Jane Bottenstein | University of Texas, Medical Branch at Galveston (original) (raw)
Papers by Jane Bottenstein
The Society for Neuroscience Abstracts, May 10, 1989
PubMed, 1981
The requirement for a serum supplement to basal culture medium to support the growth and viabilit... more The requirement for a serum supplement to basal culture medium to support the growth and viability of cells in vitro has been a major hindrance to reducing and thereby controlling experimental variables. The complexity and undefined nature of serum as well as the variability of serum lots make it difficult to reproduce data and to interpret results of experiments. This paper presents preliminary results obtained with C6 glioma cell cultures in which serum was replaced by growth factors and a hormone. A brief review of the use of serum-free defined medium for the culture of tumor cells and the potential uses of defined media that are cell-type specific are also presented.
Journal of Neurochemistry, Jun 1, 1979
Nerve growth factor (NGF) at 10 B.U./ml produced a 5‐fold increase in the concentration of cyclic... more Nerve growth factor (NGF) at 10 B.U./ml produced a 5‐fold increase in the concentration of cyclic AMP 10min after addition to freshly excised embryonic chick dorsal root ganglia (DRG). The presence of NGF at 10 B.U./ml, but not 1 B.U./ml, over a 6 h incubation in nutrient‐free medium prevented this cyclic AMP increase when the DRG were again challenged with NGF (10 B.U./ml) after the 6 h. Incubation of DRG for 6 h at 37°C without NGF did not prevent the cyclic AMP response from occurring when NGF was presented at this time. The basal cyclic AMP concentration, however, decreased by 65–75% during the course of the 6 h incubation, and the continuous presence of NGF (10 B.U./ml) was unable to prevent this. When NGF was added to 6‐h NGF‐deprived DRG, the time for maximum cyclic AMP increase decreased from 15min to 6min with increasing NGF concentrations from 1 to 50 B.U./ml, although the relative magnitude of the cyclic AMP increase was essentially the same (approx 3‐fold) at these NGF concentrations.Attempts were made to correlate the cyclic AMP response in NGF‐deprived DRG with another rapid response resulting from administration of NGF to NGF‐deprived DRG, namely, the reactivation of hexose uptake. The cyclic AMP and permeation responses both occurred on the same time scale (5‐15 min), and both responded to increasing concentrations of NGF with a decreasing time to achieve maximal effect. A temporal relationship between cyclic AMP and membrane permeability was noted, but it could not be ruled out that it might reflect difficulties in methodology and/or inadequacy in current knowledge of the underlying mechanisms.
Developmental Brain Research, Jul 1, 1990
Journal of Neuroscience Research, Apr 1, 1991
Journal of Neuroscience Research, 1982
A serum‐free defined medium has been formulated that supports proliferation and morphologic diffe... more A serum‐free defined medium has been formulated that supports proliferation and morphologic differentiation of U‐251 MGsp human and C6‐2BD rat glioma cells. This defined medium consists of a basal medium supplemented with transferrin, fibroblast growth factor, hydrocortisone, selenium, biotin, and fibronectin (G2 medium). When U‐251 cells were plated in G2 medium on poly‐D‐lysine precoated dishes, their growth rate was 77% and final cell density was 82% of serum‐grown counterparts. The growth rate of C6 cells in G2 medium was 67% compared to cells cultured in serum supplemented medium. Although G2 medium supported the growth of human and rat glioma cells, LA‐N‐1 human neuroblastoma and WI‐38 human fibroblast cells showed no increase in cell number when grown in G2 medium compared to basal medium. A similar formulation (G3 medium), lacking fibroblast growth factor and hydrocortisone, supported the proliferation of RN‐22 rat schwannoma cells. Morphologic differences were observed between cells grown in the presence of serum and in defined media. All three glial cell lines changed from a flattened shape in serum supplemented medium to a more spherical appearance in defined medium. In addition, both U‐251 and C6 cells developed numerous processes, some reaching several cell diameters in length. These defined media will facilitate studies of the growth and differentiation of glial‐derived cells.
