Synapses are a fundamental unit of computation in the brain. Far from being passive connections between spiking neurons, synapses display striking short-term dynamics, undergo long-term changes in strength, and sculpt network-level processes in a complex manner. These synaptic dynamics, both in time and across space, may be a fundamental determinant of population-level computations and behavioral output of the brain, yet their role in neuromodulatory circuits is relatively under-explored. First, I developed and validated a set of likelihood-based inference tools to quantify the dynamics of synaptic ensemble composition throughout development. Second, I examined network computations in the serotonergic dorsal raphe nucleus through a dynamical lens, exploring the role of short-term synaptic dynamics at sparse recurrent connections, and of distinct long-range synaptic inputs, in shaping the output of spiking populations. 1. Simulation-based inference of synaptic ensembles. Functional features of synapses are typically inferred by sampling small ensembles of synapses, yet it is unclear if such subsamples exhibit biases. I developed a statistical framework to address this question, using it to demonstrate that common bulk electrical stimulation methods for characterizing the fraction of silent synapses exhibit high bias and variance, and using typical sample sizes, possess insufficient statistical power for accurate inference. I developed and validated a novel synthetic likelihood-based inference approach based on a simulator of the underlying experimental methodology. This new estimator, made available in an object-oriented Python toolbox, reduces bias and variance compared to previously reported methods, and provides a scalable method for examining synaptic dynamics throughout development. These tools were validated by targeted recording from hippocampal CA1 neurons in juvenile mice, where they reveal fundamental tradeoffs between release probability, number of synapses sampled, and statistical power. 2. Synaptic dynamics and population computations in the serotonin system. This part is comprised of two manuscripts. First, in the dorsal raphe nucleus, I uncovered slow, inhibitory recurrent interactions between serotonin neurons that are generated by local serotonin release. These connections were probabilistic, displayed striking short-term facilitation, gated the spiking output of serotonin neurons, and could be activated by long-range excitatory input from lateral habenula, representing threat signals. Targeted physiology and modeling revealed that these recurrent short-term facilitation features generated paradoxical excitation-driven inhibition in response to high-frequency habenula input. These facilitation rules additionally supported winner-take-all dynamics at the population level, providing a contrastive operation between functionally distinct serotonergic ensembles. Behaviorally, activating long-range lateral habenula input to dorsal raphe nucleus generated a transient, frequency-dependent suppression of reward anticipation consistent with these recurrent dynamics, without modulating the underlying reward association itself. These dynamics, we suggest, support sharp behavioral state transitions in changing environments. In a second manuscript, I explored the multiplexing of distinct long-range inputs in serotonergic circuits through spike synchrony. I demonstrated that a population of serotonergic neurons receives input from both lateral habenula and prefrontal cortex. These inputs produced similar subthreshold events, but prefrontal cortex triggered spikes with much higher latencies, supporting a population synchrony code for input identity. These input-specific spike timing patterns could be read out by simple linear decoders with high accuracy, suggesting they could be demultiplexed by downstream circuits receiving sparse innervation by serotonergic axons. We uncovered a novel intracellular calcium conductance in serotonergic neurons that altered the spectral characteristics of membrane voltage in a manner sufficient to generate long-latency, power law-distributed spike times, suggesting a simple dynamical origin for the production of synchronous or asynchronous spiking. This work indicates that serotonergic circuits can multiplex distinct informational streams through population spike synchrony mechanisms. Together, these investigations reveal that the dynamics of short-term facilitation and synaptic ensemble composition can act as the fundamental substrate for flexible computation by spiking networks across the brain.

Although insult of the developing noradrenergic neuronal system in the brain has been associated with redistribution of noradrenergic fiber input to various target brain regions, few studies have investigated the effects of such insults on locus coeruleus cell survival. In the present study the dorsal noradrenergic bundle was transected by means of a midbrain knife cut in rats 3 days after birth, and the effects of this lesion were determined approximately 8-10 weeks later. Bymeans of an immunofluorescent histochemical procedure. it was shown that tyrosine hydroxylase-containing fibers and dopamine β-hydroxylase-containing fibers were markedly reduced in number in the neocortex and hippocampus - regions anterograde to the site of axonal transection. It was further demonstrated that the number of fluorescent fibers coursing through the dorsal bundle was similarly reduced. Sprouting of noradrenergic fibers in the brainstem and cerebellum accompanied the above alterations. When locus coeruleus cell number was determined by counting Cresyl violet-stained nucleoli in serial sections it was found that dorsal bundle transection produced a loss of 17% of the cells of the coeruleus. By dividing the counts for each nucleus into fifths, it was additionally found that approximately 20-25% of those cells comprising the midportion of the nucleus, along a rostrocaudal axis, were the ones destroyed by axonal transection. These findings indicate that a neonatal lesion of the dorsal bundle produces a loss of cells in the midportion of the nucleus locus coeruleus, and that this effect is associated with noradrenergic neuronal hyperinnervation of the brainstem and cerebellum.

