SLC6A3 coding variant Ala559Val found in two autism probands alters dopamine transporter function and trafficking.

Emerging evidence associates dysfunction in the dopamine (DA) transporter (DAT) with the pathophysiology of autism spectrum disorder (ASD). The human DAT (hDAT; SLC6A3) rare variant with an Ala to Val substitution at amino acid 559 (hDAT A559V) was previously reported in individuals with bipolar disorder or attention-deficit hyperactivity disorder (ADHD). We have demonstrated that this variant is hyper-phosphorylated at the amino (N)-terminal serine (Ser) residues and promotes an anomalous DA efflux phenotype. Here, we report the novel identification of hDAT A559V in two unrelated ASD subjects and provide the first mechanistic description of its impaired trafficking phenotype. DAT surface expression is dynamically regulated by DAT substrates including the psychostimulant amphetamine (AMPH), which causes hDAT trafficking away from the plasma membrane. The integrity of DAT trafficking directly impacts DA transport capacity and therefore dopaminergic neurotransmission. Here, we show that hDAT A559V is resistant to AMPH-induced cell surface redistribution. This unique trafficking phenotype is conferred by altered protein kinase C ß (PKCß) activity. Cells expressing hDAT A559V exhibit constitutively elevated PKCß activity, inhibition of which restores the AMPH-induced hDAT A559V membrane redistribution. Mechanistically, we link the inability of hDAT A559V to traffic in response to AMPH to the phosphorylation of the five most distal DAT N-terminal Ser. Mutation of these N-terminal Ser to Ala restores AMPH-induced trafficking. Furthermore, hDAT A559V has a diminished ability to transport AMPH, and therefore lacks AMPH-induced DA efflux. Pharmacological inhibition of PKCß or Ser to Ala substitution in the hDAT A559V background restores AMPH-induced DA efflux while promoting intracellular AMPH accumulation. Although hDAT A559V is a rare variant, it has been found in multiple probands with neuropsychiatric disorders associated with imbalances in DA neurotransmission, including ADHD, bipolar disorder, and now ASD. These findings provide valuable insight into a new cellular phenotype (altered hDAT trafficking) supporting dysregulated DA function in these disorders. They also provide a novel potential target (PKCß) for therapeutic interventions in individuals with ASD.

De novo mutation in the dopamine transporter gene associates dopamine dysfunction with autism spectrum disorder.

De novo genetic variation is an important class of risk factors for autism spectrum disorder (ASD). Recently, whole-exome sequencing of ASD families has identified a novel de novo missense mutation in the human dopamine (DA) transporter (hDAT) gene, which results in a Thr to Met substitution at site 356 (hDAT T356M). The dopamine transporter (DAT) is a presynaptic membrane protein that regulates dopaminergic tone in the central nervous system by mediating the high-affinity reuptake of synaptically released DA, making it a crucial regulator of DA homeostasis. Here, we report the first functional, structural and behavioral characterization of an ASD-associated de novo mutation in the hDAT. We demonstrate that the hDAT T356M displays anomalous function, characterized as a persistent reverse transport of DA (substrate efflux). Importantly, in the bacterial homolog leucine transporter, substitution of A289 (the homologous site to T356) with a Met promotes an outward-facing conformation upon substrate binding. In the substrate-bound state, an outward-facing transporter conformation is required for substrate efflux. In Drosophila melanogaster, the expression of hDAT T356M in DA neurons-lacking Drosophila DAT leads to hyperlocomotion, a trait associated with DA dysfunction and ASD. Taken together, our findings demonstrate that alterations in DA homeostasis, mediated by aberrant DAT function, may confer risk for ASD and related neuropsychiatric conditions.

The next generation of patient education: multilingual Dental Explorer 3D.

