Métodos automatizados de segmentación para cuantificar el grosor cortical en RM en pacientes con deterioro cognitivo y enfermedad de Parkinson

El objetivo del presente estudio es encontrar la relación que existe entre los síntomas de deterioro cognitivo y el grosor cortical cerebral en una muestra de 45 pacientes con Enfermedad de Parkinson diagnosticados y tratados en la consulta externa del Servicio de Neurología del Hospital de la Santa Creu i Sant Pau de Barcelona, quienes fueron sometidos a tests neuropsicológicos que evaluaban su función mental y motora y a quienes se les realizó un estudio de Resonancia Magnética 3 Tesla para evaluar, mediante técnicas de postprocesados, su grosor cortical. Los resultados demostraron una clara disminución del grosor cortical en determinadas áreas que se correlacionaban con las funciones cognitivas afectadas en estos pacientes.

Cortical mapping of the neuronal circuits modulating the muscle tone. Introduction to the electrophysiological treatment of the spastic hand.

L’objectiu d’aquest estudi es investigar l’organització cortical junt amb la connectivitat còrtico-subcortical en subjectes sans, com a estudi preliminar. Els mapes corticals s’han fet per TMS navegada, i els punts motors obtinguts s’han exportant per estudi tractogràfic i anàlisi de las seves connexions. El coneixement precís de la localització de l’àrea cortical motora primària i les seves connexions es la base per ser utilitzada en estudis posteriors de la reorganització cortical i sub-cortical en pacients amb infart cerebral. Aquesta reorganització es deguda a la neuroplasticitat i pot ser influenciada per els efectes neuromoduladors de la estimulació cerebral no invasiva.

Influence of the in vivo deregulation of the expression of ntrk3 (trkc) on cortical development of a murine model of panic/anxiety disorder

Alteracions durant el desenvolupament cerebral produirien canvis en la connectivitat neuronal i la bioquímica cel•lular que podrien resultar en una disfunció cognitiva i/o emocional, desembocant a trastorns psiquiàtrics. Les neurotrofines intervenen en els processos del neurodesenvolupament i en la funcionalitat del cervell adult i, conseqüentment, serien bons candidats com a factors de predisposició en diverses malalties mentals. S’ha suggerit la implicació del receptor de la neurotrofina 3, TrkC, en el trastorn de pànic. Nosaltres proposem que la sobreexpressió del gen NTRK3 (TrkC) és un mediador comú dels desencadenants genètics i ambientals d’aquest trastorn. Concretament, la seva desregulació podria produir canvis estructurals i funcionals a l’escorça cerebral dels pacients pel seu paper durant l’establiment dels circuïts corticals i la neuroplasticitat a l’adult, probablement esdevenint elements de predisposició a patir atacs de pànic. Els objectius principals d’aquest treball han estat: 1/determinar la contribució específica del gen NTRK3 a les alteracions de l’escorça cerebral observades en pacients, utilitzant un model murí modificat genèticament (TgNTRK3), i 2/analitzar l’impacte específic de la sobreexpressió de NTRK3 sobre la corticogènesi durant estadis embrionaris o postnatals estudiant la neurogènesi i la neuritogènesi. Els resultats indiquen que la sobreexpressió de NTRK3 als ratolins produeix una reducció del gruix de l’escorça frontal, recapitulant la hipofrontalitat dels pacients, que comportaria una menor inhibició dels nuclis subcorticals del sistema límbic com l’amígdala, i alteracions citoarquitectòniques a l’escorça prefrontal medial que recolzen la hipòtesi del seu mal funcionament. Tanmateix, els ratolins TgNTRK3 presenten canvis estructurals a l’escorça somatosensorial, suggerint que el processament de la informació sensorial podria estar alterat, el que encara no s’ha explorat en pacients. La sobreexpressió de NTRK3 també afecta la neuritogènesi en cultius primaris corticals i modifica la resposta de les neurones a l’estimulació amb neurotrofines. Per tant, el fenotip cortical adult dels TgNTRK3 podria dependre d’alteracions durant la corticogènesi.

