In Optogenetics, cells, e.g. neurons or cardiac cells, are genetically altered to produce for example the lightsensitive protein Channelrhodopsin-2. Illuminating these cells induces action potentials or contractions and therefore allows to control electrical activity. Thus, light-induced cell stimulation can be used to gain insight to various biological processes. Many optogenetics studies, however, use only full field illumination and thus gain no local information about their specimen. But using modern spatial light modulators (SLM) in conjunction with computer-generated holograms (CGH), cells may be stimulated locally, thus enabling the research of the foundations of cell networks and cell communications. In our contribution, we present a digital holographic system for the patterned, spatially resolved stimulation of cell networks. We employ a fast ferroelectric liquid crystal on silicon SLM to display CGH at up to 1.7 kHz. With an effective working distance of 33 mm, we achieve a focus of 10 µm at a positioning accuracy of the individual foci of about 8 µm. We utilized our setup for the optogenetic stimulation of clusters of cardiac cells derived from induced pluripotent stem cells and were able to observe contractions correlated to both temporal frequency and spatial power distribution of the light incident on the cell clusters.

This Thesis contains details of the construction and characterisation of a compact apparatus for the cooling of ultracold atoms to quantum degeneracy, and their manipulation in flexible holographic optical traps. We have designed and built two iterations of this apparatus. The first version consists of a stainless steel single-cell vacuum chamber, in which we confine ⁸⁷Rb and ⁶Li or ⁷Li in a Magneto-Optical Trap. We characterise the alternative methods of pulsed atomic dispenser and Light Induced Atomic Desorption (LIAD) to rapidly vary the background pressure in the vacuum chamber with the view to enabling efficient evaporative cooling in the single chamber, loading MOTs of up to 10⁸ atoms using pulsed dispensers. The LIAD is found to be ineffective in loading large MOTs in this setup, while the pulsed dispensers method gradually increases the background pressure in the chamber over time. Based on the results of this first iteration, we designed and built a second single-chamber apparatus for cooling of ⁸⁷Rb to quantum degeneracy. The LIAD technique was used to successfully load MOTs containing 8x10⁷ atoms in this single pyrex cell with a rapidly-varying background pressure. The lifetime of an atomic cloud loaded from the MOT into a magnetic trap increased by a factor of 6 when LIAD was used. The holographic optical traps for cold atoms are generated using a Spatial Light Modulator, and we present our novel method for improving the quality of holographic light patterns to the point where they are suitable for trapping ultracold atoms using a feedback algorithm. As demonstrations of this new capability, we show power-law optical traps which provide an efficient, reversible route to Bose-Einstein Condensation and a dynamic ring trap for the investigation of superfluidity in cold atoms.

Dois tipos diferentes de hologramas (Fresnel e Fourier) foram gerados por computador. O holograma de Fresnel foi escolhido para fazer parte de um arranjo experimental que teve como objetivo estimar a rugosidade de amostras metálicas. Para isso, um novo método de aplicação de um correlator óptico foi desenvolvido. Hologramas de Fourier geralmente fazem parte do correlator óptico de VanderLugt, o qual é muito utilizado no reconhecimento de padrões. A reconstrução numérica de hologramas de Fresnel gerados por computador, "distorcidos" por superfícies metálicas (também simuladas), serviram de base para que a reconstrução óptica de um holograma de Fresnel fosse utilizada, de forma inédita, no reconhecimento de padrões para estimar a rugosidade de amostras metálicas. / Two different types of holograms (Fresnel and Fourier) have been computer-generated. The Fresnel hologram has been chosen as part of an experimental set, which meant to estimate the roughness of the metalic samples. A new method for the aplication of an optical correlator has been developed. Fourier holograms are, generally, part of the VanderLugt optical correlator, that is very used for pattern recognition. The numerical reconstruction of the computer-generated Fresnel holograms, "distorted" by metalic surfaces (simulated as well), worked as the base for the utilization, in an original way, of the optical reconstruction of a Fresnel hologram, at the pattern recognition to estimate the roughness of the metalic samples.

