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C N Svendsen

Publications and source records attributed to C N Svendsen.

At least 37 records · Page 2Linked to original sources
PubMed1999

Human neural stem cells: isolation, expansion and transplantation.

Neural stem cells, with the capacity to self renew and produce the major cell types of the brain, exist in the developing and adult rodent central nervous system (CNS). Their exact function and distribution is currently being assessed, but they represent an interesting cell population, which may be used to study factors important for the differentiation of neurons, astrocytes and oligodendrocytes. Recent evidence suggests that neural stem cells may also exist in both the developing and adult human CNS. These cells can be grown in vitro for long periods of time while retaining the potential to differentiate into nervous tissue. Significantly, many neurons can be produced from a limited number of starting cells, raising the possibility of cell replacement therapy for a wide range of neurological disorders. This review summarises this fascinating and growing field of neurobiology, with a particular focus on human tissues.

Animals↗
PubMed1998

A new method for the rapid and long term growth of human neural precursor cells.

A reliable source of human neural tissue would be of immense practical value to both neuroscientists and clinical neural transplantation trials. In this study, human precursor cells were isolated from the developing human cortex and, in the presence of both epidermal and fibroblast growth factor-2, grew in culture as sphere shaped clusters. Using traditional passaging techniques and culture mediums the rate of growth was extremely slow, and only a 12-fold expansion in total cell number could be achieved. However, when intact spheres were sectioned into quarters, rather than mechanically dissociated, cell cell contacts were maintained and cellular trauma minimised which permitted the rapid and continual growth of each individual quarter. Using this method we have achieved a 1.5 million-fold increase in precursor cell number over a period of less than 200 days. Upon differentiation by exposure to a substrate, cells migrated out from the spheres and formed a monolayer of astrocytes and neurons. No oligodendrocytes were found to develop from these human neural precursor cells at late passages when whole spheres were differentiated. This simple and novel culture method allows the rapid expansion of large numbers of non-transformed human neural precursor cells which may be of use in drug discovery, ex vivo gene therapy and clinical neural transplantation.

Automation↗
PubMed1998

Fibroblast growth factor 2 (FGF-2) promotes acquisition of epidermal growth factor (EGF) responsiveness in mouse striatal precursor cells: identification of neural precursors responding to both EGF and FGF-2.

Epidermal growth factor (EGF) and fibroblast growth factor 2 (FGF-2) induce the proliferation of neural precursor cells isolated from specific regions of the embryonic and adult brain. However, the lineage relationship between the EGF- and FGF-2-responsive cells is unknown. In this study we used phosphorylation of the transcription factor cAMP response element-binding protein as a functional readout to identify cells responding to EGF and FGF-2. In primary cultures of mouse embryonic day 14 (E14) striatum, maintained in vitro for 24 hr, 12% of the cells responded to FGF-2, whereas no response to EGF could be detected. Seventy-five percent of these FGF-2-responsive cells were beta tubulin III (TuJ1)-positive neurons, and 25% expressed nestin, a marker for neuroepithelial precursors. After growth factor treatment for 6 d, a population of nestin-positive cells responding to both EGF and FGF-2 were identified. The 6-d-old cultures also contained a small number of TuJ1-positive cells that responded to FGF-2 only. Priming of striatal cells for 24 hr with FGF-2 but not with EGF was sufficient to induce the appearance of EGF- and FGF-2 responsive cells after only 2 d in vitro. Thus, neural precursor cells from the mouse E14 striatum initially responding to FGF-2 only acquire EGF responsiveness later during in vitro development. At this stage EGF and FGF-2 act on the same cells. The acquisition of EGF responsiveness is promoted by FGF-2.

Animals↗
PubMed1998

Dehydroepiandrosterone (DHEA) and DHEA-sulfate (DHEAS) protect hippocampal neurons against excitatory amino acid-induced neurotoxicity.

