Engineering brain activity patterns by neuromodulator polytherapy for treatment of disorders.

Conventional drug screens and treatments often ignore the underlying complexity of brain network dysfunctions, resulting in suboptimal outcomes. Here we ask whether we can correct abnormal functional connectivity of the entire brain by identifying and combining multiple neuromodulators that perturb connectivity in complementary ways. Our approach avoids the combinatorial complexity of screening all drug combinations.

Combinatorial programming of human neuronal progenitors using magnetically-guided stoichiometric mRNA delivery.

Identification of optimal transcription factor expression patterns to direct cellular differentiation along a desired pathway presents significant challenges. We demonstrate massively combinatorial screening of temporally-varying mRNA transcription factors to direct differentiation of neural progenitor cells using a dynamically-reconfigurable magnetically-guided spotting technology for localizing mRNA, enabling experiments on millimetre size spots. In addition, we present a time-interleaved delivery method that dramatically reduces fluctuations in the delivered transcription factor copy numbers per cell.

Brain activity patterns in high-throughput electrophysiology screen predict both drug efficacies and side effects.

Neurological drugs are often associated with serious side effects, yet drug screens typically focus only on efficacy. We demonstrate a novel paradigm utilizing high-throughput in vivo electrophysiology and brain activity patterns (BAPs). A platform with high sensitivity records local field potentials (LFPs) simultaneously from many zebrafish larvae over extended periods.

Automated deep-phenotyping of the vertebrate brain.

Here, we describe an automated platform suitable for large-scale deep-phenotyping of zebrafish mutant lines, which uses optical projection tomography to rapidly image brain-specific gene expression patterns in 3D at cellular resolution. Registration algorithms and correlation analysis are then used to compare 3D expression patterns, to automatically detect all statistically significant alterations in mutants, and to map them onto a brain atlas. Automated deep-phenotyping of a mutation in the master transcriptional regulator not only detects all known phenotypes but also uncovers important novel neural deficits that were overlooked in previous studies.

Organ-targeted high-throughput in vivo biologics screen identifies materials for RNA delivery.

Therapies based on biologics involving delivery of proteins, DNA, and RNA are currently among the most promising approaches. However, although large combinatorial libraries of biologics and delivery vehicles can be readily synthesized, there are currently no means to rapidly characterize them in vivo using animal models. Here, we demonstrate high-throughput in vivo screening of biologics and delivery vehicles by automated delivery into target tissues of small vertebrates with developed organs.

Studying apoptosis in the Zebrafish.

Zebrafish (Danio rerio) have been extensively used to study apoptotic cell death during normal development and under a wide range of experimental manipulations. A number of features make zebrafish a particularly powerful model organism: (1) embryos are small in size, develop rapidly outside the mother, and are optically transparent; (2) tools are readily available for rapid knockdown and overexpression of genes; and (3) embryos can be arrayed into multiwell plates and are permeable to a wide range of drugs and small molecules. The molecular machinery underlying the intrinsic and extrinsic apoptosis pathways appears to be highly conserved between zebrafish and mammals.

High-throughput hyperdimensional vertebrate phenotyping.

Most gene mutations and biologically active molecules cause complex responses in animals that cannot be predicted by cell culture models. Yet animal studies remain too slow and their analyses are often limited to only a few readouts. Here we demonstrate high-throughput optical projection tomography with micrometre resolution and hyperdimensional screening of entire vertebrates in tens of seconds using a simple fluidic system.

The zebrafish as a model organism for the study of apoptosis.

Apoptosis plays important roles in embryogenesis, tissue homeostasis, and immune system regulation. The zebrafish (Danio rerio) is a powerful vertebrate model organism that has been extensively used to study apoptotic cell death during normal development and under conditions of cellular stress. In the past 5 years, a detailed picture has begun to emerge of the molecular underpinnings of the cell-intrinsic and the cell-extrinsic apoptosis signaling pathways in zebrafish.

The use of in vivo zebrafish assays in drug toxicity screening.

Anecdotal evidence has long suggested that zebrafish may be a good model to predict toxicity of human drugs. As summarized in this review, several groups have recently conducted systematic evaluations of zebrafish toxicity end points using large numbers of pharmacologically relevant compounds. Assays of particular interest include those for cardiotoxicity, ototoxicity, seizure liability, developmental toxicity and gastrointestinal motility.

Noggin is required for correct guidance of dorsal root ganglion axons.

Members of the bone morphogenetic protein family of secreted protein signals have been implicated as axon guidance cues for specific neurons in Caenorhabditis elegans and in mammals. We have examined axonal pathfinding in mice lacking the secreted bone morphogenetic protein antagonist Noggin. We have found defects in projection of several groups of neurons, including the initial ascending projections from the dorsal root ganglia, motor axons innervating the distal forelimb, and cranial nerve VII.

Effects of heterodimerization and proteolytic processing on Derrière and Nodal activity: implications for mesoderm induction in Xenopus.

Derrière is a recently discovered member of the TGFbeta superfamily that can induce mesoderm in explant assays and is expressed at the right time and location to mediate mesoderm induction in response to VegT during Xenopus embryogenesis. We show that the ability of Derrière to induce dorsal or ventral mesoderm depends strictly on the location of expression and that a dominant-negative Derrière cleavage mutant completely blocks all mesoderm formation when ectopically expressed. This differs from the activity of similar Xnr2 cleavage mutant constructs, which are secreted and retain signaling activity.