Regular Stimulation Research Articles

Effects of inter-stimulus interval (ISI) duration on the N1 and P2 components of the auditory event-related potential

The N1 and P2 components of the event-related potential are relevant markers in the processing of auditory information, indicating the presence of several acoustic phenomena, such as pure tones or speech sounds. In addition, the expression of these components seems to be sensitive to diverse experimental variations. The main purpose of the present investigation was to explore the role of inter-stimulus interval (ISI) on the N1 and P2 responses, considering two widely used experimental paradigms: a single tone task (1000Hz sound repeated in a fixed rhythm) and an auditory oddball (80% of the stimuli were equal to the sound used in the single tone and the remaining were a 1500Hz tone). Both tasks had four different conditions, and each one tested a fixed value of ISI (600, 1000, 3000, or 6000ms). A sample of 22 participants performed these tasks, while an EEG was recorded, in order to examine the maximum amplitude of the N1 and P2 components. Analysis of the stimuli in the single tone task and the frequent tones in the oddball task revealed a similar outcome for both tasks and for both components: N1 and P2 amplitudes were enhanced in conditions with longer ISIs regardless of task. This response pattern emphasizes the dependence of both the N1 and P2 components on the ISI, especially in a scenario of repetitive and regular stimulation. The absence of task effects suggests that the ISI effect reported may depend on refractory mechanisms rather than being due to habituation effects.

Chronic deep cerebellar stimulation promotes long-term potentiation, microstructural plasticity, and reorganization of perilesional cortical representation in a rodent model.

Control over postinjury CNS plasticity is a major frontier of science that, if conquered, would open new avenues for treatment of neurological disorders. Here we investigate the functional, physiological, and structural changes in the cerebral cortex associated with chronic deep brain stimulation of the cerebellar output, a treatment approach that has been shown to improve postischemia motor recovery in a rodent model of cortical infarcts. Long-Evans rats were pretrained on the pasta-matrix retrieval task, followed by induction of focal cortical ischemia and implantation of a macroelectrode in the contralesional lateral cerebellar nucleus. Animals were assigned to one of three treatment groups pseudorandomly to balance severity of poststroke motor deficits: REGULAR stimulation, BURST stimulation, or SHAM. Treatment initiated 2 weeks post surgery and continued for 5 weeks. At the end, animals were randomly selected for perilesional intracortical microstimulation mapping and tissue sampling for Western blot analysis or contributed tissue for 3D electron microscopy. Evidence of enhanced cortical plasticity with therapeutically effective stimulation is shown, marked by greater perilesional reorganization in stimulation- treated animals versus SHAM. BURST stimulation was significantly effective for promoting distal forepaw cortical representation. Stimulation-treated animals showed a twofold increase in synaptic density compared with SHAM. In addition, treated animals demonstrated increased expression of synaptic markers of long-term potentiation and plasticity, including synaptophysin, NMDAR1, CaMKII, and PSD95. These findings provide a critical foundation of how deep cerebellar stimulation may guide plastic reparative reorganization after nonprogressive brain injury and indicate strong translational potential.

Alteration of endometrial receptivity in rats with ovarian hyperstimulation syndrome

It is unclear if the higher pregnancy rate in patients who experience ovarian hyperstimulation syndrome (OHSS) indicates that OHSS is favourable for embryo implantation, or if patients should be maintained in a hyperstimulation state in order to increase the success rate of embryo transplantation. We developed an animal model to determine the endometrial receptivity in rats with OHSS. Endometrial mRNA levels of ER, PR, HOXA10 and LIF were determined by semi-quantitative PCR and ER, PR, HOXA10, LIF and integrin αv β3 protein levels were determined by Western blotting. Development of pinopodes in the hyperstimulation group was slightly delayed, while in the regular stimulation group, development was significantly inhibited. mRNA transcription in the regular stimulation group was lower, while transcription in the hyperstimulation group was not different from controls. Protein expressions were lowest in the regular stimulation. We conclude that OHSS is associated with favourable endometrial receptivity, similar to that seen in a normal cycle, and receptivity that is increased relative to that seen with a routine stimulation protocol.

