archives@tulane.edu / Mycobacterium tuberculosis (Mtb) remains the single largest infectious disease killer of man worldwide. The non-human primate (NHP) model, including the Indian Rhesus macaque, is particularly valuable for the study of this disease because they fully recapitulate the pathological and immunological responses, can be co-infected with Simian Immunodeficiency Virus to model lentivirus synergism, and provide ideal candidates to study novel vaccine and drug development. However, while much has been elucidated over the past centuries in regards to host immunity, bacterial responses, and granuloma formation, little remains known about histomorphologic differences between active tuberculosis (ATBI) and latent tuberculosis (ATBI) disease states. Differentiation between these disease states, in humans or in NHPs, is based on clinical parameters, and there are currently no established methods for detecting morphologic differences between these conditions at the microscopic level. The aim of this study was to develop and validate a novel approach for assessment of pulmonary pathology in Rhesus macaques experimentally-infected with M. tuberculosis alone or in the setting of SIV co-infection. Archival lung samples from experimentally-infected macaques were assessed by blinded pathologists to determine differences in a series of pathological parameters based on previous experiments. Interobserver agreement and repeatability was good between pathologists. Significant differences were observed in several pathology categories, with ATBI animals having a greater likelihood of increased alveolar macropahges, type II pneumocyte hyperplasia, perivasculitis, vasculitis/lymphangitis, and consolidation in comparison to LTBI animals. SIV co-infection increased the likelihood of perivasculitis and lymphangitis/vasculitis in both ATBI and LTBI animals. SIV co-infection also increased alveolar macrophages and type II pneumocyte hyperplasia in LTBI animals. Immunofluorescence was used to confirm the presence of Mtb bacilli within the perivascular inflammation. A similar grading system approach was used in 2 additional studies examining reactivation of ATBI in the setting of SIV coinfection unrelated to CD4+ T cell depletion and to evaluate pulmonary pathology changes in the setting of the use of an attenuated vaccine in SIV co-infected animals with similarly significant results. This grading scheme provides a valuable and desperately needed adjunctive assessment tool for evaluation of pulmonary pathology changes in the NHP model of pulmonary tuberculosis. / 1 / Denae N. LoBato

archives@tulane.edu / The goal of this project was to examine patterns of craniofacial ontogeny in three species of baboons, Papio anubis (n = 55), P. cynocephalus (n = 43), and P. ursinus (n = 42), and three species of macaques, Macaca. cyclopis (n = 34), M. fascicularis (n = 55), and M. mulatta (n = 59) to determine the degree to which they exhibited similarities and differences in ontogeny, and then to apply those findings to Neandertals (n = 12) and modern humans (n = 42) to better understand ontogenetic variation within Homo. Macaques and baboons were chosen as model species because, like Neandertals and modern humans, they have relatively large geographic ranges, and this study aimed to investigate whether that had any impact on shifts in ontogeny. First, virtual 3D models of each individual were created in Agisoft Photoscan. Then 3D coordinates from 39 type I and type II landmarks representing the entire cranium from each individual were collected in Stratovan Checkpoint. Missing landmarks were estimated in R and statistical analyses were conducted in Paleontological Statistics (PAST). The results indicate that Neandertals and modern humans share parallel postnatal growth trajectories, which is in agreement with existing literature. However, there was some indication of non-parallel trajectories among baboons and macaques. Specifically, in some, though not all, aspects of shape, P. anubis and P. ursinus have divergent trajectories, as does M. cyclopis compared to both M. fascicularis and M. mulatta. One possible explanation for these differences lies in their geographic ranges. Although there is interspecific overlap in baboon ranges and baboons are known to hybridize, P. anubis and P. ursinus ranges do not touch. Similarly, although there is overlap in the ranges of M. fascicularis and M. mulatta and evidence of interbreeding between the two species, M. cyclopis is isolated on the island of Taiwan. Thus, while additional data, especially in terms of larger juvenile sample sizes are needed to confirm this pattern, the results of this study suggest possible subtle divergences in ontogeny of species whose ranges do not overlap and who therefore do not hybridize in their natural environments. / 1 / Whitney Karriger

