Journal of Neuroscience Methods

Does combing the scalp reduce scalp electrode impedances?

Publication Date: 2010 Mar 4 PMID: 20211649
Authors: Mahajan, Y. - McArthur, G.
Journal: J Neurosci Methods

Electrical activity from the human brain can be recorded via electrodes on the scalp. It is important to reduce the impedance of each electrode to minimize unwanted noise in the recording. Electrode impedance can be improved by abrading the skin to remove dead skin cells. In this experiment, we tested if abrading the skin by combing the scalp leads to a significant reduction in electrode impedance. We compared the mean electrode impedance values of 20 subjects whose scalps were combed prior to electrode cap placement, with 20 subjects whose scalps were not combed. Combing significantly reduced the impedances at central, right, and left areas of the scalp. This finding supports the use of scalp combing to reduce the time and subject discomfort that can be associated with placing scalp electrodes. This is particularly important for experiments testing children.

post to: CiteULike

Ultrasound Imaging of Regenerating Rat Sciatic Nerves In Situ.

Publication Date: 2010 Mar 4 PMID: 20211648
Authors: Kuffler, D. P.
Journal: J Neurosci Methods

Following clinical or veterinary peripheral nerve trauma, it is critical to localize the site of nerve injury, determine its type, whether a crush, maceration or transection, which will indicate whether and where surgical intervention is required, and subsequently to follow the process of axon regeneration. Typical surface ultrasound probes provide resolution of more than 1mm, sufficient detail for clinically relevant data from tissue in situ, such as heart valves, organs and fetal development. Higher resolution ultrasound nerve imaging yields data to the fascicular level and allows the following of the anatomical course of a nerve, but does not allow imaging of single axons or even groups of axons, which is required to study the process of axon regeneration, neurological recovery and other important clinical and basic science questions. More significant data could be acquired with even higher frequency, and therefore higher resolution, ultrasound imaging. The present study, using a rat sciatic nerve lesion model, was performed to determine whether a new ultrasound imaging device with 30mum resolution would allow imaging of nerve anatomy and regenerating axons, and whether the data collected from a nerve in situ was the same as when the nerve was surgically exposed. Although the increased ultrasound resolution provided good anatomical detail on the location and type of nerve damage was nearly identical for nerves in situ and when exposed, the resolution was insufficient for imaging regenerating axons. Thus, an even higher resolution ultrasound device is required to allow noninvasive imaging of axons in situ.

post to: CiteULike

Prion infection of differentiated Neurospheres.

Publication Date: 2010 Mar 2 PMID: 20206206
Authors: Herva, M. E. - Relano-Gines, A. - Villa, A. - Torres, J. M.
Journal: J Neurosci Methods

Until now only a few cell lines have been proved able to propagate prions and only limited prion strains have been replicated in cell models. Neurosphere lines isolated from the brains of mice at embryonic day 14 grow as aggregates and contain CNS stem cells. Others authors have previously reported that cultured neurospheres expressing cellular prion protein (PrP(C)) can be infected with prions. As potential neural progenitors the neurosphere cultures are supposed to differentiate into neurons and astrocytes which represent the main cell types infected by prions in vivo. Here we study the ability of undifferentiated and differentiated neurospheres to replicate several prion strains. Neurosphere cultures were isolated from 129/ola, FVB, Prnp(0/0) and Tga20 mice, which overexpress murine PrP. We were not able to detect PrP(res) accumulation in dividing neurosphere cultures after prion exposure to two different mouse adapted scrapie inocula (RML and 22L). In contrast, with differentiated neurosphere cultures expressing PrP(C) (129/ola, FVB and Tga20) a successful PrP(Res) amplification was observed in very short time experiments when infected with the same inocula, implying that cell differentiation improve prion replication in these cultured cells. The mouse BSE adapted inocula (301C) was not amplified in these neurosphere cultures neither before nor after differentiation, suggesting that these cell cultures showed a differential prion strain susceptibility. These results suggest that differentiated neurosphere cultures can complement prion bioassays in mouse models.

post to: CiteULike

Topology-corrected segmentation and local intensity estimates for improved partial volume classification of brain cortex in MRI.

Publication Date: 2010 Mar 1 PMID: 20193712
Authors: Rueda, A. - Acosta, O. - Couprie, M. - Bourgeat, P. - Fripp, J. - Dowson, N. - Romero, E. - Salvado, O.
Journal: J Neurosci Methods

In magnetic resonance imaging (MRI), accuracy and precision with which brain structures may be quantified are frequently affected by the partial volume (PV) effect. PV is due to the limited spatial resolution of MRI compared to the size of anatomical structures. Accurate classification of mixed voxels and correct estimation of the proportion of each pure tissue (fractional content) may help to increase the precision of cortical thickness estimation in regions where this measure is particularly difficult, such as deep sulci. The contribution of this work is twofold: on the one hand, we propose a new method to label voxels and compute tissue fractional content, integrating a mechanism for detecting sulci with topology preserving operators. On the other hand, we improve the computation of the fractional content of mixed voxels using local estimation of pure tissue intensity means. Accuracy and precision were assessed using simulated and real MR data and comparison with other existing approaches demonstrated the benefits of our method. Significant improvements in gray matter (GM) classification and cortical thickness estimation were brought by the topology correction. The fractional content root mean squared error diminished by 6.3% (p<0.01) on simulated data. The reproducibility error decreased by 8.8% (p<0.001) and the Jaccard similarity measure increased by 3.5% on real data. Furthermore, compared with manually guided expert segmentations, the similarity measure was improved by 12.0% (p<0.001). Thickness estimation with the proposed method showed a higher reproducibility compared with the measure performed after partial volume classification using other methods.

post to: CiteULike

Phase-dependent audiometry with low-frequency masking revisited.

Publication Date: 2010 Feb 19 PMID: 20171987
Authors: Rahne, T. - Rasinski, C. - Neumann, K.
Journal: J Neurosci Methods

Low-frequency masking is a psychoacoustical phenomenon, describing the modulation of a high-frequency probe tone burst by a low-frequency masker tone. The probe tone threshold is increased, if the probe tone is presented at a low-frequency phases around 90 degrees and 270 degrees . At these phases, the low-frequency masker tone induces a displacement of the basilar membrane of the inner ear which modulates the sensitivity of the inner hair cells. Measuring the modulation depth is partially applied in clinical routine to diagnose the endolymphatic hydrops. Although the modulation depth differs between normal ears and those which reveal an endolymphatic hydrops, the significance of these tests seems debatable. Here, we describe a new experimental setup, completely consisting of commercially available devices. Further, a user interface was developed to enable the application in the clinical routine. The experimental setup was approved with ten normal hearing listeners. All reveal a modulation of the probe stimulus threshold by different phases of the low-frequency masker stimulus. With this experimental setup, custom-made modifications of the essential parameters are feasible. This would be a contribution to solve open questions on the clinical relevance of the low-frequency masking phenomenon.

post to: CiteULike

Corticosterone urinalysis and nicotinic receptor modulation in rats.

Publication Date: 2010 Feb 19 PMID: 20171986
Authors: Loomis, S. - Gilmour, G.
Journal: J Neurosci Methods

A routine method of measuring circulating corticosterone (CORT) levels in rats involves sampling of plasma from cannulated animals. However, being somewhat invasive, this method can potentially be confounded by its inherently stressful nature. This study investigated the feasibility of measuring corticosterone using a non-invasive sampling method from voided urine of male rats. Reliability was assessed pharmacologically with nicotinic compounds previously demonstrated to modulate plasma glucocorticoid levels. Nicotine (0.1-1mg/kgsc) dose-dependently increased corticosterone levels in rat urine at 30-70min following administration. The short-lived nature of this elevation was confirmed as CORT levels measured 6 and 24h later were shown to have returned to basal levels. Both basal and nicotine-induced (0.5mg/kg sc) elevations in urinary CORT were consistent between groups of animals with weights ranging from 200 to 400g. The magnitude of urinary CORT elevation induced by nicotine (0.5mg/kgsc) was found to be similar to that induced by a forced swim stressor in male Lister Hooded, Sprague Dawley and Fischer 344 rats. The pharmacological specificity of this effect was confirmed as the nicotinic acetylcholine receptor (nAChR) antagonist mecamylamine (0.05-0.5mg/kg sc) dose-dependently reversed the effects of nicotine (0.5mg/kg sc) on urinary CORT. Finally, the alpha(4)beta(2)-subunit preferring agonist TC-2559 induced a dose-dependent increase in CORT, whereas alpha(7)- and beta(4)-subunit preferring ligands had no effect, suggestive of the potential for differential involvement of nicotinic receptor subtypes in the mediation of this response. In conclusion, urinary corticosterone sampling in rats represents a robust assay sensitive to experimental manipulations of both pharmacological and behavioural relevances.

post to: CiteULike

From foraging to operant conditioning: A new computer-controlled Skinner box to study free-flying nectar gathering behavior in bees.

Publication Date: 2010 Feb 19 PMID: 20171985
Authors: Sokolowski, M. B. - Abramson, C. I.
Journal: J Neurosci Methods

The experimental study of nectar foraging behavior in free-flying bees requires the use of automated devices to control solution delivery and measure dependent variables associated with nectar gathering. We describe a new computer-controlled artificial flower and provide calibration data to measure the precision of the apparatus. Our device is similar to a "Skinner box" and we present data of an experiment where various amounts of a 50% sugar solution are presented randomly to individual bees. These data show large individual variations among subjects across several dependent variables. Finally, we discuss possible applications of our device to problems in behavioral sciences.

post to: CiteULike

Imaging fluid flow and cilia beating pattern in Xenopus brain ventricles.

Publication Date: 2010 Feb 19 PMID: 20171984
Authors: Miskevich, F.
Journal: J Neurosci Methods

Brain development and health depends upon the efficient movement of the cerebrospinal fluid inside of brain ventricles. When disrupted either through mutation, disease, or physiological damage, brain function becomes significantly impaired. Here I present a simple method of following cerebrospinal fluid circulation in Xenopus tadpoles using fluorescent microspheres which can be applied to imaging fluid circulation in any transparent embryo. In particular, cilia may be labeled with these microspheres to study their dynamics and movement patterns in vivo while simultaneously measuring bulk fluid flow. This technique will facilitate the analysis of fluid dynamics in developing embryos and aid in understanding the regulation of cilia dependent fluid flow in vivo.

post to: CiteULike

The conundrum of sensitization when recording from nociceptors.

Publication Date: 2010 Feb 18 PMID: 20171245
Authors: Bove, G. M. - Dilley, A.
Journal: J Neurosci Methods

Nociceptors are sensory neurons that detect harmful, or potentially harmful, stimuli, and can become sensitized following injury or repetitive stimulation. When sensitized, nociceptors often exhibit activity in the absence of apparent or additional stimulation, called ongoing (or spontaneous) activity (OA). In this report, we provide evidence that OA in nociceptors can be caused by the stimuli typically used to identify and characterize the neuron, which must by definition be noxious and therefore potentially sensitizing. Such OA caused by the experimental methodology can confound interpretation. In our nerve inflammation model, OA can potentially arise from multiple sites, including the lesion site and the receptive field. We provide evidence that the OA rate recorded during these experiments may be related to the site and cause of OA generation. We suggest that there are two types of OA, characterized by their rates. Very slow rates of ongoing activity (<0.2Hz) are likely to arise from the receptive field and may indicate sensitization during the experiment. Faster rates are likely to arise from the nerve trunk, i.e. the neuritis, or the neuronal cell body. Without appropriate methodological consideration, interpretations of results from such studies of nociceptor function may be methodologically confounded.

post to: CiteULike

Performance of juvenile baboons on neuropsychological tests assessing associative learning, motivation and attention.

Publication Date: 2010 Feb 17 PMID: 20170676
Authors: Zurcher, N. R. - Rodriguez, J. S. - Jenkins, S. L. - Keenan, K. - Bartlett, T. Q. - McDonald, T. J. - Nathanielsz, P. W. - Nijland, M. J.
Journal: J Neurosci Methods

The CANTAB (Cambridge Neuropsychological Test Automated Battery), a system developed for human neuropsychological testing, has previously been used to assess cognitive function in two species of nonhuman primates, common marmoset monkeys and rhesus macaques. We describe the application of the system to the juvenile baboon, a nonhuman primate species offering specific investigative advantages. Juvenile baboons were trained and tested on a progressive ratio task to assess motivation, simple discrimination and simple reversal tasks to assess associative learning, and intra- and extra-dimensional set-shifting tasks to assess selective attention and attentional set-shifting, respectively. Study subjects were 8 juvenile baboons (Papio sp.), 4 females and 4 males aged 3.0+/-0.1 (mean+SEM) years and weight 8.2+/-0.4kg. All baboons were easily trained, readily learned the neuropsychological tests and exhibited a stable performance. Applying a method such as the CANTAB has significant implications for expanding on the translational utility of the baboon in studies of neurodevelopment.

post to: CiteULike

Multiple ERbeta antisera label in ERbeta knockout and null mouse tissues.