P 116. Influence of transorbital alternating current stimulation on post-traumatic neuronal morphology and death

Transorbital Alternating Current Stimulation (RTACS) has been studied in clinical trials for rehabilitation on patients with lesions in the visual system and significant beneficial effects have been demonstrated. In animal studies it appeared that RTACS may also induce neuroprotection early post lesion. To further investigate this effect we used In Vivo Confocal Neuroimaging (ICON) of the retina to follow individual cells with microscopic (cellular) resolution in live animals and by this to analyse influences of RTACS on neuronal survival and morphology. We investigated whether RTACS treatment early after traumatic axonal lesion increased the survival of Retinal Ganglion Cells (RGC) and whether the protective effect was sustainable over 2–3 weeks. Furthermore, we quantified the influence of RTACS on trauma-induced swelling and shrinkage. We also analysed neuronal activity with an indicator of cellular free calcium. As post-traumatic neuronal death develops over weeks, these experiments will indicate which pathophysiological phase is most responsive to RTACS and what may be the mechanisms underlying protection. Fluorescent markers were injected into the superior colliculus of rats and baseline ICON was conducted one week afterwards. We then performed optic nerve crush according to standard protocol. RTACS (biphasic square wave pulses; 1 ms/phase; 100–200 μA) was applied immediately after the crush and then several stimulation schedules were tested: (i) rare stimulation (2×) (ii) regular stimulation (5×; 2 weeks) (iii) regular, sustained stimulation (7×; 3 weeks). With ICON we quantified the number of surviving cells at different post-lesional time points and in study (i) and (iii) we also measured the cell size. In experiment (iii) we used a marker of intracellular free calcium (Oregon Green BAPTA) and quantified the fluorescent intensity which is an indicator of neuronal activity. Our experiments suggest that RTACS increased survival of RGC when applied directly after traumatic damage. However, we did not see sustained neuroprotection in study (i) and (ii) but a very late protective effect in experiment (iii) [Fig. 1]. This indicates that for long-term neuroprotection frequent stimulations are necessary. Morphological analyses showed that RTACS decreases the trauma-induced morphological changes (swelling/shrinkage) and that intracellular Ca concentrations are influenced after the first week in which the death rate is highest. RTACS protects neurons early after a traumatic axonal lesion and there is a significant effect on cell morphology which suggests that the RGC do react less to a traumatic challenge when they are stimulated with biphasic currents. However, the effect seems to be only sustained when the treatment is continued. In this case, also the very delayed neuronal death can be reduced. Interestingly, neuronal activity is affected after the first week of massive cell loss in surviving neurons and we hypothesize that this may be associated with post-lesional neuronal plasticity.

Evaluation of Baseline Cortisol, Endogenous ACTH, and Cortisol/ACTH Ratio to Monitor Trilostane Treatment in Dogs with Pituitary‐Dependent Hypercortisolism

The effectiveness of trilostane treatment is currently monitored by regular ACTH stimulation tests, which are time-consuming and expensive. Therefore, a monitoring system without a stimulation protocol and with less client expense would be preferable. The aim of our study was to evaluate if baseline cortisol, endogenous ACTH (ACTH) concentration or the baseline cortisol to ACTH ratio (cortisol/ACTH ratio) could replace the ACTH stimulation test. Forty trilostane-treated dogs with pituitary-dependent hypercortisolism (PDH) were included in this prospective study. A total of 148 ACTH stimulation tests and 77 ACTH concentrations and cortisol/ACTH ratios were analyzed. Control of cortisol release was classified according to cortisol concentration after ACTH administration as excessive (<1.5 μg/dL; group 1), adequate (1.5-5.4 μg/dL; group 2), or inadequate (>5.4 μg/dL; group 3). Baseline cortisol concentrations had considerable overlap between excessively, adequately, and inadequately controlled dogs. Only baseline cortisol >4.4 μg/dL (in 12% of tests) was a reliable diagnosis of inadequate control. Endogenous ACTH concentrations did not differ between groups. The overlap of the cortisol/ACTH ratio between groups was large. Correct classification was only possible if the cortisol/ACTH ratio was >15, which occurred in 4% of tests. To monitor trilostane treatment the ACTH stimulation test cannot be replaced by baseline cortisol, ACTH concentration, or the cortisol/ACTH ratio.