Primates, including humans, explore their visual environment with sequences of gaze fixations interrupted by saccadic eye movements that re-orient the fovea to objects of interest. This visual behavior is thought to involve two separate processes. First, the current foveal image is analyzed and the next object of interest is selected as a saccade target. Second, previously examined objects are retained to prevent their re-examination. Visual behavior has been studied successfully using the visual search paradigm, in which subjects locate a unique target stimulus from amongst multiple distracting stimuli. Models of visual search posit that the process of saccade target selection is guided by a visual salience map. This map receives both stimulus-driven and goal-directed inputs to form representations of visual objects, and a competition between those representations is played out to determine the next saccade target. Neurophysiological studies using nonhuman primates have suggested that the salience map is distributed across a network of brain areas that includes the midbrain superior colliculus (SC). These studies, however, have not ruled out the possibility that selective activity for a saccade target may instead be related to the preparation of the saccade. Moreover, not much is known about the selection of a saccade target beyond the first in a sequence of gaze fixations. Finally, the mechanisms underlying the process of saccade target retention are not well understood. In this thesis, I will investigate the role of the primate SC in visual behavior by recording the activity of single neurons while monkeys perform visual search tasks. The major findings will describe 1) how SC sensory-motor neurons instantiate the visual salience map; 2) how this salience map is dynamically updated so that saccade targets are retained; and 3) how multiple representations on this salience map are processed in parallel for saccade target selection. Given SC’s role in the control of visual behavior and its position within the network involved in cognitive processes, these findings have important implications for our understanding of the neural basis of human cognition and of its dysfunctions in disease states. / Thesis (Ph.D, Neuroscience Studies) -- Queen's University, 2010-12-22 09:52:20.143

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Cerebral blood flow dynamics is an essential component for preserving cerebral integrity. Cerebral blood flow abnormalities are often seen in patients with central nervous system pathologies such as epilepsy, migraine, Alzheimer's Disease, vascular dementia, stroke, and even HIV/AIDS. There is increasing clinical and experimental evidence implicating cerebral hypoperfusion during ageing. The determination of cerebral perfusion has therefore become an important objective in physiological, pathological, pharmacological, and clinical investigations. The knowledge of regional cerebral blood flow further provides useful diagnostic information and/or data for a better understanding of the complex clinical presentations in patients with neurological and psychiatric disorders. Several cerebrovasoactive drugs have found application in the clinical setting of cerebrovascular diseases such as migraine and dementia. Due to the similarities between humans and non-human primates with respect to their brains, both structurally and behaviourally, numerous studies have been conducted and several non-human primate models have been developed for physiological, pathological, pharmacological, and clinical studies, amongst others in Parkinson's disease and diabetes. The relatively large size of the Cape baboon Papio Ursinus with a weight of 27-30 kg for a large male, makes this primate especially suitable for in vivo brain studies using radiotracers and Single Photon Emission Computed Tomography (SPECT). The main aim of the current study was therefore to develop a suitable radiotracer (99m Tc-hexamethylpropylene amine oxime (HMPAO) or 99m Tc_ethyl_cysteinatedimer (ECD) or 123l-iodoamphetamine (IMP)) for adapted in vivo cerebral blood flow measurements in a non-human primate (Papio ursinus) as an investigative model. The model was to be validated and applied in various drug studies for the evaluation of pharmacological interventions. The study design made use of split-dose methodology, whereby the radiopharmaceutical (radiotracer) was administered twice during each study. The first administration was injected soon after the induction of the anaesthesia, and was followed by the first SPECT data acquisition. The second administration of the radioligand, a double dose of radioactivity with respect to the first radioligand injection, was done at a specific time during