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G D Maxwell

Publications and source records attributed to G D Maxwell.

34 records · Page 2Linked to original sources
PubMed1988

Adrenergic development of neural crest cells grown in a defined medium under a reconstituted basement-membrane-like matrix.

The embryonic neural crest of vertebrates is the source of a wide variety of adult cell types. We have demonstrated previously that the presence of a reconstituted basement-membrane-like (RBM) gel overlay can dramatically stimulate the development of adrenergic cells in neural crest cultures grown in a complex medium containing horse serum and chick embryo extract. In the present experiments we have analyzed the differentiation of neural crest cells grown in a defined medium with an RBM gel overlay. We found that the presence of the RBM gel promoted the development of catecholamine-containing (CA+) cells in neural crest cultures grown in defined medium compared to cultures grown in this same medium in the absence of the gel. The number of CA+ cells which developed in cultures grown in defined medium in the presence of the RBM gel overlay was similar to that seen in cultures grown in complex medium in the absence of the RBM gel.

Adrenergic Fibers↗
PubMed1988

Analysis of the development of cellular subsets present in the neural crest using cell sorting and cell culture.

We have tested the hypothesis that developmentally significant cellular subsets are present in the early stages of neural crest ontogenesis. Cultured quail trunk neural crest cells probed with the monoclonal antibodies HNK-1 and R24 exhibited heterogeneous staining patterns. Fluorescence-activated cell sorting was used to isolate the HNK-1+ and HNK-1- cell populations at 2 days in vitro. When these cell populations were cultured, the HNK-1+ sorted cells differentiated into melanocytes, unpigmented cells, and numerous catecholamine-positive (CA+) cells. In contrast, the HNK-1- sorted cells gave rise to melanocytes and unpigmented cells, but few, if any, CA+ cells. When neural crest cells at 2 days in vitro were labeled with R24 and sorted, both the R24+ the R24- sorted cell populations produced numerous CA+ cell, melanocytes, and unpigmented cells. These results provide evidence for the existence of developmental preferences in some subsets of neural crest cells early in embryogenesis.

Animals↗
PubMed1987

Exogenous basement-membrane-like matrix stimulates adrenergic development in avian neural crest cultures.

The development of quail trunk neural crest cultures was dramatically altered when the cultures were overlaid with a gel of reconstituted basement membrane (RBM) components derived from the Engelbreth-Holm-Swarm sarcoma. In the presence of the RBM gel overlay, the number of catecholamine-positive (CA+) cells that developed was increased 50-fold, while the final number of melanocytes and total cells was only half that seen in the control cultures. The presence of the RBM gel overlay did not alter the time of onset of differentiation of the CA+ cells or melanocytes. The stimulation of CA+ cell number was not observed with type IV collagen substrates, laminin substrates or type I collagen gel overlays with or without added laminin. The stimulation of CA+ cell development was dependent on initial plating density. The number of CA+ cells that developed in the presence of the RBM gel was proportional to the initial plating density at 80-320 cells mm-2, whereas no CA+ cells were observed below 20 cells mm-2 and only a few CA+ cells were detected at 40 cells mm-2. There was, however, extensive cell division and differentiation of melanocytes and unpigmented cells at the lower initial plating densities. When the RBM gel was used as a substrate, rather than as an overlay, a striking rearrangement of cells into interconnected strands was observed. After several days in culture, melanocytes, CA+ cells and unpigmented cells were present in these strands. These results indicate that molecules associated with a reconstituted basement-membrane-like matrix are a potent stimulatory influence on adrenergic development and also act to inhibit the production of other cell types in neural crest cultures.

Animals↗
PubMed1987

Phenotypic properties of catecholamine-positive cells that differentiate in avian neural crest cultures.

