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O Isacson

Publications and source records attributed to O Isacson.

142 records · Page 8Linked to original sources
PubMed1985

Monitoring of cell viability in suspensions of embryonic CNS tissue and its use as a criterion for intracerebral graft survival.

Neuronal cell suspensions, prepared by trypsination and mechanical disruption from embryonic CNS tissue, are currently used for intracerebral neuronal grafting to deep brain sites. In the present study the viability of suspended neurons from different brain regions has been monitored with a fluorescent vital stain, and studied as a function of time after dissociation and age of the donor rat embryos. Subsequently, the validity of the in vitro viability rates as a criterion for the in vivo survivability of each individual suspension was tested for suspensions prepared from the developing mesencephalon, rich in dopamine-containing neurons. The results indicate that mechanically dissociated embryonic CNS neurons remain viable for several hours in a simple glucose-saline solution at room temperature. The in vitro viability scores declined faster in suspensions prepared from mesencephalon than in those prepared from telencephalon (striatum and basal forebrain), and they declined faster in suspensions prepared from older embryos. The fetal cells were sensitive to the mechanical trauma caused by excessive pipetting, and tissue from older embryos seemed generally more vulnerable in the trypsination-dissociation procedure. The grafting experiments showed a good correlation between the in vitro cell viability counts and in vivo neuronal survival after grafting, indicating that the vital stain, at least under certain conditions, can be used as a simple and practical routine test to check and standardize cell suspensions to be used in intracerebral grafting experiments.

Animals↗
PubMed1985

Neural grafting in a rat model of Huntington's disease: progressive neurochemical changes after neostriatal ibotenate lesions and striatal tissue grafting.

The acute and long-term changes following large neostriatal ibotenic acid lesions and intrastriatal striatal neuronal grafting have been studied neurochemically by determinations of the gamma-aminobutyrate (GABA) and acetylcholine-synthetic enzymes, glutamate decarboxylase and choline acetyltransferase, and of dopamine and its primary acidic metabolite, 3,4-dihydroxyphenylacetic acid. The neurochemical data have been matched with estimates of tissue volume changes and striatal graft development through tissue weight and protein content analysis and histological volumetric measurements. Injections of 20 micrograms ibotenic acid, deposited over four injection sites in the head of the caudate-putamen, had by one week caused a 70-85% reduction in glutamate decarboxylase and choline acetyltransferase activity. With time there was a progressive recovery of the specific enzyme activities in the lesioned areas (expressed per microgram protein or per mg wet weight) to 40-60% of control levels as determined at 20 weeks post-lesion in the caudate-putamen. This increase was, however, largely if not exclusively due to the long-term shrinkage of the lesioned caudate-putamen, amounting to 50-70% at 20 weeks. Thus, the total glutamate decarboxylase and choline acetyltransferase activity levels in the lesioned caudate-putamen remained virtually unchanged (between 15 and 25% of control) over the 20 week experimental period. Glutamate decarboxylase activity was also markedly reduced (35-70%) in the two primary striatal projection areas, globus pallidus and substantia nigra, and there were no signs of recovery over time. Striatal dopamine levels, which were acutely unaffected by the lesion, showed a slow decline so that the total dopamine content in the area was reduced by about 80% at 20 weeks. Suspended striatal neurons obtained from the striatal primordia of 14-15-day-old rat fetuses, injected into the previously lesioned caudate-putamen, survived and established a new striatum-like structure at the site of the ibotenic acid lesion. The final volume of the graft tissue reached up to about 10 mm3 in volume and reduced striatal atrophy on average from about 50 to 70% of normal control in the rats with lesions to about 30-40% in the animals with grafts. In the rats with grafts, there was a significant recovery of glutamate decarboxylase and choline acetyltransferase activities not only in the lesioned caudate-putamen, containing the graft (from 20-25% to 40-50%, when expressed as total enzyme activity levels), but also the glutamate decarboxylase activity in the globus pallidus, a structure located at a distance from the graft.(ABSTRACT TRUNCATED AT 400 WORDS)

3,4-Dihydroxyphenylacetic Acid↗
PubMedDate not supplied

Functional neuronal replacement by grafted striatal neurones in the ibotenic acid-lesioned rat striatum.