Publication Date: 2010 Feb 16 PMID: 20170675
Authors: Snyder, M. A. - Smejkalova, T. - Forlano, P. M. - Woolley, C. S.
Journal: J Neurosci Methods

In the process of characterizing a custom-made affinity-purified antiserum for estrogen receptor beta (ERbeta), ck5912, we used a number of common tests for specificity of ck5912 along with that of 8 commercially available ERbeta antisera: Affinity Bioreagents PA1-310B, Invitrogen D7N, Upstate 06-629, Santa Cruz H150, Y19, L20, 1531, and Abcam 9.88. We tested their recognition of recombinant ERbeta (rERbeta) versus rERalpha, ERbeta versus ERalpha transfected into cell lines, as well as labeling in wildtype (WT) versus estrogen receptor beta knockout (betaERKO) and null (ERbeta(ST)(L-/L-)) mouse ovary, hypothalamus, and hippocampus. To our surprise, we found that while most of these antisera passed some tests, giving the initial impression of specificity, western blot analysis showed that all of them recognized apparently identical protein bands in WT, betaERKO and ERbeta(ST)(L-/L-) tissues. We share these results with the goal of helping other researchers avoid pitfalls in interpretation that could come from use of these ERbeta antisera.

post to: CiteULike

A near-infrared calibration method suitable for quantification of broadband data in humans.

Publication Date: 2010 Feb 13 PMID: 20156483
Authors: Zhang, Q. - Srinivasan, S. - Wu, Y. - Natah, S. - Dunn, J. F.
Journal: J Neurosci Methods

Broadband near-infrared spectroscopy (bNIRS) is a powerful non-invasive technique for the measurement of hemoglobin. bNIRS systems are relatively simple to construct compared with many near-infrared instruments since they operate on the principle of continuous wave. The advantage of the broadband method is the capacity to model the spectra and to use "the second differential method" to quantify deoxyhemoglobin (HHb). An "anoxia pulse" method can be applied to quantify total haemoglobin (tHb) and tissue oxygen saturation (S(t)O(2)). A disadvantage is that this calibration method is not suitable for application in humans. In this study, we compared the "anoxia pulse" method with "graded hypoxia" method, which can be applied for human studies, to quantify tHb and S(t)O(2). The values obtained with the two methods were respectively (tHb=47.8+/-2.8 and 49.4+/-7.7muM, mean+/-S.D., n=8) and (S(t)O(2)=72.8+/-3.7% and 73.2+/-5.7%, mean+/-S.D., n=8). There was no significant difference (p<0.05) between the two methods, indicating that the graded hypoxia method could be used for quantification of tHb and S(t)O(2) in human subjects.

post to: CiteULike

Optimizing Drosophila olfactory learning with a semi-automated training device.

Publication Date: 2010 Feb 12 PMID: 20153774
Authors: Murakami, S. - Dan, C. - Zagaeski, B. - Maeyama, Y. - Kunes, S. - Tabata, T.
Journal: J Neurosci Methods

Drosophila olfactory aversive conditioning has served as a powerful model system with which to elucidate the molecular and neuronal mechanisms underlying memory formation. In the typical protocol, flies are exposed to a constant odor stream while receiving a pulsed electric shock in the conditioning tube of a manual apparatus. We have devised a simple, low-cost semi-automated conditioning apparatus that computationally controls the delivery of odor and shock. A semiconductor-based odor sensor is employed to monitor the change of odor concentration in the training tube. The system thus allows electric shocks to be precisely matched with odor concentration in the training tube. We found that short-term memory performance was improved with a pulsed odor flow protocol, in which odor is presented in short pulses, each paired with electric shock, rather than as a constant flow. The effect of pulsed odor flow might be ascribed to the phenomenon of 'conditioned approach', where approach toward an odor is induced when the electric shock is presented before odor pulse ends. Our data shows that the system is applicable to the study of olfactory memory formation and to the examination of conditioning parameters at a level of detail not practical with a manual apparatus.

post to: CiteULike

An organotypic culture model to study nigro-striatal degeneration.

Publication Date: 2010 Feb 11 PMID: 20153372
Authors: Cavaliere, F. - Vicente, E. S. - Matute, C.
Journal: J Neurosci Methods

Functional and reliable in vitro models of Parkinson's disease (PD) are valuable for studying mechanisms of dopaminergic degeneration before proceeding to animal testing. At present, all in vitro models involve substitute cell types and thus their direct relevance to PD is questionable. Here, we describe an organotypic culture model which conserves the 3D architecture of the nigro-striatal pathway, together with the subventricular zone and cerebral cortex, and recapitulates a specific pattern of dopaminergic degeneration which is the principal hallmark of PD. The organotypic culture is kept in vitro for up to 12 days and dopaminergic degeneration is induced by the simple cutting of dopaminergic fibers. This organotypic model represents a rapid and useful method (30min/pup for preparation and up to 12 days of cultivation) to investigate in vitro the mechanisms underlying neuronal death and protection, as well as neurogenesis and repair after nigro-striatal neurodegeneration.

post to: CiteULike

Improved signal processing approaches in an offline simulation of a hybrid brain-computer interface.

Publication Date: 2010 Feb 11 PMID: 20153371
Authors: Brunner, C. - Allison, B. Z. - Krusienski, D. J. - Kaiser, V. - Muller-Putz, G. R. - Pfurtscheller, G. - Neuper, C.
Journal: J Neurosci Methods

In a conventional brain-computer interface (BCI) system, users perform mental tasks that yield specific patterns of brain activity. A pattern recognition system determines which brain activity pattern a user is producing and thereby infers the user's mental task, allowing users to send messages or commands through brain activity alone. Unfortunately, despite extensive research to improve classification accuracy, BCIs almost always exhibit errors, which are sometimes so severe that effective communication is impossible. We recently introduced a new idea to improve accuracy, especially for users with poor performance. In an offline simulation of a "hybrid" BCI, subjects performed two mental tasks independently and then simultaneously. This hybrid BCI could use two different types of brain signals common in BCIs - event-related desynchronization (ERD) and steady-state evoked potentials (SSEPs). This study suggested that such a hybrid BCI is feasible. Here, we re-analyzed the data from our initial study. We explored eight different signal processing methods that aimed to improve classification and further assess both the causes and the extent of the benefits of the hybrid condition. Most analyses showed that the improved methods described here yielded a statistically significant improvement over our initial study. Some of these improvements could be relevant to conventional BCIs as well. Moreover, the number of illiterates could be reduced with the hybrid condition. Results are also discussed in terms of dual task interference and relevance to protocol design in hybrid BCIs.

post to: CiteULike

A bio-friendly and economical technique for chronic implantation of multiple microelectrode arrays.

Publication Date: 2010 Feb 11 PMID: 20153370
Authors: Chhatbar, P. Y. - von Kraus, L. M. - Semework, M. - Francis, J. T.
Journal: J Neurosci Methods

Many neurophysiological experiments on rodents and non-human primates involve the implantation of more than one multi-electrode array to record from many regions of the brain. So called 'floating' microelectrode arrays are implanted in cortical regions of interest and are coupled via a flexible cable to their connectors which are fixed to the skull by a cement cap or a titanium pedestal, such as the Cereport system, which has been approved for human use. The use of bone cement has several disadvantages including the creation of infection prone areas at the interface with the skull and surrounding skin. Alternatively, the more biocompatible Cereport has a limited carrying capacity and is far more expensive. In this paper, we describe a new implantation technique, which combines the biocompatibility of titanium, a high carrying capacity with a minimal skull footprint, and a decreased chance of infection, all in a relatively inexpensive package. This technique utilizes an in-house fabricated 'Nesting Platform' (NP), mounted on a titanium headpost to hold multiple connectors above the skin, making the headpost the only transcutaneous object. The use of delrin, a durable, lightweight and easily machinable material, allows easy customization of the NP for a wide variety of floating electrodes and their connectors. The ultimate result is a longer survival time with superior neural recordings that can potentially last longer than with traditional implantation techniques.

post to: CiteULike

Microsurgical approach to the spinal canal in rats.

Publication Date: 2010 Feb 11 PMID: 20153369
Authors: Gierthmuehlen, M. - Freiman, T. - V Elverfeldt, D. - Kaminsky, J.
Journal: J Neurosci Methods

The spinal cord of the rat has become a widely used model for biodynamic, pharmaceutical and neurological experiments. However, no standard procedure to approach the spinal cord in rats has been published in detail. We present a description of a dorsal approach to the spine, spinal canal and myelon of the rat. This approach provides sufficient exposure of the neural structures to perform extended microsurgery at the spinal nerve-roots, the lateral and dorsal myelon and vertebral structures under a surgical microscope. Perioperative management, anaesthesia and anatomical landmarks are discussed and common pitfalls are described.

post to: CiteULike

Multiple serial picture presentation with millisecond resolution using a three-way LC-shutter-tachistoscope.

Publication Date: 2010 Mar 30 PMID: 20122963
Authors: Fischmeister, F. P. - Leodolter, U. - Windischberger, C. - Kasess, C. H. - Schopf, V. - Moser, E. - Bauer, H.
Journal: J Neurosci Methods

Throughout recent years there has been an increasing interest in studying unconscious visual processes. Such conditions of unawareness are typically achieved by either a sufficient reduction of the stimulus presentation time or visual masking. However, there are growing concerns about the reliability of the presentation devices used. As all these devices show great variability in presentation parameters, the processing of visual stimuli becomes dependent on the display-device, e.g. minimal changes in the physical stimulus properties may have an enormous impact on stimulus processing by the sensory system and on the actual experience of the stimulus. Here we present a custom-built three-way LC-shutter-tachistoscope which allows experimental setups with both, precise and reliable stimulus delivery, and millisecond resolution. This tachistoscope consists of three LCD-projectors equipped with zoom lenses to enable stimulus presentation via a built-in mirror-system onto a back projection screen from an adjacent room. Two high-speed liquid crystal shutters are mounted serially in front of each projector to control the stimulus duration. To verify the intended properties empirically, different sequences of presentation times were performed while changes in optical power were measured using a photoreceiver. The obtained results demonstrate that interfering variabilities in stimulus parameters and stimulus rendering are markedly reduced. Together with the possibility to collect external signals and to send trigger-signals to other devices, this tachistoscope represents a highly flexible and easy to set up research tool not only for the study of unconscious processing in the brain but for vision research in general.

post to: CiteULike

Gain of the human dura in vivo and its effects on invasive brain signal feature detection.

Publication Date: 2010 Mar 30 PMID: 20109492
Authors: Torres Valderrama, A. - Oostenveld, R. - Vansteensel, M. J. - Huiskamp, G. M. - Ramsey, N. F.
Journal: J Neurosci Methods

Invasive brain signal recordings generally rely on bioelectrodes implanted on the cortex underneath the dura. Subdural recordings have strong advantages in terms of bandwidth, spatial resolution and signal quality. However, subdural electrodes also have the drawback of compromising the long-term stability of such implants and heighten the risk of infection. Epidurally implanted electrodes might provide a viable alternative to subdural electrodes, offering a compromise between signal quality and invasiveness. Determining the feasibility of epidural electrode implantation for e.g., clinical research, brain-computer interfacing (BCI) and cognitive experiments, requires the characterization of the electrical properties of the dura, and its effect on signal feature detection. In this paper we report measurements of brain signal attenuation by the human dura in vivo. In addition, we use signal detection theory to study how the presence of the dura between the sources and the recording electrodes affects signal power features in motor BCI experiments. For noise levels typical of clinical brain signal recording equipment, we observed no detrimental effects on signal feature detection due to the dura. Subdural recordings were found to be more robust with respect to increased instrumentation noise level as compared to their epidural counterpart nonetheless. Our findings suggest that epidural electrode implantation is a viable alternative to subdural implants from the feature detection viewpoint.

post to: CiteULike

Measurement of anti-Abeta1-42 antibodies in intravenous immunoglobulin with indirect ELISA: The problem of nonspecific binding.