The A3 Imperative (Anything, Anywhere, Anytime) that has left its mark on our information and knowledge society also characterizes the healthcare sector: we see "informed patients" who always consult the Web prior to their visit to the physician or dentist. The problem is that the knowledge concerning their suspected disease is often superficial. It is the task of the treatment provider to make factual information available and to discuss diagnostic aspects and therapeutic concepts with the patient, competently and based on the merits of the individual case. Dentistry is particularly affected by the online information trend, because the available restorative options cover a broad therapeutic spectrum with many conceivable alternatives that present a highly complex picture. Against this background, a dedicated three-dimensional multimedia software program was developed that visualizes all relevant individual dental treatment options in 3D as appropriate to the patient's oral status, actively supporting chairside communication. A 2D and 3D database containing more than 20,000 image and video files was created that visualizes--in several languages--the status of the individual patient and the planned restorative treatment. With this far-ranging concept, the process of patient-shared or participatory decision-making has been raised to new qualitative levels.

A closer look at amphetamine-induced reverse transport and trafficking of the dopamine and norepinephrine transporters.

Amphetamine (AMPH) and its derivatives are regularly used in the treatment of a wide array of disorders such as attention-deficit hyperactivity disorder (ADHD), obesity, traumatic brain injury, and narcolepsy (Prog Neurobiol 75:406-433, 2005; J Am Med Assoc 105:2051-2054, 1935; J Am Acad Child Adolesc Psychiatry 41:514-521, 2002; Neuron 43:261-269, 2004; Annu Rev Pharmacol Toxicol 47:681-698, 2007; Drugs Aging 21:67-79, 2004). Despite the important medicinal role for AMPH, it is more widely known for its psychostimulant and addictive properties as a drug of abuse. The primary molecular targets of AMPH are both the vesicular monoamine transporters (VMATs) and plasma membrane monoamine-dopamine (DA), norepinephrine (NE), and serotonin (5-HT)-transporters. The rewarding and addicting properties of AMPH rely on its ability to act as a substrate for these transporters and ultimately increase extracellular levels of monoamines. AMPH achieves this elevation in extracellular levels of neurotransmitter by inducing synaptic vesicle depletion, which increases intracellular monoamine levels, and also by promoting reverse transport (efflux) through plasma membrane monoamine transporters (J Biol Chem 237:2311-2317, 1962; Med Exp Int J Exp Med 6:47-53, 1962; Neuron 19:1271-1283, 1997; J Physiol 144:314-336, 1958; J Neurosci 18:1979-1986, 1998; Science 237:1219-1223, 1987; J Neurosc 15:4102-4108, 1995). This review will focus on two important aspects of AMPH-induced regulation of the plasma membrane monoamine transporters-transporter mediated monoamine efflux and transporter trafficking.

Stoned B mediates sorting of integral synaptic vesicle proteins.

A continuous supply of fusion-competent synaptic vesicles is essential for sustainable neurotransmission. Drosophila mutations of the dicistronic stoned locus disrupt normal vesicle cycling and cause functional deficits in synaptic transmission. Although both Stoned A and B proteins putatively participate in reconstituting synaptic vesicles, their precise function is still unclear. Here we investigate the effects of progressive depletion of Stoned B protein (STNB) on the release properties of neuromuscular synapses using a novel set of synthetic stnB hypomorphic alleles. Decreasing neuronal STNB expression to < or =35% of wild-type level causes a strong reduction in excitatory junctional current amplitude at low stimulation frequencies and a marked slowing in synaptic depression during high-frequency stimulation, suggesting vesicle depletion is attenuated by decreased release probability. Recovery from synaptic depression after prolonged stimulation is also decelerated in mutants, indicating a delayed recovery of fusion-ready vesicles. These phenotypes appear not to be due to a diminished vesicle population, since the docked vesicle pool is ultrastructurally unaffected, and the total number of vesicles is only slightly reduced in these hypomorphs, unlike lethal stoned mutants. Therefore, we conclude that STNB not only functions as an essential component of the endocytic complex for vesicle reconstitution, as previously proposed, but also regulates the competence of recycled vesicles to undergo fusion. In support of such role of STNB, synaptic levels of the vesicular glutamate transporter (vGLUT) and synaptotagmin-1 are strongly reduced with diminishing STNB function, while other synaptic proteins are largely unaffected. We conclude that STNB organizes the endocytic sorting of a subset of integral synaptic vesicle proteins thereby regulating the fusion-competence of the recycled vesicle.