In rats, striatal neuronal destruction by so-called excitotoxic amino acids, kainic acid or ibotenic acid (IA) produce neuropathological and neurochemical changes in the basal ganglia which resemble those seen in patients with Huntington's chorea. Such lesioned animals show a behavioural syndrome which is reminiscent of the cardinal symptoms of the disease, accompanied by a substantial increase in local cerebral metabolic activity in several striatal target structures within the extrapyramidal motor system. The study was designed to explore the potential of grafted fetal striatal neurones implanted into the IA-lesioned striatum to compensate for the structural, neurochemical, metabolic and behavioural defects of IA-lesioned rats. Extending previous studies, we report here that such striatal implants can significantly ameliorate the lesion-induced locomotor hyperactivity and at least partly normalize the metabolic hyperactivity in the extrapyramidal neuronal system.

Animals↗
PubMedDate not supplied

Intracerebral grafting of embryonic neural cells into the adult host brain: an overview of the cell suspension method and its application.

An overview is presented of general principles for intracerebral grafting of embryonic brain tissue to the adult mammalian brain. Special reference is made to the development and use of the dissociated neuronal cell suspension method. Examples are drawn primarily from experiments where embryonic ventral mesencephalon is transplanted to adult striatum and embryonic septal-diagonal band area is transplanted to the hippocampal formation. Results related to the cell viability in vitro and in vivo and to axonal outgrowth are the main focuses of this overview.

Acetylcholinesterase↗
PubMedDate not supplied

Porcine xenografts in Parkinson's disease and Huntington's disease patients: preliminary results.

The observation that fetal neurons are able to survive and function when transplanted into the adult brain fostered the development of cellular therapy as a promising approach to achieve neuronal replacement for treatment of diseases of the adult central nervous system. This approach has been demonstrated to be efficacious in patients with Parkinson's disease after transplantation of human fetal neurons. The use of human fetal tissue is limited by ethical, infectious, regulatory, and practical concerns. Other mammalian fetal neural tissue could serve as an alternative cell source. Pigs are a reasonable source of fetal neuronal tissue because of their brain size, large litters, and the extensive experience in rearing them in captivity under controlled conditions. In Phase I studies porcine fetal neural cells grafted unilaterally into Parkinson's disease (PD) and Huntington's disease (HD) patients are being evaluated for safety and efficacy. Clinical improvement of 19% has been observed in the Unified Parkinson's Disease Rating Scale "off" state scores in 10 PD patients assessed 12 months after unilateral striatal transplantation of 12 million fetal porcine ventral mesencephalic (VM) cells. Several patients have improved more than 30%. In a single autopsied PD patient some porcine fetal VM cells were observed to survive 7 months after transplantation. Twelve HD patients have shown a favorable safety profile and no change in total functional capacity score 1 year after unilateral striatal placement of up to 24 million fetal porcine striatal cells. Xenotransplantation of fetal porcine neurons is a promising approach to delivery of healthy neurons to the CNS. The major challenges to the successful use of xenogeneic fetal neuronal cells in neurodegenerative diseases appear to be minimizing immune-mediated rejection, management of the risk of xenotic (cross-species) infections, and the accurate assessment of clinical outcome of diseases that are slowly progressive.

Adult↗
PubMedDate not supplied

Neurotrophic factors NGF and FGF-2 alter levels of huntingtin (IT15) in striatal neuronal cell cultures.

A mutation of the human IT15 gene is responsible for Huntington's disease (HD) and the causative factor in the major neuronal loss observed in the striatum. The growth factors basic fibroblast growth factor (FGF-2), nerve growth factor (NGF), and brain-derived neurotrophic factor (BDNF) improve survival and promote differentiation of striatal neurons, as well as exert a neuroprotective effect when such neurons are challenged with metabolic toxins or excitatory amino acids. Using Western blotting and striatal cell cultures, we found that FGF-2 increased the level of huntingtin in a dose-dependent fashion, whereas NGF decreased huntingtin expression. The neurotrophic factor-specific, dose-dependent effect on striatal levels of huntingtin may be relevant to understanding the normal role of IT15 and developing new therapies against the disease provoking mutated IT15.

Animals↗
PubMedDate not supplied

Extensive axonal and glial fiber growth from fetal porcine cortical xenografts in the adult rat cortex.

