Chez les organismes complexes, comme les mammifères, les séquences de régulation génomique, dispersées sur les chromosomes, peuvent interagir à l'intérieur de l'espace nucléaire pour effectuer des actions coordonnées de régulations géniques. La méthylation de l'ADN et les modifications post-traductionnelles des histones, en combinaison avec des séquences de régulation, des facteurs protéiques et des ARNs non codants, conduisent à une organisation supérieure de la chromatine spécifique du type cellulaire. Cependant, l'organisation et la dynamique de la chromatine in vivo à l'échelle supérieure à celle du nucléosome reste encore largement méconnues. L'objectif général des travaux de notre équipe est d'élucider l'organisation de la chromatine à l'échelle supranucléosomale et sa dynamique in vivo, dans différents contextes physiologiques ou pathologiques, afin de comprendre leurs participations au contrôle et à la coordination de l'expression des gènes chez les mammifères. Notre hypothèse de travail est que certains compartiments nucléaires permettent un confinement de contacts chromatiniens spécifiques facilitant les régulations génomiques. L'objectif principal de mon travail de thèse était de développer une nouvelle méthode, simple et directe, permettant de cartographier et d'analyser les régions du génome murin qui sont associées aux compartiments nucléaires importants pour la régulation de l'expression des gènes (lamine nucléaire, les nucléoles, usines à transcription ou corps de Cajal). Le principe de notre méthode repose sur des traitements à haut sel de noyaux cellulaires transcriptionnellement actifs. Des séquençages à haut-débit permettent ensuite d'identifier les régions génomiques retenues dans les complexes nucléaires ainsi rendus d'insolubles. Elle a donc été appelée HRS-Seq : High-salt Recovered Sequences-sequencing (séquençage de séquences récupérées à haut-sel). Mon programme de travail s'est déroulé en 4 étapes distinctes : 1- la mise en œuvre et l'amélioration de la partie expérimentale (test HRS), 2- l'adaptation des techniques de séquençage à haut-débit à notre méthode (collaboration avec L. Journot, H. Parrinello, E. Dubois), 3 – l'application d'une analyse statistiques adéquate afin d'identifier les HRS (collaboration avec C. Reynes et R. Sabatier, statisticiens) et 4- l'analyse bio-informatique de ces régions destinée à les cartographier et à les caractériser (collaboration avec J. Mozziconacci et A. Cournac).Dans un premier temps, nous avons utilisé la méthode HRS-seq sur des noyaux de cellules de foie de souris. L'analyse bioinformatique des HRS nous a permis de réaliser la toute première cartographie de ces régions chez la souris et de découvrir leurs principales caractéristiques. Les régions HRS peuvent être classées en deux catégories distinctes : Les HRS riches en AT sont fortement associées à la lamine nucléaire, tandis que celles riches en GC sont associées aux régions géniques. La présence exceptionnelle, parmi cette dernière catégorie, des gènes codant pour les protéines d'histones, indique que le test HRS permet la rétention des Corps des Loci d'Histones (HLB – Histone Locus Body), un type spécifique de corps de Cajal. De plus, grâce à une analyse croisée avec des données de Hi-C disponibles dans la littérature, nous avons pu montrer que les HRS présentent entre-elles une haute probabilité de contact dans l'espace tridimensionnel du noyau, et qu'elles sont fortement enrichies en certaines séquences répétées (gènes des ARNt). L'ensemble de ces résultats nous permet de valider expérimentalement notre méthode. Dans un second temps, nous avons appliqué cette méthode à 3 autres types cellulaires : des cellules souches embryonnaires, des cellules progénitrices neurales et des neurones (collaboration avec T. Bouschet). Le but de ce travail est de déterminer comment les régions HRS évoluent au cours de la différentiation cellulaire. Les analyses statistiques et bioinformatiques sont en cours. / In complex organisms like mammals, regulatory sequences, dispersed on the chromosomes, can interact together within the nuclear space to tightly coordinate gene expression. DNA methylation and post-translational histone modifications combine with regulatory sequences, proteic factors and non-coding RNA, to provide cell-type specific patterns of higher-order chromatin organization. However, the in vivo organization of the mammalian chromatin beyond the simple nucleosomal array remains largely enigmatic. The general objective of our group is to elucidate the in vivo organization and dynamic of the chromatin at the supranucleosomal scale in diverse physiological and pathological contexts, in order to better understand