A absorção intestinal de fármacos (HIA, na sigla em inglês para human intestinal absorption) é um dos processos farmacocinéticos mais críticos para a ação de medicamentos administrados por via oral. Modelos in vitro baseados em células que mimetizam os enterócitos do intestino humano (células Caco-2), ou aqueles que utilizam membranas articifiais são muito utilizados no estudo da permeabilidade celular de moléculas bioativas. No processo de planejamento de fármacos, estas técnicas são frequentemente empregadas em paralelo com estratégias computacionais na avaliação de propriedades físico-químicas e farmacocinéticas. Nessa tese de doutorado foram desenvolvidos modelos computacionais para a predição da permeabilidade celular de compostos bioativos em células Caco-2 e em membrana artificial paralela (PAMPA, na sigla em inglês para Parallel Artificial Membrane Permeability Assay). Os modelos foram gerados através do método baseado em fragmentos moleculares holograma QSAR (HQSAR). Os indicadores estatísticos obtidos indicam a boa capacidade de correlação dos modelos Caco-2 e PAMPA para os dados utilizados na modelagem e o alto poder de predição destes modelos para a permeabilidade de um conjunto teste de compostos. Adicionalmente, os modelos foram avaliados quanto a sua capacidade preditiva frente a dois conjuntos de validação, constituídos por novas moléculas que foram planejadas e testadas experimentalmente durante o desenvolvimento deste trabalho. Para estes conjuntos, os modelos demonstraram alta capacidade de predição da permebildade tanto em células Caco-2 quanto em ensaios PAMPA. O processo de validação experimental foi essencial para a confirmação da robustez dos modelos desenvolvidos, que serão incorporados à base de dados de propriedades farmacocinéticas PK/DB (http://www.pkdb.ifsc.usp.br/), contribuindo desta forma para o seu aprimoramento. Os métodos computacionais e experimentais utilizados neste projeto permitiram o desenvolvimento de ferramentas úteis para a avaliação da absorção de fármacos administrados por via oral e para o planejamento de moléculas bioativas com propriedades farmacocinéticas otimizadas. / Human intestinal absorption is a critical pharmacokinetics-related parameter for orally given drugs. In vitro models based on Caco-2 cells, which mimetize enterocytes, or those based on the parallel artificial membrane permeability assay (PAMPA) are broadly used in permeability studies of bioactive molecules. In drug discovery projects, these techniques are simultaneously used with computational methods to evaluate physico-chemical and pharmacokinetics properties. In this study, computational models to predict the cellular permeability of bioactive compounds in Caco-2 cells and in PAMPA assays were developed. The models were constructed by using the fragment-based method Hologram Quantitative Structure Activity Relationships (HQSAR). The obtained statistical indexes indicated the suitable correlation ability of the Caco-2 and PAMPA models for the employed data set, and the high predictive power for selected test set compounds. In addition, the HQSAR models were evaluated regarding their predictive ability against two validation sets, which are comprised by novel molecules that were designed and experimentally tested during this study. For these compounds, both the Caco-2 and PAMPA models demonstrated high predictive power. The process of experimental validation was an essential step for confirming the robustness of the developed models, which will be incorporated into the database of pharmacokinetic properties PK/DB (http://www.pkdb.ifsc.usp.br/), thus contributing to its improvement. The developed HQSAR models will be incorporated to the PK/DB pharmacokinetics database (http://www.pkdb.ifsc.usp.br/), thus contributing to its improvement. The molecular modeling and experimental methods employed in this study allowed for the development o robust tools for evaluating intestinal drug absorption and for the design of bioactive molecules with improved pharmacokinetics profiles.

