Parvalbumin interneurons (PV-INs) are the largest subpopulation of GABAergic neurons. Recent evidence suggests transcallosal projections of PV-INs within primary motor, visual and auditory cortices. Whether transcallosal PV-INs feature prominently, and the extent to which their local influence affects global excitatory activity is unknown. We created a novel imaging system and mouse line for all-optical neuronal probing and readout to map local and global interactions between excitatory neurons and PV-INs over the cortex in awake mice. PV-INs extend inhibitory influences spanning several mm over the cortex and into more distant, ipsilateral regions. Further, we show region-specific interhemispheric inhibitory influences of PV-INs.
Recent developments in optogenetics allow for quick and minimally invasive methods of mapping functional brain circuits in animal models. DeepLabCut (DLC), a toolbox for markerless pose estimation, offers the ability to track features in three-dimensions. We demonstrate a hybrid method utilizing DLC and light-based, optogenetic motor mapping to concurrently localize motor representations of multiple limbs in mice. Our results suggest that behaviorally-relevant, motor movements involving multiple limbs reside in overlapping cortical representations of each limb. Applications of this technique include characterizing recovery of finer, articulated movements of affected limbs after stroke, or mapping brain network activity during naturalistic behavior.
Psychedelics, which effects appear to be mediated via seratonin-2A receptors(5-HT2AR), are a promising therapeutic. Recent human studies have shown drastic changes in functional network organization but may be confounded due to Serotonin’s vasoconstriction effects. We aimed to determine if 5-HT2AR agonism (via Lisuride), alone, differentially affect cortical neuronal and hemodynamic activity, functional connectivity (FC), and neurovascular coupling (NVC). Eight Thy1-jRGECO1a mice were imaged for 60-minutes under awake, resting-state conditions using wide-field optical imaging. Lisuride significantly reduced neuronal and hemodynamic bilateral FC and activity in a region- and frequency-band-specific manner. Furthermore, delayed NVC was observed. Results suggest 5-HT2AR agonism alters hemodynamic activity, FC and NVC. Future work will evaluate acute effects of commonly used psychedelics on brain function and differentiate hallucinatory vs non-hallucinatory 5-HT2A receptor activation.
Understanding how different central nervous system diseases affect different components of neurovascular coupling will allow for linking changes in neural or metabolic dysfunction to changes in hemodynamic signaling upon which blood-based imaging methods rely. We developed a dual fluorophore imaging system for simultaneous, high-speed mapping of neural, metabolic, and hemodynamic activity. Proof-of-concept measurements of spontaneous and stimulus-evoked dynamics are presented in awake and anesthetized mice. This flexible hardware platform allows for integrating optogenetic stimulation for all optical neural circuit interrogation and readout, and for examining the interaction between multiple cell populations.
Recent developments in optogenetics allow for quick and minimally invasive methods of mapping functional brain circuits in animal models. DeepLabCut (DLC), a toolbox for markerless pose estimation, offers the ability to track features in 3-dimensions. We demonstrate a hybrid method utilizing DLC and light-based, optogenetic motor mapping to concurrently localize motor representations of multiple limbs in mice. Our results suggest that behaviorally-relevant, motor movements involving multiple limbs reside in overlapping cortical representations of each limb. Applications of this technique include characterizing recovery of finer, articulated movements of affected limbs after stroke, or mapping brain network activity during naturalistic behavior.
Understanding how different diseases of the central nervous system affect neurovascular coupling will allow for linking changes in neural or metabolic dysfunction to changes in hemodynamic signaling upon which blood-based imaging methods rely. We developed a dual fluorophore imaging system for simultaneous, high-speed mapping of neural, metabolic, and hemodynamic activity. Proof-of-concept measurements of spontaneous and stimulus-evoked dynamics are presented in awake and anesthetized mice. This flexible hardware platform allows for integrating optogenetic stimulation for all optical neural circuit interrogation and readout, and for examining the interaction between multiple cell populations.
