[Evaluation of the survival of bone marrow mononucleate cells transplanted in a rat model of striatal lesion with quinolinic acid]. / Evaluacion de la supervivencia de las células mononucleadas de la médula ósea trasplantadas en un modelo de ratas con lesión estriatal por ácido quinolínico.

INTRODUCTION: Transplant is one of the alternatives available for the treatment of neurodegenerative diseases aimed at replacing the cells lost during the course of the disease. One promising source of cells for the development of transplants could be the mononucleate cells from bone marrow. AIMS. The purpose of this study was to study the capacity of bone marrow mononucleate cells to survive the transplant process, and to search for a method that enables tracking of these cells in vivo once they have been implanted. MATERIALS AND METHODS: Bone marrow mononucleate cells were extracted from the femur of rats by means of a Ficoll-Hypaque gradient. The cells under study were modified genetically with an adenovirus that expresses the PFV or which are marked with Hoechst dye. The marked cells were implanted in the striatum of rats with lesions caused by quinolinic acid. RESULTS: The viability of the genetically modified cells was low, whereas that of the cells marked with Hoechst dye was above 90%. The implanted cells survived the transplant at least a month and dispersed away from the site of entry towards the corpus callosum and cortex. CONCLUSIONS: We consider that the use of Hoechst dye offers more advantages for tracking these cells in vivo. Mononucleate cells have a number of characteristics that allow them to be included as candidate sources of cells for the treatment of neurodegenerative diseases.

Evaluación de la supervivencia de las células mononucleadas de la médula ósea trasplantadas en un modelo de ratas con lesión estriatal por ácido quinolínico / Evaluation of the survival of bone marrow mononucleate cells transplanted in a rat model of striatal lesion with quinolinic acid

Introducción. El trasplante es una de las alternativas para el tratamiento de enfermedades neurodegenerativas, y está encaminado hacia el reemplazo de las células perdidas durante el desarrollo de la enfermedad. Una fuente celular prometedora para el desarrollo de los trasplantes podrían ser las células mononucleadas de la médula ósea. Objetivo. Estudiar la capacidad de las células mononucleadas de la médula ósea de sobrevivir al trasplante y buscar un método que permita el seguimiento de estas células in vivo una vez implantadas. Materiales y métodos. Las células mononucleadas fueron extraídas del fémur de ratas mediante un gradiente de Ficoll-Hypaque. Las células objeto de estudio fueron modificadas genéticamente con un adenovirus que expresa la PFV o marcadas con el reactivo de Hoechst. Las células marcadas se implantaron en el estriado de ratas lesionadas con ácido quinolínico. Resultados. La viabilidad de las células modificadas genéticamente fue baja, mientras que la de las células marcadas con el reactivo de Hoechst fue superior al 90%. Las células implantadas sobrevivieron al trasplante al menos un mes y se dispersaron desde el sitio de entrada hacia el cuerpo calloso y la corteza. Conclusiones. Consideramos más ventajoso el uso del reactivo de Hoechst para el seguimiento de estas células in vivo. Las células mononucleadas tienen características que les permiten formar parte de las fuentes celulares candidatas para el tratamiento de las enfermedades neurodegenerativas

Introduction. Transplant is one of the alternatives available for the treatment of neurodegenerative diseases aimed at replacing the cells lost during the course of the disease. One promising source of cells for the development of transplants could be the mononucleate cells from bone marrow. Aims. The purpose of this study was to study the capacity of bone marrow mononucleate cells to survive the transplant process, and to search for a method that enables tracking of these cells in vivo once they have been implanted. Materials and methods. Bone marrow mononucleate cells were extracted from the femur of rats by means of a Ficoll-Hypaque gradient. The cells under study were modified genetically with an adenovirus that expresses the PFV or which are marked with Hoechst dye. The marked cells were implanted in the striatum of rats with lesions caused by quinolinic acid. Results. The viability of the genetically modified cells was low, whereas that of the cells marked with Hoechst dye was above 90%. The implanted cells survived the transplant at least a month and dispersed away from the site of entry towards the corpus callosum and cortex. Conclusions. We consider that the use of Hoechst dye offers more advantages for tracking these cells in vivo. Mononucleate cells have a number of characteristics that allow them to be included as candidate sources of cells for the treatment of neurodegenerative diseases

[Stromal cell transplant in the 6-OHDA lesion model]. / Trasplante de células estromales en el modelo de lesión por 6-OHDA.

