Developmental Expression of Kv Potassium Channels at the Axon Initial Segment of Cultured Hippocampal Neurons

Axonal outgrowth and the formation of the axon initial segment (AIS) are early events in the acquisition of neuronal polarity. The AIS is characterized by a high concentration of voltage-dependent sodium and potassium channels. However, the specific ion channel subunits present and their precise localization in this axonal subdomain vary both during development and among the types of neurons, probably determining their firing characteristics in response to stimulation. Here, we characterize the developmental expression of different subfamilies of voltage-gated potassium channels in the AISs of cultured mouse hippocampal neurons, including subunits Kv1.2, Kv2.2 and Kv7.2. In contrast to the early appearance of voltage-gated sodium channels and the Kv7.2 subunit at the AIS, Kv1.2 and Kv2.2 subunits were tethered at the AIS only after 10 days in vitro. Interestingly, we observed different patterns of Kv1.2 and Kv2.2 subunit expression, with each confined to distinct neuronal populations. The accumulation of Kv1.2 and Kv2.2 subunits at the AIS was dependent on ankyrin G tethering, it was not affected by disruption of the actin cytoskeleton and it was resistant to detergent extraction, as described previously for other AIS proteins. This distribution of potassium channels in the AIS further emphasizes the heterogeneity of this structure in different neuronal populations, as proposed previously, and suggests corresponding differences in action potential regulation.

Aircraft flight trajectory deviation in the standard instrument departures. Operators penalties.

In the recent years many problems are emerging due to the aircraft noise on the airport surrounding areas. The solution to this problem is not easy considering that the neighbourhood asks for the reduction of the number of aircraft operations and the airlines ask for a growing demand in the number of operations in the major airports. So the airport and regulatory authorities try to get a solution imposing a fine to the aircraft which its actual trajectory differs from the nominal one more than a lateral deviation. But, which is the value of this deviation?. The current situation is that many operators have to pay a lot of money for exceeding a deviation which has been established without operational criteria. This paper presents the results of a research program which is being carried out by the authors which aims to determine the "delta" deviation to be used for this purpose. In addition it is proposed a customized method per SID and per airport to be used for determining the maximum allowed lateral deviation by which if the aircraft is within it, then none fine will be imposed.

AUTOFLY-Aid: Flight Deck Automation Support with Dynamic 4D Trajectory Management for Responsive and Adaptive Airborne Collision Avoidance

AUTOFLY-Aid Project aims to develop and demonstrate novel automation support algorithms and tools to the flight crew for flight critical collision avoidance using “dynamic 4D trajectory management”. The automation support system is envisioned to improve the primary shortcomings of TCAS, and to aid the pilot through add-on avionics/head-up displays and reality augmentation devices in dynamically evolving collision avoidance scenarios. The main theoretical innovative and novel concepts to be developed by AUTOFLY-Aid project are a) design and development of the mathematical models of the full composite airspace picture from the flight deck’s perspective, as seen/measured/informed by the aircraft flying in SESAR 2020, b) design and development of a dynamic trajectory planning algorithm that can generate at real-time (on the order of seconds) flyable (i.e. dynamically and performance-wise feasible) alternative trajectories across the evolving stochastic composite airspace picture (which includes new conflicts, blunder risks, terrain and weather limitations) and c) development and testing of the Collision Avoidance Automation Support System on a Boeing 737 NG FNPT II Flight Simulator with synthetic vision and reality augmentation while providing the flight crew with quantified and visual understanding of collision risks in terms of time and directions and countermeasures.

Iron distribution through the developmental stages of Medicago truncatula nodules.

