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Background Tissue-clearing techniques have recently been developed to make tissues transparent for three-dimensional (3D) imaging at different scales, including single-cell resolution. However, current tissue-clearing workflows have several disadvantages, including complex protocols, time-consuming application, and fluorescence quenching. Additionally, they can be used mainly for clearing larger-volume samples, preventing wide and easy applicability in conventional experimental approaches. In this study, we aimed to develop a versatile, fast, and convenient method for clearing thin and semi-thick samples, which can be used for three-dimensional imaging of experimental or even clinical samples. Results We developed an alkaline solution (AKS) containing a combination of 2,2′-thiodiethanol (TDE), DMSO, D-sorbitol, and Tris for tissue clearing, as the alkaline environment is suitable for maintaining the fluorescence of most commonly used fluorescence protein GFP and its variants, and tested its clearing effect on samples from mice and human brains. We assessed the clearing speed, the preservation of fluorescence protein and dyes, and the imaging depth and quality. The results showed that AKS treatment rapidly cleared 300-μm-thick brain slices and 1-mm-thick slices from different organs within 5 min and 1 h, respectively. Moreover, AKS was compatible with a variety of fluorescence proteins and dyes. Most importantly, AKS enhanced the fluorescence of YFP, in contrast to the majority of existing tissue-clearing methods which reduce the fluorescence intensity of fluorescent proteins. Using AKS, we performed long-time high-resolution imaging of weak fluorescent protein-labelled tissues, long-distance fibre tracking, larger-scale 3D imaging and cell counting of the entire brain area, neural circuit tracing, 3D neuromorphic reconstruction, and 3D histopathology imaging. Conclusions AKS can be used for simple and rapid clearing of samples from mice and human brains and is widely compatible with a variety of fluorescent dyes. Therefore, AKS has great potential to be used as a broad tissue-clearing reagent for biological optical imaging, especially for time-sensitive experiments.

Clinical reports suggest a potential link between excess retinoids and development of depression. Although it has been shown that all-trans retinoic acid (ATRA) administration induces behavioral changes, further insight into how ATRA is involved is lacking. The hippocampus seems to be a major target of retinoids, and abnormal synaptic plasticity of the hippocampus is involved in depression. We examined two genes associated with synaptic function, discs large homolog 2 (DLG2), and synapse differentiation-inducing gene protein 1 (SynDIG1) in terms of hippocampal expression and correlation with behavior. Three different doses of ATRA were injected into young mice and 10 mg/kg ATRA was found to induce depression-like behavior. In the hippocampus, DLG2 mRNA was significantly decreased by ATRA. mRNA levels were positively correlated with central area duration and distance in the open-field test. Increased SynDIG1 mRNA levels were observed. There was a negative correlation between SynDIG1 mRNA levels and mobility time in the forced swimming test. Retinoic acid receptor γ mRNA was significantly positively correlated with DLG2 and negatively correlated with SynDIG1. To summarize, ATRA administration induced anxiety- and depression-like behavior accompanied by a decreased expression of DLG2 and an increased expression of SynDIG1. Moreover, DLG2 was correlated with anxiety-like behavior and SynDIG1 was correlated with depression-like behavior. These results might constitute a novel target underlying ATRA-induced anxiety- and depression-like behavior.

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Obtaining fine neuron morphology and connections data is extraordinarily useful in understanding the brain’s functionality. Golgi staining is a widely used method for revealing neuronal morphology. However, Golgi-Cox-stained tissue is difficult to image in three dimensions and lacks cell-type specificity, limiting its use in neuronal circuit studies. Here, we describe an expansion-based method for rapidly clearing Golgi-Cox-stained tissue. The results show that 1 mm thick Golgi-Cox-stained tissue can be cleared within 6 hours with a well preserved Golgi-Cox-stained signal. At the same time, we found for the first time that the cleared Golgi-Cox-stained samples were compatible with three-dimensional (3D) immunostaining and multi-round immunostaining. By combining the Golgi-Cox staining with tissue clearing and immunostaining, Golgi-Cox-stained tissue could be used for large-volume 3D imaging, identification of cell types of Golgi-Cox-stained cells, and reconstruction of the neural circuits at dendritic spines level. More importantly, these methods could also be applied to samples from human brains, providing a tool for analyzing the neuronal circuit of the human brain.

The stainless steel lined clad pipe (SSLCP) can be widely used in different fields. Industrial SSLCP are generally manufactured by the cold hydraulic expansion (CHE) method which requires high hydraulic pressure and good sealing property. The inter-layer bonding force is difficult to obtain by the CHE when there is little difference between the yield strengths of inner and outer pipes. Furthermore, the hydraulic expansion process nearly can not be achieved when the yield strength of inner pipe is higher than that of outer pipe. The thermal hydraulic expansion (THE) method can overcome these difficulties. In this paper, the graph method was adopted to analyze the THE principles for "inner strong and outer weak (ISOW)" and "outer strong and inner weak (OSIW)" pipes. The effective hydraulic expansion criterion conditions of SSLCPs were proposed and can be used as basis of clad pipe material matching and forming process selection. Through the stress and strain analysis of inner and outer pipes during THE process, the deformation coordination conditions of elastic deformation and thermal deformation of the inner and outer pipe were established. The correlation between the residual contact pressure p*c, the hydraulic expansion pressure pi and the effective temperature difference ?Te were derived. The calculation formula of the maximum expansion pressure pimax and the minimum expansion pressure pimin was obtained. The maximum heating temperature of outer pipe was also derived. Furthermore, the finite element analysis (FEA) method was adopted and simulated results verify the feasibility and applicability of the theory study.