Simultaneous morphological and functional imaging of the honeybee’s brain by two-photon microscopy

Thanks to its rather simply structured but highly performing brain, the honeybee (Apis mellifera) is an important model for neurobiological studies. Therefore there is a great need for new functional imaging modalities adapted to this species. Herein we give a detailed report on the development and performance of a platform for in vivo functional and morphological imaging of the honeybee’s brain, focusing on its primary olfactory centres, the antennal lobes (ALs). The experimental setup consists of a two-photon microscope combined with a synchronized odour stimulus generator. Our imaging platform allows to simultaneously obtain both morphological measurements of the ALs functional units, the glomeruli, and in vivo calcium recording of their neural activity. We were able to record the characteristic glomerular response maps to odour stimuli applied to the bee’s antennae. Our approach offers several advantages over the commonly used conventional fluorescence microscopy. Two-photon microscopy provides substantial enhancement in both spatial and temporal resolutions, while minimizing photo damage. Calcium recordings show a more than fourfold improvement in the functional signal with respect to the techniques available up to now. Finally, the extended penetration depth, thanks to the infrared excitation, allows the functional imaging of profound glomeruli which have not been optically accessible up to now

Detecting magnetically guided atoms with an optical cavity

We show that a low finesse cavity can be efficient for detecting neutral atoms. The low finesse can be compensated for by decreasing the mode waist of the cavity. We have used a near concentric resonator with a beam waist of 12$\mu$m and a finesse of only 1100 to detect magnetically guided Rb atoms with a detection sensitivity of 0.1 atom in the mode volume. For future experiments on single atom detection and cavity QED applications, it should be very beneficial to use miniaturized optical resonator integrated on atom chips.Comment: To appear in Optics Letter

Single atom detection in low finesse cavities

Im Rahmen dieser Doktorarbeit wurde der Grundstein gelegt für die Integration mikrooptischer Elements auf Atomchips. Dies beinhaltete theoretische und experimentelle Untersuchungen zur Möglichkeit der Detektion einzelner Atome mit Hilfe von optischen Resonatoren niedriger Güte. Hierfür wurde ein theoretisches Model zur Beschreibung des gekoppelten Systems Atom-Resonator entwickelt. Angewandt auf einen optischen Faserresonator auf dem Atomchip, sagt dieses Model das Erreichen eines Signal-zu-Rausch-Verhältnisses von über 30 für die Einzelatomdetektion voraus, für Meßintervalle von 10µs. Des weiteren wurde ein Experiment mit kalten Rubidium Atomen aufgebaut, das den ersten Atomchip mit integriertem Faserresonator aufnehmen wird. Eine Serie von Experimenten mit einem makroskopischen Testresonator, der in seinen Eigenschaften einem Faserresonantor gleicht, wurde durchgeführt. Indem man den Resonator sehr nah am konzentrischen Punkt betreibt, kann dessen Fokusbreite auf 12µm reduziert werden, bei einer Finesse von 1200. Diese Parameter liegen in der gleichen Größenordnung wie die eines Faserresonators. Experimente mit Atomen, die frei durch den Resonator fallen oder mit Hilfe eines magnetischen Leiters hindurchgeführt werden, bestätigten die Voraussagen der Theorie bezüglich des Einflusses der Atome auf das Resonatortransmissionssignal

In-vivo two-photon imaging of the honey bee antennal lobe

Due to the honey bee's importance as a simple neural model, there is a great need for new functional imaging modalities. Herein we report on the use of two-photon microscopy for in-vivo functional and morphological imaging of the honey bee's olfactory system focusing on its primary centers, the antennal lobes (ALs). Our imaging platform allows for simultaneously obtaining both morphological measurements of the AL and in-vivo calcium recording of neural activities. By applying external odor stimuli to the bee's antennas, we were able to record the characteristic odor response maps. Compared to previous works where conventional fluorescence microscopy is used, our approach offers all the typical advantages of multi-photon imaging, providing substantial enhancement in both spatial and temporal resolutions while minimizing photo-damages and autofluorescence contribution with a four-fold improvement in the functional signal. Moreover, the multi-photon associated extended penetration depth allows for functional imaging within profound glomeruli.Comment: 3 pages, 3 figure