HRP applied to cut dorsal and ventral roots of the cat sacral spinal cord labeled afferent axons with swellings in close apposition to labeled preganglionic neurons (PGNs) in the sacral parasympathetic nucleus. Electron microscopy allowed characterization of synaptic contacts between afferents and PGNs. The results suggest that both the dorsal and ventral root afferents can directly activate autonomic preganglionic neurons.

1. Although the ORL1 receptor is clearly located within the spinal cord, the functional signalling mechanism of the ORL1 receptor in the spinal cord has not been clearly documented. The present study was then to investigate the guanine nucleotide binding protein (G-protein) activation mediated through by the ORL1 receptor in the mouse spinal cord, measuring the modulation of guanosine-5'-o-(3-[35S]-thio) triphosphate ([35S]-GTPγS) binding by the putative endogenous ligand nociceptin, also referred as orphanin FQ. We also studied the anatomical distribution of nociceptin-like immunoreactivity and nociceptin-stimulated [35S]-GTPγS autoradiography in the spinal cord. 2. Immunohistochemical staining of mouse spinal cord sections revealed a dense plexus of nociceptin-like immunoreactive fibres in the superficial layers of the dorsal horn throughout the entire length of the spinal cord. In addition, networks of fibres were seen projecting from the lateral border of the dorsal horn to the lateral grey matter and around the central canal. 3. In vitro [35S]-GTPγS autoradiography showed high levels of nociceptin-stimulated [35S]-GTPγS binding in the superficial layers of the mouse dorsal horn and around the central canal, corresponding to the areas where nociceptin-like immunoreactive fibres were concentrated. 4. In [35S]-GTPγS membrane assay, nociceptin increased [35S]-GTPγS binding of mouse spinal cord membranes in a concentration-dependent and saturable manner, affording maximal stimulation of 64.1 ± 2.4%. This effect was markedly inhibited by the specific ORL1 receptor antagonist [Phe1ψ (CH2-NH) Gly2] nociceptin (1-13) NH2. None of the μ-, δ-, and κ-opioid and other G-protein-coupled receptor antagonists had a significant effect on basal or nociceptin-stimulated [35S]-GTPγS binding. 5. These findings suggest that nociceptin-containing fibres terminate in the superficial layers of the dorsal horn and the central canal and that nociceptin released in these areas may selectively stimulate the ORL1 receptor to activate G-protein. Furthermore, the unique pattern of G-protein activation in the present study provide additional evidence that nociceptin is distinct from the μ-, δ- or κ-opioid system.

Pax3 has been used as a valuable marker in research aimed at understanding tissue interactions involved in trigeminal ophthalmic (opV) placode development. While Pax3 expression coincides with opV neuron specification, the function of Pax3 in these cells has not previously been investigated. Splotch mutant mice (which lack Pax3) have a reduced trigeminal ganglion; however it is not clear whether this reduction is due to neural crest or placode cells. We have used electroporation in the chick model system to block or ectopically express Pax3 at key times in opV placode development. Using several markers of placode cell differentiation, we have determined the experimental effects manipulating Pax3. Blocking placodal Pax3 with gene specific morpholinos resulted in a loss of migratory placode cells, and a downregulation of all opV placode markers in targeted cells. Ectopic expression of Pax3, either within the placode domain or in adjacent cranial ectoderm, resulted in the upregulation of some but not all placode markers. We conclude that opV placodal Pax3 expression is required for normal placode cell development, and hypothesize that its expression must be tightly regulated in order for placode cells to fully differentiate. The precise role of Pax3 and Pax7 in the restriction and differentiation of dorsal interneuron progenitors has been difficult to assess due to the many additional factors involved in specification and patterning of the neural tube. We have used electroporation in the chick model system to ectopically express Pax3 and Pax7 unilaterally in the neural tube. Using several markers for differentiation of ventral and dorsal neuronal progenitors, we have experimentally determined the effects of Pax3 and Pax7 ventrally and dorsally. Ectopic expression of these transcription factors in the ventral neural tube resulted in the loss of motorneurons. Though mis-expression did not qualitatively affect commissural neurons as assayed by neurofilament staining, ectopic expression of Pax3 and Pax7 in the dorsal neural tube stopped dorsal interneuron progenitors from differentiating. We conclude that Pax3 and Pax7 expression is sufficient to restrict ventral neuron identity. We also hypothesize that downregulation of these transcription factors in the dorsal neural tube is required for normal dorsal interneuron differentiation.

The relatively few serotonin (5-HT) neurons located in the Dorsal Raphe Nucleus (DRN) give rise to an extensive axonal network modulating a wide-range of brain functions and behaviors. In turn, the DRN receives inputs from several brain regions and therefore exhibits the characteristics of a hub network. While recent technological advancements have provided an unprecedented look at the neurobiology of the DRN, important knowledge gaps remain in understanding how the constellation of synaptic inputs to this region confers 5-HT neurons their unique coding features. As a first step towards characterizing the DRN's input processing strategy, we set out to explore the landscape of dendritic operation operating in DRN 5-HT neurons. Using multi-photon microscopy and in vitro electrophysical recordings, we conducted a morphological and electrophysiological survey of 5-HT neurons where we identified two structurally and morphologically distinct types of glutamatergic synapses both expressing small NMDAR-mediated conductance. Our initial findings provide valuable insights on local rules that govern how synaptic inputs to the DRN are being processed to ultimately confer 5-HT neurons their unique coding features.

