Janaka Senarathna

Miniaturized microscopes or ‘miniscopes’ for neuroimaging in freely behaving animals mostly operate over short durations (24 h) imaging, remote operation and simultaneous characterization of multiple neurophysiological variables such as neuronal activity, blood flow, blood volume, oxygenation and cellular dynamics (a capability that we call ‘neurosurveillance’). Thus, we developed the ‘CloudScope’, a cloud-based multicontrast miniscope for autonomous neurosurveillance in freely behaving animals. Its cloud-based architecture enables global remote operation and continuous acquisition of multicontrast images over CNS disease model life cycles. We demonstrate CloudScope’s neurosurveillance capabilities in predicting behavior from 24-h neuroimaging data with deep learning (DL), characterizing neurovascular changes during natural behavior, seizure-induced neurovascular disruptions, and in vivo cellular and microvascular phenotyping of brain tumor microenvironments. Finally, CloudScope’s architecture enables ‘time-shared’ imaging, which potentially reduces animal use. Collectively, CloudScope’s neurosurveillance capabilities in conjunction with CNS disease models establish a new paradigm for characterizing their etiology and evolution.

Extracorporeal cardiopulmonary resuscitation (ECPR) is increasingly used for refractory cardiac arrest, yet neurologic outcome is often limited by acute brain injury (ABI) secondary to disrupted cerebrovascular autoregulation (CVAR). We established a proof-of-concept porcine model that combines veno-arterial ECMO with dual-mode optical neuromonitoring. After 10 minutes of untreated ventricular fibrillation, ECMO support was initiated, and a cranial window enabled real-time laser speckle contrast imaging (LSCI) of cerebral blood flow (CBF) and intrinsic optical signal (IOS) mapping of cerebral blood volume (CBV). Mean arterial pressure (MAP) was recorded continuously, and vessel-specific hemoglobin-volume index (HVx) was calculated to quantify CVAR. Cortical vessels became pressure-passive—HVx \texorpdfstring{$\geq$}{>=} 0.3, indicating loss of autoregulation—during the initial minutes of reperfusion and again when ECMO pump speed was deliberately reduced to 75\% and 50\% of baseline. The impairment was heterogeneous and reversible, with adjacent vessels showing either intact (HVx \texorpdfstring{$\leq$}{< =} 0) or impaired CVAR in the same epoch. Neither pump flow nor systemic MAP reliably mirrored these microvascular changes: at several time points, CBF rose while pump output and MAP fell, and vice versa. Despite technical challenges (motion artifact, coagulopathy, lack of invasive flow validation), this single-subject study demonstrates the feasibility of simultaneous LSCI and IOS during ECPR.

Recent advances in low-power wireless-capable system-on-chips (SoCs) have accelerated diverse Internet of Things (IoT) applications, encompassing wearables, asset monitoring, and more. Concurrently, the field of neuroimaging has experienced escalating demand for lightweight, untethered, low-power systems capable of imaging in small animals. This article explores the feasibility of using a low-power asset monitoring system as the basis of a new architecture for fluorescence and hemodynamic contrast-based wireless functional imaging. The core system architecture hinges on the fusion of a Bluetooth Low Energy (BLE) 5.2 SoC and a low-power 560 × 560, 8-bit monochrome CMOS image sensor module. Successful integration of a multicontrast optical front-end consisting of a fluorescence channel (FL) and an intrinsic optical signal (IOS) channel resulted in the creation of a wireless microscope called ‘BLEscope’. Next, we developed a wireless (i.e., BLE) protocol to remotely operate the BLEscope via a laptop and acquire in vivo images at 1 frame per second (fps). We then conducted a comprehensive characterization of the BLEscope to assess its optical capabilities and power consumption. We report a new benchmark for continuous wireless imaging of ∼1.5 hours with a 100 mAh battery. Via the FL channel of the BLEscope, we successfully tracked the kinetics of an intravenously injected fluorescent tracer and acquired images of fluorescent brain tumor cells in vivo. Via the IOS channel, we characterized the differential response of normal and tumor-associated blood vessels to a carbogen gas inhalation challenge. When miniaturized, the BLEscope will result in a new class of low-power, implantable or wireless microscopes that could transform preclinical and clinical neuroimaging applications.

The development of new endovascular technologies for transarterial embolization has relied on animal studies to validate efficacy before clinical trials are undertaken. Because embolizations in animals and patients are primarily conducted with fluoroscopy alone, local hemodynamic changes are not assessed during testing. However, such hemodynamic metrics could be important indicators of procedure efficacy that could support improved patient outcomes, such as via the determination of procedural endpoints. The purpose of this study is to create a high-fidelity benchtop system for multiparametric (i.e., hemodynamic and imaging) assessment of transarterial embolization procedures. The benchtop system consists of a 3D printed, anatomically accurate vascular phantom; a flow loop with a cardiac output simulator; a high-speed video camera; and pressure transducers and flow meters. This system enabled us to vary the heart rate and blood pressure and to simulate clinically relevant hemodynamic states, such as healthy adult, aortic regurgitation, and hypovolemic shock. With our radiation-free angiography-mimetic imaging system, we could simultaneously assess gauge pressure and flow values during transarterial embolization. We demonstrated the feasibility of recapitulating the digital subtraction angiography workflow. Finally, we highlighted the utility of this system by characterizing the relationship between an imaging-based metric of procedural endpoint and intravascular flow. We also characterized hemodynamic changes associated with particle embolization within a branch of the hepatic artery and found them to be within reported patient data. Our benchtop vascular system was low-cost and reproduced transarterial embolization-related hemodynamic phenomena with high fidelity. We believe that this novel platform enables the characterization of patient physiology, novel catheterization devices, and techniques.