Orientador: Prof. Dr. Marcos Roberto de Rocha Gesualdi / Tese (doutorado) - Universidade Federal do ABC, Programa de Pós-Graduação em Física, 2014. / Neste trabalho, realizando uma revisão teórica e experimental da geração de feixes não difrativos a partir dos fundamentos de teoria de difração, Hologramas Gerados por Computador (CGH) e Holograa, utilizando Moduladores Espaciais de Luz (SLM). Mostramos a possibilidade de geração de múltiplos feixes não difrativos de iguais ou diferentes simetrias utilizando a reconstrução holográca em SLM. Apresentamos de forma inédita a obtenção experimental de um novo conceito de feixe não difrativo que utiliza a superposição de feixes de Bessel cujo padrão longitudinal de intensidade pode ser escolhido a priori, a chamada Frozen Wave. Finalmente realizamos uma revisão teórica das propriedades de ondas em meios não lineares Fotorrefrativos, e a partir desse estudo desenvolvemos e montamos um arranjo de Holograa Fotorrefrativa e demonstramos experimentalmente a possibilidade de gerar feixes não difrativos utilizando essa técnica. Evidenciamos através dessa técnica uma nova possibilidade de geração de feixes não difrativos e em especial para geração de Frozen Waves. / In this work, we perform a theoretical and experimental review of genration non diractive beams since difraction theory, Computing Generation Hologram (CGH) and Holography using Spatial Light Modulator (SLM). We show the possibility to generate multiple non diractive beams with the igual or dierent symetry using a holographic reconstruction in SLM. We show for the rst time the experimental observation of a new nondifractive beam concepte that use Bessel beam superpositions whose intensite longitudinal shape can be chosen a priori, known as Frozen Wave. Finally, we made a theoretical review of the nonlinear Photorefractive properties, and of from this study we developed and mont a Photorefractive holography setup to show the nondiractive beam generation through this technique. Evidencing through this technique a new possibility to generate a non diractive beams and on special case to generate Frozen Waves.

La trame générale de ces travaux de recherche concerne la modélisation et l'implantation de systèmes d'interconnexions optiques à forte bande-passante spatiale et temporelle. Nous présenterons une suite de résultats au niveau des modèles, des applications et des réalisations prototypales. L'origine de ces travaux est une étude sur les connexions optiques en espace libre permettant de modéliser des opérations de calcul optique parallèle (telles que la multiplication matricielle), débouchant sur une classe d'opérations plus générale mettant en jeu un réseau d'interconnexions non-invariantes spatialement entre plans de données. Cette étape a permis d'évaluer, tant qualitativement que quantitativement, l'apport de la synergie entre la synthèse numérique d'éléments optiques diffractants (HSO) et les modulateurs spatiaux de lumière (MSL) à cristal liquide, dans le but de réaliser des dispositifs de traitement optique spatial dynamique de l'information. L'étape suivante traite des interconnexions dynamiques entre fibres optiques monomodes, débouchant naturellement sur la conception de dispositifs d'aiguillage et de commutation pour les réseaux de transmission optiques dorsaux multiplexés en longueur d'onde (WDM). A ce niveau, les contraintes principales portent sur les temps de commutation requis −inférieurs à la milliseconde− et les capacités de plusieurs dizaines de voies spatiales indépendantes. Au niveau de l'implantation prototypale, nous avons poussé le plus loin possible le concept d'holographie digitale dynamique sur MSL à cristal liquide, aboutissant à la conception et l'intégration de plusieurs générations d'aiguilleurs de bandes optiques performants (jusqu'à 64 voies spatiales). Par la suite, devant la nécessité de développer des règles d'ingénierie sur les anneaux et les réseaux maillés WDM, je me suis impliqué dans l'évaluation et le développement d'outils de simulation photonique concernant les dispositifs de commutation et de brassage transparent de canaux optiques. Je présente enfin les axes de recherche ayant émergé plus récemment, centrés sur le dimensionnement et l'intégration de réseaux locaux utilisant l'optique comme canal de propagation sans fil à très haut débit et à impact sanitaire et environnemental réduit.

