Braitenberg Vehicles as Developmental Neurosimulation

The connection between brain and behavior is a longstanding issue in the areas of behavioral science, artificial intelligence, and neurobiology. Particularly in artificial intelligence research, behavior is generated by a black box approximating the brain. As is standard among models of artificial and biological neural networks, an analogue of the fully mature brain is presented as a blank slate. This model generates outputs and behaviors from a priori associations, yet this does not consider the realities of biological development and developmental learning. Our purpose is to model the development of an artificial organism that exhibits complex behaviors. We will introduce our approach, which is to use Braitenberg Vehicles (BVs) to model the development of an artificial nervous system. The resulting developmental BVs will generate behaviors that range from stimulus responses to group behavior that resembles collective motion. Next, we will situate this work in the domain of artificial brain networks. Then we will focus on broader themes such as embodied cognition, feedback, and emergence. Our perspective will then be exemplified by three software instantiations that demonstrate how a BV-genetic algorithm hybrid model, multisensory Hebbian learning model, and multi-agent approaches can be used to approach BV development. We introduce use cases such as optimized spatial cognition (vehicle-genetic algorithm hybrid model), hinges connecting behavioral and neural models (multisensory Hebbian learning model), and cumulative classification (multi-agent approaches). In conclusion, we will revisit concepts related to our approach and how they might guide future development.Comment: 32 pages, 8 figures, 2 table

On the role of molecular mechanisms and unequal cleavage during neurogenesis in the C. elegans C lineage

Required for neurogenesis is a family of evolutionarily conserved bHLH transcription factors known as proneural genes. However, regulation of their initial expression remains a poorly understood aspect of neurodevelopment in any model, particularly Caenorhabditis elegans. A key mechanism by which cells acquire different fates is asymmetric division and in neuronal lineages these often generate unequally sized daughters. Whether this unequal size directly affects cell fate regulation is often unknown. Indeed, the question of how control of cell size intersects with fate decisions is poorly understood in biology more generally. Taking advantage of the single-cell resolution provided by the invariant cell lineage of C. elegans, I interrogate these two fundamental biological questions in the C lineage. Expression of the proneural gene hlh-14/Ascl1 in a single branch of the lineage is required for neurogenesis of the DVC and PVR neurons and is immediately preceded by unequal cleavages. Addressing both molecular and cellular regulators I perform a 4D-lineage based genetic screen for upstream regulators of hlh-14/Ascl1 and address the effect of unequal cleavage and daughter cell size. I find that a regulator of other neuronal lineage cleavages, PIG-1/MELK, is also required in the C lineage, yet equalisation does not affect the initiation of hlh-14/Ascl1 expression. Conversely, I demonstrate that unequal cleavage and acquisition of neuronal fate in separate successive divisions are controlled by the same key regulators. The first by an upstream regulator of hlh-14, the Mediator complex kinase module let-19/Mdt-13 and the second by hlh-14 itself. Taken together the results described in this thesis suggest that rather than acting to correctly segregate initial proneural gene expression, unequal cleavages are instead co-regulated by the same factors regulating neuronal fate acquisition. This co-regulation at successive divisions thus coordinates two separable aspects of fate; acquisition of neuronal identity and correct post-mitotic embryonic cell size

Stress-Dependent Regulation Of A Major Node Of The Insulin-Like Peptide Network That Modulates Survival

