312 research outputs found
Neocortex in the Spotlight: Concepts, Questions, and Methods
No full textWhen mammals such as mice, cats, monkeys, or humans act in the world, they continually make behaviorally relevant decisions based on perceived sensory information and memorized experiences and they constantly adapt to outside challenges through learning. These cognitive capabilities largely arise from neural processing in the outermost thin sheet of the forebrain called the neocortex. Although the mammalian neocortex has been studied extensively, the astounding complexity of both its structure and dynamics has precluded a comprehensive understanding of its function so far. Higher cortical function emerges from the interplay of myriads of diverse neocortical cells, organized across multiple hierarchical levels from local neuronal networks (“microcircuits”) to communicating brain regions (“macrocircuits”). It remains elusive how these neural circuits operate—assisted by glial networks and fuelled by the vascular system—to generate intelligent behavior and ensure adequate learning. Advances in experimental methodology are essential to further unravel cortical function and in this book we highlight the rapid recent progress in optical methods for measuring and controlling neocortical dynamics, complementing classic electrophysiological approaches. In this chapter we provide a brief overview of the functional organization of the neocortex, its tissue constituents, and current concepts of neocortical dynamics. In preparation of subsequent chapters, we summarize the manifold ways photons can be used to study neocortical function, utilizing specially designed molecular tools and various imaging technologies. We conclude with a brief future outlook. Putting neocortex literally “into the spotlight” may help uncover its intriguing mysteries
Centripetal integration of past events in hippocampal astrocytes regulated by locus coeruleus
No full text<h2>Raw data of astrocytic calcium imaging</h2>
<p>Example raw data of calcium imaging of astrocytes in hippocampus of behaving mice. The example includes recording segments with spontaneous behavior and behavior-evoked astrocytic activity, as well as recording segments with optogenetic stimulation of the locus coeruleus with opto-evoked responses. The main goal of the dataset is to convey an impression of raw data quality.</p>
<h3>Raw calcium imaging data</h3>
<p>The raw calcium imaging data are in the folder 2023-05-11. Each subfolder contains data from three simultaneously acquired imaging planes as .tif files. Between recording segments, a short break was made due to limitations of the data acquisition software. Recordings have already been motion-corrected using non-rigid motion correction. Layer 1 was recorded typically around the pyramidal layer or more ventrally; Layer 2 was typically recorded at the pyramidal layer or more dorsally; Layer 3 (analyzed for the manuscript) was recorded in the <em>stratum oriens</em> of CA1. The extracted average activity across planes is saved as a .mat file in each subfolder for convenient inspection. </p>
<h3>Behavior videos</h3>
<p>Behavior videos are saved in the main folder as *.avi files. Their sequence corresponds to the calcium imaging subfolders. The behavioral videos are used to extract face/paw motion as well as pupil diameter as described in the associated manuscript.</p>
<h3>Recorded behavioral events</h3>
<p>Additional behavioral or experimental events are save in *.mat files (to be opened with MATLAB or Python's scipy.io module). These metadata include the locomotion of the animal (<em>time_running </em>and <em>running</em>), the times of sugar rewards (<em>reward_times</em>) and, when applicable, the timing and properties of optogenetic stimulation (<em>opto_times </em>and <em>opto_protocols</em>).</p>
<h3>Relative timing of behavioral and calcium data</h3>
<p>Calcium imaging data were recorded at a different framerate and with a small temporal offset compared to the behavioral videos and the behaviorally recorded events. This raw data example mainly serves to illustrate raw data quality and therefore does not include scripts to synchronize behavioral to calcium imaging data. If such a synchronization is necessary to judge the data, please get in touch with the corresponding authors (<span>[email protected]</span>).</p>
Dendritic Branch-constrained N-Methyl-d-Aspartate Receptor-mediated Spikes Drive Synaptic Plasticity in Hippocampal CA3 Pyramidal Cells
Full text linkN-methyl-d-aspartate receptor-mediated ( spikes can be causally linked to the induction of synaptic long-term potentiation (LTP) in hippocampal and cortical pyramidal cells. However, it is unclear if they regulate plasticity at a local or global scale in the dendritic tree. Here, we used dendritic patch-clamp recordings and calcium imaging to investigate the integrative properties of single dendrites of hippocampal CA3 cells. We show that local hyperpolarization of a single dendritic segment prevents NMDA spikes, their associated calcium transients, as well as LTP in a branch-specific manner. This result provides direct, causal evidence that the single dendritic branch can operate as a functional unit in regulating CA3 pyramidal cell plasticity
Little strokes fill big oaks: a simple in vivo stain of brain cells
