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Microglial cells (MGCs) are highly dynamic and have been implicated in shaping discrete neural maps in several unimodal systems. MGCs respond to numerous cues in their microenvironment, including the neuronally-expressed chemokine, fractalkine (CX3CL1), via interactions with its corresponding fractalkine receptor (CX3CR1). The present study examines microglial and CX3CL1 patterns with regard to the emerging modular-extramodular matrix organization within the lateral cortex of the inferior colliculus (LCIC). The LCIC is a multisensory shell region of the midbrain inferior colliculus where discrete compartments receive modality-specific connections. Somatosensory inputs terminate within modular confines, while auditory inputs target the surrounding matrix. Glutamic acid decarboxylase (GAD) is an established marker of LCIC modules in developing mouse. During early postnatal development, multimodal LCIC afferents segregate into discrete, neurochemically-defined compartments. Here, we analyzed neonatal GAD67-GFP and CX3CR1-GFP mice to assess: (1) whether MGCs are recruited to distinct LCIC compartments known to be undergoing active circuit assembly, and (2) if such behaviors are fractalkine signaling-dependent. MGCs colonize the nascent LCIC by birth and increase in density until postnatal day 12 (P12). At the peak critical period (P4-P8), MGCs conspicuously border emerging LCIC modules, prior to their subsequent invasion by P12. CX3CL1 expression becomes distinctly modular at P12, in keeping with the notion of fractalkine-mediated recruitment of microglia to modular centers. In CX3CR1(GFP/GFP) mice with compromised fractalkine signaling, microglial recruitment into modules is delayed. Taken together, these results suggest a potential role for microglia and fractalkine signaling in sculpting multisensory LCIC maps during an early critical period.

In the multimodal lateral cortex of the inferior colliculus (LCIC) there are two neurochemically and connectionally distinct compartments, termed modular and extra modular zones. Modular fields span LCIC layer 2 and are recipients of somatosensory afferents, while encompassing extra modular domains receive auditory inputs. Recently in developing mice we identified several markers (among them glutamic acid decarboxylase, GAD) that consistently label the same modular set, as well as a reliable extra modular marker, calretinin, (CR). Previous reports from our lab show similar modular-extra modular patterns for certain Eph-ephrin guidance members, although their precise alignment with the developing LCIC neurochemical framework has yet to be addressed. Here we confirm in the nascent LCIC complementary GAD/CR-positive compartments, and characterize the registry of EphA4 and ephrin-B2 expression patterns with respect to its emerging modular-extra modular organization. Immunocytochemical approaches in GAD67-GFP knock-in mice reveal patchy EphA4 and ephrin-B2 domains that precisely align with GAD-positive LCIC modules, and are complementary to CR-defined extra modular zones. Such patterning was detectable neonatally, yielding discrete compartments prior to hearing onset. A dense plexus of EphA4-positive fibers filled modules, surrounding labeled ephrin-B2 and GAD cell populations. The majority of observed GABAergic neurons within modular boundaries were also positive for ephrin-B2. These results suggest an early compartmentalization of the LCIC that is likely instructed in part through Eph-ephrin guidance mechanisms. The overlap of developing LCIC neurochemical and guidance patterns is discussed in the context of its seemingly segregated multimodal input-output streams.

The complex neuroanatomical connections of the inferior colliculus (IC) and its major subdivisions offer a juxtaposition of segregated processing streams with distinct organizational features. While the tonotopically layered central nucleus (CNIC) is well-documented, less is known about functional compartments in the neighboring lateral cortex (LCIC). In addition to a laminar framework, LCIC afferent-efferent patterns suggest a multimodal mosaic, consisting of a patchy modular network with surrounding extramodular domains. The present study utilizes several neurochemical markers that reveal an emerging LCIC modular-extramodular microarchitecture. In newborn and post-hearing C57BL/6J and CBA/CaJ mice, histochemical and immunocytochemical stains were performed for acetylcholinesterase (AChE), nicotinamide adenine dinucleotide phosphate-diaphorase (NADPH-d), glutamic acid decarboxylase (GAD), cytochrome oxidase (CO), and calretinin (CR). Discontinuous layer 2 modules are positive for AChE, NADPH-d, GAD, and CO throughout the rostrocaudal LCIC. While not readily apparent at birth, discrete cell clusters emerge over the first postnatal week, yielding an identifiable modular network prior to hearing onset. Modular boundaries continue to become increasingly distinct with age, as surrounding extramodular fields remain largely negative for each marker. Alignment of modular markers in serial sections suggests each highlight the same periodic patchy network throughout the nascent LCIC. In contrast, CR patterns appear complementary, preferentially staining extramodular LCIC zones. Double-labeling experiments confirm that NADPH-d, the most consistent developmental modular marker, and CR label separate, non-overlapping LCIC compartments. Determining how this emerging modularity may align with similar LCIC patch-matrix-like Eph/ephrin guidance patterns, and how each interface with, and potentially influence developing multimodal LCIC projection configurations is discussed. This article is protected by copyright. All rights reserved.

The multimodal lateral cortex of the inferior colliculus (LCIC) exhibits a modular-extramodular micro-organization that is evident early in development. In addition to a set of neurochemical markers that reliably highlight its modular-extramodular organization (e.g. modules: GAD67-positive, extramodular zones: calretinin-positive, CR), mature projection patterns suggest that major LCIC afferents recognize and adhere to such a framework. In adult mice, distinct afferent projections appear segregated, with somatosensory inputs targeting LCIC modules and auditory inputs surrounding extramodular fields. Currently lacking is an understanding regarding the development and shaping of multimodal LCIC afferents with respect to its emerging modular-extramodular microarchitecture. Combining living slice tract-tracing and immunocytochemical approaches in GAD67-GFP knock-in mice, the present study characterizes the critical period of projection shaping for LCIC auditory afferents arising from its neighboring central nucleus (CNIC). Both crossed and uncrossed projection patterns exhibit LCIC extramodular mapping characteristics that emerge from initially diffuse distributions. Projection mismatch with GAD-defined modules and alignment with encompassing extramodular zones becomes increasingly clear over the early postnatal period (birth to postnatal day 12). CNIC inputs terminate almost exclusively in extramodular zones that express CR. These findings suggest multimodal LCIC inputs may initially be sparse and intermingle, prior to segregation into distinct processing streams. Future experiments are needed to determine the likely complex interactions and mechanisms (e.g. activity-dependent and independent) responsible for shaping early modality-specific LCIC circuits.

