Optical imaging has become a powerful tool for studying brains in vivo. The opacity of adult brains makes microendoscopy, with an optical probe such as a gradient index (GRIN) lens embedded into brain tissue to provide optical relay, the method of choice for imaging neurons and neural activity in deeply buried brain structures. Incorporating a Bessel focus scanning module into two-photon fluorescence microendoscopy, we extended the excitation focus axially and improved its lateral resolution. Scanning the Bessel focus in 2D, we imaged volumes of neurons at high-throughput while resolving fine structures such as synaptic terminals. We applied this approach to the volumetric anatomical imaging of dendritic spines and axonal boutons in the mouse hippocampus, and functional imaging of GABAergic neurons in the mouse lateral hypothalamus in vivo.
Almost all data needed to evaluate the conclusions in the paper are present in the paper or the supplementary materials; Raw image data for Figs. 2, 4 & 9 are available from Dryad, 10.5061/dryad.pr4t978
- Guanghan Meng
- Yajie Liang
- Wan-chen Jiang
- Rongwen Lu
- Joshua Tate Dudman
- Na Ji
- Guanghan Meng
- Na Ji
- Sarah Sarsfield
- Yeka Aponte
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Animal experimentation: All animal experiments were conducted according to the United States National Institutes of Health guidelines for animal research. Procedures and protocols were approved by the Institutional Animal Care and Use Committee at Janelia Research Campus, Howard Hughes Medical Institute (protocol number: 16-147)
- David Kleinfeld, University of California, San Diego, United States
© 2019, Meng et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
Mathys et al. conducted the first single-nucleus RNA-seq (snRNA-seq) study of Alzheimer’s disease (AD) (Mathys et al., 2019). With bulk RNA-seq, changes in gene expression across cell types can be lost, potentially masking the differentially expressed genes (DEGs) across different cell types. Through the use of single-cell techniques, the authors benefitted from increased resolution with the potential to uncover cell type-specific DEGs in AD for the first time. However, there were limitations in both their data processing and quality control and their differential expression analysis. Here, we correct these issues and use best-practice approaches to snRNA-seq differential expression, resulting in 549 times fewer DEGs at a false discovery rate of 0.05. Thus, this study highlights the impact of quality control and differential analysis methods on the discovery of disease-associated genes and aims to refocus the AD research field away from spuriously identified genes.
The strength of a fear memory significantly influences whether it drives adaptive or maladaptive behavior in the future. Yet, how mild and strong fear memories differ in underlying biology is not well understood. We hypothesized that this distinction may not be exclusively the result of changes within specific brain regions, but rather the outcome of collective changes in connectivity across multiple regions within the neural network. To test this, rats were fear conditioned in protocols of varying intensities to generate mild or strong memories. Neuronal activation driven by recall was measured using c-fos immunohistochemistry in 12 brain regions implicated in fear learning and memory. The interregional coordinated brain activity was computed and graph-based functional networks were generated to compare how mild and strong fear memories differ at the systems level. Our results show that mild fear recall is supported by a well-connected brain network with small-world properties in which the amygdala is well-positioned to be modulated by other regions. In contrast, this connectivity is disrupted in strong fear memories and the amygdala is isolated from other regions. These findings indicate that the neural systems underlying mild and strong fear memories differ, with implications for understanding and treating disorders of fear dysregulation.