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1.
Exp Brain Res ; 235(3): 731-742, 2017 03.
Article in English | MEDLINE | ID: mdl-27866262

ABSTRACT

The formation of oligomers and aggregates of overexpressed or mutant α-synuclein play a role in the degeneration of dopaminergic neurons in Parkinson's disease by causing dysfunction of mitochondria, reflected in their disturbed mobility and production of ROS. The mode of action and mechanisms underlying this mitochondrial impairment is still unclear. We have induced stable expression of wild-type, A30P or A53T α-synuclein in neuronally differentiated SH-SY5Y neuroblastoma cells and studied anterograde and retrograde mitochondrial trafficking in this cell model for Parkinson's disease. In contrast to wild-type and A30P, A53T α-synuclein significantly inhibited mitochondrial trafficking, at first retrogradely and in a later stage anterogradely. Accordingly, A53T α-synuclein also caused the highest increase in ROS production in the dysmobilized mitochondria in comparison to wild-type or A30P α-synuclein. Treatment with NAP, the eight amino acid peptide identified as the active component of activity-dependent neuroprotective protein (ADNP), completely annihilated the adverse effects of A53T on mitochondrial dynamics. Our results reveal that A53T α-synuclein (oligomers or aggregates) leads to the inhibition of mitochondrial trafficking, which can be rescued by NAP, suggesting the involvement of microtubule disruption in the pathophysiology of Parkinson's disease.


Subject(s)
Mitochondria/drug effects , Mitochondria/genetics , Oligopeptides/pharmacology , alpha-Synuclein/genetics , alpha-Synuclein/metabolism , Alanine/genetics , Cell Differentiation/drug effects , Cell Differentiation/genetics , Cell Line, Tumor , Gene Expression Regulation/drug effects , Gene Expression Regulation/genetics , Humans , Kymography , Luminescent Proteins/genetics , Luminescent Proteins/metabolism , Membrane Potential, Mitochondrial/drug effects , Membrane Potential, Mitochondrial/genetics , Microtubule-Associated Proteins/genetics , Microtubule-Associated Proteins/metabolism , Mitochondria/pathology , Neuroblastoma/pathology , Neuroblastoma/ultrastructure , RNA, Messenger/metabolism , Reactive Oxygen Species/metabolism , Threonine/genetics , Transfection , alpha-Synuclein/chemistry
2.
Neuropathol Appl Neurobiol ; 35(1): 89-102, 2009 Feb.
Article in English | MEDLINE | ID: mdl-19187061

ABSTRACT

AIMS: Previous studies on the therapeutic time window for intravascular administration of bone marrow stem cells (BMSCs) after stroke have shown that early intervention (from 3 h after onset) in the middle cerebral artery occlusion (MCAO) rat model is the most effective approach to reduce ischaemic lesion size. We have confirmed these observations but noticed that 2 weeks after transplantation, almost none of the grafted BMSCs could be detected in or around the lesion. The present experiments aimed to assess the fate and kinetics of intravascularly injected BMSCs shortly after administration in correlation to the development of the ischaemic lesion after MCAO. METHODS: We administered a syngeneic suspension of complete (haematopoietic and mesenchymal) BMSCs via the carotid artery to rats at 2 h after MCAO onset. We examined the distribution and tissue location of BMSCs within the first 24 h after arterial administration by perfusion-fixating rats and performing immunohistochemical analysis at different time points. RESULTS: The vast majority (>95%) of BMSCs appeared to become trapped in the spleen shortly after injection. Six hours after implantation, together with the appearance of activated microglia, the first BMSCs could be detected in and around the lesion; their number gradually increased during the first 12 h after implantation but started to decrease at 24 h. The implanted BMSCs were surrounded by activated and phagocytotic microglia. CONCLUSION: Our results show that ischaemic lesion size reduction can already be achieved by the early transient presence at the lesion site of intravascularly implanted BMSCs, possibly mediated via activated microglia.


