Human Embryonic Stem Cell-Derived Immature Midbrain Dopaminergic Neurons Transplanted in Parkinsonian Monkeys.

Human embryonic stem cells (hESCs) differentiate into specialized cells, including midbrain dopaminergic neurons (DANs), and Non-human primates (NHPs) injected with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine develop some alterations observed in Parkinson's disease (PD) patients. Here, we obtained well-characterized DANs from hESCs and transplanted them into two parkinsonian monkeys to assess their behavioral and imaging changes. DANs from hESCs expressed dopaminergic markers, generated action potentials, and released dopamine (DA) in vitro. These neurons were transplanted bilaterally into the putamen of parkinsonian NHPs, and using magnetic resonance imaging techniques, we calculated the fractional anisotropy (FA) and mean diffusivity (MD), both employed for the first time for these purposes, to detect in vivo axonal and cellular density changes in the brain. Likewise, positron-emission tomography scans were performed to evaluate grafted DANs. Histological analyses identified grafted DANs, which were quantified stereologically. After grafting, animals showed signs of partially improved motor behavior in some of the HALLWAY motor tasks. Improvement in motor evaluations was inversely correlated with increases in bilateral FA. MD did not correlate with behavior but presented a negative correlation with FA. We also found higher 11C-DTBZ binding in positron-emission tomography scans associated with grafts. Higher DA levels measured by microdialysis after stimulation with a high-potassium solution or amphetamine were present in grafted animals after ten months, which has not been previously reported. Postmortem analysis of NHP brains showed that transplanted DANs survived in the putamen long-term, without developing tumors, in immunosuppressed animals. Although these results need to be confirmed with larger groups of NHPs, our molecular, behavioral, biochemical, and imaging findings support the integration and survival of human DANs in this pre-clinical PD model.

Terapia de extinción y estimulación magnética transcraneal para el tratamiento de trastornos de ansiedad / Extinction therapy and transcranial magnetic stimulation for the treatment of anxiety disorders

Resumen Objetivo: Describir los factores que pueden determinar la reducción de los síntomas en el trastorno de ansiedad generalizada y trastorno por estrés postraumático, mediante estimulación magnética transcraneal en combinación con terapia de extinción. Material y Métodos: Se realizó una búsqueda en bases de datos (Cochrane, EBSCO, Pubmed, Sciencedirect y Wiley), con las palabras clave "transcranial magnetic stimulation", "human", "fear extinction". Los criterios de selección incluyen estudios en humanos, tratamientos con terapia de extinción y EMT, en donde se registre la conductancia de la piel como variable de respuesta. Resultados: Existe poca investigación que cumpla con los criterios de la presente revisión bibliográfica. Se obtuvieron 5 artículos enfocados en el tratamiento de síntomas como el miedo y la recurrencia de recuerdos traumáticos. Los protocolos de estimulación son heterogéneos, la frecuencia de estimulación va de 1 Hz a 30 Hz. La estimulación de alta frecuencia fue la más utilizada. La duración máxima de los efectos reportados fue de 1 mes. Conclusiones: La EMT junto con la terapia de extinción como tratamiento para TEPT y TAG es un campo de estudio que requiere de más investigación. Los resultados sobre su eficacia no son concluyentes, el tamaño de muestra es pequeño y es necesario identificar qué protocolos son eficaces a largo plazo. Los estudios clínicos con pacientes que presenten estos trastornos son relevantes para conocer los efectos de aquellos protocolos que han sido exitosos en pacientes sanos (condicionados al miedo).

Abstract Objective: To describe the factors that can determine the reduction of symptoms in generalized anxiety and posttraumatic stress disorders by transcranial magnetic stimulation in combination with extinction therapy. Material and methods: A bibliographic review was conducted in databases (Cochrane, EBSCO, PubMed, ScienceDirect y Wiley), using the keywords: "transcranial magnetic stimulation", "human" and "fear extinction". A selection of clinical trials that used extinction therapy plus TMS and the skin conductance as variable quantified was made. Results: Five articles focused on the treatment of symptoms, like fear and recurrence of traumatic memories were obtained. There is little research on the topic. Stimulation protocols are heterogeneous between studies (stimulation frequency ranges from 1 to 30 Hz). Most of the studies reviewed reported the use of high-frequency stimulation. The maximum duration of therapeutic effects reported was one month. Conclusions: TMS and extinction therapy as a treatment for PTSD and GAD has a growing research field. Effectiveness results are not conclusive, sample sizes are small, and studies do not focus on which protocols are effective in the long-term. New studies that include patients with diagnosed PTSD and GAD are relevant to assess the protocols that have already been successful in healthy patients (fear-conditioned).