Publication Date: 2010 Mar 30 PMID: 20097229
Authors: Klaver, A. C. - Patrias, L. M. - Coffey, M. P. - Finke, J. M. - Loeffler, D. A.
Journal: J Neurosci Methods

Improvement in cognitive scores in patients with Alzheimer's disease (AD) has been reported in two trials in which intravenous immunoglobulin (IvIg) preparations were administered. IvIg's benefits in AD patients have been suggested to be due to antibodies to amyloid-beta (Abeta). Our previous study using indirect enzyme-linked immunosorbent assay (ELISA) indicated that much of IvIg's apparent binding to Abeta1-42 is nonspecific; it is detectable even when IvIg is incubated on "specificity controls" (bovine serum albumin [BSA] and Abeta reverse sequence Abeta42-1) rather than Abeta1-42. The objective of this study was to evaluate procedures that might reduce this nonspecific binding. The IvIg preparation used was Gamunex (Talecris Biotherapeutics). Multiple blocking agents were evaluated, but even the most effective blocker only reduced nonspecific binding by 48%. Dissociating Gamunex's antibody-antigen complexes had no effect on specific binding when Abeta42-1 was used as the specificity control, although it increased this binding when BSA was the specificity control. Decreasing Gamunex's dilution from 1:1,500 to 1:500 resulted in a slight (7.4%) but significant (p=0.027) increase in specific binding. Using a sandwich ELISA to measure Gamunex's anti-Abeta antibodies resulted in even less specific binding to Abeta1-42 than with the indirect ELISA. Despite Gamunex's low percentage of specific binding to Abeta1-42, it inhibited Abeta oligomer formation. We conclude that, when anti-Abeta antibodies in IvIg are measured by indirect ELISA, extensive nonspecific binding occurs despite procedures taken to prevent it. This must be subtracted from total binding to accurately measure specific anti-Abeta antibody concentrations.

post to: CiteULike

A new method to isolate microglia from adult mice and culture them for an extended period of time.

Publication Date: 2010 Mar 30 PMID: 20097228
Authors: Moussaud, S. - Draheim, H. J.
Journal: J Neurosci Methods

As the major immuno-competent cells of the brain, microglia are highly implicated in neuro-protection as well as in neurodegeneration. Therefore, they are of key interest for research on numerous CNS diseases. Currently, to model inflammation in the brain, microglial cell lines or primary microglia prepared from embryonic or neo-natal rodents are widely used. However, these in vitro microglial models are not suitable for research in the field of neuro-degenerative diseases where aging is a crucial parameter. Only a few in vitro studies on aged microglia have been published so far, most of which use ex vivo microglia which cannot be kept in culture for prolonged periods of time. In the present study, we provide a new approach which allows the isolation and culture of an almost pure population of microglia from adult mouse brains. The isolation is based on a procedure which combines density separation and a subsequent culture selection process. After these steps, microglia form a non-adherent floating cell layer that can be easily and repeatedly harvested and replated. This method is simple and allows for a comparatively high yield and purity of adult microglial cells. The collected primary adult microglia proliferate and can be kept in culture for extended periods of time. We compared the primary adult microglia to primary microglia from neo-natal mice as well as to the C8-B4 microglial cell line. We found that adult microglia have similar, but not identical, immuno-phenotypic, functional and electrophysiological characteristics to the other in vitro models.

post to: CiteULike

Adaptive cyclic physiologic noise modeling and correction in functional MRI.

Publication Date: 2010 Mar 30 PMID: 20096307
Authors: Beall, E. B.
Journal: J Neurosci Methods

Physiologic noise in BOLD-weighted MRI data is known to be a significant source of the variance, reducing the statistical power and specificity in fMRI and functional connectivity analyses. We show a dramatic improvement on current noise correction methods in both fMRI and fcMRI data that avoids overfitting. The traditional noise model is a Fourier series expansion superimposed on the periodicity of parallel measured breathing and cardiac cycles. Correction using this model results in removal of variance matching the periodicity of the physiologic cycles. Using this framework allows easy modeling of noise. However, using a large number of regressors comes at the cost of removing variance unrelated to physiologic noise, such as variance due to the signal of functional interest (overfitting the data). It is our hypothesis that there are a small variety of fits that describe all of the significantly coupled physiologic noise. If this is true, we can replace a large number of regressors used in the model with a smaller number of the fitted regressors and thereby account for the noise sources with a smaller reduction in variance of interest. We describe these extensions and demonstrate that we can preserve variance in the data unrelated to physiologic noise while removing physiologic noise equivalently, resulting in data with a higher effective SNR than with current corrections techniques. Our results demonstrate a significant improvement in the sensitivity of fMRI (up to a 17% increase in activation volume for fMRI compared with higher order traditional noise correction) and functional connectivity analyses.

post to: CiteULike

Automated imaging of neuronal activity in freely behaving Caenorhabditis elegans.

Publication Date: 2010 Mar 30 PMID: 20096306
Authors: Ben Arous, J. - Tanizawa, Y. - Rabinowitch, I. - Chatenay, D. - Schafer, W. R.
Journal: J Neurosci Methods

In order to understand how neuronal circuits control locomotory patterns it is necessary to record neuronal activity of freely behaving animals. Here, using a new automated system for simultaneous recording of behavior and neuronal activity in freely moving Caenorhabditis elegans on standard agar plates, we show that spontaneous reversals from forward to backward locomotion reflect precisely the activity of the AVA command interneurons. We also witness spontaneous activity transients in the PLM sensory neurons during free behavior of the worm in standard conditions. We show that these activity transients are coupled to short spontaneous forward accelerations of the worm.

post to: CiteULike

Robust movement segmentation by combining multiple sources of information.

Publication Date: 2010 Mar 30 PMID: 20096305
Authors: Schot, W. D. - Brenner, E. - Smeets, J. B.
Journal: J Neurosci Methods

One of the first steps in analyzing kinematic data is determining the beginning and end of movement segments. This is often done automatically on the basis of one parameter (such as a speed minimum) and subsequently corrections are made if visual inspection of other kinematic parameters suggests that the obtained value was incorrect. We argue that in many cases it is impossible to find a satisfactory endpoint for all possible movement segments within an experiment using a single parameter because the intuition about the end of a segment is based on multiple criteria. Therefore by taking the maximum of an objective function based on multiple sources of information one can find the best possible time point to call the endpoint. We will demonstrate that this Multiple Sources of Information method (MSI-method) for finding endpoints performs better than conventional methods and that it is robust against arbitrary choices made by the researcher. Using it reduces the chance of introducing biases and eliminates the need for subjective corrections. Although we will take goal directed upper limb motion as an example throughout this paper, it should be stressed that the method could be applied to a wide variety of movements.

post to: CiteULike

The dolognawmeter: A novel instrument and assay to quantify nociception in rodent models of orofacial pain.

Publication Date: 2010 Mar 30 PMID: 20096303
Authors: Dolan, J. C. - Lam, D. K. - Achdjian, S. H. - Schmidt, B. L.
Journal: J Neurosci Methods

Rodent pain models play an important role in understanding the mechanisms of nociception and have accelerated the search for new treatment approaches for pain. Creating an objective metric for orofacial nociception in these models presents significant technical obstacles. No animal assay accurately measures pain-induced orofacial dysfunction that is directly comparable to human orofacial dysfunction. We developed and validated a high throughput, objective, operant, nociceptive animal assay, and an instrument to perform the assay termed the dolognawmeter, for evaluation of conditions known to elicit orofacial pain in humans. Using the device our assay quantifies gnawing function in the mouse. We quantified a behavioral index of nociception and demonstrated blockade of nociception in three models of orofacial pain: (1) TMJ inflammation, (2) masticatory myositis, and (3) head and neck cancer. This assay will be useful in the study of nociceptive mediators involved in the development and progression of orofacial pain conditions and it will also provide a unique tool for development and assessment of new therapeutic approaches.

post to: CiteULike

Improving the saccade peak velocity measurement for detecting fatigue.

Publication Date: 2010 Mar 30 PMID: 20083140
Authors: Hirvonen, K. - Puttonen, S. - Gould, K. - Korpela, J. - Koefoed, V. F. - Muller, K.
Journal: J Neurosci Methods

The aim of the study was to compare saccadic peak velocity (SPV) values measured with video based Fitness Impairment Tester (FIT) and electro-oculography (EOG) during prolonged wakefulness. We tested different numbers of saccades and two saccade paradigms to improve the EOG measurements for detecting fatigue. The SPVs were measured from 11 fast patrol boat navigators with FIT and EOG every sixth hour until 54h. Subjective sleepiness was assessed with the Karolinska Sleepiness Scale. EOG was measured using an overlap and a gap paradigm and the data was divided into sequential five 20-saccade blocks and cumulative blocks of 20, 40, 60, 80, and 100 saccades. Compared to the gap paradigm, the overlap paradigm produced a higher number of analyzable saccades for a given measurement time. The shorter measurements (20-40 saccades) appeared to be more sensitive for fatigue, whereas the longer measurements (60-100 saccades) were more sensitive to time spent on the task. Thus, the optimal number of saccades varies also depending on the research question. The EOG method was more sensitive to fatigue than FIT. The FIT values measured after 30 and 36h of wakefulness did not differ significantly from the baseline values, while subjective sleepiness and the EOG values showed that the participants were significantly less alert at these time points. The EOG measurements can be improved for detecting fatigue by using the overlap saccade paradigm. The SPV values measured with the EOG method appear to be somewhat more sensitive in detecting fatigue than the FIT method.

post to: CiteULike

Coping with spatial attention in real space: A low-cost portable testing system for the investigation of visuo-spatial processing in the human brain.

Publication Date: 2010 Mar 30 PMID: 20079374
Authors: Wolfe, B. - Rushmore, R. J. - Valero-Cabre, A.
Journal: J Neurosci Methods

While two-dimensional stimuli may be easily presented with any computer, an apparatus which allows a range of stimuli to be presented in three dimensions is not easily or cheaply available to researchers or clinicians. To fill this gap, we have developed the Realspace Testing System (RTS) which addresses the need for a flexible and multimodal stimulus presentation system capable of displaying stimuli in a three-dimensional space with a high degree of temporal accuracy. The RTS is able to control 26 channels of visual or audio stimuli, to send trigger pulses during each trial to external devices, such as a transcranial magnetic stimulator, and to record subject responses during the testing sessions. The RTS is flexible, portable and can be used in laboratory or clinical settings as required while being built at a low cost using off the shelf components. We have tested the RTS by performing an exploratory experiment on the role of right posterior parietal cortex in visuo-spatial processing in conjunction with online transcranial magnetic stimulation (TMS) and verified that the system can accurately present stimuli as needed while triggering a TMS pulse during each trial at the required time. The RTS could be appealing and useful to a range of researchers or clinicians who may choose to use it much as we have designed it, or use it in its current state as a starting point to customize their stimulus control systems in real space.

post to: CiteULike

Intrinsic signal optical imaging of brain function using short stimulus delivery intervals.

Publication Date: 2010 Mar 30 PMID: 20079373
Authors: Chen-Bee, C. H. - Agoncillo, T. - Lay, C. C. - Frostig, R. D.
Journal: J Neurosci Methods

Intrinsic signal optical imaging (ISOI) can be used to map cortical function and organization. Because its detected signal lasts 10+s consisting of three phases, trials are typically collected using a long (tens of seconds) stimulus delivery interval (SDI) at the expense of efficiency, even when interested in mapping only the first signal phase (e.g., ISOI initial dip). It is unclear how the activity profile can change when stimuli are delivered at shorter intervals, and whether a short SDI can be implemented to improve efficiency. The goals of the present study are twofold: characterize the ISOI activity profile when multiple stimuli are delivered at 4s intervals, and determine whether successful mapping can be attained from trials collected using an SDI of 4s (offering >10x increase in efficiency). Our results indicate that four stimuli delivered 4s apart evoke an activity profile different from the triphasic signal, consisting of signal dips in a series at the same frequency as the stimuli despite a strong rise in signal prior to the 2nd to 4th stimuli. Visualization of such signal dips is dependent on using a baseline immediately prior to every stimulus. Use of the 4-s SDI is confirmed to successfully map activity with a similar location in peak activity and increased areal extent and peak magnitude compared to using a long SDI. Additional experiments were performed to begin addressing issues such as SDI temporal jittering, response magnitude as a function of SDI duration, and application for successful mapping of cortical function topography.

post to: CiteULike

Tail nerve electrical stimulation induces body weight-supported stepping in rats with spinal cord injury.