Paramount to symbiotic nitrogen fixation (SNF) is the synthesis of a number of metalloenzymes that use iron as a critical component of their catalytical core. Since this process is carried out by endosymbiotic rhizobia living in legume root nodules, the mechanisms involved in iron delivery to the rhizobia-containing cells are critical for SNF. In order to gain insight into iron transport to the nodule, we have used synchrotron-based X-ray fluorescence to determine the spatio-temporal distribution of this metal in nodules of the legume Medicago truncatula with hitherto unattained sensitivity and resolution. The data support a model in which iron is released from the vasculature into the apoplast of the infection/differentiation zone of the nodule (zone II). The infected cell subsequently takes up this apoplastic iron and delivers it to the symbiosome and the secretory system to synthesize ferroproteins. Upon senescence, iron is relocated to the vasculature to be reused by the shoot. These observations highlight the important role of yet to be discovered metal transporters in iron compartmentalization in the nodule and in the recovery of an essential and scarce nutrient for flowering and seed production.

Contributions to trajectory prediction theory and its application to arrival management for air traffic control

La gestión del tráfico aéreo (Air Traffic Management, ATM) está experimentando un cambio de paradigma hacia las denominadas operaciones basadas trayectoria. Bajo dicho paradigma se modifica el papel de los controladores de tráfico aéreo desde una operativa basada su intervención táctica continuada hacia una labor de supervisión a más largo plazo. Esto se apoya en la creciente confianza en las soluciones aportadas por las herramientas automatizadas de soporte a la decisión más modernas. Para dar soporte a este concepto, se precisa una importante inversión para el desarrollo, junto con la adquisición de nuevos equipos en tierra y embarcados, que permitan la sincronización precisa de la visión de la trayectoria, basada en el intercambio de información entre ambos actores. Durante los últimos 30 a 40 años las aerolíneas han generado uno de los menores retornos de la inversión de entre todas las industrias. Sin beneficios tangibles, la industria aérea tiene dificultades para atraer el capital requerido para su modernización, lo que retrasa la implantación de dichas mejoras. Esta tesis tiene como objetivo responder a la pregunta de si las capacidades actualmente instaladas en las aeronaves comerciales se pueden aplicar para lograr la sincronización de la trayectoria con el nivel de calidad requerido. Además, se analiza en ella si, conjuntamente con mejoras en las herramientas de predicción trayectorias instaladas en tierra en para facilitar la gestión de las arribadas, dichas capacidades permiten obtener los beneficios esperados en el marco de las operaciones basadas en trayectoria. Esto podría proporcionar un incentivo para futuras actualizaciones de la aviónica que podrían llevar a mejoras adicionales. El concepto operacional propuesto en esta tesis tiene como objetivo permitir que los aviones sean pilotados de una manera consistente con las técnicas actuales de vuelo optimizado. Se permite a las aeronaves que desciendan en el denominado “modo de ángulo de descenso gestionado” (path-managed mode), que es el preferido por la mayoría de las compañías aéreas, debido a que conlleva un reducido consumo de combustible. El problema de este modo es que en él no se controla de forma activa el tiempo de llegada al punto de interés. En nuestro concepto operacional, la incertidumbre temporal se gestiona en mediante de la medición del tiempo en puntos estratégicamente escogidos a lo largo de la trayectoria de la aeronave, y permitiendo la modificación por el control de tierra de la velocidad de la aeronave. Aunque la base del concepto es la gestión de las ordenes de velocidad que se proporcionan al piloto, para ser capaces de operar con los niveles de equipamiento típicos actualmente, dicho concepto también constituye un marco en el que la aviónica más avanzada (por ejemplo, que permita el control por el FMS del tiempo de llegada) puede integrarse de forma natural, una vez que esta tecnología este instalada. Además de gestionar la incertidumbre temporal a través de la medición en múltiples puntos, se intenta reducir dicha incertidumbre al mínimo mediante la mejora de las herramienta de predicción de la trayectoria en tierra. En esta tesis se presenta una novedosa descomposición del proceso de predicción de trayectorias en dos etapas. Dicha descomposición permite integrar adecuadamente los datos de la trayectoria de referencia calculada por el Flight Management System (FMS), disponibles usando Futuro Sistema de Navegación Aérea (FANS), en el sistema de predicción de trayectorias en tierra. FANS es un equipo presente en los aviones comerciales de fuselaje ancho actualmente en la producción, e incluso algunos aviones de fuselaje estrecho pueden tener instalada avionica FANS. Además de informar automáticamente de la posición de la aeronave, FANS permite proporcionar (parte de) la trayectoria de referencia en poder de los FMS, pero la explotación de esta capacidad para la mejora de la predicción de trayectorias no se ha estudiado en profundidad en el pasado. La predicción en dos etapas proporciona una solución adecuada al problema de sincronización de trayectorias aire-tierra dado que permite la sincronización de las dimensiones controladas por el sistema de guiado utilizando la información de la trayectoria de referencia proporcionada mediante FANS, y también facilita la mejora en la predicción de las dimensiones abiertas restantes usado un modelo del guiado que explota los modelos meteorológicos mejorados disponibles en tierra. Este proceso de predicción de la trayectoria de dos etapas se aplicó a una muestra de 438 vuelos reales que realizaron un descenso continuo (sin intervención del controlador) con destino Melbourne. Dichos vuelos son de aeronaves del modelo Boeing 737-800, si bien la metodología descrita es extrapolable a otros tipos de aeronave. El método propuesto de predicción de trayectorias permite una mejora en la desviación estándar del error de la estimación del tiempo de