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Biallelic pathogenic variants in PLPBP (formerly called PROSC) have recently been shown to cause a novel form of vitamin B6-dependent epilepsy, the pathophysiological basis of which is poorly understood. When left untreated, the disease can progress to status epilepticus and death in infancy. Here we present 12 previously undescribed patients and six novel pathogenic variants in PLPBP. Suspected clinical diagnoses prior to identification of PLPBP variants included mitochondrial encephalopathy (two patients), folinic acid-responsive epilepsy (one patient) and a movement disorder compatible with AADC deficiency (one patient). The encoded protein, PLPHP is believed to be crucial for B6 homeostasis. We modelled the pathogenicity of the variants and developed a clinical severity scoring system. The most severe phenotypes were associated with variants leading to loss of function of PLPBP or significantly affecting protein stability/PLP-binding. To explore the pathophysiology of this disease further, we developed the first zebrafish model of PLPHP deficiency using CRISPR/Cas9. Our model recapitulates the disease, with plpbp(-/-) larvae showing behavioural, biochemical, and electrophysiological signs of seizure activity by 10 days post-fertilization and early death by 16 days post-fertilization. Treatment with pyridoxine significantly improved the epileptic phenotype and extended lifespan in plpbp(-/-) animals. Larvae had disruptions in amino acid metabolism as well as GABA and catecholamine biosynthesis, indicating impairment of PLP-dependent enzymatic activities. Using mass spectrometry, we observed significant B6 vitamer level changes in plpbp(-/-) zebrafish, patient fibroblasts and PLPHP-deficient HEK293 cells. Additional studies in human cells and yeast provide the first empirical evidence that PLPHP is localized in mitochondria and may play a role in mitochondrial metabolism. These models provide new insights into disease mechanisms and can serve as a platform for drug discovery.

It has long been established that the spinal cord is able to produce locomotor activity on its own. Despite extensive research identifying and describing the involvement of multiple spinal neuron populations that are part of the spinal locomotor circuit, the manner in which these different components act together to precisely control the rhythm and the pattern of activation of muscles during locomotion remains largely undetermined. We sought to shed light on how the components of spinal locomotor circuits interact to produce robust locomotion using a developmental approach in zebrafish larvae. We used electrophysiological techniques to observe how the rhythm generation mechanism developed while the fish was transitioning from an early form of swimming to a more mature swimming behaviour. In the process we were able to highlight fundamental changes in the organization of spinal locomotor circuits as its operation moves from a pacemaker-based architecture relying on intrinsic properties of neurons to a network oscillator-based architecture relying on synaptic connectivity to generate proper rhythm driving the fish tail beats. Additionally, we revealed that this transition occurred at different times along the spinal cord progressing in a caudorostral direction. By combining these experimental observations with already published insights we were able to propose models of spinal locomotor circuits reproducing the successive locomotor behaviours encountered through development. By incrementing supplementing the circuit model in a manner that reflected biological processes by which the nervous system maturates (neurogenesis, synaptic connectivity refinement and maturation of intrinsic properties) we mirrored the natural development of the spinal locomotor circuit. This series of successively constructed models permitted us to pinpoint possible roles of specific neural populations for swimming behaviour as well as eventual targets and mechanism of actions of neuromodulators (serotonin and dopamine). In the process, I further provided testable hypotheses for future inquiries. Overall, the experimental findings in combination with the modeling work are an important step forward in fully understanding how the spinal cord generates swimming movements in zebrafish.

This thesis develops a method to optimize the inventory-related performance of a VMI (Vendor Managed Inventory) in retailing environments, for two successive products with overlapping lifecycles, under stock-dependent demand. The study refers to existing research in different fields, such as inventory policies and VMIs, retailing of substitutable products, stock-dependent demands and product lifecycle management. However, no study encompassing all these research streams could be found, and thus an abductive approach is developed, aiming at building new theory. The thesis draws upon a case study at Sony Germany, which has started an ambitious VMI initiative with the largest retailer of consumer electronics products in Germany. First, stock and sales data available, along with insights collected among employees, is used to understand the key profit-drivers when lifecycles of the two products overlap, and a set of indicators is propounded to measure them. The pattern of sales in relation to inventories of the two products is studied, and results in the validation of the stock-dependent demand assumption, with demand following a Poisson distribution of parameter λ [on-hand stock]. Demand for the new product is also found to be negatively affected by stock levels of the old product, in an exponential way. Second, a model is built and a scenario- based simulation embedding the patterns previously established is performed to evaluate different strategies to steer the stock levels in the VMI. Scenarios are evaluated against the proposed set of indicators, but no scenario is found to over perform consistently the others. Nevertheless interesting patterns emerge and it is possible to relate the performance observed to specificities of scenarios and product contexts. Using the patterns identified, a set of guidelines is suggested, along with proposals for practical implementation.