Impaired fast-spiking, suppressed cortical inhibition and increased susceptibility to seizures in mice lacking Kv3.2 K+ channel proteins

Voltage-gated K+ channels of the Kv3 subfamily have unusual electrophysiological properties, including activation at very depolarized voltages (positive to −10 mV) and very fast deactivation rates, suggesting special roles in neuronal excitability. In the brain, Kv3 channels are prominently expressed in select neuronal populations, which include fast-spiking (FS) GABAergic interneurons of the neocortex, hippocampus, and caudate, as well as other high-frequency firing neurons. Although evidence points to a key role in high-frequency firing, a definitive understanding of the function of these channels has been hampered by a lack of selective pharmacological tools. We therefore generated mouse lines in which one of the Kv3 genes, Kv3.2, was disrupted by gene-targeting methods. Whole-cell electrophysiological recording showed that the ability to fire spikes at high frequencies was impaired in immunocytochemically identified FS interneurons of deep cortical layers (5-6) in which Kv3.2 proteins are normally prominent. No such impairment was found for FS neurons of superficial layers (2-4) in which Kv3.2 proteins are normally only weakly expressed. These data directly support the hypothesis that Kv3 channels are necessary for high-frequency firing. Moreover, we found that Kv3.2 −/− mice showed specific alterations in their cortical EEG patterns and an increased susceptibility to epileptic seizures consistent with an impairment of cortical inhibitory mechanisms. This implies that, rather than producing hyperexcitability of the inhibitory interneurons, Kv3.2 channel elimination suppresses their activity. These data suggest that normal cortical operations depend on the ability of inhibitory interneurons to generate high-frequency firing.

Regenerating cortical connections in a dish: the entorhino-hippocampal organotypic slice co-culture as tool for pharmacological screening of molecules promoting axon regeneration

We present a method for using long-term organotypic slice co-cultures of the entorhino-hippocampal formation to analyze the axon-regenerative properties of a determined compound. The culture method is based on the membrane interphase method, which is easy to perform and is generally reproducible. The degree of axonal regeneration after treatment in lesioned cultures can be seen directly using green fluorescent protein (GFP) transgenic mice or by axon tracing and histological methods. Possible changes in cell morphology after pharmacological treatment can be determined easily by focal in vitro electroporation. The well-preserved cytoarchitectonics in the co-culture facilitate the analysis of identified cells or regenerating axons. The protocol takes up to a month.

Cortistain is expressed in a distinct subset of cortical interneurons

Cortistatin is a presumptive neuropeptide that shares 11 of its 14 amino acids with somatostatin. In contrast to somatostatin, administration of cortistatin into the rat brain ventricles specifically enhances slow wave sleep, apparently by antagonizing the effects of acetylcholine on cortical excitability. Here we show that preprocortistatin mRNA is expressed in a subset of GABAergic cells in the cortex and hippocampus that partially overlap with those containing somatostatin. A significant percentage of cortistatin-positive neurons is also positive for parvalbumin. In contrast, no colocalization was found between cortistatin and calretinin, cholecystokinin, or vasoactive intestinal peptide. During development there is a transient increase in cortistatin-expressing cells in the second postnatal week in all cortical areas and in the dentate gyrus. A transient expression of preprocortistatin mRNA in the hilar region at P16 is paralleled by electrophysiological changes in dentate granule cells. Together, these observations suggest mechanisms by which cortistatin may regulate cortical activity.

Neurites regrowth of cortical neurons by GSK3b inhibition independently of Nogo Receptor 1

Lesioned axons do not regenerate in the adult mammalian central nervous system, owing to the overexpression of inhibitory molecules such as myelin-derived proteins or chondroitin sulphate proteoglycans. In order to overcome axon inhibition, strategies based on extrinsic and intrinsic treatments have been developed. For myelin-associated inhibition, blockage with NEP1-40, receptor bodies or IN-1 antibodies has been used. In addition, endogenous blockage of cell signalling mechanisms induced by myelin-associated proteins is a potential tool for overcoming axon inhibitory signals. We examined the participation of glycogen synthase kinase 3 (GSK3) and ERK1/2 in axon regeneration failure in lesioned cortical neurons. We also investigated whether pharmacological blockage of GSK3 and ERK1/2 activities facilitates regeneration after myelin-directed inhibition in two models: i) cerebellar granule cells and ii) lesioned entorhino-hippocampal pathway in slice cultures, and whether the regenerative effects are mediated by Nogo Receptor 1 (NgR1). We demonstrate that, in contrast to ERK1/2 inhibition, the pharmacological treatment of GSK3 inhibition strongly facilitated regrowth of cerebellar granule neurons over myelin independently of NgR1. Lastly these regenerative effects were corroborated in the lesioned EHP in NgR1 -/- mutant mice. These results provide new findings for the development of new assays and strategies to enhance axon regeneration in injured cortical connections.