Recent developments in optogenetics allow for quick and minimally-invasive methods of studying functional brain organization in animal models. DeepLabCut (DLC), a toolbox for markerless pose estimation, offers the ability to track user-defined features in 3-dimensions with human level accuracy. We demonstrate a hybrid method utilizing DLC and optogenetic motor mapping to localize the movements of multiple modalities to the mouse cortex. We outline a pipeline to map and characterize multiple motor representations in anesthetized and awake mice. Furthermore, we identify behaviorally-relevant motor movements of multiple limbs reside in overlapping cortical representations of the respective limbs.
Tumors in the central nervous system (CNS) can produce significant behavior deficits. These deficits significantly reduce the quality of life of long-term brain tumor survivors. Motor deficits can be the main presenting symptoms that brings a brain tumor patient into the hospital. Additionally, motor deficits can either improve or worsen after treatment. Its poorly understood how tumor phenotype (e.g infiltrative or circumscribed) contributes to the development and evolution of motor deficits. Resting state functional connectivity magnetic resonance imaging (rsfc-MRI) could help determine a relationship between brain function and tumor phenotype. rsfc-MRI provides measurements of functional connectivity (FC), which is a measure of brain activity and has been shown to correlate with neurological function (e.g. cognitive and motor deficit) in many disease states. Additional studies are needed before rsfc-MRI can be included in the standard of care for brain tumor patients because it is largely unknown how important clinical factors, such as tumor location and tumor burden (e.g. tumor volume), modulate the association between FC measurements and neurological performance. However, human studies are unlikely to have the statistical power to determine the relationships between FC, clinical factors, and neurological function because it is difficult to enroll many human subjects while controlling for a factor. Fortunately, mice can be used to control for a factor in many genetically identical specimens. Therefore, FC assessments need to be applied to studies in mouse models of brain tumors to assess the interactions be specific clinical factors, FC measurements and neurological function. FC measurements are infrequently obtained in mouse models because it is technically difficult to perform rsfc-MRI on the small mouse brain. Fortunately, functional connectivity optical intrinsic signal imaging (fcOISI) can be used obtain FC measurements in mice. In this study, we investigated how tumor phenotype modulates the relationship between tumor burden measures and motor function. We injected in two groups of mice with human brain tumor cell lines that exhibit different growth phenotypes. We assessed the motor function of the mice by their performance on a battery of sensorimotor tests. We determined tumor burden with bioluminescence and MRI and assessed FC with fcOISI. Lastly, we performed linear regression analysis to determine how these measured factors interact in the prediction of performance on the motor tests.
Modulation of brain state, e.g., by anesthesia, alters the correlation structure of spontaneous activity, especially in the delta band. This effect has largely been attributed to the ∼1 Hz slow oscillation that is characteristic of anesthesia and nonrapid eye movement (NREM) sleep. However, the effect of the slow oscillation on correlation structures and the spectral content of spontaneous activity across brain states (including NREM) has not been comprehensively examined. Further, discrepancies between activity dynamics observed with hemoglobin versus calcium (GCaMP6) imaging have not been reconciled. Lastly, whether the slow oscillation replaces functional connectivity (FC) patterns typical of the alert state, or superimposes on them, remains unclear. Here, we use wide-field calcium imaging to study spontaneous cortical activity in awake, anesthetized, and naturally sleeping mice. We find modest brain state-dependent changes in infraslow correlations but larger changes in GCaMP6 delta correlations. Principal component analysis of GCaMP6 sleep/anesthesia data in the delta band revealed that the slow oscillation is largely confined to the first three components. Removal of these components revealed a correlation structure strikingly similar to that observed during wake. These results indicate that, during NREM sleep/anesthesia, the slow oscillation superimposes onto a canonical FC architecture.