Where, what, why: Mr. Q on the Web.

John Naisbitt, in his 1982 book "Megatrends," postulated an important characteristic of our information and knowledge society: "We are drowning in information but starved for knowledge." Today, in the age of the Information and Knowledge society, we are faced with this problem every single day, because Web searches and information selection are highly time-consuming activities. Internet search engines attempt to employ intelligent search algorithms in order to optimize their search results. Nevertheless, the question remains how "qualitative knowledge" can be selected, ie, knowledge needed for supporting decisions in medicine and dentistry. Semantic search engines are one current approach to this problem. For this reason, a project entitled "Mr. Q, your personal Web Assistant" has been initiated and will be introduced in this paper.

[A web-based e-learning tool in academic teaching of trauma surgery. First experiences and evaluation results]. / Webbasiertes E-Learning-Tool in der unfallchirurgischen Lehre. Erste Erfahrungen und Evaluationsergebnisse.

There are lots of possibilities for universities to offer contents of teaching to students by the Internet. Often the students can download slides or a special lecture note from the intranet of the university. Another way is to make a movie of the lecture and post this lecture movie on the Internet. In the Hanover Medical School we employed an alternative. It was developed by the Trauma Surgery Clinic and the Institute of Medical Informatics at the Hanover Medical School. Our goal was to use just one web-based content resource for the lecture and for the work at home. The Institute of Medical Informatics used a web-based content management system (CMS) Schoolbook to implement this e-learning application.Since October 2005 the Trauma Surgery Schoolbook has been used in the lecture on trauma surgery in all terms, and we evaluated the academic year 2005/2006. The results of the evaluation showed us that the students were very interested in using this e-learning application. The possibility to reinforce the learning material at home is a good chance for the students. Also the organisation of lectures was improved because the materials were all in one place. The lecturer needs to learn several new tasks, but we also got a positive response. Our experiences of the last academic year showed that it was a good way to use one web-based content resource for teaching and learning in the context of a lecture.

eLearning in education and advanced training in neuroradiology: introduction of a web-based teaching and learning application.

INTRODUCTION: New information technologies offer the possibility of major improvements in the professional education and advanced training of physicians. The web-based, multimedia teaching and learning application Schoolbook has been created and utilized for neuroradiology. METHODS: Schoolbook is technically based as a content management system and is realized in a LAMP environment. The content is generated with the help of the developed system and stored in a database. The layout is defined by a PHP application, and the webpages are generated from the system. RESULTS: Schoolbook is realized as an authoring tool so that it can be integrated into daily practice. This enables the teacher to autonomously process the content into the web-based application which is used for lectures, seminars and self-study. A multimedia case library is the central building block of Schoolbook for neuroradiology, whereby the learner is provided with original diagnostic and therapeutic data from numerous individual cases. The user can put individual emphasis on key learning points as there are various ways to work with the case histories. Besides the case-based way of teaching and learning, a systematically structured way of dealing with the content is available. CONCLUSION: eLearning offers various opportunities for teaching and learning in academic and scientific as well as in economic contexts. Web-based applications such as Schoolbook may be beneficial not only for basic university education but also for the realization of international educational programmes such as the European Master of Medical Science with a major in neuroradiology.

Evaluation of surface and volume rendering in 3D-CT of facial fractures.