llegada al punto de interés, que es un 30% menor que la que obtiene el FMS. Dicha trayectoria prevista mejorada se puede utilizar para establecer la secuencia de arribadas y para la asignación de las franjas horarias para cada aterrizaje (slots). Sobre la base del slot asignado, se determina un perfil de velocidades que permita cumplir con dicho slot con un impacto mínimo en la eficiencia del vuelo. En la tesis se propone un nuevo algoritmo que determina las velocidades requeridas sin necesidad de un proceso iterativo de búsqueda sobre el sistema de predicción de trayectorias. El algoritmo se basa en una parametrización inteligente del proceso de predicción de la trayectoria, que permite relacionar el tiempo estimado de llegada con una función polinómica. Resolviendo dicho polinomio para el tiempo de llegada deseado, se obtiene de forma natural el perfil de velocidades optimo para cumplir con dicho tiempo de llegada sin comprometer la eficiencia. El diseño de los sistemas de gestión de arribadas propuesto en esta tesis aprovecha la aviónica y los sistemas de comunicación instalados de un modo mucho más eficiente, proporcionando valor añadido para la industria. Por tanto, la solución es compatible con la transición hacia los sistemas de aviónica avanzados que están desarrollándose actualmente. Los beneficios que se obtengan a lo largo de dicha transición son un incentivo para inversiones subsiguientes en la aviónica y en los sistemas de control de tráfico en tierra. ABSTRACT Air traffic management (ATM) is undergoing a paradigm shift towards trajectory based operations where the role of an air traffic controller evolves from that of continuous intervention towards supervision, as decision making is improved based on increased confidence in the solutions provided by advanced automation. To support this concept, significant investment for the development and acquisition of new equipment is required on the ground as well as in the air, to facilitate the high degree of trajectory synchronisation and information exchange required. Over the past 30-40 years the airline industry has generated one of the lowest returns on invested capital among all industries. Without tangible benefits realised, the airline industry may find it difficult to attract the required investment capital and delay acquiring equipment needed to realise the concept of trajectory based operations. In response to these challenges facing the modernisation of ATM, this thesis aims to answer the question whether existing aircraft capabilities can be applied to achieve sufficient trajectory synchronisation and improvements to ground-based trajectory prediction in support of the arrival management process, to realise some of the benefits envisioned under trajectory based operations, and to provide an incentive for further avionics upgrades. The proposed operational concept aims to permit aircraft to operate in a manner consistent with current optimal aircraft operating techniques. It allows aircraft to descend in the fuel efficient path managed mode as preferred by a majority of airlines, with arrival time not actively controlled by the airborne automation. The temporal uncertainty is managed through metering at strategically chosen points along the aircraft’s trajectory with primary use of speed advisories. While the focus is on speed advisories to support all aircraft and different levels of equipage, the concept also constitutes a framework in which advanced avionics as airborne time-of-arrival control can be integrated once this technology is widely available. In addition to managing temporal uncertainty through metering at multiple points, this temporal uncertainty is minimised by improving the supporting trajectory prediction capability. A novel two-stage trajectory prediction process is presented to adequately integrate aircraft trajectory data available through Future Air Navigation Systems (FANS) into the ground-based trajectory predictor. FANS is standard equipment on any wide-body aircraft in production today, and some single-aisle aircraft are easily capable of being fitted with FANS. In addition to automatic position reporting, FANS provides the ability to provide (part of) the reference trajectory held by the aircraft’s Flight Management System (FMS), but this capability has yet been widely overlooked. The two-stage process provides a ‘best of both world’s’ solution to the air-ground synchronisation problem by synchronising with the FMS reference trajectory those dimensions controlled by the guidance mode, and improving on the prediction of the remaining open dimensions by exploiting the high resolution meteorological forecast available to a ground-based system. The two-stage trajectory prediction process was applied to a sample of 438 FANS-equipped Boeing 737-800 flights into Melbourne conducting a continuous descent free from ATC intervention, and can be extrapolated to other types of aircraft. Trajectories predicted through the two-stage approach provided estimated time of arrivals with a 30% reduction in standard deviation of the error compared to estimated time of arrival calculated by the FMS. This improved predicted trajectory can subsequently be used to set the sequence and allocate landing slots. Based on the allocated landing slot, the proposed system calculates a speed schedule for the aircraft to meet this landing slot at minimal flight efficiency impact. A novel algorithm is presented that determines this speed schedule without requiring an iterative process in which multiple calls to a trajectory predictor need to be made. The algorithm is based on parameterisation of the trajectory prediction process, allowing the estimate time of arrival to be represented by a polynomial function of the speed schedule, providing an analytical solution to the speed schedule required to meet a set arrival time. The arrival management solution proposed in this thesis leverages the use of existing avionics and communications systems resulting in new value for industry for current investment. The solution therefore supports a transition concept from mixed equipage towards advanced avionics currently under development. Benefits realised under this transition may provide an incentive for ongoing investment in avionics.

