Tilt in Place Microscopy: a Simple, Low-Cost Solution to Image Neural Responses to Body Rotations.

Animals use information about gravity and other destabilizing forces to balance and navigate through their environment. Measuring how brains respond to these forces requires considerable technical knowledge and/or financial resources. We present a simple alternative-Tilt In Place Microscopy (TIPM), a low-cost and noninvasive way to measure neural activity following rapid changes in body orientation. Here, we used TIPM to study vestibulospinal neurons in larval zebrafish during and immediately after roll tilts. Vestibulospinal neurons responded with reliable increases in activity that varied as a function of ipsilateral tilt amplitude. TIPM differentiated tonic (i.e., sustained tilt) from phasic responses, revealing coarse topography of stimulus sensitivity in the lateral vestibular nucleus. Neuronal variability across repeated sessions was minor relative to trial-to-trial variability, allowing us to use TIPM for longitudinal studies of the same neurons across two developmental time points. There, we observed global increases in response strength and systematic changes in the neural representation of stimulus direction. Our data extend classical characterization of the body tilt representation by vestibulospinal neurons and establish the utility of TIPM to study the neural basis of balance, especially in developing animals.SIGNIFICANCE STATEMENT Vestibular sensation influences everything from navigation to interoception. Here, we detail a straightforward, validated, and nearly universal approach to image how the nervous system senses and responds to body tilts. We use our new method to replicate and expand on past findings of tilt sensing by a conserved population of spinal-projecting vestibular neurons. The simplicity and broad compatibility of our approach will democratize the study of the response of the brain to destabilization, particularly across development.

Assessment of thoracic spinal cord electrophysiological activity through magnetoneurography.

OBJECTIVE: Noninvasive and detailed visualization of electrophysiological activity in the thoracic spinal cord through magnetoneurography. METHODS: In five healthy volunteers, magnetic fields around current flowing in the thoracic spinal cord after alternating unilateral and synchronized bilateral sciatic nerve stimulation were measured using a magnetoneurograph system with superconductive quantum interference device biomagnetometers. The current distribution was obtained from the magnetic data by spatial filtering and visualized by superimposing it on the X-ray image. Conduction velocity was calculated using the peak latency of the current waveforms. RESULTS: A sufficiently high magnetic signal intensity and signal-to-noise ratio were obtained in all participants after synchronized bilateral sciatic nerve stimulation. Leading and trailing components along the spinal canal and inward components flowing into the depolarization site ascended to the upper thoracic spine. Conduction velocity of the inward current in the whole thoracic spine was 42.4 m/s. CONCLUSIONS: Visualization of electrophysiological activity in the thoracic spinal cord was achieved through magnetoneurography and a new method for synchronized bilateral sciatic nerve stimulation. Magnetoneurography is expected to be a useful modality in functional assessment of thoracic myelopathy. SIGNIFICANCE: This is the first report to use magnetoneurography to noninvasively visualize electrophysiological activity in the thoracic spinal cord in detail.

Thoracic Spinal Cord Hemisection Surgery and Open-Field Locomotor Assessment in the Rat.

Spinal cord injury (SCI) causes disturbances in motor, sensory, and autonomic function below the level of the lesion. Experimental animal models are valuable tools to understand the neural mechanisms involved in locomotor recovery after SCI and to design therapies for clinical populations. There are several experimental SCI models including contusion, compression, and transection injuries that are used in a wide variety of species. A hemisection involves the unilateral transection of the spinal cord and disrupts all ascending and descending tracts on one side only. Spinal hemisection produces a highly selective and reproducible injury in comparison to contusion or compression techniques that is useful for investigating neural plasticity in spared and damaged pathways associated with functional recovery. We present a detailed step-by-step protocol for performing a thoracic hemisection at the T8 vertebral level in the rat that results in an initial paralysis of the hindlimb on the side of the lesion with graded spontaneous recovery of locomotor function over several weeks. We also provide a locomotor scoring protocol to assess functional recovery in the open-field. The locomotor assessment provides a linear recovery profile and can be performed both early and repeatedly after injury in order to accurately screen animals for appropriate time points in which to conduct more specialized behavioral testing. The hemisection technique presented can be readily adapted to other transection models and species, and the locomotor assessment can be used in a variety of SCI and other injury models to score locomotor function.