how they are involved in the maintenance and coordination of gene expression in mammals. Our working hypothesis is that some nuclear compartments are confining specific chromatin contacts in order to facilitate genomic regulations. The principal objective of my thesis was to develop a novel straightforward method to map and to characterize genomic regions that are associated, in the mouse, with nuclear compartments that are important for gene regulation (nuclear lamina, nucleolus, transcription factories, Cajal bodies). The principle of our method is based on high-salt treatments of transcriptionally active cell nuclei. High-throughput sequencings then allow to identify the genomic regions that are retained in the resulting insoluble nuclear complexes. We thus named this method the HRS-seq (High-salt Recovered Sequences-sequencing). My working program was divided into 4 steps: 1- the improvement of the experimental procedure (HRS assay), 2- the adaptation of the NGS techniques to our method (collaboration with L. Journot, H. Parrinello, E. Dubois), 3- the use of an adequate statistical analysis in order to identify the HRS (Collaboration with C. Reynes and R. Sabatier, statisticians), 4- the bioinformatics analysis of these regions in order to map and to characterize them (collaboration with J. Mozziconacci and A. Cournac). We first used the HRS-seq method on mouse liver cells. The bioinformatics analysis allowed us to obtain the first global profiling of HRS in the mouse and to discover their essential characteristics. The HRS can be classified into two categories: the AT-rich HRS are linked to lamina associated domains, while GC-rich HRS are strongly associated to genes. The presence of histone genes amongst this latter category suggests that the Histone Locus Bodies (HLBs), a specific type of Cajal's body, is retained in the HRS assay. Furthermore, thanks to a cross-analysis with Hi-C data available in international databases, we have shown that the HRS display a high contact probability in the tri-dimensional space of the nucleus and that they are highly enriched in some specific repeat sequences (tRNA genes). Globally, these results allow us to validate the experimental approach used in the HRS-seq method. In a second time, we have applied this method to 3 other cell types: mouse embryonic stem cells, neural progenitor cells and neurons (collaboration with T. Bouschet). The aim of this work is to determine how the HRS regions are regulated during cell differentiation. Statistical and bio-informatics analyses are in progress.

Made available in DSpace on 2014-10-09T12:35:41Z (GMT). No. of bitstreams: 0 / Made available in DSpace on 2014-10-09T14:03:56Z (GMT). No. of bitstreams: 0 / Este trabalho apresenta dois modelos baseados em análise compartimental: modelo Animal e modelo Humano, usando imagens obtidas com mini gama câmera e dados de pacientes (obtidos da literatura) para a determinação das constantes cinéticas na biodistribuição do 177Lu-DOTATATO para três espécies animais (rato Wistar, hamster armênio e hamster sírio) e para o Humano. Os estudos de biodistribuição consideraram duas fases: a Fase 1, governada pela transferência do sangue para os órgãos e a Fase 2 governada pela excreção renal. As constantes cinéticas calculadas a partir dos dados obtidos com os animais foram usadas para a construção de escalas alométricas para prever a biodistribuição do radiofármaco 177Lu-DOTATATO empregando relações de massa, metabolismo, longevidade e parâmetros fisiológicos. Os resultados de extrapolação foram comparados com resultados calculados diretamente para os pacientes de PRRT (Peptide Receptor Radiotherapy) usando o modelo Humano. As constantes cinéticas calculadas a partir dos dados obtidos com humanos foram usadas para estimativa de dose em pacientes de PRRT considerando 26 órgãos e tecidos considerados pelo método MIRD. Os resultados de dosimetria foram concordantes com aqueles disponíveis na literatura. Para a Fase 1, as relações alométricas para as constantes cinéticas dos principais órgãos envolvidos no metabolismo e excreção do 177Lu-DOTATATO - fígado, rins e bexiga - mostraram boa correlação na projeção por massa, por metabolismo e por parâmetros fisiológicos. Para a Fase 2, apenas as constantes do sangue para os rins e do sangue para o fígado apresentaram boa correlação. Considerando o efeito de bloqueio da excreção renal pelo uso de anestésico, não se justificam tempos de medição maiores que 40 minutos para estudos in vivo, em pequenos animais. Dessa maneira, os resultados obtidos com as medições da Fase 1 da biodistribuição do 177Lu-DOTATATO em animais se apresentam como suficientes para estabelecer relações alométricas para estimativa de dose em pacientes submetidos à PRRT. / Tese (Doutoramento) / IPEN/T / Instituto de Pesquisas Energeticas e Nucleares - IPEN-CNEN/SP

