Previously we reported that neural stem cells (NSCs) embedded in fibrin matrices containing 9 growth factors and an anti-apoptotic agent, survived and completely filled sites of spinal cord injury. In the current study, we examine whether the number of factors can be reduced while optimizing NSC survival and filling of the lesion site. NSCs derived from embryonic day 14 F344 rat spinal cord (expressing green fluorescent protein, GFP) were embedded in fibrin matrices containing a defined growth factor cocktail (1 to 4 factors among BDNF, bFGF, VEGF and calpain inhibitor, or original 10 factor combination). Grafts were made into a C5 lateral hemisection of wild- type adult F344 rats two weeks post-injury (N=3-6 per group, total 9 groups). Graft survival was assessed 2 weeks post-grafting. A 4 factor cocktail resulted in graft survival, neuronal differentiation, and filling of lesion site that was equivalent or superior to the 10 factor cocktail. The use of fewer than 4 growth factors, including single growth factors, also frequently but less consistently resulted in NSC survival and fill of lesion site. The effect of fewer growth factors on axon extension from the graft into the host cord is currently under analysis. Collectively, these findings suggest that excellent neural stem cell engraftment and survival can be achieved with a reduced growth factor cocktail, enhancing clinical practicality

Lensless imaging provides opportunities to design imaging systems free from the constraints imposed by traditional camera architectures. Thanks to advances in imaging hardware, fabrication techniques, and new algorithms, researchers have recently developed lensless imaging systems that are extremely compact, lightweight or able to image higher-dimensional quantities. Here we review these recent advances and describe the design principles and their effects that one should consider when developing and using lensless imaging systems.

Hydra vulgaris is an emerging model organism for neuroscience due to its small size, transparency, genetic tractability, and regenerative nervous system; however, fundamental properties of its sensorimotor behaviors remain unknown. Here, we use microfluidic devices combined with fluorescent calcium imaging and surgical resectioning to study how the diffuse nervous system coordinates Hydra's mechanosensory response. Mechanical stimuli cause animals to contract, and we find this response relies on at least two distinct networks of neurons in the oral and aboral regions of the animal. Different activity patterns arise in these networks depending on whether the animal is contracting spontaneously or contracting in response to mechanical stimulation. Together, these findings improve our understanding of how Hydra's diffuse nervous system coordinates sensorimotor behaviors. These insights help reveal how sensory information is processed in an animal with a diffuse, radially symmetric neural architecture unlike the dense, bilaterally symmetric nervous systems found in most model organisms.

Understanding how neural circuits are regenerated following injury is a fundamental question in neuroscience. Hydra is a powerful model for studying this process because it has a simple neural circuit structure, significant and reproducible regenerative abilities, and established methods for creating transgenics with cell-type-specific expression. While Hydra is a long-standing model for regeneration and development, little is known about how neural activity and behavior is restored following significant injury. In this study, we ask if regenerating neurons terminally differentiate prior to reforming functional neural circuits, or if neural circuits regenerate first and then guide the constituent naive cells toward their terminal fate. To address this question, we developed a dual-expression transgenic Hydra line that expresses a cell-type-specific red fluorescent protein (tdTomato) in ec5 peduncle neurons, and a calcium indicator (GCaMP7s) in all neurons. With this transgenic line, we can simultaneously record neural activity and track the reappearance of the terminally-differentiated ec5 neurons. Using SCAPE (Swept Confocally Aligned Planar Excitation) microscopy, we monitored both calcium activity and expression of tdTomato-positive neurons in 3D with single-cell resolution during regeneration of Hydra's aboral end. The synchronized neural activity associated with a regenerated neural circuit was observed approximately 4 to 8 hours after expression of tdTomato in ec5 neurons. These data suggest that regenerating ec5 neurons undergo terminal differentiation prior to re-establishing their functional role in the nervous system. The combination of dynamic imaging of neural activity and gene expression during regeneration make Hydra a powerful model system for understanding the key molecular and functional processes involved in neural regeneration following injury.

Axon regeneration after spinal cord injury (SCI) is attenuated by growth inhibitory molecules associated with myelin. We report that rat myelin stimulated the growth of axons emerging from rat neural progenitor cells (NPCs) transplanted into sites of SCI in adult rat recipients. When plated on a myelin substrate, neurite outgrowth from rat NPCs and from human induced pluripotent stem cell (iPSC)-derived neural stem cells (NSCs) was enhanced threefold. In vivo, rat NPCs and human iPSC-derived NSCs extended greater numbers of axons through adult central nervous system white matter than through gray matter and preferentially associated with rat host myelin. Mechanistic investigations excluded Nogo receptor signaling as a mediator of stem cell-derived axon growth in response to myelin. Transcriptomic screens of rodent NPCs identified the cell adhesion molecule neuronal growth regulator 1 (Negr1) as one mediator of permissive axon-myelin interactions. The stimulatory effect of myelin-associated proteins on rodent NPCs was developmentally regulated and involved direct activation of the extracellular signal-regulated kinase (ERK). The stimulatory effects of myelin on NPC/NSC axon outgrowth should be investigated further and could potentially be exploited for neural repair after SCI.