Control of hypothalamic neuroendocrine interactions

El hipotálamo juega en los mamíferos superiores un papel central en la integración de funciones vitales como la regulación del metabolismo energético global, la saciedad y el hambre, el control de la presión sanguínea y la temperatura corporal, la sed, hidratación y metabolismo salino del organismo, y las funciones testiculares y ováricas, entre otras. Muchas de estas funciones neuroendocrinas se realizan mediante el control del funcionamiento de la hipófisis, utilizando un complejo sistema de retroalimentación que modula la secreción de una gran variedad de hormonas hipofisarias con efectos sistémicos de vital importancia, incluyendo las hormonas tiroideas o la hormona del crecimiento, entre otras. El hipotálamo consta de aproximadamente una docena de subestructuras, conocidas como núcleos hipotalámicos, que se encargan de controlar los diversos procesos. Hasta muy recientemente no ha sido posible evaluar la función hipotalámica directamente in vivo, un aspecto que se resolvía mediante procedimientos indirectos como la determinación de cambios en el peso corporal, eliminación de líquidos, alteraciones en la termorregulación o desequilibrios en el perfil de hormonas en sangre. En esta revisión describiremos toda una nueva serie de métodos de imagen no invasiva para la evaluación directa de la función hipotalámica y su impacto potencial en nuestro conocimiento actual de la regulación de las interacciones neuroendocrinas, con especial referencia a la regulación hipotalámica del apetito in vivo.Este trabajo ha sido financiado en parte por las ayudas: SAF-2008-01327, SAF2011-23622 del Ministerio de Economía y Competitividad y S2010/BMD-2349 de la Comunidad de Madrid concedidas a Sebastián Cerdán y la beca predoctoral BES 2009-027615 del Ministerio de Economía y Competitividad concedida a Blanca Lizarbe.Peer Reviewe

Neuroimaging Evidence of Major Morpho-Anatomical and Functional Abnormalities in the BTBR T+TF/J Mouse Model of Autism

BTBR T+tf/J (BTBR) mice display prominent behavioural deficits analogous to the defining symptoms of autism, a feature that has prompted a widespread use of the model in preclinical autism research. Because neuro-behavioural traits are described with respect to reference populations, multiple investigators have examined and described the behaviour of BTBR mice against that exhibited by C57BL/6J (B6), a mouse line characterised by high sociability and low self-grooming. In an attempt to probe the translational relevance of this comparison for autism research, we used Magnetic Resonance Imaging (MRI) to map in both strain multiple morpho-anatomical and functional neuroimaging readouts that have been extensively used in patient populations. Diffusion tensor tractography confirmed previous reports of callosal agenesis and lack of hippocampal commissure in BTBR mice, and revealed a concomitant rostro-caudal reorganisation of major cortical white matter bundles. Intact inter-hemispheric tracts were found in the anterior commissure, ventro-medial thalamus, and in a strain-specific white matter formation located above the third ventricle. BTBR also exhibited decreased fronto-cortical, occipital and thalamic gray matter volume and widespread reductions in cortical thickness with respect to control B6 mice. Foci of increased gray matter volume and thickness were observed in the medial prefrontal and insular cortex. Mapping of resting-state brain activity using cerebral blood volume weighted fMRI revealed reduced cortico-thalamic function together with foci of increased activity in the hypothalamus and dorsal hippocampus of BTBR mice. Collectively, our results show pronounced functional and structural abnormalities in the brain of BTBR mice with respect to control B6 mice. The large and widespread white and gray matter abnormalities observed do not appear to be representative of the neuroanatomical alterations typically observed in autistic patients. The presence of reduced fronto-cortical metabolism is of potential translational relevance, as this feature recapitulates previously-reported clinical observations

