Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0030567 (Parkinson's disease)
 63,064 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We studied how stimulation of protein kinase C and cAMP-dependent protein kinases affect the development of mesencephalic dopaminergic neurons in primary cell cultures derived from fetal rats at embryonic day E14. The effects of compounds which activate these second messenger systems were compared to those of basic fibroblast growth factor (bFGF) and insulin-like growth factor I (IGF-I). In mesencephalic cultures, there was a continuous loss of dopaminergic neurons. Despite this decline in cell number, neurotransmitter uptake per neuron increased with time, indicating that the surviving dopaminergic neurons continued their biochemical differentiation while others degenerated. IGF-I and bFGF did not affect the number of dopaminergic neurons. However, dopamine uptake per neuron was significantly higher in bFGF and IGF-I treated cultures, suggesting that these factors stimulated differentiation. Protein kinase C and cAMP-dependent protein kinases were not involved in mediating the effects of bFGF and IGF-I. Treatment of cultures with phorbol esters did not affect dopamine uptake, whereas elevated levels of intracellular cAMP resulted in an increase in dopamine uptake which was additive to that elicited by bFGF or IGF-I. Further analysis revealed that exposure of mesencephalic cultures to dibutyryl cAMP (dbcAMP) during the first 3 days after plating increased the survival of dopaminergic neurons, whereas prolonged treatment attenuated the development of the dopamine uptake system. Moreover, cyclic AMP, but not bFGF, was able to prevent the degeneration of dopaminergic neurons induced by 1-methyl-4-phenyl-pyridinium ion (MPP+), the active metabolite of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). The results suggest that increased intracellular levels of cAMP protect dopaminergic neurons in situations of stress like the process of dissociation and plating or the exposure to neurotoxic compounds. Our results reveal novel possibilities for the treatment of Parkinson's disease.
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PMID:Cyclic AMP, but not basic FGF, increases the in vitro survival of mesencephalic dopaminergic neurons and protects them from MPP(+)-induced degeneration. 135 86

The intracerebral transplantation of embryonic dopaminergic nigral neurons, although relatively successful, leads to a fairly low yield of surviving cells. Many factors may influence the viability of dopaminergic grafts and one of these is the preparation of the tissue prior to transplantation. We have investigated the effects of different steps during the preparation and storage of embryonic rat nigral cell suspensions on their subsequent survival at a variety of different time points using a combination of techniques and studies. For studies concerned with the first 24 h we employed vital stains, in the period covering the next 7 days we used in vitro cultures, and in the long term experiment we used in vivo grafts. The results suggest that nigral cell suspensions may remain sufficiently viable for grafting for much longer periods than previously reported. In addition a number of parameters which affect cell survival have been characterised, including the age of the embryonic donor tissue, the use of proteolytic enzymes and the trituration procedure used during the preparation of the suspension. The optimal preparation technique, therefore, uses E13-E14 embryos with the dissected ventral mesencephalon being incubated in purified 0.1% trypsin solutions for 60 min and triturated using a flame polished Pasteur pipette. This may have important implications in improving intracerebral transplantation for Parkinson's disease.
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PMID:A comparative study of preparation techniques for improving the viability of nigral grafts using vital stains, in vitro cultures, and in vivo grafts. 753 99

Neural transplantation in experimental Parkinsonism has so far focused on the ectopic placement of fetal ventral mesencephalic (VM) neurons into the dopamine-denervated caudate-putamen. VM grafts are effective in restoring dopamine neurotransmission in the grafted caudate-putamen and in partial amelioration of behavioral deficits. Recent pharmacological and physiological data have provided strong evidence that dopamine released from dendrites of the substantia nigra pars compacta (SNc) neurons within the pars reticulata (SNr) plays an important role in the regulation of the basal ganglia output pathways. Using a novel microtransplantation approach, multiple small cell suspension grafts (250 nl) derived from the VM of E14 rat embryos were implanted into the SNr of unilaterally 6-hydroxydopamine-lesioned rats. Behavioral changes in drug-induced rotation asymmetry were monitored for up to 14 weeks postgrafting, followed by a quantitative assessment and correlation of tyrosine hydroxylase (TH)-positive cell survival. The reduction in rotational asymmetry caused by the intranigral VM grafts was 64% for SKF 38393 (D1 agonist), 54% for apomorphine (mixed D1 and D2 agonist), and 67% for quinpirole (D2 agonist) when compared to a control spinal cord graft group. By contrast, amphetamine-induced rotation was completely unaffected. The correlation between number of TH-positive cells and behavioral compensation was highest for the D1 agonist (R = -0.729), though clear-cut also for the mixed D1/D2 agonist apomorphine (R = -0.664) and the D2 agonist quinpirole (R = -0.642). Favorable morphological features of the VM micrografts included extensive migration of the dopaminergic neurons into the host SNr and the formation of dense patches of dendrite-like TH-positive terminal networks within the SNr. The results demonstrate a novel pattern of behavioral recovery induced by intranigral VM transplants in the rat Parkinson model. This may have important implications for the understanding of how the nigrostriatal dopamine system influences motor control in the basal ganglia as well as for the development of optimal transplantation strategies in Parkinson's disease.
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PMID:Intranigral fetal dopamine grafts induce behavioral compensation in the rat Parkinson model. 791 16