The role of auditory transient and deviance processing in distraction of task performance: a combined behavioral and event-related brain potential study

Distraction of goal-oriented performance by a sudden change in the auditory environment is an everyday life experience. Different types of changes can be distracting, including a sudden onset of a transient sound and a slight deviation of otherwise regular auditory background stimulation. With regard to deviance detection, it is assumed that slight changes in a continuous sequence of auditory stimuli are detected by a predictive coding mechanisms and it has been demonstrated that this mechanism is capable of distracting ongoing task performance. In contrast, it is open whether transient detection—which does not rely on predictive coding mechanisms—can trigger behavioral distraction, too. In the present study, the effect of rare auditory changes on visual task performance is tested in an auditory-visual cross-modal distraction paradigm. The rare changes are either embedded within a continuous standard stimulation (triggering deviance detection) or are presented within an otherwise silent situation (triggering transient detection). In the event-related brain potentials, deviants elicited the mismatch negativity (MMN) while transients elicited an enhanced N1 component, mirroring pre-attentive change detection in both conditions but on the basis of different neuro-cognitive processes. These sensory components are followed by attention related ERP components including the P3a and the reorienting negativity (RON). This demonstrates that both types of changes trigger switches of attention. Finally, distraction of task performance is observable, too, but the impact of deviants is higher compared to transients. These findings suggest different routes of distraction allowing for the automatic processing of a wide range of potentially relevant changes in the environment as a pre-requisite for adaptive behavior.

Detection of Near-Threshold Sounds is Independent of EEG Phase in Common Frequency Bands.

Low-frequency oscillations in the electroencephalogram (EEG) are thought to reflect periodic excitability changes of large neural networks. Consistent with this notion, detection probability of near-threshold somatosensory, visual, and auditory targets has been reported to co-vary with the phase of oscillations in the EEG. In audition, entrainment of δ-oscillations to the periodic occurrence of sounds has been suggested to function as a mechanism of attentional selection. Here, we examine in humans whether the detection of brief near-threshold sounds in quiet depends on the phase of EEG oscillations. When stimuli were presented at irregular intervals, we did not find a systematic relationship between detection probability and phase. When stimuli were presented at regular intervals (2-s), reaction times were significantly shorter and we observed phase entrainment of EEG oscillations corresponding to the frequency of stimulus presentation (0.5 Hz), revealing an adjustment of the system to the regular stimulation. The amplitude of the entrained oscillation was higher for hits than for misses, suggesting a link between entrainment and stimulus detection. However, detection was independent of phase at frequencies ≥1 Hz. Furthermore, we show that when the data are analyzed using acausal, though common, algorithms, an apparent “entrainment” of the δ-phase to presented stimuli emerges and detection probability appears to depend on δ-phase, similar to reports in the literature. We show that these effects are artifacts from phase distortion at stimulus onset by contamination with the event-related potential, which differs markedly for hits and misses. This highlights the need to carefully deal with this common problem, since otherwise it might bias and mislead this exciting field of research.

Multiple Cholinergic Signaling Pathways in Pituitary Gonadotrophs

Acetylcholine (ACh) has been established as a paracrine factor in the anterior pituitary gland, but the receptors mediating ACh action and the cell types bearing these receptors have not been identified. Our results showed that the expression of the nicotinic subunits mRNAs followed the order β2 > β1 = α9 > α4 in cultured rat pituitary cells. The expression of the subunits in immortalized LβT2 mouse gonadotrophs followed the order β2 > α4 = α1. M4 > M3 muscarinic receptor mRNA were also identified in pituitary and LβT2 cells. The treatment of cultured pituitary cells with GnRH down-regulated the expression of α9 and α4 mRNAs, without affecting the expression of M3 and M4 receptor mRNAs, and ACh did not alter the expression of GnRH receptor mRNA. We also performed double immunostaining to show the expression of β2-subunit and M4 receptor proteins in gonadotrophs. Functional nicotinic channels capable of generating an inward current, facilitation of electrical activity, and Ca(2+) influx were identified in single gonadotrophs and LβT2 cells. In both cell types, the M3 receptor-mediated, phospholipase C-dependent Ca(2+) mobilization activated an outward apamin-sensitive K(+) current and caused hyperpolarization. The activation of M4 receptors by ACh inhibited cAMP production and GnRH-induced LH release in a pertussis toxin-sensitive manner. We concluded that multiple cholinergic receptors are expressed in gonadotrophs and that the main secretory action of ACh is inhibitory through M4 receptor-mediated down-regulation of cAMP production. The expression of nicotinic receptors in vitro compensates for the lack of regular GnRH stimulation of gonadotrophs.