Analysis of the geometric altimetry to support aircraft optimal trajectories within the future 4d trajectory management

Over the past few years, the common practice within air traffic management has been that commercial aircraft fly by following a set of predefined routes to reach their destination. Currently, aircraft operators are requesting more flexibility to fly according to their prefer- ences, in order to achieve their business objectives. Due to this reason, much research effort is being invested in developing different techniques which evaluate aircraft optimal trajectory and traffic synchronisation. Also, the inefficient use of the airspace using barometric altitude overall in the landing and takeoff phases or in Continuous Descent Approach (CDA) trajectories where currently it is necessary introduce the necessary reference setting (QNH or QFE). To solve this problem and to permit a better airspace management born the interest of this research. Where the main goals will be to evaluate the impact, weakness and strength of the use of geometrical altitude instead of the use of barometric altitude. Moreover, this dissertation propose the design a simplified trajectory simulator which is able to predict aircraft trajectories. The model is based on a three degrees of freedom aircraft point mass model that can adapt aircraft performance data from Base of Aircraft Data, and meteorological information. A feature of this trajectory simulator is to support the improvement of the strategic and pre-tactical trajectory planning in the future Air Traffic Management. To this end, the error of the tool (aircraft Trajectory Simulator) is measured by comparing its performance variables with actual flown trajectories obtained from Flight Data Recorder information. The trajectory simulator is validated by analysing the performance of different type of aircraft and considering different routes. A fuel consumption estimation error was identified and a correction is proposed for each type of aircraft model. In the future Air Traffic Management (ATM) system, the trajectory becomes the fundamental element of a new set of operating procedures collectively referred to as Trajectory-Based Operations (TBO). Thus, governmental institutions, academia, and industry have shown a renewed interest for the application of trajectory optimisation techniques in com- mercial aviation. The trajectory optimisation problem can be solved using optimal control methods. In this research we present and discuss the existing methods for solving optimal control problems focusing on direct collocation, which has received recent attention by the scientific community. In particular, two families of collocation methods are analysed, i.e., Hermite-Legendre-Gauss-Lobatto collocation and the pseudospectral collocation. They are first compared based on a benchmark case study: the minimum fuel trajectory problem with fixed arrival time. For the sake of scalability to more realistic problems, the different meth- ods are also tested based on a real Airbus 319 El Cairo-Madrid flight. Results show that pseudospectral collocation, which has shown to be numerically more accurate and computa- tionally much faster, is suitable for the type of problems arising in trajectory optimisation with application to ATM. Fast and accurate optimal trajectory can contribute properly to achieve the new challenges of the future ATM. As atmosphere uncertainties are one of the most important issues in the trajectory plan- ning, the final objective of this dissertation is to have a magnitude order of how different is the fuel consumption under different atmosphere condition. Is important to note that in the strategic phase planning the optimal trajectories are determined by meteorological predictions which differ from the moment of the flight. The optimal trajectories have shown savings of at least 500 [kg] in the majority of the atmosphere condition (different pressure, and temperature at Mean Sea Level, and different lapse rate temperature) with respect to the conventional procedure simulated at the same atmosphere condition.This results show that the implementation of optimal profiles are beneficial under the current Air traffic Management (ATM).