Magnetic resonance imaging of appetite-induced hypothalamic activity

Tesis doctoral inédita leida en la Universidad Autónoma de Madrid, Facultad de Ciencias, Departamento de Física de la Materia Condensada. Fecha de lectura: 26-10-2013La obesidad es un síndrome pandémico que subyace a las enfermedades más prevalentes y mórbidas en los países desarrollados. Es resultado de un desequilibrio en la regulación del apetito y del gasto energético; dos mecanismos que están fundamentalmente controlados por el hipotálamo. Las técnicas de imagen por resonancia magnética constituyen una herramienta excelente de evaluación anatómica y funcional del cerebro, en condiciones fisiológicas y patológicas, proporcionando un constante creciente tipo de aplicaciones. La tesis que aquí presento se ha centrado en el desarrollo de nuevas estrategias para evaluar de forma no invasiva los procesos de regulación cerebral del apetito en ratones y seres humanos, utilizando técnicas de resonancia magnética pesada en difusión. El Capítulo 1 proporciona una introducción a los principios básicos de la imagen por resonancia magnética, así como alguna de sus aplicaciones, recopilando las principales nociones fisiológicas de los mecanismos hipotalámicos de la regulación del apetito. También incluye s una breve compilación de las técnicas de neuroimagen más utilizadas en la evaluación de procesos relacionados con el apetito. En el Capítulo 2 describo el desarrollo y la implementación de una nueva técnica de imagen funcional basada en difusión para la detección de la actividad hipotalámica por apetito, en ratones y en seres humanos. El Capítulo 3 está dedicado a la aplicación de esta técnica a cuatro situaciones experimentales distintas, diseñadas para evaluar la respuesta específica de los núcleos hipotalámicos en procesos en los que la regulación del apetito y el balance energético están alterados. Finalmente, en el Capítulo 4 muestro la comparación del uso de la metodología analizada con distintos modelos de difusión- con los resultados obtenidos mediante la aplicación de una técnica funcional más convencional, ambas aplicadas al estudio de los efectos hipotalámicos de la administración de glucosa a ratones ayunados. En conclusión, mi Tesis doctoral demuestra que la técnica de imagen de resonancia magnética pesada en difusión proporciona un instrumento nuevo y robusto para el estudio de la regulación del apetito de forma no invasiva.Obesity is a pandemic syndrome that underlies the most morbid and prevalent diseases in developed countries. It results from an imbalance in appetite regulation and energy expenditure, two processes that are fundamentally controlled by the hypothalamus. Magnetic Resonance Imaging methods are excellently endowed to assess brain anatomy and function, under physiological and pathological conditions, providing an always increasing array of approaches. In this dissertation, I will introduce a collection of new strategies to evaluate non-invasively appetite regulation in the brain of mice and humans, based in the use of diffusion weighted magnetic resonance imaging methods. Chapter 1 introduces general concepts on the magnetic resonance imaging phenomenon and its applications, reviewing the key physiological mechanisms supporting hypothalamic appetite regulation. A short compilation of the most common neuroimaging techniques used to evaluate appetiterelated processes is also included. Chapter 2 describes the development and implementation of a new functional diffusion weighted imaging method applied to the detection of hypothalamic activity by fasting in mice and humans. Chapter 3 covers four different experimental manipulations designed to probe the role of specific hypothalamic nuclei in the regulation of appetite control and energy balance, under conditions where these are intentionally altered. Finally, Chapter 4 compares the use of the methodology analysed with different models of diffusion- with the results obtained with a more conventional functional imaging technique, both applied to the paradigm of glucose administration to fasting mice. In conclusion, this dissertation demonstrates that diffusion weighted magnetic resonance imaging methods provide a novel and useful approach to investigate appetite regulation non-invasively