Systematic study of the effects of stimulus parameters and stimulus location on afterdischarges elicited by electrical stimulation in the rat

Electrical brain stimulation is used in a variety of clinical situations, including cortical mapping for epilepsy surgery, cortical stimulation therapy to terminate seizure activity in the cortex, and in deep brain stimulation therapy. However, the effects of stimulus parameters are not fully understood. In this study, we systematically tested the impact of various stimulation parameters on the generation of motor symptoms and afterdischarges (ADs). Focal electrical stimulation was delivered at subdural cortical, intracortical, and hippocampal sites in a rat model. The effects of stimulus parameter on the generation of motor symptoms and on the occurrence of ADs were examined. The effect of stimulus irregularity was tested using random or regular 50Hz stimulation through subdural electrodes. Hippocampal stimulation produced ADs at lower thresholds than neocortical stimulation. Hippocampal stimulation also produced significantly longer ADs. Both in hippocampal and cortical stimulation, when the total current was kept constant with changing pulse width, the threshold for motor symptom or AD was lowest between 50 and 100Hz and higher at both low and high frequencies. However, if the pulse width was fixed, the threshold did not increase above 100Hz and it apparently continued to decrease through 800Hz even if the difference did not reach statistical significance. There was no significant difference between random and regular stimulation. Overall, these results indicate that electrode location and several stimulus parameters including frequency, pulse width, and total electricity are important in electrical stimulation to produce motor symptoms and ADs.

Atrial arrhythmia, triggering events and conduction abnormalities in isolated murine RyR2‐P2328S hearts

RyR2 mutations are associated with catecholaminergic polymorphic tachycardia, a condition characterized by ventricular and atrial arrhythmias. The present experiments investigate the atrial electrophysiology of homozygotic murine RyR2-P2328S (RyR2(S/S)) hearts for ectopic triggering events and for conduction abnormalities that might provide a re-entrant substrate. Electrocardiograph recordings were made from regularly stimulated RyR2(S/S) and wild type (WT) hearts, perfused using a novel modified Langendorff preparation. This permitted the simultaneous use of either floating intracellular microelectrodes to measure action potential (AP) parameters, or a multielectrode array to measure epicardial conduction velocity (CV). RyR2(S/S) showed frequent sustained tachyarrhythmias, delayed afterdepolarizations and ectopic APs, increased interatrial conduction delays, reduced epicardial CVs and reduced maximum rates of AP depolarization ((dV/dt)(max)), despite similar effective refractory periods, AP durations and AP amplitudes. Effective interatrial CVs and (dV/dt)(max) values of APs following ectopic (S2) stimulation were lower than those of APs following regular stimulation and decreased with shortening S1S2 intervals. However, although RyR2(S/S) atria showed arrhythmias over a wider range of S1S2 intervals, the interatrial CV and (dV/dt)(max) of S2 APs provoking such arrhythmias were similar in RyR2(S/S) and WT. These results suggest that abnormal intracellular Ca(2+) homoeostasis produces both arrhythmic triggers and a slow-conducting arrhythmic substrate in RyR2(S/S) atria. A similar mechanism might also contribute to arrhythmogenesis in other conditions, associated with diastolic Ca(2+) release, such as atrial fibrillation.

Improved efficacy of temporally non-regular deep brain stimulation in Parkinson's disease

High frequency deep brain stimulation is an effective therapy for motor symptoms in Parkinson's disease. However, the relative clinical efficacy of regular versus non-regular temporal patterns of stimulation in Parkinson's disease remains unclear. To determine the temporal characteristics of non-regular temporal patterns of stimulation important for the treatment of Parkinson's disease, we compared the efficacy of temporally regular stimulation with four non-regular patterns of stimulation in subjects with Parkinson's disease using an alternating finger tapping task. The patterns of stimulation were also evaluated in a biophysical model of the parkinsonian basal ganglia that exhibited prominent oscillatory activity in the beta frequency range. The temporal patterns of stimulation differentially improved motor task performance. Three of the non-regular patterns of stimulation improved performance of the finger tapping task more than temporally regular stimulation. In the computational model all patterns of deep brain stimulation suppressed beta band oscillatory activity, and the degree of suppression was strongly correlated with the clinical efficacy across stimulation patterns. The three non-regular patterns of stimulation that improved motor performance over regular stimulation also suppressed beta band oscillatory activity in the computational model more effectively than regular stimulation. These data demonstrate that the temporal pattern of stimulation is an important consideration for the clinical efficacy of deep brain stimulation in Parkinson's disease. Furthermore, non-regular patterns of stimulation may ameliorate motor symptoms and suppress pathological rhythmic activity in the basal ganglia more effectively than regular stimulation. Therefore, non-regular patterns of deep brain stimulation may have useful clinical and experimental applications.