INTRODUCTION: A good deal of evidence currently exists to show that transplanting foetal mesencephalic tissue can produce symptomatic benefits both in patients and in disease models. Nevertheless, the technical and ethical difficulties involved in obtaining enough suitable foetal cerebral tissue have been a serious obstacle to its application. Stromal cells derived from bone marrow, due to their potential capacity to generate different types of cells, could be an ideal source of material for cell restoration in neurodegenerative diseases. AIMS: Our aim was to evaluate the effect of transplanting stromal cells derived from bone marrow on the behaviour of 6-OHDA rats, when they are inserted into the striatum. MATERIAL AND METHODS: In this study we used rats with a lesion in the substantia nigra induced by 6-hydroxydopamine, divided into several experimental groups. Rotary activity induced by D-amphetamine (5 mg/kg, intraperitoneally) was evaluated before and throughout the three months following the transplant in all the experimental groups, except in the group of healthy controls. Hemiparkinsonian rats received a total of 350 000 foetal ventral mesencephalic cells and 8 x 10(4) stromal cells/microL, which were implanted in the striatum. RESULTS AND CONCLUSIONS: Animals with stromal cells transplanted in the body of the striatum significantly reduced the number of turns induced by amphetamine (p < 0.05); yet this reduction was not greater than that induced by foetal mesencephalic cell transplants. We were also unable to demonstrate any significant improvement in the motor skills of the forelimbs.

[Lesions in the pars compacta substantiae nigra and the subthalamic nucleus modify the density of muscarinic receptors in different nuclei of the basal ganglia]. / La lesión de la sustancia negra pars compacta y del núcleo subtalámico modifica la densidad de receptores muscarínicos en distintos núcleos de los ganglios basales.

INTRODUCTION: Several studies that has focused to the dopaminergic transmission in the basal ganglia in parkinsonian condition, but only a few article has taking into account the imbalance between dopaminergic and cholinergic transmission. OBJECTIVE: To evaluate the muscarinic cholinergic receptors density in SNc and PPN in the 6-OHDA model. MATERIALS AND METHODS: Were organized five experimental groups in correspondence to the place of the lesion: I. Non treated rats, II. 6-OHDA lesion in SNc, III. 6-OHDA lesion in SNc + quinolinic acid lesion in NST, IV. Sham operated rats, V. Quinolinic acid in STN. Were obtained coronal sections of 20 microm thickness of SNc and PPN from rats and in these sections was evaluated the muscarinic receptors density through autoradiographic technique with [3H]quinuclidinylbenzilate (QNB) (1.23 nM). The muscarinic antagonist atropine (1 microM) was utilized as non-specific union. The density was evaluated in both hemispheres and the density optical was converted in fentomolas/mg of tissue with base to values obtained from tritium standards. RESULTS: Significant diminution of the muscarinic receptors density was found in the SNc ipsilateral to the 6-OHDA lesion from experimental groups II (t=2.76; p<0.05) and III (t=4.06; p<0.05). In the group V, was seen a significant increase of muscarinic receptor density in the SNc ipsilateral to the 6-OHDA lesion. The comparison between experimental groups evidenced significant differences among them (F=13.13; p<0.001) with a significant decrease in the density from SNc of groups II and III and significant increase in the density from SNc of group V in comparison of the others groups. In relation to PPN, muscarinic receptors density from right PPN ipsilateral to the 6-OHDA lesion, shown significant differences (F=3.93; p<0.01) between the experimental groups with a significant increase of this variable in the group II. CONCLUSIONS: These results signal a modification of cholinergic activity after 6-OHDA lesion. The changes in the muscarinic receptors populations located in SNc and PPN could be part of different compensatory mechanisms to attempt ameliorate the imbalance between dopaminergic and cholinergic transmission that it was installed after denervation of nigrostriatal forebrain bundle. The excitotoxic lesion of STN impose a new adjust mechanism for cell from PPN, which could be expressed in the changes of muscarinic cholinergic receptors population at the level of SNc.

