Correlation of diffusion tensor tractography and intraoperative macrostimulation during deep brain stimulation for Parkinson disease.

Object The purpose of this study was to investigate whether diffusion tensor imaging (DTI) of the corticospinal tract (CST) is a reliable surrogate for intraoperative macrostimulation through the deep brain stimulation (DBS) leads. The authors hypothesized that the distance on MRI from the DBS lead to the CST as determined by DTI would correlate with intraoperative motor thresholds from macrostimulations through the same DBS lead. Methods The authors retrospectively reviewed pre- and postoperative MRI studies and intraoperative macrostimulation recordings in 17 patients with Parkinson disease (PD) treated by DBS stimulation. Preoperative DTI tractography of the CST was coregistered with postoperative MRI studies showing the position of the DBS leads. The shortest distance and the angle from each contact of each DBS lead to the CST was automatically calculated using software-based analysis. The distance measurements calculated for each contact were evaluated with respect to the intraoperative voltage thresholds that elicited a motor response at each contact. Results There was a nonsignificant trend for voltage thresholds to increase when the distances between the DBS leads and the CST increased. There was a significant correlation between the angle and the voltage, but the correlation was weak (coefficient of correlation [R] = 0.36). Conclusions Caution needs to be exercised when using DTI tractography information to guide DBS lead placement in patients with PD. Further studies are needed to compare DTI tractography measurements with other approaches such as microelectrode recordings and conventional intraoperative MRI-guided placement of DBS leads.

Chronic deep brain stimulation in mesial temporal lobe epilepsy.

The objective of this study was to evaluate the efficiency and the effects of changes in parameters of chronic amygdala-hippocampal deep brain stimulation (AH-DBS) in mesial temporal lobe epilepsy (TLE). Eight pharmacoresistant patients, not candidates for ablative surgery, received chronic AH-DBS (130 Hz, follow-up 12-24 months): two patients with hippocampal sclerosis (HS) and six patients with non-lesional mesial TLE (NLES). The effects of stepwise increases in intensity (0-Off to 2 V) and stimulation configuration (quadripolar and bipolar), on seizure frequency and neuropsychological performance were studied. The two HS patients obtained a significant decrease (65-75%) in seizure frequency with high voltage bipolar DBS (≥1 V) or with quadripolar stimulation. Two out of six NLES patients became seizure-free, one of them without stimulation, suggesting a microlesional effect. Two NLES patients experienced reductions of seizure frequency (65-70%), whereas the remaining two showed no significant seizure reduction. Neuropsychological evaluations showed reversible memory impairments in two patients under strong stimulation only. AH-DBS showed long-term efficiency in most of the TLE patients. It is a valuable treatment option for patients who suffer from drug resistant epilepsy and who are not candidates for resective surgery. The effects of changes in the stimulation parameters suggest that a large zone of stimulation would be required in HS patients, while a limited zone of stimulation or even a microlesional effect could be sufficient in NLES patients, for whom the importance of the proximity of the electrode to the epileptogenic zone remains to be studied. Further studies are required to ascertain these latter observations.

Event-related potential analyses via single-subject topographic classification.