The Age‐dependence of atrial arrhythmogenicity in Scn5a+/−murine hearts reflects alterations in action potential propagation and recovery

1. In the present study, we investigated the effect of age on atrial electrophysiological properties in Scn5a(+/-) hearts used to model corresponding increases in atrial arrhythmic tendency in human Brugada syndrome. 2. Atrial action potential initiation, propagation and recovery were compared in young (3 month old) and aged (12 month old), wild-type (WT) and Scn5a(+/-) hearts. Multielectrode array recordings assessed the spatial propagation of intrinsic electrical activity in superfused atrial preparations, whereas bipolar electrogram recordings measured basic cycle lengths (BCL) in Langendorff preparations. The duration of electrogram activity (EGD) during regular and extrasystolic stimulation with programmed electrical stimulation provided EGD ratios and atrial effective refractory periods (AERP). Monophasic recordings measured action potential durations (APD). 3. Systematic statistical explorations for independent and interacting effects of age and the Scn5a(+/-) condition demonstrated that both young and aged Scn5a(+/-) mice exhibited slowed propagation of atrial excitation relative to corresponding WT mice, with the greatest effects in aged Scn5a(+/-) mice, which additionally exhibited increased intrinsic BCL. 4. Young Scn5a(+/-) mice exhibited greater EGD and EGD ratios, as well as APD/AERP ratios, suggesting increased arrhythmic tendency compared with WT mice. 5. Aged Scn5a(+/-) mice exhibited normal EGD, EGD ratios and APD compared to aged WT and young Scn5a(+/-), and increased AERP and smaller APD/AERP ratios compared with young Scn5a(+/-). 6. These electrophysiological findings indicate increased atrial arrhythmogenicity with maximal effects on both conduction and repolarization characteristics in young compared with aged Scn5a(+/-) mice.

Loss of Effector Function of Human Cytolytic T Lymphocytes Is Accompanied by Major Alterations in N- and O-Glycosylation

Most human tumors are not eliminated by the immune system, and therapeutic vaccination shows poor results, a fact that can be explained at least partially by an immunosuppressive tumor microenvironment that is abundant in galectin-3. On cytolytic T lymphocyte (CTL) clones, maintained in culture by regular stimulation, recently activated CTLs present low effector functions. However, these functions are restored after a short treatment with LacNAc. The latter, which is in agreement with the glycoprotein-galectin lattice concept involving reduced motility, poses the question why galectin-3 ligands improve effector functions. We employed ultrasensitive MALDI-TOF-MS on resting and recently activated CTL clones combined with various glycosidase digestions and GC-MS linkage analyses. Our results showed that compared with the resting CTLs, the N-glycans of the recently activated CTLs consisted of (i) larger LacNAc oligomers of which a significant portion was longer than four-units and (ii) more multi-antennary structures. Interestingly, our results showed that the poly-LacNAc appeared to be equally distributed on all available N-glycan branches and not selectively enriched on a specific branch. The above structural alterations in the recently activated CTLs are expected to increase the galectin-3-LacNAc lattices and multivalent interactions and, therefore, reduce the motility of surface glycoproteins, such as the T-cell receptor. These findings suggest that the loss of effector functions on CTLs may be linked to reduced motility of surface glycoproteins. In addition, our results showed that recently activated CTLs had a reduced abundance of NeuAcα2,6-linked N-glycans and an increased abundance of disialylated core 1 and monosialylated core 2 O-glycan structures.

The Relationship Between Conduction Velocity and Atrial Arrhythmogenicity under Conditions of Altered Ca2+ Homeostasis in RyR2-P2328S Murine Hearts

Atrial fibrillation (AF) is a significant contributor to mortality and health care expenditure; it is associated with abnormal sarcoplasmic reticulum (SR) Ca2+ release through cardiac ryanodine receptors. The initiation of AF requires an ectopic action potential (AP) triggered within a substrate of atria conducive to the formation of re-entry circuits. The present experiments explored whether increased diastolic SR Ca2+ release might contribute to atrial arrhythmogenicity in homozygotic gain-of-function RyR2-P2328S mice (RyR2s/s).Electrocardiographs and intracellular APs were recorded from Langendorff-perfused hearts at regular 8 Hz (S1) stimulation. RyR2s/s hearts showed depolarised resting membrane potentials that were associated with an increased incidence of ectopic APs and sustained tachyarrhythmias. During regular stimulation, the RyR2s/s showed reduced epicardial conduction velocities (CV), measured using a 64-electrode array, that corresponded with reduced S1 AP upstroke velocities ((dV/dt)max).When ectopic (S2) APs were imposed during S1 stimulation at progressively shorter S1S2 intervals, the inter-atrial CV and dV/dt(max) of the S2 APs decreased with each successive interval. Moderate reductions in S1S2 interval provoked sustained tachyarrhythmias in RyR2s/s hearts, whereas significantly shorter S1S2 intervals were required to provoke arrhythmias in WT hearts. Interestingly, the mean S1S2 interval required to provoke an arrhythmia in each variant resulted in similar inter-atrial CVs and dV/dt(max).These findings suggest that triggered activity is likely to lead to a sustained tachyarrhythmia in murine atria only when the CV of an ectopic AP is slow (<∼1.1 m/s). We have shown this is more likely to occur in RyR2s/s atria where increased diastolic SR Ca2+ release is associated with both increased ectopic APs and an intrinsically slow CV. We therefore suggest that abnormal Ca2+ homeostasis contributes to both the trigger and the re-entry substrate in sustained atrial tachyarrhythmias.

