Thursday 26 February 2004

s of the 2nd Scientific Conference "Restauración Neurológica 2004" 252 Centro Internacional de Restauración Neurológica, Ciudad de la Habana, Cuba. Instituto Cajal, CSIC, Madrid, Spain Treatment with L-DOPA is today the most efficacious, non-invasive therapy for Parkinson’s Disease. However, chronic treatment with L-DOPA induces in most of the patients the appearance of abnormal involuntary movements known as dyskinesias. The molecular mechanisms underlying these abnormal movements are unknown, although the implication of dopamine receptors as well as other neurotransmitter receptors interacting with the system is suspected. In the present work, we have studied the contribution of D1 and D2 receptor subtypes in levodopa-induced dyskinesia. In this experiment dyskinesias were induced with intermittent doses of L-DOPA in unilateral 6-OHDA-lesioned mice. Rotational tests were carried out in a rotometer system that counted completed turns (360o) only and the abnormal involuntary movements we have divided in four different types: orofacial, forelimb, locomotive and dystonia, and evaluated. In wild-type animals chronic treatment with L-DOPA induced abnormal involuntary movements that including horizontal or vertical abnormal jaw movements, ballistic movements of the contralateral forelimb and axial dystonia. We have found that inactivation of D1 dopamine receptors completely abolished orofacial dyskinesia, did not affect axial distonia and slightly reduced forelimb dyskinesia. By contrast, knock out mice lacking D2 receptors showed an increase in orofacial dyskinesia after LDOPA treatment but axial dystonia and forelimb dyskinesia were not present. In summary, our result demonstrate that D1 and D2 receptors play a differential and complementary a role in L-DOPA-induced dyskinesia and that the integrity of the D1 and D2 receptors is critical for different aspects of levodopainduced dyskinesias 26A4 Ex vivo VEGF gene transfer does not increase grafted dopamine neuron survival C.E. Sortwell, W.J. Bowers, H.J. Federoff, M.F. Fleming, D.M. Marchionini, N.M Kanaan, B. Terpstra, K. A. Steece-Collier and T.J. Collier Neurology, Rush-Presbyterian, Chicago, IL, USA; Neurology., Univ. Rochester, Rochester, NY, USA Specific conditions associated with the posttransplantation interval render grafted mesencephalic dopamine (DA) neurons susceptible to apoptotic and necrotic forms of cell death. Immediately following transplantation, grafted cells are dependent upon diffusion of oxygen and blood-borne materials from the host vasculature until they establish circulation with the host brain. Vascular endothelial growth factor (VEGF) is the primary endogenous endothelial cell mitogen involved in both vasculogenesis and angiogenesis. The present study investigates the ability of ex vivo transduction of mesencephalic reaggregates with a helper virus-free Herpes simplex virus (HSV) amplicon vector encoding VEGF to increase DA neuron survival after transplantation. Mesencephalic reaggregates were generated from E14 F344 rat pups and transduced immediately with either helper virusfree HSV-vegf, HSV-lac (MOI = 1.0 or 2.0) or no vector. Four days post-transduction (PTD4) reaggregates were analyzed for VEGF release to the culture media and levels of VEGF within the aggregates themselves using immunoassay and grafted to the denervated striatum of Fisher 344 rats. On PTD4 HSV-vegf-transduced reaggregates released at least 250 fold more VEGF protein into the culture media and contained at least 130 fold more VEGF protein than HSV-lac or non-transduced control reaggregate cultures. Although these in vitro assays yielded promising results, results of HSV-vegf ex vivo transduction of grafted mesencephalic reaggregates on behavioral and morphological outcome measures were not positive. The present results demonstrate that ex vivo gene transfer of VEGF is not successful in augmenting the survival of mesencephalic grafts. Supported by: AG21546, AG000844, Rochester Nathan Shock Center. 26A5 Administration of different doses of methylprednisolone in adult rats with traumatic spinal cord injury of different intensity and at different spinal cord levels H. Salgado-Ceballos,b, S. Torres-Castillo,c, J.L. Torres-Vera, D. Zabaleta, A. Zavala, S. OrozcoSuárez, R. Capin, A. Feria-Velasco Research Medical Unit in Neurological Diseases, CMN, S. XXI, IMSS, Proyecto Camina A.C., Metropolitan Autonomous University, Iztapalapa, CIATEJ SEP-CONACYT,Research Unit in Oncologyc Diseases, CMN, S. XXI, IMSS, Mexico city, Mexico After series of preclinical and clinical works, methylprednisolone (MP) became the standard of care in traumatic spinal cord injury (TSCI). However, in some studies MP failed to demonstrate consistent and significant effect on functional outcome. With the aim to test if the injury intensity or the injured level Abstracts of the 2nd Scientific Conference "Restauración Neurológica 2004" 253s of the 2nd Scientific Conference "Restauración Neurológica 2004" 253 could affect the response to the MP treatment, adult rats were subjected to mild (30 g/cm), moderate (75 g/cm) or severe (150 g/cm) TSCI at cervical (C6), thoracic (T9) or lumbar (L2) level using a standardized weight-drop contusion model. Groups were divided into subgroups that received vehicle or MP at 15, 30 or 45 mg/kg of body weight. Using the thiobarbituric acid reaction, malondialdehyde (MDA) levels, resulting from lipid peroxidation were obtained. Morphometric studies were done by light and electron microscopy and motor function using the BBB scale. Mild thoracic and mild and moderate lumbar injuries showed motor recovery with MP at 15mg/kg, while 30 and 45 mg/kg were as deleterious as in cervical and lumbar severe injuries. Non motor recovery was observed in moderate cervical and thoracic injuries, where the mortality was increased. In severe thoracic injuries motor recovery was observed with all doses of MP, specially with 30 mg/kg (p<0.03). However, in all groups where MP produced motor recovery by protecting axons and myelin sheaths from the secondary injury, it inhibited axonal collateral emission. MDA levels correlated with morphometric parameters and motor recovery. Response to MP treatment after TSCI seams to depend of the intensity and the injury level. 