For functional neuroimaging, existing small-animal diffuse optical tomography (DOT) systems either do not provide adequate temporal sampling rates, have sparse spatial sampling, or have limited three-dimensional fields of view. To achieve adequate frame rates (1-10 Hz), we have constructed a system using sCMOS detection-based DOT, with asymmetric measurements, with many (>10,000) detectors and fewer (<100) structured illumination patterns (using digital micromirror devices: DMDs). The system employs multiple views, involving multiple cameras and illuminators, to provide a three-dimensional field of view. To coregister the measurements with the mouse head anatomy, we developed a surface profiling method in which point illumination patterns are scanned over the mouse head and combined with calibration data to create three-dimensional point clouds and meshes representing the head. We applied this method to a 3D-printed figurine, and the resulting mesh had surface vertices whose positions deviated 0.4 ± 0.2 mm (mean ± SD) from the original "ground truth" mesh that had been employed to 3D-print the figurine. To evaluate the imaging system's resolution, field of view, and sensitivity versus depth, we placed simulated activations at different depths within a tissue model of a real mouse head imaged with our surface profiling method. Results indicate that this imaging system is sensitive to absorption changes at depths of >3 mm. In addition, a partial (one-camera, one-illuminator) version of the system successfully imaged neural activations evoked by forepaw stimulation of a live mouse.
Recent fcMRI studies examining spontaneous brain activity after stoke have revealed disrupted global patterns of functional connectivity (FC). Interestingly, acute interhemispheric homotopic FC has been shown to be predictive of recovery potential. While substantial indirect evidence also suggests that homotopic brain activity may directly impact recovery, results in humans are extremely varied. A better understanding of how activity within networks functionally-connected to lesioned tissue influences brain plasticity might improve therapeutic strategies. We combine cell-type specific optogenetic targeting with optical intrinsic signal (OIS) imaging to assess the effects of homotopic contralesional activity (specifically in excitatory CamKIIa pyramidal neurons) on FC, cortical remapping, and behavior after stroke. Thirty-one mice were housed in enriched cages for the experiment. OIS imaging was performed before, 1, and 4 weeks after photothrombosis of left forepaw somatosensory cortex (S1fp). On day 1 after stroke, 17 mice were subjected to chronic, intermittent optical stimulation of right S1fp for 10 min, 5 days/week for 4 weeks. New cortical representations of left S1fp appeared in non-stimulated mice at week 1, but not in stimulated mice (p=0.005). Evoked responses were comparable in both groups at week 4 (p=0.57). Homotopic FC between left and right S1fp regions was equally reduced in both groups (p=0.012) at week 1. However, in non-stimulated mice, behavioral performance and FC between right S1fp and left perilesional S1 cortex was significantly higher by 4 weeks compared to stimulated mice (p=0.009). Our results suggest that increased homotopic, contralesional activity in excitatory neurons negatively influences spontaneous recovery following ischemic stroke.
Functional magnetic resonance imaging (fMRI) has transformed our understanding of the brain’s functional organization. However, mapping subunits of a functional network using hemoglobin alone presents several disadvantages. Evoked and spontaneous hemodynamic fluctuations reflect ensemble activity from several populations of neurons making it difficult to discern excitatory vs inhibitory network activity. Still, blood-based methods of brain mapping remain powerful because hemoglobin provides endogenous contrast in all mammalian brains. To add greater specificity to hemoglobin assays, we integrated optical intrinsic signal(OIS) imaging with optogenetic stimulation to create an Opto-OIS mapping tool that combines the cell-specificity of optogenetics with label-free, hemoglobin imaging. Before mapping, titrated photostimuli determined which stimulus parameters elicited linear hemodynamic responses in the cortex. Optimized stimuli were then scanned over the left hemisphere to create a set of optogenetically-defined effective connectivity (Opto-EC) maps. For many sites investigated, Opto-EC maps exhibited higher spatial specificity than those determined using spontaneous hemodynamic fluctuations. For example, resting-state functional connectivity (RS-FC) patterns exhibited widespread ipsilateral connectivity while Opto-EC maps contained distinct short- and long-range constellations of ipsilateral connectivity. Further, RS-FC maps were usually symmetric about midline while Opto-EC maps displayed more heterogeneous contralateral homotopic connectivity. Both Opto-EC and RS-FC patterns were compared to mouse connectivity data from the Allen Institute. Unlike RS-FC maps, Thy1-based maps collected in awake, behaving mice closely recapitulated the connectivity structure derived using ex vivo anatomical tracer methods. Opto-OIS mapping could be a powerful tool for understanding cellular and molecular contributions to network dynamics and processing in the mouse brain.