L’acide phosphatidique (PA) est un lipide simple qui peut exister sous différentes formes. A partir des sondes que j’ai préparé en se basant sur des domaines de liaison au PA : PDE4A1, Spo20p, et OpiQ2, j’ai pu étudier la localisation subcellulaire du PA dans les cellules PC12 et les macrophages RAW264.7. Ces sondes lient différents formes de PA dans les membranes de différents compartiments subcellulaires. De plus, j’ai pu montrer qu’il y a une néosynthèse de PA et de certaines espèces de PA mono- ou bi-insaturé à la membrane plasmique lors de la stimulation de l’exocytose. Nous avons ainsi observé que la PLD1 semble être la source principale de PA dans les glandes surrénales. D’autre part, mes travaux indiquent une augmentation du niveau global de PA à la membrane plasmique et une diminution importante du PA au niveau du RE dans les macrophages après stimulation de la phagocytose frustrée. Ce qui pourrait ainsi valider le concept d’une fusion d’une partie de la membrane du RE avec la membrane plasmique lors de la phagocytose. / The phosphatidic acid (PA) is a simple lipid which may exist in various forms. I have generated probes based on PA binding domains: PDE4A1, Spo20p and OpiQ2 to study the subcellular localization of PA in PC12 cells and RAW264.7 macrophages. These probes bind different form of PA in different subcellular compartments. In addition, I show that PA and certain species mono- or bi-unsaturated of PA are synthesized at the plasma membrane upon stimulation of exocytosis. We observed that the PLD1 seems to be the main source of PA in the adrenal glands. On the other hand, my research indicates an increase in the level of PA at the plasma membrane and a significant decrease in the ER in macrophages after stimulation of phagocytosis frustrated. Thus these results could validate the concept of a fusion of a portion of the ER membrane with the plasma membrane during phagocytosis.

Les mathématiques sont comprises en tant qu’ensemble d’idées abstraites, dans le sens où le monde réel – ou plutôt réalité – n’a pas à intervenir. Pourtant, certains faits mathématiques observables dans des données expérimentales ou simulées peuvent être contre-intuitifs. La thèse est divisée en deux parties : premièrement, on étudie mathématiquement les matrices du genre celles dont nous avons discutées en biologie et finance. En particulier, nous mettons en évidence le fait contre-intuitif suivant : pour ces matrices, le vecteur propre associé à la plus haute valeur propre est très proche de la somme de chacune des lignes de la matrice, colonne par colonne. Nous discutons aussi d’applications en théorie des graphes avec bon nombre de simulations numériques. Dans un second temps, nous attaquons le problème des contacts géniques : à partir d’une carte de contact génique, un vrai défi actuel est de retrouver la structure tridimensionnelle de l’ADN. Nous proposons diverses méthodes d’analyse matricielle de données, dont une met en évidence l’existence, dans le noyau, de zones disjointes où les interactions sont de différents types. Ces zones sont des compartiments nucléaires. Avec d’autres données biologiques, nous mettons en évidence la fonction biologique de chacun de ces compartiments. Les outils d’analyses sont ceux utilisés en finance pour analyser des matrices d’auto-corrélation, ou même des séries temporelles. / Mathematics are understood as a set of abstract ideas, in the measure of the real world – or reality – has no way to intervene. However, some observable mathematical facts in experimental or simulated data can be counter-intuitive. The PhD is divided into two parts: first, we mathematically study the matrices of the same type of the ones in biology and finance. In particular, we show the following counter-intuitive fact: for these matrices, the eigenvector associated with the highest eigenvalue is close to the sum of each row, column by column. We also discuss some applications to graph theory with many numerical simulations and data analysis.On the other hand, we will face the genetic contact problem: from a contact map, a real current challenge is to find the DNA 3D-structure. We propose several matrix analysis methods, which one show disjoinct areas in the nucleus where the DNA interactions are different. These areas are nuclear compartments. With other biological features, we characterize the biological function of each of the compartments. The analysis tools are the ones already used in finance to analyze the autocorrelation matrices, or even time series.