Subject(s)
Brain Ischemia/therapy , Hematopoietic Stem Cell Transplantation , Hematopoietic Stem Cells/physiology , Animals , Apoptosis , Brain/physiopathology , Carotid Arteries , Immunohistochemistry , Infarction, Middle Cerebral Artery/therapy , Injections, Intra-Arterial , Male , Mesenchymal Stem Cell Transplantation , Mesenchymal Stem Cells/physiology , Microglia/physiology , Phagocytosis , Rats , Rats, Wistar , Spleen/physiology , Stroke/therapy
3.
Neuropathol Appl Neurobiol ; 31(6): 600-9, 2005 Dec.
Article in English | MEDLINE | ID: mdl-16281908

ABSTRACT

Feeding C57Bl/6 J mice the copper chelator cuprizone leads to selective apoptosis of mature oligodendrocytes and concomitant demyelination predominantly in the corpus callosum. The process of oligodendrocyte apoptosis in this animal model for multiple sclerosis (MS) involves early microglial activation, but no infiltration of T-lymphocytes. Therefore, this model could mimic early stages of oligodendrocyte degeneration Affected oligodendrocytes express the common neurotrophin receptor, p75(NTR), a 'stress-receptor' which under certain circumstances can induce apoptosis. Only affected oligodendrocytes in MS lesions and MS animal models express this receptor. In order to study the significance of p75(NTR) in the fate of oligodendrocytes, we have exposed wild-type as well as p75(NTR)-knockout mice to a 0.2% (w/w) cuprizone diet and performed a comparative immunohistochemical analysis of the corpus callosum at various time points. Surprisingly, our results show that the absence of p75(NTR) did not alter cuprizone-induced oligodendrocyte death (and subsequent de- or remyelination). Apparently, intracellular apoptosis pathways in adult oligodendrocytes do not require p75(NTR) activated signal transduction in the absence of T-lymphocytes and T-lymphocyte derived cytokines.


Subject(s)
Apoptosis , Demyelinating Diseases/pathology , Demyelinating Diseases/physiopathology , Oligodendroglia/pathology , Receptor, Nerve Growth Factor/genetics , Animals , Corpus Callosum/pathology , Cuprizone , Demyelinating Diseases/chemically induced , Male , Mice , Mice, Inbred C57BL , Mice, Knockout , Monoamine Oxidase Inhibitors , Nerve Regeneration , T-Lymphocytes/pathology
4.
Neurosci Res ; 49(2): 261-5, 2004 Jun.
Article in English | MEDLINE | ID: mdl-15140568

ABSTRACT

Embryonic mouse neural stem cells (NSCs) were isolated from E14 mice, multiplied in medium containing epidermal growth factor (EGF) and basic fibroblast growth factor (bFGF) and plated in laminin-coated wells in basic serum-free neurobasal medium. After 7 days in vitro, approximately 20% of the embryonic mouse NSCs developed into morphologically and biochemically fully maturated neurons, with extensive dendrites and multiple synaptic contacts. However, even after 22 days of culture, none of these neurons developed voltage-dependent sodium-channels characteristic for a functional neuron. Apparently, the morphological differentiation and the electrophysiological maturation of an embryonic mouse NSC into a neuron are independently regulated.


Subject(s)
Cell Differentiation/physiology , Neurons/cytology , Stem Cells/cytology , Animals , Cells, Cultured , Electric Stimulation/methods , Embryo, Mammalian , Epidermal Growth Factor/physiology , Fibroblast Growth Factor 2/physiology , Glial Fibrillary Acidic Protein/physiology , Immunohistochemistry/methods , Membrane Potentials/physiology , Membrane Potentials/radiation effects , Mice , Mice, Inbred C57BL , Microtubule-Associated Proteins/metabolism , Neurons/physiology , Patch-Clamp Techniques/methods , Stem Cells/physiology
6.
Article in English | MEDLINE | ID: mdl-14506941

ABSTRACT

hSOD1 (G93A) transgenic mice develop pathological changes similar to those in patients with familial amyotrophic lateral sclerosis (FALS). In particular, the progressive degeneration of motoneurons is charactered in this mouse model. One feature of stressed motoneurons in ALS and the hSOD1 mice is the induction of the p75 neurotrophin receptor, which is thought, under certain circumstances, to be a death-signaling molecule. We have studied disease progression of hSOD1 (G93A) mice in the absence of the p75NTR receptor and we monitored histological changes in the ventral spinal cord. Whereas female double transgenics showed prolonged survival, this effect was not observed in males. Improved survival in female mice was not correlated with increased motoneuronal survival, but with less astrocytic activation in lumbar ventral spinal cord, as shown by glial fibrillary acidic protein immunohistochemistry. These data suggest that p75NTR is not directly involved in the mechanism leading to motoneuron degeneration. More likely, an indirect process, presumably via regulation of astrocytes, might be responsible for the increased survival responses of female double transgenic mice.