Comparative analysis of striatal [18 F]FDOPA uptake in a partial lesion model of Parkinson's disease in rats: Ratio method versus graphical model.

Animal models of Parkinson's disease are useful to evaluate new treatments and to elucidate the etiology of the disease. Hence, it is necessary to have methods that allow quantification of their effectiveness. [18 F]FDOPA-PET (FDOPA-PET) imaging is outstanding for this purpose because of its capacity to measure changes in the dopaminergic pathway noninvasively and in vivo. Nevertheless, PET acquisition and quantification is time-consuming making it necessary to find faster ways to quantify FDOPA-PET data. This study evaluated Male Wistar rats by FDOPA, before and after being partially injured with 6-OHDA unilaterally. MicroPET scans with a duration of 120 min were acquired and Patlak reference plots were created to estimate the influx constant Kc in the striatum using the full dynamic scan data. Additionally, simple striatal-to-cerebral ratios (SCR) of short static acquisitions were computed and compared with the Kc values. Good correlation (r > 0.70) was obtained between Kc and SCR, acquired between 80-120 min after FDOPA administration with frames of 10 or 20 min and both methods were able to separate the FDOPA-uptake of healthy controls from that of the PD model (SCR -28%, Kc -71%). The present study concludes that Kc and SCR can be trustfully used to discriminate partially lesioned rats from healthy controls.

Effect of extremely low frequency magnetic fields on oxidative balance in rat brains subjected to an experimental model of chronic unpredictable mild stress.

BACKGROUND: There has been an increasing interest in researching on the effects of extremely low-frequency magnetic fields on living systems. The mechanism of action of extremely low-frequency magnetic fields on organisms has not been established. One of the hypotheses is related to induce changes in oxidative balance. In this study, we measured the effects of chronic unpredictable mild stress induced-oxidative balance of rat's brain exposed to extremely low-frequency magnetic fields. METHODS: A first experiment was conducted to find out if 14 days of chronic unpredictable mild stress caused oxidative unbalance in male Wistar rat's brain. Catalase activity, reduced glutathione concentration, and lipoperoxidation were measured in cerebrum and cerebellum. In the second experiment, we investigate the effects of 7 days extremely low-frequency magnetic fields exposure on animals stressed and unstressed. RESULTS: The main results obtained were a significant increase in the catalase activity and reduced glutathione concentration on the cerebrum of animals where the chronic unpredictable mild stress were suspended at day 14 and then exposed 7 days to extremely low-frequency magnetic fields. Interestingly, the same treatment decreases the lipoperoxidation in the cerebrum. The stressed animals that received concomitant extremely low frequency magnetic fields exposure showed an oxidative status like stressed animals by 21 days. Thus, no changes were observed on the chronic unpredictable mild stress induced-oxidative damage in the rat's cerebrum by the extremely low-frequency magnetic field exposure together with chronic unpredictable mild stress. CONCLUSIONS: The extremely low-frequency electromagnetic field exposure can partially restore the cerebrum antioxidant system of previously stressed animals.

Subchronic stress effects on vascular reactivity in C57BL/6 strain mice.