We have investigated several phenotypic features of the catecholamine-positive (CA+) cell population that develops in quail neural crest cultures. The number, spatial distribution, and morphology of CA+ and tyrosine hydroxylase-positive (TH+) cells are similar at all ages examined, suggesting that these 2 cell classes are identical. Neither CA+ nor TH+ cell bodies or processes were stained using antisera that recognize the 70 or 160 kDa subunits of chicken neurofilament protein. Other cell bodies and fibers in the cultures (which were CA- and TH-) were stained with these neurofilament antisera. The uptake and storage of 3H-norepinephrine by neural crest cultures containing CA+ cells were inhibited in the presence of desmethylimipramine and by incubation at 0 degrees C, but were unaffected by normetanephrine. Overnight treatment with reserpine eliminated histochemically detectable CA fluorescence from the cultures. Chronic reserpine treatment from day 2 to 7 in vitro prevented the appearance of CA+ cells, while normal numbers of TH+ and somatostatin-like immunoreactive (SLI) cells developed. The number and light-microscopic morphology of the CA+ cells that developed in these cultures were not dramatically altered by either exogenous NGF or 6-hydroxydopamine. Using the method of Grillo et al. (1974), we have demonstrated that the CA+ cells observed in the light microscope corresponded to cells containing abundant cytoplasmic granular vesicles (GV) characteristic of catecholamine storage granules observed in other systems. The GV diameters were quite similar in cells examined after 5, 7, 14, and 21 d in vitro. Most GV were 50-200 nm in diameter and were distributed in a unimodal manner, with the observed modal values in the range of 85-115 nm at the ages examined. The number of GV/micron2 of cytoplasmic area remained quite constant at all ages examined. These data, taken together with other available information, suggest that the CA+ cells that differentiate in our neural crest cultures resemble, in many respects, the small, intensely fluorescent cells found in autonomic ganglia and extra-adrenal chromaffin tissue of many species. At present, we do not know if the CA+ cells that differentiate in our neural crest cultures are a stable endpoint of development or whether they are a developmental intermediate in adrenergic differentiation that is normally observed only transiently during the development of avian sympathetic ganglia in vivo, but that can persist under our tissue culture conditions.

Animals↗
PubMed1986

Evidence for a novel enzymatic mechanism of neural crest cell migration on extracellular glycoconjugate matrices.

Migrating embryonic cells have high levels of cell surface galactosyltransferase (GalTase) activity. It has been proposed that GalTase participates during migration by recognizing and binding to terminal N-acetylglucosamine (GlcNAc) residues on glycoconjugates within the extracellular matrix (Shur, B. D., 1982, Dev. Biol. 91:149-162). We tested this hypothesis using migrating neural crest cells as an in vitro model system. Cell surface GalTase activity was perturbed using three independent sets of reagents, and the effects on cell migration were analyzed by time-lapse microphotography. The GalTase modifier protein, alpha-lactalbumin (alpha-LA), was used to inhibit surface GalTase binding to terminal GlcNAc residues in the underlying substrate. alpha-LA inhibited neural crest cell migration on basal lamina-like matrices in a dose-dependent manner, while under identical conditions, alpha-LA had no effect on cell migration on fibronectin. Control proteins, such as lysozyme (structurally homologous to alpha-LA) and bovine serum albumin, did not effect migration on either matrix. Second, the addition of competitive GalTase substrates significantly inhibited neural crest cell migration on basal lamina-like matrices, but as above, had no effect on migration on fibronectin. Comparable concentrations of inappropriate sugars also had no effect on cell migration. Third, addition of the GalTase catalytic substrate, UDPgalactose, produced a dose-dependent increase in the rate of cell migration. Under identical conditions, the inappropriate sugar nucleotide, UDPglucose, had no effect. Quantitative enzyme assays confirmed the presence of GalTase substrates in basal lamina matrices, their absence in fibronectin matrices, and the ability of alpha-LA to inhibit GalTase activity towards basal lamina substrates. Laminin was found to be a principle GalTase substrate in the basal lamina, and when tested in vitro, alpha-LA inhibited cell migration on laminin. Together, these experiments show that neural crest cells have at least two distinct mechanisms for interacting with the substrate during migration, one that is fibronectin-dependent and one that uses GalTase recognition of basal lamina glycoconjugates.

Animals↗
PubMed1985

Development of cells containing catecholamines and somatostatin-like immunoreactivity in neural crest cultures: relationship of DNA synthesis to phenotypic expression.