Subject(s)
Amyotrophic Lateral Sclerosis/physiopathology , Receptors, Nerve Growth Factor/genetics , Amyotrophic Lateral Sclerosis/mortality , Amyotrophic Lateral Sclerosis/pathology , Animals , Astrocytes/pathology , Cell Count , Disease Models, Animal , Female , Gene Expression , Gliosis/pathology , Male , Mice , Mice, Knockout , Motor Neurons/enzymology , Motor Neurons/pathology , Receptor, Nerve Growth Factor , Receptors, Nerve Growth Factor/metabolism , Signal Transduction , Superoxide Dismutase/metabolism , Superoxide Dismutase-1 , Survival Rate
7.
Neuroscience ; 116(3): 685-94, 2003.
Article in English | MEDLINE | ID: mdl-12573711

ABSTRACT

Amyotrophic lateral sclerosis is a lethal neurodegenerative disorder involving motoneuron loss in the cortex, brainstem and spinal cord, resulting in progressive paralysis. Aberrant neurotrophin signalling via the low affinity neurotrophin receptor p75 has been suggested to be involved in the motoneuron death by the activation of apoptotic pathways. In order to investigate the involvement of neurotrophin receptor p75 in the amyotrophic lateral sclerosis related motoneuron degeneration process, we have studied the expression of this receptor in the spinal cord of transgenic mice carrying a mutated human Cu, Zn superoxide dismutase gene. Mutations in the superoxide dismutase gene are one of the genetic causes for familiar amyotrophic lateras sclerosis and human superoxide dismutase-1 transgenic mice develop symptoms and pathology similar to those in human amyotrophic lateras sclerosis. Our study shows that in these mice, spinal motoneurons, which normally do not contain the neurotrophin receptor p75 receptor, express this receptor during the progress of the disease. Expression of the neurotrophin receptor p75 receptor coincides with the expression of activating transcription factor 3, a member of the activating transcription factor/cyclic AMP family of stress transcription factors. Only a minority of these spinal motoneurons actually showed co-expression of neurotrophin receptor p75 with caspase-3 activity, suggesting that expression of the neurotrophin receptor p75 receptor is not directly related to the execution phase of the apoptosis process.


Subject(s)
Amyotrophic Lateral Sclerosis/metabolism , Disease Models, Animal , Motor Neurons/metabolism , Receptors, Nerve Growth Factor/biosynthesis , Amyotrophic Lateral Sclerosis/genetics , Animals , Female , Gene Expression Regulation/physiology , Male , Mice , Mice, Transgenic , Receptor, Nerve Growth Factor , Receptors, Nerve Growth Factor/deficiency , Receptors, Nerve Growth Factor/genetics , Superoxide Dismutase/deficiency , Superoxide Dismutase/genetics , Superoxide Dismutase-1
8.
Exp Neurol ; 177(2): 419-27, 2002 Oct.
Article in English | MEDLINE | ID: mdl-12429188

ABSTRACT

Previous studies suggest that neurotrophins support regeneration and survival of injured motoneurons. Based on these findings, brain-derived neurotrophic factor (BDNF) has been clinically investigated for its therapeutic potential in amyotrophic lateral sclerosis (ALS), a rapidly progressing and fatal motoneuronal disease. We questioned whether imbalances of neurotrophic levels are indeed involved in the pathology of ALS. Therefore the expression of nerve growth factor (NGF), BDNF, neurotrophin-3 (NT-3), and neurotrophin-4/5 (NT-4/5) was investigated in postmortem muscle tissue of the biceps from 15 patients with neuropathologically confirmed sporadic ALS and 15 age-matched controls. Using mRNA analysis techniques and quantitative protein measurements, we have demonstrated that both mRNA and protein levels of all four neurotrophins are increased in muscle tissue of ALS patients. The production levels displayed a disease duration dependency and different expression patterns emerged for the four neurotrophins. Whereas the early phase of the disease was characterized by a strong upregulation of BDNF, levels of NGF, NT-3, and NT-4/5 gradually increased in the course of the disorder, peaking at later stages. We conclude that decreased neurotrophic support from muscle tissue is most likely not the cause of motoneuron degeneration in ALS. On the contrary, our results suggest that degenerating motoneurons in ALS are exposed to elevated levels of muscle-derived neurotrophins.