[Comparative study of bilateral lesions in the entorhinal cortex and in the fimbria fornix]. / Estudio comparativo de la lesión bilateral de corteza entorrinal y de la fimbria-fórnix.

INTRODUCTION: Numerous reports show that lesions to hippocampus afferents, such as the entorhinal cortex (EC) and the fimbria fornix (FF), exert an effect on memory in rodents. There are, however, no long term comparative studies that show which of these lesions could be most useful as a model for studies into neuroplasticity. MATERIAL AND METHODS: Young male Sprague Dawley rats were used. Bilateral electrolytic lesion was caused to the EC or the FF was damaged by transection. One, four or 12 weeks later the animals were evaluated in a Morris water maze, first with an invisible platform and then with the platform within view. The results from the two groups were compared to each other and to those obtained from healthy controls and subjects with false lesions by means of a variance analysis. RESULTS: In the test with an invisible platform, both types of lesion gave rise to serious, irreparable involvement of the spatial memory of the animals, at least up to 12 weeks after the lesion. The test with the visible platform revealed significant differences between animals with lesion to the EC evaluated at 12 weeks, which suggests the development of some visual or motor deterioration in these animals. CONCLUSIONS: Although both lesions gave rise to behavioural deterioration that was irreversible in the long term in rodents, the lesion to the FF seems to be a better model for evaluating specific effects on learning and memory, since the lesion to the EC apparently triggers additional sensory and motor involvement.

[The action of epidermal growth factor on the development of cultured striatum cells]. / Acción del factor de crecimiento epidérmico sobre el desarrollo de las células estriatales en cultivo.

INTRODUCTION: Epidermic growth factor (EGF) has a neurotrophic mitogenic effect on different cell populations in the nervous system. This is modulated by the stage of development and microenvironment of the cells. OBJECTIVE AND METHODS: In this paper we describe the action of EGF on embryonic striatum cells of a culture system dissociated from neurons and glias. The cell culture is prepared from 16-17 day rat embryos. In the system used, the cell population was cultured for 20-24 hours in a medium containing serum. This medium was later replaced by a mixture of specific nutrients and treated for 6 days with 20 mg/ml of EGF. RESULTS AND CONCLUSIONS: The substitution of serum during the initial period of development led to an appreciable reduction in the living cells in the treated cultures and in the controls. The surviving cells were mainly cellular precursors, taking into account their morphological characteristics and capacity for proliferation. The effect of EGF was seen in an increase in the number of cells and was shown to be a stimulus to the proliferation of neuronal and astrocyte precursors. The specific activity of choline acetyl-transferases determined in the cultures at 16 days showed differentiation of a cholinergic neurons subpopulation, which responded to treatment with nerve growth factor with an increase in the activity of this enzyme.

Acción del factor de crecimiento epidérmico sobre el desarrollo de las células estriatales en cultivo / Action of the epidermal growth factor on the development of striatal cell culture

Introducción. El factor de crecimiento epidérmico (FCE) ejerce un efecto neurotrófico o mitogénico sobre diferentes poblaciones celulares en el sistema nervioso, lo que está modulado por el estadío de desarrollo celular y el microambiente donde se desarrollan las células. Objetivo y métodos. En este trabajo se describe la acción del FCE sobre las células del estriado embrionario en un sistema de cultivo disociado de neuronas y glias. El cultivo de células se preparó a partir de embriones de ratas de 16-17 días. En el sistema empleado la población celular fue cultivada durante 20-24 horas en un medio que contenía suero y posteriormente se sutituyó este por una mezcla de nutrientes definidos y se trató con 20 ng/ml de FCE durante 6 días. Resultados y conclusiones. La sustitución del suero en este período inicial de desarrollo provocó una disminución apreciable de las células vivas en los cultivos tratados y en los controles. Las células sobrevivientes estaban representadas mayoritariamente por precursores celulares teniendo en cuenta sus características morfológicas y su capacidad proliferativa. El efecto del FCE se manifestó en un aumento del número de células y se demostró un estímulo de la proliferación de los precursores neuronales y de los astrocitos. La actividad específica de la colina acetiltransferasa determinada en los cultivos a los 16 días evidenció la diferenciación de una subpoblación neuronal colinérgica, la cual respondió al tratamiento con el factor de crecimiento nervioso con un aumento de la actividad de esta enzima