Dene Ringuette

Rapid monitoring of cerebral ischemia dynamics using laser-based optical imaging of blood oxygenation and flow

Abstract: Imaging blood flow or oxygenation changes using optical techniques is useful for monitoring cortical activity in healthy subjects as well as in diseased states such as stroke or epilepsy. However, in order to gain a better understanding of hemodynamics in conscious, freely moving animals, these techniques must be implemented in a small scale, portable design that is adaptable to a wearable format. We demonstrate a novel system which combines the two techniques of laser speckle contrast imaging and intrinsic optical signal imaging simultaneously, using compact laser sources, to monitor induced cortical ischemia in a full field format with high temporal acquisition rates. We further demonstrate the advantages of using combined measurements of speckle contrast and oxygenation to establish absolute flow velocities, as well as to statistically distinguish between veins and arteries. We accomplish this system using coherence reduction techniques applied to Vertical Cavity Surface Emitting Lasers (VCSELs) operating at 680, 795 and 850 nm. This system uses minimal optical components and can easily be adapted into a portable format for continuous monitoring of cortical hemodynamics.

Multi-modality optical neural imaging using coherence control of VCSELs

Neural optical imaging can evaluate cortical hemodynamic fluctuations which reflect neural activity and disease state. We evaluate the use of vertical-cavity surface-emitting lasers (VCSELs) as illumination source for simultaneous imaging of blood flow and tissue oxygenation dynamics ex vivo and in vivo and demonstrate optical imaging of blood flow changes and oxygenation changes in response to induced ischemia. Using VCSELs we show a rapid switching from a single-mode to a special multi-mode rapid current sweep operation and noise values reduced to within a factor of 40% compared to non-coherent LED illumination. These VCSELs are promising for long-term portable continuous monitoring of brain dynamics in freely moving animals.

Evaluation of laser speckle contrast imaging as an intrinsic method to monitor blood brain barrier integrity

The integrity of the blood brain barrier (BBB) can contribute to the development of many brain disorders. We evaluate laser speckle contrast imaging (LSCI) as an intrinsic modality for monitoring BBB disruptions through simultaneous fluorescence and LSCI with vertical cavity surface emitting lasers (VCSELs). We demonstrated that drug-induced BBB opening was associated with a relative change of the arterial and venous blood velocities. Cross-sectional flow velocity ratio (veins/arteries) decreased significantly in rats treated with BBB-opening drugs, ≤0.81 of initial values.

Reducing misfocus-related motion artefacts in laser speckle contrast imaging

Laser Speckle Contrast Imaging (LSCI) is a flexible, easy-to-implement technique for measuring blood flow speeds in-vivo. In order to obtain reliable quantitative data from LSCI the object must remain in the focal plane of the imaging system for the duration of the measurement session. However, since LSCI suffers from inherent frame-to-frame noise, it often requires a moving average filter to produce quantitative results. This frame-to-frame noise also makes the implementation of rapid autofocus system challenging. In this work, we demonstrate an autofocus method and system based on a novel measure of misfocus which serves as an accurate and noise-robust feedback mechanism. This measure of misfocus is shown to enable the localization of best focus with sub-depth-of-field sensitivity, yielding more accurate estimates of blood flow speeds and blood vessel diameters.

Imaging brain activity during seizures in freely behaving rats using a miniature multi-modal imaging system

We report on a miniature label-free imaging system for monitoring brain blood flow and blood oxygenation changes in awake, freely behaving rats. The device, weighing 15 grams, enables imaging in a ∼ 2 × 2 mm field of view with 4.4 μm lateral resolution and 1 - 8 Hz temporal sampling rate. The imaging is performed through a chronically-implanted cranial window that remains optically clear between 2 to > 6 weeks after the craniotomy. This imaging method is well suited for longitudinal studies of chronic models of brain diseases and disorders. In this work, it is applied to monitoring neurovascular coupling during drug-induced absence-like seizures 6 weeks following the craniotomy.

Continuous multi-modality brain imaging reveals modified neurovascular seizure response after intervention

We developed a multi-modal brain imaging system to investigate the relationship between blood flow, blood oxygenation/volume, intracellular calcium and electrographic activity during acute seizure-like events (SLEs), both before and after pharmacological intervention. Rising blood volume was highly specific to SLE-onset whereas blood flow was more correlated with all eletrographic activity. Intracellular calcium spiked between SLEs and at SLE-onset with oscillation during SLEs. Modified neurovascular and ionic SLE responses were observed after intervention and the interval between SLEs became shorter and more inconsistent. Comparison of artery and vein pulsatile flow suggest proximal interference and greater vascular leakage prior to intervention.