There are two predominant sources of taxonomically useful morphological variability in the diverse tardigrade family Echiniscidae: the internal structure and surface sculpture of the cuticular plates covering the dorsum (sculpturing) and the arrangement and morphology of the trunk appendages (chaetotaxy). However, since the appendages often exhibit intraspecific variation (they can be reduced or can develop asymmetrically), sculpturing has been considered more stable at the species level and descriptions of new echiniscid species based solely on morphology are still being published. Here, we present a case study in which a detailed analysis of the morphology and multiple genetic markers of several species of the genus Viridiscus shows that cuticular sculpture may also exhibit considerable intraspecific variation and lead to false taxonomic conclusions. In a population collected from the eastern Nearctic, in the type locality of the recently described species V. miraviridis, individuals with transitional morphotypes between those reported for V. viridissimus and V. miraviridis were found. Importantly, all morphotypes within the viridissimus-miraviridis spectrum were grouped in a single monospecific clade according to rapidly evolving markers (ITS-1, ITS-2 and COI). Given the morphological and genetic evidence, we establish V. miraviridis as a junior synonym of V. viridissimus. This study explicitly demonstrates that a lack of DNA data associated with morphological descriptions of new taxa jeopardizes the efforts to unclutter tardigrade systematics. Additionally, V. perviridis and V. viridissimus are reported from Lâm Đồng Province in southern Vietnam, which considerably broadens their known geographic ranges.

Mémoire numérisé par la Direction des bibliothèques de l'Université de Montréal.

The nine amino acid peptide vasopressin acts as a neurohormone in the periphery and a neurotransmitter/neuromodulator in the central nervous system. Historically, research on vasopressin neurons and their projections to the pituitary has helped lay the groundwork for our understanding of peptidergic neurotransmission. Currently, our research on central vasopressin projections is driving a revolution in our understanding of social behavior. Vasopressin affects a number of social behaviors from social memory to aggression to affiliative behavior, such as pair-bonding. Further, with the addition of more and more transgenic mouse models of disease states, anxiety and depression related disorders, and social behavior dysfunction, it is important now more than ever to have a clear knowledge of the mouse vasopressin system, which derives from a number of distinct nuclei within the brain. Here, I map out vasopressin immunoreactivity in the mouse brain, and delineate the subset of brain regions with gonadal steroid hormone-dependent vasopressin immunoreactivity. Such projections are thought to derive from the bed nuclei of the stria terminalis and medial amygdala in the telencephalon. Finally, based on data from mice with lesions of the suprachiasmatic nucleus, I outline the subset of regions that likely receive vasopressin from this source. Our research on the anatomy of the vasopressin system of mice and our attempts to delineate the site of origin of the many vasopressin fibers found throughout the brain suggest that a significant amount of the vasopressin innervation deriving from cells in the bed nuclei of the stria terminalis and medial amygdala project to areas in the midbrain involved in serotonin and dopamine transmission, such as the dorsal raphe and ventral tegmental area. These transmitter systems play a crucial role in the control of anxiety and depression levels as well as motivated behavior and emotional regulation. Our results strongly suggest that a direct link exists between these systems, and future plans include an examination of this possibility. It is our hope that this work will further our understanding of the role of vasopressin and other transmitter systems in the regulation of social behaviors.

The cardiovascular system of Drosophila melanogaster consists of a cardiac tube composed of myogenic cardiomyocytes and associating non-contractile pericardial cells, pumping hemolymph into the open circulatory system. The cardiac tube, known as the dorsal vessel, is embedded in a highly regulated extracellular matrix environment, required to maintain cellular integrity and cardiac function. After embryogenesis, the dorsal vessel undergoes extensive physiological changes, relying on the extracellular matrix to adapt and remodel accordingly. Three extracellular matrix proteins are investigated throughout this thesis: Type IV Collagen, Laminin and Pericardin. Due to their localization, morphology, and role in early development, the pericardial cells are candidate cells responsible for dorsal vessel extracellular matrix deposition and regulation throughout post-embryonic growth. Using immunofluorescence techniques in combination with confocal microscopy, I characterize the association between pericardial cells and extracellular matrix proteins, and quantify extracellular matrix protein deposition at the dorsal vessel throughout post-embryonic development. Gene knock-down experiments assess pericardial cell contribution to extracellular matrix synthesis and deposition at the dorsal vessel in third instar larva. Moreover, I quantify extracellular matrix protein deposition at the dorsal vessel in the absence of pericardial cells. These data suggests that pericardial cells regulate extracellular matrix protein deposition, localization and contribute to proper cardiac morphology in post-embryonic development. / Thesis / Master of Science (MSc)