Extracting structural and functional features of widely distributed biological circuits with single cell resolution via tissue clearing and delivery vectors

The scientific community has learned a great deal from imaging small and naturally transparent organisms such as nematodes and zebrafish. The consequences of genetic mutations on their organ development and survival can be visualized easily and with high-throughput at the organism-wide scale. In contrast, three-dimensional information is less accessible in mammalian subjects because the heterogeneity of light-scattering tissue elements renders their organs opaque. Likewise, genetically labeling desired circuits across mammalian bodies is prohibitively slow and costly via the transgenic route. Emerging breakthroughs in viral vector engineering, genome editing tools, and tissue clearing can render larger opaque organisms genetically tractable and transparent for whole-organ cell phenotyping, tract tracing and imaging at depth

Extracting structural and functional features of widely distributed biological circuits with single cell resolution via tissue clearing and delivery vectors

The scientific community has learned a great deal from imaging small and naturally transparent organisms such as nematodes and zebrafish. The consequences of genetic mutations on their organ development and survival can be visualized easily and with high-throughput at the organism-wide scale. In contrast, three-dimensional information is less accessible in mammalian subjects because the heterogeneity of light-scattering tissue elements renders their organs opaque. Likewise, genetically labeling desired circuits across mammalian bodies is prohibitively slow and costly via the transgenic route. Emerging breakthroughs in viral vector engineering, genome editing tools, and tissue clearing can render larger opaque organisms genetically tractable and transparent for whole-organ cell phenotyping, tract tracing and imaging at depth

Hydrogel-Tissue Chemistry: Principles and Applications

Over the past five years, a rapidly developing experimental approach has enabled high-resolution and high-content information retrieval from intact multicellular animal (metazoan) systems. New chemical and physical forms are created in the hydrogel-tissue chemistry process, and the retention and retrieval of crucial phenotypic information regarding constituent cells and molecules (and their joint interrelationships) are thereby enabled. For example, rich data sets defining both single-cell-resolution gene expression and single-cell-resolution activity during behavior can now be collected while still preserving information on three-dimensional positioning and/or brain-wide wiring of those very same neurons—even within vertebrate brains. This new approach and its variants, as applied to neuroscience, are beginning to illuminate the fundamental cellular and chemical representations of sensation, cognition, and action. More generally, reimagining metazoans as metareactants—or positionally defined three-dimensional graphs of constituent chemicals made available for ongoing functionalization, transformation, and readout—is stimulating innovation across biology and medicine

Hydrogel-Tissue Chemistry: Principles and Applications

Over the past five years, a rapidly developing experimental approach has enabled high-resolution and high-content information retrieval from intact multicellular animal (metazoan) systems. New chemical and physical forms are created in the hydrogel-tissue chemistry process, and the retention and retrieval of crucial phenotypic information regarding constituent cells and molecules (and their joint interrelationships) are thereby enabled. For example, rich data sets defining both single-cell-resolution gene expression and single-cell-resolution activity during behavior can now be collected while still preserving information on three-dimensional positioning and/or brain-wide wiring of those very same neurons—even within vertebrate brains. This new approach and its variants, as applied to neuroscience, are beginning to illuminate the fundamental cellular and chemical representations of sensation, cognition, and action. More generally, reimagining metazoans as metareactants—or positionally defined three-dimensional graphs of constituent chemicals made available for ongoing functionalization, transformation, and readout—is stimulating innovation across biology and medicine

Dorsal Raphe Dopamine Neurons Modulate Arousal and Promote Wakefulness by Salient Stimuli

Ventral midbrain dopamine (DA) is unambiguously involved in motivation and behavioral arousal, yet the contributions of other DA populations to these processes are poorly understood. Here, we demonstrate that the dorsal raphe nucleus DA neurons are critical modulators of behavioral arousal and sleep-wake patterning. Using simultaneous fiber photometry and polysomnography, we observed time-delineated dorsal raphe nucleus dopaminergic (DRNDA) activity upon exposure to arousal-evoking salient cues, irrespective of their hedonic valence. We also observed broader fluctuations of DRNDA activity across sleep-wake cycles with highest activity during wakefulness. Both endogenous DRNDA activity and optogenetically driven DRNDA activity were associated with waking from sleep, with DA signal strength predictive of wake duration. Conversely, chemogenetic inhibition opposed wakefulness and promoted NREM sleep, even in the face of salient stimuli. Therefore, the DRNDA population is a critical contributor to wake-promoting pathways and is capable of modulating sleep-wake states according to the outside environment, wherein the perception of salient stimuli prompts vigilance and arousal

