PROLIFERATION, MIGRATION, AND SURVIVAL OF CELLS IN THE TELENCEPHALON OF THE BALL PYTHON, PYTHON REGIUS

Reptiles exhibit neurogenesis throughout the brain during adulthood. However, very few studies have quantified telencephalon-wide neurogenesis in adulthood, and no studies have performed these investigations in snakes. Quantifying neurogenesis in the adult snake is essential to understanding class-wide adult neurogenesis and providing insight into the evolution of this trait. The thymidine analog 5-bromo-2’-deoxyuridine (BrdU) was used to quantify cell proliferation, migration, and survival in the ball python (Python regius). First, to determine the proper dose of BrdU for injection we subcutaneously injected 50mg/kg, 100mg/kg, and 250mg/kg into 15 adult male P. regius. We found the 250mg/kg dose marked significantly more cells than the 50mg/kg dose, but not the 100mg/kg dose. Then we subcutaneously injected 100mg/kg BrdU into 15 juvenile male P. regius at 3 different time points (2 days, 2 weeks, 2 months) prior to sacrifice to quantify proliferation, migration, and survival of cells in several telencephalic subregions. After sectioning and immunohistochemical staining, we found proliferation to be highest in the accessory olfactory bulb (AoB), retrobulbar regions (AD, AV), dorsal ventricular ridge (DVR), and dorsolateral amygdala/lateral amygdala (DLA/LA). Of the proliferating cells, the proportions of cells that migrated after 2 weeks were highest in the ventral lateral region (VL), anterior medial and lateral cortices (aMC, aLC), and anterior NS (aNS). After 2 months, the highest proportional survival was in the AoB, aLC, aMC, aNS, DVR, and ventral medial regions (VM). Regions involved in long-term functions like spatial memory may require less proliferation and longer survival, while regions involved in short-term functions undergo more proliferation with higher relative attrition

Genetic and experiential effects on dopaminergic systems

textSuccessful reproduction requires the coordination of relevant sensory inputs with motivational and motor systems primed by sex steroid hormones to produce an appropriate hierarchical sequence of movements. Both behavioral and neural phenotypes can be altered by social interactions that, in turn, can produce long term changes in cellular activity and signaling, neural circuitry, and sexual behavior. There is considerable variability in the type and direction of neural and behavioral change in response to social interactions, and the degree of plasticity may depend on intrinsic or genetic individual differences. Dopaminergic systems modulate the expression of social and sexual behaviors in a number of vertebrate species and intrinsic differences in dopaminergic systems may underlie intrinsic individual differences in the display of sexual behavior. Here, I present data on how social interactions, genotype, and steroid hormones can affect dopamine synthesis in limbic and midbrain nuclei. I investigated this in three model systems including knockout mice and two related species of whiptail lizard. The knockout mice have a targeted deletion of the progesterone receptor and display higher mount and intromission frequencies than wild-type males. Male whiptail lizards (Cnemidophorus inornatus) have natural variation in the display of courtship behaviors: some males are more sexually vigorous than others. Finally, individuals of the parthenogenetic species C. uniparens, which arose from two hybridization events involving the sexual species C. inornatus, display both maleand female-like sexual behaviors depending on reproductive state. In contrast, C. inornatus females only display receptive behavior, and this only during when preovulatory. In all three species, individuals that displayed greater levels of mounting behaviors had greater numbers of cells expressing tyrosine hydroxylase, the rate-limiting enzyme in dopamine synthesis, in the substantia nigra pars compacta. In addition, in male whiptail lizards and the related parthenogen, dopamine production in the dorsal hypothalamus was correlated with the propensity to display mounting behaviors. Dopamine can increase the display of mounting behavior in mice as well as in male and parthenogenetic whiptail lizards. My dissertation indicates that not only is dopamine sufficient to elicit mounting behavior, but differences in dopamine production may contribute to individual differences in behavioral phenotype.Biological Sciences, School o