26A6 The administration of two new derivates of xanthines (A15ET and A15BU) reversed the locomotor impairments in rats with dopaminergic lesion Carmen Parra, Miriam Alonso, Rogelio Maxil, Alejandro Muñoz, Jesús Sandoval, Daniel Limón Laboratorio de Neurofarmacologia, Facultad de Ciencias Químicas-BUAP, Centro de Investigaciones FCQ-Benemérita Universidad Antónima de Puebla. Puebla, México In Parkinson’s disease therapy is symptomatic. The L-dopa like is alternative in this pathology when is administrated by long time. Because of it is necessary to research new drugs about this pathology. One of them are the A2A antagonists that improve some Parkinson motor behavior in patient. In our University new derivates of xanthines like A15Et and A15Bu have been synthesized. The aim of this work was to evaluate the effect of new A2A antagonist (Al5Et and A15Bu) on motor behavior in rats with dopaminergic lesion. One group the rats were injected with 2 ml of 6-OHDA(8mg/ml) in SNc. The lesioned groups were evaluated in the drug induced turning behavior model, and in this condition were administrated A15Et or A15Bu (1mg/kg). The A15Bu was also evaluated in the star case test. Others groups of intact rats were evaluated in locomotors activity in and cataleptic behavior. We found that new A2A antagonist A15Et and A15Bu decreased 50% and 25% the turning behavior 70 and 80 minutes after administration. In the staircase the ability of reach pellets improved in 98%, the catalepsy behavior was reversed by injection of A15Bu 50% and did not modify the locomotor activity in intact rats. These data suggests that A15Et and A15Bu improve the motor function of rats with dopaminergic lesion. 26A7 Calpain inhibitor prevents axonal degeneration J.N. Hokoç, L.A Couto; M.S. Narciso, and A.M.B. Martinez Instituto de Biofísica Carlos Chagas Filho and Departamento de Embriologia e Histologia, ICB, Universidade Federal do Rio de Janeiro, Rio de Janeiro, Brazil The ultrastructural change that characterizes the onset of Wallerian degeneration is the disintegration of axoplasmic microtubules and neurofilaments, followed by myelin breakdown. The mechanism underlying such processes is an increase in the amount of intracellular calcium, leading to activation of proteases called calpains. The aim of this study is to evaluate whether nerve fibers can be preserved by the use of calpain inhibitor, after optic nerve crush. The left optic nerve of eight opossums (Didelphis aurita) was crushed (Group A) and four of them received the calpain inhibitor-2 that remained in the crushed area (Group B). Right optic nerves were used as control. After 96 hrs the nerves were dissected and processed for electron microscopy. Group A presented degenerating fibers, besides disorganization of the optic nerve structure. Group B maintained the structure of the optic nerve, which was organized in fascicles, thus preventing the dispersion of the fibers. The quantitative analysis showed that the Group B presented more normal fibers (25.36 ± 7.83) than Group A (21.47 ± 7.27), p<0.05, and less degenerating fibers (7.65 ± 2.80) compared to group A (10.22 ± 4.33), p<0.001). We also calculated the G ratio (axonal area/fiber area), which showed that the Group B presented G ratio (0.45 ± 0.29), close to normal values (0.44 ± 0.19). However Group A presented the largest G ratio (0.85 ± 13.57), when compared to the other two groups (p<0.01). In conclusion, our findings suggest that calpain inhibitor is capable to provide neuroprotection after a crush lesion. Abstracts of the 2nd Scientific Conference "Restauración Neurológica 2004" 254s of the 2nd Scientific Conference "Restauración Neurológica 2004" 254 26A8 Striatal trophic factor pleiotrophin augments dopamine neuron survival and neurite outgrowth Deanna M. Marchionini, Elin Lehrmann, William J. Freed and Timothy J. Collier Rush University Medical Center, Dept. Neurological Sciences, Chicago, Illinois, USA, National Institute on Drug Abuse, Baltimore, Maryland, USA Transplantation of embryonic ventral mesencephalic (VM) tissue is an experimental therapy for Parkinson’s disease. Both survival of grafted neurons and reinnervation of the striatum are essential for functional recovery. Our aim was to identify genes/factors that are upregulated during time points critical to nigrostriatal development and synaptogenesis. We utilized a microarray to profile gene expression in the rat E15 lateral ganglionic eminence, P1 and adult striatum. We identified genes that are upregulated during development, and tested their ability to promote dopamine neuron survival and neurite outgrowth in culture. Microisland cultures of E14 rat VM were exposed to pleiotrophin. On day 4 in vitro, cells were processed for tyrosine hydroxylase immunoreactivity (THir). Cell counts of THir neurons yielded 114.7 ± 8.0, 113.9 ± 11.0 and 134.3 ± 4.5% of control for cells treated with 10, 50 and 100 ng/ml pleiotrophin, respectively. Furthermore, neurite outgrowth was significantly increased in pleiotrophin treated cultures, 158.5 ± 21.3, 150.2 ± 18.1, 180.2 ± 20.1% of control for cells treated with 10, 50 and 100 ng/ml, respectively (p<0.05). Pleiotrophin may be an important cue in the establishment of nigrostriatal circuitry. Grafting of embryonic dopamine neurons in conjunction with pleiotrophin may promote reinnervation and functional recovery in Parkinson’s disease. Molecular mechanisms of neurodegenera-