the study, which took into account the pharmacodynamics of the drug. A second SPECT data acquisition followed subsequently. The drugs that were included in the study were acetazolamide, a carbonic acid anhydrase inhibitor (often used in nuclear medicine to determine cerebral reserve); sumaptriptan, a 5-HT (serotonin) agonist used for treatment of migraine; sodium valproate (an anti-epileptic drug); nimodipine, a calcium channel blocker and nitro-glycerine, a vasodilator used for angina. Arterial blood pressures were recorded from a catheter in the femoral artery and heart rates were concurrently monitored. The split-dose method was successfully applied to develop a non-human primate cerebral blood flow model under anaesthesia. The model showed differences in cerebral perfusion of the different anaesthesia regimes. These anaesthesia data sets were suitable as control/baseline results for drug intervention studies. Acetazolamide evaluation through the split-dose method in the baboon confirmed the sensitivity of the model by presenting comparable perfusion. This result compared to those already familiar prompted the model to be applied in pharmacological intervention studies. Subsequent results of these investigations showed increases in perfusion for single drug nimodipine treatment (25%). However, nimodipine attenuated the increases in perfusion when administered in combination with acetazolamide. Sumatriptan was able to decrease and normalise the increased perfusion after long duration anaesthesia. Decreased cerebral blood flow was observed for combinations of nimodipine with sodium valproate suggesting drug-drug interaction with important clinical implications. Similar decreases were found also for sumatriptan and nitro-glycerine when administered in combination with nimodipine. Studies with the various tracers (99m Tc_HMPAO or 99m Tc_ECD or 123l_IMP) showed clear differences in the perfusion data, confirming variation in the biochemical performance of the tracers. These differences, if not taken into consideration, caution for inappropriate clinical conclusions and subsequent erroneous therapeutic decisions. Improvement of radiotracer efficacy was subsequently attempted through application of the cyclodextrine complexation approach. Although cyciodextrine technology did not markedly improve the brain disposition of the 99m Tc-ECD, protection of the tracer against degradation was demonstrated. This study encouraged further exploration of this method for protection of the tracer against chemical and metabolic degradation. The current study was aimed to develop and effectively apply a non-human primate model with nuclear medicine technology for cerebral blood flow determinations after pharmacological interventions. This was achieved through the split-dose method and dedicated computer programming, which yielded a successful model with the non-human primate under anaesthesia. The model was validated with the application of acetazolamide to confirm familiar cerebrovascular reserve results, indicating that the model is sensitive to CBF changes. The model was also effectively applied in several pharmacological intervention studies, whereby cerebropharmacodynamics of selected drugs were investigated and established. This unique model of a non-human primate, Papio ursinus for cerebral blood flow determinations has served pharmacological research successfully during the past 12 years and could do so in the future, with scope to investigate new frontiers with improved technologies. / Thesis (Ph.D. (Pharmacology))--North-West University, Potchefstroom Campus, 2004.

Cerebral blood flow dynamics is an essential component for preserving cerebral integrity. Cerebral blood flow abnormalities are often seen in patients with central nervous system pathologies such as epilepsy, migraine, Alzheimer's Disease, vascular dementia, stroke, and even HIV/AIDS. There is increasing clinical and experimental evidence implicating cerebral hypoperfusion during ageing. The determination of cerebral perfusion has therefore become an important objective in physiological, pathological, pharmacological, and clinical investigations. The knowledge of regional cerebral blood flow further provides useful diagnostic information and/or data for a better understanding of the complex clinical presentations in patients with neurological and psychiatric disorders. Several cerebrovasoactive drugs have found application in the clinical setting of cerebrovascular diseases such as migraine and dementia. Due to the similarities between humans and non-human primates with respect to their brains, both structurally and behaviourally, numerous studies have been conducted and several non-human primate models have been developed for physiological, pathological, pharmacological, and