Gliomas are known to cause significant changes in normal brain function that lead to cognitive deficits. Disruptions in resting state networks (RSNs) are thought to underlie these changes. However, investigating the effects of glioma growth on RSNs in humans is complicated by the heterogeneity in lesion size, type, and location across subjects. In this study, we evaluated the effects of tumor growth on RSNs over time in a controlled mouse model of glioma growth. Methods: Glioma cells (5x104-105 U87s) were stereotactically injected into the forepaw somatosensory cortex of adult nude mice (n=5). Disruptions in RSNs were evaluated weekly with functional connectivity optical intrinsic signal imaging (fcOIS). Tumor growth was monitored with MRI and weekly bioluminescence imaging (BLI). In order to characterize how tumor growth affected different RSNs over time, we calculated a number of functional connectivity (fc) metrics, including homotopic (bilateral) connectivity, spatial similarity, and node degree. Results: Deficits in fc initiate near the lesion, and over a period of several weeks, extend more globally. The reductions in spatial similarity were found to strongly correlate with the BLI signal indicating that increased tumor size is associated with increased RSN disruption. Conclusions: We have shown that fcOIS is capable of detecting alterations in mouse RSNs due to brain tumor growth. A better understanding of how RSN disruption contributes to the development of cognitive deficits in brain tumor patients may lead to better patient risk stratification and consequently improved cognitive outcomes.
Optical intrinsic signal (OIS) imaging has been a powerful tool for capturing functional brain hemodynamics in rodents. Recent wide field-of-view implementations of OIS have provided efficient maps of functional connectivity from spontaneous brain activity in mice. However, OIS requires scalp retraction and is limited to superficial cortical tissues. Diffuse optical tomography (DOT) techniques provide noninvasive imaging, but previous DOT systems for rodent neuroimaging have been limited either by sparse spatial sampling or by slow speed. Here, we develop a DOT system with asymmetric source–detector sampling that combines the high-density spatial sampling (0.4 mm) detection of a scientific complementary metal-oxide-semiconductor camera with the rapid (2 Hz) imaging of a few (<50) structured illumination (SI) patterns. Analysis techniques are developed to take advantage of the system’s flexibility and optimize trade-offs among spatial sampling, imaging speed, and signal-to-noise ratio. An effective source–detector separation for the SI patterns was developed and compared with light intensity for a quantitative assessment of data quality. The light fall-off versus effective distance was also used for in situ empirical optimization of our light model. We demonstrated the feasibility of this technique by noninvasively mapping the functional response in the somatosensory cortex of the mouse following electrical stimulation of the forepaw.
Resting-state functional connectivity (RSFC) imaging is an emerging neuroimaging approach that aims to identify spontaneous cerebral hemodynamic fluctuations and their associated functional connections. Clinical studies have demonstrated that RSFC is altered in brain disorders such as stroke, Alzheimer’s, autism, and epilepsy. However, conventional neuroimaging modalities cannot easily be applied to mice, the most widely used model species for human brain disease studies. For instance, functional magnetic resonance imaging (fMRI) of mice requires a very high magnetic field to obtain a sufficient signal-to-noise ratio and spatial resolution. Functional connectivity mapping with optical intrinsic signal imaging (fcOIS) is an alternative method. Due to the diffusion of light in tissue, the spatial resolution of fcOIS is limited, and experiments have been performed using an exposed skull preparation. In this study, we show for the first time, the use of photoacoustic computed tomography (PACT) to noninvasively image resting-state functional connectivity in the mouse brain, with a large field of view and a high spatial resolution. Bilateral correlations were observed in eight regions, as well as several subregions. These findings agreed well with the Paxinos mouse brain atlas. This study showed that PACT is a promising, non-invasive modality for small-animal functional brain imaging.