From locomotor behavior to cerebellum evolution and development in squamate models

Locomotor behavior, the entire set of movements an individual utilizes to modify its spatial location in time, is a crucial attribute of an organism’s life. Though not responsible for movement initiation or rhythmic locomotor pattern generation, the cerebellum, an ancient and functionally conserved feature of the vertebrate brain, plays a key role in many aspects of motor performance. Variations in its morphology, relative size and cortical organization, likely resulting from divergent developmental programs, have been observed even in closely related vertebrate species, often reflecting a tight linkage between cerebellar organization and functional demands associated with ecologically relevant factors and distinct behavioral traits. Taking advantage of the extraordinary ecomorphological diversity of squamates (lizards and snakes) and adopting a multidisciplinary approach, this thesis explores the impact of locomotor behavior on squamate brain, particularly on different levels of cerebellar biological organization, and investigates cerebellar morphogenesis in two squamate species to gain insights on the developmental mechanisms potentially responsible for squamate cerebellar divergence. Along with significant variations in cerebellar morphology and relative size across squamates, this thesis first highlights a wide heterogeneity in Purkinje cell (PC) spatial layout as well as in gene expression pattern, all correlating with specific locomotor behaviors, unveiling unique relationships between a major evolutionary transition and organ specialization in vertebrates. At the developmental level, the thesis indicates that developmental features considered, so far, exclusive hallmarks of avian and mammalian cerebellogenesis characterize squamate cerebellar morphogenesis. Furthermore, the thesis suggests that variations in the spatiotemporal patterning of different cerebellar neurons could be, at least partially, at the base of the large phenotypic diversification of the squamate cerebellum. Finally, this thesis reveals that squamates provide an important framework to expand our knowledge on organ system-ecology relationships and central nervous system (CNS) development and evolution in vertebrates.Eliöiden toimintaan liittyy oleellisena osana niiden kyky liikkua, eli siirtyä paikasta toiseen erilaisten ruuminosien liikkeiden avulla. Pikkuaivot (cerebellum) ovat hyvin oleellinen osa selkärankaisten liikkeen säätelyä, ja niiden toiminta onkin säilynyt peruspiirteiltään samana selkärankaisten evoluution aikana. Vaikka pikkuaivojen rooliin ei kuulu liikkeen aloittaminen tai rytmisen liikkeen tahdin säätely, niillä on huomattava rooli muussa liikkeen säätelyssä. Tähän lukeutuvat esimerkiksi liikkeiden oppiminen ja korjaaminen. Pikkuaivoissa esiintyy hyvin paljon lajien välistä vaihtelua, mikä johtuu todennäköisesti yksilönkehityksen ja sen säätelyn eroavaisuuksista eri lajeilla. Eroja on havaittavissa niin pikkuaivojen morfologiassa, suhteellisessa koossa kuin myös niiden kuorikerroksen rakenteessa, usein jopa lähisukuisten lajien välillä. Nämä eroavaisuudet heijastelevatkin usein eläinten erilaisia toiminnallisia tarpeita, liittyen varsinkin käyttäytymispiirteisiin sekä muihin niiden ekologiaan linkittyviin tekijöihin. Suomumatelijoilla (liskoilla ja käärmeillä) on huomattava laaja kirjos erilaisia ekomorfologioita ja liikkumistapoja. Tämä väitöskirja keskittyykin selvittämään liikkumistapojen vaikutusta suomumatelijoiden aivoihin sekä yleisesti että erityisesti pikkuaivoja tarkastellen. Huomio keskittyy pikkuaivoissa sekä kokonaiskuvan muodostamiseen niiden rakenteesta että niiden yksilönkehitykseen. Yksilönkehityksen suhteen vertailussa ovat kaksi eri suomumatelijoiden edustajaa mahdollisten yksilönkehityksen muutosten mekanismien selvittämiseksi. Väitöskirjatyössä havaittiin suomumatelijoilla merkittävää pikkuaivojen morfologian ja suhteellisen koon lajienvälistä vaihtelua. Tämän lisäksi työn aikana havaittiin huomattavia eroja pikkuaivojen niin sanottujen Purkinjen solujen järjestäytymisessä sekä eri geenien luennassa erilaista liikkumistyyppiä edustavien lajien välillä. Purkinjen solujen järjestäytymisen ja geeniluennan havaittiin myös korreloivan erilaisten liikkumistyyppien kanssa, tuoden esiin mielenkiintoisen yhteyden evolutiivisten muutosten ja elinten erikoistumisen välillä. Samoin tulokset viittaavat siihen, että linnuille ja nisäkkäille ainutlaatuisiksi luultuja pikkuaivojen muodostumisen piirteitä löytyy myös suomumatelijoilta. Väitöskirjatyössä havaittiin lisäksi viitteitä suomumatelijoiden pikkuaivojen monimuotoisuuden taustalla olevista yksilönkehityksen muutoksista. Tulosten valossa on mahdollista, että pikkuaivojen neuronien kaavoituksen ajoituksen ja sijainnin muutokset voisivat ainakin osin olla syy suomumatelijoiden pikkuaivojen monimuotoisuuteen. Laajemmassa mielessä tulokset tuovat esiin myös suomumatelijoiden erittäin oleellisen roolin selkärankaisten evoluution tutkimuksessa kahdesta oleellisesta tulokulmasta: selkärankaisten keskushermoston yksilönkehityksen ja evoluution tutkimus sekä yleisemmällä tasolla elinsysteemien ja ekologian yhteyden selvittäminen