A absorção intestinal de fármacos (HIA, na sigla em inglês para human intestinal absorption) é um dos processos farmacocinéticos mais críticos para a ação de medicamentos administrados por via oral. Modelos in vitro baseados em células que mimetizam os enterócitos do intestino humano (células Caco-2), ou aqueles que utilizam membranas articifiais são muito utilizados no estudo da permeabilidade celular de moléculas bioativas. No processo de planejamento de fármacos, estas técnicas são frequentemente empregadas em paralelo com estratégias computacionais na avaliação de propriedades físico-químicas e farmacocinéticas. Nessa tese de doutorado foram desenvolvidos modelos computacionais para a predição da permeabilidade celular de compostos bioativos em células Caco-2 e em membrana artificial paralela (PAMPA, na sigla em inglês para Parallel Artificial Membrane Permeability Assay). Os modelos foram gerados através do método baseado em fragmentos moleculares holograma QSAR (HQSAR). Os indicadores estatísticos obtidos indicam a boa capacidade de correlação dos modelos Caco-2 e PAMPA para os dados utilizados na modelagem e o alto poder de predição destes modelos para a permeabilidade de um conjunto teste de compostos. Adicionalmente, os modelos foram avaliados quanto a sua capacidade preditiva frente a dois conjuntos de validação, constituídos por novas moléculas que foram planejadas e testadas experimentalmente durante o desenvolvimento deste trabalho. Para estes conjuntos, os modelos demonstraram alta capacidade de predição da permebildade tanto em células Caco-2 quanto em ensaios PAMPA. O processo de validação experimental foi essencial para a confirmação da robustez dos modelos desenvolvidos, que serão incorporados à base de dados de propriedades farmacocinéticas PK/DB (http://www.pkdb.ifsc.usp.br/), contribuindo desta forma para o seu aprimoramento. Os métodos computacionais e experimentais utilizados neste projeto permitiram o desenvolvimento de ferramentas úteis para a avaliação da absorção de fármacos administrados por via oral e para o planejamento de moléculas bioativas com propriedades farmacocinéticas otimizadas. / Human intestinal absorption is a critical pharmacokinetics-related parameter for orally given drugs. In vitro models based on Caco-2 cells, which mimetize enterocytes, or those based on the parallel artificial membrane permeability assay (PAMPA) are broadly used in permeability studies of bioactive molecules. In drug discovery projects, these techniques are simultaneously used with computational methods to evaluate physico-chemical and pharmacokinetics properties. In this study, computational models to predict the cellular permeability of bioactive compounds in Caco-2 cells and in PAMPA assays were developed. The models were constructed by using the fragment-based method Hologram Quantitative Structure Activity Relationships (HQSAR). The obtained statistical indexes indicated the suitable correlation ability of the Caco-2 and PAMPA models for the employed data set, and the high predictive power for selected test set compounds. In addition, the HQSAR models were evaluated regarding their predictive ability against two validation sets, which are comprised by novel molecules that were designed and experimentally tested during this study. For these compounds, both the Caco-2 and PAMPA models demonstrated high predictive power. The process of experimental validation was an essential step for confirming the robustness of the developed models, which will be incorporated into the database of pharmacokinetic properties PK/DB (http://www.pkdb.ifsc.usp.br/), thus contributing to its improvement. The developed HQSAR models will be incorporated to the PK/DB pharmacokinetics database (http://www.pkdb.ifsc.usp.br/), thus contributing to its improvement. The molecular modeling and experimental methods employed in this study allowed for the development o robust tools for evaluating intestinal drug absorption and for the design of bioactive molecules with improved pharmacokinetics profiles.