DHEA, together with DHEAS, is the most abundant steroid in the blood of young adult humans. Levels in humans decline with age and during certain types of illness or stress. We have found that DHEA(S) can prevent or reduce the neurotoxic actions in the hippocampus of the glutamate agonists N-methyl-D-aspartic acid (NMDA) both in vitro and in vivo or alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) and kainic acid in vitro. Pre-treatment with DHEA (10-100 nM for 6-8 h) protected primary hippocampal cultures from embryonic day 18 (E18) embryos against NMDA-induced toxicity (0.1, 1, 10, and 50 mM). DHEA added either with NMDA (1 mM) or 1 h later had lesser, but still significant, protective actions. DHEAS also reduced NMDA-induced toxicity (1 mM), although the lowest effective dose of DHEAS (100 nM) was higher than that of DHEA (10 nM). DHEA (100 nM) protected cultured neurons against the neurotoxic actions of either AMPA (25 microM) or kainic acid (1 mM) as well. In vivo, s.c. pellets of DHEA, which resulted in plasma levels that resembled those in young adult humans, protected hippocampal CA1/2 neurons against unilateral infusions of 5 or 10 nmol of NMDA. Because the release of glutamate has been implicated in the neural damage after cerebral ischemia and other neural insults, these results suggest that decreased DHEA levels may contribute significantly to the increased vulnerability of the aging or stressed human brain to such damage.

Animals↗
PubMed1998

Heparin, but not other proteoglycans potentiates the mitogenic effects of FGF-2 on mesencephalic precursor cells.

There is increasing evidence that the proteoglycan heparin plays a critical role in the regulation of the activity of FGF-2 by either interacting with its receptor or modifying its stability and functioning. In this study precursor cells were isolated from the rat E14 ventral mesencephalon and cultured as free floating spheres in FGF-2 alone or in combination with heparin or other related proteoglycans, including chondroitin sulfate, keratin sulfate, dermatan sulfate, or hyaluronic acid. Our results show the mitogenic effects of FGF-2 could be potentiated by heparin but not the other four proteoglycans. Sodium chlorate, which blocks the cells ability to sulfate its proteoglycans, was shown to reduce the mitogenic effects of FGF-2 alone to below that of control levels, suggesting that endogenous sulfated molecules are required for the FGF-2 effects on mesencephalic precursors. Cells expanded for 7 days with either FGF-2 or FGF-2 + heparin were plated onto a substrate and allowed to differentiate for a further 7 days in the absence of growth factors. Approximately 6% of the precursors developed into neurons whether grown with or without heparin and none were positive for TH, a marker for dopamine neurons. However, there was a significant decrease in the number of astrocytes developing from cultures grown in FGF-2 + heparin when compared to FGF-2 alone. Interestingly we could not find an EGF responsive cell in the mesencephalon at this embryonic age in the absence or presence of heparin. However, there was a synergistic effect of combining EGF + FGF-2, which could be potentiated by heparin. We conclude that heparin, but not other closely related proteoglycans, is vital for the growth of FGF-2-responsive mesencephalic neural precursors.

Animals↗
PubMed1997

Restricted growth potential of rat neural precursors as compared to mouse.

Epidermal growth factor (EGF) responsive precursors isolated from the developing mouse striatum could be continually expanded in culture as free-floating spheres of cells for over 50 days. Under identical conditions, EGF-responsive precursors from the developing rat striatum could only be expanded for between 21 and 28 days, after which crisis ensued and there was a reduction in cell number at each passage. The outer regions of 28-day-old rat spheres contained a heterogeneous population of both dividing and dying cells while the cores were full of dying cells, many of which showed features consistent with apoptosis. Fibroblast growth factor-2 (FGF-2) alone did not lead to an expansion in rat striatal precursor cell number under the conditions used here. EGF combined with FGF-2 acted synergistically on cell growth, but did not prevent the final senescence and death of the rat precursors.

Animals↗
PubMed1997

Co-expression of MAP-2 and GFAP in cells developing from rat EGF responsive precursor cells.

In this study we have performed a detailed analysis of EGF-responsive precursors as they develop into neurons and astrocytes using antibodies to nestin, microtubule-associated protein 2 (MAP-2c and MAP-2ab) and glial fibriallary acidic protein (GFAP). Surprisingly, at early time points, most GFAP-positive cells also stained for MAP-2c, and we postulate that this may be a normal stage of astroglial development. At 7 days most of the cells had developed into astrocytes and MAP-2ab-positive cells only represented 5% of the total neuronal population. This study shows that (i) MAP-2c is a marker for early precursors, (ii) the majority of cells developing from. EGF-responsive precursors develop into glia and (iii) only a small population of cells arising from expanded populations of EGF-responsive precursors develop into neurons expressing MAP-2ab. Thus, certain critical signals important for full neuronal differentiation may be missing from this system.