Eph-ephrin interactions guide topographic mapping and pattern formation in a variety of systems. In contrast to other sensory pathways, their precise role in the assembly of central auditory circuits remains poorly understood. The auditory midbrain, or inferior colliculus (IC) is an intriguing structure for exploring guidance of patterned projections as adjacent subdivisions exhibit distinct organizational features. The central nucleus of the IC (CNIC) and deep aspects of its neighboring lateral cortex (LCIC, Layer 3) are tonotopically-organized and receive layered inputs from primarily downstream auditory sources. While less is known about more superficial aspects of the LCIC, its inputs are multimodal, lack a clear tonotopic order, and appear discontinuous, terminating in modular, patch/matrix-like distributions. Here we utilize X-Gal staining approaches in lacZ mutant mice (ephrin-B2, -B3, and EphA4) to reveal EphA-ephrinB expression patterns in the nascent IC during the period of projection shaping that precedes hearing onset. We also report early postnatal protein expression in the cochlear nuclei, the superior olivary complex, the nuclei of the lateral lemniscus, and relevant midline structures. Continuous ephrin-B2 and EphA4 expression gradients exist along frequency axes of the CNIC and LCIC Layer 3. In contrast, more superficial LCIC localization is not graded, but confined to a series of discrete ephrin-B2 and EphA4-positive Layer 2 modules. While heavily expressed in the midline, much of the auditory brainstem is devoid of ephrin-B3, including the CNIC, LCIC Layer 2 modular fields, the dorsal nucleus of the lateral lemniscus (DNLL), as well as much of the superior olivary complex and cochlear nuclei. Ephrin-B3 LCIC expression appears complementary to that of ephrin-B2 and EphA4, with protein most concentrated in presumptive extramodular zones. Described tonotopic gradients and seemingly complementary modular/extramodular patterns suggest Eph-ephrin guidance in establishing juxtaposed continuous and discrete neural maps in the developing IC prior to experience.

We investigated the perception of two mechanoreceptive modalities alone and in combination: main effects and interaction between auditory and somatosensory stimulation in mice. Fifteen C57BL/6J mice between the ages of 1 and 6 months were tested three times each. Experimental design roughly followed published procedures using pre-pulse inhibition (PPI) of the acoustic startle response, except pre-pulses included vibration of the test chamber as well as soft sounds. Auditory pre-pulses were 80 dB broadband noises of 4, 9, 25, or 45 ms duration. Vibrations were of the same duration but of different frequencies (500, 460, 360, and 220 Hz). Pre-pulse inhibition increased with duration of the auditory pre-pulses, as expected. There was significant PPI to some but not all vibrotactile pre-pulses. Multimodal PPI was approximately additive (no significant auditory-by-somatosensory interaction). PPI increased more with age to somatosensory than to auditory pre-pulses. Future studies of multi-modal psychophysics in various mouse mutants could lend support to more mechanistic studies of neural specificity and possibly autism, tinnitus, and PTSD.

Soil to plant Transfer Factors (TFs) are commonly used to estimate the food chain transfer of chemical elements. The entry of trace contaminants, such as potentially toxic trace elements (PTEs), from soil, into human food chains, is controlled in the long term by their uptake by plant roots. Some PTEs in the soil-plant system were studied through the TFs in an olive orchard in southern Italy. Soil samples and olive leaves were collected and analysed for Cd, Co, Cr, Cu, Mn, Ni, Pb, Zn by using Inductively Coupled Plasma-Mass Spectrometry (ICP-MS). In addition, pH, electrical conductivity, organic matter, clay content and cation exchange capacity were measured in soil samples. Factor of trace elements from soil to olive plants showed average value

The routine molecular test for spinal muscular atrophy (SMA) diagnosis is based on the detection of a homozygous deletion of exons 7 and 8 of the telomeric copy of the survival motor neuron gene (SMN1). The presence of the centromeric copy of the SMN gene (SMN2) does not allow the detection of the hemizygous absence of the SMN1 gene, which characterizes the disease carriers. The demand for a quantitative SMN1 test is permanently growing because there is a high incidence of carriers. The disease is severe and to date there are no effective pharmacological treatments. Here, we present a non-radioactive assay based on real time quantitative polymerase chain reaction. We analyzed eight SMA patients, 14 SMA relatives and 50 health individuals from Southern Italy by real time quantitative method in order to identify haploid deletion occurring in SMA carriers. SMN1 copy number was determined by the comparative threshold cycle method (ΔΔCt). The results confirmed the deletion in all homozygous patients and permitted an evaluation of the number of alleles in the healthy carriers. This method is fast, reproducible, and enables us to discriminate carriers from healthy homozygous, which is impossible with normal techniques.

Aims To determine the retinal nerve fibre layer (RNFL) thickness at which visual field (VF) damage becomes detectable and associated with structural loss. Methods In a prospective cross-sectional study, 72 healthy and 40 glaucoma subjects (one eye per subject) recruited from an academic institution had VF examinations and spectral domain optical coherence tomography (SD-OCT) optic disc cube scans (Humphrey field analyser and Cirrus HD-OCT, respectively). Comparison of global mean and sectoral RNFL thicknesses with VF threshold values showed a plateau of threshold values at high RNFL thicknesses and a sharp decrease at lower RNFL thicknesses. A ‘broken stick’ statistical model was fitted to global and sectoral data to estimate the RNFL thickness ‘tipping point’ where the VF threshold values become associated with the structural measurements. The slope for the association between structure and function was computed for data above and below the tipping point. Results The mean RNFL thickness threshold for VF loss was 75.3 mm (95% CI: 68.9 to 81.8), reflecting a 17.3% RNFL thickness loss from age-matched normative value. Above the tipping point, the slope for RNFL thickness and threshold value was 0.03 dB/mm (CI: � 0.02 to 0.08) and below the tipping point, it was 0.28 dB/mm (CI: 0.18 to 0.38); the difference between the slopes was statistically significant (p

