Sub-cortical sources of the somatosensory pathway are hypoactive in migraine interictally:a Functional Source Separation analysis

Background: Recent morpho-functional evidence pointed out that abnormalities in the thalamus could play a major role in the expression of migraine neurophysiological and clinical correlates. Whether this phenomenon is primary or secondary to its functional disconnection from the brainstem remains to be determined. We used a Functional Source Separation algorithm of EEG signal to extract the activity of the different neuronal pools recruited at different latencies along the somatosensory pathway in interictal migraine without aura (MO) patients. Methods: Twenty MO patients and 20 healthy volunteers (HV) underwent EEG recording. Four ad-hoc functional constraints, two sub-cortical (FS14 at brainstem and FS16 at thalamic level) and two cortical (FS20 radial and FS22 tangential parietal sources), were used to extract the activity of successive stages of somatosensory information processing in response to the separate left and right median nerve electric stimulation. A band-pass digital filter (450-750 Hz) was applied offline in order to extract high-frequency oscillatory (HFO) activity from the broadband EEG signal. Results: In both stimulated sides, significant reduced sub-cortical brainstem (FS14) and thalamic (FS16) HFO activations characterized MO patients when compared with HV. No difference emerged in the two cortical HFO activations between the two groups. Conclusions: Present results are the first neurophysiological evidence supporting the hypothesis that a functional disconnection of the thalamus from the subcortical monoaminergic system may underline the interictal cortical abnormal information processing in migraine. Further studies are needed to investigate the precise directional connectivity across the entire primary subcortical and cortical somatosensory pathway in interictal MO. Written informed consent to publication was obtained from the patient(s).

Stimulus Integration and Parsing in the Primate Auditory Midbrain

Integrating information from multiple sources is a crucial function of the brain. Examples of such integration include multiple stimuli of different modalties, such as visual and auditory, multiple stimuli of the same modality, such as auditory and auditory, and integrating stimuli from the sensory organs (i.e. ears) with stimuli delivered from brain-machine interfaces.

The overall aim of this body of work is to empirically examine stimulus integration in these three domains to inform our broader understanding of how and when the brain combines information from multiple sources.

First, I examine visually-guided auditory, a problem with implications for the general problem in learning of how the brain determines what lesson to learn (and what lessons not to learn). For example, sound localization is a behavior that is partially learned with the aid of vision. This process requires correctly matching a visual location to that of a sound. This is an intrinsically circular problem when sound location is itself uncertain and the visual scene is rife with possible visual matches. Here, we develop a simple paradigm using visual guidance of sound localization to gain insight into how the brain confronts this type of circularity. We tested two competing hypotheses. 1: The brain guides sound location learning based on the synchrony or simultaneity of auditory-visual stimuli, potentially involving a Hebbian associative mechanism. 2: The brain uses a ‘guess and check’ heuristic in which visual feedback that is obtained after an eye movement to a sound alters future performance, perhaps by recruiting the brain’s reward-related circuitry. We assessed the effects of exposure to visual stimuli spatially mismatched from sounds on performance of an interleaved auditory-only saccade task. We found that when humans and monkeys were provided the visual stimulus asynchronously with the sound but as feedback to an auditory-guided saccade, they shifted their subsequent auditory-only performance toward the direction of the visual cue by 1.3-1.7 degrees, or 22-28% of the original 6 degree visual-auditory mismatch. In contrast when the visual stimulus was presented synchronously with the sound but extinguished too quickly to provide this feedback, there was little change in subsequent auditory-only performance. Our results suggest that the outcome of our own actions is vital to localizing sounds correctly. Contrary to previous expectations, visual calibration of auditory space does not appear to require visual-auditory associations based on synchrony/simultaneity.

My next line of research examines how electrical stimulation of the inferior colliculus influences perception of sounds in a nonhuman primate. The central nucleus of the inferior colliculus is the major ascending relay of auditory information before it reaches the forebrain, and thus an ideal target for understanding low-level information processing prior to the forebrain, as almost all auditory signals pass through the central nucleus of the inferior colliculus before reaching the forebrain. Thus, the inferior colliculus is the ideal structure to examine to understand the format of the inputs into the forebrain and, by extension, the processing of auditory scenes that occurs in the brainstem. Therefore, the inferior colliculus was an attractive target for understanding stimulus integration in the ascending auditory pathway.