Subject(s)
Amyotrophic Lateral Sclerosis/metabolism , Muscle, Skeletal/metabolism , Nerve Growth Factors/metabolism , Adult , Aged , Amyotrophic Lateral Sclerosis/pathology , Brain-Derived Neurotrophic Factor/genetics , Brain-Derived Neurotrophic Factor/metabolism , Disease Progression , Enzyme-Linked Immunosorbent Assay , Female , Humans , In Situ Hybridization , Male , Middle Aged , Muscle, Skeletal/chemistry , Muscle, Skeletal/pathology , Nerve Growth Factor/genetics , Nerve Growth Factor/metabolism , Nerve Growth Factors/genetics , Neurotrophin 3/genetics , Neurotrophin 3/metabolism , RNA, Messenger/analysis , RNA, Messenger/metabolism , Reverse Transcriptase Polymerase Chain Reaction
9.
J Neuroimmunol ; 118(2): 203-11, 2001 Aug 30.
Article in English | MEDLINE | ID: mdl-11498255

ABSTRACT

The expression of interleukin-1beta was examined in dorsal root ganglion (DRG) neurons from adult rats using non-radioactive in situ hybridization and immunocytochemistry. At all spinal levels, approximately 70% of the DRG neurons appeared to express IL-1beta mRNA; about 80% of these DRG neurons actually appeared to produce the IL-1beta protein at markedly varying levels. The expression of IL-1beta was found in large as well as in intermediate diameter sensory neurons but only sporadically in the population of small sensory neurons. The population of IL-1beta immunopositive sensory neurons included most of the large calretinin-positive Ia afferents, but only a few of the small substance P/CGRP positive sensory neurons. In situ hybridization staining for the detection of type 1 IL-1 receptor showed expression of this receptor by most of the sensory neurons as well as by supportive glial-like cells, presumably satellite cells. The functional significance of IL-1beta in the DRG neurons needs to be elucidated, but we speculate that IL-1beta produced by DRG neurons may be an auto/paracrine signalling molecule in sensory transmission.


Subject(s)
Ganglia, Spinal/metabolism , Interleukin-1/biosynthesis , Neurons, Afferent/metabolism , Animals , Calbindin 2 , Calcitonin Gene-Related Peptide/biosynthesis , Cell Size , Cytoplasm/metabolism , Cytoplasm/ultrastructure , Female , Ganglia, Spinal/cytology , Immunohistochemistry , In Situ Hybridization , Interleukin-1/genetics , Male , Microscopy, Immunoelectron , Neuroglia/cytology , Neuroglia/metabolism , Neurons, Afferent/cytology , Polyribosomes/metabolism , Polyribosomes/ultrastructure , RNA, Messenger/biosynthesis , Rats , Receptors, Interleukin-1/biosynthesis , Receptors, Interleukin-1 Type I , S100 Calcium Binding Protein G/biosynthesis , Substance P/biosynthesis
10.
Histochem Cell Biol ; 116(6): 545-51, 2001 Dec.
Article in English | MEDLINE | ID: mdl-11810196

ABSTRACT

A non-radioactive in situ hybridisation method is described for the detection of low intramuscular levels of brain-derived neurotrophic factor (BDNF) mRNA at the electron microscope level. Application of high-grade silver-gold intensification of the diaminobenzidine end product of in situ hybridisation revealed that in adult rat muscle the constitutive expression of muscular BDNF is confined to the myofibres; satellite cells, Schwann cells, endothelial cells, fibroblasts or axons do not appear to contribute to BDNF production in normal muscle. Although muscular BDNF is a neurotrophic factor for innervating motoneurons and supposedly released only at the motor endplates, the production of BDNF mRNA appears to occur along the entire length of the myofibres and is not confined to nuclei in the postsynaptic regions.