MRI Studies of Appetite Centre Function in Rodents

Many different regions of the brain are involved in appetite control. A full understanding of their function and interaction requires studying neuronal activity at high resolution simultaneously in space and time. Two Magnetic Resonance Imaging (MRI) methods can potentially achieve this goal. Manganese-Enhanced (MEMRI) uses the accumulation of administered Mn[2+], which is paramagnetic (hence MRI visible) and taken up by active neurons through voltage-gated Ca[2+] channels during action potentials. Haemodynamic methods use one or more of many MRI-visible changes that occur to circulating blood in a brain region when it changes activity. These include blood-oxygenation level dependent (BOLD) and cerebral blood volume weighted (CBV) MRI. The aim of this project was to further develop, adapt and then use these methods to study the effects on neuronal activity of stimuli related to appetite and energy balance. The majority of work went towards adapting MEMRI for this. Amongst many tested changes, improvements were made to the MRI acquisition protocol (specifically using fast spin echo rather than spin-echo acquisition) to make it more sensitive to Mn-induced signal changes, increase spatial coverage from partial to whole brain and rostro-caudal spatial resolution from 1 to 0.4mm, all while maintaining the same temporal resolution. Most importantly, the neuroimaging analysis framework used in haemodynamic functional MRI was adapted for use with MEMRI. This included the adaptation of spatial normalization software to handle Mn-sensitive T[1]-weighted images dominated by non-brain tissue rather than brain dominated T[2]/T*[2]-weighted images, and the generation of a signal change model for use in GLM. This enabled much more objective, reproducible and less laborious data analysis than with previous hand drawn ROIs. Attempts were made to use BOLD- and CBV-fMRI to study the effects of potent, appetite-modulating gut hormones on appetite, though these failed to produce a response

A novel receive-only liquid nitrogen (LN2)-cooled RF coil for high-resolution in vivo imaging on a 3-Tesla whole-body scanner

The design and operation of a receive-only liquid nitrogen (LN2)-cooled coil and cryostat suitable for medical imaging on a 3-T whole-body magnetic resonance scanner is presented. The coil size, optimized for murine imaging, was determined by using electromagnetic (EM) simulations. This process is therefore easier and more cost effective than building a range of coils. A nonmagnetic cryostat suitable for small-animal imaging was developed having good vacuum and cryogenic temperature performance. The LN2-cooled probe had an active detuning circuit allowing the use with the scanner's built-in body coil. External tuning and matching was adopted to allow for changes to the coil due to temperature and loading. The performance of the probe was evaluated by comparison of signal-to-noise ratio (SNR) with the same radio-frequency RF) coil operating at room temperature (RT). The performance of the RF coil at RT was also benchmarked against a commercial surface coil with a similar dimension to ensure a fair SNR comparison. The cryogenic coil achieved a 1.6- to twofold SNR gain for several different medical imaging applications: For mouse-brain imaging, a 100-mu m resolution was achieved in an imaging time of 3.5 min with an SNR of 25-40, revealing fine anatomical details unseen at lower resolutions for the same time. For heavier loading conditions, such as imaging of the hind legs and liver, the SNR enhancement was slightly reduced to 1.6-fold. The observed SNR was in good agreement with the expected SNR gain correlated with the loaded-quality factor of RF coils from the EM simulations. With the aid of this end-user-friendly and economically attractive cryogenic RF coil, the enhanced SNR available can be used to improve resolution or reduce the duration of individual scans in a number of biomedical applications

Functional and structural substrates of increased dosage of Grik4 gene elucidated using multi-modal MRI