Central processing of complex auditory tasks requires elaborate circuitry. The auditory midbrain, or inferior colliculus (IC), epitomizes such precise organization, where converging inputs form discrete, tonotopically-arranged axonal layers. Previously in rat, we established that shaping of multiple afferent patterns in the IC central nucleus (CNIC) occurs prior to experience. This study implicates an Eph receptor tyrosine kinase and a corresponding ephrin ligand in signaling this early topographic registry. We report that EphA4 and ephrin-B2 expression patterns in the neonatal rat and mouse IC correlate temporally and spatially with that of developing axonal layers. DiI-labeling confirms projections arising from the lateral superior olive (LSO) form frequency-specific layers within the ipsilateral and contralateral mouse CNIC, as has been described in other species. Immunohistochemistry (EphA4 and ephrin-B2) and ephrin-B2 lacZ histochemistry reveal clear gradients in expression across the tonotopic axis, with most concentrated labeling observed in high-frequency, ventromedial aspects of the CNIC. Discrete patches of labeling were also discernible in the external cortex of the IC (ECIC; EphA4 patches in rat, ephrin-B2 patches in mouse). Observed gradients in the CNIC and compartmentalized ECIC expression persisted through the first postnatal week, before becoming less intense and more homogeneously distributed by the functional onset of hearing. EphA4 and ephrin-B2-positive neurons were evident in several auditory brainstem nuclei known to send patterened inputs to the IC. These findings suggest the involvement of cell-cell EphA4 and ephrin-B2 signaling in establishing order in the developing IC.

Purpose: To evaluate, within ocular imaging scans of acceptable quality as determined by manufacturers’ guidelines, the effects of image quality on glaucoma discrimination capabilities. Methods: One hundred and four healthy and 75 glaucomatous eyes from the Advanced Imaging in Glaucoma Study (AIGS) were imaged with GDx-VCC, HRT II, and StratusOCT. Quality score (QS≥8), pixel standard deviation (SD≤50), and signal strength (SS≥5) were used as quality parameter cutoffs, respectively. GDx nerve fiber indicator (NFI), and HRT Moorfields regression analysis (MRA) classifications and OCT mean retinal nerve fiber layer (RNFL) thickness were used as the discriminatory parameters. Logistic regression models were used to model the dichotomous clinical classification (healthy vs. glaucoma) as a function of image quality parameters and discriminatory parameters. Results: Quality parameter covariates were statistically non¬-significant for GDx and HRT but had an inverse effect on OCT in predicting disease (higher SS had lower probability of glaucoma). Age was a significant covariate for GDx and HRT, but not OCT, while ethnicity and interaction between the image quality and the institute where scans were acquired were significant covariates in the OCT models. Conclusion: Within scans considered to be acceptable based on the manufacturers’ recommended limits, scan quality does not affect the discriminating ability of global discriminatory parameters of GDx, HRT but have an effect on OCT measurements.

Aims: The purpose of this study was to compare the day-to-day reproducibility of optical coherence tomography (OCT; StratusOCT, Carl Zeiss Meditec, Dublin, CA) measurements of retinal nerve-fibre layer (RNFL) measurements at time points 1 year apart. Methods: One eye in each of 11 healthy subjects was examined using the StratusOCT fast RNFL scan protocol. Three fast RNFL scans with signal strength ⩾7 were obtained on each of 3 days within a month. This protocol was repeated after 12 months. A linear mixed effects model fitted to the nested data was used to compute the variance components. Results: The square root of the variance component that was attributed to the differences between subjects was 7.17 μm in 2005 and 7.28 μm in 2006. The square roots of the variance component due to differences between days within a single subject were 1.95 μm and 1.50 μm, respectively, and for within day within a single subject were 2.51 μm and 2.55 μm, respectively. There were no statistically significant differences for any variance component between the two testing occasions. Conclusions: Measurement error variance remains similar from year to year. Day and scan variance component values obtained in a cohort study may be safely applied for prediction of long-term reproducibility.

To compare stereometric parameters and classification results from the Heidelberg Retina Tomograph version 2 (HRT2); HRT3; and HRT3 Glaucoma Probability Score (GPS), an automated method of obtaining optic nerve head analysis without the need for manual definition of disc margin.Retrospective cross-sectional study.Five hundred four eyes from 281 consecutive subjects (glaucoma, glaucoma suspect, and healthy) evaluated in a glaucoma clinic.All participants had HRT2 scanning of the optic nerve head. Inclusion criteria were scans with good centration and focus, even illumination, an overall quality score by HRT3 of acceptable or better, and standard deviation50 mum. A Bland-Altman analysis was used for the comparison of HRT2 and HRT3. From these results, calibration equations were determined to permit conversion of the measurements between devices. The agreement between HRT2 and HRT3 Moorfields regression analysis (MRA) and HRT3 GPS classification methods was measured using kappa statistics.Heidelberg Retina Tomograph version 2 and HRT3 stereometric parameters, MRA, and global GPS.There was a statistically significant difference between HRT2 and HRT3 global disc area, rim area, cup area, rim volume, cup volume, height variation contour, and retinal nerve fiber layer cross-sectional area stereometric parameters. All of those parameters were smaller using HRT3, due to a manufacturer-reported horizontal scaling error of 4% in HRT2 that was corrected in HRT3. kappas for agreement were 0.60 between classifications (within normal limits, borderline, and outside normal limits) of MRA by HRT2 and HRT3 and 0.47 between HRT3 MRA and GPS.The HRT3 generally provided smaller stereometric disc measurements than HRT2. There was no clear conversion between HRT3 and GPS parameters, as the 2 methods for measuring the stereometric parameters differ.

The central nucleus of the inferior colliculus (IC) is a laminated structure that receives multiple converging afferent projections. These projections terminate in a layered arrangement and are aligned with dendritic arbors of the predominant disc-shaped neurons, forming fibrodendritic laminae. Within this structural framework, inputs terminate in a precise manner, establishing a mosaic of partially overlapping domains that likely define functional compartments. Although several of these patterned inputs have been described in the adult, relatively little is known about their organization prior to hearing onset. The present study used the lipophilic carbocyanine dyes DiI and DiD to examine the ipsilateral and contralateral projections from the lateral superior olivary (LSO) nucleus to the IC in a developmental series of paraformaldehyde-fixed kitten tissue. By birth, the crossed and uncrossed projections had reached the IC and were distributed across the frequency axis of the central nucleus. At this earliest postnatal stage, projections already exhibited a characteristic banded arrangement similar to that described in the adult. The heaviest terminal fields of the two inputs were always complementary in nature, with the ipsilateral input appearing slightly denser. This early arrangement of interdigitating ipsilateral and contralateral LSO axonal bands that occupy adjacent sublayers supports the idea that the initial establishment of this highly organized mosaic of inputs that defines distinct synaptic domains within the IC occurs largely in the absence of auditory experience. Potential developmental mechanisms that may shape these highly ordered inputs prior to hearing onset are discussed.