Cholinergic Mesopontine Signals Govern Locomotion and Reward through Dissociable Midbrain Pathways

The mesopontine tegmentum, including the pedunculopontine and laterodorsal tegmental nuclei (PPN and LDT), provides major cholinergic inputs to midbrain and regulates locomotion and reward. To delineate the underlying projection-specific circuit mechanisms, we employed optogenetics to control mesopontine cholinergic neurons at somata and at divergent projections within distinct midbrain areas. Bidirectional manipulation of PPN cholinergic cell bodies exerted opposing effects on locomotor behavior and reinforcement learning. These motor and reward effects were separable via limiting photostimulation to PPN cholinergic terminals in the ventral substantia nigra pars compacta (vSNc) or to the ventral tegmental area (VTA), respectively. LDT cholinergic neurons also form connections with vSNc and VTA neurons; however, although photo-excitation of LDT cholinergic terminals in the VTA caused positive reinforcement, LDT-to-vSNc modulation did not alter locomotion or reward. Therefore, the selective targeting of projection-specific mesopontine cholinergic pathways may offer increased benefit in treating movement and addiction disorders

Single-Cell Phenotyping within Transparent Intact Tissue through Whole-Body Clearing

Understanding the structure-function relationships at cellular, circuit, and organ-wide scale requires 3D anatomical and phenotypical maps, currently unavailable for many organs across species. At the root of this knowledge gap is the absence of a method that enables whole-organ imaging. Herein, we present techniques for tissue clearing in which whole organs and bodies are rendered macromolecule-permeable and optically transparent, thereby exposing their cellular structure with intact connectivity. We describe PACT (passive clarity technique), a protocol for passive tissue clearing and immunostaining of intact organs; RIMS (refractive index matching solution), a mounting media for imaging thick tissue; and PARS (perfusion-assisted agent release in situ), a method for whole-body clearing and immunolabeling. We show that in rodents PACT, RIMS, and PARS are compatible with endogenous-fluorescence, immunohistochemistry, RNA single-molecule FISH, long-term storage, and microscopy with cellular and subcellular resolution. These methods are applicable for high-resolution, high-content mapping and phenotyping of normal and pathological elements within intact organs and bodies

Whole-Brain Analysis of Cells and Circuits by Tissue Clearing and Light-Sheet Microscopy

In this photo essay, we present a sampling of technologies from laboratories at the forefront of whole-brain clearing and imaging for high-resolution analysis of cell populations and neuronal circuits. The data presented here were provided for the eponymous Mini-Symposium presented at the Society for Neuroscience's 2018 annual meeting

Optimisation of the ActWELL lifestyle intervention programme for women attending routine NHS breast screening clinics

Acknowledgements The authors would like to acknowledge the assistance of Amy Hickman, and Eluned Hughes from Breast Cancer Now, for guidance on practical intervention perspectives, and Jill Hampton in manuscript preparation. Thanks also to all the members of our public advisory group. The Health Services Research Unit, University of Aberdeen, receives core funding from Scottish Government Chief Scientist Office. Funding This work was supported by The Scottish Government, grant number BC/Screening/17/01. The funders provided independent referee reports, which guided some of the study parameters (as described in the text). The funders have read this manuscript. In-kind support was given by Breast Cancer Now for facilitating this study.Peer reviewedPublisher PD

Developing core principles for sharing Information about potential intervention benefits and harms in patient information leaflets using a modified Delphi Survey

Patients need to be informed about potential risks of taking part in clinical trials. A problem is that there are no standards telling researchers or ethics committees how these risks need to be communicated. Our background research suggests that the way harms are communicated can actually increase the risks of the harms occurring. Over half of our sample of 250,000 patients who took placebo pills (like sugar pills) in clinical trials reported some negative side effect (like pain or nausea but also more serious things). The aim of this study is to understand what information about trial harms and benefits stakeholders consider to be important for ‘principled participant information leaflets’ or ‘PrinciPILs’ to contain. The stakeholders will include patients, ethics committee members, industry representatives, medico-legal experts, psychologists, and trial managers