Interstitial cells of Cajal mediate inhibitory neurotransmission in the stomach.

The structural relationships between interstitial cells of Cajal (ICC), varicose nerve fibers, and smooth muscle cells in the gastrointestinal tract have led to the suggestion that ICC may be involved in or mediate enteric neurotransmission. We characterized the distribution of ICC in the murine stomach and found two distinct classes on the basis of morphology and immunoreactivity to antibodies against c-Kit receptors. ICC with multiple processes formed a network in the myenteric plexus region from corpus to pylorus. Spindle-shaped ICC were found within the circular and longitudinal muscle layers (IC-IM) throughout the stomach. The density of these cells was greatest in the proximal stomach. IC-IM ran along nerve fibers and were closely associated with nerve terminals and adjacent smooth muscle cells. IC-IM failed to develop in mice with mutations in c-kit. Therefore, we used W/W(V) mutants to test whether IC-IM mediate neural inputs in muscles of the gastric fundus. The distribution of inhibitory nerves in the stomachs of c-kit mutants was normal, but NO-dependent inhibitory neuro-regulation was greatly reduced. Smooth muscle tissues of W/W(V) mutants relaxed in response to exogenous sodium nitroprusside, but the membrane potential effects of sodium nitroprusside were attenuated. These data suggest that IC-IM play a critical serial role in NO-dependent neurotransmission: the cellular mechanism(s) responsible for transducing NO into electrical responses may be expressed in IC-IM. Loss of these cells causes loss of electrical responsiveness and greatly reduces responses to nitrergic nerve stimulation.

P2Y1 purinergic receptors in sensory neurons: contribution to touch-induced impulse generation.

Somatic sensation requires the conversion of physical stimuli into the depolarization of distal nerve endings. A single cRNA derived from sensory neurons renders Xenopus laevis oocytes mechanosensitive and is found to encode a P2Y1 purinergic receptor. P2Y1 mRNA is concentrated in large-fiber dorsal root ganglion neurons. In contrast, P2X3 mRNA is localized to small-fiber sensory neurons and produces less mechanosensitivity in oocytes. The frequency of touch-induced action potentials from frog sensory nerve fibers is increased by the presence of P2 receptor agonists at the peripheral nerve ending and is decreased by the presence of P2 antagonists. P2X-selective agents do not have these effects. The release of ATP into the extracellular space and the activation of peripheral P2Y1 receptors appear to participate in the generation of sensory action potentials by light touch.

Differential distribution of two ATP-gated channels (P2X receptors) determined by immunocytochemistry.

Several P2X receptor subunits were recently cloned; of these, one was cloned from the rat vas deferens (P2X1) and another from pheochromocytoma (PC12) cells differentiated with nerve growth factor (P2X2). Peptides corresponding to the C-terminal portions of the predicted receptor proteins (P2X1 391-399 and P2X2 460-472) were used to generate antisera in rabbits. The specificities of antisera were determined by staining human embryonic kidney cells stably transfected with either P2X1 or P2X2 receptors and by absorption controls with the cognate peptides. In the vas deferens and the ileal submucosa, P2X1 immunoreactivity (ir) was restricted to smooth muscle, whereas P2X2-ir was restricted to neurons and their processes. Chromaffin cells of the adrenal medulla and PC12 cells contained both P2X1- and P2X2-ir. P2X1-ir was also found in smooth muscle cells of the bladder, cardiac myocytes, and nerve fibers and terminals in the superficial dorsal horn of the spinal cord. In contrast, P2X2-ir was observed in scattered cells of the anterior pituitary, neurons in the hypothalamic arcuate and paraventricular nuclei, and catecholaminergic neurons in the olfactory bulb, the substantia nigra, ventral tegmental area, and locus coeruleus. A plexus of nerve fibers and terminals in the nucleus of the solitary tract contained P2X2-ir. This staining disappeared after nodose ganglionectomy, consistent with a presynaptic function. The location of the P2X1 subunit in smooth muscle is consistent with its role as a postjunctional receptor in autonomic transmission, while in neurons, these receptors appear in both postsynaptic and presynaptic locations.

Multineuronal codes in retinal signaling.