BACKGROUND AND AIMS: There is a close relationship between psychosocial stress and the development of cardiovascular diseases. It has been reported that there are different alterations in endothelial function in this relationship. However, results obtained in different experimental stress models are controversial. Herein, we studied the effects of subchronic stress induced by movement restraint on several cardiovascular responses and plasma corticosterone concentration in male adult mice. METHODS: Experiments were performed in adult male mice of C57BL/6 strain. Animals were allocated into three groups: Control group A, without manipulation; Control group B, with manipulation (quantitation of blood pressure); and Experimental group, with quantitation of blood pressure and exposure to movement restraint. In vivo, heart rate and blood pressure were registered. In vitro, in aortic rings, vascular reactivity was analyzed. Additionally, plasma corticosterone concentration was quantified. RESULTS: In vivo, subchronic stress did not produce changes on heart rate either on blood pressure. In vitro, aortic rings with and without endothelium from control group B and experimental group showed: 1) a significant decrease in the maximal tension developed in response to phenylephrine; 2) this decrease was reverted by L-NAME. However, aortic rings from all groups, developed the same tension in response to high K+ solution. In aortic rings from animals of the experimental group, an increase in the maximal relaxation induced by carbachol was observed. This relaxation was prevented and/or reversed by L-NAME. Plasma corticosterone concentration was higher in the experimental group than that in the control group A. CONCLUSIONS: Exposition to subchronic movement restraint did not produce alterations in neurovegetative responses in this strain mice. But according to vasomotor responses observed, the results suggest that this subchronic stress model induces an increase in the synthesis/release of nitric oxide, both from endothelial cells and vascular smooth muscle. In accordance with the aforementioned results, we propose that C57BL/6 mice strain is sensitive to subchronic movement restraint stress model.

Extremely low frequency electromagnetic field exposure and restraint stress induce changes on the brain lipid profile of Wistar rats.

BACKGROUND: Exposure to electromagnetic fields can affect human health, damaging tissues and cell homeostasis. Stress modulates neuronal responses and composition of brain lipids. The aim of this study was to evaluate the effects of chronic extremely low frequency electromagnetic field (ELF-EMF) exposure, restraint stress (RS) or both (RS + ELF-EMF) on lipid profile and lipid peroxidation in Wistar rat brain. METHODS: Twenty-four young male Wistar rats were allocated into four groups: control, RS, ELF-EMF exposure, and RS + ELF-EMF for 21 days. After treatment, rats were euthanized, the blood was obtained for quantitate plasma corticosterone concentration and their brains were dissected in cortex, cerebellum and subcortical structures for cholesterol, triacylglycerols, total free fatty acids, and thiobarbituric acid reactive substances (TBARS) analysis. In addition, fatty acid methyl esters (FAMEs) were identified by gas chromatography. RESULTS: Increased values of plasma corticosterone were found in RS and ELF-EMF exposed groups (p < 0.05), this effect was higher in RS + ELF-EMF group (p < 0.05, vs. control group). Chronic ELF-EMF exposure increased total lipids in cerebellum, and total cholesterol in cortex, but decreased polar lipids in cortex. In subcortical structures, increased concentrations of non-esterified fatty acids were observed in RS + ELF-EMF group. FAMEs analysis revealed a decrease of polyunsaturated fatty acids of cerebellum and increases of subcortical structures in the ELF-EMF exposed rats. TBARS concentration in lipids was increased in all treated groups compared to control group, particularly in cortex and cerebellum regions. CONCLUSIONS: These findings suggest that chronic exposure to ELF-EMF is similar to physiological stress, and induce changes on brain lipid profile.

Effect of Intermediate-Frequency Repetitive Transcranial Magnetic Stimulation on Recovery following Traumatic Brain Injury in Rats.

Traumatic brain injury (TBI) represents a significant public health concern and has been associated with high rates of morbidity and mortality. Although several research groups have proposed the use of repetitive transcranial magnetic stimulation (rTMS) to enhance neuroprotection and recovery in patients with TBI, few studies have obtained sufficient evidence regarding its effects in this population. Therefore, we aimed to analyze the effect of intermediate-frequency rTMS (2 Hz) on behavioral and histological recovery following TBI in rats. Male Wistar rats were divided into six groups: three groups without TBI (no manipulation, movement restriction plus sham rTMS, and movement restriction plus rTMS) and three groups subjected to TBI (TBI only, TBI plus movement restriction and sham rTMS, and TBI plus movement restriction and rTMS). The movement restriction groups were included so that rTMS could be applied without anesthesia. Our results indicate that the restriction of movement and sham rTMS per se promotes recovery, as measured using a neurobehavioral scale, although rTMS was associated with faster and superior recovery. We also observed that TBI caused alterations in the CA1 and CA3 subregions of the hippocampus, which are partly restored by movement restriction and rTMS. Our findings indicated that movement restriction prevents damage caused by TBI and that intermediate-frequency rTMS promotes behavioral and histologic recovery after TBI.

Effect of acute extremely low frequency electromagnetic field exposure on the antioxidant status and lipid levels in rat brain.