Publication Date: 2010 Mar 30 PMID: 20079372
Authors: Zhang, S. X. - Huang, F. - Gates, M. - White, J. - Holmberg, E. G.
Journal: J Neurosci Methods

Walking or stepping has been considered the result from the activation of the central pattern generator (CPG). In most patients with spinal cord injury (SCI) the CPG is undamaged. To date, there are no noninvasive approaches for activating the CPG. Recently we developed a noninvasive technique, tail nerve electrical stimulation (TANES), which can induce positive hind limb movement of SCI rats. The purpose of this study is to introduce the novel technique and examine the effect of TANES on CPG activation. A 25mm contusion injury was produced at spinal cord T10 of female, adult Long-Evans rats by using the NYU impactor device. Rats received TANES ( approximately 40mA at 4kHz) 7 weeks after injury. During TANES all injured rats demonstrated active body weight-supported stepping of hind limbs with left-right alternation and occasional front-hind coordination, resulting in significant, temporary increase in BBB scores (p<0.01). However, there is no response to TANES from rats with L2 transection, consistent with other reports that the CPG may be located at L1-2. S1 transection negatively implies the key role of TANES in CPG activation. The TANES not only renders paralyzed rats with a technique-induced ability to walk via activating CPG, but also is likely to be used for locomotor training. It has more beneficial effects for physical training over other training paradigms including treadmill training and invasive functional electrical stimulation. Therefore the TANES may have considerable potential for achieving improvement of functional recovery in animal models and a similar method may be suggested for human study.

post to: CiteULike

AnimatLab: A 3D graphics environment for neuromechanical simulations.

Publication Date: 2010 Mar 30 PMID: 20074588
Authors: Cofer, D. - Cymbalyuk, G. - Reid, J. - Zhu, Y. - Heitler, W. J. - Edwards, D. H.
Journal: J Neurosci Methods

The nervous systems of animals evolved to exert dynamic control of behavior in response to the needs of the animal and changing signals from the environment. To understand the mechanisms of dynamic control requires a means of predicting how individual neural and body elements will interact to produce the performance of the entire system. AnimatLab is a software tool that provides an approach to this problem through computer simulation. AnimatLab enables a computational model of an animal's body to be constructed from simple building blocks, situated in a virtual 3D world subject to the laws of physics, and controlled by the activity of a multicellular, multicompartment neural circuit. Sensor receptors on the body surface and inside the body respond to external and internal signals and then excite central neurons, while motor neurons activate Hill muscle models that span the joints and generate movement. AnimatLab provides a common neuromechanical simulation environment in which to construct and test models of any skeletal animal, vertebrate or invertebrate. The use of AnimatLab is demonstrated in a neuromechanical simulation of human arm flexion and the myotactic and contact-withdrawal reflexes.

post to: CiteULike

Accurate measurement of junctional conductance between electrically coupled cells with dual whole-cell voltage-clamp under conditions of high series resistance.

Publication Date: 2010 Mar 15 PMID: 20074587
Authors: Hartveit, E. - Veruki, M. L.
Journal: J Neurosci Methods

Accurate measurement of the junctional conductance (G(j)) between electrically coupled cells can provide important information about the functional properties of coupling. With the development of tight-seal, whole-cell recording, it became possible to use dual, single-electrode voltage-clamp recording from pairs of small cells to measure G(j). Experiments that require reduced perturbation of the intracellular environment can be performed with high-resistance pipettes or the perforated-patch technique, but an accompanying increase in series resistance (R(s)) compromises voltage-clamp control and reduces the accuracy of G(j) measurements. Here, we present a detailed analysis of methodologies available for accurate determination of steady-state G(j) and related parameters under conditions of high R(s), using continuous or discontinuous single-electrode voltage-clamp (CSEVC or DSEVC) amplifiers to quantify the parameters of different equivalent electrical circuit model cells. Both types of amplifiers can provide accurate measurements of G(j), with errors less than 5% for a wide range of R(s) and G(j) values. However, CSEVC amplifiers need to be combined with R(s)-compensation or mathematical correction for the effects of nonzero R(s) and finite membrane resistance (R(m)). R(s)-compensation is difficult for higher values of R(s) and leads to instability that can damage the recorded cells. Mathematical correction for R(s) and R(m) yields highly accurate results, but depends on accurate estimates of R(s) throughout an experiment. DSEVC amplifiers display very accurate measurements over a larger range of R(s) values than CSEVC amplifiers and have the advantage that knowledge of R(s) is unnecessary, suggesting that they are preferable for long-duration experiments and/or recordings with high R(s).

post to: CiteULike

Regional convection-enhanced delivery of gadolinium-labeled albumin in the rat hippocampus in vivo.

Publication Date: 2010 Mar 15 PMID: 20067808
Authors: Astary, G. W. - Kantorovich, S. - Carney, P. R. - Mareci, T. H. - Sarntinoranont, M.
Journal: J Neurosci Methods

Convection-enhanced delivery (CED) has emerged as a promising method of targeted drug delivery for treating central nervous system (CNS) disorders, but the influence of brain structure on infusate distribution is unclear. We have utilized this approach to study extracellular transport and distribution of a contrast agent in the hippocampus, a complex structure susceptible to CNS disorders. The magnetic resonance (MR) contrast agent diethylene triamene penta-acetic acid chelated gadolinium-labeled albumin (Gd-albumin), tagged with Evans blue dye, was directly infused (V(i)=5 microl) into the dorsal and ventral hippocampus of seven male Sprague-Dawley rats. The final distribution profile of the contrast agent, a product of CED and limited diffusion, was observed in vivo using high-resolution T1-weighted MR imaging at 11.1T. Dense cell layers, such as the granule cell layer of the dentate gyrus and the pyramidal cell layer of CA1, appeared to be barriers to transport of the tracer. Three-dimensional distribution shape and volume (V(d)) differences, between the dorsal and ventral hippocampus infusions, were determined from the MR images using a semi-automatic segmentation routine (dorsal V(d)=23.4+/-1.8 microl, ventral V(d)=36.4+/-5.1 microl). Finer structural detail of the hippocampus was obtained using a combination of histological analysis and fluorescence imaging. This study demonstrates that CED has the potential to target all regions of the hippocampus and that tracer distribution is influenced by infusion site, underlying structure and circuitry, and extent of backflow. Therefore, CED, combined with high-resolution MR imaging, may be a useful strategy for delivering therapeutics for the treatment of CNS disorders affecting the hippocampus.

post to: CiteULike

Laser microdissection and pressure catapulting: Combining LMPC with IHC to investigate NMDA receptor subunit composition.

Publication Date: 2010 Mar 30 PMID: 20067807
Authors: Brinkmann, H. - Curry, J. A.
Journal: J Neurosci Methods

Laser microdissection and pressure catapulting (LMPC) has allowed investigators the opportunity to look at enriched cell populations from heterogeneous tissues. Selection of cells is routinely based upon morphological or histological criteria. The inclusion of an immunohistochemistry step prior to LMPC potentially allows investigators to classify cell populations both morphologically and functionally. Practical limitations with regard to RNA integrity following IHC steps have meant that the value that this combination brings has been reduced. Here, through the use of RNA preserving buffers, we have been able to successfully label, dissect and extract RNA from NK1 expressing cells. RT-PCR was carried out to confirm the expression of NK1, NR2A and NR2B subunit expression. Our data confirmed the expression of NK1 in cells labelled with the antibody, and the absence of expression in cells absent of staining. As well as this, relative expression of both NR2A and NR2B was determined. Furthermore, the RNA was of high enough quality to allow these methods to be used for studies involving RNA amplification steps, such as microarray analysis.

post to: CiteULike

Non-contact measurement of linear external dimensions of the mouse eye.

Publication Date: 2010 Mar 30 PMID: 20067806
Authors: Wisard, J. - Chrenek, M. A. - Wright, C. - Dalal, N. - Pardue, M. T. - Boatright, J. H. - Nickerson, J. M.
Journal: J Neurosci Methods

Biometric analyses of quantitative traits in eyes of mice can reveal abnormalities related to refractive or ocular development. Due to the small size of the mouse eye, highly accurate and precise measurements are needed to detect meaningful differences. We sought a non-contact measuring technique to obtain highly accurate and precise linear dimensions of the mouse eye. Laser micrometry was validated with gauge block standards. Simple procedures to measure eye dimensions on three axes were devised. Mouse eyes from C57BL/6J and rd10 on a C57BL/6J background were dissected and extraocular muscle and fat removed. External eye dimensions of axial length (anterior-posterior (A-P) axis) and equatorial diameter (superior-inferior (S-I) and nasal-temporal (N-T) axes) were obtained with a laser micrometer. Several approaches to prevent or ameliorate evaporation due to room air were employed. The resolution of the laser micrometer was less than 0.77mum, and it provided accurate and precise non-contact measurements of eye dimensions on three axes. External dimensions of the eye strongly correlated with eye weight. The N-T and S-I dimensions of the eye correlated with each other most closely from among the 28 pair-wise combinations of the several parameters that were collected. The equatorial axis measurements correlated well from the right and left eye of each mouse. The A-P measurements did not correlate or correlated poorly in each pair of eyes. The instrument is well suited for the measurement of enucleated eyes and other structures from most commonly used species in experimental vision research and ophthalmology.

post to: CiteULike

TTC staining of damaged brain areas after MCA occlusion in the rat does not constrict quantitative gene and protein analyses.

Publication Date: 2010 Mar 15 PMID: 20064557
Authors: Kramer, M. - Dang, J. - Baertling, F. - Denecke, B. - Clarner, T. - Kirsch, C. - Beyer, C. - Kipp, M.
Journal: J Neurosci Methods

In models of ischemic stroke, TTC (2,3,5-triphenyltetrazolium chloride) staining is commonly applied for the fast and reliable visualization of hypoxic brain tissue and for defining the size of cerebral infarction and penumbra. Deciphering molecular processes of pathogenesis within the penumbra is of particular interest for the development of therapeutic strategies. The aim of this study was to assess whether TTC-stained tissues can easily and in a reliable quantitative manner be processed for further molecular and biochemical analyses. We applied phenol-based RNA isolation, protein lysis by conventional RIPA buffer, and combined RNA/protein isolation with NucleoSpinRNA/Protein-Kit. Gene and protein expression analyses were performed by RT-rtPCR and Western-blotting. Middle cerebral arteria occlusion (MCAO) in rats was performed following a standardized experimental procedure. After MCAO, TTC staining revealed massive cell death in cortical and sub-cortical areas. TTC processing did not affect the quality of tissue RNA and protein. The expression of housekeeping and regulatory genes and proteins revealed no difference between control and TTC-stained groups. The expression of known stroke-regulated genes such as TNFalpha and IL1beta revealed similar induction profiles after TTC staining as described in the literature. TTC staining allows the precise delineation of lesioned and primarily non-lesioned brain areas for subsequent dissection of selected tissue pieces for molecular analysis. Our study demonstrates that TTC-stained tissues in stroke animal models can be used for quantitative gene and protein expression analyses without constriction. Pathomechanisms of ongoing tissue damage within the penumbra region can now be investigated in detail.

post to: CiteULike

Facial muscle activation patterns in healthy male humans: a multi-channel surface EMG study.

Publication Date: 2010 Mar 15 PMID: 20064556
Authors: Schumann, N. P. - Bongers, K. - Guntinas-Lichius, O. - Scholle, H. C.
Journal: J Neurosci Methods

In order to accurately characterize essential muscle activity during facial movements a new surface EMG (SEMG) technique was introduced and applied. Results represent reference data of healthy persons for future diagnostic purposes. In 30 healthy males monopolar electromyograms of the facial muscles were simultaneously recorded from 48 bilateral-symmetrically applied small surface electrodes while performing 29 facial movements of high clinical relevance. Mean SEMG amplitudes were quantified by power spectral analysis, normalized and presented as movement-related SEMG profiles. The mean SEMG amplitudes increased significantly in response to facial movements. Critical values of the movement-related SEMG amplitude increase were ascertained, valid for 90% of all examined subjects. The mean SEMG amplitudes differed between the performed facial movements, the examined muscles, and intramuscularly between lateral-medial and superior-inferior electrode positions, but not systematically between right and left side of face. The results show that the interplay between individual facial muscles and intramuscularly between their functional subunits is more differentiated than was previously estimated. With the presented facial SEMG technique the produced SEMG profiles are highly relevant for better planning of facial movement restoration. Based on the established reference data, this method can be used to objectively evaluate a facial paresis and to monitor changes during the course of disease and treatment. To easily apply the method, a reduction of electrode positions is intended after the clinical evaluation.

post to: CiteULike

Calibration of the stereological estimation of the number of myelinated axons in the rat sciatic nerve: a multicenter study.