clinical studies, amongst others in Parkinson's disease and diabetes. The relatively large size of the Cape baboon Papio Ursinus with a weight of 27-30 kg for a large male, makes this primate especially suitable for in vivo brain studies using radiotracers and Single Photon Emission Computed Tomography (SPECT). The main aim of the current study was therefore to develop a suitable radiotracer (99m Tc-hexamethylpropylene amine oxime (HMPAO) or 99m Tc_ethyl_cysteinatedimer (ECD) or 123l-iodoamphetamine (IMP)) for adapted in vivo cerebral blood flow measurements in a non-human primate (Papio ursinus) as an investigative model. The model was to be validated and applied in various drug studies for the evaluation of pharmacological interventions. The study design made use of split-dose methodology, whereby the radiopharmaceutical (radiotracer) was administered twice during each study. The first administration was injected soon after the induction of the anaesthesia, and was followed by the first SPECT data acquisition. The second administration of the radioligand, a double dose of radioactivity with respect to the first radioligand injection, was done at a specific time during the study, which took into account the pharmacodynamics of the drug. A second SPECT data acquisition followed subsequently. The drugs that were included in the study were acetazolamide, a carbonic acid anhydrase inhibitor (often used in nuclear medicine to determine cerebral reserve); sumaptriptan, a 5-HT (serotonin) agonist used for treatment of migraine; sodium valproate (an anti-epileptic drug); nimodipine, a calcium channel blocker and nitro-glycerine, a vasodilator used for angina. Arterial blood pressures were recorded from a catheter in the femoral artery and heart rates were concurrently monitored. The split-dose method was successfully applied to develop a non-human primate cerebral blood flow model under anaesthesia. The model showed differences in cerebral perfusion of the different anaesthesia regimes. These anaesthesia data sets were suitable as control/baseline results for drug intervention studies. Acetazolamide evaluation through the split-dose method in the baboon confirmed the sensitivity of the model by presenting comparable perfusion. This result compared to those already familiar prompted the model to be applied in pharmacological intervention studies. Subsequent results of these investigations showed increases in perfusion for single drug nimodipine treatment (25%). However, nimodipine attenuated the increases in perfusion when administered in combination with acetazolamide. Sumatriptan was able to decrease and normalise the increased perfusion after long duration anaesthesia. Decreased cerebral blood flow was observed for combinations of nimodipine with sodium valproate suggesting drug-drug interaction with important clinical implications. Similar decreases were found also for sumatriptan and nitro-glycerine when administered in combination with nimodipine. Studies with the various tracers (99m Tc_HMPAO or 99m Tc_ECD or 123l_IMP) showed clear differences in the perfusion data, confirming variation in the biochemical performance of the tracers. These differences, if not taken into consideration, caution for inappropriate clinical conclusions and subsequent erroneous therapeutic decisions. Improvement of radiotracer efficacy was subsequently attempted through application of the cyclodextrine complexation approach. Although cyciodextrine technology did not markedly improve the brain disposition of the 99m Tc-ECD, protection of the tracer against degradation was demonstrated. This study encouraged further exploration of this method for protection of the tracer against chemical and metabolic degradation. The current study was aimed to develop and effectively apply a non-human primate model with nuclear medicine technology for cerebral blood flow determinations after pharmacological interventions. This was achieved through the split-dose method and dedicated computer programming, which yielded a successful model with the non-human primate under anaesthesia. The model was validated with the application of acetazolamide to confirm familiar cerebrovascular reserve results, indicating that the model is sensitive to CBF changes. The model was also effectively applied in several pharmacological intervention studies, whereby cerebropharmacodynamics of selected drugs were investigated and established. This unique model of a non-human primate, Papio ursinus for cerebral blood flow determinations has served pharmacological research successfully during the past 12 years and could do so in the future, with scope to investigate new frontiers with improved technologies. / Thesis (Ph.D. (Pharmacology))--North-West University, Potchefstroom Campus, 2004.