The goal of our work is to understand the mechanisms which regulate the differentiation of embryonic neural crest cells into a number of adult cell types, including several classes of neurons. As one aspect of this analysis, the relationship between DNA synthesis and the ontogeny of cells with catecholamines and somatostatin-like immunoreactivity (SLI) in neural crest cell cultures has been investigated. Most of the precursors of the catecholamine- and SLI-positive cells carry out DNA synthesis. As these cells differentiate, their ability to carry out DNA synthesis declines. However, a small percentage of cells continue to synthesize DNA after they become catecholamine or SLI positive. There is no apparent difference between the temporal pattern of DNA synthesis in the precursors of catecholamine-positive cells with SLI and those without SLI. Thus, the time of withdrawal from the cell cycle does not distinguish the lineage of cells that are catecholamine and SLI positive from those that are catecholamine positive and SLI negative.

Animals↗
PubMed1985

Specific cellular expression of monoamine oxidase B during early stages of quail embryogenesis.

Monoamine oxidase-B (MAO-B) is one of two distinct molecular forms of MAO that in part regulate the cellular levels of biogenic amines. In order to determine whether discrete cell populations known to express aminergic properties in the vertebrate embryo also express MAO-B, the distribution of MAO-B-immunoreactive cells was examined in early developing quail embryos. Two major patterns of staining emerge. First, tissues known to express several aminergic characteristics are initially MAO-B positive at early stages of development and continue to express immunoreactivity through Zacchei stage 20, the oldest stage examined. These include cells of the sympathetic and some cranial and trunk sensory ganglia (beginning at stage 13), the pancreas (stage 14), and the brain stem raphe (stage 14). Second, other structures that contain or accumulate biogenic amines are transiently MAO-B immunopositive during early stages of development. These tissues include extraembryonic yolk sac and presumptive gut endoderm (with most intense staining between stages 8 and 13), the ventral trunk neural tube (stages 14 and 16), and the notochord (stages 8-10). MAO-B immunoreactivity disappears from these regions at different stages, and none are MAO-B positive by stage 19-20. Other structures without known aminergic properties during early development also stain; these include liver (stage 20), mesenchymal cells that surround the Wolffian duct and lung buds (stages 14 and 18), and endothelial cells surrounding the dorsal aorta (stages 14 and 20). In general, however, MAO-B appears to be distributed in embryonic tissues that can use this enzyme to regulate biogenic amine levels either transiently or during initial phenotypic expression of aminergic traits.

Animals↗
PubMed1984

Somatostatin-like immunoreactivity is expressed in neural crest cultures.

Embryonic neural crest cells give rise to many adult cell types, including some neurons that contain the neuroactive peptide somatostatin. The development of somatostatin-like immunoreactive cells has been investigated in quail neural crest cultures. Somatostatin-like immunoreactive cells were first detected in such cultures after 5 days in vitro by immunocytochemistry. The number of immunoreactive cells and the intensity of the immunoreactivity increased dramatically between 5 and 6 days in vitro. The immunoreactivity was present in a small subset of cells in the culture. It was seen in the perikaryon and cellular processes, but was absent from the nucleus. Immunoreactive cells with and without processes were observed. Somatostatin immunoreactivity was absent if the anti-somatostatin serum was preincubated in the presence of somatostatin. A sequential procedure for detecting catecholamine-containing cells histochemically followed by immunocytochemistry for somatostatin-like immunoreactivity showed that about 20% of the cells which contained catecholamines also contained somatostatin-like immunoreactivity. Greater than 90% of the somatostatin-containing cells in this analysis also contained catecholamines. Somatostatin-like immunoreactivity was also detected and quantitated by radioimmunoassay. The somatostatin-like immunoreactivity developed in vitro in medium that contained horse serum and chick embryo extract, but in the absence of other embryonic cell types such as notochord and somitic mesenchyme. These in vitro results together with prior in vivo data indicate that expression of somatostatin-like immunoreactivity is an early event in the development of some neural crest cells which also express adrenergic properties.

Animals↗
PubMed1984

Development of somatostatin-like immunoreactivity in embryonic sympathetic ganglia.