Submitted by Fabio Sobreira Campos da Costa (fabio.sobreira@ufpe.br) on 2016-08-31T14:27:00Z No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) Dissertação Mestrado LUCYANA GALINDO ARCOVERDE VAZ.pdf: 1541733 bytes, checksum: 075c4e7b9b3f5f39a8de2b68a1df2f24 (MD5) / Made available in DSpace on 2016-08-31T14:27:00Z (GMT). No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) Dissertação Mestrado LUCYANA GALINDO ARCOVERDE VAZ.pdf: 1541733 bytes, checksum: 075c4e7b9b3f5f39a8de2b68a1df2f24 (MD5) Previous issue date: 2016-03-29 / CAPEs / A ação ergogênica da cafeína sobre a capacidade anaeróbia (CAN) está relacionada à melhora do desempenho. Entretanto, os estudos que investigaram o efeito da cafeína sobre a capacidade anaeróbia utilizaram apenas o máximo déficit acumulado de oxigênio (MAOD) como método de estimativa. Avaliar o efeito do consumo agudo de cafeína sobre a CAN estimada através dos principais métodos de estimativa, MAOD, potência crítica (ponto de intercepção y da função trabalho/tempo), fase rápida de recuperação do consumo de oxigênio + concentração de lactato (3 compartimentos) e eficiência mecânica grossa (EF), que por apresentarem medidas representativas da CAN, detectam igualmente os possíveis efeitos do consumo agudo de cafeína sobre o metabolismo anaeróbio. Nove participantes realizaram um teste incremental no cicloergômetro para determinação do primeiro limiar ventilatório (LV1) e do consumo máximo de oxigênio (VO2máx). A partir da carga referente ao LV1 foram calculadas a carga de aquecimento (90% do LV1), e as cargas dos testes experimentais. Foram doze testes experimentais, sendo seis com cafeína e seis placebo, em um desenho duplo-cego, contrabalanceado e controlado por placebo. A contribuição anaeróbia foi calculada pelos principais métodos de estimativa. Os achados mostraram que os métodos não detectam a mesma medida de CAN, logo, eles não são equivalentes na estimativa. Em relação ao efeito da cafeína sobre a estimativa sim, não houve diferença entre cafeína e placebo, exceto MAOD, que teve uma medida mais alta na situação cafeína. Além disso, não houve diferença no tempo de exaustão entre cafeína e placebo na carga supramáxima. Os métodos de estimativa não são equivalentes, eles não apresentam a mesma medida da CAN. A cafeína influenciou um dos métodos, MAOD, e isso levou a uma superestimativa da medida, quando corrigido pela estimativa do placebo, não apresentou mais diferença. Além disso, a cafeína não promoveu melhora no desempenho na carga supramáxima. / The caffeine ergogenic action on anaerobic capacity (AC) is related to performance’s improvement. However, some studies used only the maximal accumulated oxygen deficit (MAOD) as estimative method to investigate the effect of caffeine on anaerobic capacity. Evaluate the effect of acute caffeine consumption over the estimated AC through the main estimative methods, MAOD, critical power (interception point y work function/time), fast recovery phase of oxygen consumption + lactate concentration (3 compartments) and gross mechanical efficiency (GME), which by presenting representative measures of AC also detect the possible effects of acute caffeine consumption on the anaerobic metabolism. Nine participants performed an incremental test on a cycle ergometer to determine the first ventilatory threshold (VT1) and the maximal oxygen uptake (VO2max). From VT1 power output were calculated warm up load (90% of VT1), and the loads of the experimental trials. Twelve experimental trails were performed, being six after caffeine and six after placebo in a double-blind, placebo-controlled and counterbalanced design. AC was calculated by main estimative methods. The findings of the present study showed that AC was not detected similarly by the estimative methods, which might assume that they are not equivalent in the estimative. However, MAOD was higher in after caffeine ingestion when compared to placebo. There were no differences between trials in the other estimative methods. In addition, the time to exhaustion was not different between caffeine and placebo in supramaximal exercise. The estimative methods are not equivalent to measure AC. However, caffeine influenced MAOD, which probably overestimate the measurement. However, when corrected by the placebo estimative, no differences between trials was observed. Furthermore, caffeine was not efficient to improve the performance at supramaximal exercise.

