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Mouse optical imaging for understanding resting-state functional connectivity in human fMRI
摘要： Resting-state functional connectivity (FC), which measures the temporal correlation of spontaneous hemodynamic activity between distant brain areas, is a widely accepted method in functional magnetic resonance imaging (fMRI) to assess the connectome of healthy and diseased human brains. A common assumption underlying FC is that it reflects the temporal structure of large-scale neuronal activity that is converted into large-scale hemodynamic activity. However, direct observation of such relationship has been difficult. In this commentary, we describe our recent progress regarding this topic. Recently, transgenic mice that express a genetically encoded calcium indicator (GCaMP) in neocortical neurons are enabling the optical recording of neuronal activity in large-scale with high spatiotemporal resolution. Using these mice, we devised a method to simultaneously monitor neuronal and hemodynamic activity and addressed some key issues related to the neuronal basis of FC. We propose that many important questions about human resting-state fMRI can be answered using GCaMP expressing transgenic mice as a model system.
关键词： mouse,calcium imaging,functional connectivity,fMRI,resting-state
更新于2025-09-23 15:23:52
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Retinal Characterization of the Thy1-GCaMP3 Transgenic Mouse Line After Optic Nerve Transection
摘要： PURPOSE. GCaMP3 is a genetically encoded calcium indicator for monitoring intracellular calcium dynamics. We characterized the expression pattern and functional properties of GCaMP3 in the Thy1-GCaMP3 transgenic mouse retina. METHODS. To determine the specificity of GCaMP3 expression, Thy1-GCaMP3 (B6; CBA-Tg(Thy1-GCaMP3)6Gfng/J) retinas were processed for immunohistochemistry with anti-green fluorescent protein (anti-GFP, to enhance GCaMP3 fluorescence), anti-RBPMS (retinal ganglion cell [RGC]–specific marker), and antibodies against amacrine cell markers (ChAT, GABA, GAD67, syntaxin). Calcium imaging was used to characterize functional responses of GCaMP3-expressing (GCaMP+) cells by recording calcium transients evoked by superfusion of kainic acid (KA; 10, 50, or 100 μM). In a subset of animals, optic nerve transection (ONT) was performed 3, 5, or 7 days prior to calcium imaging. RESULTS. GFP immunoreactivity colocalized with RBPMS but not amacrine cell markers in both ONT and non-ONT (control) groups. Calcium transients evoked by KA were reduced after ONT (50 μM KA; ΔF/F0 [SD]; control: 1.00 [0.67], day 3: 0.50 [0.35], day 5: 0.31 [0.28], day 7: 0.35 [0.36]; P < 0.05 versus control). There was also a decrease in the number of GCaMP3+ cells after ONT (cells/mm2 [SD]; control: 2198 [453], day 3: 2224 [643], day 5: 1383 [375], day 7: 913 [178]; P < 0.05). Furthermore, the proportion of GCaMP3+ cells that responded to KA decreased after ONT (50 μM KA, 97%, 54%, 47%, and 58%; control, 3, 5, and 7 days, respectively). CONCLUSIONS. Following ONT, functional RGC responses are lost prior to the loss of RGC somata, suggesting that anatomical markers of RGCs may underestimate the extent of RGC dysfunction.
关键词： GCaMP3,retinal ganglion cells,Thy1-GCaMP3,calcium imaging,optic nerve transection
更新于2025-09-23 15:22:29
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Electrical activation of degenerated photoreceptors in blind mouse retina elicited network-mediated responses in different types of ganglion cells
摘要： Electrical (e-) stimulation is explored in schemes to rescue the vision of blind people, e.g. those affected by Retinitis Pigmentosa (RP). We e-activated subretinally the surviving degenerated photoreceptors (d-Phrs) of the rd1 mouse (RP model) and evoked visual responses in the blind retina. The e-stimulation was applied with a single platinum/iridium electrode. The d-Phrs (calcium-imaging) and ganglion cells (GC) activity (MEA-recording) were recorded in simultaneous multilayer recordings. The findings of this study confirm that the d-Phrs responded to e-stimulation and modulated the retinal network-activity. The application of blockers revealed that the synaptic interactions were dependent on voltage-gated calcium channels and mediated by the transmitters glutamate and GABA. Moreover, the gap junctions coupled networks promoted the lateral-spread of the e-evoked activity in the outer (~60 μm) and inner (~120 μm) retina. The activated GCs were identified as subtypes of the ON, OFF and ON-OFF classes. In conclusion, d-Phrs are the ideal interface partners for implants to elicit enhanced visual responses at higher temporal and spatial resolution. Furthermore, the retina’s intact circuity at the onset of complete blindness makes it a tempting target when considering the implantation of implants into young patients to provide a seamless transition from blinding to chip-aided vision.