No full textHigh-resolution functional imaging of neural activity in vivo relies on appropriate labeling methods. In this issue of Neuron, Nagayama et al. introduce a simple procedure for staining subsets of neurons with organic calcium indicator dyes via local electroporation. Neuronal populations are sparsely labeled, preserving the ability to resolve calcium signals in dendrites and synaptic structures
Calcium indicator loading of neurons using single-cell electroporation
Full text linkStudies of subcellular Ca(2+) signaling rely on methods for labeling cells with fluorescent Ca(2+) indicator dyes. In this study, we demonstrate the use of single-cell electroporation for Ca(2+) indicator loading of individual neurons and small neuronal networks in rat neocortex in vitro and in vivo. Brief voltage pulses were delivered through glass pipettes positioned close to target cells. This approach resulted in reliable and rapid (within seconds) loading of somata and subsequent complete labeling of dendritic and axonal arborizations. By using simultaneous whole-cell recordings in brain slices, we directly addressed the effect of electroporation on neurons. Cell viability was high (about 85%) with recovery from the membrane permeabilization occurring within a minute. Electrical properties of recovered cells were indistinguishable before and after electroporation. In addition, Ca(2+) transients with normal appearance could be evoked in dendrites, spines, and axonal boutons of electroporated cells. Using negative-stains of somata, targeted single-cell electroporation was equally applicable in vivo. We conclude that electroporation is a simple approach that permits Ca(2+) indicator loading of multiple cells with low background staining within a short amount of time, which makes it especially well suited for functional imaging of subcellular Ca(2+) dynamics in small neuronal networks
Image_1_iDISCO+ for the Study of Neuroimmune Architecture of the Rat Auditory Brainstem.TIF
Full text linkThe lower stations of the auditory system display a complex anatomy. The inner ear labyrinth is composed of several interconnecting membranous structures encased in cavities of the temporal bone, and the cerebellopontine angle contains fragile structures such as meningeal folds, the choroid plexus (CP), and highly variable vascular formations. For this reason, most histological studies of the auditory system have either focused on the inner ear or the CNS by physically detaching the temporal bone from the brainstem. However, several studies of neuroimmune interactions have pinpointed the importance of structures such as meninges and CP; in the auditory system, an immune function has also been suggested for inner ear structures such as the endolymphatic duct (ED) and sac. All these structures are thin, fragile, and have complex 3D shapes. In order to study the immune cell populations located on these structures and their relevance to the inner ear and auditory brainstem in health and disease, we obtained a clarified-decalcified preparation of the rat hindbrain still attached to the intact temporal bone. This preparation may be immunolabeled using a clearing protocol (based on iDISCO+) to show location and functional state of immune cells. The observed macrophage distribution suggests the presence of CP-mediated communication pathways between the inner ear and the cochlear nuclei.</p
Chronic imaging of cortical sensory map dynamics using a genetically encoded calcium indicator
No full textAbstract In vivo optical imaging can reveal the dynamics of large-scale cortical activity, but methods for chronic recording are limited. Here we present a technique for long-term investigation of cortical map dynamics using wide-field ratiometric fluorescence imaging of the genetically encoded calcium indicator (GECI) Yellow Cameleon 3.60. We find that wide-field GECI signals report sensory-evoked activity in anaesthetized mouse somatosensory cortex with high sensitivity and spatiotemporal precision, and furthermore, can be measured repeatedly in separate imaging sessions over multiple weeks. This method opens new possibilities for the longitudinal study of stability and plasticity of cortical sensory representations
Brain-wide microstrokes affect the stability of memory circuits in the hippocampus
No full textCognitive deficits affect over 70% of stroke survivors, yet the mechanisms by which multiple small ischemic events contribute to cognitive decline remain poorly understood. In this study, we employed chronic two-photon calcium imaging to longitudinally track the fate of individual neurons in the hippocampus of mice navigating a virtual reality environment, both before and after inducing brain-wide microstrokes. Our findings reveal that, under normal conditions, hippocampal neurons exhibit varying degrees of stability in their spatial memory coding. However, microstrokes disrupted this functional network architecture, leading to cognitive impairments. Notably, the preservation of stable coding place cells, along with the stability, precision, and persistence of the hippocampal network, was strongly predictive of cognitive outcomes. Mice with more synchronously active place cells near important locations demonstrated recovery from cognitive impairment. This study uncovers critical cellular responses and network alterations following brain injury, providing a foundation for novel therapeutic strategies preventing cognitive decline
3D laser-scanning techniques for two-photon calcium imaging of neural network dynamics in vivo
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