Dependence of sub-micron vaterite container release properties on pH and ionic strength of the surrounding solution

We report on the synthesis and characterization of porous monodisperse vaterite containers with controllable average sizes from 400 nm to 10 mu m. Possible release strategies of enclosed substances via recrystallization or by pH-change are presented. As a model experiment, a fluorescent marker was encapsulated and imaged by two-photon microscopy to monitor the dye release. The release process was found to be controllable via the immersion medium's properties. Release times can be further tuned by covering the containers with additional polymer layers, creating a flexible system with promising perspectives for pharmaceutical applications

A tunable narrowband entangled photon pair source for resonant single-photon single-atom interaction

We present a tunable, frequency-stabilized, narrow-bandwidth source of frequency-degenerate, entangled photon pairs. The source is based on spontaneous parametric downconversion (SPDC) in periodically-poled KTiOPO4 (PPKTP). Its wavelength can be stabilized to 850 or 854 nm, thus allowing to address two D-P transitions in 40Ca+ ions. Its output bandwidth of 22 MHz coincides with the absorption bandwidth of the calcium ions. Its spectral power density is 1.0 generated pairs/(s MHz mW).Comment: 3 pages, 3 figure

Side differences during odour processing in the honey bee brain

Trabalho de projecto de mestrado em Medicina (Pneumologia), apresentado à Faculdade de Medicina da Universidade de CoimbraA pneumonia adquirida na comunidade (PAC) é uma doença que revela morbilidade e mortalidade significativas. Os doentes mais graves necessitam de hospitalização e por vezes reúnem critérios de gravidade que condicionam o seu internamento na unidade de cuidados intensivos (UCI), constituindo uma das entidades mais frequentes nestes serviços. Apesar de controverso, o termo “severa” é frequentemente atribuído à doença que requer internamento na UCI, constituindo a definição mais comum. Pelos recursos exigidos com o internamento quer ao nível da enfermaria, quer na UCI, reconhece-se como uma entidade com impacto socioeconómico significativo, daí a importância de escalonar correctamente os doentes que reúnem critérios de severidade e que necessitam de internamento. Objectivo do trabalho Propõe-se a realização de uma revisão da literatura que permita elucidar à luz dos conhecimentos actuais diversos aspectos relativos a esta entidade, designadamente clarificar a sua definição, caracterizando aspectos particulares da sua prevalência e história natural, e congregar diferentes directivas já existentes visando definir claramente o grau de severidade, o que determinará um melhor prognóstico e rentabilização de recursos pela instituição precoce da terapêutica adequada. Pretende-se ainda um esclarecimento quanto à identificação sistematizada dos doentes que reúnem critérios para admissão em UCI bem como quanto às implicações em termos de prognóstico resultante da condição de severidade. Desenvolvimento Para definir a PAC severa (PACS) há que atender a parâmetros que incluem a fisiopatogenia, os factores risco/co-morbilidades associados e o processo diagnóstico em termos clínicos, PAC Severa: Definição, Particularidades e Factores Preditivos para Admissão em Unidade de Cuidados Intensivos R Ferreira, S Freitas 3 laboratoriais (diagnóstico etiológico) e imagiológicos. Desenvolveram-se vários scores/índices de gravidade para escalonar apropriadamente os doentes que requerem internamento. A sua aplicação estratifica a severidade e prediz o prognóstico. Vários factores estão implicados no prognóstico, nomeadamente a maior duração do internamento ou um timing de admissão mais tardio. Avaliou-se igualmente a utilidade prognóstica de biomarcadores como a pró-calcitonina, proteína C reactiva, algumas interleucinas e o cortisol plasmático, bem como o status socioeconómico, variabilidade genética, toma de anti-inflamatórios não esteróides, ventilação mecânica e o seguimento das guidelines. Conclusões A definição mais consensual de PACS continua a ser a que implica internamento em UCI, apresentando-se o Streptococcus pneumoniae como o agente etiológico mais frequente. Dos diversos scores de gravidade disponíveis, as directivas IDSA/ATS 2007 revelaram-se como as mais equilibradas, pelo que a sua aplicação se traduziu num impacto prognóstico positivo significativo. Em termos futuros, prevê-se que o desenvolvimento da investigação na área dos biomarcadores inflamatórios poderá incrementar a acuidade diagnóstica e prognóstica nesta patologiaThe community-acquired pneumonia (CAP) is a disease responsible for significant morbidity and mortality. The most critical patients require hospitalization and sometimes meet severity criteria that determine their admission in the intensive care unit (ICU), being one of the most common situations in these services. Although controversial, the term "severe" is often attributed to the disease that requires admission to ICU, being the most common definition. Given the amount of resources it requires, not only in the ward but also in the ICU, the CAP is recognized as an entity with significant economic impact, hence the importance of properly scaling the patients who meet the criteria for severe CAP and require hospitalization. This article seeks to review the literature to clarify the current knowledge over several aspects of this entity, including its definition, its prevalence and natural history, and to sum up various existing directives in order to clearly define the degree of severity, which determines the prognosis and allows a faster appropriate response as well as a better resources management. This review also aims at establishing a method of identifying the patients that meet the ICU admission criteria and at assessing the prognosis implications of their severity level. To define severe CAP (SCAP) one should consider parameters that include the pathophysiology, risk factors/comorbidities and associated diagnostic elements such as clinical, laboratory (etiology) and imaging findings. Several severity scores were developed to scale appropriately patients requiring hospitalization. Their application stratifies the severity and predicts prognosis. Several factors are implicated in worse prognosis, including length of stay in ICU and delayed admission. The prognostic value of biomarkers such as procalcitonin, C-reactive protein, plasma cortisol and some interleukins, as well as socioeconomic status, genetic variability, anti-inflammatory drugs intake, mechanical ventilation and monitoring of guidelines was also assessed. In conclusion, the most commonly agreed definition of SCAP continues to be a CAP that requires hospitalization in ICU, being the Streptococcus pneumoniae the most frequent etiologic agent. Among the various severity scores available, the directives IDSA / ATS 2007 turned out to be the most balanced and its application has resulted in a significant positive impact on prognosis. It is expected that future research on inflammatory biomarkers may improve the diagnostic accuracy and prognosis in this diseas