Cerebral vascular autoregulation is impaired following resuscitation from cardiac arrest (CA), and its quantification may allow assessing CA-induced brain injury. However, hyperemia occurring immediately post-resuscitation limits the application of most metrics that quantify autoregulation. Therefore, to characterize autoregulation during this critical period, we developed three novel metrics based on how the cerebrovascular resistance (CVR) covaries with changes in cerebral perfusion pressure (CPP): (i) θ_CVR, which quantifies the CVR vs CPP gradient, (ii) a CVR-based transfer function analysis, and (iii) CVRx, the correlation coefficient between CPP and CVR. We tested these metrics in a model of asphyxia induced CA and resuscitation using seven adult male Wistar rats. Mean arterial pressure (MAP) and cortical blood flow recorded for 30 min post-resuscitation via arterial cannulation and laser speckle contrast imaging, were used as surrogates of CPP and cerebral blood flow (CBF), while CVR was computed as the CPP/CBF ratio. Using our metrics, we found that the status of cerebral vascular autoregulation altered substantially during hyperemia, with changes spread throughout the 0–0.05 Hz frequency band. Our metrics push the boundary of how soon autoregulation can be assessed, and if validated against outcome markers, may help develop a reliable metric of brain injury post-resuscitation.

Abnormal tumor hemodynamics are a critical determinant of a tumor’s microenvironment (TME), and profoundly affect drug delivery, therapeutic efficacy and the emergence of drug and radio-resistance. Since multiple hemodynamic variables can simultaneously exhibit transient and spatiotemporally heterogeneous behavior, there is an exigent need for analysis tools that employ multiple variables to characterize the anomalous hemodynamics within the TME. To address this, we developed a new toolkit called HemoSYS for quantifying the hemodynamic landscape within angiogenic microenvironments. It employs multivariable time-series data such as in vivo tumor blood flow (BF), blood volume (BV) and intravascular oxygen saturation (Hb_sat) acquired concurrently using a wide-field multicontrast optical imaging system. The HemoSYS toolkit consists of propagation, clustering, coupling, perturbation and Fourier analysis modules. We demonstrate the utility of each module for characterizing the in vivo hemodynamic landscape of an orthotropic breast cancer model. With HemoSYS, we successfully described: (i) the propagation dynamics of acute hypoxia; (ii) the initiation and dissolution of distinct hemodynamic niches; (iii) tumor blood flow regulation via local vasomotion; (iv) the hemodynamic response to a systemic perturbation with carbogen gas; and (v) frequency domain analysis of hemodynamic heterogeneity in the TME. HemoSYS (freely downloadable via the internet) enables vascular phenotyping from multicontrast in vivo optical imaging data. Its modular design also enables characterization of non-tumor hemodynamics (e.g. brain), other preclinical disease models (e.g. stroke), vascular-targeted therapeutics, and hemodynamic data from other imaging modalities (e.g. MRI).

Traditional optical imaging systems are designed for imaging with a single contrast mechanism, and therefore can interrogate only a single neurophysiologic variable. However, the biological complexity underlying neurophysiological function and its alteration in neurodegenerative diseases, requires the simultaneous interrogation of multiple neurophysiologic variables to arrive at a better understanding. Today’s multicontrast optical imaging systems satisfy this need, but suffer from some inherent limitations. Owing to the need to integrate multiple contrast mechanisms, these imaging systems tend to be benchtop-based and unportable, often requiring animals to be anesthetized, custom built and expensive. This limits their widespread adoption. Miniaturization, although technically challenging, remains a potential solution to these limitations. To address this unmet need, here we present the design considerations and practical guidelines for building inexpensive, miniaturized, and portable multicontrast optical neuroimaging systems that allow comprehensive interrogation of brain function in freely behaving rodents. We then showcase an example tri-contrast miniature neuroimaging system and demonstrate the implementation of our guidelines. We conclude by demonstrating the utility of such a miniature multicontrast neuroimaging system by interrogating in an awake rodent the tumor extent, angiogenic vascular sprouting, flow establishment in the newly formed vessels, as well as anomalies in resting-state microvascular fluctuations in a preclinical model of brain tumor progression.

Neurovascular coupling, cerebrovascular remodeling and hemodynamic changes are critical to brain function, and dysregulated in neuropathologies such as brain tumors. Interrogating these phenomena in freely behaving animals requires a portable microscope with multiple optical contrast mechanisms. Therefore, we developed a miniaturized microscope with: a fluorescence (FL) channel for imaging neural activity (e.g., GCaMP) or fluorescent cancer cells (e.g., 9L-GFP); an intrinsic optical signal (IOS) channel for imaging hemoglobin absorption (i.e., cerebral blood volume); and a laser speckle contrast (LSC) channel for imaging perfusion (i.e., cerebral blood flow). Following extensive validation, we demonstrate the microscope’s capabilities via experiments in unanesthetized murine brains that include: (i) multi-contrast imaging of neurovascular changes following auditory stimulation; (ii) wide-area tonotopic mapping; (iii) EEG-synchronized imaging during anesthesia recovery; and (iv) microvascular connectivity mapping over the life-cycle of a brain tumor. This affordable, flexible, plug-and-play microscope heralds a new era in functional imaging of freely behaving animals.