Le rôle des cellules dopaminergiques dans la locomotion induite par l'olfaction chez la lamproie

La détection de molécules chimiques par l'odorat est importante pour guider le comportement des animaux. Chez la lamproie marine, Petromyzon marinus, l'olfaction est vitale pour plusieurs fonctions telles que l’alimentation, l’évitement des prédateurs et la reproduction. Les différents comportements olfactifs de la lamproie sont les mieux caractérisés parmi tous les vertébrés aquatiques et ils font l’objet du premier chapitre de l’introduction. Les circuits du cerveau responsables des mouvements produits lors de la détection de stimuli olfactifs ont été examinés chez la lamproie. Des études récentes révèlent qu’il existe deux organes olfactifs périphériques ayant des projections parallèles qui innervent des parties distinctes du bulbe olfactif (BO). Dans les deux cas, le signal olfactif atteint éventuellement les cellules réticulospinales (RS), qui activent les réseaux locomoteurs spinaux. La littérature portant sur ces circuits neuronaux est décrite dans le deuxième chapitre introductif. Le substrat neuronal par lequel le signal olfactif est transmis aux cellules RS n'est pas complètement caractérisé mais des données du laboratoire Dubuc suggèrent que le tubercule postérieur (TP) serait une cible importante des projections du BO. Puisque cette région contient des neurones dopaminergiques (DA) impliqués dans le contrôle moteur, l’objectif principal de cette thèse était de déterminer leur rôle dans le traitement du signal olfactif et la production de locomotion. Nos résultats ont permis de caractériser l'innervation DA du BO de la lamproie et d’observer que les neurones DA du TP projettent à la partie médiane du BO chez les animaux de stade larvaire et adulte. De plus, l’activation de récepteurs D2 dans cette région diminue la transmission du signal olfactif aux cellules RS. Dans le reste du BO, des neurones DA apparaissent au stade adulte. Ces observations sont rapportées dans le premier chapitre des résultats. Puisque les neurones DA du TP peuvent moduler la transmission olfactomotrice au niveau du BO, ils pourraient aussi jouer un rôle via leurs projections connues vers le tronc cérébral. Le deuxième chapitre des résultats se penche donc sur l’implication du TP dans le relai de l’information olfactive au système moteur. L’étude des projections du BO montre que les neurones DA sont ciblés, incluant ceux qui projettent à la région locomotrice mésencéphalique (RLM), responsable de l’initiation et du contrôle de la locomotion. Aussi, la stimulation olfactive active des neurones du TP qui projettent à la RLM. Dans une préparation dont la tête est fixée mais le corps peut se déplacer, la stimulation olfactive induit de la nage en recrutant simultanément le TP et les cellules RS. Nous montrons aussi que le TP est recruté durant la nage survenant spontanément, ce qui indique que cette région joue un rôle important dans le contrôle locomoteur. Cette thèse révèle que les neurones DA du TP peuvent 1) être activés par la détection d’odeurs et ensuite 2) moduler la transmission au niveau du BO ainsi que 3) recruter la RLM pour produire un épisode de nage. Ces données suggèrent qu’ils