Notre laboratoire a développé une stratégie d'isolement de fragments d'anticorps recombinants à partir de primates non humains (Macaca fascicularis) immunisés, en utilisant la technologie des phages. Dans le cadre de cette thèse, une comparaison des séquences d'anticorps de macaques (Macaca Mulatta) et d'anticorps humains a toutefois montré que les anticorps des deux espèces présentent des différences qui rendent souhaitable une étape d'humanisation des anticorps de macaques. Cette stratégie a été utilisée dans le cadre du projet Européen AntiBotABE (www.antibotabe.com) et l'étape de criblage a été adaptée pour isoler des scFv neutralisant de façon croisée les toxines botuliques BoNT/B des sous-types B1 et B2, en utilisant séquentiellement l'holotoxine BoNT/B1 et un fragment recombinant représentant la région C-terminale de la chaîne lourde de BoNT/B2. Le meilleur scFv ciblant les régions C-terminales des chaînes lourdes de BoNT/B1 et BoNT/B2, B2-7, a montré une bonne capacité de neutralisation de BoNT/B1 et BoNT/B2 dans le test ex vivo de paralysie hémidiaphragmatique. Les régions charpentes du scFv B2-7 ont un pourcentage d'identité élevé (80 %) avec leurs homologues humains. Des scFv neutralisant BoNT/A1 en ciblant sa chaîne légère ont aussi été isolés, dont le scFv le plus efficace, 2H8, induit une diminution de 50% de l'activité endopeptidasique à une concentration correspondant à un rapport molaire 2H8/BoNT/A1 de 64000. Les régions charpentes de 2H8 ont également un pourcentage d'identité élevée (88%) avec leurs homologues humains. La versatilité de cette stratégie en fait un outil permettant l'isolement de nombreux autres fragments d'anticorps à visée thérapeutique. / Our laboratory has developed a strategy to isolate recombinant antibody fragments technology from immunized non human primates (Macaca fascicularis) by phage display. In the course of the present thesis, a comparison between macaque (Macaca mulatta) and human antibody sequences has demonstrated that antibodies of the two species are different. This difference makes the humanization of macaque antibodies desirable. The strategy was used in the framework of the European AntiBotABE project, and the screening was adapted to isolate antibody fragments cross neutralizing the B1 and B2 subtypes of botulinum B neurotoxin, by using sequentially the holotoxin BoNT/B1 and a recombinant fragment representing the C-terminal region of the heavy chain of BoNTB2. The best scFv targeting the C-terminal region of BoNT/B1 and BoNTB2 heavy chains, B2-7, demonstrated a high capacity to neutralize BoNT/B1 and BoNT/B2 in the ex vivo hemidiaphragmatic assay. A high identity (80%) between the framework regions of B2-7 and their human homologs was observed. ScFvs neutralizing BoNT/A1 by targeting its light chain were also isolated and among them, the scFv 2H8 induced a decrease of 50% in the endopeptidase activity at a concentration corresponding to a molar ratio of 2H8/BoNT/A1 of 64000. A high identity (88%) between the framework regions of 2H8 and their human homologs was also observed. Our strategy can be used to isolate other therapeutic antibody fragments.

La compréhension des mécanismes impliqués dans les maladies neurodégénératives ou développementales ainsi que la mise en place de nouvelles approches thérapeutiques reposent sur l’utilisation de modèles expérimentaux pertinents et de techniques d’imagerie adaptées. Dans ce contexte, l’IRM est un outil de choix pour l’exploration anatomique in vivo dans la mesure où elle permet d’effectuer un suivi longitudinal. Le succès translationnel des thérapies du laboratoire au patient repose sur une bonne caractérisation des modèles et une continuité des biomarqueurs utilisés. Or, si l'IRM est disponible en préclinique et en clinique, les outils d'analyse sont peu « génériques ». Au cours de cette thèse, en s'inspirant des travaux menés chez l'Homme, nous avons développé et validé des outils automatiques de segmentation des structures neuroanatomiques chez le Macaque. La méthode proposée repose sur la mise en registre avec l'IRM du sujet d'un atlas digital probabiliste suivi de l'optimisation d'un modèle statistique par mélanges de gaussiennes et champs aléatoires de Markov. Elle a été validée chez un ensemble de sujets sains adultes puis mise en application dans le contexte du développement néonatal normal du cerveau. Afin de poser les bases d'une évaluation permettant une comparaison des biomarqueurs IRM avec les biomarqueurs post mortem de référence, nous avons également mis au point une chaîne de traitement permettant la reconstruction 3D de volumes histologiques du cerveau de Macaque et l'avons appliqué à la caractérisation du contraste IRM au cours d'une greffe de cellules souches après lésion excitotoxique. / Understanding the mechanisms involved in neurodegenerative or developmental diseases and designing new therapeutic approaches are based on the use of relevant experimental models as well as appropriate imaging techniques. In this context, MRI is a prominent tool for in vivo investigation as it allows for longitudinal follow-up. Successful translation from bench to bedside calls for well-characterized models as well as transferable biomarkers. Yet, despite the existence of both clinical and preclinical scanners, analysis tools are hardly translational. In this work, inspired by standards developed in Humans, we've built and validated tools for the automated segmentation of neuroanatomical structures in the Macaque. This method is based on the registration of a digital probabilistic atlas followed by the fitting of a statistical model consisting of a gaussian mixture and Markov random fields. It was first validated in healthy adults and then applied to the study of neonatal brain development. Furthermore, to pave the way for comparisons with gold standard post mortem biomarkers, we developed a pipeline for the automated 3D reconstruction of histological volumes that we applied to the characterization of MRI contrast in a stem-cell graft following an excitotoxic lesion.