Cryopreservation may allow long-term storage of fetal ventral mesencephalon (VM) for transplantation in patients suffering from Parkinson's disease (PD). We investigated whether the polymer methylcellulose protects fetal rat VM during cryopreservation in liquid nitrogen and improves survival and function of this tissue as intrastriatal suspension grafts in the 6-hydroxydopamine (6-OHDA) rat model. VM tissue fragments (E14-E15) were either immediately dissociated and grafted as a cell suspension (FRESH) or cryopreserved under controlled conditions for 7 days in a conventional cryoprotective medium (CRYO) or a medium containing 0.1% methylcellulose (mCRYO) and then dissociated and grafted. Rats from the cryo-groups showed only limited behavioral compensation in contrast to complete compensation observed in rats from the FRESH group. Cryopreservation of fetal rat VM decreased the viability of cell suspensions in vitro to about 70%, survival of grafted tyrosine hydroxylase-immunoreactive (TH-IR) neurons to 11% and 20%, and transplant volume to 8% and 17% (mCRYO and CRYO, respectively, compared to FRESH). The addition of 0.1% methylcellulose to tissue fragments during freezing did neither improve in vitro viability nor survival of TH-IR neurons nor behavioral compensation when compared to the control CRYO group. These results suggest that methylcellulose failed to improve survival of cryopreserved dopaminergic ventral mesencephalic neurons.
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PMID:Methylcellulose during cryopreservation of ventral mesencephalic tissue fragments fails to improve survival and function of cell suspension grafts. 869 78

The search for specific neurotrophic factors that will eventually be used to reduce or arrest the rate of degeneration of dopaminergic neurons in Parkinson's disease is being pursued by first testing the ability of putative compounds to increase the survival of dopaminergic neurons in primary cultures of the fetal, ventral mesencephalon. This research has intensified in recent years. The experimental procedures used by different laboratories in these studies differ widely, and meaningful comparisons of the results obtained are accordingly difficult to make. Some important experimental variables include the age of the fetal tissue used; the dissection technique used to isolate the ventral mesencephalon; the percentage of dopaminergic neurons present in the culture initially; handling of the tissue during dissection; the technique used to disperse the cells; the use of serum; the technique of plating the cells; the attachment factors used; detachment and loss of cells during the staining procedure; the age of the cultures at the time of analysis; the uneven distribution of cells at the time of analysis and the use of imaging techniques in the analysis. We show that when the E14 rat embryo is used, it is possible to consistently obtain a culture with 20% of tyrosine hydroxylase-positive neurons. Neither the plating density in the range of 7.8 x 10(3) to 1.25 x 10(5) cells/cm2, nor the percentage of serum in the growth medium affected the percentage of cells that expressed TH initially, at 4 or 12 h after plating. When the cells were plated as 25 microliters droplets, called microislands (area approximately 12.5 mm2), and allowed to attach before additional growth medium was added, cell density remained uniform at the center of the microisland for the duration of the culture. Restriction of the analysis of cell survival to the center of the microisland therefore helped to decrease the variability in counting that could occur when cells are dispersed over a larger area. In contrast, in an 8-well chamber slide or 35 mm petri dish, in which the whole area is plated, cell density was consistently higher at the edge (edge effect), versus the centre, by a factor of about three. The use of microisland cultures also has the additional benefit of increasing by a factor of about five the number of individual cultures that can be set up per liter, and a proportionate reduction in the number of animals used per experiment. When the percentage of serum in the growth medium was 0% always, or 10% for the first 12 h, and 0% thereafter, or 10% always, the number of TH-pos neurons per field (using a x 20 objective, column factor 1.25; area 320 microns2) after 5 days in culture (DIV5) was < 1,3-8 and 14-22, respectively. Under the same experimental conditions, the number of neurons (MAP2-positive) per field was 5-8, 18-30 and 45-65 (N = 10 in all cases), respectively. Serum deprivation therefore has a highly deleterious effect on neuronal survival in culture. We suggest that cultures that were exposed to serum at any stage of the experiment, should not be referred to as "serum-free', since even a brief exposure to serum exerts a protective effect on neurons, and especially on dopaminergic neurons. Instead, the percentage and kind of serum used, the exact usage, and the duration of exposure of the cells to serum should be stated. Finally, it is suggested that where possible, an imaging system with manual count and journaling capabilities be used in the analysis. The methods described are illustrated by dose-response curves of the neurotrophic effects of BDNF, NGF-beta and IL-6 versus percentage survival on dopaminergic neurons, when grown in serum-free medium throughout.
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PMID:Standardized methods to bioassay neurotrophic factors for dopaminergic neurons. 884 22