Moreover, understanding the relationship between the auditory selectivity of neurons and their contribution to perception is critical to the design of effective auditory brain prosthetics. These prosthetics seek to mimic natural activity patterns to achieve desired perceptual outcomes. We measured the contribution of inferior colliculus (IC) sites to perception using combined recording and electrical stimulation. Monkeys performed a frequency-based discrimination task, reporting whether a probe sound was higher or lower in frequency than a reference sound. Stimulation pulses were paired with the probe sound on 50% of trials (0.5-80 µA, 100-300 Hz, n=172 IC locations in 3 rhesus monkeys). Electrical stimulation tended to bias the animals’ judgments in a fashion that was coarsely but significantly correlated with the best frequency of the stimulation site in comparison to the reference frequency employed in the task. Although there was considerable variability in the effects of stimulation (including impairments in performance and shifts in performance away from the direction predicted based on the site’s response properties), the results indicate that stimulation of the IC can evoke percepts correlated with the frequency tuning properties of the IC. Consistent with the implications of recent human studies, the main avenue for improvement for the auditory midbrain implant suggested by our findings is to increase the number and spatial extent of electrodes, to increase the size of the region that can be electrically activated and provide a greater range of evoked percepts.

My next line of research employs a frequency-tagging approach to examine the extent to which multiple sound sources are combined (or segregated) in the nonhuman primate inferior colliculus. In the single-sound case, most inferior colliculus neurons respond and entrain to sounds in a very broad region of space, and many are entirely spatially insensitive, so it is unknown how the neurons will respond to a situation with more than one sound. I use multiple AM stimuli of different frequencies, which the inferior colliculus represents using a spike timing code. This allows me to measure spike timing in the inferior colliculus to determine which sound source is responsible for neural activity in an auditory scene containing multiple sounds. Using this approach, I find that the same neurons that are tuned to broad regions of space in the single sound condition become dramatically more selective in the dual sound condition, preferentially entraining spikes to stimuli from a smaller region of space. I will examine the possibility that there may be a conceptual linkage between this finding and the finding of receptive field shifts in the visual system.

In chapter 5, I will comment on these findings more generally, compare them to existing theoretical models, and discuss what these results tell us about processing in the central nervous system in a multi-stimulus situation. My results suggest that the brain is flexible in its processing and can adapt its integration schema to fit the available cues and the demands of the task.

A Novel Mouse Model of Diffuse Intrinsic Pontine Glioma Initiated in Pax3-Expressing Cells.

Diffuse intrinsic pontine glioma (DIPG) is a rare and incurable brain tumor that arises predominately in children and involves the pons, a structure that along with the midbrain and medulla makes up the brainstem. We have previously developed genetically engineered mouse models of brainstem glioma using the RCAS/Tv-a system by targeting PDGF-B overexpression, p53 loss, and H3.3K27M mutation to Nestin-expressing brainstem progenitor cells of the neonatal mouse. Here we describe a novel mouse model targeting these same genetic alterations to Pax3-expressing cells, which in the neonatal mouse pons consist of a Pax3+/Nestin+/Sox2+ population lining the fourth ventricle and a Pax3+/NeuN+ parenchymal population. Injection of RCAS-PDGF-B into the brainstem of Pax3-Tv-a mice at postnatal day 3 results in 40% of mice developing asymptomatic low-grade glioma. A mixture of low- and high-grade glioma results from injection of Pax3-Tv-a;p53(fl/fl) mice with RCAS-PDGF-B and RCAS-Cre, with or without RCAS-H3.3K27M. These tumors are Ki67+, Nestin+, Olig2+, and largely GFAP- and can arise anywhere within the brainstem, including the classic DIPG location of the ventral pons. Expression of the H3.3K27M mutation reduces overall H3K27me3 as compared with tumors without the mutation, similar to what has been previously shown in human and mouse tumors. Thus, we have generated a novel genetically engineered mouse model of DIPG, which faithfully recapitulates the human disease and represents a novel platform with which to study the biology and treatment of this deadly disease.

Pre-Clinical Models of Diffuse Intrinsic Pontine Glioma.

Diffuse intrinsic pontine glioma (DIPG) is a rare and incurable brain tumor that arises in the brainstem of children predominantly between the ages of 6 and 8. Its intricate morphology and involvement of normal pons tissue precludes surgical resection, and the standard of care today remains fractionated radiation alone. In the past 30 years, there have been no significant advances made in the treatment of DIPG. This is largely because we lack good models of DIPG and therefore have little biological basis for treatment. In recent years, however, due to increased biopsy and acquisition of autopsy specimens, research is beginning to unravel the genetic and epigenetic drivers of DIPG. Insight gleaned from these studies has led to improvements in approaches to both model these tumors in the lab and to potentially treat them in the clinic. This review will detail the initial strides toward modeling DIPG in animals, which included allograft and xenograft rodent models using non-DIPG glioma cells. Important advances in the field came with the development of in vitro cell and in vivo xenograft models derived directly from autopsy material of DIPG patients or from human embryonic stem cells. Finally, we will summarize the progress made in the development of genetically engineered mouse models of DIPG. Cooperation of studies incorporating all of these modeling systems to both investigate the unique mechanisms of gliomagenesis in the brainstem and to test potential novel therapeutic agents in a preclinical setting will result in improvement in treatments for DIPG patients.