Submitted by Luciana Ferreira (lucgeral@gmail.com) on 2017-04-12T12:11:16Z No. of bitstreams: 2 Dissertação - Tássia Andrielle Ponciano - 2017.pdf: 4132236 bytes, checksum: a03360d4ac30e9053e4834beac2d2830 (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) / Approved for entry into archive by Luciana Ferreira (lucgeral@gmail.com) on 2017-04-12T12:11:41Z (GMT) No. of bitstreams: 2 Dissertação - Tássia Andrielle Ponciano - 2017.pdf: 4132236 bytes, checksum: a03360d4ac30e9053e4834beac2d2830 (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) / Made available in DSpace on 2017-04-12T12:11:41Z (GMT). No. of bitstreams: 2 Dissertação - Tássia Andrielle Ponciano - 2017.pdf: 4132236 bytes, checksum: a03360d4ac30e9053e4834beac2d2830 (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Previous issue date: 2017-03-16 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES / The region of Vão do Paranã, inserted in the northeast (2000), is one of the three main centers of endemism in the Cerrado and goes through a process of fragmentation, which determines the ratings of potential and environmental vulnerabilities to promote appropriate policies of occupation. The study proposed here has as reference the integrated analysis of the landscape using the morphopedological compartments, to evaluate the dynamics of landscape structure in the microregion of Vão do Paranã. The methodology was based on the interpretation of Landsat images for temporal analysis (1984-2015) the use and soil coverage, with analysis of landscape structure through metrics and evaluation of morphopedological compartments. The results indicate that the characteristics associated morphopedological compartments the dynamics of occupation of the region respond by different forms of occupation of this area and the way it is structured the landscape in the region. Along the evolution of the use, the remaining vegetation concentrated in the eastern portion, coinciding with areas of emphasis more bustling, where currently the UC's; in the western portion, where the areas are more flat, there is a higher concentration of anthropic uses and high levels of fragmentation of the landscape. Although the agricultural expansion process has not yet strengthened in the region, the remaining areas were converted to 8.44% in anthropic areas, the pasture aptitude is present, however the agricultural areas have been growing exponentially in all compartments. The compartments favored, by relief, for anthropic occupation correspond to CMP I and III. The other compartments are indicated for the maintenance of environmental conservation. / A região do Vão do Paranã, inserida no nordeste goiano, é um dos três principais centros de endemismo do Cerrado e passa por um processo intenso de fragmentação, o que condiciona a avaliações de potencialidades e vulnerabilidades socioambientais que favoreçam políticas adequadas de ocupação. O estudo aqui proposto tem como referência a análise integrada da paisagem utilizando a morfocompartimentação, para avaliar a dinâmica da estrutura da paisagem na microrregião do Vão do Paranã. A metodologia fundamentou-se na interpretação de imagens Landsat para análise temporal (1984-2015) do uso e cobertura do solo, com análise de estrutura da paisagem por meio de métricas e avaliação dos compartimentos morfopedológicos. Os resultados indicam, que as características morfopedológicas associadas à dinâmica de ocupação da região respondem pelas diferentes formas de ocupação desta área e pela forma a qual está estruturada a paisagem na região. Ao longo da evolução do uso, a vegetação remanescente concentra se na porção leste, coincidindo com áreas de relevo mais movimentado, onde atualmente se encontram as UC´s; na porção oeste, onde as áreas são mais planas, há maior concentração dos usos antrópicos e elevados níveis de fragmentação da paisagem. Apesar do processo de expansão agrícola ainda não ter se fortalecido na região, as áreas de remanescente foram convertidas em 8,44% em áreas antropizadas, a aptidão para pastagem é presente, contudo as áreas de agricultura vêm crescendo exponencialmente em todos os compartimentos. Os compartimentos favorecidos, pelo relevo, para ocupação antrópica correspondem aos CMP I e III. Os demais compartimentos são indicados para manutenção da conservação ambiental.

