Samuel Bélanger

Real-time diffuse optical tomography based on structured illumination

A new optical acquisition scheme based on a pair of digital micromirror devices is developed and applied to three-dimensional tomographic imaging of turbid media. By using pairs of illumination-detection patterns with a single detector, we were able to perform high-resolution quantitative volumetric imaging of absorption heterogeneities embedded in optically thick samples. Additionally, a tomographic reconstruction algorithm was implemented on a graphical processor unit to provide optical reconstructions at a frame rate of 2 Hz. The structured illumination method proposed in this work has significant cost advantages over camera systems, as only a single detector is required. This configuration also has the potential to increase frame rate.

Investigating the correlation between white matter and microvasculature changes in aging using large scale optical coherence tomography and confocal fluorescence imaging combined with tissue sectioning

Here, we present a serial OCT/confocal scanner for histological study of the mouse brain. Three axis linear stages combined with a sectioning vibratome allows to cut thru the entire biological tissue and to image every section at a microscopic resolution. After acquisition, each OCT volume and confocal image is re-stitched with adjacent acquisitions to obtain a reconstructed, digital volume of the imaged tissue. This imaging platform was used to investigate correlations between white matter and microvasculature changes in aging mice. Three age groups were used in this study (4, 12, 24 months). At sacrifice, mice were transcardially perfused with a FITC containing gel. The dual imaging capability of the system allowed to reveal different contrast information: OCT imaging reveals changes in refractive indices giving contrast between white and grey matter in the mouse brain, while transcardial perfusion of a FITC shows microsvasculature in the brain with confocal imaging.

Effects of anesthesia on the cerebral capillary blood flow in young and old mice

Despite recent findings on the possible role of age-related cerebral microvasculature changes in cognition decline, previous studies of capillary blood flow in aging (using animal models) are scarce and limited to anesthetized conditions. Since anesthesia can have different effects in young and old animals, it may introduce a confounding effect in aging studies. The present study aimed to eliminate the potential confound introduced by anesthesia by measuring capillary blood flow parameters in both awake conditions and under isoflurane anesthesia. We used 2-photon laser scanning fluorescence microscopy to measure capillary diameter, red blood cell velocity and flux, hematocrit and capillary volumetric flow in individual capillaries in the barrel cortex of 6- and 24-month old C57Bl/6 mice. It was observed that microvascular properties are significantly affected by anesthesia leading to different trends in capillary blood flow parameters with aging when measured under awake or anesthetized conditions. The findings in this study suggest taking extra care in interpreting aging studies from anesthetized animals.

Astrocytic endfoot Ca2+ correlates with parenchymal vessel responses during 4-AP induced epilepsy: An in vivo two-photon lifetime microscopy study:

Neurovascular coupling (NVC) underlying the local increase in blood flow during neural activity forms the basis of functional brain imaging and is altered in epilepsy. Because astrocytic calcium (Ca2+) signaling is involved in NVC, this study investigates the role of this pathway in epilepsy. Here, we exploit 4-AP induced epileptic events to show that absolute Ca2+ concentration in cortical astrocyte endfeet in vivo correlates with the diameter of precapillary arterioles during neural activity. We simultaneously monitored free Ca2+ concentration in astrocytic endfeet with the Ca2+-sensitive indicator OGB-1 and diameter of adjacent arterioles in the somatosensory cortex of adult mice by two-photon fluorescence lifetime measurements following 4-AP injection. Our results reveal that, regardless of the mechanism by which astrocytic endfoot Ca2+ was elevated during epileptic events, increases in Ca2+ associated with vasodilation for each individual ictal event in the focus. In the remote area, increases in Ca2+ correlated with vasoconstriction at the onset of seizure and vasodilation during the later part of the seizure. Furthermore, a slow increase in absolute Ca2+ with time following multiple seizures was observed, which in turn, correlated with a trend of arteriolar constriction both at the epileptic focus and remote areas.

Correlation of hemodynamic and fluorescence signals under resting state conditions in mice's barrel field cortex

Both neurons and astrocytes are known to affect local vascular response in the brain following neuronal activity. In order to differentiate the contributions of each cell type to the hemodynamic response during stimulation and resting state, intrinsic optical signal (IOI) was recorded synchronized with fluorescence imaging of calcium concentration sensitive dye Oregon Green BAPTA-1 AM. By changing the stimulation parameters (frequency and duration), it was possible to individually promote neuronal and glial responses and to compare them to levels of oxy (HbO), deoxy (HbR) and total (HbT) hemoglobin concentrations. Finally, resting state recordings were done to investigate the possible correlation between hemoglobin fluctuation and calcium transients, based on different frequency bands associated either with neuronal or glial activity.