Developmental Biology, Jul 1, 1981
Experimental Cell Research, Oct 1, 1980
Experimental Cell Research, 1980
Proceedings of the National Academy of Sciences of the United States of America, 1979
Journal of Neurobiology, 2004
We report that cell survival after neurite transection in a mammalian neuronal model (cultured B1... more We report that cell survival after neurite transection in a mammalian neuronal model (cultured B104 cells) critically depends on somal [Ca2+]i, a novel result that reconciles separate long‐standing observations that somal survival decreases with more‐proximal axonal transections and that increased somal Ca2+ is cytotoxic. Using fluorescence microscopy, we demonstrate that extracellular Ca2+ at the site of plasmalemmal transection is necessary to form a plasmalemmal barrier, and that other divalent ions (Ba2+, Mg2+) do not play a major role. We also show that extracellular Ca2+, rather than injury per se, initiates the formation of a plasmalemmal barrier and that a transient increase in somal [Ca2+]i significantly decreases the percentage of cells that survive neurite transection. Furthermore, we show that the increased somal [Ca2+]i and decreased cell survival following proximal transections are not due to less frequent or slower plasmalemmal sealing or Ca2+ entry through plasmalemmal Na+ and Ca2+ channels. Rather, the increased somal [Ca2+]i and lethality of proximal neurite injuries may be due to the decreased path length/increased diameter for Ca2+ entering the transection site to reach the soma. A ryanodine block of Ca2+ release from internal stores before transection has no effect on cell survival; however, a ryanodine‐ or thapsigargin‐induced buildup of somal [Ca2+]i before transection markedly reduces cell survival, suggesting a minor involvement of Ca2+‐induced release from internal stores. Finally, we show that cell survival following proximal injuries can be enhanced by increasing intracellular Ca2+ buffering capacity with BAPTA to prevent the increase in somal [Ca2+]i. © 2004 Wiley Periodicals, Inc. J Neurobiol 60: 137–153, 2004
Elsevier eBooks, 1983
Publisher Summary This chapter discusses factors known to influence the survival and growth of vi... more Publisher Summary This chapter discusses factors known to influence the survival and growth of virtually all vertebrate cells as well as those of special significance for neural cells. To sustain the survival or division of most vertebrate cells in vitro , a nutrient medium must satisfy the requirements for a balanced salt solution of appropriate osmolarity, buffered hydrogen ion concentration, carbohydrate source, amino acids, vitamins, trace elements, and growth factors. A few cell types can survive in basal medium alone, but the continued survival or division of most cells still requires supplementation with serum or other undefined additives. Under normal conditions in vivo , the brain, unlike most other tissue, derives its energy almost exclusively from the oxidation of glucose. The ability to select for specific cell types from a mixed culture is a clear benefit of using defined medium, as is the suppression of fibroblast overgrowth, even in cultures where the selectivity is not absolute.
Glia, 1988
Enriched preparations of oligodendrocytes, isolated either from adult bovine brain or from 30‐day... more Enriched preparations of oligodendrocytes, isolated either from adult bovine brain or from 30‐day‐old rat brain, eventually yield cultures in MEM‐15% calf serum that contain, in addition to oligodendrocytes, proliferating astrocytes and variable numbers of fibroblast‐like cells. If these cultures are switched to a serum‐free defined medium during the 1st week, mixed cultures containing only oligodendrocytes and astrocytes are obtained. Bovine cultures can be replated and purified by selective adhesion to yield cultures that are < 99% astrocytes; similar procedures were not successful with rat cultures. Cytoskeletal preparations of the purified astrocyte cultures from mature bovine brain contain both vimentin and glial fibrillary acidic protein (GFAP), but vimentin is by far the major intermediate filament protein. Thus, the intermediate filament composition of these astrocytes is similar to that of astrocytes in primary cultures obtained from neonatal rat brain. Immunofluorescent studies of these cultures at 24 hr in vitro show that there are no GFAP+ cells in cultures of either species; the bovine cultures contain < 95% GC+ cells; and the rat cultures contain 90% GC+ cells. After a few days in vitro flat cells appear that are vimentin+/GFAP–/GC–. In serum‐free medium these cells eventually become vimentin+/GFAP+. We propose that the astrocytes that grow in these cultures arise from a population of glial precursor cells, which are present even in adult brain and are isolated together with oligodendroglia, and that they do not derive from contaminating mature astrocytes. Thus, the astrocytes in our cultures may have the same origin as astrocytes grown in culture from dissociated neonatal brain.
Journal of Neuroscience Research, Jul 1, 1988
Life Sciences, Aug 1, 1978
Humana Press eBooks, Nov 14, 2003
ABSTRACT The technique of culturing cells derived from the nervous system has been in use for ove... more ABSTRACT The technique of culturing cells derived from the nervous system has been in use for over 80 years, with the objective of simplifying the experimental system to provide readily manipulatable models of neural function. These preparations are useful for testing hypotheses relevant to cell adhesion, motility, survival, proliferation, longevity, and expression of cell type-specific properties. Many significant modifications of these methods have resulted over the years, as new data have emerged from cell biology and neurobiology studies (see reviews by Bottenstein, 1983a,Bottenstein, 1985,Bottenstein, 1988).