BACKGROUND AND AIMS: It is generally accepted that electromagnetic fields (EMF) can exert biological effects; however, the mechanisms by which EMF elicits responses are still unknown. The present study was designed to assess the immediate effects of acute EMF exposure, movement restriction, and the combination of both on the antioxidant systems and lipid content in the whole brain of rat. METHODS: Thirty two male Wistar rats were arranged in four groups: control, EMF exposed, movement restrained (MR), and EMF + MR for 2 h. Rats were then sacrificed and their brains analyzed for superoxide dismutase and catalase activities, reduced glutathione, nitric oxide, total cholesterol, and triacylglycerol levels, as well as plasma corticosterone concentrations. RESULTS: Acute exposure to EMF induces reduction in catalase and superoxide dismutase activities, whereas the combination of EMF + MR also decreases both reduced glutathione and nitric oxide levels. Our results show that the acute exposure to EMF does not induce elevation of stress-hormone corticosterone but impairs the antioxidant status in rat brain. CONCLUSIONS: Plasma corticosterone concentration and antioxidant data indicate that the acute exposure to EMF appears to be a mild stressor that leads to some adaptive responses due to the activation of systems controlling the brain oxidative balance.

Effects of acute electromagnetic field exposure and movement restraint on antioxidant system in liver, heart, kidney and plasma of Wistar rats: a preliminary report.

PURPOSE: The aim of the present study was to evaluate the early effects of acute (2 h) exposure to extremely low frequency electromagnetic fields (ELF-EMF), as well as movement restraint (MR) and the combination of both on the antioxidant systems in the plasma, liver, kidney, and heart of rats. MATERIALS AND METHODS: Twenty-four adult male Wistar rats were divided in two groups, restrained and unrestrained. The restrained animals were confined into an acrylic tube for 120 min. Half of the animals of each group were exposed to ELF-EMF (60 Hz, 2.4 mT) during the period of restriction. Immediately after treatment, reduced glutathione (GSH), catalase (CAT), superoxide dismutase (SOD), and thiobarbituric acid reactive substances (TBARS) were measured in tissues. RESULTS: GSH concentration was significantly lower in the heart of all experimental animals when compared to the control group; furthermore, the decrease was higher in the liver of restrained animals. SOD activity was lower in the plasma of restrained and EMF exposed animals compared to unrestrained rats. There were no significant differences in CAT activity and TBARS levels among all the experimental groups vs. the control group. CONCLUSION: Two hours of 60 Hz EMF exposure might immediately alter the metabolism of free radicals, decreasing SOD activity in plasma and GSH content in heart and kidney, but does not induce immediate lipid peroxidation. Oxidative stress induced by movement restraint was stronger than that produced by EMF.

Neurite outgrowth on chromaffin cells applying extremely low frequency magnetic fields by permanent magnets.

BACKGROUND AND AIMS: There is an increasing interest about the effects of electromagnetic fields on health and clinical applications. Electromagnetic fields have been shown to promote differentiation and regeneration of many tissues. The purpose of the present study was to evaluate if a magnetic field (MF) varying in time is able to induce neurite outgrowth in cultured chromaffin cells. For this reason, a stimulation system was developed in order to generate a magnetic field, using permanent magnets as a supply. METHODS: In this investigation we used a pair of permanent ferrite magnets. These were mounted in a mechanical system in which both magnets rotate around a culture Petri dish. The stimulation device was designed at Centro de Investigación y de Estudios Avanzados, Avanzados del IPN, Mexico City. Primary cultures of chromaffin cells were stimulated with a magnetic field of 6.4 mT and 4, 7, 10 or 12Hz (2h daily, during a 7-day period). After treatment, percentage of neurite outgrowth was calculated. RESULTS: Our results show that the magnetic fields produced by rotating permanent magnets induced neurite outgrowth on chromaffin cells at 7 and 10Hz. CONCLUSIONS: The present study provides evidence that MFs varying in time (7 and 10Hz) induce neurite outgrowth in chromaffin cells. These studies will contribute to elucidate the effect of noninvasive MF stimulus in order to apply it in future regeneration therapies. Also, the device designed could be used for different kind of cells and may work as a model for future clinical devices.

Effects of whole body exposure to extremely low frequency electromagnetic fields (ELF-EMF) on serum and liver lipid levels, in the rat.