Virtually nothing is known about the ontogeny of substantia nigra, pars reticulata projections to the midbrain superior colliculus, even though this pathway is critical for the basal ganglia modulation of midbrain-mediated visuomotor behaviors. The present studies used the lipophilic carbocyanine dyes 1,1'-dioctodecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate and 1,1'-dioctodecyl-3,3,3',3'-tetramethylindodi, 4-chlorobenzenesulfonate salt to examine the crossed and uncrossed nigrotectal projections in neonatal cats, from parturition to 14 days postnatal (the technical limits of the tracing technique). In retrograde experiments, paired placement of the dyes in each superior colliculus produced numerous retrogradely-labeled nigrotectal neurons, with the uncrossed neurons far out numbering their crossed counterparts. No double-labeled neurons were observed, indicating that crossed and uncrossed nigrotectal neurons are segregated at birth. In anterograde experiments, dye placements into each substantia nigra, pars reticulata resulted in an iterative series of labeled patches, aligned medial-to-lateral across the intermediate and deep superior colliculus, a pattern reminiscent of the adult. Uncrossed neonatal axons had simple linear morphologies with few branch points; by contrast, crossed axons displayed more extensive terminal arbors that were distributed diffusely throughout the rostrocaudal extent of the contralateral superior colliculus In the final series of experiments, one dye was placed unilaterally in the substantia nigra, pars reticulata, while the second dye was positioned in the predorsal bundle, in order to bilaterally label superior colliculus output neurons. Although both crossed and uncrossed axons appeared to have contacted superior colliculus output neurons, crossed axons preferentially targeted the soma and proximal dendrites, whereas uncrossed terminals were distributed more distally. Throughout this early postnatal period, no significant changes in cellular morphologies or gross modification of terminal projection patterns were observed; however, the presence of growth cones in even the oldest animals studied suggests that the refinement of the nigrotectal projections extends well into postnatal life. Nevertheless, the segregation of crossed and uncrossed nigrotectal neurons into a highly organized afferent mosaic that has established synaptic contacts with superior colliculus output neurons indicates that many of the salient features characterizing nigrotectal projections are established prior to the onset of visual experience.

Charcot–Marie-Tooth type 1A is caused by a 1.5 Mb DNA duplication in the 17p12 chromosomal region encompassing the peripheral myelin protein 22 gene. In the present study, we compared the Real-Time PCR with the other methods currently used for the diagnosis of Charcot–Marie-Tooth. By using a combination of junction fragment PCR, analysis of microsatellite markers, and pulsed field gel electrophoresis, we identified 76 unrelated patients with 17p12 duplication. In these patients, junction fragment PCR detected 63% of cases of duplication, the microsatellite markers method revealed 74%, while the combined use of microsatellite markers and junction fragment PCR revealed 91% of cases of Charcot–Marie-Tooth type 1A. Pulsed field gel electrophoresis detected 100% of the cases with duplication, even in presence of atypical 17p12 duplication. Real-Time PCR detected 100% of the cases with Charcot–Marie-Tooth type 1A and was comparable to pulsed field gel electrophoresis. However, in contrast to pulsed field gel electrophoresis, Real-Time PCR does not need fresh blood, minimizes diagnosis time and cost, and thus can be easily used for the molecular diagnosis of Charcot–Marie-Tooth type 1A.

Aims: To study the ophthalmological manifestations in individuals with the typical features of neurofibromatosis type 1 (NF1) circumscribed to one or more body segments, usually referred to as segmental NF1. Methods: Visual acuity and colour tests, visual field examination, slit lamp biomicroscopy of the anterior segment, and a detailed examination of the retina by indirect ophthalmoscopy were performed at diagnosis and follow up in 72 consecutive subjects (29 males, 43 females; aged 1–64 years; mean age 14.6 years) seen at the university departments of paediatrics in Catania and Rome, Italy, during years 1990–2003, who had in restricted body areas: (1) typical pigmentary manifestations of NF1 (cafe au lait spots and freckling) only (n = 48); (2) NF1 pigmentary manifestations and neurofibromas alone (n = 2); (3) neurofibromas only (n = 15); and (4) plexiform neurofibromas only (n = 7). Results: None of the 72 patients had Lisch nodules in the iris irrespective of age at eye examination or hypertelorism (a “minor” NF1 feature) and none developed typical associated ophthalmological NF1 complications. An additional child had an isolated optic pathways glioma (OPG), which behaved both biologically and radiographically as an NF1 associated OPG. Conclusions: This represents the first systematic study reporting on eye involvement in the largest series of individuals at different ages having segmental NF1. As one of the postulated mechanisms to explain segmental NF1 is somatic mosaicism for the NF1 gene (so far demonstrated only in two patients) the present findings could be explained either by the fact that the eye is too far from the mutated area with NF1 lesions in most cases or by the NF1 (or other “predisposing” or “cooperating”) gene mutation restricted to too few cellular clones or to tissues embryologically different from the eye.

To provide further insights into the distribution and spectrum of mutations at the MECP2 locus, 21 patients from Sicily were analyzed: 14 with classical RTT syndrome and seven with variant forms. Mutations in MECP2 were found in 10 of 14 classical Rett patients (71.4%), and in three of seven cases with a variant form (42.8%). Two novel mutations, not found in 100 control chromosomes, were detected.