BS-SEM evaluation of the tisular interactions between cortical bone and calcium-phosphate covered titanium implants

The improvement of the reliability of the contact between the osseous tissues and the implant materials has been tested by recovering the metallic implants with ceramic materials, usually calcium phosphates. In our study, the calcium phosphate recovering layers were deposited by means of a pulsed-laser deposition technique. Our aim was to to evaluate the tissue interactions established between cortical bone and titanium implants covered by five different layers, ranging from amorphous calcium phosphate to crystalline hydroxyapatite, obtained by altering the parameters of the laser ablation process. The surgical protocol of the study consisted in the simultaneous implantation of the five types of implants in both the tibial dyaphisis of three Beagle dogs, sacrificed respectively one, two and three months after the last surgical procedures. After the sacrifice, the samples were submitted to a scheduled procedure of embedding in plastic polymers without prior decalcification, in order to perform the ultrastructural studies: scanning microscopy with secondary and backscattered electrons (BS-SEM). Our observations show that both in terms of the calcified tissues appearing as a response to the presence of the different coatings and of time of recovering, the implants coated with crystalline calcium phosphate layers by laser ablation present a better result than the amorphous-calcium-phosphate-coated implants. Moreover, the constant presence of chondroid tissue, related with the mechanical induction by forces applied on the recovering area, strongly suggests that the mechanisms implied in osteointegration are related to endomembranous, rather than endochondral ossification processes

BS-SEM evaluation of the tisular interactions between cortical bone and calcium-phosphate covered titanium implants

The improvement of the reliability of the contact between the osseous tissues and the implant materials has been tested by recovering the metallic implants with ceramic materials, usually calcium phosphates. In our study, the calcium phosphate recovering layers were deposited by means of a pulsed-laser deposition technique. Our aim was to to evaluate the tissue interactions established between cortical bone and titanium implants covered by five different layers, ranging from amorphous calcium phosphate to crystalline hydroxyapatite, obtained by altering the parameters of the laser ablation process. The surgical protocol of the study consisted in the simultaneous implantation of the five types of implants in both the tibial dyaphisis of three Beagle dogs, sacrificed respectively one, two and three months after the last surgical procedures. After the sacrifice, the samples were submitted to a scheduled procedure of embedding in plastic polymers without prior decalcification, in order to perform the ultrastructural studies: scanning microscopy with secondary and backscattered electrons (BS-SEM). Our observations show that both in terms of the calcified tissues appearing as a response to the presence of the different coatings and of time of recovering, the implants coated with crystalline calcium phosphate layers by laser ablation present a better result than the amorphous-calcium-phosphate-coated implants. Moreover, the constant presence of chondroid tissue, related with the mechanical induction by forces applied on the recovering area, strongly suggests that the mechanisms implied in osteointegration are related to endomembranous, rather than endochondral ossification processes

BS-SEM evaluation of the tisular interactions between cortical bone and calcium-phosphate covered titanium implants

The improvement of the reliability of the contact between the osseous tissues and the implant materials has been tested by recovering the metallic implants with ceramic materials, usually calcium phosphates. In our study, the calcium phosphate recovering layers were deposited by means of a pulsed-laser deposition technique. Our aim was to to evaluate the tissue interactions established between cortical bone and titanium implants covered by five different layers, ranging from amorphous calcium phosphate to crystalline hydroxyapatite, obtained by altering the parameters of the laser ablation process. The surgical protocol of the study consisted in the simultaneous implantation of the five types of implants in both the tibial dyaphisis of three Beagle dogs, sacrificed respectively one, two and three months after the last surgical procedures. After the sacrifice, the samples were submitted to a scheduled procedure of embedding in plastic polymers without prior decalcification, in order to perform the ultrastructural studies: scanning microscopy with secondary and backscattered electrons (BS-SEM). Our observations show that both in terms of the calcified tissues appearing as a response to the presence of the different coatings and of time of recovering, the implants coated with crystalline calcium phosphate layers by laser ablation present a better result than the amorphous-calcium-phosphate-coated implants. Moreover, the constant presence of chondroid tissue, related with the mechanical induction by forces applied on the recovering area, strongly suggests that the mechanisms implied in osteointegration are related to endomembranous, rather than endochondral ossification processes