Axonal growth from cortically placed fetal neural transplants to subcortical targets in adult hosts has been difficult to demonstrate and is assumed to be minimal; however, experiments using xenogeneic neural grafts of either human or porcine fetal tissues into the adult rat striatum, mesencephalon, and spinal cord have demonstrated the capability for long-distance axonal growth. This study reports similar results for porcine cortical xenografts placed in the adult rat cerebral cortex and compares these findings with results from cortical allografts. Adult rats that previously received unilateral cortical lesions by an oblique intracortical stereotaxic injection of quinolinic acid, were implanted with suspensions of either E14 rat or E38 xenogeneic porcine fetal cortical cells. Xenografted rats were immunosuppressed by cyclosporin A. The corpus callosum was intact in all cases and grafts were confined to the overlying cortex. After a 31-34 wk posttransplant survival period, acetylcholinesterase (AChE) staining and tyrosine hydroxylase (TH) immunocytochemistry revealed that both allo- and xenografts received host afferents. Retrograde tracer injections into the ipsilateral striatum and cerebral peduncle in allografted animals failed to show any axonal growth to either subcortical target. Using a porcine-specific axonal marker in xenografted animals, we found graft axons in white matter tracts (corpus callosum, internal capsule, cingulum bundle, and medial forebrain bundle) and within the caudate-putamen and both the ipsilateral and contralateral cerebral cortex. Graft axons were not found in the thalamus, midbrain, or spinal cord. In addition, using an antibody to porcine glial fibers, we observed more extensive graft glial fiber growth into the same host fiber tracts, as far caudally as the cerebral peduncle, but not into gray matter targets outside the cortex. These results demonstrate that porcine cortical xenograft axons and glia can extend from lesioned cerebral cortex to cortical and subcortical targets in the adult rat brain. These findings are relevant for prospects of repairing cortical damage and obtaining functional recovery.

Acetylcholinesterase↗
PubMedDate not supplied

Development of the human striatum: implications for fetal striatal transplantation in the treatment of Huntington's disease.

Fetal neural transplantation has recently been demonstrated to ameliorate motor and other behavioral deficits in animals models of Huntington's disease, and reconstruct many of the damaged striatal circuits. However, there has been significant variability in the histological appearance of these grafts, most likely related to differences of the regions of dissection of the donor tissue. Selective dissection and transplantation of the lateral ventricular eminence in rodents has resulted in grafts consisting of primarily striatal-like tissue. This data, combined with data from our own and other laboratories has led to a description of the development of human striatum, with a particular emphasis on the relevance of human striatal development to the field of fetal tissue transplantation for the treatment of Huntington's disease. If the goal of transplantation is to graft GABAergic striatal projection neurons, it is our impression that optimal grafting results will occur when transplants are derived from the lateral ventricular eminence and the lateral aspect of the body of the ventricular eminence anterior to the foramen of Monro. Optimal results are likely to occur when donor ages range from Stage 19 to 23, with possible graft success when donor age extends to as late as postovulatory week 22.

Fetal Tissue Transplantation↗
PubMedDate not supplied

Embryonic stem cells differentiated in vitro as a novel source of cells for transplantation.

The controlled differentiation of mouse embryonic stem (ES) cells into near homogeneous populations of both neurons and skeletal muscle cells that can survive and function in vivo after transplantation is reported. We show that treatment of pluripotent ES cells with retinoic acid (RA) and dimethylsulfoxide (DMSO) induce differentiation of these cells into highly enriched populations of gamma-aminobutyric acid (GABA) expressing neurons and skeletal myoblasts, respectively. For neuronal differentiation, RA alone is sufficient to induce ES cells to differentiate into neuronal cells that show properties of postmitotic neurons both in vitro and in vivo. In vivo function of RA-induced neuronal cells was demonstrated by transplantation into the quinolinic acid lesioned striatum of rats (a rat model for Huntington's disease), where cells integrated and survived for up to 6 wk. The response of embryonic stem cells to DMSO to form muscle was less dramatic than that observed for RA. DMSO-induced ES cells formed mixed populations of muscle cells composed of cardiac, smooth, and skeletal muscle instead of homogeneous populations of a single muscle cell type. To determine whether the response of ES cells to DMSO induction could be further controlled, ES cells were stably transfected with a gene coding for the muscle-specific regulatory factor, MyoD. When induced with DMSO, ES cells constitutively expressing high levels of MyoD differentiated exclusively into skeletal myoblasts (no cardiac or smooth muscle cells) that fused to form myotubes capable of spontaneous contraction. Thus, the specific muscle cell type formed was controlled by the expression of MyoD. These results provided evidence that the specific cell type formed (whether it be muscle, neuronal, or other cell types) can be controlled in vitro. Further, these results demonstrated that ES cells can provide a source of multiple differentiated cell types that can be used for transplantation.