An assessment of the role of the falx cerebri and tentorium cerebelli in the cranium of the cat (Felis silvestris catus).

The falx cerebri and the tentorium cerebelli are two projections of the dura mater in the cranial cavity which ossify to varying degrees in some mammalian species. The idea that the ossification of these structures may be necessary to support the loads arising during feeding has been proposed and dismissed in the past, but never tested quantitatively. To address this, a biomechanical model of a domestic cat (Felis silvestris catus) skull was created and the material properties of the falx and tentorium were varied for a series of loading regimes incorporating the main masticatory and neck muscles during biting. Under these loading conditions, ossification of the falx cerebri does not have a significant impact on the stress in the cranial bones. In the case of the tentorium, however, a localized increase in stress was observed in the parietal and temporal bones, including the tympanic bulla, when a non-ossified tentorium was modelled. These effects were consistent across the different analyses, irrespective of loading regime. The results suggest that ossification of the tentorium cerebelli may play a minor role during feeding activities by decreasing the stress in the back of the skull.

In vivo assessment of spinal cord elasticity using shear wave ultrasound in dogs.

OBJECTIVE: Evaluation of living tissue elasticity has wide applications in disease characterization and prognosis prediction. Few previous ex vivo attempts have been made to characterize spinal cord elasticity (SCE). Recently, tissue elasticity assessment has been clinically feasible using ultrasound shear wave elastography (SWE). The current study aims to characterize SCE in healthy dogs, in vivo, utilizing SWE, and to address SCE changes during compression. METHODS: Ten Greyhound dogs (mean age 14 months; mean weight 14.3 kg) were anesthetized and tracheally intubated, with hemodynamic and neurological monitoring. A 3-level, midcervical laminectomy was performed. SCE was assessed at baseline. Next, 8- and 13-mm balloon compressions were sequentially applied ventral to the spinal cord. RESULTS: The mean SCE was 18.5 ± 7 kPa. Elasticity of the central canal, pia mater, and dura mater were 21.7 ± 9.6 kPa, 26.1 ± 14.8 kPa, and 63.2 ± 11.5 kPa, respectively. As expected, the spinal cord demonstrated less elasticity than the dura mater (p < 0.0001) and pia mater (trend toward significance p = 0.08). Notably, the 13-mm balloon compression resulted in a stiffer spinal cord than at baseline (233 ± 73 kPa versus 18.5 ± 7 kPa, p < 0.0001) and 8-mm balloon compression (233 ± 73 kPa versus 185 ± 68 kPa, p < 0.048). CONCLUSIONS: In vivo SCE evaluation using SWE is feasible and comparable to earlier reports, as demonstrated by physical sectioning of the spinal cord. The compressed spinal cord is stiffer than a free spinal cord, with a linear increase in SCE with increasing mechanical compression. Knowledge of the biomechanical properties of the spinal cord including SCE has potential implications for disease management and prognosis.

Interactions between brain and spinal cord mediate value effects in nocebo hyperalgesia.

Value information about a drug, such as the price tag, can strongly affect its therapeutic effect. We discovered that value information influences adverse treatment outcomes in humans even in the absence of an active substance. Labeling an inert treatment as expensive medication led to stronger nocebo hyperalgesia than labeling it as cheap medication. This effect was mediated by neural interactions between cortex, brainstem, and spinal cord. In particular, activity in the prefrontal cortex mediated the effect of value on nocebo hyperalgesia. Value furthermore modulated coupling between prefrontal areas, brainstem, and spinal cord, which might represent a flexible mechanism through which higher-cognitive representations, such as value, can modulate early pain processing.

Cortical and spinal assessment - a comparative study using encephalography and the nociceptive withdrawal reflex.

