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Proceedings of the National Academy of... Jan 2020Tuberous Sclerosis Complex (TSC) is a rare genetic disease that manifests with early symptoms, including cortical malformations, childhood epilepsy, and TSC-associated...
Tuberous Sclerosis Complex (TSC) is a rare genetic disease that manifests with early symptoms, including cortical malformations, childhood epilepsy, and TSC-associated neuropsychiatric disorders (TANDs). Cortical malformations arise during embryonic development and have been linked to childhood epilepsy before, but the underlying mechanisms of this relationship remain insufficiently understood. Zebrafish have emerged as a convenient model to study elementary neurodevelopment; however, without in-depth functional analysis, the Tsc2-deficient zebrafish line cannot be used for studies of TANDs or new drug screening. In this study, we found that the lack of Tsc2 in zebrafish resulted in heterotopias and hyperactivation of the mTorC1 pathway in pallial regions, which are homologous to the mammalian cortex. We observed commissural thinning that was responsible for brain dysconnectivity, recapitulating TSC pathology in human patients. The lack of Tsc2 also delayed axonal development and caused aberrant tract fasciculation, corresponding to the abnormal expression of genes involved in axon navigation. The mutants underwent epileptogenesis that resulted in nonmotor seizures and exhibited increased anxiety-like behavior. We further mapped discrete parameters of locomotor activity to epilepsy-like and anxiety-like behaviors, which were rescued by reducing tyrosine receptor kinase B (TrkB) signaling. Moreover, in contrast to treatment with vigabatrin and rapamycin, TrkB inhibition rescued brain dysconnectivity and anxiety-like behavior. These data reveal that commissural thinning results in the aberrant regulation of anxiety, providing a mechanistic link between brain anatomy and human TANDs. Our findings also implicate TrkB signaling in the complex pathology of TSC and reveal a therapeutic target.
Topics: Animals; Anxiety; Disease Models, Animal; Epilepsy; Female; Humans; Intracellular Signaling Peptides and Proteins; Mechanistic Target of Rapamycin Complex 1; Receptor, trkB; Seizures; Tuberous Sclerosis; Zebrafish; Zebrafish Proteins
PubMed: 31932427
DOI: 10.1073/pnas.1910834117 -
International Journal of Molecular... Dec 2019The G protein-coupled cannabinoid receptors type 1 (CB1R) and type 2 (CB2R), and their endocannabinoid (eCBs) ligands, have been implicated in several aspects of brain...
The G protein-coupled cannabinoid receptors type 1 (CB1R) and type 2 (CB2R), and their endocannabinoid (eCBs) ligands, have been implicated in several aspects of brain wiring during development. Here we aim to assess whether interfering with CB1R affects development, neuritogenesis and pathfinding of GnRH and AgRP neurons, forebrain neurons that control respectively reproduction and appetite. We pharmacologically and genetically interfered with CB1R in zebrafish strains with fluorescently labeled GnRH3 and the AgRP1 neurons. By applying CB1R antagonists we observed a reduced number of GnRH3 neurons, fiber misrouting and altered fasciculation. Similar phenotypes were observed by CB1R knockdown. Interfering with CB1R also resulted in a reduced number, misrouting and poor fasciculation of the AgRP1 neuron's axonal projections. Using a bioinformatic approach followed by qPCR validation, we have attempted to link CB1R functions with known guidance and fasciculation proteins. The search identified stathmin-2, a protein controlling microtubule dynamics, previously demonstrated to be coexpressed with CB1R and now shown to be downregulated upon interference with CB1R in zebrafish. Together, these results raise the likely possibility that embryonic exposure to low doses of CB1R-interfering compounds could impact on the development of the neuroendocrine systems controlling sexual maturation, reproduction and food intake.
Topics: Agouti-Related Protein; Animals; Axons; Benzoxazines; Embryo, Nonmammalian; Embryonic Development; Gonadotropin-Releasing Hormone; Morpholines; Morpholinos; Naphthalenes; Neurons; Pyrrolidonecarboxylic Acid; Receptor, Cannabinoid, CB1; Zebrafish; Zebrafish Proteins
PubMed: 31881740
DOI: 10.3390/ijms21010168 -
Trends in Biochemical Sciences Jan 2020Netrin is a prototypical axon guidance cue. Structural studies have revealed how netrin interacts with the deleted in colorectal cancer (DCC) receptor, other receptors,... (Review)
Review
Netrin is a prototypical axon guidance cue. Structural studies have revealed how netrin interacts with the deleted in colorectal cancer (DCC) receptor, other receptors, and co-factors for signaling. Recently, genetic studies suggested that netrin is involved in neuronal haptotaxis, which requires a reversible adhesion process. Structural data indicate that netrin can also mediate trans-adhesion between apposing cells decorated with its receptors on the condition that the auxiliary guidance cue draxin is present. Here, we propose that netrin is involved in conditional adhesion, a reversible and localized process that can contribute to cell adhesion and migration. We suggest that netrin-mediated adhesion and signaling are linked, and that local environmental factors in the ventricular zone, the floor plate, or other tissues coordinate its function.
Topics: Animals; Cell Adhesion; DCC Receptor; Humans; Netrins; Signal Transduction
PubMed: 31704057
DOI: 10.1016/j.tibs.2019.10.005 -
Journal of Tissue Engineering and... Nov 2019The central feature of peripheral motor axons is their remarkable lengths as they project from a motor neuron residing in the spinal cord to distant target muscle....
The central feature of peripheral motor axons is their remarkable lengths as they project from a motor neuron residing in the spinal cord to distant target muscle. However, current in vitro models have not replicated this feature owing to challenges in generating motor axon tracts beyond a few millimeters in length. To address this, we have developed a novel combination of microtissue engineering and mechanically assisted growth techniques to create long-projecting centimeter-scale motor axon tracts. Here, primary motor neurons were isolated from rat spinal cords and induced to form engineered microspheres via forced aggregation in custom microwells. This technique yielded healthy motor neurons projecting dense, fasciculated axonal tracts. Within our custom-built mechanobioreactors, motor neuron culture conditions, neuronal/axonal architecture, and mechanical growth conditions were optimized to generate parameters for robust and efficient stretch growth of motor axons. We found that axons projecting from motor neuron aggregates were able to tolerate displacement rates at least 10 times greater than those by axons projecting from dissociated motor neurons. The growth and structural characteristics of these stretch-grown motor axons were compared with that of benchmark stretch-grown sensory axons, revealing increased motor axon fasciculation. Finally, motor axons were integrated with myocytes and stretch grown to create novel long-projecting axonal-myocyte constructs that recreate characteristic dimensions of native nerve-muscle anatomy. This is the first demonstration of mechanical elongation of spinal motor axons and may have applications as anatomically inspired in vitro testbeds or as tissue-engineered living scaffolds for targeted axon tract reconstruction following nervous system injury or disease.
Topics: Animals; Axons; Bioreactors; Motor Neurons; Rats; Rats, Sprague-Dawley
PubMed: 31469944
DOI: 10.1002/term.2955