The visual world is presented to the brain through patterns of action potentials in the population of optic nerve fibers. Single-neuron recordings show that each retinal ganglion cell has a spatially restricted receptive field, a limited integration time, and a characteristic spectral sensitivity. Collectively, these response properties define the visual message conveyed by that neuron's action potentials. Since the size of the optic nerve is strictly constrained, one expects the retina to generate a highly efficient representation of the visual scene. By contrast, the receptive fields of nearby ganglion cells often overlap, suggesting great redundancy among the retinal output signals. Recent multineuron recordings may help resolve this paradox. They reveal concerted firing patterns among ganglion cells, in which small groups of nearby neurons fire synchronously with delays of only a few milliseconds. As there are many more such firing patterns than ganglion cells, such a distributed code might allow the retina to compress a large number of distinct visual messages into a small number of optic nerve fibers. This paper will review the evidence for a distributed coding scheme in the retinal output. The performance limits of such codes are analyzed with simple examples, illustrating that they allow a powerful trade-off between spatial and temporal resolution.

Localization of vasopressin mRNA and immunoreactivity in pituicytes of pituitary stalk-transected rats after osmotic stimulation.

The presence of [arginine] vasopressin (AVP) mRNA and AVP immunoreactivity in pituicytes of the neural lobe (NL) of intact and pituitary stalk-transected rats, with and without osmotic stimulation, was examined. AVP mRNA was analyzed by Northern blotting, as well as by in situ hybridization in combination with immunocytochemistry using anti-glial fibrillary acidic protein (GFAP) as a marker for pituicytes. In intact rats, a poly(A) tail-truncated 0.62-kb AVP mRNA was detected in the NL and was found to increase 10-fold with 7 days of continuous salt loading. Morphological analysis of the NL of 7-day salt-loaded rats revealed the presence of AVP mRNA in a significant number of GFAP-positive pituicytes in the NL and in areas most probably containing nerve fibers. Eight days after pituitary stalk transection the NL AVP mRNA diminished in animals given water to drink, whereas in those given 2% saline for 18 h followed by 6 h of water, a treatment repeated on 6 successive days beginning 2 days after surgery, the 0.62-kb AVP mRNA was present. The AVP mRNA in the pituitary stalk-transected, salt-loaded rats showed an exclusive cellular distribution in the NL, indicative of localization in pituicytes. Immunoelectron microscopy showed the presence of AVP immunoreactivity in a subpopulation of pituicytes 7 and 10 days after pituitary stalk transection in salt-loaded animals, when almost all AVP fibers had disappeared from the NL. These data show that a subset of pituicytes in the NL is activated to synthesize AVP mRNA and AVP in response to osmotic stimulation.

Phosphorylated α-synuclein-immunoreactive retinal neuronal elements in Parkinson's disease subjects

Visual symptoms are relatively common in Parkinson's disease (PD) and optical coherence tomography has indicated possible retinal thinning. Accumulation of aggregated α-synuclein is thought to be a central pathogenic event in the PD brain but there have not as yet been reports of retinal synucleinopathy. Retinal wholemounts were prepared from subjects with a primary clinicopathological diagnosis of PD (N = 9), dementia with Lewy bodies (DLB; N = 3), Alzheimer's disease (N = 3), progressive supranuclear palsy (N = 2) as well as elderly normal control subjects (N = 4). These were immunohistochemically stained with an antibody against α-synuclein phosphorylated at serine 129, which is a specific molecular marker of synucleinopathy. Phosphorylated α-synuclein-immunoreactive (p-syn IR) nerve fibers were present in 7/9 PD subjects and in 1/3 DLB subjects; these were sparsely distributed and superficially located near or at the inner retinal surface. The fibers were either long and straight or branching, often with multiple en-passant varicosities along their length. The straight fibers most often had an orientation that was radial with respect to the optic disk. Together, these features are suggestive of either retinopetal/centrifugal fibers or of ganglion cell axons. In one PD subject there were sparse p-syn IR neuronal cell bodies with dendritic morphology suggestive of G19 retinal ganglion cells or intrinsically photosensitive ganglion cells. There were no stained nerve fibers or other specific staining in any of the non-PD or non-DLB subjects. It is possible that at least some of the observed visual function impairments in PD subjects might be due to α-synucleinopathy.