Publication Date: 2010 Mar 15 PMID: 20064555
Authors: Kaplan, S. - Geuna, S. - Ronchi, G. - Ulkay, M. B. - von Bartheld, C. S.
Journal: J Neurosci Methods

Several sources of variability can affect stereological estimates. Here we measured the impact of potential sources of variability on numerical stereological estimates of myelinated axons in the adult rat sciatic nerve. Besides biological variation, parameters tested included two variations of stereological methods (unbiased counting frame versus 2D-disector), two sampling schemes (few large versus frequent small sampling boxes), and workstations with varying degrees of sophistication. All estimates were validated against exhaustive counts of the same nerve cross sections to obtain calibrated true numbers of myelinated axons (gold standard). In addition, we quantified errors in particle identification by comparing light microscopic and electron microscopic images of selected consecutive sections. Biological variation was 15.6%. There was no significant difference between the two stereological approaches or workstations used, but sampling schemes with few large samples yielded larger differences (20.7+/-3.7% SEM) of estimates from true values, while frequent small samples showed significantly smaller differences (12.7+/-1.9% SEM). Particle identification was accurate in 94% of cases (range: 89-98%). The most common identification error was due to profiles of Schwann cell nuclei mimicking profiles of small myelinated nerve fibers. We recommend sampling frequent small rather than few large areas, and conclude that workstations with basic stereological equipment are sufficient to obtain accurate estimates. Electron microscopic verification showed that particle misidentification had a surprisingly variable and large impact of up to 11%, corresponding to 2/3 of the biological variation (15.6%). Thus, errors in particle identification require further attention, and we provide a simple nerve fiber recognition test to assist investigators with self-testing and training.

post to: CiteULike

A novel decalcification method for adult rodent bone for histological analysis of peripheral-central nervous system connections.

Publication Date: 2010 Mar 15 PMID: 20043948
Authors: Begum, F. - Zhu, W. - Namaka, M. P. - Frost, E. E.
Journal: J Neurosci Methods

Histological analysis of bone encased tissue is severely hampered by technical difficulties associated with sectioning calcified tissue. Cryosectioning of bone is possible but requires significant technical adaptation and expensive materials and is often time-consuming. Some decalcifying reagents in common use result in successful cryosectioning in less time but the integrity of the soft tissue of the spinal column is often compromised during processing. This can result in significant loss of cellular detail. In order to find a method that would allow cryosectioning of the bone without loss of structural integrity of the underlying soft tissue we assessed the efficacy of four different decalcifying reagents with respect to their effects on the cellular structure of the myelin of the grey and white matter of the spinal cord. The antigenic integrity of the spinal cord white matter was evaluated using tissue structural integrity and quality of myelin basic protein immunostaining. The result of this research shows that 6% TCA not only decalcifies intact spinal column suitably for cryosectioning but does so without compromising the antigenic integrity of the tissue. The ease of application, speed of processing and a favorable cost-effective profile were secondary benefits noted with the use of the 6% TCA decalcifying solution. The ability to utilize a decalcifying solution that allows for both histomorphometry and immunohistochemistry in the same spinal column segment represents a novel technique that will provide new insights into pathophysiological aspects and therapeutic approaches ispinal cord damage or disease.

post to: CiteULike

Detection of brain-derived neurotrophic factor (BDNF) in rat blood and brain preparations using ELISA: pitfalls and solutions.

Publication Date: 2010 Mar 15 PMID: 20043947
Authors: Elfving, B. - Plougmann, P. H. - Wegener, G.
Journal: J Neurosci Methods

Quantification of endogenous brain-derived neurotrophic factor (BDNF) can be performed with an enzyme-linked immunosorbent assay (ELISA). Although BDNF has been determined in blood and brain preparations in numerous studies with ELISA kits, the methodological issues regarding measurements of BDNF concentrations in the blood and particularly in the brain have only been superficially investigated. We aimed at validating and optimizing the BDNF ELISA kit with respect to measurements in rat blood and brain samples. We found that the pre-analytical conditions were critical for plasma samples, but not serum or whole blood samples. The intra- and inter-assay variation and the accuracy and yield of the BDNF ELISA kit in rat serum and brain tissue were conducted with the optimal dilutions of frontal cortex and hippocampus extract. The optimal dilutions of frontal cortex and hippocampus extracts were determined to be 20 and 120 times, and we established that the intra-assay coefficient of variation (CV%) was 8 in hippocampus and 2 in frontal cortex and serum. The inter-assay variation was also low with a CV% of 11 or less in hippocampus, frontal cortex, and serum. Finally, we found that the accuracy and yield of the BDNF measurements were high in serum and low in hippocampus and frontal cortex. We conclude that the BDNF ELISA kit determines serum BDNF accurately and with high reproducibility. Furthermore it can be used for measurement of BDNF in rat brain preparations when particular precautions are taken and in particular with care regarding the dilution of the brain tissue samples.

post to: CiteULike

Selection of reference genes for gene expression studies in rat oligodendrocytes using quantitative real time PCR.

Publication Date: 2010 Mar 15 PMID: 20036692
Authors: Nelissen, K. - Smeets, K. - Mulder, M. - Hendriks, J. J. - Ameloot, M.
Journal: J Neurosci Methods

Quantitative real time polymerase chain reaction (qPCR) has become a widely used tool to examine gene expression levels. Reliable quantification, however, depends on a proper normalization strategy. Normalization with multiple reference genes is becoming the standard, although the most suitable reference genes depend on the applied treatment as well as the tissue or cell type studied. In this study the stability of various reference genes was investigated in cultures of oligodendrocytes derived from either mature or neonatal rats, the latter also in the presence of the liver X receptor (LXR) agonist. The expression stability of ten commonly used reference genes (HPRT, GAPDH, 18S, ActB, CycA, Tbp, Rpl13A, YWHAZ, HMBS, Pgk1) was analyzed using geNorm and NormFinder. When comparing the different types of cell cultures, Rpl13A, CycA, Pgk1 and YWHAZ were identified as most stable genes. After LXR agonist treatment, CycA, Pgk1 and Rpl13A were found to be the most stable by both geNorm and NormFinder. HMBS and the commonly used housekeeping genes GAPDH and 18S turned out to be the most variable according to geNorm and NormFinder. In conclusion, the use of multiple reference genes, instead of only one, in qPCR experiments with rat oligodendrocytes is strongly advised and standard housekeeping genes such as GAPDH and 18S are not recommended as they appear to be relatively unstable under the experimental conditions used. Reference gene selection should always be performed for each individual experiment, since useful reference genes are very specific for every situation.

post to: CiteULike

Semi-automatic stereotactic coordinate identification algorithm for routine localization of Deep Brain Stimulation electrodes.

Publication Date: 2010 Mar 15 PMID: 20036691
Authors: Hebb, A. O. - Miller, K. J.
Journal: J Neurosci Methods

Deep Brain Stimulation (DBS) is a routine therapy for movement disorders, and has several emerging indications. We present a novel protocol to define the stereotactic coordinates of metallic DBS implants that may be routinely employed for validating therapeutic anatomical targets. Patients were referred for troubleshooting or new DBS implantation. A volumetric MRI of the brain obtained prior to or during this protocol was formatted to the Anterior Commissure-Posterior Commissure (AC-PC) coordinate system. Patients underwent a CT scan of the brain in an extended Hounsfield unit (EHU) mode. A semi-automatic detection algorithm based on a Normalized Mutual Information (NMI) co-registration method was implemented to measure the AC-PC coordinates of each DBS contact. This algorithm was validated using manual DBS contact identification. Fifty MRI-CT image pairs were available in 39 patients with a total of 336 DBS electrodes. The median and mean Euclidean distance errors for automatic identification of electrode locations were 0.20mm and 0.22 mm, respectively. This method is an accurate method of localization of active DBS contacts within the sub-cortical region. As the investigational indications of DBS expand, this method may be used for verification of final implant coordinates, critical for understanding clinical benefit and comparing efficacy between subjects.

post to: CiteULike

A miniaturized chronic microelectrode drive for awake behaving head restrained mice and rats.

Publication Date: 2010 Mar 15 PMID: 20036690
Authors: Haiss, F. - Butovas, S. - Schwarz, C.
Journal: J Neurosci Methods

The present work introduces an electrode microdrive system small enough to be placed into two distinct brain areas in head-fixed mice. To meet the space constraint imposed by the size of mice and the additional presence of a head post, the size and weight of the components were minimized. In the current version, an individual microdrive moves an array of four Reitboeck type electrodes. We report about successful implantation in rats and mice using one or two microdrives. Using two of these devices in individual mice/rats, the recording of parallel single and multi-unit as well as local field potential from prefrontal, motor, somatosensory cortex and hippocampus is demonstrated. The system can be easily constructed with machinery and equipment present in most neurophysiology labs.

post to: CiteULike

Analyzing neurite outgrowth from explants by fitting ellipses.

Publication Date: 2010 Mar 15 PMID: 20036284
Authors: Haines, C. - Goodhill, G. J.
Journal: J Neurosci Methods

The establishment of appropriate connectivity in the developing nervous system depends on many molecular guidance cues. A key method for studying the response of nerve fibers to such guidance cues is to culture explants of neural tissue in three-dimensional collagen gels. However, most previous analyses of the neurite outgrowth patterns from these explants have been very simple, often measuring only one or two parameters. Here we introduce a more sophisticated method for characterizing neurite outgrowth from explants, based on fitting an ellipse to the pattern of outgrowth. This provides 5 parameters describing the outgrowth: x and y position of the center of the ellipse, the elongation, the area and the tilt. We then apply this method to a large dataset of dorsal root ganglion explants grown in the presence of precisely controlled gradients of nerve growth factor. This analysis reveals a number of new features of these data. For instance, we find that it is the position of the center of the ellipse rather than the shape of the ellipse that is correlated with the strength of the gradient. Together these results show that ellipse-fitting of explant data can give new insights into the biological processes underlying neurite guidance by molecular cues.

post to: CiteULike

Recording and online analysis of auditory steady state responses (ASSR) in Matlab.

Publication Date: 2010 Mar 15 PMID: 20036283
Authors: Bahmer, A. - Baumann, U.
Journal: J Neurosci Methods

Auditory steady state responses (ASSR) are a current research focus because of their potential use as a diagnostic tool. Research platforms are required to test user defined stimuli and algorithms for the analysis of electrophysiologic responses. Commercially available ASSR devices are not adequately flexible. To enable a larger group of scientists to pursue ASSR research, we introduce a cost-efficient and flexible ASSR setup. ASSR recording and online analysis software in Matlab (The Mathworks, Inc.) was developed for a standard PC equipped with an external professional sound card, audiometric headphones, and an EEG biosignal preamplifier.

post to: CiteULike

High resolution stereoscopic volume visualization of the mouse arginine vasopressin system.

Publication Date: 2010 Mar 15 PMID: 20036282
Authors: Clements, R. J. - Mintz, E. M. - Blank, J. L.
Journal: J Neurosci Methods

New imaging technologies have increased our capabilities to resolve three-dimensional structures from microscopic samples. Laser-scanning confocal microscopy is particularly amenable to this task because it allows the researcher to optically section biological samples, creating three-dimensional image volumes. However, a number of problems arise when studying neural tissue samples. These include data set size, physical scanning restrictions, volume registration and display. To deal with these issues, we undertook large-scale confocal scanning microscopy in order to visualize neural networks spanning multiple tissue sections. We demonstrate a technique to create and visualize a three-dimensional digital reconstruction of the hypothalamic arginine vasopressin neuroendocrine system in the male mouse. The generated three-dimensional data included a volume of tissue that measures 4.35 mm x 2.6 mm x 1.4mm with a voxel resolution of 1.2 microm. The dataset matrix included 3508 x 2072 x 700 pixels and was a composite of 19,600 optical sections. Once reconstructed into a single volume, the data is suitable for interactive stereoscopic projection. Stereoscopic imaging provides greater insight and understanding of spatial relationships in neural tissues' inherently three-dimensional structure. This technique provides a model approach for the development of data sets that can provide new and informative volume rendered views of brain structures. This study affirms the value of stereoscopic volume-based visualization in neuroscience research and education, and the feasibility of creating large-scale high resolution interactive three-dimensional reconstructions of neural tissue from microscopic imagery.

post to: CiteULike

The ERP PCA Toolkit: an open source program for advanced statistical analysis of event-related potential data.