The hypothalamus is a subcortical brain region whose limits and constituent nuclei lack consensus. The hypothalamus has been linked to emotion and different states of stress, providing critical feedback about the internal environment to the prefrontal cortex, a region known for executive function within the cortex of humans. An understanding of the developmental origin of the hypothalamus can provide a basis for defining which limits and nuclei are ontologically hypothalamic, and which are not, as well as a framework for understanding its connectional relationship with other brain regions. The Prosomeric Model (Rubenstein et al. 1994; Puelles and Rubenstein 2003; Nieuwenhuys and Puelles 2016; Puelles 2018) explains the embryological development of the central nervous system (CNS) shared by all vertebrates as a Bauplan. As a primary event, the early neural plate is patterned by intersecting longitudinal plates and transverse segments, forming a mosaic of progenitor units. The hypothalamus is specified by three prosomeres [hp1, hp2, and the acroterminal domain (At)] of the secondary prosencephalon with corresponding alar and basal plate parts, which develop apart from the diencephalon. Mounting evidence suggests that progenitor units within alar plate and basal plate parts of hp1 and hp2 give rise to distinct hypothalamic nuclei, which preserve their relative invariant positioning (topology) in the adult brain. Nonetheless, the principles of the Prosomeric Model have not been applied to the hypothalamus of adult primates. The Structural Model (Barbas 1986; Barbas and Rempel-Clower 1997) highlights the variation of laminar structure in the grey matter of the prefrontal cortex as a basis for predicting specific cortico-cortical connections. The areas of the prefrontal cortex vary along a spectrum by number of layers, laminar definition, and cellularity of those layers. The systematic laminar patterns of different areas of the prefrontal cortex seem to be associated with differential rates of development or maturation. A topographical analysis of bidirectional projections between the prefrontal cortex and the hypothalamus was previously applied using the Structural Model (Rempel-Clower and Barbas 1998). The authors found the prefrontal cortex has highly specific projections to the hypothalamus, originating mostly from limbic orbital and medial prefrontal areas, which have lower laminar definition than other prefrontal areas. In addition, the hypothalamus has relatively specific patterns of projection to the prefrontal cortex. We previously lacked an organizing principle to examine the specific pattern of connections between the hypothalamus and prefrontal cortex in adult rhesus monkey. In the present study, hypothalamic nuclei in the rhesus monkey (Macaca mulatta) were parcellated using classic architectonic boundaries and stains. The topological relations of hypothalamic nuclei and adjacent hypothalamic landmarks were then analyzed with homology across rodent and primate species to trace the origin of adult hypothalamic nuclei to the alar or basal plate components of hp1 and hp2. A novel atlas of the hypothalamus of the adult rhesus monkey was generated with developmental ontologies for each hypothalamic nucleus. This atlas was then applied to a topological analysis of the strength and pattern of connections between the hypothalamus and prefrontal cortex in the adult rhesus monkey. The result is a systematic reinterpretation of the adult hypothalamus of the rhesus monkey whose prosomeric ontology was used to study connections and neuraxial pathways linking the hypothalamus and prefrontal cortex. The convergence of the Prosomeric and Structural Models provides a framework through development to explain the structural patterns found in the adult primate cortex and hypothalamus, and the likely consequences of their disruption.