The central nucleus of the inferior colliculus (IC) is the site of convergence for nearly all ascending monaural and binaural projections. Several of these inputs, including inhibitory connections from the dorsal nucleus of the lateral lemniscus (DNLL), are highly ordered and organized into series of afferent bands or patches. Although inputs to the IC from the contralateral DNLL are present in the rat by birth [postnatal day 0 (P0)], the earliest indications of band formation are not evident until P4. Subsequently, the initially diffuse projection segregates into a pattern of bands and interband spaces, and by P12 adult-like, afferent-dense patches are established (Gabriele et al., 2000). To determine the role of the auditory periphery in the development of bands and patches before the onset of hearing (P12/P13), unilateral cochlear ablations were performed at P2 (before any evidence of banding). Rat pups were reared to P12, at which time glass pins coated with 1, 1'-dioctodecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate were placed in fixed tissue in the commissure of Probst where DNLL fibers cross the midline. The results indicate that a unilateral cochlear ablation disrupts the normal development of afferent patches in the IC. Although the crossed DNLL projections labeled via commissural dye placement always mirrored each other in P12 controls, ablation cases exhibited a consistent, bilateral asymmetry in pattern formation and relative density of the labeled projections. Possible developmental mechanisms likely to be involved in the establishment of afferent bands and patches before the onset of hearing are discussed.

Mutations in the Angiogenin gene (ANG) linked to 14q11.2 have been recently discovered to be associated with Amyotrophic Lateral Sclerosis (ALS) in Irish and Scottish populations. In our study we investigated the role of ANG gene in ALS patients from southern Italy. We found a novel mutation in the signal peptide of the ANG gene in a sporadic patient with ALS (SALS). The molecular analysis of the ANG gene also demonstrated an allelic association with the rs11701 single nucleotide polymorphism (SNP) in familial ALS (FALS) but not in SALS patients. Our finding supports the evidence that the ANG gene is involved in ALS.

Introduction TPACAP 1 est un programme d’education therapeutique en dermatologie accredite par l’ARS depuis 2011. Il concerne plus particulierement la dermatite atopique et l’urticaire chronique. L’engagement pluridisciplinaire de notre equipe en lien avec les professionnels de sante de ville nous a permis depuis 4 ans d’accompagner le patient vers d’avantage d’autonomie et de responsabilite dans la gestion de sa maladie et de son traitement. Ou en sommes-nous aujourd’hui ? Resultats L’evaluation de l’equipe a revele une cohesion pluridisciplinaire, un discours commun, un approfondissement des connaissances, une autre facon d’aborder les soins. Nous sommes parvenus a une posture de « savoir etre », un epanouissement professionnel aussi facilite par l’acquisition d’une expertise dans ces deux thematiques. Concernant l’evaluation des patients, celle-ci est globalement positive puisque nous avons atteint dans la majorite des cas les objectifs initialement fixes : – les patients sont capables d’evaluer leur etat cutane ; – d’adapter leur traitement a leur mode de vie ; – de gerer leur maladie au quotidien et en cas de crise debutante identifier les facteurs favorisants. Les patients ont aussi acquis une competence afin d’expliquer leur maladie a leur entourage, ils ont adapte leurs habitudes pour ameliorer leur qualite de vie. Par le biais de ses seances individuelles et collectives ils ont pu exprimer en toute confiance leur vecu difficile, la peur du regard des autres dans cette maladie « affichant » et ont pu trouver une ecoute attentive. Conclusion Grâce a ce programme et fort de ces 4 premieres annees d’experience, TPA CAP nous a permis d’avoir une autre vision des soins et ainsi nous avons pu faire progresser nos pratiques professionnelles. Ces rencontres entre patients et soignants ont enrichi notre experience et nous ont fait grandir mutuellement. Il est indeniable que les patients et les professionnels de sante ont chemine ensemble avec succes a travers TPA CAP.

Hereditary spastic paraplegias are neurodegenerative disorders characterized clinically by progressive spasticity of the lower limbs; they are inherited as autosomal dominant, autosomal recessive and X-linked traits. We have analyzed four autosomal recessive HSP families gathered from southern Italy. We performed genetic analysis using microsatellite markers associated with SPG5, SPG7, SPG11 and SPG14. Positive lod scores were obtained with markers located on chromosome 8. The lod scores for the four combined families were significantly higher than 3 for D8S509, D8S1102, D8S1723 and D8S260 with a maximum two-point lod score at θ = 0 of 3.99 for the marker D8S260. In one of the examined families, the haplotype analysis suggests two key recombination events demonstrating that the gene is localized in the 11 cM region flanked by markers D8S285 and D8S544, refining the ARHSP region by approximately 22 cm. We also analyzed five candidate genes localized within the HSP region: TOX, syndecan-binding-protein (SDCBP), RAB2, CA8 and PENK, but we did not find disease causing mutations.

The corneal endothelium (CE) is a single layer of cells lining the posterior face of the cornea providing metabolic functions essential for maintenance of corneal transparency. Adult CE cells lack regenerative potential, and the number of CE cells decreases throughout life. To determine whether endogenous DNA damage contributes to the age-related spontaneous loss of CE, we characterized CE in Ercc1(-/Δ) mice, which have impaired capacity to repair DNA damage and age prematurely. Eyes from 4.5- to 6-month-old Ercc1(-/Δ) mice, age-matched wild-type (WT) littermates, and old WT mice (24- to 34-month-old) were compared by spectral domain optical coherence tomography and corneal confocal microscopy. Histopathological changes in CE were further identified in paraffin tissue sections, whole-mount immunostaining, and scanning electron and transmission electron microscopy. The CE of old WT mice displayed polymorphism and polymegathism, polyploidy, decreased cell density, increased cell size, increases in Descemet's thickness, and the presence of posterior projections originating from the CE toward the anterior chamber, similar to changes documented for aging human corneas. Similar changes were observed in young adult Ercc1(-/Δ) mice CE, demonstrating spontaneous premature aging of the CE of these DNA repair-deficient mice. CD45(+) immune cells were associated with the posterior surface of CE from Ercc1(-/Δ) mice and the tissue expressed increased IL-1α, Cxcl2, and TNFα, pro-inflammatory proteins associated with senescence-associated secretory phenotype. These data provide strong experimental evidence that DNA damage can promote aging of the CE and that Ercc1(-/Δ) mice offer a rapid and accurate model to study CE pathogenesis and therapy.