Subject(s)
Brain-Derived Neurotrophic Factor/metabolism , Gene Expression/genetics , In Situ Hybridization/methods , Muscle, Skeletal/metabolism , RNA, Messenger/metabolism , Animals , Brain-Derived Neurotrophic Factor/analysis , Brain-Derived Neurotrophic Factor/genetics , Immunohistochemistry , Microscopy, Electron , Muscle, Skeletal/ultrastructure , Rats , Rats, Wistar
11.
Exp Brain Res ; 132(3): 345-50, 2000 Jun.
Article in English | MEDLINE | ID: mdl-10883382

ABSTRACT

Human fetal ventral mesencephalon tissue has been used as dopaminergic striatal implants in Parkinsonian patients, so far with variable effects. Fetuses from animals that breed in large litters, e.g., pigs, have been considered as alternative donors of dopaminergic tissue. The optimal gestational age of the porcine fetal donors has not been studied systematically. We collected ventral mesencephalic (VM) tissue from fetal pigs, embryonal ages E21, E28, E42, and E70, and examined the viability of the fetal VM cells after dissociation, the expression of tyrosine hydroxylase (TH) in culture, the presence of catecholamines, and the cellular survival and outgrowth up to 10 months after intrastriatal implantation in rats. The highest viability was found in suspensions prepared from E28 fetuses. The highest number of TH-positive cells was found in cell cultures prepared from E28 VM tissue. Explants with a gestational age of 28 and 42 days contained the largest amount of dopamine. Only E28-derived grafts showed TH-cell survival after implantation in rat striatum. Our results show that a gestational age of 28 days must be considered to be the optimal age for dopaminergic tissue derived from pig fetuses for therapeutic use as intrastriatal grafts in Parkinsonian patients.


Subject(s)
Brain Tissue Transplantation/methods , Fetal Tissue Transplantation/methods , Gestational Age , Mesencephalon/transplantation , Parkinson Disease/surgery , Animals , Brain Tissue Transplantation/standards , Cell Survival/physiology , Cells, Cultured , Dopamine/analysis , Dopamine/physiology , Female , Fetal Tissue Transplantation/standards , Graft Survival/physiology , Humans , Mesencephalon/cytology , Neurons/chemistry , Neurons/cytology , Neurons/enzymology , Parkinson Disease/physiopathology , Rats , Rats, Wistar , Reproducibility of Results , Swine , Transplantation, Heterologous , Tyrosine 3-Monooxygenase/analysis
12.
Brain Res Dev Brain Res ; 116(2): 217-22, 1999 Sep 06.
Article in English | MEDLINE | ID: mdl-10521567

ABSTRACT

The factors that regulate the ontogeny and differentiation of C1 adrenergic neurons located in the rostral ventrolateral medulla (RVLM) are completely unknown. In the present study, we have investigated the effects of a number of neurotrophic factors on the survival of E18-19 rat C1 adrenergic neurons in culture. Immunohistochemistry and reverse transcription polymerase chain reaction (RT-PCR) were used to study the expression of tyrosine hydroxylase (TH), an enzyme present in all catecholaminergic neurons, and of phenylethanolamine N-methyltransferase (PNMT), the final enzyme in the synthesis of adrenalin, as markers for the C1 RVLM neurons. Our results show that GDNF, CNTF BDNF, NT-3 and NT-4/5 increase the number of TH-immunoreactive neurons surviving in vitro. The effects of NGF, TGFbeta and bFGF were not significant. The E18-19 C1 neurons appeared to loose their ability to express PNMT in culture as examined with immunocytochemistry and RT-PCR, and none of the tested neurotrophic factors was able to sustain or induce this expression. Our results indicate that the adrenergic phenotype of C1 neurons, or the survival of these neurons, is determined by environmental factors other than the neurotrophic factors examined in this study.