Introduction: Responses to external stimuli are typically investigated by averaging peri-stimulus electroencephalography (EEG) epochs in order to derive event-related potentials (ERPs) across the electrode montage, under the assumption that signals that are related to the external stimulus are fixed in time across trials. We demonstrate the applicability of a single-trial model based on patterns of scalp topographies (De Lucia et al, 2007) that can be used for ERP analysis at the single-subject level. The model is able to classify new trials (or groups of trials) with minimal a priori hypotheses, using information derived from a training dataset. The features used for the classification (the topography of responses and their latency) can be neurophysiologically interpreted, because a difference in scalp topography indicates a different configuration of brain generators. An above chance classification accuracy on test datasets implicitly demonstrates the suitability of this model for EEG data. Methods: The data analyzed in this study were acquired from two separate visual evoked potential (VEP) experiments. The first entailed passive presentation of checkerboard stimuli to each of the four visual quadrants (hereafter, "Checkerboard Experiment") (Plomp et al, submitted). The second entailed active discrimination of novel versus repeated line drawings of common objects (hereafter, "Priming Experiment") (Murray et al, 2004). Four subjects per experiment were analyzed, using approx. 200 trials per experimental condition. These trials were randomly separated in training (90%) and testing (10%) datasets in 10 independent shuffles. In order to perform the ERP analysis we estimated the statistical distribution of voltage topographies by a Mixture of Gaussians (MofGs), which reduces our original dataset to a small number of representative voltage topographies. We then evaluated statistically the degree of presence of these template maps across trials and whether and when this was different across experimental conditions. Based on these differences, single-trials or sets of a few single-trials were classified as belonging to one or the other experimental condition. Classification performance was assessed using the Receiver Operating Characteristic (ROC) curve. Results: For the Checkerboard Experiment contrasts entailed left vs. right visual field presentations for upper and lower quadrants, separately. The average posterior probabilities, indicating the presence of the computed template maps in time and across trials revealed significant differences starting at ~60-70 ms post-stimulus. The average ROC curve area across all four subjects was 0.80 and 0.85 for upper and lower quadrants, respectively and was in all cases significantly higher than chance (unpaired t-test, p<0.0001). In the Priming Experiment, we contrasted initial versus repeated presentations of visual object stimuli. Their posterior probabilities revealed significant differences, which started at 250ms post-stimulus onset. The classification accuracy rates with single-trial test data were at chance level. We therefore considered sub-averages based on five single trials. We found that for three out of four subjects' classification rates were significantly above chance level (unpaired t-test, p<0.0001). Conclusions: The main advantage of the present approach is that it is based on topographic features that are readily interpretable along neurophysiologic lines. As these maps were previously normalized by the overall strength of the field potential on the scalp, a change in their presence across trials and between conditions forcibly reflects a change in the underlying generator configurations. The temporal periods of statistical difference between conditions were estimated for each training dataset for ten shuffles of the data. Across the ten shuffles and in both experiments, we observed a high level of consistency in the temporal periods over which the two conditions differed. With this method we are able to analyze ERPs at the single-subject level providing a novel tool to compare normal electrophysiological responses versus single cases that cannot be considered part of any cohort of subjects. This aspect promises to have a strong impact on both basic and clinical research.

Greenstick fractures of the middle third of the forearm. A prospective multi-centre study.

BACKGROUND: Greenstick fractures suffered during growth have a high risk for refracture and posttraumatic deformity, particularly at the forearm diaphysis. The use of a preemptive completion of the fracture by manipulation of the concave cortex is controversial and data supporting this approach are few. AIM: Aim of this study was to determine the factors which predispose to refracture and deformities, and to define therapeutic strategies. METHODS: We prospectively gathered clinical and radiographic data over a period of one year on greenstick fractures of the middle third of the forearm in children as part of a multi-centre study. Endpoint was a follow-up visit at one year. Radiographic deformity, state of consolidation at resumption of physical activities and refracture rate were analysed statistically (ANOVA, Student's t-test and Pearson's chi-square test) with regard to patient age, gender, fracture type, therapy and time in plaster. RESULTS: We collected the data of 103 patients (63 boys, 40 girls), average age 6.6 years (1.3-14.5 years), the vast majority of whom had a combined greenstick fracture of the radius and ulna. 6.7% of the patients sustained a refracture within 49 days (29-76) after plaster removal. They were significantly older (p=0.017) with a significantly higher incidence of manual completion of the fracture with radiographic signs of partial consolidation (p=0.025). Residual deformities were significantly smaller after completion of the fracture compared to reduction without completion (p=0.019) or plaster fixation alone (p<0.005). CONCLUSIONS: Completion of a greenstick fracture does not prevent refracture. Nevertheless, it diminishes the extent of secondary deformities in cases where the primary angulation exceeds the remodelling capacity. Prevention of refracture should include a routine radiographic follow-up 4-6 weeks after injury with continuation of plaster fixation in cases of partial consolidation.