Background Familial spinal neurofibromatosis is a form of neurofibromatosis 1 (NF1), consisting of extensive, symmetrical, histologically proven, multiple neurofibromas of the spinal roots at every level and of all major peripheral nerves sometimes associated with typical NF1 stigmata; most cases underlie NF1 gene mutations. Objectives The objectives of this study are (1) to report the findings in a set of 16-year-old monozygotic twin girls and a 14-year-old boy and (2) to review the existing literature. Methods and Results In this article, we report the cases of three children who (1) had manifested mildly different symptomatic neuropathy (twins, aged 4 years; and a boy, aged 9 years) associated with massive, symmetrical neurofibromas; (2) had few café-au-lait spots with irregular margins and pale brown pigmentation; (3) were presented with, at brain magnetic resonance imaging (MRI), bilateral, NF1-like high-signal abnormalities in the basal ganglia; (4) yielded missense NF1 gene mutations in exon 39; and (5) had unaffected parents with negative NF1 genetic testing as well as discuss 12 families and 20 sporadic and 5 additional cases that presented spinal neurofibromatosis within classical NF1 families (53 cases) that were reported in the literature. Conclusions This article presents the first report on (1) spinal neurofibromatosis in a set of affected monozygotic twins; (2) the earliest onset of the disease; and (3) the occurrence of high signal lesions in the brain at MRI. © 2013 Georg Thieme Verlag KG Stuttgart.

Estimating activity of natural radionuclides in agricultural soil is very important for the protection of public health because the released radioactivity can enter the food chain. Radioactivity measurements were carried out in two different dates (winter and summer) in agricultural soil using a GRM-260 gamma-ray spectrometer. The study area (100 m x 100 m) was an olive orchard in southern Italy. Measurements were carried out at 361 locations in January and July 2011. At the same locations, soil water content was measured to take into account the effect of soil moisture on radioactivity. A multi-Gaussian approach was used to explore and map the activity of naturally occurring radionuclides and soil water content for both seasons of measurements. The minimum radioactivity values were recorded in winter and the maximum values in summer, probably as a consequence of changes in weather and soil conditions (rainfall, soil moisture, temperature).

Neurofibromatosis type 2 (NF2) with onset before the first year of life has been anecdotally reported in the literature. We (a) prospectively (years 1997-2012) followed up three unrelated NF2 children, all harbouring NF2 gene mutations whose onset of disease was before age 1 year, and (b) systematically reviewed published reports on NF2 in the youngest age group (i.e. onset < 1 year). The present three children had (1) small (< 1 cm), bilateral vestibular schwannomas (VSs) detected (as an incidental finding) at magnetic resonance imaging (MRI) by the age of 4 to 5 months that were asymptomatic for 10 to 14 years, with sudden and rapid (< 12 months) progression in two cases at the age of 11 and 15 years, respectively; (2) development of large numbers of skin NF2 plaques mainly in atypical locations (i.e. face, hands, legs and knees), which reverted to normal skin appearance at the time of VSs progression; (3) lens opacities (n = 1) and NF2 retinal changes (n = 2) detected as early as age of 3-4 months; (4) diffuse (asymptomatic) high signal lesions at brain MRI in the periventricular regions (alike cortical dysplasia); and (5) unaffected first-degree relatives who did not harbour NF2 gene abnormalities. This represents the youngest NF2 group with the longest prospective follow-up so far reported. NF2 may present as a congenital form with bilateral VSs presenting as early as the first months of life and with natural history different to that which occurs in classical NF2.

Agricultural soil geochemistry is an important research issue because soil pollution by potentially toxic trace elements (PTEs) is of concern worldwide. High concentrations of PTEs in soil pose a threat for human health because they can be taken up into the trophic chain via assimilation by plants. Mapping the PTEs in soil is essential to delineate contamination. Geostatistical methods provide us a valuable tool to study spatial structure of the PTEs. They take into account spatial autocorrelation of data to create mathematical models of spatial correlation structures commonly expressed by variograms. The aims of this study were to quantify the spatial structure and the relationships of some PTEs (Ag, Be, Bi, Cd, Co, Cr, Cu, Ni, Pb, Sn, Tl, Zn): To do that a multivariate geostatistical approach was used. The experimental area (100 m x 100 m) was an olive orchard located in southern Italy (Calabria) where at 100 locations topsoil (0-0.20 m) samples were collected. Soil samples were ground, dried, weighed, digested in aqua regia and analyzed in laboratory for Ag, Be, Bi, Cd, Co, Cr, Cu, Ni, Pb, Sn, Tl, and Zn by Inductively Coupled Plasma Mass Spectrometry (ICP-MS). Four elements exceeded the regulatory threshold value (Limit A) in analysed soils: Be, Sn, Zn, and V, while Cr, Pb, Cu, Tl, Co, Ni and Cd were within the tolerability limits. A variographic analysis was carried out to quantify the spatial structure of PTEs and no significant difference as a function of direction was found. Mapping the spatial distribution of PTEs allowed defining their spatial relationships and delineating the potentially risky areas. Finally, stochastic images generated by the sequential Gaussian simulation were jointly combined to calculate the probability of exceeding the regulatory threshold values that might cause concern for human health.

A 34-year-old female was referred to us for a consultation of her dermatological lesions (pigmented lesions present in her oral cavity and on her right shoulder) in May 2007. These lesions had been present since childhood. Recently, the patient had developed seizures and a headache. An MRI of the brain showed the presence of two intracranial masses. The intracranial tumours were surgically removed whereas skin and mucosal lesions were biopsied. Histological findings of brain tumours were consistent with a diagnosis of “melanocytoma” while cutaneous lesions presented “benign dermal melanocytic infiltrations”. Whole brain irradiation was performed. After 3 months a new melanocytic skin lesion appeared on the scalp with histological picture similar to the other cutaneous ones. At the 5-year follow-up examination no recurrence of intracranial tumour or other skin or mucosal lesions were registered. According to the clinical and histological findings, we classify our case as a form of neurocutaneous melanosis in a young adult patient and we present it for the rarity of this syndrome, for the difficulty of the diagnosis, for the potential aggressive behaviour of intracranial lesions that necessitates a constant attentive follow-up and for the unusual feature of new developing skin lesion during the course of the disease.