Chronic stress disrupts insulin signaling, predisposing human populations to diabetes, cardiovascular disease, Alzheimer’s Disease, and other metabolic and neurological disorders, including post-traumatic disorders (PTSD). Thus, efficient recovery from stress optimizes survival. However, stress recovery in humans is difficult to study, but is much easier to dissect in model organisms. The worm genetic model Caenorhabditis elegans can switch between stressed and non-stressed states, and this switch is largely regulated by insulin signaling. Previously, the Alcedo lab proposed that insulin-like peptides (ILPs), which exist as multiple members of a protein family in both C. elegans and humans, implements a combinatorial coding strategy to control the switch between the two physiological states. The concept of combinatorial coding has led to the identification of an inter-ILP network, where one ILP, ins-6, is a major node of the network. This is consistent with ins-6 as the most pleotropic of all ILPs that have been tested. ins-6 has also been shown to be the most important ILP in promoting stress recovery in C. elegans. Because of its central role in the ILP network and in stress recovery, for my thesis I identified mechanisms through which INS-6 regulates the network and an animal’s recovery from stress. Under optimal environments, ins-6 mRNA is endogenously expressed in the cell bodies of one or two chemosensory neurons, ASI and ASJ, in the developing animal. However, upon stress-induced developmental arrest, known as dauer, ins-6 mRNA is only limited to the ASJ sensory neurons. I discovered that ins-6 mRNA from ASJ is also surprisingly transported to the axonal nerve ring bundle of stressed animals, but lost from the nerve ring after recovery from stress. Consistent with the existence of an inter-ILP network, insulin signaling regulates ins-6 mRNA transport, which also requires the activities of specific kinesins. This transport additionally depends on the untranslated regions of ins-6 mRNA, but these regions are insufficient for transport. More importantly and in collaboration with other members of the Alcedo lab, we showed that axonal ins-6 mRNA facilitates stress recovery, where high axonal ins-6 mRNA promotes faster recovery and low axonal ins-6 mRNA delays recovery. Moreover, I demonstrated the existence of axonal Golgi bodies, whose mobilization are enhanced during stress. Together my data suggest that stress stimulates the axonal transport of ILP mRNAs, which are then locally translated and packaged for secretion--a mechanism that promotes plasticity during stress and optimal stress recovery. To identify additional regulators of ins-6 mRNA, I also performed, together with other members of the lab, a forward genetic screen for mutants that alter ins-6 transcription during stress. Through whole-genome sequencing, one of the five mutants we isolated is potentially a mutation in an innexin gap junction protein. Since innexins have been shown to regulate neural activity, I tested the hypothesis that neural activity will also affect axonal ins-6 mRNA transport. Interestingly, I found that a synaptic transmission mutant, which should have low neural activity, increases axonal ins-6 mRNA and alters neurite morphology. My thesis study raises an intriguing hypothesis: stress modulates neurite activity and morphology, which in turn promote ILP mRNA transport to the axons. The axonal localization of an ILP mRNA also uncovers a novel mechanism of insulin signaling during stress. Because of the high degree of conservation between C. elegans and humans and the effects of altered insulin signaling in stressed brains, my findings should advance our understanding of how a nervous system recovers from stress. The work described in this thesis should lead to potential therapies for stress management to promote better health

Comprehensive Techniques to Study Activity-dependent and Neurogenic Mechanisms in Stem Cell-derived Neuron Models

Accurately modeling human-related neuronal phenomena remains at the forefront of neuroscience. This thesis utilizes already-established in vitro models of mouse embryonic stem cells, designing an efficient method to optogenetically stimulate neurons derived from mouse stem cells and expounds upon their scope with novel protocols to generate hippocampal neurons from human induced pluripotent stem cells. First, a novel platform for optogenetic stimulation was built and tested on mouse embryonic stem cells to demonstrate functionality of optogenetic channels in mouse embryonic stem cell-derived neurons. The device was built from 3D printed materials and validated with oscilloscopy and spectrophotometry while neurons were cultured for over 30 days in vitro and assayed first for electrical activity by electrophysiology, calcium signaling, and small molecule activation of glutamatergic receptors. When verified that both device and neurons were functional, cells were transduced with a ChannelRhodopsin variant, ChR2-eYFP-NpHR, and were stimulated over several light cycle parameters and assayed for CFOS expression. Having shown that neurons responded in an activity-dependent manner to the device, I established preliminary studies into human hippocampal embryonic neurogenesis. I derived a novel protocol to differentiate hiPSCs to hippocampal neural progenitors using small molecules and specific laminar substrates unique to the subgranular zone. Hippocampal progenitors were assayed for literature-established genetic markers including WNT7b, WNT8a, PROX1, FOXG1, and ZBTB20, and then allowed to spontaneously differentiate into neurons expressing canonical neural, synaptic, glutamatergic, and constitutive hippocampal markers. These cells were expanded over 200 days in vitro. When allowed to spontaneously differentiate or forced to differentiate under NOTCH inhibition, neuronal cultures sustained ZBTB20 and FOXG1 coexpression over the terminal differentiation path though cultures at ~200 days old did not differentiate at the same rate as cultures from ~30 days. When transplanted in vivo, human hippocampal progenitors differentiated fully after 4 months, projected toward the CA3 from the dentate gyrus, and established synaptic connections with host neurons identified by staining synaptic markers. In conclusion, several novel findings are demonstrated throughout this thesis, though the most pertinent include: 1.) mESC-derived neurons may be optogenetically stimulated by ergonomic device fabrication. 2.) Sustained or adult neurogenesis is dependent on the laminin isoform expressed in the subgranular zone. 3.) Hippocampal progenitors from human induced pluripotent stem cells behave like neurons and can be optogenetically targeted and are transplantable in vivo hippocampus in which they integrate into pre-existing hippocampal networks. Future investigations include merging activity-dependent Tau phosphorylation in mESC- and hiPSC-derived human hippocampal neurons and transplantation of human hippocampa