Publication Date: 2010 Mar 15 PMID: 20035787
Authors: Dien, J.
Journal: J Neurosci Methods

This article presents an open source Matlab program, the ERP PCA (EP) Toolkit, for facilitating the multivariate decomposition and analysis of event-related potential data. This program is intended to supplement existing ERP analysis programs by providing functions for conducting artifact correction, robust averaging, referencing and baseline correction, data editing and visualization, principal components analysis, and robust inferential statistical analysis. This program subserves three major goals: (1) optimizing analysis of noisy data, such as clinical or developmental; (2) facilitating the multivariate decomposition of ERP data into its constituent components; (3) increasing the transparency of analysis operations by providing direct visualization of the corresponding waveforms.

post to: CiteULike

Comparison of visually evoked local field potentials in isolated turtle brain: patterned versus blank stimulation.

Publication Date: 2010 Mar 15 PMID: 20034520
Authors: Luo, Q. - Lu, H. - Lu, H. - Yang, Y. - Gao, J. H.
Journal: J Neurosci Methods

Isolated turtle brain/eye preparation has recently been used as a bloodless animal model for detecting the magnetic resonance imaging (MRI) signal changes produced by visually evoked neuronal currents. The present work aims to determine whether checkerboard-patterned or full field flash (blank) stimulation should be used in order to achieve stronger neuronal responses in turtle brain/eye preparation. The knowledge gained in this study is essential for optimizing the visual stimulation methods in functional neuroimaging studies using turtle brain/eye preparation. In this study, visually evoked local field potentials (LFPs) were measured and compared in turtle visual cortex and optic tectum elicited by checkerboard and full field flash stimuli with three different inter-stimulus intervals (ISIs=5, 10, and 16s). It was found that the behavior of neuronal adaptation in the cortical and tectal LFP signals for checkerboard stimulation was comparable to flash stimulation. In addition, there was no significant difference in the LFP peak amplitudes (ISI=16s) between these two stimuli. These results indicate that the intensity of neuronal responses to checkerboard is comparable to flash stimulation. These two stimulation methods should be equivalent in functional neuroimaging studies using turtle brain/eye preparation.

post to: CiteULike

An instrumented glove for small primates.

Publication Date: 2010 Mar 15 PMID: 20034519
Authors: Overduin, S. A. - Zaheer, F. - Bizzi, E. - d'Avella, A.
Journal: J Neurosci Methods

The Cymanus is a novel flex sensor glove for measuring hand kinematics in primates. It was used to monitor 9 joints of a rhesus macaque performing a grasping task with 25 objects. Over 6 days, the monkey tolerated the glove and showed no significant impairment in performance. The sensors linearly tracked joint angles, with joint trajectories preserved over days. Angular positions discriminated objects as accurately as electromyograms recorded simultaneously from 24 arm and hand muscles, and were maximally informative of object identity at the end of reach-to-grasp. In a further validation of the glove, muscle activity controlling a joint was correlated with the joint's angular acceleration 70 ms later.

post to: CiteULike

Modular double sucrose gap apparatus for improved recording of compound action potentials from rat and mouse spinal cord white matter preparations.

Publication Date: 2010 Mar 15 PMID: 20034518
Authors: Velumian, A. A. - Wan, Y. - Samoilova, M. - Fehlings, M. G.
Journal: J Neurosci Methods

Compound action potential (CAP) recording is a powerful tool for studying the conduction properties and pharmacology of axons in multi-axonal preparations. The sucrose gap technique improves CAP recording by replacing the extracellular solution between the recording electrodes with a non-conductive sucrose solution to minimize extracellular shunting. The double sucrose gap (DSG), conferring similar advantages at the stimulation site, has been extensively used on guinea pig spinal cord white matter (WM) in vitro. Establishing the DSG methodology for WM preparations from smaller animals such as rats and mice is appealing due to their extensive use in basic and translationally oriented research. Here we describe a versatile modular DSG apparatus with rubber membrane separation of the compartments, suitable for WM strips from rat and mouse spinal cord. The small volumes of compartments (<0.1 ml) and the air-tight design allow perfusion rates of 0.5-1 ml/min with faster refreshment rates compared to commonly used 2-3 ml/min and larger compartments, providing economical usage of expensive pharmacological drugs. Our improved DSG design is particularly efficient for uncovering slower conducting, higher threshold CAP components, as demonstrated by recordings of C-wave (non-myelinated axons) in rat dorsal WM. In myelin-deficient Shiverer mice with genetically dysmyelinated axons, our DSG apparatus recordings revealed a multi-peak C-wave without preceding faster components. The improved stimulation and recording with our DSG apparatus, lowering the range of required stimulus intensities and reducing the artifact interference with recorded CAPs provide for critical technical advantages that allow for more detailed analysis of CAPs in relatively short preparations.

post to: CiteULike

Optimized cannula design and placement for convection-enhanced delivery in rat striatum.

Publication Date: 2010 Mar 15 PMID: 20026357
Authors: Yin, D. - Forsayeth, J. - Bankiewicz, K. S.
Journal: J Neurosci Methods

The stereotactic delivery of therapeutic agents into brain has been problematic because of reflux and leakage of the delivered agent. Good distribution of infusates by convection-enhanced delivery (CED) depends very much on cannula design, precise cannula placement and infusion rates. We have recently published cannula targeting data for the non-human primate (NHP) putamen in which we defined infusion parameters referred to as "red", "blue", and "green" zones for cannula placements that result in poor, sub-optimal and optimal volumes of distribution (Vd), respectively. Therefore, we applied our observations in NHP putamen to the rat brain. Initially, trypan blue dye was infused into agarose gels to evaluate distribution and reflux characteristics of a scaled-down cannula without step and 1-mm stepped cannula. "Stepped" means a sharp transition from a wider stent to a narrower tip; thus the distance of the cannula tip to the larger diameter attachment defines the step distance. Reflux was contained with the stepped design even with an infusion rate of 3.0 microl/min and large infusion volumes in the agarose gel study. Infusions of a recombinant growth factor, GDNF, into rat striatum demonstrated that the presence of a 1-mm stepped cannula prevented reflux and resulted in excellent distribution of GDNF in the striatum. We conclude that a stepped cannula with a 1-mm tip is important for achieving reliable distribution of infused agents in rat brain. It should be considered when local therapies such as gene transfer, local protein administration or cellular replacement are evaluated in rodent models.

post to: CiteULike

Development of animal model for vasculatic neuropathy: Induction by ischemic-reperfusion in the rat femoral artery.

Publication Date: 2010 Feb 15 PMID: 20026113
Authors: Muthuraman, A. - Ramesh, M. - Sood, S.
Journal: J Neurosci Methods

Ischemic-reperfusion (I/R) is common in various pathological conditions like diabetic complication, complex regional pain syndrome type II (CRPS II), necrotizing vascular occlusive disease and trauma. We have developed an animal model of ischemic-reperfusion injury induced nociceptive sensory neuropathy in rats. The model was validated after 2, 4 and 6h of ischemia followed by prolonged reperfusion. The sensory behavioral assessment revealed thermal and mechanical hyperalgesia in paw and in tail which expressed the peripheral and central neuropathic pain respectively. We observed a decrease in the serum IL-10 and nerve conduction velocity and increase in the serum nitrate, malondialdehyde (MDA) and TNF-alpha levels in the 4 and 6h I/R groups in biochemical and electrophysiological evaluations. Histopathological study had revealed the decrease in nerve fiber density in the moderate and severe I/R groups. We selected the moderate (4h) ischemic-reperfusion injury as beneficial model because of the good correlation with clinical status for the development of neuropathy in human associated with severe pain disorders. This model can be used to explore pathophysiological mechanisms implied in the genesis of neuropathic pain and also to evaluate the new analgesic agents, peripheral neuro-vasoactive substances and neuroprotective drugs.

post to: CiteULike

Technique for collection of cerebrospinal fluid from the cisterna magna in rat.

Publication Date: 2010 Mar 15 PMID: 20005255
Authors: Pegg, C. C. - He, C. - Stroink, A. R. - Kattner, K. A. - Wang, C. X.
Journal: J Neurosci Methods

Cerebrospinal fluid (CSF) is in direct contact with the extracellular space in the central nervous system (CNS), and biological changes in the brain can be reflected in CSF. In the present article, a procedure for collection of CSF in rats is described. The technique quickly and reliably yields large quantities of CSF (50-150 microl) in rats. More importantly, blood contamination of the CSF is avoided. Furthermore, detections of ATP and interleukin (IL)-1 beta in the CSF have been carried out. ATP concentration in the CSF samples was between 8.3 and 15.8 nM, with an average of 10.5+/-0.83 nM (mean+/-SEM). The concentrations of IL-1beta were below the detection limit in the CSF in the laminectomy control rats, but it increased to 0.26+/-0.07 ng/ml at 1h after spinal cord injury. This technique offers an alternative method to surgical cannulation for the collection of CSF in rats.

post to: CiteULike

Neurolucida Lucivid versus Neurolucida camera: A quantitative and qualitative comparison of three-dimensional neuronal reconstructions.

Publication Date: 2010 Feb 15 PMID: 19963008
Authors: Anderson, K. - Yamamoto, E. - Kaplan, J. - Hannan, M. - Jacobs, B.
Journal: J Neurosci Methods

A critical issue in quantitative neuromorphology is the accuracy and subsequent reliability of the tracing techniques employed to characterize neuronal components. Historically, the camera lucida was the only option for such investigations. In 1987, MBF Bioscience, Inc. (Williston, VT) developed the integrative Neurolucida computer-microscope system, replacing the camera lucida drawing tube with a Lucivid cathode ray tube, thereby allowing computer overlays directly on the view through microscope oculars. Subsequent advances in digital cameras have allowed the Lucivid system to be replaced so that microscope images can be traced by viewing the digital image on a computer monitor. Indeed, the camera systems now outsell Lucivid systems 9 to 1 (J. Glaser, personal communication, 08/2008). Nevertheless, researchers seldom note which of these configurations are being used (which may confound the accuracy of data sharing), and there have been no published comparisons of the Lucivid and camera configurations. The present study thus assesses the relative accuracy of these two hardware configurations by examining reconstructions of human pyramidal neurons. We report significant differences with respect to dendritic spines, with the camera estimates of spine counts being greater than those obtained with the Lucivid system. Potential underlying reasons (e.g., magnification, illumination, and resolution, as well as observer ergonomic differences between the two systems) for these quantitative findings are explored here, along with qualitative observations on the relative strengths of each configuration.

post to: CiteULike

Genetically modified canine Schwann cells--In vitro and in vivo evaluation of their suitability for peripheral nerve tissue engineering.

Publication Date: 2010 Feb 15 PMID: 19962404
Authors: Schmitte, R. - Tipold, A. - Stein, V. M. - Schenk, H. - Flieshardt, C. - Grothe, C. - Haastert, K.
Journal: J Neurosci Methods

After peripheral nerve injury, Schwann cells (SC) guarantee for a regeneration-promoting milieu and are crucially involved in axonal regeneration. For extended nerve defects, bridging with an autologous nerve transplant is the gold standard therapy. Artificial biohybrid nerve transplants which combine a synthetic conduit with autologous SC genetically modified to express regeneration-promoting proteins may provide an alternative therapy to autotransplantation. The dog seems to be an ideal translational animal model for new treatment strategies. In the present study, utilizing a new transfection protocol, we transplanted enhanced green fluorescent protein (EGFP)-expressing adult canine SC (cSC) into a 5mm epineural pouch in the sciatic nerve of adult rats (n=9). The epineurial pouch technique serves as proof of principle to follow the fate of the transplanted cSC for up to 14 days after surgery. Fluorescence microscopy and immunohistochemistry revealed survival and integration of EGFP-expressing cSC into the regenerating host nerve tissue. We demonstrate that transplanted cSC contribute to the formation of bands of Bungner and are located in close vicinity to growth-associated protein-43 (GAP-43) expressing regenerating nerve fibers. This provides first evidence that transplanted genetically modified Schwann cells do successfully integrate into the host tissue where they could actively contribute to the regeneration process.

post to: CiteULike

A criterion for signal-based selection of wavelets for denoising intrafascicular nerve recordings.