In cortical neurons HDAC3 suppresses RD4-dependent SMRT export

The transcriptional corepressor SMRT controls neuronal responsiveness of several transcription factors and can regulate neuroprotective and neurogenic pathways. SMRT is a multi-domain protein that complexes with HDAC3 as well as being capable of interactions with HDACs 1, 4, 5 and 7. We previously showed that in rat cortical neurons, nuclear localisation of SMRT requires histone deacetylase activity: Inhibition of class I/II HDACs by treatment with trichostatin A (TSA) causes redistribution of SMRT to the cytoplasm, and potentiates the activation of SMRT-repressed nuclear receptors. Here we have sought to identify the HDAC(s) and region(s) of SMRT responsible for anchoring it in the nucleus under normal circumstances and for mediating nuclear export following HDAC inhibition. We show that in rat cortical neurons SMRT export can be triggered by treatment with the class I-preferring HDAC inhibitor valproate and the HDAC2/3-selective inhibitor apicidin, and by HDAC3 knockdown, implicating HDAC3 activity as being required to maintain SMRT in the nucleus. HDAC3 interaction with SMRT's deacetylation activation domain (DAD) is known to be important for activation of HDAC3 deacetylase function. Consistent with a role for HDAC3 activity in promoting SMRT nuclear localization, we found that inactivation of SMRT's DAD by deletion or point mutation triggered partial redistribution of SMRT to the cytoplasm. We also investigated whether other regions of SMRT were involved in mediating nuclear export following HDAC inhibition. TSA- and valproate-induced SMRT export was strongly impaired by deletion of its repression domain-4 (RD4). Furthermore, over-expression of a region of SMRT containing the RD4 region suppressed TSA-induced export of full-length SMRT. Collectively these data support a model whereby SMRT's RD4 region can recruit factors capable of mediating nuclear export of SMRT, but whose function and/or recruitment is suppressed by HDAC3 activity. Furthermore, they underline the fact that HDAC inhibitors can cause reorganization and redistribution of corepressor complexes.

The natural hallucinogen 5-MeO-DMT, component of Ayahuasca, disrupts cortical function in rats: reversal by antipsychotic drugs

5-Methoxy-N,N-dimethyltryptamine (5-MeO-DMT) is a natural hallucinogen component of Ayahuasca, an Amazonian beverage traditionally used for ritual, religious and healing purposes that is being increasingly used for recreational purposes in US and Europe. 5MeO-DMT is of potential interest for schizophrenia research owing to its hallucinogenic properties. Two other psychotomimetic agents, phencyclidine and 2,5-dimethoxy-4-iodo-phenylisopropylamine (DOI), markedly disrupt neuronal activity and reduce the power of low frequency cortical oscillations (<4 Hz, LFCO) in rodent medial prefrontal cortex (mPFC). Here we examined the effect of 5-MeO-DMT on cortical function and its potential reversal by antipsychotic drugs. Moreover, regional brain activity was assessed by blood-oxygen level dependent (BOLD) functional magnetic resonance imaging (fMRI). 5-MeO-DMT disrupted mPFC activity, increasing and decreasing the discharge of 51 and 35% of the recorded pyramidal neurons, and reducing (−31%) the power of LFCO. The latter effect depended on 5-HT1A and 5-HT2A receptor activation and was reversed by haloperidol, clozapine, risperidone, and the mGlu2/3 agonist LY379268. Likewise, 5-MeO-DMT decreased BOLD responses in visual cortex (V1) and mPFC. The disruption of cortical activity induced by 5-MeO-DMT resembles that produced by phencyclidine and DOI. This, together with the reversal by antipsychotic drugs, suggests that the observed cortical alterations are related to the psychotomimetic action of 5-MeO-DMT. Overall, the present model may help to understand the neurobiological basis of hallucinations and to identify new targets in antipsychotic drug development.