Genetic control of photoreceptor terminal differentiation in Drosophila melanogaster

Why do photoreceptors differentiate in the eye? Though simple, biologically this is an important question, and it may prove complex to answer. To present a bigger picture: animals have evolved a diversity of highly specialised sensory organs, which they use to obtain information from their environment and thus survive. These organs contain different types of receptor neurons. For example, there are chemoreceptors in the labellum and in the antennae of insects, or mechanoreceptors in the inner ear of vertebrates... and each of these types of receptor neurons specifically possesses the molecular machinery to detect and transduce stimuli from one particular sensory modality. In the case of the eye, it contains photoreceptor neurons, which are specialised in light detection. Neither photoreceptors nor most of the components of the phototransduction cascade appear commonly outside the eye. Therefore, what are mechanisms that ensure that photoreceptors differentiate correctly in the eye, and not in other body parts? To start answering this question, first it might be useful to understand the early-acting process of eye field specification. This depends on a group of transcription factors that are collectively called the ‘retinal determination network’ (RDN), and work in combination with each other to confer eye identity to the developing, multipotent tissue. RDN genes are both necessary and sufficient for eye formation in different animal species, from Drosophila to vertebrates, and they tend to act through an evolutionarily conserved sequence of transcriptional events. First in this sequence, following the Drosophila nomenclature, the transcription factor Eyeless activates the expression of sine oculis and eyes absent. Then, Sine oculis and Eyes absent form a heterodimer and direct eye formation. Despite the importance of the RDN, until recently, little was known about its targets, or about the molecular mechanisms by which it coordinates eye development. In particular, how does it instruct photoreceptor differentiation? Our work suggests that a key step in this process is coordinated by the zinc finger transcription factor glass, which is a direct target of Sine oculis. While previous literature has shown that the Glass protein is primarily expressed in photoreceptors, its role in these cells was not known because it was believed that glass mutant photoreceptor precursors died during metamorphosis. Contrary to former studies, we demonstrate that glass mutant photoreceptor precursors survive and are present in the adult retina, but fail to mature as functional photoreceptors. Importantly, we have found that Glass is required for the expression of virtually all the proteins that are involved in the phototransduction cascade, and thus glass mutant flies are blind. Consistent with this, ectopic expression of Glass is able to induce some phototransduction components in the brain. Another step in the formation of photoreceptors is regulated by the homeodomain transcription factor Hazy, which is a direct target of Glass. While we show that both Glass and Hazy act synergistically to induce the expression of phototransduction proteins, we have also found that Glass can initiate the expression of most of the components of the phototransduction machinery in a Hazy-independent manner, and that hazy mutant flies only fail to detect white light after they are older than five days. Glass seems to be both required and sufficient for the expression of Hazy, and inducing Hazy in the retina partly rescues the glass mutant phenotype. Taken together, our results show a transcriptional link between the RDN and the expression of the proteins that adult Drosophila photoreceptors need to sense light, placing Glass at a key position in this developmental process. Finally, we compare the expression pattern of Glass in Drosophila and in the annelid Platynereis, and discuss the possibility that Glass plays an evolutionarily conserved role across different phyla.Warum bilden sich Fotorezeptoren gerade im Auge aus? Obwohl diese Frage einfach erscheint, ist sie aus biologischer Sicht doch sehr bedeutend und bedarf eventuell einer komplexen