Animals↗
PubMedDate not supplied

Cell-mediated delivery of brain-derived neurotrophic factor enhances dopamine levels in an MPP+ rat model of substantia nigra degeneration.

Brain-derived neurotrophic factor (BDNF) promotes the survival of fetal mesencephalic dopaminergic cells and protects dopaminergic neurons against the toxicity of MPP+ in vitro. Supranigral implantation of fibroblasts genetically engineered to secrete BDNF attenuates the loss of substantia nigra pars compacta (SNc) dopaminergic neurons associated with striatal infusion of MPP+ in the adult rat. Using this MPP+ rat model of nigral degeneration, we evaluated the neurochemical effects of supranigral, cell-mediated delivery of BDNF on substantia nigra (SN) dopamine (DA) content and turnover. Genetically engineered BDNF-secreting fibroblasts (approximately 12 ng BDNF/24 h) were implanted dorsal to the SN 7 days prior to striatal MPP+ administration. The present results demonstrate that BDNF-secreting fibroblasts, as compared to control fibroblasts, enhance SN DA levels ipsilateral as well as contralateral to the graft without altering DA turnover. This augmentation of DA levels suggests that local neurotrophic factor delivery by genetically engineered cells may provide a therapeutic strategy for preventing neuronal death or enhancing neuronal function in neurodegenerative diseases characterized by dopaminergic neuronal dysfunction, such as Parkinson's disease.

1-Methyl-4-phenylpyridinium↗
PubMedDate not supplied

Differential dissection of the rat E16 ventral mesencephalon and survival and reinnervation of the 6-OHDA-lesioned striatum by a subset of aldehyde dehydrogenase-positive TH neurons.

The retinoic acid-generating enzyme, aldehyde dehydrogenase (AHD), is expressed in a subpopulation of dopaminergic neurons found in the substantia nigra. Using AHD and tyrosine hydroxylase (TH) as immunohistochemical markers, we determined whether differential dissection of the embryonic (E16) ventral mesencephalon (VM) into its lateral and medial portions contributed equally to the number of TH cells surviving transplantation, if grafted AHD/TH neurons reinnervate the host striatum according to their normal projection patterns, and examined the functional recovery caused by the implanted cells as assessed by amphetamine-induced rotation in a 6-OHDA-lesioned model of Parkinson's disease. The embryonic tissue was transplanted as solid pieces injected via a 20-gauge lumbar puncture needle into the center of the deafferented striatum. Groups received either one complete ventral mesencephalic piece (VM), two medial pieces of ventral mesencephalic tissue (MVM), or two lateral pieces of ventral mesencephalic tissue (LVM). Both VM and MVM groups showed a significant decrease in amphetamine-induced rotation over time and, there was no difference in the degree of reduction observed between the two groups. Histological evaluation of the transplants revealed a much larger total number of surviving TH cells in grafts from the VM and MVM groups compared to the LVM group. Surviving AHD/TH neurons were found in all groups. Whereas TH staining of the transplanted striatum displayed a halo of graft-derived fibers all around the transplant and integration of these fibers into the host neuropil, AHD staining showed a preferential reinnervation of the dorsolateral striatum corresponding to the normal projection pattern of AHD/TH neurons. In summary, selective dissection of the embryonic ventral mesencephalon is possible, functional recovery as assessed by amphetamine-induced rotation in animals transplanted with MVM is similar to that seen in animals grafted with VM, and AHD/TH neurons have a selective reinnervation pattern in the PD transplantation paradigm. These findings may have implications for the grafting of fetal mesencephalic tissue in PD patients.

Age Factors↗
PubMedDate not supplied

Pig fetal septal neurons implanted into the hippocampus of aged or cholinergic deafferented rats grow axons and form cross-species synapses in appropriate target regions.