This thesis deals with the relationship between property rights and the protection of environmental compartments. The thesis is divided into chapters corresponding to individual compartments, namely soil, minerals, energy, water, air, organisms and ecosystems. Each chapter defines given compartment and then describes its current Czech regulation. This thesis doesn't deal with all legislation in the area of environmental protection, but only with provisions related to ownership, namely whether given compartment can be an object of property rights and if possible whether its owner's treatment of this compartment can be restricted in the interest of its protection. Some chapters are further divided into sub-chapters due to quantity of laws concerning corresponding environmental compartment, e.g. the chapter about ownership of organisms differentiates animals from other organisms and further divides them into animals in captivity and free-living ones. Chapters about soil, minerals, water and organisms are ended with partial conclusions summarising my knowledge and thoughts about their respective topics, while the conclusion of the thesis as whole represents combination of synthesis of those partial conclusions and my opinions on overall legal regulation of property rights to environmetal compartments.

Cette étude cherche à évaluer le Débit de la Filtration Glomérulaire (DFG) dans le rein humain par IRM. L'estimation de ce paramètre quantitatif nécessite le suivi de la cinétique intrarénale de Gd après son injection en bolus. Pour atteindre cet objectif, nous avons développé sous IDEA Siemens une séquence SR-FLASH strictement pondérée en T1 capable de suivre en dynamique l'évolution du signal RMN après l'injection d'un bolus de produit de contraste. Cette séquence possède un codage de phase centré permettant de déterminer le contraste au début de la période d'acquisition de l'image. Nous avons également mis en oeuvre une séquence d'inversion IR-FLASH (avec codage de phase centré) permettant la mesure de la relaxation longitudinale en l'absence de produit de contraste, ce paramètre étant indispensable à la conversion de l'intensité du signal en concentration. Sachant que la relation entre l'intensité du signal RMN et la concentration de Gd n'est pas linéaire, nous avons proposé deux méthodes originales et malgré tout rapides et robustes pour convertir le signal RMN en concentration locale de Gd. Ceci nous a permis d'évaluer l'évolution de la concentration dans les deux reins et dans l'aorte au cours des premiers passages du produit. L'ajustement des ces mesures de concentration sur les équations décrivant une modélisation bicompartimentale de la fonction rénale a permis de calculer le DFG de chaque rein dans une population de cinq sujets possédant un fonctionnement rénal normal. / This study seeks to assess the Glomerular Filtration Rate (GFR) in the human kidney by MRI. To quantitatively estimate this parameter requires monitoring of the intrarenal kinetics of Gd after its bolus injection. To achieve this goal, we have developed under IDEA Siemens a SR-FLASH T1-weighted MRI sequence which can follow dynamic NMR signal changes after a bolus injection of Gd. This sequence has a centric phase-encoding scheme, and thus the image contrast was determined at the beginning of the acquisition period. Subsequently, we have implemented an IR-FLASH (with centric phase-encoding scheme) sequence to measure the longitudinal relaxation time in the absence of any injection of Gd ; this parameter is required to convert NMR signal intensities into Gd concentrations. knowing that the relation between the NMR signal intensities and the Gd concentrations is not linear, we have proposed two novel methods and yet fast and robust for conversion of the NMR signal intensities into local Gd concentration. This allowed us to estimate the temporal evolution of Gd concentrations in both kidneys and aorta. The fit of these concentrations measurements by a two-compartments model describing the function of the kidney allowed us to calculate the GFR of each kidney in a population of five subjects with normal renal function.