Grik4 is the gene responsible for encoding the high-affinity GluK4 subunit of the kainate receptors. Increased dosage of this subunit in the forebrain was linked to an increased level of anxiety, lack of social communication, and depression. On the synaptic level, abnormal synaptic transmission was also reported. The manifestations of this abnormal expression have not been investigated at the circuit level, nor the correlations between those circuits and the abnormal patterns of the behavior previously reported. In this line of work, we aspired to use different non-invasive magnetic resonance imaging (MRI) modalities to elucidate any disturbance that might stem from the increased dosage of Grik4 and how those changes might explain the abnormal behaviors. MRI offers a noninvasive way to look into the intact brain in vivo. Resting-state functional MRI casts light on how the brain function at rest on the network level and has the capability to detect any anomalies that might occur within or between those networks. On the microstructural level, the diffusion MRI is concerned with the underlying features of the tissues, using the diffusion of water molecules as a proxy for that end. Moving more macroscopically, using structural scans, voxel-based morphometry can detect subtle differences in the morphology of the different brain structures. We recorded videos of our animals performing two tasks that have long been linked to anxiety, the open field and the plus-maze tests before acquiring structural and functional scans. Lastly, we recorded blood-oxygenationlevel dependent (BOLD) signals in a different set of animals during electrical stimulation of specific white matter tracts in order to investigate how neuronal activity propagates. Our analysis showed a vast spectrum of changes in the transgenic group relative to the animals in the control group. On the resting-state networks level, we observed an increase in the within-network strength spanning different structures such as the hippocampus, some regions of the cortex, and the hypothalamus. The increased internal coherence or strength in the networks contrasted with a significant reduction in between-networks connectivity for some regions such as parts of the cortex and the hypothalamus, suggesting long-range network decorrelation. Supporting this idea, major white matter (WM) tracts, such as the corpus callosum and the hippocampal commissure, suffered from substantial changes compatible with an important reduction in myelination and/or a decrease in the mean axonal diameter. Macrostructurally speaking, the overexpression of GluK4 subunit had a bimodal effect, with expansion in some cortical areas in the transgenic animals accompanied by a shrinkage in the subcortical regions. Upon stimulating the brain with an electrical current, we noticed a difference in activity propagation between the two hemispheres. In transgenic animals, the evoked activity remained more confined to the stimulated hemisphere, again consistent with an impaired long-range connectivity. The structural changes both, at the micro and macro level, were in tight correlation with different aspects of the behavior including markers of anxiety such as the time spent in the open arms vs the closed arms in the plus-maze test and the time spent in the center vs the corners in the open field test. Our findings reveal how the disruption of kainate receptors, or more globally the glutamate receptors, and the abnormal synaptic transmission can translate into brain-wide changes in connectivity and alter the functional equilibrium between macro-and mesoscopic networks. The postsynaptic enhancement previously reported in the transgenic animals was here reflected in the BOLD signal and measured as an increase in the within-network strength. Importantly, the correlations between the structural changes and the behavior help to put the developmental changes and their behavioral ramifications into context. RESUMEN Grik4 es el gen responsable de codificar la subunidad GluK4 de alta afinidad de los receptores de kainato. El aumento de la dosis de esta subunidad en el prosencéfalo se relacionó con un mayor nivel de ansiedad, falta de comunicación social y depresión. A nivel sináptico, también se informó una transmisión sináptica anormal. Las manifestaciones de esta expresión anormal no se han investigado a nivel de circuito, ni las correlaciones entre esos circuitos y los patrones anormales de la conducta previamente informada. En esta línea de trabajo, aspiramos a utilizar diferentes modalidades de imágenes por resonancia magnética (MRI) no invasivas para dilucidar cualquier alteración que pudiera derivarse del aumento de la dosis de Grik4 y cómo esos cambios podrían explicar los comportamientos anormales. La resonancia magnética ofrece una forma no invasiva de observar el cerebro intacto in vivo. La resonancia magnética funcional en estado de reposo arroja luz sobre cómo funciona el cerebro en reposo en el nivel de la red y tiene la capacidad de detectar cualquier anomalía que pueda ocurrir dentro o entre esas redes. En el nivel microestructural, la resonancia magnética de difusión se ocupa de las características subyacentes de los tejidos utilizando la difusión de moléculas de agua como un proxy para ese fin. Moviéndose más macroscópicamente, utilizando escaneos estructurales, la morfometría basada en vóxeles puede detectar diferencias sutiles en la morfología de las diferentes estructuras cerebrales. Grabamos videos de nuestros animales realizando dos tareas que durante mucho tiempo se han relacionado con la ansiedad, el campo abierto y las pruebas de laberinto positivo antes de adquirir escaneos estructurales y funcionales. Por último, registramos señales dependientes del nivel de oxigenación de la sangre (BOLD) en un grupo diferente de animales durante la estimulación eléctrica de tractos específicos de materia blanca para investigar cómo se propaga la actividad neuronal. Nuestro análisis mostró un amplio espectro de cambios en el grupo transgénico en relación con los animales en el grupo de control. En el nivel de las redes de estado de reposo, observamos un aumento en la fuerza dentro de la red que abarca diferentes estructuras como el hipocampo, algunas regiones de la corteza y el hipotálamo. La mayor coherencia interna o fuerza en las redes contrastó con una reducción significativa en la conectividad entre redes para algunas regiones como partes de la corteza y el hipotálamo, lo que sugiere una descorrelación de redes de largo alcance. Apoyando esta idea, los grandes tractos de materia blanca (WM), como el cuerpo calloso y la comisura del hipocampo, sufrieron cambios sustanciales compatibles con una importante reducción de la mielinización y / o una disminución del diámetro axonal medio. Macroestructuralmente hablando, la sobreexpresión de la subunidad GluK4 tuvo un efecto bimodal, con expansión en algunas áreas corticales en los animales transgénicos acompañada de una contracción en las regiones subcorticales. Al estimular el cerebro con una corriente eléctrica, notamos una diferencia en la propagación de la actividad entre las dos hemiesferas. En los animales transgénicos, la actividad evocada permaneció más confinada al hemisferio estimulado, de nuevo consistente con una conectividad de largo alcance deteriorada. Los cambios estructurales, tanto a nivel micro como macro, estaban en estrecha correlación con diferentes aspectos de la conducta, incluidos marcadores de ansiedad como el tiempo pasado con los brazos abiertos frente a los brazos cerrados en la prueba del laberinto positivo y el tiempo pasado en el centro vs las esquinas en la prueba de campo abierto. Nuestros hallazgos revelan cómo la interrupción de los receptores de kainato, o más globalmente los receptores de glutamato, y la transmisión sináptica anormal pueden traducirse en cambios de conectividad en todo el cerebro y alterar el equilibrio funcional entre las redes macro y mesoscópicas. La mejora postsináptica informada anteriormente en los animales transgénicos se reflejó aquí en la señal BOLD y se midió como un aumento en la fuerza dentro de la red. Es importante destacar que las correlaciones entre los cambios estructurales y elcomportamiento ayudan a contextualizar los cambios en el desarrollo y sus ramificaciones conductuales