Effect of the pulse trajectory on ultrasonic fluid velocity measurement

In a general situation a non-uniform velocity field gives rise to a shift of the otherwise straight acoustic pulse trajectory between the transmitter and receiver transducers of a sonic anemometer. The aim of this paper is to determine the effects of trajectory shifts on the velocity as measured by the sonic anemometer. This determination has been accomplished by developing a mathematical model of the measuring process carried out by sonic anemometers; a model which includes the non-straight trajectory effect. The problem is solved by small perturbation techniques, based on the relevant small parameter of the problem, the Mach number of the reference flow, M. As part of the solution, a general analytical expression for the deviations of the computed measured speed from the nominal speed has been obtained. The correction terms of both the transit time and of the measured speed are of M 2 order in rotational velocity field. The method has been applied to three simple, paradigmatic flows: one-directional horizontal and vertical shear flows, and mixed with a uniform horizontal flow.

Developmental emergence of different forms of neuromodulation in Aplysia sensory neurons

The capacity for neuromodulation and biophysical plasticity is a defining feature of most mature neuronal cell types. In several cases, modulation at the level of the individual neuron has been causally linked to changes in the functional output of a neuronal circuit and subsequent adaptive changes in the organism’s behavioral responses. Understanding how such capacity for neuromodulation develops therefore may provide insights into the mechanisms both of neuronal development and learning and memory. We have examined the development of multiple forms of neuromodulation triggered by a common neurotransmitter, serotonin, in the pleural sensory neurons of Aplysia californica. We have found that multiple signaling cascades within a single neuron develop sequentially, with some being expressed only very late in development. In addition, our data suggest a model in which, within a single neuromodulatory pathway, the elements of the signaling cascade are developmentally expressed in a “retrograde” manner with the ionic channel that is modulated appearing early in development, functional elements in the second messenger cascade appearing later, and finally, coupling of the second messenger cascade to the serotonin receptor appearing quite late. These studies provide the characterization of the development of neuromodulation at the level of an identified cell type and offer insights into the potential roles of neuromodulatory processes in development and adult plasticity.