Brain Research, Sep 1, 2006
Developmental Brain Research, Sep 1, 1989
The Society for Neuroscience Abstracts, May 10, 1989
PubMed, 1981
The requirement for a serum supplement to basal culture medium to support the growth and viabilit... more The requirement for a serum supplement to basal culture medium to support the growth and viability of cells in vitro has been a major hindrance to reducing and thereby controlling experimental variables. The complexity and undefined nature of serum as well as the variability of serum lots make it difficult to reproduce data and to interpret results of experiments. This paper presents preliminary results obtained with C6 glioma cell cultures in which serum was replaced by growth factors and a hormone. A brief review of the use of serum-free defined medium for the culture of tumor cells and the potential uses of defined media that are cell-type specific are also presented.
Journal of Neurochemistry, Jun 1, 1979
Nerve growth factor (NGF) at 10 B.U./ml produced a 5‐fold increase in the concentration of cyclic... more Nerve growth factor (NGF) at 10 B.U./ml produced a 5‐fold increase in the concentration of cyclic AMP 10min after addition to freshly excised embryonic chick dorsal root ganglia (DRG). The presence of NGF at 10 B.U./ml, but not 1 B.U./ml, over a 6 h incubation in nutrient‐free medium prevented this cyclic AMP increase when the DRG were again challenged with NGF (10 B.U./ml) after the 6 h. Incubation of DRG for 6 h at 37°C without NGF did not prevent the cyclic AMP response from occurring when NGF was presented at this time. The basal cyclic AMP concentration, however, decreased by 65–75% during the course of the 6 h incubation, and the continuous presence of NGF (10 B.U./ml) was unable to prevent this. When NGF was added to 6‐h NGF‐deprived DRG, the time for maximum cyclic AMP increase decreased from 15min to 6min with increasing NGF concentrations from 1 to 50 B.U./ml, although the relative magnitude of the cyclic AMP increase was essentially the same (approx 3‐fold) at these NGF concentrations.Attempts were made to correlate the cyclic AMP response in NGF‐deprived DRG with another rapid response resulting from administration of NGF to NGF‐deprived DRG, namely, the reactivation of hexose uptake. The cyclic AMP and permeation responses both occurred on the same time scale (5‐15 min), and both responded to increasing concentrations of NGF with a decreasing time to achieve maximal effect. A temporal relationship between cyclic AMP and membrane permeability was noted, but it could not be ruled out that it might reflect difficulties in methodology and/or inadequacy in current knowledge of the underlying mechanisms.
Developmental Brain Research, Jul 1, 1990
Journal of Neuroscience Research, Apr 1, 1991
Journal of Neuroscience Research, 1982
A serum‐free defined medium has been formulated that supports proliferation and morphologic diffe... more A serum‐free defined medium has been formulated that supports proliferation and morphologic differentiation of U‐251 MGsp human and C6‐2BD rat glioma cells. This defined medium consists of a basal medium supplemented with transferrin, fibroblast growth factor, hydrocortisone, selenium, biotin, and fibronectin (G2 medium). When U‐251 cells were plated in G2 medium on poly‐D‐lysine precoated dishes, their growth rate was 77% and final cell density was 82% of serum‐grown counterparts. The growth rate of C6 cells in G2 medium was 67% compared to cells cultured in serum supplemented medium. Although G2 medium supported the growth of human and rat glioma cells, LA‐N‐1 human neuroblastoma and WI‐38 human fibroblast cells showed no increase in cell number when grown in G2 medium compared to basal medium. A similar formulation (G3 medium), lacking fibroblast growth factor and hydrocortisone, supported the proliferation of RN‐22 rat schwannoma cells. Morphologic differences were observed between cells grown in the presence of serum and in defined media. All three glial cell lines changed from a flattened shape in serum supplemented medium to a more spherical appearance in defined medium. In addition, both U‐251 and C6 cells developed numerous processes, some reaching several cell diameters in length. These defined media will facilitate studies of the growth and differentiation of glial‐derived cells.
