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Combined multi-well production method has irreplaceable advantages over single well, especially in the future industrialization of natural gas hydrate regarded as a highly efficient and clean potential alternative energy source. Researching the effectiveness of combined multi-well production contributes to optimizing production plans and well designs, thereby improving overall production efficiency. In this study, utilizing a self-developed large-size two-dimensional natural gas hydrate production simulation apparatus to conduct stepwise depressurization production experiments under three types of well, including single horizontal well, single vertical well, and combined horizontal well and vertical well. The investigation focused on the impact of well types on reservoir responses and gas production characteristics at the planar scale during depressurization production. The findings reveal that during the constant pressure stage of dual-well combined production, pressure can be maintained stable, but maintaining stability in temperature is challenging due to the comprehensive impact of factors such as the distribution of hydrates in the reactor, hydrate reformation, and heat transfer efficiency. Throughout the process of dual-well combined production, combined production effect is observed. In the early and middle stages of production, the two wells complement each other, exhibiting a synergistic effect. In the later stages of production, combined production effect manifests as optimization of gas production behavior, whereby the gas production behavior is optimized to be more conducive to further production of hydrate.

Neural progenitor cells generated from human induced pluripotent stem cells (hiPSCs) are the forefront of ″brain-on-chip″ investigations. Viable and functional hiPSC-derived neuronal networks are shaping powerful in vitromodels for evaluating the normal and abnormal formation of cortical circuits, understanding the underlying disease mechanisms, and investigating the response to drugs. They therefore represent a desirable instrument for both the scientific community and the pharmacological industry. However, culture conditions required for the full functional maturation of individual neurons and networks are still unidentified. It has been recognized that three-dimensional (3D) culture conditions can better emulate in vivoneuronal tissue development compared to 2D cultures and thus provide a more desirable in vitroapproach. In this paper, we present the design and implementation of a 3D scaffold platform that supports and promotes intricate neuronal network development. 3D scaffolds were produced through direct laser writing by two-photon polymerization (2PP), a high-resolution 3D laser microstructuring technology, using the biocompatible and nondegradable photoreactive resin Dental LT Clear (DClear). Neurons developed and interconnected on a 3D environment shaped by vertically stacked scaffold layers. The developed networks could support different cell types. Starting at the day 50 of 3D culture, neuronal progenitor cells could develop into cortical projection neurons (CNPs) of all six layers, different types of inhibitory neurons, and glia. Additionally and in contrast to 2D conditions, 3D scaffolds supported the long-term culturing of neuronal networks over the course of 120 days. Network health and functionality were probed through calcium imaging, which revealed a strong spontaneous neuronal activity that combined individual and collective events. Taken together, our results highlight advanced microstructured 3D scaffolds as a reliable platform for the 3D in vitromodeling of neuronal functions.

Efficient derivation of human cerebral neocortical neural stem cells (NSCs) and functional neurons from pluripotent stem cells (PSCs) facilitates functional studies of human cerebral cortex development, disease modeling and drug discovery. Here we provide a detailed protocol for directing the differentiation of human embryonic stem cells (hESCs) and induced pluripotent stem cells (iPSCs) to all classes of cortical projection neurons. We demonstrate an 80-d, three-stage process that recapitulates cortical development, in which human PSCs (hPSCs) first differentiate to cortical stem and progenitor cells that then generate cortical projection neurons in a stereotypical temporal order before maturing to actively fire action potentials, undergo synaptogenesis and form neural circuits in vitro. Methods to characterize cortical neuron identity and synapse formation are described.

Cultured cerebral cortex neurons generated from human Down syndrome induced pluripotent stem cells rapidly develop Alzheimer’s disease pathologies.