary Team Care Pathway.

Failed back surgery syndrome (FBSS) is a major, worldwide health problem that generates considerable expense for healthcare systems. A number of controversial issues concerning the management of FBSS are regularly debated, but no clear consensus has been reached. This pitfall is the result of lack of a standardized care pathway due to insufficient characterization of underlying pathophysiological mechanisms, which are essential to identify in order to offer appropriate treatment, and the paucity of evidence of treatment outcomes. In an attempt to address the challenges and barriers in the clinical management of FBSS, an international panel of physicians with a special interest in FBSS established the Chronic Back and Leg Pain (CBLP) Network with the primary intention to provide recommendations through consensus on how to optimize outcomes. In the first of a series of two papers, a definition of FBSS was delineated with specification of criteria for patient assessment and identification of appropriate evaluation tools in order to choose the right treatment options. In this second paper, we present a proposal of a standardized care pathway aiming to guide clinicians in their decision-making on how to optimize their management of FBSS patients. The utilization of a multidisciplinary approach is emphasized to ensure that care is provided in a uniform manner to reduce variation in practice and improve patient outcomes.

Optimizing the Management and Outcomes of Failed Back Surgery Syndrome: A Consensus Statement on Definition and Outlines for Patient Assessment.

Failed back surgery syndrome (FBSS) is a controversial term for identifying patients affected by new, recurrent, or persistent pain in the low back and/or legs following spinal surgery. The lack of a comprehensive standardized care pathway compromises the appropriate management of FBSS patients, which is associated with a heavy financial burden. An international panel of spine surgeons, neurosurgeons, and pain specialists with a particular interest in FBSS established the chronic back and leg pain (CBLP) network with the aim of addressing the challenges and barriers in the clinical management of FBSS patients by building a common transdisciplinary vision. Based on literature reviews, additional input from clinical expertise of multiple professional disciplines, and consensus among its members, the network attempted to provide recommendations on the management of patients with FBSS utilizing a multidisciplinary team (MDT) approach. The presentation of this work has been divided in two separate parts to enhance its clarity. This first paper, in favour of selecting appropriate validated tools to improve the FBSS patient assessment, focuses on FBSS taxonomy and its clinical implications for evaluation. Concise recommendations for assessment, treatment, and outcome evaluation using a MDT approach would be an important resource for specialists and nonspecialist clinicians who manage patients with FBSS, to improve decision-making, reduce variation in practice, and optimize treatment outcomes in this difficult-to-treat population.

Evaluation of the Effectiveness of Percutaneous Octapolar Leads in Pain Treatment with Spinal Cord Stimulation of Patients with Failed Back Surgery Syndrome During a 1-Year Follow-Up: A Prospective Multicenter International Study.

OBJECTIVE: To evaluate the effectiveness and safety of percutaneous octapolar (8-contact) leads in spinal cord stimulation (SCS) treatment of failed back surgery syndrome (FBSS) patients who have not reached their therapy goals with other treatment interventions. METHODS: Our prospective, multicenter, open-label, nonrandomized study included 93 patients ≥ 18 years of age suffering from chronic (≥ 6 months), intractable pain predominantly in the legs. Patients implanted with octapolar lead(s) and a neurostimulator after a successful test trial were followed for 12 months. Patients provided self-reported data on change in visual analog scale (VAS) score for leg pain (primary outcome) and low back pain, quality of life (EuroQol 5 dimensions [EQ-5D] index), sleep, medication use, and paresthesia coverage (secondary outcomes). Adverse events and preferred stimulation settings were documented. RESULTS: Eighty-one (87%) patients had a successful SCS trial. Patients reported significantly improved leg pain relief; average VAS score was 72 ± 17 prior to SCS treatment and 32 ± 24 at 12 months (P < 0.001). Significant decrease in back pain (P < 0.001), improvement in quality of life (P < 0.001), and improvement in sleep (P < 0.05) was observed. Sixty-three percent and 40% of patients were responders (≥ 50% pain reduction) on leg and back pain, respectively, after 12 months. A decrease in medication use was seen for antidepressants and anticonvulsants. Eighty-eight percent of the patients managed with 1 or 2 programs for optimal effect and paresthesia coverage. Twenty-five SCS-related adverse events were registered in 22 patients (24%). Surgical revision due to lead displacement or dysfunction was needed in 6 (6%) of the patients. CONCLUSIONS: Use of percutaneous octapolar SCS leads gives significant long-term pain relief and improvement in quality of life and sleep in FBSS patients. The outcomes are better than reported on 4-contact leads and indicate that the progress in SCS technology that has taken place during the past decade correlates with therapy improvements.

Uptake of the antisecretory factor peptide AF-16 in rat blood and cerebrospinal fluid and effects on elevated intracranial pressure.