The GTPase RalA regulates different steps of the secretory process in pancreatic beta-cells.

BACKGROUND: RalA and RalB are multifuntional GTPases involved in a variety of cellular processes including proliferation, oncogenic transformation and membrane trafficking. Here we investigated the mechanisms leading to activation of Ral proteins in pancreatic beta-cells and analyzed the impact on different steps of the insulin-secretory process. METHODOLOGY/PRINCIPAL FINDINGS: We found that RalA is the predominant isoform expressed in pancreatic islets and insulin-secreting cell lines. Silencing of this GTPase in INS-1E cells by RNA interference led to a decrease in secretagogue-induced insulin release. Real-time measurements by fluorescence resonance energy transfer revealed that RalA activation in response to secretagogues occurs within 3-5 min and reaches a plateau after 10-15 min. The activation of the GTPase is triggered by increases in intracellular Ca2+ and cAMP and is prevented by the L-type voltage-gated Ca2+ channel blocker Nifedipine and by the protein kinase A inhibitor H89. Defective insulin release in cells lacking RalA is associated with a decrease in the secretory granules docked at the plasma membrane detected by Total Internal Reflection Fluorescence microscopy and with a strong impairment in Phospholipase D1 activation in response to secretagogues. RalA was found to be activated by RalGDS and to be severely hampered upon silencing of this GDP/GTP exchange factor. Accordingly, INS-1E cells lacking RalGDS displayed a reduction in hormone secretion induced by secretagogues and in the number of insulin-containing granules docked at the plasma membrane. CONCLUSIONS/SIGNIFICANCE: Taken together, our data indicate that RalA activation elicited by the exchange factor RalGDS in response to a rise in intracellular Ca2+ and cAMP controls hormone release from pancreatic beta-cell by coordinating the execution of different events in the secretory pathway.

Spontaneous pre-stimulus fluctuations in the activity of right fronto-parietal areas influence inhibitory control performance.

Inhibitory control refers to the ability to suppress planned or ongoing cognitive or motor processes. Electrophysiological indices of inhibitory control failure have been found to manifest even before the presentation of the stimuli triggering the inhibition, suggesting that pre-stimulus brain-states modulate inhibition performance. However, previous electrophysiological investigations on the state-dependency of inhibitory control were based on averaged event-related potentials (ERPs), a method eliminating the variability in the ongoing brain activity not time-locked to the event of interest. These studies thus left unresolved whether spontaneous variations in the brain-state immediately preceding unpredictable inhibition-triggering stimuli also influence inhibitory control performance. To address this question, we applied single-trial EEG topographic analyses on the time interval immediately preceding NoGo stimuli in conditions where the responses to NoGo trials were correctly inhibited [correct rejection (CR)] vs. committed [false alarms (FAs)] during an auditory spatial Go/NoGo task. We found a specific configuration of the EEG voltage field manifesting more frequently before correctly inhibited responses to NoGo stimuli than before FAs. There was no evidence for an EEG topography occurring more frequently before FAs than before CR. The visualization of distributed electrical source estimations of the EEG topography preceding successful response inhibition suggested that it resulted from the activity of a right fronto-parietal brain network. Our results suggest that the fluctuations in the ongoing brain activity immediately preceding stimulus presentation contribute to the behavioral outcomes during an inhibitory control task. Our results further suggest that the state-dependency of sensory-cognitive processing might not only concern perceptual processes, but also high-order, top-down inhibitory control mechanisms.