occupent une position-clé dans l’intégration sensorimotrice des stimuli olfactifs puisqu’ils encodent à la fois de l’information sensorielle et motrice.The detection of chemical molecules by smell is important in guiding the behavior of animals. In the sea lamprey, Petromyzon marinus, olfaction is vital for several functions such as feeding, predator avoidance and reproduction. The various olfactory behaviors of the lamprey are the best characterized among all aquatic vertebrates and they were reviewed in the first chapter of the introduction. The brain circuitry responsible for producing movement upon sensing olfactory stimuli has been examined in lamprey. Recent studies revealed that there are two peripheral olfactory epithelia with parallel projections that reach distinct parts of the olfactory bulb (OB). In both cases, the olfactory signal eventually reaches reticulospinal (RS) cells, which activate the locomotor networks of the spinal cord. The literature describing these neural circuits is thoroughly reviewed in the second chapter of the introduction. The neuronal substrate by which the olfactory signal is transmitted to RS cells is not fully characterized, but data from the Dubuc laboratory suggest that the posterior tubercle (PT) may be an important target for OB projections. Since this region contains dopaminergic (DA) neurons involved in motor control, the main objective of this thesis was to determine their role in olfactory signal processing and the production of locomotion. Our results have allowed to characterize the DA innervation of the lamprey OB and show that DA neurons of the PT send projections to the medial part of the OB in larval and adult animals. In addition, the activation of D2 receptors in this region decreases the transmission of the olfactory signal to RS cells. In the rest of the OB, DA neurons appear in adult animals. These observations are reported in the first chapter of the Results. Since DA neurons of the PT can modulate olfactory-motor transmission at the level of the OB, they could also play a role through existing descending projections to the brainstem. Thus, in the second chapter of the Results, we studied the involvement of the PT in the relay of olfactory information to the motor system. The analysis of OB projections shows that DA neurons are targeted, including those that project to the mesencephalic locomotor region (MLR), which is responsible for initiating and controlling locomotion. Moreover, olfactory stimulation activates PT neurons that project to the MLR. In a head-fixed preparation in which the body moves, olfactory stimulation induces swimming simultaneously with PT and RS cell activity. We also show that the PT is recruited during spontaneously occurring swimming, which indicates that this region plays an important role in locomotor control. This thesis reveals that DA neurons in the PT can 1) be activated following odorant detection and then 2) modulate the transmission at the level of the OB as well as 3) recruit the MLR to produce a swimming episode. These data suggest that they occupy a key position in the sensorimotor integration of olfactory stimuli since they encode both sensory and motor information