Evolutionary compromises in ecological adaptation: urea and ammonia tolerance in Drosophila suzukii and Drosophila melanogaster

The invasive species Drosophila suzukii has evolved morphological and behavioral adaptations to lay eggs under the skin of fresh fruits. This results in severe damage of a wide range of small fruits, making this species a serious agricultural and economical threat. Drosophila suzukii females typically lay few eggs per fruit, preferring not infested fruits. Hence, larvae are exposed to a reduced amount of nitrogenous waste. Differently, the innocuous Drosophila melanogaster lays eggs on fermented fruits already infested by conspecifics, with larvae developing in a crowded environment with accumulation of nitrogenous waste such as ammonia and urea. The observed differences in oviposition site and larval ecological niche suggest that these species might differ in behavioral and physiological mechanisms used to cope with nitrogenous waste. We investigated how different concentrations of ammonia and urea affect fecundity and larval development in both species. Females and larvae of D. suzukii showed greater sensitivity to high concentration of both compounds, with a dramatic decrease in fecundity and egg viability. To better understand the pathways underlying these differences, we evaluated the effect on ornithine aminotransferase and glutathione-S-transferase, two enzymes involved in nitrogen metabolism and stress response that are expressed during larval development. Both ammonia and urea significantly reduced the expression of these enzymes in D. suzukii compared to D. melanogaster. This manifests how the ecological shift of D. suzukii to fresh fruit resulted in less efficient detoxifying and excretory mechanisms, with important implications for evolutionary biology and applied research