Publication Date: 2010 Feb 15 PMID: 19962403
Authors: Kamavuako, E. N. - Jensen, W. - Yoshida, K. - Kurstjens, M. - Farina, D.
Journal: J Neurosci Methods

In this paper we propose a novel method for denoising intrafascicular nerve signals with the aim of improving action potential (AP) detection. The method is based on the stationary wavelet transform and thresholding of the wavelet coefficients. Since the choice of the mother wavelet substantially impact the performance, a criterion is proposed for selecting the optimal wavelet. The criterion for selection was based on the root mean square of the average of the output signal triggered by the detected APs. The mother wavelet was parameterized through the scaling filter, which allowed optimization through the proposed criterion. The method was tested on simulated signals and on experimental neural recordings. Experimental signals were recorded from the tibial branch of the sciatic nerve of three anaesthetized New Zealand white rabbits during controlled muscle stretches. The simulation results showed that the proposed method had an equivalent effect on AP detection performance (percentage of correct detection at 6 dB signal-to-noise ratio, mean+/-SD, 95.3+/-5.2%) to the a-posteriori choice of the best wavelet (96.1+/-3.6). Moreover, the AP detection after the proposed denoising method resulted in a correlation of 0.94+/-0.02 between the estimated spike rate and the muscle length. Therefore, the study proposes an effective method for selecting the optimal mother wavelet for denoising neural signals with the aim of improving AP detection.

post to: CiteULike

A MATLAB toolbox for Granger causal connectivity analysis.

Publication Date: 2010 Feb 15 PMID: 19961876
Authors: Seth, A. K.
Journal: J Neurosci Methods

Assessing directed functional connectivity from time series data is a key challenge in neuroscience. One approach to this problem leverages a combination of Granger causality analysis and network theory. This article describes a freely available MATLAB toolbox--'Granger causal connectivity analysis' (GCCA)--which provides a core set of methods for performing this analysis on a variety of neuroscience data types including neuroelectric, neuromagnetic, functional MRI, and other neural signals. The toolbox includes core functions for Granger causality analysis of multivariate steady-state and event-related data, functions to preprocess data, assess statistical significance and validate results, and to compute and display network-level indices of causal connectivity including 'causal density' and 'causal flow'. The toolbox is deliberately small, enabling its easy assimilation into the repertoire of researchers. It is however readily extensible given proficiency with the MATLAB language.

post to: CiteULike

Signalling effect of NIR pulsed lasers on axonal growth.

Publication Date: 2010 Feb 15 PMID: 19945486
Authors: Mathew, M. - Amat-Roldan, I. - Andres, R. - Santos, S. I. - Artigas, D. - Soriano, E. - Loza-Alvarez, P.
Journal: J Neurosci Methods

In this work we show that a pulsed laser light placed at a distance is able to modulate the growth of axons of primary neuronal cell cultures. In our experiments continuous wave (CW), chopped CW and modelocked fs (FS) laser light was focused through a microscope objective to a point placed at a distance of about 15 microm from the growth cone. We found that CW light does not produce any significant influence on the axon growth. In contrast, when using pulsed light (chopped CW light or FS pulses), the beam was able to modify the trajectory of the axons, attracting approximately 45% of the observed cases to the beam spot. Such effect could possibly indicate the capacity of neurons to interpret the pulsating NIR light as the source of other nearby cells, resulting in extension of processes in the direction of the source.

post to: CiteULike

A cognitive neuroscience-based computerized battery for efficient measurement of individual differences: Standardization and initial construct validation.

Publication Date: 2010 Mar 30 PMID: 19945485
Authors: Gur, R. C. - Richard, J. - Hughett, P. - Calkins, M. E. - Macy, L. - Bilker, W. B. - Brensinger, C. - Gur, R. E.
Journal: J Neurosci Methods

There is increased need for efficient computerized methods to collect reliable data on a range of cognitive domains that can be linked to specific brain systems. Such need arises in functional neuroimaging studies, where individual differences in cognitive performance are variables of interest or serve as confounds. In genetic studies of complex behavior, which require particularly large samples, such trait measures can serve as endophenotypes. Traditional neuropsychological tests, based on clinical pathological correlations, are protracted, require extensive training in administration and scoring, and leave lengthy paper trails (double-entry for analysis). We present a computerized battery that takes an average of 1h and provides measures of accuracy and speed on 9 neurocognitive domains. They are cognitive neuroscience-based in that they have been linked experimentally to specific brain systems with functional neuroimaging studies. We describe the process of translating tasks used in functional neuroimaging to tests for assessing individual differences. Data are presented on each test with samples ranging from 139 (81 female) to 536 (311 female) of carefully screened healthy individuals ranging in age from 18 to 84. Item consistency was established with acceptable to high Cronbach alpha coefficients. Inter-item correlations were moderate to high within domain and low to nil across domains, indicating construct validity. Initial criterion validity was demonstrated by sensitivity to sex differences and the effects of age, education and parental education. These results encourage the use of this battery in studies needing an efficient assessment of major neurocognitive domains such as multi-site genetic studies and clinical trials.

post to: CiteULike

A mechanomyographic frequency-based fatigue threshold test.

Publication Date: 2010 Mar 15 PMID: 19945484
Authors: Hendrix, C. R. - Housh, T. J. - Zuniga, J. M. - Camic, C. L. - Mielke, M. - Johnson, G. O. - Schmidt, R. J.
Journal: J Neurosci Methods

Theoretically, the mechanomyographic (MMG) mean power frequency fatigue threshold (MMG MPF(FT)) describes the maximal isometric torque that can be maintained for an extended period of time with no change in the global firing rate of the unfused, activated motor units. PURPOSE: The purposes of this study were twofold: (1) to determine if the mathematical model for estimating the electromyographic (EMG) MPF(FT) from the frequency of the EMG signal was applicable to the frequency domain of the MMG signal to estimate a new fatigue threshold called the MMG MPF(FT); and (2) to compare the mean torque levels derived from the MMG MPF(FT) test for the vastus lateralis (VL), vastus medialis (VM), and rectus femoris (RF) muscles during isometric leg extension muscle actions. METHODS: Nine adults (4 men and 5 women; mean+/-S.D. age=21.6+/-1.2 years) performed three or four continuous, fatiguing, isometric muscle actions of the leg extensors at 30, 45, 60, and 75% of maximum voluntary isometric contraction (MVIC) to exhaustion. Surface MMG signals were recorded from the VL, VM, and RF muscles during each fatiguing isometric muscle action. The MMG MPF(FT) was defined as the y-intercept of the isometric torque versus slope coefficient (MMG MPF versus time) plot. RESULTS: There were no significant differences among the MMG MPF(FT) values for the VL, VM, and RF (34.8+/-23.4, 32.1+/-16.1, and 31.6+/-15.2 Nm, respectively) muscles. CONCLUSION: The MMG MPF(FT) test may provide a non-invasive method to examine the effects of various interventions on the global motor unit firing rate during isometric muscle actions.

post to: CiteULike

In vivo gene delivery to the postnatal ferret cerebral cortex by DNA electroporation.

Publication Date: 2010 Feb 15 PMID: 19944720
Authors: Borrell, V.
Journal: J Neurosci Methods

Ferrets have been extensively used to unravel the neural mechanisms of coding and processing of visual information, and also to identify the developmental mechanisms underlying the emergence of such a complex and fine-tuned neural system. In recent years numerous tools have been generated that allow studying neural systems with unprecedented power. Unfortunately, because many of these tools are genetically encoded, they are having a limited impact on research involving "non-genetic" species, like ferret, cat and monkey. Here I show how in vivo electroporation can be performed in postnatal ferret kits to deliver genetic constructs to pyramidal neurons of the cerebral cortex. Electroporation of GFP- and DsRed-encoding plasmids results in labeling of cortical progenitors first, then migrating neurons, and finally differentiating neurons and their processes. This technique also allows for the genetic manipulation of cortical development in the ferret, as illustrated by electroporation of a dominant-negative form of Cdk5. In the mature brain of electroporated animals, expression of reporter genes reveals the detailed morphological traits of cortical pyramids, including their axonal and dendritic arborization, and dendritic spines. I also show that postnatal electroporation can be used for the transfection of a massive cortical territory, or it can be specifically directed to a subset of cortical areas, and even only to a few scattered pyramids along the cortical mantle. In vivo electroporation of postnatal ferrets is therefore an effective, rapid, simple and highly versatile method for delivering genetic constructs to this animal, optimal for both developmental studies and adult anatomical/functional studies.

post to: CiteULike

Sensory driven multi-neuronal activity and associative learning monitored in an intact CNS on a multielectrode array.

Publication Date: 2010 Feb 15 PMID: 19941897
Authors: Harris, C. A. - Passaro, P. A. - Kemenes, I. - Kemenes, G. - O'Shea, M.
Journal: J Neurosci Methods

The neuronal network controlling feeding behavior in the CNS of the mollusc Lymnaea stagnalis has been extensively investigated using intracellular microelectrodes. Using microelectrodes however it has not been possible to record from large numbers of neurons simultaneously and therefore little is known about the population coding properties of the feeding network. Neither can the relationships between feeding and neuronal networks controlling other behaviors be easily analyzed with microelectrodes. Here we describe a multielectrode array (MEA) technique for recording action potentials simultaneously from up to 60 electrodes on the intact CNS. The preparation consists of the whole CNS connected by sensory nerves to the chemosensory epithelia of the lip and esophagus. From the buccal ganglia, the region of the CNS containing the feeding central pattern generator (CPG), a rhythmic pattern of activity characteristic of feeding was readily induced either by depolarizing an identified feeding-command neuron (the CV1a) or by perfusing the chemosensory epithelia with sucrose, a gustatory stimulus known to activate feeding. Activity induced by sucrose is not restricted to the buccal ganglia but is distributed widely throughout the CNS, notably in ganglia controlling locomotion, a behavior that must be coordinated with feeding. The MEA also enabled us to record electrophysiological consequences of the associative conditioning of feeding behavior. The results suggest that MEA recording from an intact CNS enables distributed, multiple-source neural activity to be analyzed in the context of biologically relevant behavior, behavioral coordination and behavioral plasticity.

post to: CiteULike

Correction of low-frequency physiological noise from the resting state BOLD fMRI--Effect on ICA default mode analysis at 1.5 T.

Publication Date: 2010 Feb 15 PMID: 19941896
Authors: Starck, T. - Remes, J. - Nikkinen, J. - Tervonen, O. - Kiviniemi, V.
Journal: J Neurosci Methods

Confounding low-frequency fluctuation (LFF) physiological noise is a concern for functional connectivity analyses in blood oxygen level-dependent (BOLD) functional magnetic resonance imaging (fMRI). Using estimates of LFF physiological noise derived from measured cardiac and respiration signals, noise can be filtered from the time series thus improving the results of functional connectivity analysis. The ability of spatial independent component analysis (ICA) to separate LFF physiological noise from the default mode network (DMN), which overlap each other spatially and occur at similar frequencies, has remained an open question. We aimed to define the net effect of physiological correction for spatial ICA DMN detection at 1.5 T by statistically testing obtained ICASSO centrotype DMN maps before and after physiological correction. Comparisons with 21 subjects were performed for ICA model orders 20, 30 and 40 and no statistically significant spatial difference was found after physiological correction, although slight DMN reduction in precuneus or sagittal sinus was detected in all dimensionalities. A confounding factor in the analysis is the susceptibility of the ICA decomposition for data changes yielding different DMN splitting between and after physiological correction conditions without comparable true change in the data. This issue is mitigated at higher ICA model orders. The results suggest that subject-level DMN can for some subjects be optimized by physiological correction, but on the group-level this contribution is minor.

post to: CiteULike

It is darkness and not light: Depression-like behaviors of diurnal unstriped Nile grass rats maintained under a short photoperiod schedule.

