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Skin suturing and cortical surface viral infusion improves imaging of neuronal ensemble activity with head-mounted miniature microscopes.

ABSTRACT: BACKGROUND:In vivo optical imaging of neural activity provides important insights into brain functions at the single-cell level. Cranial windows and virally delivered calcium indicators are commonly used for imaging cortical activity through two-photon microscopes in head-fixed animals. Recently, head-mounted one-photon microscopes have been developed for freely behaving animals. However, minimizing tissue damage from the virus injection procedure and maintaining window clarity for imaging can be technically challenging. NEW METHOD:We used a wide-diameter glass pipette at the cortical surface for infusing the viral calcium reporter AAV-GCaMP6 into the cortex. After infusion, the scalp skin over the implanted optical window was sutured to facilitate postoperative recovery. The sutured scalp was removed approximately two weeks later and a miniature microscope was attached above the window to image neuronal activity in freely moving mice. RESULTS:We found that cortical surface virus infusion efficiently labeled neurons in superficial layers, and scalp skin suturing helped to maintain the long-term clarity of optical windows. As a result, several hundred neurons could be recorded in freely moving animals. COMPARISON WITH EXISTING METHODS:Compared to intracortical virus injection and open-scalp postoperative recovery, our methods minimized tissue damage and dura overgrowth underneath the optical window, and significantly increased the experimental success rate and the yield of identified neurons. CONCLUSION:Our improved cranial surgery technique allows for high-yield calcium imaging of cortical neurons with head-mounted microscopes in freely behaving animals. This technique may be beneficial for other optical applications such as two-photon microscopy, multi-site imaging, and optogenetic modulation.

SUBMITTER: Li X

PROVIDER: S-EPMC5650079 | biostudies-literature | 2017 Nov

REPOSITORIES: biostudies-literature

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Skin suturing and cortical surface viral infusion improves imaging of neuronal ensemble activity with head-mounted miniature microscopes.

Li Xinjian X Cao Vania Y VY Zhang Wenyu W Mastwal Surjeet S SS Liu Qing Q Otte Stephani S Wang Kuan Hong KH

Journal of neuroscience methods 20170819

<h4>Background</h4>In vivo optical imaging of neural activity provides important insights into brain functions at the single-cell level. Cranial windows and virally delivered calcium indicators are commonly used for imaging cortical activity through two-photon microscopes in head-fixed animals. Recently, head-mounted one-photon microscopes have been developed for freely behaving animals. However, minimizing tissue damage from the virus injection procedure and maintaining window clarity for imagi ...[more]

PMID: 28830724

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Project description:SignificanceIntrinsic optical signals (IOS) generated in the cortical tissue as a result of various interacting metabolic processes are used extensively to elucidate the underlying mechanisms that govern neurovascular coupling. However, current IOS measurements still often rely on bulky, tabletop imaging systems, and there remains a dearth of studies in freely moving subjects. Lightweight, miniature head-mounted imaging devices provide unique opportunities for investigating cortical dynamics in small animals under a variety of naturalistic behavioral settings.AimThe aim of this work was to monitor IOS in the somatosensory cortex of wild-type mice by developing a lightweight, biocompatible imaging device that readily lends itself to animal experiments in freely moving conditions.ApproachHerein we describe a method for realizing long-term IOS imaging in mice using a 0.54-g, compact, CMOS-based, head-mounted imager. The two-part module, consisting of a tethered sensor plate and a base plate, allows facile assembly prior to imaging sessions and disassembly when the sensor is not in use. LEDs integrated into the device were chosen to illuminate the cortical mantle at two different wavelengths in the visible regime (λcenter: 535 and 625 nm) for monitoring volume- and oxygenation state-dependent changes in the IOS, respectively. To test whether the system can detect robust cortical responses, we recorded sensory-evoked IOS from mechanical stimulation of the hindlimbs (HL) of anesthetized mice in both acute and long-term implantation conditions.ResultsCortical IOS recordings in the primary somatosensory cortex hindlimb receptive field (S1HL) of anesthetized mice under green and red LED illumination revealed robust, multiphasic profiles that were time-locked to the mechanical stimulation of the contralateral plantar hindpaw. Similar intrinsic signal profiles observed in S1HL at 40 days postimplantation demonstrated the viability of the approach for long-term imaging. Immunohistochemical analysis showed that the brain tissue did not exhibit appreciable immune response due to the device implantation and operation. A proof-of-principle imaging session in a freely behaving mouse showed minimal locomotor impediment for the animal and also enabled estimation of blood flow speed.ConclusionsWe demonstrate the utility of a miniature cortical imaging device for monitoring IOS and related hemodynamic processes in both anesthetized and freely moving mice, cueing potential for applications to some neuroscientific studies of sensation and naturalistic behavior.