Quantitative analysis of immunolabeling for serotonin and for glutamate transporters after administration of imipramine and citalopram

Serotonin (5-hydroxytryptamine, 5-HT) is an amine neurotransmitter derived from tryptophan and is important in brain systems regulating mood, emotional behavior, and sleep. Selective serotonin reuptake inhibitor (SSRI) drugs are used to treat disorders such as depression, stress, eating disorders, autism, and schizophrenia. It is thought that these drugs act to prolong the action of 5-HT by blocking reuptake. This may lead to decreased 5-HT content in the nerve fibers themselves; however, this has not previously been directly demonstrated. We have studied the effects of administration of two drugs, imipramine and citalopram, on levels of 5-HT in nerve fibers in the murine brain. Quantitative analysis of the areal density of 5-HT fibers throughout the brain was performed using ImageJ software. While a high density of fibers was observed in mid- and hind-brain regions and areas such as thalamus and hypothalamus, densities were far lower in areas such as cortex, where SSRIs might be thought to exert their actions. As anticipated, imipramine and citalopram produced a decline in 5-HT levels in nerve fibers, but the result was not uniform. Areas such as inferior colliculus showed significant reduction whereas little, if any, change was observed in the adjacent superior colliculus. The reason for, and significance of, this regionality is unclear. It has been proposed that serotonin effects in the brain might be linked to changes in glutamatergic transmission. Extracellular glutamate levels are regulated primarily by glial glutamate transporters. Qualitative evaluation of glutamate transporter immunolabeling in cortex of control and drug-treated mice revealed no discernable difference in intensity of glutamate transporter immunoreactivity. These data suggest that changes in intracellular and extracellular levels of serotonin do not cause concomitant changes in astroglial glutamate transporter expression, and thus cannot represent a mechanism for the delayed efficacy of antidepressants when administered clinically. © 2005 Elsevier B.V. All rights reserved.

The number, morphology, and distribution of retinal ganglion cells and optic axons in the Australian lungfish Neoceratodus forsteri (Krefft 1870)

Australian lungfish Neoceratodus forsteri may be the closest living relative to the first tetrapods and yet little is known about their retinal ganglion cells. This study reveals that lungfish possess a heterogeneous population of ganglion cells distributed in a horizontal streak across the retinal meridian, which is formed early in development and maintained through to adult stages. The number and complement of both ganglion cells and a population of putative amacrine cells within the ganglion cell layer are examined using retrograde labelling from the optic nerve and transmission electron-microscopic analysis of axons within the optic nerve. At least four types of retinal ganglion cells are present and lie predominantly within a thin ganglion cell layer, although two subpopulations are identified, one within the inner plexiform and the other within the inner nuclear layer. A subpopulation of retinal ganglion cells comprising up to 7% or the total population are significantly larger (> 400 mu m(2)) and are characterized as giant or alpha-like cells. Up to 44% of cells within the retinal ganglion cell layer represent a population of presumed amacrine cells. The optic nerve is heavily fasciculated and the proportion of myelinated axons increases with body length from 17% in subadults to 74% in adults. Spatial resolving power, based on ganglion cell spacing, is low (1.6-1.9 cycles deg(-1), n = 2) and does not significantly increase with growth. This represents the first detailed study of retinal ganglion cells in sarcopterygian fish, and reveals that, despite variation amongst animal groups, trends in ganglion cell density distribution and characteristics of cell types were defined early in vertebrate evolution.

The presence of pleiotrophin in the human intervertebral disc is associated with increased vascularization:an immunohistologic study