BACKGROUND: Standardized objective methods to assess the analgesic effects of opioids, enable identification of underlying mechanisms of drug actions in the central nervous system. Opioids may exert their effect on both cortical and spinal levels. In this study actions of morphine at both levels were investigated, followed by analysis of a possible correlation between the cortical processing and spinal transmission. METHODS: The study was conducted after a double-blinded, two-way crossover design in thirty-nine healthy participants. Each participant received 30mg morphine or placebo as oral solution in randomized order. The electroencephalogram (EEG) was recorded during rest and during immersion of the hand into ice-water. Electrical stimulation of the sole of the foot was used to elicit the nociceptive withdrawal reflex and the reflex amplitude was recorded. RESULTS: Data from thirty subjects was included in the data analysis. There was no change in the activity in resting EEG (P>0.05) after morphine administration as compared to placebo. During cold pressor stimulation, morphine significantly lowered the relative activity in the delta (1-4Hz) band (P=0.03) and increased the activity in the alpha (8-12Hz) band (P=0.001) as compared to placebo. The reflex amplitudes significantly decreased after morphine administration (P=0.047) as compared to placebo. There was no correlation between individual EEG changes during cold pressor stimulation and the decrease in the reflex amplitude after morphine administration (P>0.05). CONCLUSIONS: Cold pressor EEG and the nociceptive reflex were more sensitive to morphine analgesia than resting EEG and can be used as standardized objective methods to assess opioid effects. However, no correlation between the analgesic effect of morphine on the spinal and cortical assessments could be demonstrated.

Regeneration of Zebrafish CNS: Adult Neurogenesis.

Regeneration in the animal kingdom is one of the most fascinating problems that have allowed scientists to address many issues of fundamental importance in basic biology. However, we came to know that the regenerative capability may vary across different species. Among vertebrates, fish and amphibians are capable of regenerating a variety of complex organs through epimorphosis. Zebrafish is an excellent animal model, which can repair several organs like damaged retina, severed spinal cord, injured brain and heart, and amputated fins. The focus of the present paper is on spinal cord regeneration in adult zebrafish. We intend to discuss our current understanding of the cellular and molecular mechanism(s) that allows formation of proliferating progenitors and controls neurogenesis, which involve changes in epigenetic and transcription programs. Unlike mammals, zebrafish retains radial glia, a nonneuronal cell type in their adult central nervous system. Injury induced proliferation involves radial glia which proliferate, transcribe embryonic genes, and can give rise to new neurons. Recent technological development of exquisite molecular tools in zebrafish, such as cell ablation, lineage analysis, and novel and substantial microarray, together with advancement in stem cell biology, allowed us to investigate how progenitor cells contribute to the generation of appropriate structures and various underlying mechanisms like reprogramming.

Magnetic resonance imaging assessment of spinal cord and cauda equina motion in supine patients with spinal metastases planned for spine stereotactic body radiation therapy.

PURPOSE: To assess motion of the spinal cord and cauda equina, which are critical neural tissues (CNT), which is important when evaluating the planning organ-at-risk margin required for stereotactic body radiation therapy. METHODS AND MATERIALS: We analyzed CNT motion in 65 patients with spinal metastases (11 cervical, 39 thoracic, and 24 lumbar spinal segments) in the supine position using dynamic axial and sagittal magnetic resonance imaging (dMRI, 3T Verio, Siemens) over a 137-second interval. Motion was segregated according to physiologic cardiorespiratory oscillatory motion (characterized by the average root mean square deviation) and random bulk shifts associated with gross patient motion (characterized by the range). Displacement was evaluated in the anteroposterior (AP), lateral (LR), and superior-inferior (SI) directions by use of a correlation coefficient template matching algorithm, with quantification of random motion measure error over 3 separate trials. Statistical significance was defined according to P<.05. RESULTS: In the AP, LR, and SI directions, significant oscillatory motion was observed in 39.2%, 35.1%, and 10.8% of spinal segments, respectively, and significant bulk motions in all cases. The median oscillatory CNT motions in the AP, LR, and SI directions were 0.16 mm, 0.17 mm, and 0.44 mm, respectively, and the maximal statistically significant oscillatory motions were 0.39 mm, 0.41 mm, and 0.77 mm, respectively. The median bulk displacements in the AP, LR, and SI directions were 0.51 mm, 0.59 mm, and 0.66 mm, and the maximal statistically significant displacements were 2.21 mm, 2.87 mm, and 3.90 mm, respectively. In the AP, LR, and SI directions, bulk displacements were greater than 1.5 mm in 5.4%, 9.0%, and 14.9% of spinal segments, respectively. No significant differences in axial motion were observed according to cord level or cauda equina. CONCLUSIONS: Oscillatory CNT motion was observed to be relatively minor. Our results support the importance of controlling bulk patient motion and the practice of applying a planning organ-at-risk margin.