Premotor Mechanisms for Orofacial Coordination

The mouth, throat, and face contain numerous muscles that participate in a large variety of orofacial behaviors. The jaw and tongue can move independently, and thus require a high degree of coordination among the muscles that move them to prevent self-injury. However, different orofacial behaviors require distinct patterns of coordination between these muscles. The method through which motor control circuitry might coordinate this activity has yet to be determined. Electrophysiological, immunohistochemical, and retrograde tracing studies have attempted to identify populations of premotor neurons which directly send information to orofacial motoneurons in an effort to identify sources of coordination. Yet these studies have not provided a complete picture of the population of neurons which monosynaptically connect to jaw and tongue motoneurons. Additionally, while many of these studies have suggested that premotor neurons projecting to multiple motor pools may play a role in coordination of orofacial muscles, no clear functional roles for these neurons in the coordination of natural orofacial movements has been identified.

In this dissertation, I took advantage of the recently developed monosynaptic rabies virus to trace the premotor circuits for the jaw-closing masseter muscle and tongue-protruding genioglossus muscle in the neonatal mouse, uncovering novel premotor inputs in the brainstem. Furthermore, these studies identified a set of neurons which form boutons onto motor neurons in multiple motor pools, providing a premotor substrate for orofacial coordination. I then combined a retrogradely traveling lentivirus with a split-intein mediated split-Cre recombinase system to isolate and manipulate a population of neurons which project to both left and right jaw-closing motor nuclei. I found that these bilaterally projecting neurons also innervate multiple other orofacial motor nuclei, premotor regions, and midbrain regions implicated in motor control. I anatomically and physiologically characterized these neurons and used optogenetic and chemicogenetic approaches to assess their role in natural jaw-closing behavior, specifically with reference to bilateral masseter muscle electromyogram (EMG) activity. These studies identified a population of bilaterally projecting neurons in the supratrigeminal nucleus as essential for maintenance of an appropriate level of masseter activation during natural chewing behavior in the freely moving mouse. Moreover, these studies uncovered two distinct roles of supratrigeminal bilaterally projecting neurons in bilaterally synchronized activation of masseter muscles, and active balancing of bilateral masseter muscle tone against an excitatory input. Together, these studies identify neurons which project to multiple motor nuclei as a mechanism by which the brain coordinates orofacial muscles during natural behavior.

miRNAs as therapeutic agents in neurodegeneration, a pilot study

L’échec des différents essais cliniques souligne la nécessité de développer des nouvelles thérapies pour la maladie d’Alzheimer (MA), la cause la plus commune de démence. Les microARNs (miARNs) sont les ARNs non-codants les plus étudiés et ils jouent un rôle important dans la modulation de l’expression des gènes et de multiples voies de signalisation. Des études antérieures, dont celles de mon laboratoire d’accueil, ont permis de développer l’hypothèse que certains membres de la famille miR-15/107 (c.-à-d. miR-15ab, miR-16, miR-195, miR-424, and miR-497) pourraient être utilisés comme agents thérapeutiques dans MA. En effet, cette famille avait le potentiel de réguler de multiples gènes associés à MA, tels que la protéine précurseur de l’amyloïde (APP), la β-secrétase (BACE1), et la protéine Tau. Tel que démontré dans ce projet de thèse, j’ai choisi miR-16 comme cible thérapeutique potentielle pour MA parmi tous les membres de la famille. L’essai luciférase dans ce projet confirme que miR-16 peut réguler simultanément APP et BACE1, directement par une interaction avec la région non-codante en 3’ de l’ARNm). Notamment, nous observons aussi une réduction de la production des peptides amyloïdes et de la phosphorylation de Tau après une augmentation de miR-16 en cellule. J’ai ensuite validé mes résultats in vivo dans la souris en utilisant une méthode de livraison de miR-16 via une pompe osmotique implanté dans le cerveau. Dans ce cas, l’expression des protéines d’intérêts (APP, BACE1, Tau) a été mesurée par immunobuvardage et PCR à temps réel. Après validation, ces résultats ont été complémentés par une étude protéomique (iTRAQ) du tronc cérébral et de l’hippocampe, deux régions associées à la maladie. Ces données m’ont permis d’identifier d’autres protéines régulées par miR-16 in vivo, incluant α-Synucléine, Transferrine receptor1, et SRm300. Une autre observation intéressante : les voies régulées par miR-16 in vivo sont directement en lien avec le stress oxydatif et la neurodégénération. En résumé, ce travail démontre l’efficacité et la faisabilité d’utiliser un miARN comme outil thérapeutique pour la maladie d’Alzheimer. Ces résultats rentrent dans un cadre plus vaste de découvrir de nouvelles cibles pour MA, et en particulier la forme sporadique de la maladie qui représente plus de 95% de tous les cas. Évidemment, la découverte d’une molécule pouvant cibler simultanément les deux pathologies de la maladie (plaques amyloïdes et hyper phosphorylation de tau) est nouvelle et intéressante, et ce domaine de recherche ouvre la porte aux autres petits ARNs non-codants dans MA et les maladies neurodégénératives connexes.