Subject(s)
Catecholamines/physiology , Medulla Oblongata/drug effects , Nerve Growth Factors/pharmacology , Neurons/drug effects , Animals , Cell Differentiation/drug effects , Embryo, Mammalian/cytology , Embryo, Mammalian/drug effects , Embryo, Mammalian/metabolism , Immunohistochemistry , Medulla Oblongata/cytology , Medulla Oblongata/embryology , Rats , Rats, Wistar
13.
J Physiol Paris ; 93(1-2): 87-96, 1999.
Article in English | MEDLINE | ID: mdl-10084712

ABSTRACT

A general survey is given of old as well as more recent findings concerning matches between electrophysiological properties of motoneurones and contractile properties of their muscle fibres. Mechanisms for creating and maintaining such matches are discussed. It is pointed out that it is not sufficient to describe the variation of functional motoneurone characteristics simply in terms of 'fast' or 'slow': all properties seem continuously graded and there is cytochemical evidence for several, seemingly independent parameters of functional specialisation.


Subject(s)
Motor Neurons/physiology , Muscle Fibers, Skeletal/physiology , Adaptation, Physiological , Animals , Calcium/metabolism , Electric Conductivity , Histocytochemistry , Homeostasis , Humans , Muscle Contraction/physiology
14.
Neurosci Lett ; 236(1): 41-4, 1997 Oct 24.
Article in English | MEDLINE | ID: mdl-9404947

ABSTRACT

We have studied the regulation of the expression of neurotrophin-3 (NT-3) mRNA in neonatal and adult rat muscle spindles after denervation and after denervation followed by reinnervation. Denervation of the intrafusal fibres did not result in an upregulation of the NT-3 mRNA expression but decreased this expression below the detection limit of the in situ hybridization method. Reinnervation of intrafusal fibres restored the NT-3 mRNA expression. The results suggest that the expression of NT-3 mRNA in postnatal muscle spindles is controlled by neuronal factors. The intrafusal fibre derived NT-3 may act as an instructive, feedback messenger for innervating neurons and may play a role in stabilizing and maintaining functional neuromuscular connections.


Subject(s)
Muscle Fibers, Skeletal/metabolism , Muscle Spindles/metabolism , Nerve Growth Factors/genetics , Nerve Regeneration/physiology , RNA, Messenger/biosynthesis , Animals , Animals, Newborn , Denervation , Nerve Crush , Neurotrophin 3 , Rats , Rats, Wistar
15.
Brain Res ; 755(2): 319-25, 1997 May 02.
Article in English | MEDLINE | ID: mdl-9175899

ABSTRACT

The ultrastructure and distribution of dopaminergic boutons within the rat mesencephalic trigeminal (Me5) nucleus was examined with the use of electronmicroscopic immunocytochemistry. A total of 5102 boutons, comprising axosomatic and axodendritic synaptic terminals as well as non-synaptic boutons (or varicosities), located in the ventrocaudal portion of Me5 was analysed. Approximately 20% of these boutons were dopamine-immunoreactive. Morphological analysis showed that the dopaminergic synaptic terminals, axodendritic as well as axosomatic, were exclusively of the S- and G-bouton type; they contained, respectively, small spherical vesicles or small pleomorphic vesicles in combination with large granular dense-cored vesicles. All dopaminergic varicosities in the Me5 were of the G-bouton type. Quantitative analysis revealed that most of the dopaminergic synaptic terminals in the Me5 nucleus contacted dendrites, while only a minority (12%) contacted Me5 somata. This dopaminergic somatic input comprised about half (52%) of the total axosomatic input on Me5 neurons. The present results and previous findings with respect to the prominent serotonergic component of the axosomatic input to Me5 neurons indicate that dopamine and serotonin account for most of the axosomatic input in the ventrocaudal part of the Me5 nucleus. In fact, the present results seem to support previous observations regarding the existence of a population of afferent neurons in which dopamine and serotonin are colocalized.