Study Design. An immunohistological study of surgical specimens of human intervertebral disc.Objective.To examine the presence of pleiotrophin in diseased or damaged intervertebral disc tissue and the association between its presence and the extent of tissue vascularization and innervation.Summary of Background Data. Increased levels of pleiotrophin, a growth and differentiation factor that is active in various pathophysiologic processes, including angiogenesis, has been associated with osteoarthritic changes of human articular cartilage. The association between pleiotrophin expression and pathologic conditions of the human intervertebral disc is unknown.Methods. Specimens of human lumbar intervertebral discs, obtained following surgical discectomy, were divided into 3 groups: nondegenerated discs (n = 7), degenerated discs (n = 6), and prolapsed discs (n = 11). Serial tissue sections of each specimen were immunostained to determine the presence of pleiotrophin, blood vessels (CD34-positive endothelial cells), and nerves (neurofilament 200 kDa [NF200]-positive nerve fibers).Results. Pleiotrophin immunoreactivity was seen in disc cells, endothelial cells, and in the extracellular matrix in most specimens of intervertebral disc but was most prevalent in vascularized tissue in prolapsed discs. There was a significant correlation between the presence of pleiotrophin-positive disc cells and that of CD34-positive blood vessels. NF200-positive nerves were seen in vascularized areas of more degenerated discs, but nerves did not appear to codistribute with blood vessels or pleiotrophin positivity in prolapsed discs.Conclusions. Pleiotrophin is present in pathologic human intervertebral discs, and its prevalence and distribution suggest that it may play a role in neovascularization of diseased or damaged disc tissue.

The effect of Gaussian noise on the threshold, dynamic range, and loudness of analogue cochlear implant stimuli

The deliberate addition of Gaussian noise to cochlear implant signals has previously been proposed to enhance the time coding of signals by the cochlear nerve. Potentially, the addition of an inaudible level of noise could also have secondary benefits: it could lower the threshold to the information-bearing signal, and by desynchronization of nerve discharges, it could increase the level at which the information-bearing signal becomes uncomfortable. Both these effects would lead to an increased dynamic range, which might be expected to enhance speech comprehension and make the choice of cochlear implant compression parameters less critical (as with a wider dynamic range, small changes in the parameters would have less effect on loudness). The hypothesized secondary effects were investigated with eight users of the Clarion cochlear implant; the stimulation was analogue and monopolar. For presentations in noise, noise at 95% of the threshold level was applied simultaneously and independently to all the electrodes. The noise was found in two-alternative forced-choice (2AFC) experiments to decrease the threshold to sinusoidal stimuli (100 Hz, 1 kHz, 5 kHz) by about 2.0 dB and increase the dynamic range by 0.7 dB. Furthermore, in 2AFC loudness balance experiments, noise was found to decrease the loudness of moderate to intense stimuli. This suggests that loudness is partially coded by the degree of phase-locking of cochlear nerve fibers. The overall gain in dynamic range was modest, and more complex noise strategies, for example, using inhibition between the noise sources, may be required to get a clinically useful benefit. © 2006 Association for Research in Otolaryngology.

Quantitative Analysis of Kilohertz-Frequency Neurostimulation

Mainstream electrical stimulation therapies, e.g., spinal cord stimulation (SCS) and deep brain stimulation, use pulse trains that are delivered at rates no higher than 200 Hz. In recent years, stimulation of nerve fibers using kilohertz-frequency (KHF) signals has received increased attention due to the potential to penetrate deeper in the tissue and to the ability to block conduction of action potentials. As well, there are a growing number of clinical applications that use KHF waveforms, including transcutaneous electrical stimulation (TES) for overactive bladder and SCS for chronic pain. However, there is a lack of fundamental understanding of the mechanisms of action of KHF stimulation. The goal of this research was to analyze quantitatively KHF neurostimulation.

We implemented a multilayer volume conductor model of TES including dispersion and capacitive effects, and we validated the model with in vitro measurements in a phantom constructed from dispersive materials. We quantified the effects of frequency on the distribution of potentials and fiber excitation. We also quantified the effects of a novel transdermal amplitude modulated signal (TAMS) consisting of a non-zero offset sinusoidal carrier modulated by a square-pulse train. The model revealed that high-frequency signals generated larger potentials at depth than did low frequencies, but this did not translate into lower stimulation thresholds. Both TAMS and conventional rectangular pulses activated more superficial fibers in addition to the deeper, target fibers, and at no frequency did we observe an inversion of the strength-distance relationship. In addition, we performed in vivo experiments and applied direct stimulation to the sciatic nerve of cats and rats. We measured electromyogram and compound action potential activity evoked by pulses, TAMS and modified versions of TAMS in which we varied the amplitude of the carrier. Nerve fiber activation using TAMS showed no difference with respect to activation with conventional pulse for carrier frequencies of 20 kHz and higher, regardless the size of the carrier. Therefore, TAMS with carrier frequencies >20 kHz does not offer any advantage over conventional pulses, even with larger amplitudes of the carrier, and this has implications for design of waveforms for efficient and effective TES.