Animals↗
PubMed1997

Long-term survival of human central nervous system progenitor cells transplanted into a rat model of Parkinson's disease.

Progenitor cells were isolated from the developing human central nervous system (CNS), induced to divide using a combination of epidermal growth factor and fibroblast growth factor-2, and then transplanted into the striatum of adult rats with unilateral dopaminergic lesions. Large grafts were found at 2 weeks survival which contained many undifferentiated cells, some of which were migrating into the host striatum. However, by 20 weeks survival, only a thin strip of cells remained at the graft core while a large number of migrating astrocytes labeled with a human-specific antibody could be seen throughout the striatum. Fully differentiated graft-derived neurons, also labeled with a human-specific antibody, were seen close to the transplant site in some animals. A number of these neurons expressed tyrosine hydroxylase and were sufficient to partially ameliorate lesion-induced behavioral deficits in two animals. These results show that expanded populations of human CNS progenitor cells maintained in a proliferative state in culture can migrate and differentiate into both neurons and astrocytes following intracerebral grafting. As such these cells may have potential for development as an alternative source of tissue for neural transplantation in degenerative diseases.

Amphetamine↗
PubMed1996

GDNF enhances dopaminergic cell survival and fibre outgrowth in embryonic nigral grafts.

Two groups of rats with unilateral 6OHDA lesions received either intrastriatal suspension grafts of embryonic ventral mesencephalon or sham grafts. Three subgroups of each of these received intrastriatal infusions of 1000 ng or 500 ng glial cell-line derived trophic factor (GDNF) or vehicle alone for 10 consecutive days. There was a highly significant dose-dependent effect of GDNF both on the number of TH-positive cells surviving in the grafts and on the density of fibre outgrowth. All grafted rats showed rapid compensation of amphetamine-induced rotation compared with rats with sham grafts. GDNF may provide a powerful tool to enhance the survival and maturation of dopaminergic neurones within mesencephalic transplants.

Animals↗
PubMed1996

Survival and differentiation of rat and human epidermal growth factor-responsive precursor cells following grafting into the lesioned adult central nervous system.

Epidermal Growth Factor (EGF)-responsive stem cells isolated from the developing central nervous system (CNS) can be expanded exponentially in culture while retaining the ability to differentiate into neurons and glia. As such, they represent a possible source of tissue for neural transplantation, providing they can survive and mature following grafting into the adult brain. In this study we have shown that purified rat stem cells generated from either the embryonic mesencephalon or the striatum can survive grafting into the striatum of rats with either ibotenic acid or nigrostriatal dopamine lesions. However, transplanted stem cells do not survive as a large mass typical of primary embryonic CNS tissue grafts, but in contrast form thin grafts containing only a small number of surviving cells. There was no extensive migration of transplanted stem cells labeled with either the lac-z gene or bromodeoxyuridine into the host region surrounding the graft, although a small number of labeled cells were seen in the ventral striatum some distance from the site of implantation. Some of these appeared to differentiate into dopamine neurons, particularly when the developing mesencephalon was used as the starting material for generating the stem cells. EGF-responsive stem cells could also be isolated from the mesencephalon of developing human embryos and expanded in culture, but only grew in large numbers when the gestational age of the embryo was greater than 11 weeks. Purified human CNS stem cells were also transplanted into immunosuppressed rats with nigrostriatal lesions and formed thin grafts similar to those seen when using rat stem cells. However, when primary cultures of human mesencephalon were grown with EGF for only 10 days and this mixture of stem cells and primary neural tissue was transplanted into the dopamine-depleted striatum, large well-formed grafts developed. These contained mostly small undifferentiated cells intermixed with a number of well-differentiated TH-positive neurons. These results show that purified populations of rat or human EGF-responsive CNS stem cells do not form large graft masses or migrate extensively into the surrounding host tissues when transplanted into the adult striatum. However, modifications of the growth conditions in vitro may lead to an improvement of their survival in vivo.