Graded and modular expressions of Eph-ephrins are known to provide positional information for the formation of topographic maps and patterning in the developing nervous system. Previously we have shown that ephrin-B2 is expressed in a continuous gradient across the tonotopic axis of the central nucleus of the inferior colliculus (CNIC), whereas patterns are discontinuous and modular in the lateral cortex of the IC (LCIC). The present study explores the involvement of ephrin-B2 signaling in the development of projections to the CNIC and LCIC arising from the lateral superior olivary nuclei (LSO) prior to hearing onset. Anterograde and retrograde fluorescent tracing methods in neonatal fixed tissue preparations were used to compare topographic mapping and the establishment of LSO layers/modules in wild-type and ephrin-B2(lacZ/+) mice (severely compromised reverse signaling). At birth, pioneer LSO axons occupy the ipsilateral IC in both groups but are delayed contralaterally in ephrin-B2(lacZ/+) mutants. By the onset of hearing, both wild-type and mutant projections form discernible layers bilaterally in the CNIC and modular arrangements within the ipsilateral LCIC. In contrast, ephrin-B2(lacZ/+) mice lack a reliable topography in LSO-IC projections, suggesting that fully functional ephrin-B2 reverse signaling is required for normal projection mapping. Taken together, these ephrin-B2 findings paired with known coexpression of EphA4 suggest the importance of these signaling proteins in establishing functional auditory circuits prior to experience.

We describe the clinical, neuropathological and molecular findings from a patient affected with neuronal ceroid lipofuscinosis with a juvenile onset (JNCL). She was a 9-year-old right-handed girl with a normal birth and early developmental milestones. At the age of 4 the early symptoms began. Skin biopsy showed granular osmiophilic deposits (GRODs).Because JNCL with GRODs is caused by mutations in the CNL1 gene, we performed a molecular investigation by direct sequencing of nine exons of the CNL1 gene. This analysis revealed a novel mutation in homozygous form in the exon 7 that caused an aminoacid substitution at codon 222 (Leu --> Pro). Direct sequencing of the exon 7 in both parents showed the same substitution in heterozygous form.

Retinal ganglion cell (RGC) death induced by optic nerve crush injury has been used as a model to investigate axonal degeneration of the central nervous system in mice.1–4 In adult mice, approximately 60% of RGCs are lost within 3 weeks after optic nerve crush injury, as determined by histology.1,4 Ophthalmic imaging techniques such as confocal scanning laser ophthalmoscopy (CSLO) and optical coherence tomography (OCT) can provide quantitative and detailed structural information from the retina and optic nerve head (ONH) region. In vivo images of RGCs in mice and rats after optic nerve crush have been acquired using retrograde labeling and CSLO.5–7 Recently, Leung et al.8 demonstrated the use of CSLO with Thy-1 cyan fluorescent protein expressing transgenic mice to monitor RGCs in vivo after nerve crush. The authors in these studies were able to observe the loss of RGCs in the same eyes over time, contrary to previous experiments using histology that required several different animals at each time point. OCT allows for noninvasive, in vivo imaging of ocular structures and has been used to obtain optical cross sections of the mouse retina.9–12 Time-domain OCT9,10 and spectral-domain OCT (SD-OCT)13–17 imaging have been used to observe retinal thinning in mouse models of retinal degeneration. One study used SD-OCT to image axotomized rats and manually obtained measurements of retinal layers in six cross-sectional images.18 This approach, however, is prone to subjective bias both in the selection of individual cross-sections and in the manual segmentation of retinal layers. To fully benefit from the high resolution and rapid scanning provided by SD-OCT, it would be desirable to obtain automated measurements from the area of interest. The goal of this study, therefore, was to investigate the longitudinal effect of optic nerve crush injury on total retinal thickness (TRT) in adult mice using automated segmentation analysis of SD-OCT images.

Measurements of human Schlemm's canal (SC) have been limited to histologic sections. The purpose of this study was to demonstrate noninvasive measurements of aqueous outflow (AO) structures in the human eye, examining regional variation in cross-sectional SC areas (on/off collector channel [CC] ostia [SC/CC] and nasal/temporal) in the eyes of living humans.SC was imaged by spectral-domain optical coherence tomography with a 200-nm bandwidth light source. Both eyes of 21 healthy subjects and one glaucomatous eye of three subjects were imaged nasally and temporally. Contrast and magnification were adjusted to maximize visualization. Cross-sectional SC on and off SC/CC was traced three times by two independent masked observers using ImageJ (ImageJ 1.40g, http://rsb.info.nih.gov/ij/ Wayne Rasband, developer, National Institutes of Health, Bethesda, MD). The mean SC area was recorded. A linear mixed-effects model was used to analyze eye, nasal/temporal laterality, and SC area on or off SC/CC.SC area was significantly larger on SC/CCs than off (12,890 vs. 7,391 micorm(2), P0.0001) and was significantly larger on the nasal side than on the temporal (10,983 vs. 8,308 micorm(2), P = 0.009). SC areas were significantly smaller in glaucoma patients than in normal subjects, whether pooled (P = 0.0073) or grouped by on (P = 0.0215) or off (P = 0.0114) SC/CC.Aqueous outflow structures, including SC and CCs, can be noninvasively assessed in the human eye. These measurements will be useful in physiological studies of AO and will be clinically useful in the determination of the impact of glaucoma therapies on IOP as well as presurgical planning.

Three dimensional (3D) ophthalmic imaging using optical coherence tomography (OCT) has revolutionized assessment of the eye, the retina in particular. Recent technological improvements have made the acquisition of 3D-OCT datasets feasible. However, while volumetric data can improve disease diagnosis and follow-up, novel image analysis techniques are now necessary in order to process the dense 3D-OCT dataset. Fundamental software improvements include methods for correcting subject eye motion, segmenting structures or volumes of interest, extracting relevant data post hoc and signal averaging to improve delineation of retinal layers. In addition, innovative methods for image display, such as C-mode sectioning, provide a unique viewing perspective and may improve interpretation of OCT images of pathologic structures. While all of these methods are being developed, most remain in an immature state. This review describes the current status of 3D-OCT scanning and interpretation, and discusses the need for standardization of clinical protocols as well as the potential benefits of 3D-OCT scanning that could come when software methods for fully exploiting these rich data sets are available clinically. The implications of new image analysis approaches include improved reproducibility of measurements garnered from 3D-OCT, which may then help improve disease discrimination and progression detection. In addition, 3D-OCT offers the potential for preoperative surgical planning and intraoperative surgical guidance.