OBJECTIVES: Three-dimensional computed tomography (3D-CT) of facial fractures has been reported as beneficial using surface (SR) and volume rendering (VR). There are controversial statements concerning the preferable algorithm. The purpose of this study was to evaluate and compare SR and VR for clinical 3D-CT in facial fractures on an experimental basis. METHODS: Multislice CT was obtained in 22 patients with facial fractures using two data acquisition protocols. Five SR and VR post-processing protocols were applied. Five assessors independently evaluated the quality of visualization of the fracture gap and dislocated fragments as well as the overall image quality using a five-point rating scale. The potential benefit of the 3D-images for radiological diagnosis and presentation was evaluated. The influence of the data acquisition protocol was analysed. RESULTS: SR in general achieved better evaluation scores than VR at corresponding thresholds. Variation of evaluation scores for all criteria was found for SR and VR depending on the segmentation threshold. Apart from the overall image quality no significant influence of the data acquisition technique was found for the evaluated criteria. CONCLUSIONS: SR provided sufficient and time efficient means for 3D-visualization of facial fractures in this study. No diagnostic benefit of VR over SR was found.

Use of 2D histograms for volume rendering of multidetector CT data: development of a graphical user interface.

RATIONALE AND OBJECTIVES: Direct volume rendering reveals 3D information on anatomic structures without preprocessing the data. This increases the interest in this technique as a diagnostic tool. A fast and simple method for setting transfer functions is crucial for clinical routine work. However, this is still a complex task. Present commercial workstations are usually limited to design galleries and window/level functionality. MATERIALS AND METHODS: We present a graphical user interface for volume rendering of multidetector row CT data that permits a much more flexible specification of rendering parameters. A 2D histogram of CT density versus gradient magnitude facilitates the understanding of the spatial connections of different tissues. The incorporation of gradient magnitude into the transfer function domain allows discrimination of features of interest that are not distinguishable on CT density alone. Penetration length, color, and gradient magnitude are depicted on a stack of 2D slices according to the settings of the opacity transfer function and the viewing direction. A gallery of thumbnails with presets of transfer functions is interactively adapted if the volume is rotated or cropped. RESULTS: This allows for fast evaluation of numerous rendering protocols at once. The interface was evaluated with CT data covering skeletal trauma, pathologies of the thorax/abdomen, and CT angiography. CONCLUSION: We observed that high-quality visualizations could be obtained with reasonable interaction times. The 2D histogram and penetration length displays provided valuable insight into the dataset that made the specification of transfer functions a goal-oriented process.

The learning unit "Orthodontic set-up" as a new-media module in teaching.

The present study examines the extent to which computer-assisted learning units provided independently of place and time are used in self-study as a supplement to the classical classroom instruction of dental students. Indications as to whether such teaching modules improve training in orthodontics should be obtained from this. Attention was focussed on the implementation and evaluation of the "Orthodontic set-up" teaching module, which can be accessed in the Internet and Intranet of the university. The didactic arrangement offered classical university courses in parallel (four lectures on the subjects of occlusion, function, diagnostics, and therapy) in addition to the electronically communicated teaching contents. In addition, intensive supervision during the production of the set-up was guaranteed. The use of this multimedia learning concept was in general assessed positively by 63 surveyed students in the 2002/03 winter semester. The results revealed on the one hand the intensity of use and features of the acquisition of knowledge (use types), and on the other hand, in terms of professional relevance, the contents were found to be well explained, didactically attractive, and understandably presented. However, numerous drawbacks were also mentioned (technical and time problems; qualification deficits). The experience gained in this project should encourage more future investment in the development of alternative university didactic models.

Orcein and litmus.

Orcein was separated into 14 dyes by partition chromatography. Their constitutions were determined mainly by spectroscopy and led to formulae that are derived from 7-amino-2-phenoxazone, 7-hydroxy-2-phenoxazone, and 7-amino-2-phenoxazime, and that were confirmed by syntheses. The major constituent of litmus is assembled polymerically from 7-hydroxy-2-phenoxazone chromophores. The mechanism of formation is elucidated.