Sensorimotor adjustments after unilateral spinal cord injury in adult rats

A variety of behavioural tests were used to examine both sensory and motor function of freely behaving unilaterally spinal cord-injured and uninjured rats. The first experiment was designed to determine whether sensory and motor differences existed between uninjured Fischer, Lewis, Long-Evans, Sprague-Dawley and Wistar rats using endpoint, quantitative kinematic, and kinetic measurements. The second experiment examined differences in sensorimotor responses to cervical spinal cord hemisection in Lewis, Long-Evans and Wistar rats. For the third experiment, reflex and locomotor abilities of unilateral cervical or thoracic spinal cord hemisected Long-Evans rats were determined using endpoint, semi-quantitative kinematic, and kinetic measurements. The fourth experiment was designed to investigate the importance of the rubrospinal tract and ascending dorsal column pathways to overground locomotion. This experiment was conducted to help explain the behavioural observations made following cervical spinal cord hemisection. Furthermore, this experiment examined the effects of combined unilateral rubrospinal and dorsal column injury on overground locomotion using endpoint and kinetic measurements. Finally, the fifth experiment set out to investigate the contribution of tracts running in the ventrolateral spinal cord on overground locomotion in freely behaving Long-Evans rats. These animals were assessed using endpoint and kinetic measurements. The results of these studies revealed that motor and sensory functions are not similar for all uninjured strains of rats. Specifically, Fischer rats tend to have considerable differences in their morphological features and sensorimotor abilities compared to the other strains examined. Results from the other experiments indicate that adult freely behaving female rats develop a characteristic gait when pathways important for locomotion are injured unilaterally, regardless of strain. The rubrospinal tract and ascending dorsal column pathways appear to be important for both skilled and flat-ground locomotion as well as forelimb use while rearing. Pathways traveling within the ventrolateral pathway, however, are not necessary or sufficient for locomotion or limb useage while rearing when injured by themselves. Animals with ventrolateral spinal funiculus injuries regain normal forelimb use and skilled locomotor abilities. Injury to the ventrolateral spinal funiculus, however, results in mild (compared to rubrospinal and dorsal column injured animals) yet long-lasting locomotor changes based on ground reaction force determination. These findings are in agreement with the current opinion that there is a substantial amount of functional redundancy of pathways traveling in the ventral and ventrolateral funiculi

Noradrenergic control of spinal motor circuitry in two related amphibian species 'Xenopus laevis' and 'Rama temporaria'

1. The role of the catecholamine noradrenaline (NA) was examined during fictive swimming in Xenopus laevis tadpoles. 2. The primary effects of the amine in both embryonic and larval Xenopus was to markedly decrease motor frequency whilst simultaneously reducing rostrocaudal delays during swimming. 3. The NA-mediated modulation of swimming activity in Xenopus larvae can be reversed with phentolamine, a non-selective an adrenergic receptor antagonist, suggesting that NA may be acting through either α₁ or α₂ receptors, or a combination of both. 4. Intracellular recordings made from embryo spinal motorneurones revealed that reciprocal inhibitory glycinergic potentials are enhanced by NA. This effect is most prominent in caudal regions of the spinal cord where inhibitory synaptic drive is generally weaker. 5. NA was also found to enhance glycinergic reciprocal inhibition during swimming in larval spinal cord motomeurones. 6. Intracellular recordings, under tetrodotoxin, reveal that NA enhances the occurrence of spontaneous glycinergic inhibitory post synaptic potentials arising from the terminals of inhibitory intemeurones, suggesting that the amine is acting presynaptically to enhance evoked release of glycine during swimming. 7. The effects of NA on swimming frequency and rostrocaudal delay appear to be largely mediated through an enhancement of glycinergic reciprocal inhibition as blockade of glycine receptors with strychnine weakens the ability of the amine affect these parameters of motor output. 8. The effects of NA on motor output were also examined in embryos of the amphibian Rana temporaria. Whilst NA did not obviously affect swimming activity, the amine induced a non-rhythmic pattern of motor activity. 9. The free radical gas, nitric oxide also induced a non-rhythmic pattern of motor discharge that was remarkably similar to that elicited by NA, indicating that this neural messenger may be important for motor control

Life Sciences Program Tasks and Bibliography

This document includes information on all peer reviewed projects funded by the Office of Life and Microgravity Sciences and Applications, Life Sciences Division during fiscal year 1995. Additionally, this inaugural edition of the Task Book includes information for FY 1994 programs. This document will be published annually and made available to scientists in the space life sciences field both as a hard copy and as an interactive Internet web pag