Tremor reduction and modeled neural activity during cycling thalamic deep brain stimulation

ObjectiveThe effectiveness of deep brain stimulation (DBS) depends on both the frequency and the temporal pattern of stimulation. We quantified responses to cycling DBS with constant frequency to determine if there was a critical on and/or off time for alleviating tremor. MethodsWe measured postural tremor in 10 subjects with thalamic DBS and quantified neuronal entropy in a network model of Vim thalamic DBS. We tested 12 combinations of cycling on/off times that maintained the same average frequency of 125Hz, four constant frequency settings, and baseline. ResultsTremor and neural firing pattern entropy decreased as the percent on time increased from 50% to 100%. Cycling with stimulation on for at least 60% of the time was as effective as regular stimulation. All cycling settings reduced the firing pattern entropy of model neurons from the no stimulation condition by regularizing pathological firing patterns, either through synaptically-mediated inhibition or axon excitation. ConclusionsThese results indicate that pauses present in cycling stimulation decreased its effectiveness in suppressing tremor, and that changes in the amount of tremor suppression were strongly correlated with changes in the firing pattern entropy of model neurons. SignificanceCycling stimulation may reduce power consumption during clinical DBS, and thereby increase the battery life of the implanted pulse generator.

Neuron-restrictive silencer factor is not required for the antiepileptic effect of the ketogenic diet

The ketogenic diet (KD) has been used as an effective antiepileptic treatment for nearly a century. Inhibition of glycolysis and increased levels of ketone bodies are both known to contribute to the antiepileptic effects of the KD. Neuron-restrictive silencer factor (NRSF), also known as RE-1 silencing transcription factor (REST), is implicated in the antiepileptic effects of the glycolytic inhibitor 2-deoxy-d-glucose (2DG). Glycolytic inhibition is a common feature of the KD and 2DG treatment, leading to the hypothesis that NRSF might also be involved in the antiepileptic effect of the KD. To test this hypothesis, the present study was designed to investigate the role of NRSF in the antiepileptic effect of 2DG, the KD, and acetone in vivo. Kindling was used as a model to test the antiepileptic effects of 2DG, the KD, and acetone on control and NRSF conditional knockout mice (NRSF-cKO; from the intercross of CamKIIα-iCre and NRSF exon 2 floxed mice). After recovery from electrode implantation, adult mice were stimulated twice a day at afterdischarge threshold (ADT) current intensity. In the 2DG- (500 mg/kg) and acetone- (10 mmol/kg) treated groups, drugs were injected intraperitoneally 20 min before each stimulus. In the 2DG group, mice were pretreated with intraperitoneal injections for 3 days in addition to the injections administered before the regular kindling stimulation. In the KD group, mice were fed the KD instead of a control diet until the end of stimulations. Compared with control mice, the antiepileptic effect of 2DG was abolished in NRSF-cKO mice, indicating that NRSF is required for the antiepileptic effect of 2DG. In the KD-fed group, kindling development was retarded in both control and NRSF-cKO mice. In the acetone-treated group, inhibition of kindling-induced epileptogenesis was observed in both control and NRSF-cKO mice, similar to the action of the KD. These findings imply that NRSF repression complex is not essential for the antiepileptic effect of the ketogenic diet.