Grafting of fetal ventral mesencephalon in Parkinson's disease has been extensively studied. A crucial draw back of this technique is the low survival rate of the dopaminergic neurons. It has been documented that only 5-20% of the grafted neurons survive, and to enhance graft efficacy to a satisfying level, increased cell survival is of utmost desire. In this study we have used the antioxidant tiriliazad mesylate (U-74006F) to study the effect on the survival of dopaminergic neurons after grafting. The in oculo grafting model was used and ventral mesencephalon was dissected from E14-E15 rat fetuses in Hanks' balanced salt solution (HBSS), in Dulbecco's modified Eagle medium (DMEM), or in 0.3, 3.0, or 30 microM U-74006F diluted in DMEM. The tissue was then inserted into the anterior chamber of the eye. Some of the transplants were further treated with intraocular injections of 3 or 30 microM U-74006F (5 microliters) weekly for 2 weeks. Quantification of tyrosine hydroxylase (TH)-immunoreactive profiles revealed that in transplants treated with U-74006F at dissection only, no change in the number of TH-positive neurons was found. Pretreatment of 0.3 microM U-74006F during dissection combined with intraocular injections of U-74006F after grafting, on the other hand, resulted in a dose-dependent enhancement of survival of TH-positive neurons. Dissection in, and intraocular treatment with, 3 microM U-74006F resulted in a significantly enhanced survival of TH-positive neurons whereas using U-74006F at a concentration of 30 microM did not change the cell survival compared to solely DMEM-treated grafts. Thus, 30 microM was interpreted to be an overdose. Comparing cell survival when dissected in DMEM with that dissected in HBSS showed that DMEM was clearly superior. Nerve fiber formation was most pronounced in grafts treated with 3 microM U-74006F. In conclusion, survival of TH-positive neurons is enhanced by U-74006F, which is readily available for clinical use and thus could be employed to enhance graft survival when transplanting patients suffering from Parkinson's disease.
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PMID:Tirilazad mesylate increases dopaminergic neuronal survival in the in Oculo grafting model. 939 75

Both deprenyl and rasagiline (R(+)-N-propargyl-1-aminoindane mesylate), at a concentration of 1-10 microM, increased survival in vitro of rat E14 mesencephalic dopaminergic neurons that had been primed with 10% serum for 12 h (p < 0.05). Rasagiline, but not deprenyl, also increased total neuronal (MAP2-positive) survival (p < 0.05) Under serum-free conditions, rasagiline, but not deprenyl, retained its neuroprotective action on dopaminergic neurones. GABAergic neurons were not affected by either deprenyl or rasagiline. Clorgyline, an MAO-A inhibitor, did not exert any of these effects. The protective action of rasagiline on dopaminergic neurons, even under stringent serum-free conditions, is striking, and warrants further investigation for a role in the treatment of Parkinson's disease.
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PMID:Increased survival of dopaminergic neurons by rasagiline, a monoamine oxidase B inhibitor. 955 42