We have developed a fluorescence goggle device for intraoperative oncologic imaging. With our system design, the
surgeon can directly visualize the fluorescence information from the eyepieces in real time without any additional
monitor, which can improve one's coordination and surgical accuracy. In conjunction with targeting fluorescent dyes,
the goggle device can successfully detect tumor margins and small nodules that are not obvious to naked eye. This can
potentially decrease the incidence of incomplete resection.
The specificity of molecular and functional photoacoustic (PA) images depends on the accuracy of the photoacoustic absorption spectroscopy. The PA signal is proportional to the product of the optical absorption coefficient and local light fluence; quantitative PA measurements of the optical absorption coefficient therefore require an accurate estimation of optical fluence. Light-modeling aided by diffuse optical tomography (DOT) can be used to map the required fluence and to reduce errors in traditional PA spectroscopic analysis. As a proof-of-concept, we designed a tissue-mimicking phantom to demonstrate how fluence-related artifacts in PA images can lead to misrepresentations of tissue properties. To correct for these inaccuracies, the internal fluence in the tissue phantom was estimated by using DOT to reconstruct spatial distributions of the absorption and reduced scattering coefficients of multiple targets within the phantom. The derived fluence map, which only consisted of low spatial frequency components, was used to correct PA images of the phantom. Once calibrated to a known absorber, this method reduced errors in estimated absorption coefficients from 33% to 6%. These results experimentally demonstrate that combining DOT with PA imaging can significantly reduce fluence-related errors in PA images, while producing quantitatively accurate, high-resolution images of the optical absorption coefficient.
The specificity of both molecular and functional
photoacoustic (PA) images depends on the accuracy of the
photoacoustic absorption spectroscopy. Because the PA signal is
a product of both the optical absorption coefficient and the local
light fluence, quantitative PA measurements of absorption
require an accurate estimate of the optical fluence. Lightmodeling
aided by diffuse optical tomography (DOT) methods
can be used to provide the required fluence map and to reduce
errors in traditional PA spectroscopic analysis. As a proof-ofconcept,
we designed a phantom to demonstrate artifacts
commonly found in photoacoustic tomography (PAT) and how
fluence-related artifacts in PAT images can lead to
misrepresentations of tissue properties. Specifically, we show that
without accounting for fluence-related inhomogeneities in our
phantom, errors in estimates of the absorption coefficient from a
PAT image were as much as 33%. To correct for this problem,
DOT was used to reconstruct spatial distributions of the
absorption coefficients of the phantom, and along with the
surface fluence distribution from the PAT system, we calculated
the fluence everywhere in the phantom. This fluence map was
used to correct PAT images of the phantom, reducing the error in
the estimated absorption coefficient from the PAT image to less
than 5%. Thus, we demonstrate experimentally that combining
DOT with PAT can significantly reduce fluence-related errors in
PAT images, as well as produce quantitatively accurate, highresolution
images of the optical absorption coefficient.
We have developed a novel real-time intraoperative fluorescence imaging device that can detect near-infrared (NIR)
fluorescence and map sentinel lymph nodes (SLNs). In contrast to conventional imaging systems, this device is compact,
portable, and battery-operated. It is also wearable and thus allows hands-free operation of clinicians. The system directly
displays the fluorescence in its goggle eyepiece, eliminating the need for a remote monitor. Using this device in murine
lymphatic mapping, the SLNs stained with indocyanine green (ICG) can be readily detected. Fluorescence-guided SLN
resection under the new device was performed with ease. Ex vivo examination of resected tissues also revealed high
fluorescence level in the SLNs. Histology further confirmed the lymphatic nature of the resected SLNs.
Access to the requested content is limited to institutions that have purchased or subscribe to SPIE eBooks.
You are receiving this notice because your organization may not have SPIE eBooks access.*
*Shibboleth/Open Athens users─please
sign in
to access your institution's subscriptions.
To obtain this item, you may purchase the complete book in print or electronic format on
SPIE.org.
INSTITUTIONAL Select your institution to access the SPIE Digital Library.
PERSONAL Sign in with your SPIE account to access your personal subscriptions or to use specific features such as save to my library, sign up for alerts, save searches, etc.