The role of LEKTI in fate determination of keratinocyte stem cells

SPINK5 (serine protease inhibitor Kazal-type 5) encodes the putative proteinase inhibitor LEKTI (lympho-epithelial Kazal-type related inhibitor). In skin, LEKTI expression is restricted to the stratum granulosum of the epidermis and the inner root sheath of hair follicles. Mutations that create premature termination codons in SPINK5 have been reported as the cause of Netherton syndrome (NS), a human autosomal recessive disorder characterized by congenital ichthyosis with defective cornification, a specific hair shaft defect known as trichorrexis invaginata or 'bamboo hair', and severe atopic manifestations, including atopic dermatitis and hayfever. Althought recombinant human LEKTI inhibits a battery of serine proteases including plasmin, trypsin, subtilisin A, cathepsin G, and elastase, the precise role of LEKTI in the physiopathology of NS remains unclear. Spink5−/− mice display a NS-like phenotype. Surprisingly, a psoriasis-like hyperplasia, basement membrane breakdown followed by evagination of spindle-shaped epidermal cells into the dermal compartment, and the presence of numerous sweat gland-like structures were also observed when the skin of Spink5−/− newborn mice, which die at birth, was transplanted onto the back of nude mice. Collectively, these observations suggest that LEKTI may play a role on cell proliferation and stem cell fate. Our current work aims at elucidating the mechanisms by which LEKTI impact these biological processes. Using keratinocyte stem cells obtained from NS patients, we have identified LEKTI as a regulator node in several signaling pathways involved in stem cell behavior.

A role for mitochondria in NLRP3 inflammasome activation.

An inflammatory response initiated by the NLRP3 inflammasome is triggered by a variety of situations of host 'danger', including infection and metabolic dysregulation. Previous studies suggested that NLRP3 inflammasome activity is negatively regulated by autophagy and positively regulated by reactive oxygen species (ROS) derived from an uncharacterized organelle. Here we show that mitophagy/autophagy blockade leads to the accumulation of damaged, ROS-generating mitochondria, and this in turn activates the NLRP3 inflammasome. Resting NLRP3 localizes to endoplasmic reticulum structures, whereas on inflammasome activation both NLRP3 and its adaptor ASC redistribute to the perinuclear space where they co-localize with endoplasmic reticulum and mitochondria organelle clusters. Notably, both ROS generation and inflammasome activation are suppressed when mitochondrial activity is dysregulated by inhibition of the voltage-dependent anion channel. This indicates that NLRP3 inflammasome senses mitochondrial dysfunction and may explain the frequent association of mitochondrial damage with inflammatory diseases.

Traitement des fractures de l'enfant par enclouage centro-médullaire élastique stable (ECMES) [Treatment of pediatric fractures by intramedullary stable elastic pinning]

During the past decade several new techniques for the treatment of children's fractures respecting the specificity of the growing bone have been described. The goal of all these techniques was to mechanically stabilise the fracture however to preserve a certain instability of the fracture gap itself inducing early callus formation and subsequent consolidation. The dynamic external fixation as well as the elastic stable intramedullary pinning have become accepted means in the treatment of long bone fractures in the paediatric age group. We report our experience of the last seven years with the intramedullary pinning of 105 fractures. Eighty-four were fractures of the femur, 9 of the humerus, 8 of the forearm, and a further 4 of the tibial shaft. The intramedullary elastic pinning represents a simple technique which supports or even enhances the natural process of fracture healing of the growing bone. The method is not very invasive, is cost effective, and allows short hospitalisation. Early physical activity is guaranteed due to early consolidation of the fracture. Complications are rare and the final orthopedic and cosmetic outcome is excellent.

