Spectroscopie résolue en temps par continuum femtoseconde. Applications en neurobiologie

International audienceLa spectroscopie résolue en temps utilisant un laser blanc femtoseconde est appliquée à la mesure in vivo des principaux absorbeurs du cerveau. Après génération adéquate du continuum de lumière blanche femtoseconde (50mW/[580-756nm] à 1 Hz), cette source se propage dans la calvaria, les méninges et le cortex chez le rat anesthésié. La transmission est étudiée sur 7mm de distance entre l impact laser et la fibre optique de collection. Le signal transmis est analysé dans la fenêtre 580-760nm, par un spectromètre couplé à une caméra à balayage de fente permettant la décorrélation de l absorption et de la diffusion

Spectroscopie résolue en temps par laser blanc femtoseconde pour l'exploration fonctionnelle in vivo du métabolisme énergétique cérébral

Les principaux chromophores endogènes d'intérêt sont les transporteurs chimiques d'hydrogène et les transporteurs chimiques d'oxygène/électron. Dans la fenêtre spectrale visible et proche infrarouge, bande spectrale de forte pénétration des tissus par la lumière, les premiers transporteurs sont accessibles avec la spectroscopie de fluorescence et les seconds avec la spectroscopie d'absorption des tissus. L'objectif de cette thèse est de réaliser la mesure non invasive in vivo des indices métaboliques par spectroscopie d'absorption des molécules d'oxy- et de déoxy- hémoglobine. Les travaux se sont orientés vers le développement d'une instrumentation ultra-brève novatrice en biophotonique et la construction de modèles mathématiques des tissus sondés. L'approche expérimentale couple la transillumination par laser blanc femtoseconde kilohertz avec une camera à balayage de fente monocoup en mode comptage de photo-électron unique. Ceci autorise l'analyse des coefficients d'absorption et de diffusion sur une large fenêtre spectrale (180 nm), sans balayage. Le spectroscope en milieu diffus est validé in vitro dans des solutions lactés additionnées de différents colorants et in vivo chez le rat de laboratoire et l'oiseau chanteur diamant mandarin. Les mesures montrent la capacité du dispositif à réaliser le suivi temporel des indices métaboliques lors d'évènements biologiques comme l'hypercapnie en normoxie. Afin de relier les coefficients optiques aux paramètres physiologiques pertinents, l'étude de la distribution tri-dimensionnelle des absorbeurs des tissus est nécessaire. La modélisation de l'architecture vasculaire corticale (compartiment contenant les hémoglobines) est réalisée chez le rat. Les premiers résultats montrent la nécessité de tenir compte de cette organisation " space-filling " lors de l'interprétation des coefficients optiques, notamment par le biais de la théorie mathématique de l'homogénéisationThe main tissue endogen chromophores are the chemical carriers of hydrogen and oxygen/electron. In the visible and infrared spectral window, where the light is strongly penetrating, the first carriers are accessible with fluorescence spectroscopy and the second with absorption spectroscopy. The purpose is the realization of the in vivo non invasive measurement of metabolism indexes by absorption spectroscopy of oxy- and deoxy- hemoglobin. The works are based on the development of an new ultrashort device in biophotonic and the construction of a mathematical representation of probed tissues. The experimental approach couples a kilohertz white laser transillumination with a single shot streak camera in single photo-electron counting mode. It allows the analyze of the absorption coefficient and reduced scattering coefficient on a broad spectral window (180 nm), without scanning. The spectroscope for scattering media is validated in vitro into dye added milk solutions and validated in vivo into the laboratory rat and into the Zebra finch songbird. The measurements show the ability of the apparatus to realize temporal follow of metabolism indexes during biological events as normoxic hypercapnia. In order to link optical coefficients to relevant physiological parameters, the study of the three dimension distribution of tissue absorbers is necessary. The cortical angioarchitecture is realized for the rat. The first results show the necessity to take into account this "space-filling" organization for optical coefficients interpretation, notably by means of mathematical theory of homogenizationST ETIENNE-BU Sciences (422182103) / SudocSudocFranceF