4′,6-Diamidine-2-phenylindole (DAPI) is a common fluorochrome that is able to bind to deoxyribonucleic acid (DNA) with distinct, sequence-dependent enhancement of fluorescence. This work presents the synthesis of a new multifunctional compound that includes the fluorescent dye as a ⁹⁹ᵐTechnetium (⁹⁹ᵐTc) carrier. A new technique for the bioconjugation of DAPI with 6-hydrazinonicotinic acid (HYNIC) through an amide linkage was developed. The radiolabelling was performed with HYNIC as a chelator and N-IJ2-hydroxy-1,1-bisIJhydroxymethyl)ethyl)glycine (tricine) as a coligand. Furthermore, experimental evidence showed that ⁹⁹ᵐTc complexes with DAPI as DNA-binding moieties are detectable in living Fischer rat thyroid follicular cell line 5 (FRTL5) and their nuclei. The investigations indicated further that the new HYNIC-DAPI derivative is able to interact with double-stranded DNA. This establishes the possibility of locating ⁹⁹ᵐTc in close proximity to biological structures of living cells, of which especially the genetic information-carrying cell compartments are at the centre of interest. In this context, further investigations are related to the radiotoxic effects of DNA-bound ⁹⁹ᵐTc-HYNIC-DAPI derivatives and dosimetric calculations.

Transcription factors of the helix-loop-helix (HLH) gene family are widespread in the animal kingdom. Among them, members of HLH subfamilies such as ASCL, Neurogenin, NeuroD, COE, Atonal, Oligo, NSCL, Hairy/E(spl) and Hey (here referred to as neural HLH genes) have been shown to be fundamental for the development of the nervous system. They are expressed at different time periods of neuronal differentiation, from the specification of ectoderm towards a neural lineage, to the ultimate differentiation of neurons. Few HLH genes have been identified in the lamprey; however, considering the wide diversity of HLH gene subfamilies in metazoans, including vertebrates, it is very likely that lampreys possess a large repertoire of HLH genes in their genome. In the present study, the identification of several HLH genes in the lamprey genome, as well as the isolation and expression of different lamprey neural HLH genes is reported. As expected, a wide repertoire of HLH genes was identified in the sea lamprey (Petromyzon marinus) genome. On the other hand, the identification and expression analysis of different neural HLH genes of the ASCL, Neurogenin, COE and Hairy/E(spl) in the brook lamprey Lampetra planeri showed an overall conservation with other vertebrates, both at the sequence and expression pattern levels. In addition, novel features of the lamprey nervous system are revealed, such as the identification of possible new sensory cranial placodes in pharyngeal arches. Furthermore, these genes can serve as molecular markers for different cranial placodes and dorsal root ganglia (DRG), and their expression also highlights the presence of a ventricular zone in the brain and spinal cord, along with a complementary marginal zone. Finally, with the use of a Notch pathway inhibitor in developing L. planeri embryos, the regulation of expression of the isolated genes by the Notch signaling pathway was shown to be generally conserved between lampreys and gnathostomes in the spinal cord. This functional study also revealed that the lamprey spinal cord likely presents an independent developmental programme from the brain. All together, the present study shows that the analysis of neural HLH genes represents an excellent tool to understand the lamprey nervous system.