BACKGROUND: AF-16 is a 16-amino-acid-long peptide derived from the amino-terminal part of the endogenous protein, antisecretory factor (AF). AF-16 in vivo has been shown to regulate dysfunctions in the water and ion transport system under various pathological conditions and also to counteract experimentally increased tissue pressure. METHODS: Rats were subjected to a cryogenic brain injury in order to increase the intracranial pressure (ICP). The distribution of AF-16 in blood and CSF after intravenous or intranasal administration was determined in injured and control rats. ICP was monitored in freely moving, awake rats, by means of an epidural pressure transducer catheter connected to a wireless device placed subcutaneously on the skull. The continuous ICP registrations were achieved by means of telemetry. RESULTS: Intranasal administration of AF-16 resulted in a significantly higher CSF concentrations of AF-16 in injured than in control rats, 1.3 versus 0.6 ng/ml, whereas no difference between injured and control rats was seen when AF-16 was given intravenously. Rats subjected to cryogenic brain injury developed gradually increasing ICP levels. Intranasal administration of AF-16 suppressed the increased ICP to normal values within 30 min. CONCLUSION: Optimal AF-16 concentrations in CSF are achieved after intranasal administration in rats subjected to a cryogenic brain injury. The ability of AF-16 to suppress an increased ICP was manifested.

The appropriate use of neurostimulation of the spinal cord and peripheral nervous system for the treatment of chronic pain and ischemic diseases: the Neuromodulation Appropriateness Consensus Committee.

INTRODUCTION: The Neuromodulation Appropriateness Consensus Committee (NACC) of the International Neuromodulation Society (INS) evaluated evidence regarding the safety and efficacy of neurostimulation to treat chronic pain, chronic critical limb ischemia, and refractory angina and recommended appropriate clinical applications. METHODS: The NACC used literature reviews, expert opinion, clinical experience, and individual research. Authors consulted the Practice Parameters for the Use of Spinal Cord Stimulation in the Treatment of Neuropathic Pain (2006), systematic reviews (1984 to 2013), and prospective and randomized controlled trials (2005 to 2013) identified through PubMed, EMBASE, and Google Scholar. RESULTS: Neurostimulation is relatively safe because of its minimally invasive and reversible characteristics. Comparison with medical management is difficult, as patients considered for neurostimulation have failed conservative management. Unlike alternative therapies, neurostimulation is not associated with medication-related side effects and has enduring effect. Device-related complications are not uncommon; however, the incidence is becoming less frequent as technology progresses and surgical skills improve. Randomized controlled studies support the efficacy of spinal cord stimulation in treating failed back surgery syndrome and complex regional pain syndrome. Similar studies of neurostimulation for peripheral neuropathic pain, postamputation pain, postherpetic neuralgia, and other causes of nerve injury are needed. International guidelines recommend spinal cord stimulation to treat refractory angina; other indications, such as congestive heart failure, are being investigated. CONCLUSIONS: Appropriate neurostimulation is safe and effective in some chronic pain conditions. Technological refinements and clinical evidence will continue to expand its use. The NACC seeks to facilitate the efficacy and safety of neurostimulation.

The peptide AF-16 and the AF protein counteract intracranial hypertension.

Intracranial hypertension develops after, for example, trauma, stroke and brain inflammation, and contributes to increased morbidity, mortality, and persistent neuropsychiatric sequelae. Nonsurgical therapy offers limited relief. We investigated whether the peptide AF-16 and the endogenous protein Antisecretory Factor (AF) counteracted abnormal fluid transfer by cells, and lowered raised intracranial pressure (ICP). Adult rats, infected with an encephalitogenic Herpes simplex virus (HSV-1), developed after 5 days' sickness of increasing severity. AF-16 rescued all rats while vehicle treatment only saved 20%. AF-16 from day 4 reduced the ICP in HSV-1-infected rats from 30.7 to 14.6 mmHg and all survived without sequelae. A standardised closed head brain injury in rats raised the ICP. Continuous and intermittent AF-16 kept ICP at an almost normal level. A single dose of AF-16 maintained the raised ICP after a TBI lowered during 3-9 h. The AF protein, enriched in egg yolk, similarly lowered the post-traumatically raised ICP in rats. AF-16 also lowered the ICP in rabbits with diffuse brain injury. We conclude that the peptide AF-16 and the AF protein offer new approaches to treat raised ICP with no side effects.

The peptide AF-16 decreases high interstitial fluid pressure in solid tumors.