Revue du traitement chirurgical de la pseudarthrose de la diaphyse fémorale

Ce travail consiste en l'analyse et la revue des différentes prises en charge de la pseudarthrose de la diaphyse fémorale dans un groupe nommé « Echantillon CHUV » formé par 16 patients. Les patients ont tous été opérés au Centre Hospitalier Universitaire Vaudois, CHUV, entre 2008 et 2011 pour cure de pseudarthrose. Une présentation succincte des fractures de la diaphyse fémorale ainsi que de la pseudarthrose de la diaphyse fémorale consiste en la première partie du rapport. La deuxième partie est une revue des cas des patients de l'échantillon CHUV. Des paramètres tels que le mode initial de fracture, la durée effective de la pseudarthrose, le nombre de révisions avant la consolidation ou la répartition des pseudarthroses selon leur type font partie des différents éléments caractéristiques répertoriés puis analysés dans cette revue. 25 cures de pseudarthroses sont effectuées sur les 16 patients. Les techniques de fixations les plus utilisées sont la fixation par plaque après réduction ouverte (ORIF, 64%, 16 cures sur 25) et l'enclouage centromédullaire (24%, 6 cures sur 25). L'utilisation de substituts osseux est très souvent complémentaire à la refixation mécanique lors des cures de pseudarthrose (recours à la greffe osseuse dans 72% des cures). Le taux d'union après la première cure de pseudarthrose s'établit à 63% et il monte à 100% après l'ensemble des cures. Le retour à une fonction adéquate du membre inférieur est obtenu chez 14 des 16 patients (1 patient est traité par arthrodèse du genou sur pseudarthrose du tiers distal de la diaphyse fémorale et 1 patient tétraplégique est traité par résection de la tête et du col fémoral sur pseudarthrose du tiers proximal de la diaphyse fémorale). La revue quantifie la durée de l'invalidité causée par la pseudarthrose de la diaphyse fémorale : la longueur totale du traitement avant union est d'au minimum 12 mois pour 75% des patients. 8 patients sur 16 ont un temps avant union supérieur à 20 mois. La pseudarthrose de la diaphyse fémorale doit être considérée comme une complication grave. La localisation au niveau du fémur réduit voire abolit l'autonomie de marche du patient et limite ses activités de la vie quotidienne. Elle entrave le retour au travail et réduit terriblement la qualité de vie pendant souvent plus d'une année. L'ensemble des symptômes ont un effet dévastateur sur la rééducation et peuvent parfois laisser d'importantes séquelles physiques ou psychologiques sur le long terme. Pour ces raisons, le traitement doit être adapté du mieux possible à la pseudarthrose du patient afin d'augmenter les chances de réussite de la cure.

Protonation controls ASIC1a activity via coordinated movements in multiple domains.

Acid-sensing ion channels (ASICs) are neuronal Na(+)-conducting channels activated by extracellular acidification. ASICs are involved in pain sensation, expression of fear, and neurodegeneration after ischemic stroke. Functional ASICs are composed of three identical or homologous subunits, whose extracellular part has a handlike structure. Currently, it is unclear how protonation of residues in extracellular domains controls ASIC activity. Knowledge of these mechanisms would allow a rational development of drugs acting on ASICs. Protonation may induce conformational changes that control the position of the channel gate. We used voltage-clamp fluorometry with fluorophores attached to residues in different domains of ASIC1a to detect conformational changes. Comparison of the timing of fluorescence and current signals identified residues involved in movements that preceded desensitization and may therefore be associated with channel opening or early steps leading to desensitization. Other residues participated in movements intimately linked to desensitization and recovery from desensitization. Fluorescence signals of all mutants were detected at more alkaline pH than ionic currents. Their midpoint of pH dependence was close to that of steady-state desensitization, whereas the steepness of the pH fluorescence relationship was closer to that of current activation. A sequence of movements was observed upon acidification, and its backward movements during recovery from desensitization occurred in the reverse order, indicating that the individual steps are interdependent. Furthermore, the fluorescence signal of some labeled residues in the finger domain was strongly quenched by a Trp residue in the neighboring β-ball domain. Upon channel activation, their fluorescence intensity increased, indicating that the finger moved away from the β ball. This extensive analysis of activity-dependent conformational changes in ASICs sheds new light on the mechanisms by which protonation controls ASIC activity.

Synapses on motoneuron dendrites in the brachial section of the frog spinal cord: a computer-aided electron microscopic study of cobalt-filled cells.