Control of food intake and energy expenditure by Nos1 neurons of the paraventricular hypothalamus.

The paraventricular nucleus of the hypothalamus (PVH) contains a heterogeneous cluster of Sim1-expressing cell types that comprise a major autonomic output nucleus and play critical roles in the control of food intake and energy homeostasis. The roles of specific PVH neuronal subtypes in energy balance have yet to be defined, however. The PVH contains nitric oxide synthase-1 (Nos1)-expressing (Nos1(PVH)) neurons of unknown function; these represent a subset of the larger population of Sim1-expressing PVH (Sim1(PVH)) neurons. To determine the role of Nos1(PVH) neurons in energy balance, we used Cre-dependent viral vectors to both map their efferent projections and test their functional output in mice. Here we show that Nos1(PVH) neurons project to hindbrain and spinal cord regions important for food intake and energy expenditure control. Moreover, pharmacogenetic activation of Nos1(PVH) neurons suppresses feeding to a similar extent as Sim1(PVH) neurons, and increases energy expenditure and activity. Furthermore, we found that oxytocin-expressing PVH neurons (OXT(PVH)) are a subset of Nos1(PVH) neurons. OXT(PVH) cells project to preganglionic, sympathetic neurons in the thoracic spinal cord and increase energy expenditure upon activation, though not to the same extent as Nos1(PVH) neurons; their activation fails to alter feeding, however. Thus, Nos1(PVH) neurons promote negative energy balance through changes in feeding and energy expenditure, whereas OXT(PVH) neurons regulate energy expenditure alone, suggesting a crucial role for non-OXT Nos1(PVH) neurons in feeding regulation.

Assessment of data acquisition parameters, and analysis techniques for noise reduction in spinal cord fMRI data.

PURPOSE: The purpose of this work is to characterize the noise in spinal cord functional MRI, assess current methods aimed at reducing noise, and optimize imaging parameters. METHODS: Functional MRI data were acquired at multiple echo times and the contrast-to-noise ratio (CNR) was calculated. Independently, the repetition time was systematically varied with and without parallel imaging, to maximize BOLD sensitivity and minimize type I errors. Noise in the images was characterized by examining the frequency spectrum, and investigating whether autocorrelations exist. The efficacy of several physiological noise reduction methods in both null (no stimuli) and task (thermal pain paradigm) data was also assessed. Finally, our previous normalization methods were extended. RESULTS: The echo time with the highest functional CNR at 3 Tesla is at roughly 75msec. Parallel imaging reduced the variance and the presence of autocorrelations, however the BOLD response in task data was more robust in data acquired without parallel imaging. Model-free based approaches further increased the detection of active voxels in the task data. Finally, inter-subject registration was improved. CONCLUSIONS: Results from this study provide a rigorous characterization of the properties of the noise and assessment of data acquisition and analysis methods for spinal cord and brainstem fMRI.

Outcomes of percutaneous and paddle lead implantation for spinal cord stimulation: a comparative analysis of complications, reoperation rates, and health-care costs.