The anatomical specificity of axon growth from fetal pig septal xenografts was studied by transplanting septal cells from E30-35 pig fetuses into cholinergic deafferented (192-IgG-saporin-infused) rats or into aged rats (> 18 months). Cell suspensions (100,000 cells/microl) were injected bilaterally into the dorsal and ventral hippocampus of immunosuppressed rats (10 mg/kg/day cyclosporine A). To assess axonal growth and synapse formation, acetylcholinesterase histochemistry, an antibody to choline acetyltransferase (ChAT), and three pig-positive/rat-negative antibodies: bovine 70kD neurofilament (NF70), human low-affinity NGF receptor (hNGFr), and human synaptobrevin (hSB) were used. In rats with surviving grafts at 6 months, NF70 axonal labeling was more extensive than either ChAT or hNGFr labeling. All three markers demonstrated graft axons extending selectively through the hippocampal CA fields and the molecular layer of the dentate gyrus. Graft axons did not extend into adjacent entorhinal cortex or neocortex. The distribution of pig hSB-positive synapses correlated with AChE-positive fiber outgrowth in to the host. Electron microscopic analysis of hSB-immunostained hippocampal sections revealed pig presynaptic terminals in contact with normal rat postsynaptic structures in the CA fields and the dentate gyrus. These data demonstrate target-appropriate growth of pig cholinergic axons and the formation of cross-species synapses in the deafferented or aged rat hippocampus.

Afferent Pathways↗
PubMedDate not supplied

Morris water maze analysis of 192-IgG-saporin-lesioned rats and porcine cholinergic transplants to the hippocampus.

Adults rats were lesioned with 192-IgG-saporin, an immunotoxin that targets cholinergic neurons in the basal forebrain expressing the low-affinity nerve growth factor receptor (p75). One month later, rats received E30-35 porcine cholinergic neurons bilaterally into the hippocampus, and were tested in the Morris water maze and the passive avoidance task 4.5-6 months after transplantation (in two experiments, rats were retested in the water maze) followed by histological and cellular analyses. The 192-IgG-saporin-lesioned animals displayed clear cognitive deficits in the Morris water maze. In all experiments the lesioned animals had spatial probe deficits on day 5 testing. A large variance was found among the transplanted animals, with individual animals exhibiting improved performance, but little overall improvement when compared to lesion-alone animals as a group. The relationships between behavioral performance and graft cholinergic factors were established by histological analyses. Grafted animals exhibited an increase in cholinergic innervation of the dentate gyrus (DG) region of the dorsal hippocampus when compared to lesion-alone animals. There was a significant correlation between the level of cholinergic innervation in the dentate gyrus and spatial navigation performance (latency and spatial probe) in the Morris water maze task. These data provide evidence of memory and spatial deficits following cholinergic denervation, and of target-specific growth of xenogeneic cholinergic neurons into the hippocampus. The lack of a clear treatment (transplant) effect in the behavioral measures leads us to believe that functional restoration of cognitive function would require cholinergic reinnervation of both the hippocampus and the neocortex in this 192-IgG-saporin animal model.

Acetylcholine↗
PubMedDate not supplied

Distribution of degeneration of cholinergic neurons in the septum following axotomy in different portions of the fimbria-fornix: a correlation between degree of cell loss and proximity of neuronal somata to the lesion.

The degree of cell loss of immunohistochemically stained cholinergic neurons was quantitatively compared in equivalent regions of the septum in three groups of animals with lesions transecting their axons at different levels in the fimbria-fornix (FF). Locations of different septal regions and of FF lesions were defined according to their distances from fixed anatomical reference points. Individual animals all exhibited a gradient of cholinergic cell loss, such that the severity of cell loss diminished progressively in the rostral and ventral directions as the distance from the lesion increased beyond a certain point. Comparison of animals with FF lesions in different locations showed that this gradient of cell loss shifts in the caudo-rostral and dorso-ventral directions within the septal complex in direct relation to the proximity of the axotomizing FF lesion. These findings suggest that among septal cholinergic neurons there is a fixed spatial relationship between the distance of neuronal somata from an axotomizing FF lesion and the likelihood of neuronal loss in response to axotomy. This relationship could not be defined precisely using the material studied, but simple geometrical calculations showed that within 2500 microns of the lesion cell loss is always pronounced (less than 30 per cent detectable cells vs controls) and beyond 4000 microns cell loss is generally not severe (greater than 80 per cent detectable cells). Between these two distances cell loss diminishes in a gradiated manner. Thus, relatively small differences (1 mm) in the location of FF lesions can lead to marked differences in the severity of degeneration observed in certain equivalent regions of the septum. The findings have implications both for attempts to understand the causes underlying retrograde neuronal degeneration and for studies investigating means of preventing retrograde degeneration of cholinergic septal neurons.

Animals↗