Dois tipos diferentes de hologramas (Fresnel e Fourier) foram gerados por computador. O holograma de Fresnel foi escolhido para fazer parte de um arranjo experimental que teve como objetivo estimar a rugosidade de amostras metálicas. Para isso, um novo método de aplicação de um correlator óptico foi desenvolvido. Hologramas de Fourier geralmente fazem parte do correlator óptico de VanderLugt, o qual é muito utilizado no reconhecimento de padrões. A reconstrução numérica de hologramas de Fresnel gerados por computador, "distorcidos" por superfícies metálicas (também simuladas), serviram de base para que a reconstrução óptica de um holograma de Fresnel fosse utilizada, de forma inédita, no reconhecimento de padrões para estimar a rugosidade de amostras metálicas. / Two different types of holograms (Fresnel and Fourier) have been computer-generated. The Fresnel hologram has been chosen as part of an experimental set, which meant to estimate the roughness of the metalic samples. A new method for the aplication of an optical correlator has been developed. Fourier holograms are, generally, part of the VanderLugt optical correlator, that is very used for pattern recognition. The numerical reconstruction of the computer-generated Fresnel holograms, "distorted" by metalic surfaces (simulated as well), worked as the base for the utilization, in an original way, of the optical reconstruction of a Fresnel hologram, at the pattern recognition to estimate the roughness of the metalic samples.

The main objective of this thesis is the development of new processing techniques for digital holograms. The present work is part of the HoloFlow project that intends to integrate the DHM technology for the monitoring of water quality. Different tools for an automated analysis of digital holograms have been developed to detect, refocus and classify particles in continuous fluid flows. A detailed study of the refocusing criterion permits to determine its dependencies and to quantify its robustness. An automated detection procedure has been developed to determine automatically the 3D positions of organisms flowing in the experiment volume. Two detection techniques are proposed: a usual method based on a global threshold and a new robust and generic method based on propagation matrices, allowing to considerably increase the amount of detected organisms (up to 95 %) and the reliability of the detection. To handle the case of aggregates of particles commonly encountered when working with large concentrations, a new separation procedure, based on a complete analysis of the evolution of the focus planes, has been proposed. This method allows the separation aggregates up to an overlapping area of around 80 %. These processing tools have been used to classify organisms where the use of the full interferometric information of species enables high classifier performances to be reached (higher than 93 %). / Doctorat en Sciences de l'ingénieur / info:eu-repo/semantics/nonPublished

[ES] El tratamiento de enfermedades neurológicas está muy limitado por la ineficiente penetración de los fármacos en el tejido cerebral dañado debido a la barrera hematoencefálica (BHE), lo que imposibilita mejorar la salud del paciente. La BHE es un mecanismo de protección natural para evitar la difusión de agentes potencialmente peligrosas para el sistema nervioso central. No obstante, la BHE se puede inhibir mediante ultrasonidos focalizados e inyección de microburbujas de forma segura, localizada y transitoria, una tecnología empleada mundialmente. La principal ventaja es su carácter no invasivo, siendo así muy atractiva y cómoda para el paciente. Normalmente, la zona cerebral enferma se trata en su parte central empleando un único foco. Sin embargo, enfermedades como el Alzheimer o el Parkinson requieren un tratamiento sobre estructuras de geometría compleja y tamaño elevado, situadas en ambos hemisferios cerebrales. Por tanto, la tecnología actual está muy limitada al no cumplir dichos requisitos. Esta tesis doctoral tiene como objetivo el desarrollo de una técnica novedosa, basada en hologramas acústicos, para resolver las limitaciones presentes en los tratamientos neurológicos empleando ultrasonidos. Se estudian las lentes acústicas holográficas impresas en 3D, que acopladas a un transductor mono-elemento, permiten el control preciso del frente de onda ultrasónico tanto para (1) compensar las distorsiones que sufre el haz hasta alcanzar el cerebro, como (2) focalizarlo simultáneamente en regiones múltiples y de geometría compleja o formando de vórtices acústicos, proporcionando así efectividad en tiempo y coste. Por ello, la investigación desarrollada en esta tesis abre un camino prometedor en el campo de