Rôle de DSCAM dans le développement normal du cortex moteur et de la voie corticospinale

DSCAM est exprimé dans le cortex lors du développement et sa mutation altère l’arborisation dendritique des neurones pyramidaux du cortex moteur. Considérant que les souris DSCAM2J possèdent des problèmes posturaux et locomoteurs, nous émettons l’hypothèse que DSCAM est impliqué dans le fonctionnement normal du cortex moteur et de la voie corticospinale. Comparées aux souris contrôles, les souris DSCAM2J vont présenter des problèmes moteurs à basse vitesse et enjamber un obstacle presque normalement à vitesse intermédiaire. Le traçage antérograde de la voie corticospinale révèle un patron d’innervation normal dans le tronc cérébrale et la moelle épinière. Des microstimulations intracorticale du cortex moteur évoque des réponses électromyographiques dans les membres à un seuil et une latence plus élevé. Par contre, une stimulation de la voie corticospinale dans la médulla évoque des réponses électromyographies à un seuil et une latence similaire entre les deux groupes, suggérant une réduction de l’excitabilité du cortex moteur.

Real-time functional magnetic resonance imaging neurofeedback in motor neurorehabilitation

Purpose of review Recent developments in functional magnetic resonance imaging (fMRI) have catalyzed a new field of translational neuroscience. Using fMRI to monitor the aspects of task-related changes in neural activation or brain connectivity, investigators can offer feedback of simple or complex neural signals/patterns back to the participant on a quasireal-time basis [real-time-fMRI-based neurofeedback (rt-fMRI-NF)]. Here, we introduce some background methodology of the new developments in this field and give a perspective on how they may be used in neurorehabilitation in the future. Recent findings The development of rt-fMRI-NF has been used to promote self-regulation of activity in several brain regions and networks. In addition, and unlike other noninvasive techniques, rt-fMRI-NF can access specific subcortical regions and in principle any region that can be monitored using fMRI including the cerebellum, brainstem and spinal cord. In Parkinson’s disease and stroke, rt-fMRI-NF has been demonstrated to alter neural activity after the self-regulation training was completed and to modify specific behaviours. Summary Future exploitation of rt-fMRI-NF could be used to induce neuroplasticity in brain networks that are involved in certain neurological conditions. However, currently, the use of rt-fMRI-NF in randomized, controlled clinical trials is in its infancy.