Subject(s)
Dopamine/analysis , Mesencephalon/chemistry , Trigeminal Nuclei/chemistry , Animals , Immunohistochemistry , Male , Mesencephalon/ultrastructure , Microscopy, Immunoelectron , Rats , Rats, Wistar , Trigeminal Nuclei/ultrastructure
16.
J Chem Neuroanat ; 11(1): 57-65, 1996 Jul.
Article in English | MEDLINE | ID: mdl-8841889

ABSTRACT

We have examined the expression of calreticulin in rat spinal motoneurons in order to reveal the occurrence and distribution of Ca2(+)-storage organelles in these neurons. Calreticulin, the non-muscle equivalent of calsequestrin, is the low-affinity, high-capacity calcium-binding protein responsible for intracompartmental Ca2(+)-storage in a number of different cell types. The results of the present immunohistochemical study show that all spinal motoneurons express calreticulin at approximately the same level; no significant differences in cytoplasmic immunostaining intensity were observed between different motoneuron pools or between small and large spinal motoneurons. Immunoelectron microscopy revealed that the intracellular localization of calreticulin within spinal motoneurons was confined to the endoplasmic reticulum and to spherical or pleiomorphic, frequently 'coated' vesicles with a diameter ranging between 120 and 150 nm. Some of these vesicles may represent the so-called calciosomes, the intracellular Ca2(+)-storage vesicles described in liver cells and in cerebellar Purkinje cells. The molecular components responsible for the uptake and release of Ca2+ from the Ca2(+)-storage organelles in spinal motoneurons still remain to be identified.


Subject(s)
Calcium-Binding Proteins/biosynthesis , Motor Neurons/metabolism , Ribonucleoproteins/biosynthesis , Spinal Cord/metabolism , Animals , Calcium/metabolism , Calreticulin , Female , Immunoenzyme Techniques , Motor Neurons/ultrastructure , Organelles/metabolism , Rats , Rats, Wistar , Spinal Cord/ultrastructure
17.
Acta Anat (Basel) ; 155(1): 50-6, 1996.
Article in English | MEDLINE | ID: mdl-8811115

ABSTRACT

With the use of postembedding electron-microscopic immunogold cytochemistry, the vesicular distribution of serotonin within serotonergic synaptic terminals in the mesencephalic trigeminal nucleus was determined in order to obtain further insight into the mechanisms and functional significance of serotonin release to these jaw muscle spindle afferent neurons. Immunogold labelling was restricted to the previously described type I and type II terminals. The distribution of immunogold particles over the synaptic terminals indicated that serotonin was stored in small round or pleomorphic (RSV) vesicles and in large granular dense-cored (DSV) vesicles. The amount of serotonin present in the DSV vesicles, which are generally considered to contain colocalized neuropeptides, showed a high degree of variation. These DSV vesicles were usually located at some distance from the synaptic area of the terminals suggesting that they represent a nondirectly releasable vesicle pool. The serotonergic RSV vesicles were, in general, irregularly distributed over the terminals. However, in about 19% of the analyzed labelled synaptic terminals serotonergic RSV vesicles were clustered together near their release site at the synaptic cleft. These synaptic terminals may represent highly active serotonergic synapses with a high release frequency and a high reuptake level, resulting in a dense concentration of vesicles containing a high amount of serotonin near the synaptic cleft.


Subject(s)
Immunohistochemistry , Mesencephalon/chemistry , Presynaptic Terminals/chemistry , Serotonin/metabolism , Trigeminal Nuclei/ultrastructure , Animals , Cytoplasmic Granules/ultrastructure , Male , Mesencephalon/metabolism , Mesencephalon/ultrastructure , Microscopy, Electron , Presynaptic Terminals/metabolism , Presynaptic Terminals/ultrastructure , Rats , Rats, Wistar , Serotonin/immunology , Synaptic Vesicles/chemistry , Synaptic Vesicles/ultrastructure , Trigeminal Nuclei/metabolism
18.
Restor Neurol Neurosci ; 7(4): 199-205, 1995 Jan 01.
Article in English | MEDLINE | ID: mdl-21551790

ABSTRACT

The objective of this study was to develop an optimal dissection procedure for fetal porcine ventral mesencephalon (VM) grafts and to characterize the cellular composition of such an explant, in particular with respect to the dopaminergic and GABAergic components. We have used a monolayer cell culture system to study and identify the various VM cell types. The in vitro development of the fetal VM cells and the effect of the addition of brain-derived neurotrophic factor (BDNF) was investigated during a culture period of 5 days. Extracellular dopamine levels were measured by means of high performance liquid chromatography (HPLC) with electrochemical detection (LCEC). Our results indicate that the ratio of dopaminergic to GABAergic neurons changed in favour of the dopaminergic component when a more selective dissection technique was used. Although addition of BDNF to the cultures appeared to exert trophic influences on all the cellular components of pig fetal VM, this effect was most pronounced on the TH-positive cells. Highest extracellular DA levels were found in the VM culture with the addition of BDNF and when a more selective dissection method was used. Our in vitro findings suggest that porcine fetal dopaminergic cells retain their potential for development and outgrowth after proper explantation and dissociation. Anticipating on the results of ongoing transplantation studies in rat, they suggest that pig fetal VM can be a suitable alternative for the use of fetal human VM as a graft for Parkinson's disease.