Publication Date: 2010 Feb 15 PMID: 19932714
Authors: Ashkenazy-Frolinger, T. - Kronfeld-Schor, N. - Juetten, J. - Einat, H.
Journal: J Neurosci Methods

Circadian rhythms are strongly implicated in affective disorders and some recent studies suggested that diurnal rodents might be advantageous model animals for them. In line with this possibility, previous work demonstrated that in the diurnal fat sand rat, short photoperiod conditions result in depression- and anxiety-like behavioral phenotype that is relieved with bright light treatment. To further explore the possibility of using diurnal species as model animals for affective disorders, the present study examined the effects of short photoperiod schedule in an additional diurnal rodent, the unstriped Nile grass rat. Results indicate that 6 weeks short photoperiod (5 h light/19 h dark) regimen induced depression-like behavior in the forced swim test and the saccharin preference test compared with animals maintained in a neutral photoperiod regimen (12 h light/12 h dark). No effects were shown in the light/dark box model of anxiety or in a test for spontaneous activity. These results demonstrate that photoperiod manipulations in diurnal rodents induce affective-like behavioral change and support the possibility that diurnal rodents might provide a good potential as model animals for depression spectrum disorders.

post to: CiteULike

Avoiding the ballistocardiogram (BCG) artifact of EEG data acquired simultaneously with fMRI by pulse-triggered presentation of stimuli.

Publication Date: 2010 Feb 15 PMID: 19931564
Authors: Ertl, M. - Kirsch, V. - Leicht, G. - Karch, S. - Olbrich, S. - Reiser, M. - Hegerl, U. - Pogarell, O. - Mulert, C.
Journal: J Neurosci Methods

Simultaneous acquisition of electroencephalography (EEG) and functional magnetic resonance imaging (fMRI) data could offer a much deeper understanding of brain function, e.g. in the analysis of tempo-spatial dynamics of brain activity in cognitive processing. However, more sophisticated analysis methods such as single-trial coupling of EEG and fMRI are often handicapped by the limited quality of EEGs acquired in the MRI scanner. In particular, the ballistocardiogram (BCG) artifact is still a relevant problem. Methods that are currently available typically remove the BCG artifact either in post-recording or real-time signal processing. Here, we would like to suggest a new strategy to avoid BCG artifacts during data acquisition. In our proposal, stimuli are presented pulse-triggered (PT), thus avoiding interference of BCG artifacts with the evoked potentials investigated during EEG recording. This method is based on the observation that the main influence of the BCG artifact is generally limited to the time interval of 150-500 ms post-QRS complex. Based on real measurements, we simulated different signal presentation methods relative to the onset of the BCG artifact for 14 subjects. Stimuli were either presented independently of the BCG artifact or pulse-triggered at fixed time-points (280 ms, 480 ms and 680 ms post-QRS complex) and with a jitter (short: 120 ms or long: 240 ms). In combination with an averaged artifact subtraction method signal distortion was reduced at best by 47%.

post to: CiteULike

Characterization of trial-to-trial fluctuations in local field potentials recorded in cerebral cortex of awake behaving macaque.

Publication Date: 2010 Feb 15 PMID: 19931563
Authors: Menzer, D. L. - Bokil, H. - Ryou, J. W. - Schiff, N. D. - Purpura, K. P. - Mitra, P. P.
Journal: J Neurosci Methods

In analyzing neurophysiologic data, individual experimental trials are usually assumed to be statistically independent. However, many studies employing functional imaging and electrophysiology have shown that brain activity during behavioral tasks includes temporally correlated trial-to-trial fluctuations. This could lead to spurious results in statistical significance tests used to compare data from different interleaved behavioral conditions presented throughout an experiment. We characterize trial-to-trial fluctuations in local field potentials recorded from the frontal cortex of a macaque monkey performing an oculomotor delayed response task. Our analysis identifies slow fluctuations (<0.1 Hz) of spectral power in 22/27 recording sessions. These trial-to-trial fluctuations are non-Gaussian, and call into question the statistical utility of standard trial shuffling. We compare our results with evidence for slow fluctuations in human functional imaging studies and other electrophysiologic studies in nonhuman primates.

post to: CiteULike

Use of fNIRS to assess resting state functional connectivity.

Publication Date: 2010 Feb 15 PMID: 19931310
Authors: Lu, C. M. - Zhang, Y. J. - Biswal, B. B. - Zang, Y. F. - Peng, D. L. - Zhu, C. Z.
Journal: J Neurosci Methods

Recently, resting state functional connectivity (RSFC) studies based on fMRI and EEG/MEG have provided valuable insight into the intrinsic functional architecture of the human brain. However, whether functional near infrared spectroscopy (fNIRS), a suitable imaging method for infant and patient populations, can be used to examine RSFC remains elusive. Using an ETG-4000 Optical Topography System, the present study measured 29 adult subjects (14 females) over the sensorimotor and auditory cortexes during a resting session and a motor-localizer task session. The RSFC maps were computed by seed-based correlation analysis and data-driven cluster analysis. The results from both analyses showed robust RSFC maps, which were not only consistent with the localizer task-related activation results, but also those of previous fMRI findings. Moreover, the strong consistency between the seed-based correlation analysis and the data-driven cluster analysis further validated the use of fNIRS to assess RSFC. The potential influence of a specific low-frequency filtering range (0.04-0.15 Hz and 0.01-0.08 Hz) and three fNIRS parameters (oxy-Hb, deoxy-Hb, and total-Hb) on RSFC results were also examined.

post to: CiteULike

A combined method of laser capture microdissection and X-Gal histology to analyze gene expression in c-Fos-specific neurons.

Publication Date: 2010 Feb 15 PMID: 19925827
Authors: Kwon, B. - Houpt, T. A.
Journal: J Neurosci Methods

c-Fos is a member of the activator protein 1 family that regulates transcription of target genes. c-Fos is transiently induced in specific regions of the brain after a variety of external stimuli including learning and memory formation. Analysis of gene expression in c-Fos-expressing cells of the brain may help identify target genes that play important roles in synaptic strength or neuronal morphology. In the present study, we developed a combined method of laser capture microdissection and 5-bromo-4-chloro-3-indoly-beta-D-galactopyranosidase (X-Gal) histology to analyze gene expression in stimulus-induced c-Fos-positive cells. Using transgenic mice carrying a c-fos-lacZ fusion gene, c-Fos-positive cells were easily identified by measuring of beta-galactosidase (beta-Gal) activity. To establish the fidelity of the reporter transgene, the time course of endogenous c-Fos and the c-fos-lacZ transgene expression in the amygdala induced by LiCl administration was investigated by immunohistochemistry and X-Gal staining. LiCl increased the numbers of c-Fos- and beta-Gal-positive cells in the central and basolateral amygdala of the transgenic mice. To ensure that RNA was preserved in X-Gal stained tissue sections, different fixations were examined, with the conclusion that ethanol fixation was best for both RNA preservation and X-Gal staining quality. Finally, in combining X-Gal staining, single-cell LCM and RT-PCR, we confirmed mRNA expression of endogenous c-fos and beta-actin genes in LiCl-induced beta-Gal-positive cells in the CeA, cortex and hippocampus. Combining LCM and transgenic reporter genes provides a powerful tool with which to investigate tissue- or cell-specific gene expression.

post to: CiteULike

Correlation of locomotor activity and brain infarction in rats with transient focal ischemia.

Publication Date: 2010 Feb 15 PMID: 19917312
Authors: Shen, H. - Wang, Y.
Journal: J Neurosci Methods

Occlusion of middle cerebral artery (MCAo) is commonly used to generate stroke in experimental animals. Different behavioral assays have been used to evaluate the severity of lesions or recovery after treatment in this model. In this study, we examined the correlation between the size of infarction and locomotor activity after transient MCAo in adult Sprague-Dawley rats. The right middle cerebral artery was occluded for 30-90-min by ligation with a 10-O suture. At 2 days after MCAo, animals were individually placed in a 3-D infrared activity chamber for 1h to measure their horizontal, vertical, stereotypic and rotational activity. Body asymmetry was examined using an elevated body swing test. Volume of infarction was measured by triphenyltetrazolium chloride staining. We found that there is a significant correlation between the volume of infarction and vertical movement, such as vertical activity, vertical movement number and vertical movement time. There is also a significant correlation between the infarction volume and body asymmetry. In conclusion, our data suggest that vertical locomotor activity and body asymmetry are useful behavioral indices to estimate the size of lesioning in rodents after MCAo.

post to: CiteULike

A simple protocol for assessing inter-trial and inter-examiner reliability for two noninvasive measures of limb muscle strength.

Publication Date: 2010 Feb 15 PMID: 19917311
Authors: Carlson, C. G. - Rutter, J. - Bledsoe, C. - Singh, R. - Hoff, H. - Bruemmer, K. - Sesti, J. - Gatti, F. - Berge, J. - McCarthy, L.
Journal: J Neurosci Methods

Noninvasive measures of limb muscle strength are quite useful in preclinical translational studies that use mouse models of muscle disease, peripheral nerve disease, and movement disorders. The present study uses a simple protocol for assessing both inter-trial and inter-examiner reliability for two noninvasive methods of assessing limb strength in dystrophic (mdx) and wild type mice. One method, termed the whole body tension (WBT) method or escape test, measures the total phasic pulling tension exerted by the fore- and hindlimbs while a mouse attempts to escape into a darkened tube. Another procedure, termed the four limb wire grid holding test, measures the minimal amount of sustained tension (physical impulse) exerted by the fore- and hindlimbs while the mouse hangs suspended in an upside-down position. A comparison of the two methods revealed significant inter-trial and inter-examiner correlations in each procedure, although the WBT procedure consistently produced higher correlations than the four limb wire grid holding test. Inter-trial reliability for each test was higher than inter-examiner reliability, indicating that each longitudinal series of tests is best performed by a single investigator. The holding test also did not consistently detect differences between wild type and mdx populations at ages greater than 4 months. These results demonstrate the utility of a simple protocol for assessing the reliability of noninvasive tests that measure limb strength, and should be useful in comparing different functional measures in a broad range of translational studies.

post to: CiteULike

Alteration of Methamphetamine-induced stereotypic behaviour in transgenic mice expressing HIV-1 envelope protein gp120.

Publication Date: 2010 Feb 15 PMID: 19917310
Authors: Roberts, A. J. - Maung, R. - Sejbuk, N. E. - Ake, C. - Kaul, M.
Journal: J Neurosci Methods

The use of drugs for recreational purposes, in particular Methamphetamine, is associated with an increased risk of infection with human immunodeficiency virus (HIV)-1. HIV-1 infection in turn can lead to HIV-associated neurological disorders (HAND) that range from mild cognitive and motor impairment to HIV-associated dementia (HAD). Interestingly, post mortem brain specimens from HAD patients and transgenic (tg) mice expressing the viral envelope protein gp120 in the central nervous system display similar neuropathological signs. In HIV patients, the use of Methamphetamine appears to aggravate neurocognitive alterations. In the present study, we injected HIV/gp120tg mice and non-transgenic littermate control animals with Methamphetamine dissolved in Saline or Saline vehicle and assessed locomotion and stereotyped behaviour. We found that HIVgp120-transgenic mice differ significantly from non-transgenic controls in certain domains of their behavioural response to Methamphetamine. Thus this experimental model system may be useful to further study the mechanistic interaction of both the viral envelope protein and the psychostimulant drug in behavioural alterations and neurodegenerative disease.

post to: CiteULike

Brain penetration of local anaesthetics in the rat: Implications for experimental neuroscience.

Publication Date: 2010 Feb 15 PMID: 19917309
Authors: Ferrari, L. - Crestan, V. - Sabattini, G. - Vinco, F. - Fontana, S. - Gozzi, A.
Journal: J Neurosci Methods

Multiple experimental neuroscience techniques rely on the use of general anaesthesia to minimize the discomfort associated to animal restraint and to achieve a more effective control of relevant physiological parameters. In order to minimise potential interference on brain neuronal activity, such studies are typically conducted at low anaesthetic doses. This practice is often coupled to peripheral infiltration of local anaesthetics to provide supplementary analgesia and prevent sub-threshold activation of pain pathways that may confound central measurements of brain function. However, little is known of the effect of peripheral anaesthesia on central measurements of brain activity in small laboratory animal species. In order to begin to address this question, we measured total and free brain exposure of five different local anaesthetics following subcutaneous infiltration of analgesic doses in a surgical protocol widely used in rodent neuroimaging and electrophysiology studies. Notably, all the anaesthetics exhibited detectable total and free brain concentrations at all the time points examined. Lidocaine and mepivacaine showed the highest free brain exposures (>525 ng/g), followed by bupivacaine and ropivacaine (>70 ng/g). The ester-type local anaesthetic tetracaine produced the lowest free brain exposure (<8.6 ng/g). Our data suggest that peripheral administration of local anaesthetics in small laboratory animals could result in pharmacologically active brain exposures that might influence and confound central measurements of brain function. The use of the ester-type anaesthetic tetracaine produced considerably lower brain exposure, and may represent a viable experimental option when local anaesthesia is required.

post to: CiteULike