Diffusion kurtosis imaging detects the time-dependent progress of pathological changes in the oral rotenone mouse model of Parkinson's disease

Clinical diagnosis of Parkinson's disease (PD) occurs typically when a substantial proportion of dopaminergic neurons in the substantia nigra (SN) already died, and the first motor symptoms appear. Therefore, tools enabling the early diagnosis of PD are essential to identify early-stage PD patients in which neuroprotective treatments could have a significant impact. Here, we test the utility and sensitivity of the diffusion kurtosis imaging (DKI) in detecting progressive microstructural changes in several brain regions of mice exposed to chronic intragastric administration of rotenone, a mouse model that mimics the spatiotemporal progression of PD-like pathology from the ENS to the SN as described by Braak's staging. Our results show that DKI, especially kurtosis, can detect the progression of pathology-associated changes throughout the CNS. Increases in mean kurtosis were first observed in the dorsal motor nucleus of the vagus (DMV) after 2 months of exposure to rotenone and before the loss of dopaminergic neurons in the SN occurred. Remarkably, we also show that limited exposure to rotenone for 2 months is enough to trigger the progression of the disease in the absence of the environmental toxin, thus suggesting that once the first pathological changes in one region appear, they can self-perpetuate and progress within the CNS. Overall, our results show that DKI can be a useful radiological marker for the early detection and monitoring of PD pathology progression in patients with the potential to improve the clinical diagnosis and the development of neuroprotective treatments. (Figure presented.). © 2021 International Society for Neurochemistr