Optical Systems in Ultrafast Biophotonics

International audienceIn the field of biophotonics the main goals are the control and processing of in vivo biological tissues and the monitoring of biomolecule dynamics. Two particular pitfalls are present: the dynamic multiscale organization and the photostress of the medium. Until now the state of the art of the pico-femtosecond systems designed to these applications shows that the changing laser technology has been only used as an add-on. Our approach is based on a bottom-up procedure and on the medium-centered knowledge. The range of neurobiological applications of ultrafast photonics extends from TRP (time-resolved propagation) to linear and non-linear TRE (time-resolved emission). The device combines a one kilohertz chirp pulse amplification laser system and a single shot streak camera. For discrete wavelength applications (TRE), the set-up is a SHG/OPG/OPA3/SHG design. In the case of TRP, the beam is focused into pure water to generate a white light continuum. After propagation through tissue, a single-shot streak camera with single photo-electron counting capability performs the picosecond time-resolved spectroscopy of the collected photons. Depending on the acceptable level of photostress, the integration time can extend from 33ms up to several minutes with a real-time control of the jitter and time drifts. The meaning of the TRE spectro-temporal image is particularly detailed in the 450-480nm excitation window in regards to the contributions of mitochondrial flavoproteins. This optical system fulfills the reliability and the sensitivity, conditions required for measuring opto-electronic quantities from freely moving animal at low irradiation

<title><emph type="1">In vivo </emph>brain spectroscopy with femtosecond white light continuum</title>

International audienceOur purpose is to spectrally probe the main brain absorbers. The determination of their spatial distribution remains a challenge. According to anatomical data, the proposed 3D model of the rat pial-cortical vascular networks is divided into three parts: (1) the pial vessels could be approximated by a dense layer of around 250 micrometers depth; (2) the penetrating vessels repartition is described as periodic hexagonal prisms with three modules; (3) the capillary network is modelized using a periodic tiling of polyhedron with a density of 817mm.mm-3 and a branching pattern of 10000mm-3. With anaesthetized rats under stereotaxic conditions, in vivo time-resolved brain spectroscopy experiments are presented. The setup is designed to allow broadband time-resolved spectroscopy using a streak camera. A femtosecond white light continuum is produced by focusing 800nm pulses (0.5mJ, 1kHz, 150fs) in an adapted third order non linear medium. In the case of water, the spectrum expands over 380-780nm with an efficiency of 20 percent. Mathematical homogenization techniques could be applied to the radiative transfer equation with this geometrical vascular architecture and might be useful to analyze in depth time-resolved spectroscopy of such complex media

In vivo and noninvasive measurement of a songbird head's optical properties.

International audienceBy assessing the cerebral blood volume and the hemoglobin oxygen saturation level, near-infrared spectroscopy (NIRS) probes brain oxygenation, which reflects cerebral activity. To develop a noninvasive method monitoring the brain of a songbird, we use an original NIRS device, i.e., a white laser coupled with an ultrafast spectrotemporal detector of optical signals without wavelength scanning. We perform in vivo measurements of the absorption coefficient and the reduced scattering coefficient of the caudal nidopallium area of the head of a songbird (the zebra finch)

Functional white-laser imaging to study brain oxygen uncoupling/recoupling in songbirds

Contrary to the intense debate about brain oxygen dynamics and its uncoupling in mammals, very little is known in birds. In zebra finches, picosecond optical tomography with a white laser and a streak camera can measure in vivo oxyhemoglobin (HbO2) and deoxyhemoglobin (Hb) concentration changes following physiologic stimulation (familiar calls and songs). Picosecond optical tomography showed sufficient submicromolar sensitivity to resolve the fast changes in the hippocampus and auditory forebrain areas with 250 μm resolution. The time course is composed of (1) an early 2-second-long event with a significant decrease in Hb and HbO2 levels of −0.7 and −0.9 μmol/L, respectively, (2) a subsequent increase in blood oxygen availability with a plateau of HbO2 (+0.3 μmol/L), and (3) pronounced vasodilatation events immediately after the end of the stimulus. One of the findings of our study is the direct link between blood oxygen level-dependent signals previously published in birds and our results. Furthermore, the early vasoconstriction event and poststimulus ringing seem to be more pronounced in birds than in mammals. These results in birds, tachymetabolic vertebrates with a long lifespan, can potentially yield new insights, e.g., into brain aging

Diffuse optical tomography with an amplified ultrafast laser and a single-shot streak camera: application to real-time in vivo songbird neuro-imaging

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Functional white-laser imaging to study brain oxygen uncoupling/recoupling in songbirds

International audienceThis corrects the article "Functional white-laser imaging to study brain oxygen uncoupling/recoupling in songbirds" in volume 31 on page 393.Following the publication of this article, the authors noted the following errors:(1) Two authors were omitted from the author list. The correct and complete author names appear above. The affiliation for Drs Vignal and Mathevon is the Université de St-Etienne, CNRS, ENES/CNPS UMR8195, Saint-Etienne, France.(2) The grants should have included the following note: ‘These experiments were supported by the Program ‘Emergence' of the Région Rhône-Alpes and the Agence Nationale de la Recherche (Project ‘Birds' voices, ANR-06-BLAN-0293-01)'