Modified Pseudocontinent Perineal Colostomy: A Special Technique

Innovative techniques created to restore gastrointestinal perineal continuity after abdominoperineal resection in patients with anorectal cancer include pseudocontinent perineal colostomy, in which the colon is pulled to the perineum and wrapped with a sleeve of stretched colon segment to act as a new sphincter. We investigated perineal reconstruction with a modified pseudocontinent perineal colostomy technique. Prospective cohort study. Tertiary care university hospital in Egypt. Patients with T2 or T3 anorectal cancer invading the sphincter who underwent Miles abdominoperineal resection and immediate total pelvic reconstruction between 2003 and 2007. Reconstruction consisted of a vertical rectus abdominis myocutaneous flap with modified perineal colostomy pulled through the flap to add the high-pressure zone of the flap to that of the colostomy and to create a persistent new anorectal angle. Early and late complications were recorded. Functional results were evaluated at regular intervals by questionnaire, physical examination, and balloon manometry. Continence was graded according to Kirwan. Satisfaction with continence was assessed by questionnaire. A total of 14 patients (3 women) were included. Tumors were adenocarcinoma (n = 11), squamous-cell carcinoma (n = 2), and melanoma (n = 1). Complete (R0) resection was achieved in all patients without perioperative deaths, major postoperative morbidity, or conversion to permanent iliac colostomy. Early postoperative complications (perineal wound infection, flap dehiscence, and partial perineal stoma necrosis) occurred in the first 4 patients. Late complications occurred in 7 patients, with mucosal prolapse in 3, stomal stricture in 4, and tumor recurrence in 1. Fecal continence progressed consistently with time, and by the end of the first year 8 patients (57%) had complete continence (grade A), 5 (36%) were continent with minor soiling (grade C), and 1 (7%) still had major soiling (grade D). After 6 months, 9 patients (64%) were satisfied with continence; after 1 year, 13 patients (93%) were satisfied. Regular enemas were necessary during the first year to improve soiling, and 8 patients (57%) were not in need after that. At 37 months median follow-up, 8 of 9 evaluable patients (89%) were satisfied with continence (grade A) without regular enemas. This was a preliminary observational study with no control group. Total orthotopic pelvic reconstruction with autologous tissue transposition to rebuild the principle anorectal continence elements is feasible with minor complications, and is oncologically safe. This new technique offered high continence satisfaction independent of regular enemas and electrical stimulation.

Long-term Training with Electrical Stimulation Regulates Muscle Gene Expression and Bone Architecture after Spinal Cord Injury

Long-term training with electrical stimulation regulates muscle gene expression and bone architecture after SCI: Adams CM, Suneja M, McHenry CM, Littmann AL, Dudley-Javoroski S, Shields RK, Physical Therapy and Rehabilitation Science, Carver College of Medicine, University of Iowa Background: Musculoskeletal deterioration is a serious secondary complication associated with paralysis from spinal cord injury (SCI). PURPOSE: We present the long-term effects of regular electrical stimulation training on muscle gene regulation and bone architecture after SCI. METHODS: Individuals with complete paralysis from SCI trained the calf muscle (soleus) or the thigh muscle (quadricep) during novel resistive tasks. The electrical stimulation protocol induced 2-4 bouts of 60-120 isometric contractions at a 15-20 Hz stimulation. After at least 2 years of unilateral training, we obtained muscle biopsies from the calf (soleus) and thigh muscles (vastus lateralis) of (trained and untrained) subjects with SCI. We then used Affymetrix human exon 1.0 arrays to identify mRNAs that were ≥ 1.5 fold increased or decreased in the atrophied untrained muscles. We next examined the trabecular bone using mDCT to quantify the sites and quality of the trabecular lattice. RESULTS: Out of >17,000 transcripts examined, in the untrained limb 51 mRNAs were increased including mRNAs encoding fast-twitch isoforms of myosin and troponin, as well as the mRNA encoding myostatin. Fifty-four mRNAs were decreased, including mRNAs encoding slow-twitch isoforms of myosin and troponin, 11 enzymes for oxidative phosphorylation, as well as PGC1α, a transcriptional co-activator that promotes the slow twitch, oxidative phenotype and inhibits FoxO3-mediated muscle atrophy. Training increased the mRNAs encoding slow-twitch isoforms and isoforms associated with glucose metabolism. The bone trabecular lattice was denser with distinct rods and plates within the trained femur and tibia. CONCLUSIONS: These findings support that thirty minutes/day of electrical stimulation stress induced significant muscle molecular adaptations and prevented the deterioration of bone after SCI. Funded by Nielsen Foundation and National Institutes of Health (R01-NR-01028505; R01-HD-04583233; R01HD062507).