Developmental Biology, Jul 1, 1981
Experimental Cell Research, Oct 1, 1980
Experimental Cell Research, 1980
Proceedings of the National Academy of Sciences of the United States of America, 1979
Journal of Neurobiology, 2004
We report that cell survival after neurite transection in a mammalian neuronal model (cultured B1... more We report that cell survival after neurite transection in a mammalian neuronal model (cultured B104 cells) critically depends on somal [Ca2+]i, a novel result that reconciles separate long‐standing observations that somal survival decreases with more‐proximal axonal transections and that increased somal Ca2+ is cytotoxic. Using fluorescence microscopy, we demonstrate that extracellular Ca2+ at the site of plasmalemmal transection is necessary to form a plasmalemmal barrier, and that other divalent ions (Ba2+, Mg2+) do not play a major role. We also show that extracellular Ca2+, rather than injury per se, initiates the formation of a plasmalemmal barrier and that a transient increase in somal [Ca2+]i significantly decreases the percentage of cells that survive neurite transection. Furthermore, we show that the increased somal [Ca2+]i and decreased cell survival following proximal transections are not due to less frequent or slower plasmalemmal sealing or Ca2+ entry through plasmalemmal Na+ and Ca2+ channels. Rather, the increased somal [Ca2+]i and lethality of proximal neurite injuries may be due to the decreased path length/increased diameter for Ca2+ entering the transection site to reach the soma. A ryanodine block of Ca2+ release from internal stores before transection has no effect on cell survival; however, a ryanodine‐ or thapsigargin‐induced buildup of somal [Ca2+]i before transection markedly reduces cell survival, suggesting a minor involvement of Ca2+‐induced release from internal stores. Finally, we show that cell survival following proximal injuries can be enhanced by increasing intracellular Ca2+ buffering capacity with BAPTA to prevent the increase in somal [Ca2+]i. © 2004 Wiley Periodicals, Inc. J Neurobiol 60: 137–153, 2004
Elsevier eBooks, 1983
Publisher Summary This chapter discusses factors known to influence the survival and growth of vi... more Publisher Summary This chapter discusses factors known to influence the survival and growth of virtually all vertebrate cells as well as those of special significance for neural cells. To sustain the survival or division of most vertebrate cells in vitro , a nutrient medium must satisfy the requirements for a balanced salt solution of appropriate osmolarity, buffered hydrogen ion concentration, carbohydrate source, amino acids, vitamins, trace elements, and growth factors. A few cell types can survive in basal medium alone, but the continued survival or division of most cells still requires supplementation with serum or other undefined additives. Under normal conditions in vivo , the brain, unlike most other tissue, derives its energy almost exclusively from the oxidation of glucose. The ability to select for specific cell types from a mixed culture is a clear benefit of using defined medium, as is the suppression of fibroblast overgrowth, even in cultures where the selectivity is not absolute.
Glia, 1988
Enriched preparations of oligodendrocytes, isolated either from adult bovine brain or from 30‐day... more Enriched preparations of oligodendrocytes, isolated either from adult bovine brain or from 30‐day‐old rat brain, eventually yield cultures in MEM‐15% calf serum that contain, in addition to oligodendrocytes, proliferating astrocytes and variable numbers of fibroblast‐like cells. If these cultures are switched to a serum‐free defined medium during the 1st week, mixed cultures containing only oligodendrocytes and astrocytes are obtained. Bovine cultures can be replated and purified by selective adhesion to yield cultures that are < 99% astrocytes; similar procedures were not successful with rat cultures. Cytoskeletal preparations of the purified astrocyte cultures from mature bovine brain contain both vimentin and glial fibrillary acidic protein (GFAP), but vimentin is by far the major intermediate filament protein. Thus, the intermediate filament composition of these astrocytes is similar to that of astrocytes in primary cultures obtained from neonatal rat brain. Immunofluorescent studies of these cultures at 24 hr in vitro show that there are no GFAP+ cells in cultures of either species; the bovine cultures contain < 95% GC+ cells; and the rat cultures contain 90% GC+ cells. After a few days in vitro flat cells appear that are vimentin+/GFAP–/GC–. In serum‐free medium these cells eventually become vimentin+/GFAP+. We propose that the astrocytes that grow in these cultures arise from a population of glial precursor cells, which are present even in adult brain and are isolated together with oligodendroglia, and that they do not derive from contaminating mature astrocytes. Thus, the astrocytes in our cultures may have the same origin as astrocytes grown in culture from dissociated neonatal brain.
Journal of Neuroscience Research, Jul 1, 1988
Life Sciences, Aug 1, 1978
Humana Press eBooks, Nov 14, 2003
ABSTRACT The technique of culturing cells derived from the nervous system has been in use for ove... more ABSTRACT The technique of culturing cells derived from the nervous system has been in use for over 80 years, with the objective of simplifying the experimental system to provide readily manipulatable models of neural function. These preparations are useful for testing hypotheses relevant to cell adhesion, motility, survival, proliferation, longevity, and expression of cell type-specific properties. Many significant modifications of these methods have resulted over the years, as new data have emerged from cell biology and neurobiology studies (see reviews by Bottenstein, 1983a,Bottenstein, 1985,Bottenstein, 1988).
Brain Research, Sep 1, 2006
Developmental Brain Research, Sep 1, 1989