Multi-modal in vivo imaging of brain blood oxygenation, blood flow and neural calcium dynamics during acute seizures

Dysfunction of the vascular endothelium has been implicated in the development of epilepsy. To better understand the relation between vascular function and seizure and provide a foundation for interpreting results from functional imaging in chronic disease models, we investigate the relationship between intracellular calcium dynamics and local cerebral blood flow and blood oxygen saturation during acute seizure-like events and pharmacological seizure rescue. To probe the relation between the aforementioned physiological markers in an acute model of epilepsy in rats, we integrated three different optical modalities together with electrophysiological recordings: Laser speckle contrast imaging (LSCI) was used to study changes in flow speeds, Intrinsic optical signal imaging (IOSI) was used to monitor changes in oxygenated, de-oxygenated, and total hemoglobin concentration, and Calcium-sensitive dye imaging was used to monitor intracellular calcium dynamics. We designed a dedicated cortical flow chamber to remove superficial blood and dye resulting from the injection procedure, which reduced spurious artifacts. The near infrared light used for IOSI and LSCI was delivered via a light pipe integrated with the flow chamber to minimize the effect of fluid surface movement on illumination stability. Calcium-sensitive dye was injected via a glass electrode used for recording the local field potential. Our system allowed us to observe and correlate increases in intracellular calcium, blood flow and blood volume during seizure-like events and provide a quantitative analysis of neurovascular coupling changes associated with seizure rescue via injection of an anti-convulsive agent.

Chronic monitoring of cortical hemodynamics in behaving, freely-moving rats using a miniaturized head-mounted optical microscope

Growing interest within the neurophysiology community in assessing healthy and pathological brain activity in animals that are awake and freely-behaving has triggered the need for optical systems that are suitable for such longitudinal studies. In this work we report label-free multi-modal imaging of cortical hemodynamics in the somatosensory cortex of awake, freely-behaving rats, using a novel head-mounted miniature optical microscope. The microscope employs vertical cavity surface emitting lasers (VCSELs) at three distinct wavelengths (680 nm, 795 nm, and 850 nm) to provide measurements of four hemodynamic markers: blood flow speeds, HbO, HbR, and total Hb concentration, across a > 2 mm field of view. Blood flow speeds are extracted using Laser Speckle Contrast Imaging (LSCI), while oxygenation measurements are performed using Intrinsic Optical Signal Imaging (IOSI). Longitudinal measurements on the same animal are made possible over the course of > 6 weeks using a chronic window that is surgically implanted into the skull. We use the device to examine changes in blood flow and blood oxygenation in superficial cortical blood vessels and tissue in response to drug-induced absence-like seizures, correlating motor behavior with changes in blood flow and blood oxygenation in the brain.

Multi-modality Optical Imaging of Temporal and Spatial Dynamics During in vivo Seizure-like Activity

A multi-modal imaging system was used to analyze the spatiotemporal evolution of correlates to neural activity in a uniform cortical seizure model. We observed wave-like propagation during seizures and subtle spatial variation during drug interference.

Miniature device for chronic, label-free multi-modal optical imaging of cortical hemodynamics in rats

We report on a novel miniature head-mounted imaging system for simultaneous optical recording of brain blood flow and changes in brain blood oxygenation in a rat. Measurements of blood flow speeds are accomplished using Laser Speckle Contrast Imaging (LSCI) technique, while changes in blood oxygenation are measured via Intrinsic Optical Signal Imaging (IOSI) technique. A single multi-wavelength (wavelength = 680, 795, 850 nm) package of vertical cavity surface emitting lasers (VCSELs) is used as the sole brain illumination source. VCSELs enable rapid toggling between wavelengths, and between high-coherence and low-coherence modes, necessary for LSCI and IOSI, respectively. The combination of a miniature light source and a small 10-bit CCD camera sensor lead to a sub-20 g device mass. The miniature imaging system, including the lens, camera, and illumination lasers, is packaged as a module, and is mounted on a chronic implanted observation window that is surgically placed in the skull, allowing for repeated measurements and removal of the imaging system from the rats head after the imaging session. The imaging system allows for a 2mm-diameter field of view and a resolution of 7.4 µm. It will allow neurophysiologists to correlate standard behavioural assays to neurovascular response in animal models, and thus enrich their understanding of neurovascular coupling dynamics of brain disorders and diseases such as stroke and epilepsy.