Glial cell line-derived neurotrophic factor (GDNF) is known as a potent neurotrophic factor for dopaminergic neurons. Since adeno-associated virus (AAV) vector is a suitable vehicle for gene transfer into neurons, rat E14 mesencephalic cells were transduced with an AAV vector expressing GDNF. When compared with mock transduction, a larger number of dopaminergic neurons survived in AAV-GDNF-transduced cultures (234% and 325% of controls at 1 and 2 weeks, respectively; P < 0.01). Furthermore, the dopaminergic neurons in the latter cultures grew more prominent neurites than those in the former. These findings suggest that AAV vector-mediated GDNF gene transfer may prevent dopaminergic neuron death, and is therefore a logical approach for the treatment of Parkinson's disease.
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PMID:Prevention of dopaminergic neuron death by adeno-associated virus vector-mediated GDNF gene transfer in rat mesencephalic cells in vitro. 966 64

Among the dopaminergic neurons in substantia nigra pars compacta and in the ventral tegmental area, subpopulations express the calcium-binding proteins calbindin (CB) and calretinin (CR), and the CB-containing neurons are supposed to be less prone to degeneration in Parkinson's disease. Glial cell line-derived neurotrophic factor (GDNF) is a potent survival factor for nigrostriatal dopaminergic neurons. Using free-floating roller-tube (FFRT) cultures derived from fetal rat (E14) ventral mesencephalon we found that GDNF (10 ng/ml) significantly increased the number of surviving tyrosine hydroxylase (TH)-immunoreactive neurons. The possible effects of GDNF treatment on CB-immunoreactive (CB-ir) and CR-ir neurons in such cultures were examined in the present study. The neuronal cell densities were measured by quantifying the numbers of CB-ir and CR-ir neurons in areas of sections through the most extensive parts of the spherical cultures. In 4-day-old and 8-day-old cultures GDNF treatment increased the density of CB-ir neurons by 50% and 59%, respectively. Partial co-existence of TH and CB was shown using the method of double immunolabeling. The density of CR-containing neurons was unaffected by GDNF treatment as confirmed by Western blotting for CR. Parallel effects of GDNF treatment were obtained for cultures of human fetal ventral mesencephalon (8 weeks postconception). In conclusion, our findings identify GDNF as a potent factor for fetal rat and human nigral CB-ir neurons able to promote their survival in culture. Referring to a suggested neuroprotective role of CB, the results may be of relevance in the context of neuronal transplantation of patients suffering from severe Parkinson's disease.
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PMID:GDNF increases the density of cells containing calbindin but not of cells containing calretinin in cultured rat and human fetal nigral tissue. 1033 73

Transplantation using fetal nigral grafts has been performed by various groups worldwide in over 200 Parkinson's disease (PD) patients in an attempt to restore dopaminergic (DA) input to the striatum. However, the proportion of the implanted DA neurons that survives, whether using suspension, partially dissociated, or solid grafts, is small, often as low as 5 to 10%, which is insufficient to allow a full functional recovery. A significant proportion of the transplanted neurons in animal models of PD has been shown to die via apoptosis, but the reason for this is unclear. Since the methods used to prepare donor tissue for neural transplantation and in vitro culture are identical, we have looked at the time course of DA neuron loss following cell suspension preparation using an in vitro assay system and considered whether the procedures used may, in part, be responsible for the poor DA neuron survival. Primary dissociated cultures of E14 rat ventral mesencephala were incubated for different periods in serum-containing and serum-free media. After fixation, the TUNEL method, as well as ethidium bromide and acridine orange, were used to detect apoptosis, and DA neurons were localized immunocytochemically. Results showed that most apoptosis occurred during the first 24 h and that 50% of the DA neurons were lost in the first 8 h. Double-immunofluorescent labeling confirmed the presence of TUNEL+ve nuclei within DA neurons. There was no difference in either the extent or rate of loss between the serum-containing and serum-free medium during the first 32 h. We suggest, therefore, that existing methods used to prepare cell suspensions probably induce apoptosis and may need to be modified in order to increase the survival of DA neurons.
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PMID:Apoptosis in primary cultures of E14 rat ventral mesencephala: time course of dopaminergic cell death and implications for neural transplantation. 1063 Jan 93


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