We developed a double cable model of a dorsal column (DC) fiber to quantify the responses of DC fibers to a novel KHF-SCS signal. We validated the model using in vivo recordings of the strength-duration relationship and the recovery cycle of single DC fibers. We coupled the fiber model to a model of SCS in human and applied the KHF-SCS signal to quantify thresholds for activation and conduction block for different fiber diameters at different locations in the DCs. Activation and block thresholds increased sharply as the fibers were placed deeper in the DCs, and decreased for larger diameter fibers. Activation thresholds were > 5 mA in all cases and up to five times higher than for conventional (~ 50 Hz) SCS. For fibers exhibiting persistent activation, the degree of synchronization of the firing activity to the KHF-SCS signal, as quantified using the vector strength, was low for a broad amplitude range, and the dissimilarity between the activities in pairs of fibers, as quantified using the spike time distance, was high and decreased for more closely positioned fibers. Conduction block thresholds were higher than 30 mA for all fiber diameters at any depth and well above the amplitudes used clinically (0.5 – 5 mA). KHF-SCS appears to activate few, large, superficial fibers, and the activated fibers fire asynchronously to the stimulation signal and to other activated fibers.

The outcomes of this work contribute to the understanding of KHF neurostimulation by establishing the importance of the tissue filtering properties on the distribution of potentials, assessing quantitatively the impact of KHF stimulation on nerve fiber excitation, and developing and validating a detailed model of a DC fiber to characterize the effects of KHF stimulation on DC axons. The results have implications for design of waveforms for efficient and effective nerve fiber stimulation in the peripheral and central nervous system.

Évaluation objective de la séquence de récupération des fibres sensitives de la troisième division du nerf trijumeau suite à une ostéotomie sagittale mandibulaire bilatérale (OSMB)

Le projet de recherche est une étude prospective consistant à déterminer la séquence de récupération des fibres sensitives de la troisième division du nerf trijumeau (V3) suite à une ostéotomie sagittale mandibulaire bilatérale (OSMB). Dix-neuf sujets ont été recrutés entre les mois de mars et septembre 2008. Tous ont eu une chirurgie orthognathique d’OSMB afin de corriger une malocclusion. La sensibilité dans le territoire cutané innervé par V3 de chacun des sujets a été évaluée en pré-opératoire de même qu’à cinq autres reprises en post-opératoire (2, 4, 20, 36 et 52 semaines). Deux méthodes d’évaluation objectives de la récupération nerveuse sensitive ont été utilisées. La première consistait à utiliser un appareil nommé Neurometer afin de déterminer l’intensité minimale de courant électrique (Current Perception Threshold [CPT]) pouvant être ressentie spécifiquement par chacun des trois types de fibres nerveuses sensitives (A-Bêta, A-Delta et C) dans le territoire cutané de V3. La deuxième méthode consistait à utiliser les monofilaments de Semmes-Weinstein afin de déterminer le seuil minimal de pression (Von Frey) pouvant être ressenti dans le même territoire cutané. De plus, lors de chacun des rendez-vous post-opératoires, il a été demandé à chaque sujet de quantifier subjectivement sa sensibilité à l’aide d’une échelle visuelle analogue. Cela a permis de corréler les valeurs de CPT, les seuils de perception de la pression et l’évaluation subjective que le patient a de sa propre sensibilité. Il a été démontré que la séquence de récupération des fibres sensitives de V3 suite à une OSMB est la suivante : les fibres A-Delta récupèrent en premier, suivies des fibres C puis des fibres A-Bêta.