la biomedicina que permitirá mejorar los tratamientos neurológicos, además de aplicaciones en neuroestimulación o ablación térmica del tejido. / [CA] El tractament de malalties neurològiques està molt limitat per la ineficient penetració del fàrmac en el teixit cerebral danyat a causa de la barrera hematoencefàlica (BHE), i així no és possible una millora de salut del pacient. La BHE és un mecanisme de protecció natural per a evitar la difusió d'agents potencialment perillosos per al Sistema Nervios Central. No obstant això, aquesta barrera es pot inhibir mitjancant una tecnologia emprada mundialment basada en ultrasons focalitzats i injeccio de microbombolles. El principal avantatge és el seu caràcter no invasiu, sent així molt atractiva i còmoda per al pacient, i permet obrir la BHE de manera segura, localitzada i transitòria. Normalment, la zona cerebral malalta es tracta en la seua part central, emprant un unic focus. No obstant això, malalties com l'Alzheimer o el Parkinson requereixen un tractament al llarg d'estructures de geometria complexa i grandària elevada, situades en tots dos hemisferis cerebrals. Per tant, la tecnologia actual està fortament limitada al no complir amb aquests requeriments. Aquesta tesi doctoral està enfocada a investigar i desenvolupar una tècnica nova, basada en hologrames acústics, per a solucionar les limitacions presents en els tractaments neurològics. Una lent acústica holograca de baix cost impresa en 3D acoblada a un transductor d'element simple permet el control precs del front d'ona ultrasònic punt per a (1) compensar les distorsions que pateix el feix en el seu camí cap al cervell, i (2) focalització simultània del feix en regions multiples i de geometria complexa, proporcionant aix un tractament efectiu en temps i cost. Per això, la investigació desenvolupada en aquesta tesi demostra la possibilitat de realitzar qualsevol tractament neurològic, a més d'aplicacions en la neuroestimulació o l'ablació tèrmica dins del camp biomèdic. / [EN] Treatments for neurological diseases are strongly limited by the inefficient penetration of therapeutic drugs into the diseased brain due to the blood-brain barrier (BBB), and therefore no health improvement can be achieved. In fact, the BBB is a protection mechanism of the human body to avoid the diffusion of potentially dangerous agents into the central nervous system. Nevertheless, this barrier can be successfully inhibited by using a worldwide spread technology based on microbubble-enhanced focused ultrasound. Its main advantage is its non-invasive nature, thus defining a patient-friendly clinical procedure that allows to disrupt the BBB in a safe, local and transient manner. Conventionally, the diseased brain structure has been targeted in its center, with a single focus. However, Alzheimer's or Parkinson's Diseases do require that ultrasound is delivered to entire, complex-geometry and large-volume structures located at both hemispheres of the brain. Therefore, current technology presents several limitations as it does not fulfill these requirements. This doctoral thesis aims to develop a novel technique based on using focused ultrasound acoustic holograms to solve the existing limitations to treat neurological diseases. In this dissertation, we study 3D-printed holographic acoustic lenses coupled to a single-element transducer that allow to accurately control the acoustic wavefront to both (1) compensate distortions suffered by the beam in its path to the brain, and (2) simultaneous focusing in multiple and complex-geometry structures or acoustic vortex generation, providing a time- and cost- efficient procedure. Therefore, the research carried out throughout this thesis opens a promising path in the biomedical field to improve the treatment for neurological diseases, neurostimulation or tissue ablation applications. / Acknowledgments to the Spanish institution Generalitat Valenciana, which funding grant allowed me to develop this doctoral thesis, and as well funded my research stay at Columbia University. The development of the entire thesis was supported through grant Nª. ACIF/2017/045. Particularly, the research carried out in Chapter 3 and Chapter 4 was possible thanks to and supported through grant BEFPI/2019/075. Action co-financied by the Agència Valenciana de la Innovació through grant INNVAL10/19/016 and by the European Union through the Programa Operativo del Fondo Europeo de Desarrollo Regional (FEDER) of the Comunitat Valenciana 2014-2020 (IDIFEDER/2018/022). / Jiménez Gambín, S. (2021). Transcranial Ultrasound Holograms for the Blood-Brain Barrier Opening [Tesis doctoral]. Universitat Politècnica de València. https://doi.org/10.4995/Thesis/10251/171373 / TESIS