Comparison of the performance of two depth-resolved optical imaging systems: laminar optical tomography and spatially modulated imaging

The objective of this work is to compare quantitatively the imaging capabilities of a laminar optical tomography (LOT) system with those of a spatially modulated imaging (SMI) system. LOT is a three dimensional optical imaging technique that achieves depth sensitivity by measuring multiple-scattered light at different source-detector separations. The SMI method is based on spatially modulated illumination-detection patterns, which encode both optical properties and depth information. In this work, simulation studies are carried out at different noise levels, to obtain the figures of merit of tomographic reconstructions for both systems. Experiments on phantoms are performed to demonstrate the validity of the numerical results.

Compromised microvascular oxygen delivery increases brain tissue vulnerability with age

Despite the possible role of impaired cerebral tissue oxygenation in age-related cognition decline, much is still unknown about the changes in brain tissue pO2 with age. Using a detailed investigation of the age-related changes in cerebral tissue oxygenation in the barrel cortex of healthy, awake aged mice, we demonstrate decreased arteriolar and tissue pO2 with age. These changes are exacerbated after middle-age. We further uncovered evidence of the presence of hypoxic micro-pockets in the cortex of awake old mice. Our data suggests that from young to middle-age, a well-regulated capillary oxygen supply maintains the oxygen availability in cerebral tissue, despite decreased tissue pO2 next to arterioles. After middle-age, due to decreased hematocrit, reduced capillary density and higher capillary transit time heterogeneity, the capillary network fails to compensate for larger decreases in arterial pO2. The substantial decrease in brain tissue pO2, and the presence of hypoxic micro-pockets after middle-age are of significant importance, as these factors may be related to cognitive decline in elderly people.

In vivo measurement of astrocytic endfoot Ca2+ and parenchymal vessel responses during 4-AP induced epilepsy using two-photon fluorescence lifetime microscopy

Neurovascular coupling (NVC) is defined as a local increase in cerebral blood flow in response to neuronal activity, it forms the basis of functional brain imaging and is altered during epilepsy. Because astrocytic calcium signaling (Ca2+) has been involved in the response of parenchymal vessels, this study investigates the role of this pathway during epilepsy. We exploit 4-Aminopyridine (4-AP) induced epileptic seizures to show that absolute Ca2+ concentration in astrocytic endfeet correlates with the changes in diameter of parenchymal vessels during neural activity in vivo. A two-photon laser scanning fluorescence lifetime microscopy was developed to simultaneously monitor free Ca2+ concentration in astrocytic endfeet with the calcium-sensitive indicator Oregon Green 488 BAPTA-1 (OGB-1) and the diameter of parenchymal vessels in the somatosensory cortex of mice following 4-AP injection. Our results reveal that the resting Ca2+ concentration in glial cells was spatially heterogeneous and that resting Ca2+ concentration in somatic regions was significantly higher than in endfoot regions. Moreover, following 4-AP injection in the somatosensory cortex of mice, we observed increases of Ca2+ in astrocytic endfeet associated with vasodilation of parenchymal vessels for each individual ictal event in the epileptic focus. However, vasodilation was seen to be inhibited by increase in absolute resting Ca2+ concentration. Our results suggest a role for baseline astrocytic Ca2+ concentration in vasodilation.

Retinal Vessel Segmentation from a Hyperspectral Camera Images

In this paper, a vessel segmentation method from hyperspectral retinal images based on the Multi-Scale Line Detection algorithm is proposed. The method consists in combining segmentation information from several consecutive images obtained at specific wavelengths around the green channel to produce an accurate segmentation of the retinal vessel network. Images obtained from six subjects were used to evaluate the performance of the proposed method. Preliminary results suggest a potential advantage of combining multispectral information instead of using only the green channel in segmenting retinal blood vessels.

Application of compressed sensing to optical tomography

A concrete experimental application of compressed sensing is described. It uses a pair of digital micromirrors to perform diffuse optical tomography. Specific constraints arise due to the requirement of rapid imaging, which entails using of binary illumination patterns. Benefits and limitations of compressed sensing towards reaching these goals are discussed with preliminary results.