Animals↗
PubMed1996

Reduced retrograde labelling with fluorescent tracer accompanies neuronal atrophy of basal forebrain cholinergic neurons in aged rats.

During ageing, basal forebrain cholinergic neurons are prone to degeneration for unknown reasons. In this study we morphometrically evaluated the retrograde labelling of basal forebrain neurons obtained after injection of FluoroGold into multiple sites in the cerebral neocortex in aged (24-33 months) as compared with young adult (four to six months) male Sprague-Dawley rats. In addition, we looked for differences in the distribution of degenerative changes in topographic subdivisions of the basal forebrain cholinergic complex of neurons identified by immunohistochemical detection of the cholinergic markers choline acetyltransferase or low-affinity neurotrophin receptor. After injection of FluoroGold into the cerebral neocortex, the number of retrogradely labelled neurons in the horizontal diagonal band/ substantia innominata and basal nucleus was significantly lower in aged rats, by 41% and 48%, respectively. In aged rats injected with FluoroGold as well as in non-injected aged rats, the numbers of neurons immunoreactive for choline acetyltransferase and low-affinity neurotrophin receptor were significantly lower, by 23-27% in the basal forebrain system as a whole, with no significant difference in the degree of decline amongst different subdivisions (i.e. medial septum, diagonal band, substantia innominata and basal nucleus). The ratios of the number of neurons labelled with FluoroGold as compared with the number of neurons immunoreactive for either cholinergic marker were significantly lower in aged rats, by 32-37%, indicating that the decline in the number of neurons retrogradely transporting tracer was greater than the decline in the number of immunoreactive neurons in aged animals. Immunoreactive as well as retrogradely labelled neurons showed a significant shrinkage of cell surface area of 6-13% in different subdivisions of the basal forebrain cholinergic system in aged rats. These findings confirm significant loss and atrophy of basal forebrain cholinergic neurons in aged rats, and demonstrate significantly reduced retrograde labelling of these neurons with fluorescent tracer applied to their target cortex. This reduced retrograde labelling suggests an impairment of either uptake or retrograde transport mechanisms in these neurons in aged rats. Such an impairment may contribute to the degenerative changes of basal forebrain cholinergic neurons observed in ageing and age-related degenerative conditions such as Alzheimer's disease.

Acetylcholine↗
PubMed1996

Do central nervous system neurons require target-derived neurotrophic support for survival throughout adult life and aging?

Many growth factors that have effects on neurons are present in the developing and mature central nervous system where their functions are poorly understood. In the peripheral nervous system, target-derived growth factors can regulate the survival of developing afferent neurons. Recent studies suggest that neurotrophins derived from target neurons may also regulate the survival of afferent basal forebrain cholinergic neurons during development of the central nervous system. However, in mature animals, these cholinergic neurons do not appear to require target-derived neuroptrophins for survival. Similar findings have also been reported for dopaminergic neurons of the substantia nigra. Although further studies are required, available experimental evidence suggests that target-derived growth factors may influence neuronal phenotypes such as axonal sprouting or transmitter production instead of survival in mature animals.

Aging↗
PubMed1996

Bilateral striatal lesions impair retention of an operant test of short-term memory.

It has been previously shown that lesions of the dorsal striatum can disrupt performance on a variety of cognitive tasks related to prefrontal cortex function. In order to extend these studies, we have compared the effects of bilateral striatal lesions on retention of an operant test of short-term memory using a delayed matching to position task. Rats were initially pretrained on the matching task. Then, one group of animals received ibotenic acid lesions in the dorsal striatum and a second group received sham lesions. The striatal lesions induced marked deficits on delayed matching performance when the rats were retested one week following surgery. The delay-independent pattern of deficit observed does not suggest a primary impairment in short-term memory function. The lesion induced deficits were temporary, with recovery seen in the lesion group back to a control level of performance in 2 additional test sessions 5 and 10 weeks post-lesion. A follow up study investigating the time course of the deficit suggests that the recovery seen in performance measures cannot be attributed to the elapse of time per se, but rather is due to re-learning. In addition to the performance deficits in the operant task, the lesioned rats exhibited marked nocturnal locomotor hyperactivity. The results indicate that bilateral striatal lesions induce marked deficits in retention of the delayed matching to position task. They suggest a key role for the neostriatum in the execution of pre-learned responses, but do not exclude the possibility of additional involvement in short-term memory functions under certain testing conditions.