Antwort. Allgemein lässt sich sagen, dass Tiere eine Vielfalt von hoch spezialisierten Sinnesorganen entwickelt haben, durch die sie Informationen aus ihrer Umwelt aufnehmen und auf diese Weise ihr Überleben sichern. Diese Organe enthalten verschiedene Arten von Rezeptorneuronen. Zum Beispiel gibt es Chemorezeptoren im Labellum und in den Antennen der Insekten, oder Mechanorezeptoren im Innenohr von Wirbeltieren... und jedes dieser Rezeptorneuronen besitzt eine spezifische molekulare Maschinerie, um Reize einer bestimmten Sinnesmodalität wahrzunehmen und umzuwandeln. Beim Auge sind es Fotorezeptorneuronen, die auf die Wahrnehmung von Lichtreizen spezialisiert sind. Weder die Fotorezeptoren noch die meisten der Komponenten der Fototransduktionskaskade kommen außerhalb des Auges vor. Welche Mechanismen sind demzufolge ausschlaggebend, damit sich Fotorezeptoren im Auge und nicht in anderen Körperteilen entwickeln? Um diese Frage zu beantworten, ist es zunächst wichtig die frühen Mechanismen der Augenspezifizierung zu verstehen. Diese erfolgt unter Einfluss einer Gruppe von Transkriptionsfaktoren, die als „Retinales Determinations Netzwerk“ (RDN) bezeichnet werden. Diese Transkriptionsfaktoren interagieren, um aus dem sich entwickelnden multipotenten Gewebe ein Sehorgan zu bilden. RDN-Gene sind für die Augenentwicklung verschiedener Tierarten, von Drosophila bis zu Wirbeltieren, sowohl notwendig als auch ausreichend. Sie agieren durch eine evolutionär konservierte Sequenz transkriptioneller Mechanismen. An erster Stelle dieser Sequenz, nach der Drosophila Nomenklatur, aktiviert der Transkriptionsfaktor Eyeless die Expression von sine oculis und eyes absent. Anschließend bilden Sine Oculis und Eyes absent ein Heterodimer und induzieren die Entwicklung des Auges. Trotz der Bedeutung des RDNs war bis vor Kurzem nur sehr wenig über seinen Zweck oder die molekularen Mechanismen durch die es die Augenentwicklung koordiniert, bekannt. Vor allem stellt sich die Frage, wie es die Differenzierung der Fotorezeptoren reguliert? Unsere Arbeit legt nahe, dass ein wesentlicher Schritt in diesem Prozess durch den Zinkfinger-Transkriptionsfaktor glass koordiniert wird. Dabei handelt es sich um ein direktes Zielgen von Sine oculis. Obwohl in früheren wissenschaftlichen Arbeiten belegt wurde, dass das Glass-Protein in erster Linie in Fotorezeptoren exprimiert wird, war seine Rolle in diesen Zellen nicht bekannt, da angenommen wurde, dass Fotorezeptoren von glass Mutanten während der Metamorphose absterben. Im Gegensatz zu früheren Studien belegen wir das Überleben der Fotorezeptor-Vorläuferzellen von glass Mutanten und ihre Präsenz in der Retina adulter Fliegen, wobei sie jedoch nicht zu funktionsfähigen Fotorezeptoren heranreifen. Insbesondere konnten wir zeigen, dass Glass für die Expression fast aller Proteine, die in der Fototransduktionskaskade involviert sind, erforderlich ist. Daher sind glass Mutanten blind. In Übereinstimmung mit diesen Erkenntnissen bewirkt die ektopische Expression von Glass die Induktion einiger Komponenten der Fototransduktion im Gehirn. Ein weiterer Schritt in der Bildung von Fotorezeptoren wird reguliert durch den Homeodomänen-Transkriptionsfaktor Hazy, der ein direktes Ziel von Glass ist. Wir zeigen zum einen die synergetische Wirkung von Glass und Hazy bei der Expression von Fototransduktionsproteinen, zum anderen belegen wir, dass Glass die meisten Komponenten der Fototransduktionsmaschinerie unabhängig von Hazy induzieren kann, und dass hazy Mutanten ab dem Alter von fünf Tagen weißes Licht nicht mehr wahrnehmen können. Glass scheint notwendig und ausreichend für die Expression von Hazy zu sein und die Induktion von Hazy in der Retina rettet teilweise den Phänotyp von glass Mutanten. Insgesamt beweisen unsere Ergebnisse einen transkriptionellen Zusammenhang zwischen dem RDN und der Expression von Proteinen, die in Fotorezeptoren von adulten Drosophila Fliegen notwendig sind um Licht wahrzunehmen. Bei diesem Entwicklungsprozess hat Glass eine Schlüsselposition. Schließlich vergleichen wir die Expressionsmuster von Glass in Drosophila und im Anneliden Platynereis und diskutieren die Möglichkeit, dass Glass eine evolutionär konservierte Rolle über verschiedene Phyla hinweg spielt