Time domain optical coherence tomography (TD-OCT) has been used commonly in clinical practice, producing a large inventory of circular scan data for retinal nerve fiber layer (RNFL) assessment. Spectral domain (SD)-OCT produces three-dimensional (3-D) data volumes. The purpose of this study was to create a robust technique that makes TD-OCT circular scan RNFL thickness measurements comparable with those from 3-D SD-OCT volumes.Eleven eyes of 11 healthy subjects and 7 eyes of 7 subjects with glaucoma were enrolled. Each eye was scanned with one centered and eight displaced TD-OCT scanning circles. One 3-D SD-OCT cube scan was obtained at the same visit. The matching location of the TD-OCT scanning circle was automatically detected within the corresponding 3-D SD-OCT scan. Algorithm performance was assessed by estimating the difference between the detected scanning circle location on 3-D SD-OCT volume and the TD-OCT circle location. Global and sectoral RNFL thickness measurement errors between the two devices were also compared.The difference (95% confidence interval) in scanning circle center locations between TD- and SD-OCT was 2.3 (1.5-3.2) pixels (69.0 [45.0-96.0] microm on the retina) for healthy eyes and 3.1 (2.0-4.1) pixels (93.0 [60.0-123.0] microm on the retina) for glaucomatous eyes. The absolute RNFL thickness measurement difference was significantly smaller with the matched scanning circle.Scan location matching may bridge the gap in RNFL thickness measurements between TD-OCT circular scan data and 3-D SD-OCT scan data, providing follow-up comparability across the two generations of OCTs.

Mutations in the SPG4 gene are the most common causes of hereditary spastic paraplegia (HSP) accounting for up to 40% of autosomal dominant (AD) forms and 12-18% of sporadic cases. The phenotype associated with HSP due to mutations in the SPG4 gene tends to be pure. There is increasing evidence, however, of patients with complicated forms of spastic paraplegia in which SPG4 mutations were identified. A cohort of 38 unrelated Italian patients with spastic paraplegia, of which 24 had a clear dominant inheritance and 14 were apparently sporadic, were screened for mutations in the SPG4 gene.We identified 11 different mutations, six of which were novel (p.Glu143GlyfsX8, p.Tyr415X, p.Asp548Asn, c.1656_1664delinsTGACCT, c.1688-3C>G and c.*2G>T) and two exon deletions previously reported. The overall rate of SPG4 gene mutation in our patients was 36.8% (14/38); in AD-HSP we observed a mutation frequency of 45.8% (11/24), in sporadic cases the frequency was 21.4% (3/14). Furthermore, we found a mutational rate of 22.2% (2/9) and 41.4% (12/29) in the complicated and pure forms, respectively. The results underlie the importance of genetic testing in all affected individuals.

TAR-DNA-binding protein 43 (TDP-43) has recently been identified as the major pathological protein in abnormal inclusions in neurons and glial cells in sporadic amyotrophic lateral sclerosis (SALS) and SOD1 negative familial cases with amyotrophic lateral sclerosis (FALS). TDP-43 is evolutionarily conserved, consisting of two RNA recognition motifs and a glycine-rich C-terminal domain. It is involved in the regulation of expression and splicing, and in other cellular processes such as microRNA biogenesis, apoptosis and cell division.1 2 Starting in early 2008, dominant mutations in TARDBP gene have been reported by several groups as a primary cause of ALS. To date, a total of 30 mutations of TARDBP have been reported not only in SOD1 -negative FALS cases (∼3%) but also in SALS cases (∼1.5%). All but one of the mutations identified (D169G) reside in exon 6 of the TARDBP gene, which encodes for the C-terminal glycine-rich domain of TDP-43. All of these mutations are dominantly inherited missense changes with the exception of a truncating mutation (Y374X) at the extreme C terminus of the protein.3 In this study, in order to investigate the presence and frequency of TARDBP mutations in a cohort of 310 south Italian patients affected by ALS, we performed a mutational screening of the exon 4 and exon 6 of the gene in SOD1 -negative FALS and SALS patients. …

An upgraded optical coherence tomography system with integrated retinal tracker (TOCT) was developed. The upgraded system uses improved components to extend the tracking bandwidth, fully integrates the tracking hardware into the optical head of the clinical OCT system, and operates from a single software platform. The system was able to achieve transverse scan registration with sub-pixel accuracy (~10 microm). We demonstrate several advanced scan sequences with the TOCT, including composite scans averaged (co-added) from multiple B-scans taken consecutively and several hours apart, en face images collected by summing the A-scans of circular, line, and raster scans, and three-dimensional (3D) retinal maps of the fovea and optic disc. The new system achieves highly accurate OCT scan registration yielding composite images with significantly improved spatial resolution, increased signal-to-noise ratio, and reduced speckle while maintaining well-defined boundaries and sharp fine structure compared to single scans. Precise re-registration of multiple scans over separate imaging sessions demonstrates TOCT utility for longitudinal studies. En face images and 3D data cubes generated from these data reveal high fidelity image registration with tracking, despite scan durations of more than one minute.

Background/aims: To investigate retinal nerve fibre layer (RNFL) thickness measurement reproducibility using conventional time-domain optical coherence tomography (TD-OCT) and spectral-domain OCT (SD-OCT), and to evaluate two methods defining the optic nerve head (ONH) centring: Centred Each Time (CET) vs Centred Once (CO), in terms of RNFL thickness measurement variability on SD-OCT. Methods: Twenty-seven eyes (14 healthy subjects) had three circumpapillary scans with TD-OCT and three raster scans (three-dimensional or 3D image data) around ONH with SD-OCT. SD-OCT images were analysed in two ways: (1) CET: ONH centre was defined on each image separately and (2) CO: ONH centre was defined on one image and exported to other images after scan registration. After defining the ONH centre, a 3.4 mm diameter virtual circular OCT was resampled on SD-OCT images to mimic the conventional circumpapillary RNFL thickness measurements taken with TD-OCT. Results: CET and CO showed statistically significantly better reproducibility than TD-OCT except for 11:00 with CET. CET and CO methods showed similar reproducibility. Conclusions: SD-OCT 3D cube data generally showed better RNFL measurement reproducibility than TD-OCT. The choice of ONH centring methods did not affect RNFL measurement reproducibility.