Animals↗
PubMed1995

The effects of bilateral striatal lesions on the acquisition of an operant test of short term memory.

It has been previously shown that lesions of the dorsal striatum can disrupt performance on a variety of cognitive tasks related to frontal cortex function. In order to extend these studies, we have investigated the effects of bilateral striatal lesions on the acquisition of an operant test of short term memory in the delayed non-matching to position paradigm. The animals received either ibotenic acid or saline control injections into the dorsal striatum prior to training on the non-matching task. Striatal lesions retarded acquisition of the task, although with further training the lesioned rats achieved a similar level of asymptotic performance to the control animals. The lesioned rats also exhibited marked nocturnal locomotor hyperactivity when tested under conditions of food deprivation, but not when tested satiated. The results indicate that bilateral striatal lesions induce mild deficits in the acquisition of the discrimination rules involved in performance of the delayed non-matching to position task. The present study does not support a role for the neostriatum in the specific mediation of short term memory in a operant DNMTP test.

Animals↗
PubMed1995

Increased survival of rat EGF-generated CNS precursor cells using B27 supplemented medium.

Previous studies suggest that a population of precursor cells from the developing and adult mouse striatum can be expanded in culture using serum-free, N2-supplemented medium and mitogenic factors such as epidermal growth factor (EGF). Here we show that EGF-responsive precursor cells from embryonic rat striatum and mesencephalon can also be expanded in culture, incorporate bromodeoxy uridine (BrDU) and develop into spheres that either adhere to the surface of the culture dish or float freely in the medium. Addition of B27, a medium supplement that increases neuronal survival in primary CNS cultures, resulted in a tenfold increase in the number of proliferating cells in vitro over the first week. The effects of B27-supplemented medium on precursor cell survival were only seen when primary cultures were used, such that dividing cells grown in B27 for 1 week could then be transferred to either B27 or N2 medium and show similar survival and division rates in response to EGF. After 1, 2 or 4 weeks of growth in B27-supplemented medium, dissociated precursor cells from either striatal or mesencephalic cultures could be differentiated when exposed to a poly-l-lysine-coated substrate in serum and EGF-free medium supplemented with B27. These cells then matured into a mixed culture containing neurons (approximately 35% of cells), astrocytes (approximately 44% of cells), and oligodendrocytes (approximately 10% of cells), based on immunocytochemical staining with microtuble-associated protein (MAP2), glial fibriallary acidic protein and galactocerebrosidase. When whole spheres of precursor cells were allowed to differentiate, every one examined was found to generate neurons, astrocytes and oligodendrocytes in similar proportions.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1994

Death of developing septal cholinergic neurons following NGF withdrawal in vitro: protection by protein synthesis inhibition.

Fetal septal neurons were grown in vitro under glass coverslips. This sandwich culture method significantly increased general neuronal survival, reduced glial proliferation, and permitted the removal of serum from the growth medium after 5 d in vitro. Thereafter, a simple, and completely defined, medium was used, and the effects of NGF, NGF withdrawal, and protein synthesis inhibition were examined on septal cholinergic neurons. NGF added to septal cultures at the time of plating resulted in a threefold increase in the number of cholinergic neurons seen at 14 d in vitro but had no effect on the survival of non-cholinergic cells. Cholinergic neurons identified by staining for AChE, ChAT, and p75NGFR could be maintained in serum-free, NGF-supplemented medium for over 40 d. When NGF was removed and NGF antibodies added to 14-d-old cultures, less than 30% of cholinergic neurons survived a further 4 d, but when NGF was similarly withdrawn from 35-d-old cultures, over 75% of cholinergic neurons survived. Reapplication of NGF after 3 but not after 12 or more hours of NGF withdrawal from 14-d-old cultures prevented the death of most cholinergic neurons. When NGF was withdrawn from 14-d-old cultures in the presence of the protein synthesis inhibitor cycloheximide, over 75% of the cholinergic neurons survived. These findings suggest that septal cholinergic neurons are dependent on NGF for survival only during a critical period of development and that growth factor-regulated developmental cell death may occur in CNS neurons by activation of programmed cell death requiring protein synthesis.

Animals↗