OBJECTIVES: Spinal cord stimulation (SCS) is a well-established modality for the treatment of chronic pain, and can utilize percutaneous or paddle leads. While percutaneous leads are less invasive, they have been shown to have higher lead migration rates. In this study, we compared the long-term outcomes and health-care costs associated with paddle and percutaneous lead implantation. MATERIALS AND METHODS: We utilized the MarketScan data base to examine patients who underwent percutaneous or paddle lead SCS system implantation from 2000 to 2009. Outcomes including complications, reoperation rates, and health-care costs were evaluated in propensity score matched cohorts using univariate and multivariate analyses. RESULTS: The study cohort was comprised of 13,774 patients. At 90 days following the initial procedure, patients in the SCS paddle group were more likely to develop a postoperative complication than patients receiving percutaneous systems (3.4% vs. 2.2%, p = 0.0005). Two-year (6.3% vs. 3.5%, p = 0.0056) and long-term (five+ years) (22.9% vs. 8.5%, p < 0.0008) reoperation rates were significantly higher in those with percutaneous lead systems. However, long-term health-care costs were similar for those receiving paddle and percutaneous leads ($169,768 vs. $186,139, p = 0.30). CONCLUSIONS: While the implantation of paddle leads is associated with slightly higher initial postoperative complications, these leads are associated with significantly lower long-term reoperation rates. Nonetheless, long-term health-care costs are similar between paddle and percutaneous leads. Additional improvements in SCS technologies that address the shortcomings of current systems are needed to reduce the risk of reoperation due to hardware failure. Further study is required to evaluate the efficacy of newer percutaneous and paddle SCS systems and examine their comparative outcomes.

Assessment of cord dorsum potentials from caudal nerves in anesthetized clinically normal adult dogs without or during neuromuscular blockade.

OBJECTIVE: To assess the feasibility of measuring cord dorsum potentials (CDPs) in anesthetized clinically normal dogs after caudal nerve stimulation, determine the intervertebral site of maximum amplitude and best waveform of the CDP, and evaluate the effects of neuromuscular blockade. ANIMALS: 8 male and 4 female dogs (age, 1 to 5 years). PROCEDURES: Dogs were anesthetized, and CDPs were recorded via needles placed on the dorsal lamina at intervertebral spaces L1-2 through L7-S1. Caudal nerves were stimulated with monopolar electrodes inserted laterally to the level of the caudal vertebrae. Dogs were tested without and during neuromuscular blockade induced with atracurium besylate. The CDP latency and amplitude were determined from the largest amplitude tracings. RESULTS: CDPs were recorded in 11 of 12 dogs without neuromuscular blockade and in all dogs during neuromuscular blockade. The CDP was largest and most isolated at the L4-5 intervertebral space (3 dogs) or the L5-6 intervertebral space (9 dogs); this site corresponded to the segment of insertion of the first caudal nerve. Onset latencies ranged from 2.0 to 4.7 milliseconds, and there was no effect of neuromuscular blockade on latencies. Amplitudes of the CDPs were highly variable for both experimental conditions. CONCLUSIONS AND CLINICAL RELEVANCE: CDPs were recorded from all dogs tested in the study; neuromuscular blockade was not critical for successful CDP recording but reduced muscle artifact. This technique may be useful as a tool to assess the caudal nerve roots in dogs suspected of having compressive lumbosacral disease or myelopathies affecting the lumbar intumescence.

Evidence-based guideline update: intraoperative spinal monitoring with somatosensory and transcranial electrical motor evoked potentials: report of the Therapeutics and Technology Assessment Subcommittee of the American Academy of Neurology and the American Clinical Neurophysiology Society.

OBJECTIVE: To evaluate whether spinal cord intraoperative monitoring (IOM) with somatosensory and transcranial electrical motor evoked potentials (EPs) predicts adverse surgical outcomes. METHODS: A panel of experts reviewed the results of a comprehensive literature search and identified published studies relevant to the clinical question. These studies were classified according to the evidence-based methodology of the American Academy of Neurology. Objective outcomes of postoperative onset of paraparesis, paraplegia, and quadriplegia were used because no randomized or masked studies were available. RESULTS AND RECOMMENDATIONS: Four Class I and 8 Class II studies met inclusion criteria for analysis. The 4 Class I studies and 7 of the 8 Class II studies reached significance in showing that paraparesis, paraplegia, and quadriplegia occurred in the IOM patients with EP changes compared with the IOM group without EP changes. All studies were consistent in showing all occurrences of paraparesis, paraplegia, and quadriplegia in the IOM patients with EP changes, with no occurrences of paraparesis, paraplegia, and quadriplegia in patients without EP changes. In the Class I studies, 16%-40% of the IOM patients with EP changes developed postoperative-onset paraparesis, paraplegia, or quadriplegia. IOM is established as effective to predict an increased risk of the adverse outcomes of paraparesis, paraplegia, and quadriplegia in spinal surgery (4 Class I and 7 Class II studies). Surgeons and other members of the operating team should be alerted to the increased risk of severe adverse neurologic outcomes in patients with important IOM changes (Level A).