Organisation chimioanatomique des afférences pallidales chez le primate

L’élucidation de la position qu’occupent les projections sérotoninergique (5-HT), cholinergique (ACh) et dopaminergique (DA) du tronc cérébral dans l’organisation anatomofonctionelle du globus pallidus externe (GPe) et interne (GPi) au sein des ganglions de la base chez le primate est primordiale à la compréhension de ce système neuronal hautement complexe impliqué dans le contrôle du comportement moteur. Les travaux de recherche consolidés dans la présente thèse rapportent les résultats principalement obtenus chez le singe écureuil (Saimiri sciureus) à l’aide de marquages immunohistochimiques et de quantifications stéréologiques servant à évaluer la distribution régionale et les caractéristiques ultrastructurales des varicosités axonales 5-HT, ACh et DA observées dans le pallidum. Nos données ont permis l’éloboration d’un nouveau modèle du neurone pallidal en tenant compte de la hiérarchie et des caractéristiques neurochimiques de ses entrées synaptiques. Ainsi, l’analyse quantitative en microscopie optique révèle que le GPe et le GPi reçoivent des innervations 5-HT, ACh et DA de densités variables et distribuées de façon hétérogène. Plus particulièrement, le GPe est innervé par 600 000 varicosités 5-HT/mm3 de tissu, 500 000 varicosités ACh/mm3 et 170 000 varicosités DA/mm3. En revanche, le GPi reçoit 600 000 varicosités 5-HT/mm3, 250 000 varicosités ACh/mm3 et 190 000 varicosités DA/mm3. De plus, la 5-HT, l’ACh et la DA ciblent préférentiellement les secteurs correspondant aux territoires fonctionnels associatifs et limbiques du pallidum, suggérant un rôle de ces projections dans la planification du comportement moteur ainsi que dans la régulation de l’attention et de l’humeur. Nos analyses en microscopie électronique révèlent que très peu de ces varicosités axonales établissent un contact synaptique, puisque plus de 70% des varicosités 5-HT, ACh et DA sont complètement dépouvues de jonction synaptique. Ainsi, bien que la 5-HT, l’ACh et la DA seraient en mesure de moduler directement les neurones pallidaux grâce à la transmission synaptique, leur plus grande influence s’opérerait par la transmission volumique, permettant d’influencer à la fois les neurones pallidaux et leurs afférences, principalement du striatum et noyau subthalamique. L’ensemble de ces résultats indique que les projections 5-HT, ACh et DA du tronc cérébral agissent de concert avec les afférences plus robustes en provenance du striatum et du noyau subthalamique. Ces nouvelles données neuroanatomiques positionnent le tronc cérébral en tant qu’acteur important dans l’organisation anatomique et fonctionnelle du pallidum chez le primate et doivent être prises en considération dans l’élaboration de nouvelles approches thérapeutiques visant à contrer les processus neurodégénératifs qui affectent les ganglions de la base, tel que la maladie de Parkinson.

Two Cases of Arnold-Chiari Malformation with Respiratory Failure

Arnold–Chiari malformation is defined as downward displacement of the brainstem and cerebellum through the foramen magnum. It has different clinical presentations and four subtypes. It is known that downward migration of posterior fossa components through the foramen magnum and associated lower cranial nerve palsy and brainstem compression can cause respiratory failure. Acute respiratory failure could mark the onset of the disease. Posterior fossa decompression performed to treat primary disease can improve the central sleep abnormalities. As respiratory failure is rarely seen, this paper presents two cases of Arnold–Chiari malformation with respiratory failure.

Double-trouble for respiratory control in Pompe disease

A commentary on ‘Hypoglossal neuropathology and respiratory activity in Pompe mice’, by Lee, K.-Z., Qiu, K., Sandhu, M. S., Elmullah, M. K., Falk, D. J., Lane, M. A., Reier, P. J., Byrne, B. J., and Fuller, D. D. (2011). Front. Physiol. 2:31. doi: 10.3389/fphys.2011.00031.

Development of metabotropic glutamate receptors from trigeminal nuclei to barrel cortex in postnatal mouse

Expression patterns of group I (mGluR1α and mGluR5)and group II (mGluR2/3) metabotropic glutamate receptor subtypes were examined immunocytochemically in the trigeminal system of mice during the first 3 weeks of postnatal development, when somatotopic whisker representations are sequentially established from brainstem through thalamus to cerebral cortex. Immunostaining for all three epitopes formed whisker-related patterns in the trigeminal nuclei from postnatal day (P) 0, in the ventral posterior thalamic nucleus from P2, and in the posteromedial barrel subfield of somatosensory cortex (SI) from P4. The appearance of whisker-related patterns was preceded by increased levels of immunostaining of the neuropil, which subsequently declined from the trigeminal nuclei upward. In SI, mGluR1α-positive neurons were observed in all cortical layers from P2. mGluR5 was localized in neurons, glial cells, and neuropil from P2. mGluR2/3 immunostaining was distributed only in the neuropil at all ages. The three receptor subtypes showed moderate to high expression in deep layer V throughout development. Transient expression peaked in the hollows of layer IV barrels from P4 to P9, and then fell off as expression increased in supragranular layers from P14 to P21. The deep aspect of the cortical subplate (layer VIb) showed dense mGluR5 and less dense mGluR1α immunostaining throughout development. Up-regulation of expression of group I and II mGluRs is correlated with the growth and refinement of connectivity and the establishment of somatotopic patterns in the three main relay stations of the trigeminal system. This finding suggests roles for mGluRs in the early processing of sensory information and in developmental plasticity.