Interactive PC-based volume rendering of CT datasets.

In radiology, the reading of large CT volumes is a time consuming task. Interactive volume rendering (iVRT) is a promising new technique. Using dedicated hardware (VP1000, Terarecon Inc.) it can now be realized on a standard PC in a cost effective manner. For this purpose, a program built using the Visualization Toolkit with integrated functionality for the VP 1000 is used for almost real-time iVRT (8-9 frames/second). It is possible to embed opaque and translucent polygon surfaces (e.g., segmented structures). By interactively varying the opacity, color and gradient transfer functions as well as using freely placable cutting planes, the visualization can easily be adapted to different diagnostic needs.

[Personality, accentuated traits and personality disorders. A contribution to dimensional diagnosis of personality disorders]. / Persönlichkeit, akzentuierte Wesenszüge und Persönlichkeitsstörungen. Ein Beitrag zur dimensionalen Diagnostik von Persönlichkeitsstörungen.

A dimensional diagnostic system for personality disorders (PD) postulates continuous transition from normal to disordered personalities (continuity hypothesis) and universal validity of basic personality dimensions (universal hypothesis). The present study investigates the validity of Leonhard's concept of attenuated personalities that define a conceptual link between normal personality dimensions and PD. Nine possible continuous transitions between three conceptual levels (Big Five personality factors, nine attenuated personality traits, nine PD) were tested by questionnaire data obtained from a mentally healthy (n = 166) and a clinical sample (n = 78). Both samples differed significantly in nearly all variables. However, they showed substantial similarity concerning the (in)validity of single continua and the complex structure of all variables as analyzed by multidimensional scaling. The concept of attenuated personalities could be validated for six out of nine tested continua and can be recommended for application in dimensional models of personality and personality disorders.

Drosophila fragile X-related gene regulates the MAP1B homolog Futsch to control synaptic structure and function.

Fragile X mental retardation gene (FMR1) encodes an RNA binding protein that acts as a negative translational regulator. We have developed a Drosophila fragile X syndrome model using loss-of-function mutants and overexpression of the FMR1 homolog (dfxr). dfxr nulls display enlarged synaptic terminals, whereas neuronal overexpression results in fewer and larger synaptic boutons. Synaptic structural defects are accompanied by altered neurotransmission, with synapse type-specific regulation in central and peripheral synapses. These phenotypes mimic those observed in mutants of microtubule-associated Futsch. Immunoprecipitation of dFXR shows association with futsch mRNA, and Western analyses demonstrate that dFXR inversely regulates Futsch expression. dfxr futsch double mutants restore normal synaptic structure and function. We propose that dFXR acts as a translational repressor of Futsch to regulate microtubule-dependent synaptic growth and function.

Orphan kinesin NOD lacks motile properties but does possess a microtubule-stimulated ATPase activity.

NOD is a Drosophila chromosome-associated kinesin-like protein that does not fall into the chromokinesin subfamily. Although NOD lacks residues known to be critical for kinesin function, we show that microtubules activate the ATPase activity of NOD >2000-fold. Biochemical and genetic analysis of two genetically identified mutations of NOD (NOD(DTW) and NOD("DR2")) demonstrates that this allosteric activation is critical for the function of NOD in vivo. However, several lines of evidence indicate that this ATPase activity is not coupled to vectorial transport, including 1) NOD does not produce microtubule gliding; and 2) the substitution of a single amino acid in the Drosophila kinesin heavy chain with the analogous amino acid in NOD results in a drastic inhibition of motility. We suggest that the microtubule-activated ATPase activity of NOD provides transient attachments of chromosomes to microtubules rather than producing vectorial transport.

Neurocan is dispensable for brain development.