Ventricular rate adaptation: A novel, rapid, cellular-based in-vitro assay to identify proarrhythmic and torsadogenic compounds

IntroductionDelayed cardiac repolarization is an established risk factor for proarrhythmia and Torsades-de-Pointes (TdeP) that is typically measured in vitro during slow, regular stimulation. We have developed an alternative, novel, and rapid cellular-based approach for predicting drug-induced proarrhythmia that detects changes in electrical refractoriness based on mechanical responses (measured optically) during increasingly rapid trains of stimulation interspersed with pauses (mimicking the clinically observed short–long–short (SLS) stimulation sequence associated with the TdeP initiation). MethodsAcutely isolated rabbit ventricular myocytes were superfused and electrically stimulated using an accelerating pacing protocol (APP) consisting of 12 consecutive pacing segments (10 beats per segment) with incrementally faster cycle lengths; trains were separated by pauses to identify loss of stimulus capture as well as to mimic clinically observed SLS sequences. Drug effects were evaluated based on a myocyte's ability to contract during progressively faster pacing segments (rate-adaptation); the earliest rate during which the myocyte fails to respond (longest cycle length with incomplete capture (CLIC)) was used to quantify electrophysiologic effects. ResultsTorsadogenic drugs known to delay repolarization during slow stimulation prolonged CLIC and dramatically limited the ability to respond to progressively rapid stimulation. The recognized proarrhythmic compounds E-4031, cisapride, grepafloxacin, and haloperidol rapidly prolonged CLIC at and above therapeutic concentrations in a concentration-dependent manner, while negative controls (captopril, indomethacin, and loratidine) do not affect rate-adaptation. DiscussionVentricular rate adaptation represents a novel approach for rapidly detecting drugs with torsadogenic risk using rapid rhythms that are typically not employed when evaluating proarrhythmic risk. This method is well suited for detecting and avoiding potential cardiac liabilities early in drug discovery (“frontloading”) prior to final selection of candidate drugs.

In-Line-Water-Separation Prototype Development and Testing

This article, written by Assistant Technology Editor Karen Bybee, contains highlights of paper SPE 137967, ’Inline Water Separation (IWS) Field Prototype Development and Testing,’ by J.J. Xiao, SPE, and Ramsey White, Saudi Aramco, and Shoubo Wang and Luis Gomez, Multiphase Systems Integration, originally prepared for the 2010 SPE Abu Dhabi International Petroleum Exhibition and Conference, Abu Dhabi, UAE, 1-4 November. The paper has not been peer reviewed. A compact multiphase in-line-water-separation (IWS) field prototype system has been developed and tested. The system consists of a gas/liquid cylindrical cyclone (GLCC), a liquid/liquid pipe separator (LLPS), a liquid/liquid cylindrical cyclone (LLCC), and two-stage liquid/liquid hydrocyclones (LLHCs). The function of the system is to separate a significant portion of the produced water from a production stream while the remaining production fluids are sent to existing processing facilities. Introduction As oil fields mature, more water is produced with the oil and gas production. This is especially true when water injection is used to maintain reservoir pressure and increase oil recovery. A common production-water-handling strategy is centralized processing. In this scheme, production from individual wells is gathered to manifolds through flowlines and is transported to the gas-/oil-separation plant (GOSP) through large-diameter trunklines. At the GOSP, separation and processing of oil, gas, and water take place with large conventional vessels. The processed water is either disposed of at nearby water-disposal wells or sent back to the field for water reinjection into the reservoir. With increasing water production, several issues can develop: Water volume exceeds water-handling capacity of the GOSP, which can cause processed water to be off specification. The off-specification water in turn can lead to a decline in water injectivity, which may require regular stimulation treatments. Upgrade or expansion of GOSP water-handling capacity with conventional bulky separation systems can be costly. Existing gathering systems can become a bottleneck to oil production. To maintain oil production, the total fluid produced will increase as the water cut increases, and the gathering-system capacity will become a limiting factor. Laying extra flowlines or trunklines can be expensive, especially offshore. Wells can cease to produce because of increasing system backpressure as a result of the increasing water volume being transported through the gathering systems. Higher water production leads to higher chemical use (demulsifers and corrosion inhibitors) and, hence, higher processing cost. In addition, energy consumption will increase to transport large quantities of water from the field to the GOSP and then from the GOSP back for reinjection into the wells in the field. A number of methods are used to reduce the quantity of water produced to the surface. These methods include restricted production, cyclic production, mechanical or chemical water shutoff, and short-radius horizontal drilling. For new wells, the use of production equalizers and smart completions also can be viewed broadly as a way to control water production. The next-best place to handle water production is in the wellbore. However, downhole oil/water separation and reinjection still remain a challenge even after 20 years of industry research and development.