关键词： blind retina,MEA recording,subretinal implant,gap junctions,glutamate,rd1 mouse,calcium imaging,ganglion cells,GABA,electrical stimulation,Retinitis Pigmentosa,degenerated photoreceptors
更新于2025-09-19 17:15:36
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Fast objective coupled planar illumination microscopy
摘要： Among optical imaging techniques light sheet fluorescence microscopy is one of the most attractive for capturing high-speed biological dynamics unfolding in three dimensions. The technique is potentially millions of times faster than point-scanning techniques such as two-photon microscopy. However light sheet microscopes are limited by volume scanning rate and/or camera speed. We present speed-optimized Objective Coupled Planar Illumination (OCPI) microscopy, a fast light sheet technique that avoids compromising image quality or photon efficiency. Our fast scan system supports 40 Hz imaging of 700 μm-thick volumes if camera speed is sufficient. We also address the camera speed limitation by introducing Distributed Planar Imaging (DPI), a scaleable technique that parallelizes image acquisition across cameras. Finally, we demonstrate fast calcium imaging of the larval zebrafish brain and find a heartbeat-induced artifact, removable when the imaging rate exceeds 15 Hz. These advances extend the reach of fluorescence microscopy for monitoring fast processes in large volumes.
关键词： Distributed Planar Imaging,calcium imaging,light sheet fluorescence microscopy,zebrafish brain,OCPI microscopy
更新于2025-09-16 10:30:52
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Plaid Detectors in Macaque V1 Revealed by Two-Photon Calcium Imaging
摘要： Neuronal responses to one-dimensional orientations are combined to represent two-dimensional composite patterns; this plays a key role in intermediate-level vision such as texture segmentation. However, where and how the visual cortex starts to represent composite patterns, such as a plaid consisting of two superimposing gratings of different orientations, remains neurophysiologically elusive. Psychophysical and modeling evidence has suggested the existence of early neural mechanisms specialized in plaid detection [1–6], but the responses of V1 neurons to an optimally orientated grating are actually suppressed by a superimposing grating of different orientation (i.e., cross-orientation inhibition) [7, 8]. Would some other V1 neurons be plaid detectors? Here, we used two-photon calcium imaging [9] to compare the responses of V1 super?cial-layer neurons to gratings and plaids in awake macaques. We found that many non-orientation-tuned neurons responded weakly to gratings but strongly to plaids, often with plaid orientation selectivity and cross-angle selectivity. In comparison, most (～94%) orientation-tuned neurons showed more or less cross-orientation inhibition, regardless of the relative stimulus contrasts. Only a small portion (～8%) of them showed plaid facilitation at off-peak orientations. These results suggest separate subpopulations of plaid and grating responding neurons. Because most of these plaid neurons (～95%) were insensitive to motion direction, they were plaid pattern detectors, not plaid motion detectors.
关键词： Neuronal Responses,Macaque V1,Cross-Orientation Inhibition,Plaid Detectors,Two-Photon Calcium Imaging
更新于2025-09-16 10:30:52
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In Vivo Calcium Imaging of Lateral-line Hair Cells in Larval Zebrafish
摘要： Sensory hair cells are mechanoreceptors found in the inner ear that are required for hearing and balance. Hair cells are activated in response to sensory stimuli that mechanically deflect apical protrusions called hair bundles. Deflection opens mechanotransduction (MET) channels in hair bundles, leading to an influx of cations, including calcium. This cation influx depolarizes the cell and opens voltage-gated calcium channels located basally at the hair-cell presynapse. In mammals, hair cells are encased in bone, and it is challenging to functionally assess these activities in vivo. In contrast, larval zebrafish are transparent and possess an externally located lateral-line organ that contains hair cells. These hair cells are functionally and structurally similar to mammalian hair cells and can be functionally assessed in vivo. This article outlines a technique that utilizes a genetically encoded calcium indicator (GECI), GCaMP6s, to measure stimulus-evoked calcium signals in zebrafish lateral-line hair cells. GCaMP6s can be used, along with confocal imaging, to measure in vivo calcium signals at the apex and base of lateral-line hair cells. These signals provide a real-time, quantifiable readout of both mechanosensation- and presynapse-dependent calcium activities within these hair cells. These calcium signals also provide important functional information regarding how hair cells detect and transmit sensory stimuli. Overall, this technique generates useful data about relative changes in calcium activity in vivo. It is less well-suited for quantification of the absolute magnitude of calcium changes. This in vivo technique is sensitive to motion artifacts. A reasonable amount of practice and skill are required for proper positioning, immobilization, and stimulation of larvae. Ultimately, when properly executed, the protocol outlined in this article provides a powerful way to collect valuable information about the activity of hair-cells in their natural, fully integrated states within a live animal.
关键词： sensory neuroscience,Issue 141,calcium imaging,GCaMP,Neuroscience,genetically encoded indicators,hair cells,in vivo imaging,Zebrafish,lateral line,confocal imaging
更新于2025-09-04 15:30:14