BACKGROUND: [corrected] The effects of extremely low-frequency electromagnetic fields (ELF-EMF) on the blood serum and liver lipid concentrations of male Wistar rats were assessed. METHODS: Animals were exposed to a single stimulation (2 h) of ELF-EMF (60 Hz, 2.4 mT) or sham-stimulated and thereafter sacrificed at different times (24, 48 or 96 h after beginning the exposure). RESULTS: Blood lipids showed, at 48 h stimulated animals, a significant increase of cholesterol associated to high density lipoproteins (HDL-C) than those observed at any other studied time. Free fatty acid serum presented at 24 h significant increases in comparison with control group. The other serum lipids, triacylglycerols and total cholesterol did not show differences between groups, at any time evaluated. No statistical differences were shown on total lipids of the liver but total cholesterol was elevated at 24 h with a significant decrease at 96 h (p = 0.026). The ELF-EMF stimulation increased the liver content of lipoperoxides at 24 h. CONCLUSION: Single exposures to ELF-EMF increases the serum values of HDL-C, the liver content of lipoperoxides and decreases total cholesterol of the liver. The mechanisms for the effects of ELF-EMF on lipid metabolism are not well understand yet, but could be associated to the nitric oxide synthase EMF-stimulation.

Adrenalectomy and dexamethasone replacement on yawning behavior.

Yawning, a phylogenetic behavior, present in reptiles, birds and mammals, has been studied for several decades, but to date its physiological function is still unknown. The role of stress as well as several peptides and the hypothalamus has been studied in relation to its regulation. To date however, no studies has been carried out to determine the role of the adrenal glands. Therefore, yawning behavior was studied in adrenalectomized rats, who then received dexamethasone replacement. The results show that rats whose adrenal glands were removed stopped both spontaneous and apomorphine-induced yawning, while dexamethasone reverted this effect. The results are discussed in terms of the possible role of corticosterone on yawning behavior.

Exposure to extremely low-frequency electromagnetic fields improves social recognition in male rats.

The effect of exposure to low-frequency electromagnetic fields (ELF EMFs) on social recognition was studied. The test was based upon a comparison between two encounters of an adult rat and a conspecific juvenile, separated by an interexposure interval (IEI). The exposure to ELF EMF of 1 mT intensity during 2 h for 9 days increased the duration of short-term memory of adult male Wistar rats up to 300 min. These data indicate, for the first time, that ELF EMF improves social recognition memory in rats.

Cell transplantation for Parkinson's disease: present status.

1. Parkinson's disease (PD) is a neurodegenerative disorder caused by the loss of neurons in the substantia nigra pars compacta and a striatal deficiency of dopamine. PD typically affects people in late middle age and progresses slowly. In the early stages of the disease, treatment targeting the dopaminergic network is effective. However, with disease progression, transplantation is an option for repairing and replacing missing dopaminergic neurons. 2. In this review, we evaluate the tissue grafts and cellular therapies that have and are being considered. Clinical trials were originally derived from transplants of adrenal medullary chromaffin cells and embryonic nigral dopaminergic neurons in patients with PD. 3. Recently, novel molecular and cellular treatments are being utilized in animals and these include embryonic stem cells, fetal cells from pigs, or transfected cells. In spite of new molecular techniques and some 20 years of experience, the transplantation therapy for PD has today the same problems and results as the first reports which used neural fetal tissue or adrenal grafts.

Determination of dopamine-releasing protein (DARP) in cerebrospinal fluid of patients with neurological disorders

Levels of DARP in the cerebrospinal fluid (CSF) of patients having a wide variety of nerulogical disorders were determined. Neurological disorders were categorized as degenrative, demyelinating, epilepsy, trauma, hydrocephalia, inflammatory, A-V malformation, CNS neoplasia, parasitic and stroke. DARP levels were determined by an enzyme-linked immunoabsorbent assay (ELISA) using monoclonal anti-DARP antibodies. A synthetic peptide corresponding to the first 36 aa of the N-terminal of DARP was used as standard. A total of 7 non-neurological patients and 73 patients with neurological disorders were tested. The relative concentrations of DARP decreased in patients with Parkinson's diseases vs. patients with non-neurological diseases and increased in other neuropathologies such as demyelinating, hydrocephalia and A-V malformations. Data obtained suggest that changes in the percentage and concentration of DARP may correlate with certain neurological disorders, showing particularly low levels in Parkinson's disease patients