Project description:BackgroundAlthough surgical suturing is one of the most important basic skills, many medical school graduates do not acquire sufficient knowledge of it due to its lack of integration into the curriculum or a shortage of tutors. E-learning approaches attempt to address this issue but still rely on the involvement of tutors. Furthermore, the learning experience and visual-spatial ability appear to play a critical role in surgical skill acquisition. Virtual reality head-mounted displays (HMDs) could address this, but the benefits of immersive and stereoscopic learning of surgical suturing techniques are still unclear.Material and methodsIn this multi-arm randomized controlled trial, 150 novices participated. Three teaching modalities were compared: an e-learning course (monoscopic), an HMD-based course (stereoscopic, immersive), both self-directed and a tutor-led course with feedback. Suturing performance was recorded by video camera both before and after course participation (>26 h of video material) and assessed in a blinded fashion using the Objective Structured Assessment of Technical Skills (OSATS) Global Rating Score (GRS). Furthermore, the optical flow of the videos was determined using an algorithm. The number of sutures performed was counted, the visual-spatial ability was measured with the Mental Rotation Test (MRT), and courses were assessed with questionnaires.ResultsStudents' self-assessment in the HMD-based course was comparable to that of the tutor-led course and significantly better than in the e-learning course ( P =0.003). Course suitability was rated best for the tutor-led course ( x̄ =4.8), followed by the HMD-based ( x̄ =3.6) and e-learning ( x̄ =2.5) courses. The median ΔGRS between courses was comparable ( P =0.15) at 12.4 (95% CI 10.0-12.7) for the e-learning course, 14.1 (95% CI 13.0-15.0) for the HMD-based course, and 12.7 (95% CI 10.3-14.2) for the tutor-led course. However, the ΔGRS was significantly correlated with the number of sutures performed during the training session ( P =0.002), but not with visual-spatial ability ( P =0.615). Optical flow ( R2 =0.15, P <0.001) and the number of sutures performed ( R2 =0.73, P <0.001) can be used as additional measures to GRS.ConclusionThe use of HMDs with stereoscopic and immersive video provides advantages in the learning experience and should be preferred over a traditional web application for e-learning. Contrary to expectations, feedback is not necessary for novices to achieve a sufficient level in suturing; only the number of surgical sutures performed during training is a good determinant of competence improvement. Nevertheless, feedback still enhances the learning experience. Therefore, automated assessment as an alternative feedback approach could further improve self-directed learning modalities. As a next step, the data from this study could be used to develop such automated AI-based assessments.

Project description:This study offers a new method for examining the bodily, manual, and eye movements of a chimpanzee at the micro-level. A female chimpanzee wore a lightweight head-mounted eye tracker (60 Hz) on her head while engaging in daily interactions with the human experimenter. The eye tracker recorded her eye movements accurately while the chimpanzee freely moved her head, hands, and body. Three video cameras recorded the bodily and manual movements of the chimpanzee from multiple angles. We examined how the chimpanzee viewed the experimenter in this interactive setting and how the eye movements were related to the ongoing interactive contexts and actions. We prepared two experimentally defined contexts in each session: a face-to-face greeting phase upon the appearance of the experimenter in the experimental room, and a subsequent face-to-face task phase that included manual gestures and fruit rewards. Overall, the general viewing pattern of the chimpanzee, measured in terms of duration of individual fixations, length of individual saccades, and total viewing duration of the experimenter's face/body, was very similar to that observed in previous eye-tracking studies that used non-interactive situations, despite the differences in the experimental settings. However, the chimpanzee viewed the experimenter and the scene objects differently depending on the ongoing context and actions. The chimpanzee viewed the experimenter's face and body during the greeting phase, but viewed the experimenter's face and hands as well as the fruit reward during the task phase. These differences can be explained by the differential bodily/manual actions produced by the chimpanzee and the experimenter during each experimental phase (i.e., greeting gestures, task cueing). Additionally, the chimpanzee's viewing pattern varied depending on the identity of the experimenter (i.e., the chimpanzee's prior experience with the experimenter). These methods and results offer new possibilities for examining the natural gaze behavior of chimpanzees.

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Skin suturing and cortical surface viral infusion improves imaging of neuronal ensemble activity with head-mounted miniature microscopes.

Publications

Skin suturing and cortical surface viral infusion improves imaging of neuronal ensemble activity with head-mounted miniature microscopes.

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