19.
Neuroscience ; 63(4): 1125-35, 1994 Dec.
Article in English | MEDLINE | ID: mdl-7700514

ABSTRACT

The expression of neurotrophin-3 messenger RNA was studied by in situ hybridization in rat muscle spindles from the first embryonic stages of their formation until their mature appearance in adult animals. The first expression of neurotrophin-3 messenger RNA in developing muscles was observed at E19 in the firstly formed intrafusal fiber, the nuclear bag2 fiber. High levels of neurotrophin messenger RNA were found in the equatorial region of these intrafusal fibers in thin lines of cytoplasma around and between the packed-up nuclei. From E21 on, neurotrophin-3 messenger RNA was also present in the nuclear bag1 type intrafusal fiber. The expression of neurotrophin-3 messenger RNA in nuclear chain fibers, which were found in muscle spindles from day 6 after birth, was low and insignificant in comparison to the expression in the nuclear bag fibers. After completion of muscle spindle formation around the third week after birth, high levels of neurotrophin-3 messenger RNA remained present in the intrafusal fibers throughout life. During the entire period of muscle formation, examined from E15 on, as well as in mature muscles, no neurotrophin-3 messenger RNA could be detected in extrafusal fibers by in situ hybridization. The exclusive intramuscular expression of neurotrophin-3 messenger RNA in intrafusal fibers during development as well as in mature stages suggests the involvement of neurotrophin-3 in the formation and the maintenance of muscle spindles.


Subject(s)
Muscle Spindles/metabolism , Nerve Growth Factors/biosynthesis , RNA, Messenger/biosynthesis , Animals , Desmin/metabolism , Extremities/embryology , Female , Immunohistochemistry , In Situ Hybridization , Muscle Spindles/embryology , Muscle, Skeletal/embryology , Muscle, Skeletal/metabolism , Neurotrophin 3 , Pregnancy , Rats , Rats, Wistar
20.
Brain Res Dev Brain Res ; 81(1): 57-65, 1994 Aug 12.
Article in English | MEDLINE | ID: mdl-7805287

ABSTRACT

In this study we have examined the calcium-binding protein expression in rat embryonic (E16) dorsal root ganglia (DRG) neurons in vitro in the presence of neurotrophin-3 (NT-3). A comparison was made with the expression of calcium-binding proteins in DRG subpopulations that depended in vitro on nerve growth factor (NGF) or brain-derived neurotrophic factor (BDNF). Our results show that NT-3 promotes the survival of a DRG subpopulation of which over 75% expresses parvalbumin (PV). The majority of these PV-positive NT-3-dependent DRG neurons were large 'type A' neurons. Expression of calbindin-D28k (CaBP) and calretinin (Calr) in the NT-3-dependent DRG population was seen in smaller fractions (between 12 and 17%) of the surviving DRG neurons and in both type A and B neurons. The preferential expression of PV in NT-3-dependent type A neurons is unique in comparison to the expression of PV and the other calcium-binding proteins in DRG neurons surviving in vitro in the presence of NGF or BDNF.


Subject(s)
Calcium-Binding Proteins/biosynthesis , Ganglia, Spinal/metabolism , Nerve Growth Factors/physiology , Animals , Calbindin 1 , Calbindin 2 , Calbindins , Cell Size/physiology , Cell Survival/physiology , Cells, Cultured , Eye Proteins/biosynthesis , Ganglia, Spinal/ultrastructure , Immunohistochemistry , Neurites/metabolism , Neurites/ultrastructure , Neurons/metabolism , Neurons/ultrastructure , Neurotrophin 3 , Parvalbumins/biosynthesis , Rats , Rats, Wistar , S100 Calcium Binding Protein G/biosynthesis
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