Estudi de les alteracions en la capacitat neuroprotectora dels astròcits reactius en l'envelliment

En el periodo 2005-2008 hemos publicado tres artículos sobre las alteraciones de los astrocitos reactivos en el cerebro durante el envejecimiento. En el primer estudio, evaluamos la capacidad neuroprotectora de los astrocitos en un modelo experimental in vitro de envejecimiento. Los cambios en el estrés oxidativo, la captación del glutamato y la expresión proteica fueron evaluados en los astrocitos corticales de rata cultivados durante 10 y 90 días in vitro (DIV). Los astrocitos envejecidos tenían una capacidad reducida de mantener la supervivencia neuronal. Estos resultados indican que los astrocitos pueden perder parcialmente su capacidad neuroprotectora durante el envejecimiento. En el segundo estudio el factor neurotrófico derivado de la línea glial (GDNF) fue probado para observar sus efectos neurotróficos contra la atrofia neuronal que causa déficits cognitivos en la vejez. Las ratas envejecidas Fisher 344 con deficiencias en el laberinto de Morris recibieron inyecciones intrahippocampales de un vector lentiviral que codifica GDNF humano en los astrocitos o del mismo vector que codifica la proteína fluorescente verde humana como control. El GDNF secretado por los astrocitos mejoró la función de la neurona como se muestra por aumentos locales en la síntesis de los neurotransmisores acetilcolina, dopamina y serotonina. El aprendizaje espacial y la prueba de memoria demostraron un aumento significativo en las capacidades cognitivas debido a la exposición de GDNF, mientras que las ratas control mantuvieron sus resultados al nivel del azar. Estos resultados confirman el amplio espectro de la acción neurotrófica del GDNF y abre nuevas posibilidades de terapia génica para reducir la neurodegeneración asociada al envejecimiento. En el último estudio, examinamos cambios en la fosforilación de tau, el estrés oxidativo y la captación de glutamato en los cultivos primarios de astrocitos corticales de ratones neonatos de senescencia acelerada (SAMP8) y ratones resistentes a la senescencia (SAMR1). Nuestros resultados indican que las alteraciones en cultivos del astrocitos de los ratones SAMP8 son similares a las detectadas en cerebros enteros de los ratones SAMP8 de 1-5 meses de edad. Por otra parte, nuestros resultados sugieren que esta preparación in vitro es adecuada para estudiar en este modelo murino el envejecimiento temprano y sus procesos moleculares y celulares.

Estudi dels factors mol•leculars i cel•lulars implicats en les alteracions corticals de la Síndrome de down

Investigación producida a partir de una estancia en en el Instituto de Neurociencias de la Universidad Miguel Hernández entre enero y mayo del 2007. El SD o trisomía del cromosoma 21 es la aneuploidía cromosómica más frecuente y constituye la principal causa de retraso mental. Las cuestiones que aún son objeto de debate en el SD son: 1) si pueden existir, entre los genes triplicados, algunos que contribuyan de forma más importante a algunos de los fenotipos observables en SD y, 2) hasta qué punto los fenotipos observados derivan de alteraciones del neurodesarrollo o de alteraciones funcionales en el adulto. Con el fin de abordar esta cuestión nos hemos centrado en las alteraciones cognitivas del SD y hemos realizado la caracterización del papel de Dyrk1A en el desarrollo de una estructura clave para esta función: la corteza cerebral. Los resultados obtenidos muestran que la sobrexpresión de Dyrk1A produce un desajuste proliferativo dando lugar a un retraso en la formación de la subplaca, con consecuencias en la laminación de la placa cortical. Las alteraciones en la corticogénesis van a tener consecuencias en el establecimiento de la conectividad tálamo-cortical que se encuentra marcadamente retrasada. En el hipocampo, los ratones transgénicos mostraron una reducción del grosor de las capas. Estos resultados pueden ser relevantes para el SD, puesto que es similar a lo observado en fetos SD.

Hiperparatiroidismo primario en mujeres postmenopáusicas: evolución de la densidad mineral ósea tras tratamiento quirúrgico

Es realitza un estudi prospectiu observacional amb 158 dones postmenopàusiques diagnosticades d’hiperparatiroidisme primari. L’objectiu és descriure i analitzar l’evolució de la densitat mineral òssia d’aquestes dones després de ser sotmeses a tractament quirúrgic. Els resultats mostren que l’os més afectat en aquesta malaltia és l’os cortical, però el que més es recupera i de forma més ràpida després de la paratiroidectomia és l’os trabecular. Així mateix, les malaltes que tenen un hiperparatiroidisme més sever presenten una major recuperació de la densitat mineral òssia després de la cirurgia.