BACKGROUND: The high interstitial fluid pressure (IFP) in solid tumors restricts the access to nutrients, oxygen and drugs. MATERIAL AND METHODS: We investigated the ability of the peptide AF-16, involved in water and ion transfer through cell membranes, to lower the IFP in two different solid rat mammary tumors, one chemically induced, slowly growing, and the other transplantable, and rapidly progressing having high cellularity. AF-16 was administered either in the tumor capsule, intranasally or intravenously. The IFP was measured by a miniature fiber optic device. RESULTS: AF-16 significantly lowered the IFP in both the slowly and the rapidly progressing tumors, whether administrated locally or systemically. The AF-16 induced IFP reduction was maximal after 90 min, lasted at least 3 h, and returned to pretreatment levels in less than 24 h. Topical AF-16 transiently reduced the IFP in the DMBA tumors from 17.7 ± 4.2 mmHg to 8.6 ± 2.1 mmHg. CONCLUSION: We conclude that AF-16 transiently and reversibly lowered the high IFP in solid tumors during a few hours, which might translate into improved therapeutic efficacy.

Nuclear translocation and calpain-dependent reduction of Bcl-2 after neonatal cerebral hypoxia-ischemia.

Apoptosis-related mechanisms are important in the pathophysiology of hypoxic-ischemic injury in the neonatal brain. Caspases are the major executioners of apoptosis, but there are a number of upstream players that influence the cell death pathways. The Bcl-2 family proteins are important modulators of mitochondrial permeability, working either to promote or prevent apoptosis. In this study we focused on the anti-apoptotic Bcl-2 protein after neonatal cerebral hypoxia-ischemia (HI) in 8-day-old rats. Bcl-2 translocated to nuclei and accumulated there over the first 24h of reperfusion after HI, as judged by immunohistochemistry and immuno-electron microscopy. We also found that the total level of Bcl-2 decreased after HI in vivo and after ionophore challenge in cultured human neuroblastoma (IMR-32) cells in vitro. Furthermore, the Bcl-2 reduction was calpain-dependent, because it could be prevented by the calpain inhibitor CX295 both in vivo and in vitro, suggesting cross-talk between excitotoxic and apoptotic mechanisms.

The role of NGF uptake in selective vulnerability to cell death in ageing sympathetic neurons.

We have examined the hypothesis that differences in nerve growth factor (NGF) uptake and transport determine vulnerability to age-related neurodegeneration. Neurons projecting to cerebral blood vessels (CV) in aged rats are more vulnerable to age-related degeneration than those projecting to the iris. Uptake of NGF was therefore examined in sympathetic neurons projecting from the superior cervical ganglion (SCG) to CV and iris in young and old rats by treating the peripheral processes of these neurons with different doses of I125-NGF. Total uptake of I125-NGF was reduced in old CV-projecting, but not iris-projecting, neurons. Numbers of radiolabelled neurons projecting to each target were counted in sectioned ganglia. The data showed age-related reductions in numbers of labelled neurons projecting to CV, but no change in numbers of neurons projecting to the iris. Calculation of uptake of I125-NGF per neuron unexpectedly showed no major age-related differences in either of the two neuron populations. However, uptake per neuron was considerably lower for young and old CV-projecting, compared to iris-projecting, SCG neurons. We hypothesized that variations in NGF uptake might affect neuronal survival in old age. Counts of SCG neurons using a physical disector following retrograde tracing with Fluorogold confirmed the selective vulnerability of CV-projecting neurons by showing a significant 37% loss of these neurons in the period between 15 and 24 months. In contrast, there was no significant loss of iris-projecting neurons. We conclude that vulnerability to, or protection from, age-related neurodegeneration and neuronal cell death are associated with life-long low, or high, levels of NGF uptake, respectively.

Early onset of degenerative changes at nodes of Ranvier in alpha-motor axons of Cntf null (-/-) mutant mice.

The nodes of Ranvier are sites of specific interaction between Schwann cells and axons. Besides their crucial role in transmission of action potentials, the nodes of Ranvier and in particular the paranodal axon-Schwann cell networks (ASNs) are thought to function as local centers in large motor axons for removal, degradation, and disposal of organelles. In order to test whether ciliary neurotrophic factor (CNTF), which is present at high levels in the Schwann cell cytoplasm, is involved in the maintenance of these structures, we have examined lumbar ventral root nerve fibers of alpha-motor neurons by electron microscopy in 3- and 9-month-old Cntf null ((-/-)) mutant mice. Nerve fibers and nodes of Ranvier in 3-month-old Cntf(-/-) mutants appeared morphologically normal, except that ASNs were more voluminous in the mutants than in wild-type control animals at this age. In 9-month-old Cntf(-/-) animals, morphological changes, such as reduction in nerve fiber and axon diameter, myelin sheath disruption, and loss of ASNs at nodes of Ranvier, were observed. These findings suggest that endogenous CNTF, in addition to its role in promoting motor neuron survival and regeneration, is needed for long-term maintenance of alpha-motor nerve fibers. The premature loss of paranodal ASNs in animals lacking CNTF, which seems to be a defect related to a disturbed interaction in the nodal region between the axon and its myelinating Schwann cells, could impede the maintenance of a normal milieu in the motor axon, preceding more general neuronal damage.