Neurocan is a component of the extracellular matrix in brain. Due to its inhibition of neuronal adhesion and outgrowth in vitro and its expression pattern in vivo it was suggested to play an important role in axon guidance and neurite growth. To study the role of neurocan in brain development we generated neurocan-deficient mice by targeted disruption of the neurocan gene. These mice are viable and fertile and have no obvious deficits in reproduction and general performance. Brain anatomy, morphology, and ultrastructure are similar to those of wild-type mice. Perineuronal nets surrounding neurons appear largely normal. Mild deficits in synaptic plasticity may exist, as maintenance of late-phase hippocampal long-term potentiation is reduced. These data indicate that neurocan has either a redundant or a more subtle function in the development of the brain.

Telemedicine of the future: teleneuropathology.

The chief relevance of telemedicine lies in its capability to link medical practitioners and remote hospitals to larger or specialized facilities in a very fast electronic manner. This may become even more important due to current increase in subspecialization and demand for more precise diagnosis and consultation in difficult cases. A network attaching small clinics or laboratories to larger and more specialized units, and to highly specialized referral centers may improve the professional standard of health care services and education. For a wider use, a technological standardization will be required, since the existence of several types of computer and numerous image manipulation programs, have resulted in a proliferation of file formats. However, every potential user or client of telemedicine should keep in mind, that standardization also includes legal and ethical issues such as patient confidentiality and malpractice avoidance. The adoption of workable guidelines and protocols is required. Telepathology in general and teleneuropathology in particular is the practice of pathology at a distance, viewing digitized images of histological slides on a video monitor rather than directly through a light microscope. For the transmission of the digitized images from a telemicroscope to the remote diagnostic video monitor, different technologies such as ordinary telephone lines, broadband telecommunications channels, and the Internet can be used. The transmitted images may serve for primary neuropathological diagnosis, teleconsultation, quality assurance, proficiency testing, and distance learning. Static-imaging and dynamic-imaging are the two major competing technologies of telemicroscopy. Static-imaging systems appear to have levels of diagnostic accuracy which are not satisfactory for diagnostic neuropathology. On the contrary, high levels of diagnostic accuracy can be achieved using dynamic-imaging systems with the transmission of live video images in real time and by using a robotized telemicroscope with the possibility to examine the entire histological specimen under control of the remote teleneuropathologist.

Enhancement of glutamate release by L-fucose changes effects of glutamate receptor antagonists on long-term potentiation in the rat hippocampus.

In previous studies L-fucose has been shown to facilitate long-term memory formation and to enhance and prolong long-term potentiation (LTP). To search for possible presynaptic or postsynaptic mechanisms that are affected by L-fucose, we examined the effect of L-fucose on (1) inhibition of LTP induction via glutamate receptors by antagonists, (2) paired-pulse facilitation, and (3) presynaptic transmitter release. Coapplication of 0.2 mM L-fucose with the competitive N-methyl-D-aspartate (NMDA) receptor antagonist, D-2-amino-5-phosphonovalerate (AP5), or coapplication of 0.2 mM L-fucose in the presence of an inhibitor for class I/II metabotropic glutamate receptors, (S)-alpha-methyl-4-carboxyphenylglycine (MCPG), reversed LTP blockade in the CA1-region of hippocampal slices. In contrast, L-fucose had no effect on the LTP blockade by the noncompetitive NMDA ion-channel blocker (5R,10S)-(+)-5-Methyl-10, 11-dihydro-5H-dibenzo[a,d]cyclohepten-5, 10-imine hydrogen maleate (MK-801). Paired-pulse facilitation, which is a primarily presynaptic phenomenon of short-term plasticity, was decreased in the presence of 0.2 mM L-fucose. Furthermore, L-fucose enhanced the K(+)-stimulated release of [(3)H]-D-aspartate from preloaded hippocampal slices in a concentration-dependent manner. These observations demonstrate an influence of L-fucose on transmitter release that in turn can increase transmitter availability at postsynaptic glutamate receptors. This effect of L-fucose may contribute to the LTP facilitation seen in vitro and in vivo as well as to improvement in memory formation.