Assessment of physiological noise modelling methods for functional imaging of the spinal cord.

The spinal cord is the main pathway for information between the central and the peripheral nervous systems. Non-invasive functional MRI offers the possibility of studying spinal cord function and central sensitisation processes. However, imaging neural activity in the spinal cord is more difficult than in the brain. A significant challenge when dealing with such data is the influence of physiological noise (primarily cardiac and respiratory), and currently there is no standard approach to account for these effects. We have previously studied the various sources of physiological noise for spinal cord fMRI at 1.5T and proposed a physiological noise model (PNM) (Brooks et al., 2008). An alternative de-noising strategy, selective averaging filter (SAF), was proposed by Deckers et al. (2006). In this study we reviewed and implemented published physiological noise correction methods at higher field (3T) and aimed to find the optimal models for gradient-echo-based BOLD acquisitions. Two general techniques were compared: physiological noise model (PNM) and selective averaging filter (SAF), along with regressors designed to account for specific signal compartments and physiological processes: cerebrospinal fluid (CSF), motion correction (MC) parameters, heart rate (HR), respiration volume per time (RVT), and the associated cardiac and respiratory response functions. Functional responses were recorded from the cervical spinal cord of 18 healthy subjects in response to noxious thermal and non-noxious punctate stimulation. The various combinations of models and regressors were compared in three ways: the model fit residuals, regression model F-tests and the number of activated voxels. The PNM was found to outperform SAF in all three tests. Furthermore, inclusion of the CSF regressor was crucial as it explained a significant amount of signal variance in the cord and increased the number of active cord voxels. Whilst HR, RVT and MC explained additional signal (noise) variance, they were also found (in particular HR and RVT) to have a negative impact on the parameter estimates (of interest)--as they may be correlated with task conditions e.g. noxious thermal stimuli. Convolution with previously published cardiac and respiratory impulse response functions was not found to be beneficial. The other novel aspect of current study is the investigation of the influence of pre-whitening together with PNM regressors on spinal fMRI data. Pre-whitening was found to reduce non-white noise, which was not accounted for by physiological noise correction, and decrease false positive detection rates.

Spinal cord stimulation versus re-operation in patients with failed back surgery syndrome: an international multicenter randomized controlled trial (EVIDENCE study).

OBJECTIVE: This paper presents the protocol of the EVIDENCE study, a multicenter multinational randomized controlled trial to assess the effectiveness and cost-effectiveness of spinal cord stimulation (SCS) with rechargeable pulse generator versus re-operation through 36-month follow-up in patients with failed back surgery syndrome. STUDY DESIGN: Study subjects have neuropathic radicular leg pain exceeding or equaling any low back pain and meet specified entry criteria. One-to-one randomization is stratified by site and by one or more prior lumbosacral operations. The sample size of 132 subjects may be adjusted to between 100 and 200 subjects using a standard adaptive design statistical method with pre-defined rules. Crossover treatment is possible. Co-primary endpoints are proportion of subjects reporting ≥ 50% leg pain relief without crossover at 6 and at 24 months after SCS screening trial or re-operation. Insufficient pain relief constitutes failure of randomized treatment, as does crossover. Secondary endpoints include cost-effectiveness; relief of leg, back, and overall pain; change in disability and quality of life; and rate of crossover. We are collecting data on subject global impression of change, patient satisfaction with treatment, employment status, pain/paresthesia overlap, SCS programming, and adverse events. DISCUSSION: As the first multicenter randomized controlled trial of SCS versus re-operation and the first to use only rechargeable SCS pulse generators, the EVIDENCE study will provide up-to-date evidence on the treatment of failed back surgery syndrome.