Effectors of a developmental mitogen-activated protein kinase cascade revealed by expression signatures of signaling mutants

Despite the importance of mitogen-activated protein kinase (MAPK) signaling in eukaryotic biology, the mechanisms by which signaling yields phenotypic changes are poorly understood. We have combined transcriptional profiling with genetics to determine how the Kss1 MAPK signaling pathway controls dimorphic development in Saccharomyces cerevisiae. This analysis identified dozens of transcripts that are regulated by the pathway, whereas previous work had identified only a single downstream target, FLO11. One of the MAPK-regulated genes is PGU1, which encodes a secreted enzyme that hydrolyzes polygalacturonic acid, a structural barrier to microbial invasion present in the natural plant substrate of S. cerevisiae. A third key transcriptional target is the G1 cyclin gene CLN1, a morphogenetic regulator that we show to be essential for pseudohyphal growth. In contrast, the homologous CLN2 cyclin gene is dispensable for development. Thus, the Kss1 MAPK cascade programs development by coordinately modulating a cell adhesion factor, a secreted host-destroying activity, and a specialized subunit of the Cdc28 cyclin-dependent kinase.

Developmental exposure to vasopressin increases aggression in adult prairie voles

Although the biological roots of aggression have been the source of intense debate, the precise physiological mechanisms responsible for aggression remain poorly understood. In most species, aggression is more common in males than females; thus, gonadal hormones have been a focal point for research in this field. Although gonadal hormones have been shown to influence the expression of aggression, in many cases aggression can continue after castration, indicating that testicular steroids are not completely essential for the expression of aggression. Recently, the mammalian neuropeptide arginine vasopressin (AVP) has been implicated in aggression. AVP plays a particularly important role in social behavior in monogamous mammals, such as prairie voles (Microtus ochrogaster). In turn, the effects of social experiences may be mediated by neuropeptides, including AVP. For example, sexually naïve prairie voles are rarely aggressive. However, 24 h after the onset of mating, males of this species become significantly aggressive toward strangers. Likewise, in adult male prairie voles, central (intracerebroventricular) injections of AVP can significantly increase intermale aggression, suggesting a role for AVP in the expression of postcopulatory aggression in adult male prairie voles. In this paper, we demonstrate that early postnatal exposure to AVP can have long-lasting effects on the tendency to show aggression, producing levels of aggression in sexually naïve, adult male prairie voles that are comparable to those levels observed after mating. Females showed less aggression and were less responsive to exogenous AVP, but the capacity of an AVP V1a receptor antagonist to block female aggression also implicates AVP in the development of female aggression.

Desynchronization of cells on the developmental path triggers the formation of spiral waves of cAMP during Dictyostelium aggregation

Whereas it is relatively easy to account for the formation of concentric (target) waves of cAMP in the course of Dictyostelium discoideum aggregation after starvation, the origin of spiral waves remains obscure. We investigate a physiologically plausible mechanism for the spontaneous formation of spiral waves of cAMP in D. discoideum. The scenario relies on the developmental path associated with the continuous changes in the activity of enzymes such as adenylate cyclase and phosphodiesterase observed during the hours that follow starvation. These changes bring the cells successively from a nonexcitable state to an excitable state in which they relay suprathreshold cAMP pulses, and then to autonomous oscillations of cAMP, before the system returns to an excitable state. By analyzing a model for cAMP signaling based on receptor desensitization, we show that the desynchronization of cells on this developmental path triggers the formation of fully developed spirals of cAMP. Developmental paths that do not correspond to the sequence of dynamic transitions no relay-relay-oscillations-relay are less able or fail to give rise to the formation of spirals.

Self-antigen does not accelerate immature B cell apoptosis, but stimulates receptor editing as a consequence of developmental arrest

In pre-B lymphocytes, productive rearrangement of Ig light chain genes allows assembly of the B cell receptor (BCR), which selectively promotes further developmental maturation through poorly defined transmembrane signaling events. Using a novel in vitro system to study immune tolerance during development, we find that BCR reactivity to auto-antigen blocks this positive selection, preventing down-regulation of light chain gene recombination and promoting secondary light chain gene rearrangements that often alter BCR specificity, a process called receptor editing. Under these experimental conditions, self-antigen induces secondary light chain gene rearrangements in at least two-thirds of autoreactive immature B cells, but fails to accelerate cell death at this stage. These data suggest that in these cells the mechanism of immune tolerance is receptor selection rather than clonal selection.