Cobalt-labelled motoneuron dendrites of the frog spinal cord at the level of the second spinal nerve were photographed in the electron microscope from long series of ultrathin sections. Three-dimensional computer reconstructions of 120 dendrite segments were analysed. The samples were taken from two locations: proximal to cell body and distal, as defined in a transverse plane of the spinal cord. The dendrites showed highly irregular outlines with many 1-2 microns-long 'thorns' (on average 8.5 thorns per 100 microns 2 of dendritic area). Taken together, the reconstructed dendrite segments from the proximal sites had a total length of about 250 microns; those from the distal locations, 180 microns. On all segments together there were 699 synapses. Nine percent of the synapses were on thorns, and many more close to their base on the dendritic shaft. The synapses were classified in four groups. One third of the synapses were asymmetric with spherical vesicles; one half were symmetric with spherical vesicles; and one tenth were symmetric with flattened vesicles. A fourth, small class of asymmetric synapses had dense-core vesicles. The area of the active zones was large for the asymmetric synapses (median value 0.20 microns 2), and small for the symmetric ones (median value 0.10 microns 2), and the difference was significant. On average, the areas of the active zones of the synapses on thin dendrites were larger than those of synapses on large calibre dendrites. About every 4 microns 2 of dendritic area received one contact. There was a significant difference between the areas of the active zones of the synapses at the two locations. Moreover, the number per unit dendritic length was correlated with dendrite calibre. On average, the active zones covered more than 4% of the dendritic area; this value for thin dendrites was about twice as large as that of large calibre dendrites. We suggest that the larger active zones and the larger synaptic coverage of the thin dendrites compensate for the longer electrotonic distance of these synapses from the soma.

Electrically assisted ocular gene therapy.

Electrotransfer and iontophoresis are being developed as innovative non-viral gene delivery systems for the treatment of eye diseases. These two techniques rely on the use of electric current to allow for higher transfection yield of various ocular cell types in vivo. Short pulses of relatively high-intensity electric fields are used for electrotransfer delivery, whereas the iontophoresis technique is based on the application of low voltage electric current. The basic principles of these techniques and their potential therapeutic application for diseases of the anterior and posterior segments of the eye are reviewed. Iontophoresis has been found most efficient for the delivery of small nucleic acid fragments such as antisense oligonucleotides, siRNA, or ribozymes. Electrotransfer, on the other hand, is being developed for the delivery of oligonucleotides or custom designed plasmids. The wide range of strategies already validated and the potential for targeting specific types of cells confirm the promising early observations made using electrotransfer and iontophoresis. These two nonviral delivery systems are safe and can be used efficiently for targeted gene delivery to ocular tissues in vivo. At the present, their application for the treatment of ocular human diseases is nearing its final stages of adaptation and practical implementation at the bedside.

Nitric oxide reduces Cl- absorption in the mouse cortical collecting duct through an ENaC-dependent mechanism.

Since nitric oxide (NO) participates in the renal regulation of blood pressure, in part, by modulating transport of Na(+) and Cl(-) in the kidney, we asked whether NO regulates net Cl(-) flux (JCl) in the cortical collecting duct (CCD) and determined the transporter(s) that mediate NO-sensitive Cl(-) absorption. Cl(-) absorption was measured in CCDs perfused in vitro that were taken from aldosterone-treated mice. Administration of an NO donor (10 μM MAHMA NONOate) reduced JCl and transepithelial voltage (VT) both in the presence or absence of angiotensin II. However, reducing endogenous NO production by inhibiting NO synthase (100 μM N(G)-nitro-l-arginine methyl ester) increased JCl only in the presence of angiotensin II, suggesting that angiotensin II stimulates NO synthase activity. To determine the transport process that mediates NO-sensitive changes in JCl, we examined the effect of NO on JCl following either genetic ablation or chemical inhibition of transporters in the CCD. Since the application of hydrochlorothiazide (100 μM) or bafilomycin (5 nM) to the perfusate or ablation of the gene encoding pendrin did not alter NO-sensitive JCl, NO modulates JCl independent of the Na(+)-dependent Cl(-)/HCO3(-) exchanger (NDCBE, Slc4a8), the A cell apical plasma membrane H(+)-ATPase and pendrin. In contrast, both total and NO-sensitive JCl and VT were abolished with application of an epithelial Na(+) channel (ENaC) inhibitor (3 μM benzamil) to the perfusate. We conclude that NO reduces Cl(-) absorption in the CCD through a mechanism that is ENaC-dependent.