A novel mutation of AFG3L2 might cause dominant optic atrophy in patients with mild intellectual disability

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Microneurovascular free muscle transfer with cross-over nerve grafts in facial reanimation

Microneurovascular free muscle transfer with cross-over nerve grafts in facial reanimation Loss of facial symmetry and mimetic function as seen in facial paralysis has an enormous impact on the psychosocial conditions of the patients. Patients with severe long-term facial paralysis are often reanimated with a two-stage procedure combining cross-facial nerve grafting, and 6 to 8 months later with microneurovascular (MNV) muscle transfer. In this thesis, we recorded the long-term results of MNV surgery in facial paralysis and observed the possible contributing factors to final functional and aesthetic outcome after this procedure. Twenty-seven out of forty patients operated on were interviewed, and the functional outcome was graded. Magnetic resonance imaging (MRI) of MNV muscle flaps was done, and nerve graft samples (n=37) were obtained in second stage of the operation and muscle biopsies (n=18) were taken during secondary operations.. The structure of MNV muscles and nerve grafts was evaluated using histological and immunohistochemical methods ( Ki-67, anti-myosin fast, S-100, NF-200, CD-31, p75NGFR, VEGF, Flt-1, Flk-1). Statistical analysis was performed. In our studies, we found that almost two-thirds of the patients achieved good result in facial reanimation. The longer the follow-up time after muscle transfer the weaker was the muscle function. A majority of the patients (78%) defined their quality of life improved after surgery. In MRI study, the free MNV flaps were significantly smaller than originally. A correlation was found between good functional outcome and normal muscle structure in MRI. In muscle biopsies, the mean muscle fiber diameter was diminished to 40% compared to control values. Proliferative activity of satellite cells was seen in 60% of the samples and it tended to decline with an increase of follow-up time. All samples showed intramuscular innervation. Severe muscle atrophy correlated with prolonged intraoperative ischaemia. The good long-term functional outcome correlated with dominance of fast fibers in muscle grafts. In nerve grafts, the mean number of viable axons amounted to 38% of that in control samples. The grafted nerves characterized by fibrosis and regenerated axons were thinner than in control samples although they were well vascularized. A longer time between cross facial nerve grafting and biopsy sampling correlated with a higher number of viable axons. P75Nerve Growth Factor Receptor (p75NGFR) was expressed in every nerve graft sample. The expression of p75NGFR was lower in older than in younger patients. A high expression of p75NGFR was often seen with better function of the transplanted muscle. In grafted nerve Vascular Endothelial Growth Factor (VEGF) and its receptors were expressed in nervous tissue. In conclusion, most of the patients achieved good result in facial reanimation and were satisfied with the functional outcome. The mimic function was poorer in patients with longer follow-up time. MRI can be used to evaluate the structure of the microneurovascular muscle flaps. Regeneration of the muscle flaps was still going on many years after the transplantation and reinnervation was seen in all muscle samples. Grafted nerves were characterized by fibrosis and fewer, thinner axons compared to control nerves although they were well vascularized. P75NGFR and VEGF were expressed in human nerve grafts with higher intensity than in control nerves which is described for the first time.

Loss of nerve fiber layer in HIV positive patients and comparative patterns in a series of cases in Bucaramanga, Colombia

Objective: The objective of this study is to conduct a description of the features of optic neuropathy associated with Human Immunodeficiency Virus in relation to their possible incidence within our population, regarding that there is no data in our population in terms of frequency of this pathology (1,2). Methodology: Descriptive cross-sectional study of a clinical series of patients infected with human immunodeficiency virus, but AIDS, and the thickness of optic nerve´s layer of fibers studied with OCT technology (optical coherence tomography), patients were cited once captured. OCT was performed by the same observer, by taking 3 shots and picking the one with better reliability. Patients were given personally to the Ophthalmologic Foundation of Santander to conduct the review called OCT (optical coherence tomography). Results: In terms of viral load variable, we found a clear correlation in which validates the hypothesis that lower viral load means a thicker layer of fibers finding statistically significant differences for the 6 hours in right eye and 12 and 6 hours in left eye. Comparison between the known nomogram of fiber layer thickness for the population of Bucaramanga, Santander and thickness found in our sample, we note a clear decrease in the upper and lower quadrants, specifically in 7 hours and 11 hours, being more important in 7 hours, showing statistically significant differences. Conclusions: The pattern  of thinning of the nerve fiber layer in HIV positive patients without AIDS, and antiretroviral treatment type HAART, showed a statistically significant thinning targeted at 7 hours and 11 hours, being higher in first. Viral load figures have a direct relation with loss fiber layer, showing a statistically significant difference for the 6 and 12 hours.