5-HT2A and 5-HT2C receptors as hypothalamic targets of developmental programming in male rats

Funding The research reported in this publication was supported by the Biotechnology and Biological Sciences Research Council (E007821/1 to M.S.M-G, R.L.C and E00797X/1; BB/K001418 /1 to L.K.H), the British Heart Foundation (FS/09/029/27902 to S.E.O.), the UK Medical Research Council Metabolic Diseases Unit (MC_UU_12012/4 to S.E.O and MC_UU_12012/1 to G.S.H.Y), the Wellcome Trust (WT081713 and WT098012 to L.K.H), the European Union (FP7-HEALTH-266408 Full4Health to G.S.H.Y) and the Helmholtz Alliance ICEMED to G.S.H.Y.

Developmental disorder associated with increased cellular nucleotidase activity

Four unrelated patients are described with a syndrome that included developmental delay, seizures, ataxia, recurrent infections, severe language deficit, and an unusual behavioral phenotype characterized by hyperactivity, short attention span, and poor social interaction. These manifestations appeared within the first few years of life. Each patient displayed abnormalities on EEG. No unusual metabolites were found in plasma or urine, and metabolic testing was normal except for persistent hypouricosuria. Investigation of purine and pyrimidine metabolism in cultured fibroblasts derived from these patients showed normal incorporation of purine bases into nucleotides but decreased incorporation of uridine. De novo synthesis of purines and cellular phosphoribosyl pyrophosphate content also were moderately decreased. The distribution of incorporated purines and pyrimidines did not reveal a pattern suggestive of a deficient enzyme activity. Assay of individual enzymes in fibroblast lysates showed no deficiencies. However, the activity of cytosolic 5′-nucleotidase was elevated 6- to 10-fold. Based on the possibility that the observed increased catabolic activity and decreased pyrimidine salvage might be causing a deficiency of pyrimidine nucleotides, the patients were treated with oral pyrimidine nucleoside or nucleotide compounds. All patients showed remarkable improvement in speech and behavior as well as decreased seizure activity and frequency of infections. A double-blind placebo trial was undertaken to ascertain the efficacy of this supplementation regimen. Upon replacement of the supplements with placebo, all patients showed rapid regression to their pretreatment states. These observations suggest that increased nucleotide catabolism is related to the symptoms of these patients, and that the effects of this increased catabolism are reversed by administration of uridine.

Developmental changes in DNA methylation of the two tobacco pollen nuclei during maturation

Changes in DNA methylation during tobacco pollen development have been studied by confocal fluorescence microscopy using a monoclonal anti-5-methylcytosine (anti-m5C) antibody and a polyclonal anti-histone H1 (anti-histone) antibody as an internal standard. The specificity of the anti-m5C antibody was demonstrated by a titration series against both single-stranded DNA and double-stranded DNA substrates in either the methylated or unmethylated forms. The antibody was found to show similar kinetics against both double- and single-stranded DNA, and the fluorescence was proportional to the amount of DNA used. No signal was observed with unmethylated substrates. The extent of methylation of the two pollen nuclei remained approximately constant after the mitotic division that gave rise to the vegetative and generative nuclei. However, during the subsequent development of the pollen, the staining of the generative nucleus decreased until it reached a normalized value of \documentclass[12pt]{minimal} \usepackage{wasysym} \usepackage{amsmath} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \setlength{\oddsidemargin}{-69pt} \begin{document} \begin{equation*}\frac{1}{5}\end{equation*}\end{document} of that of the vegetative nucleus. The use of a confocal microscope makes these data independent of possible focusing artefacts. The anti-histone antibody was used as a control to show that, while the antibody staining directed against 5-methylcytosine changed dramatically during pollen maturation, the histone signal did not. We observed the existence of structural dimorphism amongst tobacco pollen grains, the majority having three pollen apertures and the rest with four. However, the methylation changes observed occurred to the same extent in both subclasses.