Neural crest cells are the embryonic progenitors of several adult cell types, including some neurons that contain the neuroactive peptide somatostatin. To begin to understand the control of peptide expression during neuronal ontogeny, we have investigated the development of somatostatin-like immunoreactivity (SLI) in embryonic quail paravertebral sympathetic ganglia in vivo. SLI was identified by immunohistochemistry in paraformaldehyde-fixed cryostat sections from the trunk region of quail embryos. SLI was first observed in the cells of the primary sympathetic trunks at stage 18 (Zacchei, A.M. (1961) Arch. Ital. Anat. Embriol. 66:36-62), which corresponds to embryonic day 4 (E4). The primary sympathetic trunks are the sites of the initial aggregation of neural crest cells to form the sympathetic ganglia. The SLI in these cells was located in the cytoplasm and was absent from the nucleus. SLI persisted in subsequent developmental stages as formation of the definitive sympathetic ganglia occurred. At stage 23 (E7), when the lumbosacral paravertebral sympathetic ganglia have reached their definitive location, some cells in the ganglia contained SLI, and they were often located adjacent to one another. During the later stages of embryogenesis, sympathetic ganglia can be dissected from the embryo and the SLI content determined by radioimmunoassay. The amount of SLI is several-fold higher in ganglia removed at stage 23-24 (E7-8) than in ganglia removed at stage 26-27 (E9-10) or stage 31-32 (E13-14). This decline in SLI content reflects an absolute decrease per ganglion and is not solely due to the growth of tissue devoid of SLI.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1983

Expression of the capacity to release [3H]norepinephrine by neural crest cultures.

Cultures of trunk neural crest cells from quail embryos were tested for their ability to release [3H]norepinephrine [( 3H]NE) in response to depolarization. After 7 days in vitro, exposure of the cultures to either the alkaloid veratridine or 40 mM K+ results in the evoked release of [3H]NE. The release evoked by veratridine is blocked in the presence of tetrodotoxin. The release evoked by increased K+ is blocked by the calcium antagonist cobalt. Release in response to the nicotinic cholinergic agonist 1,1-dimethyl-4-phenylpiperazine was also observed. The amount of evoked release is highly correlated with the number of histochemically demonstrable catecholamine-containing cells in a given culture. Autoradiography reveals that the radioactivity taken up by these cultures is located in a subpopulation of cells whose morphology resembles that of the histochemically detectable catecholamine-containing cell population. Whereas capacity for the release of [3H] NE is readily detectable after 7 days in vitro, it is detectable only with difficulty after 4 days in vitro. There is a greater than 6-fold increase in uptake capacity over the period of 4 to 7 days in vitro. These results demonstrate that neural crest cultures grown without their normal synaptic inputs or targets can exhibit the capacity for stimulus secretion coupling characteristic of synaptic neurotransmitter release.

Animals↗
PubMed1982

Development of neuron-specific enolase immunoreactivity in avian nervous tissue in vivo and in vitro.

Neuron-specific enolase (NSE) is a glycolytic isoenzyme that is primarily located in neurons and neuroendocrine cells. The development of NSE immunoreactivity in th avian nervous system at the level of the hind limb has been examined using immunocytochemical methods. NSE immunoreactivity is first detected in ventral horn motor neurons and dorsal root ganglion neurons at embryonic day 9-10. This is at least 2-3 days after some neurons in both these populations are capable of electrical activity. The glycogen body, a non-neuronal structure, also exhibits NSE (+) staining, but the onset of this immunoreactivity is earlier, at 8 days of embryonic development. NSE immunoreactivity was absent from the cell bodies of paravertebral sympathetic ganglia throughout development, but was present in cellular processes and terminals in the adult ganglia. NSE immunoreactivity also develops in tissue cultures containing cells of neural tube and neural crest origin.

Animals↗
PubMed1982

Synthesis an accumulation of putative neurotransmitters by cultured neural crest cells.

The events mediating the differentiation of embryonic neural crest cells into several types of neurons are incompletely understood. In order to probe one aspect of this differentiation, we have examined the capacity of cultured quail trunk neural crest cells to synthesize, from radioactive precursors, and store several putative neurotransmitter compounds. These neural crest cultures develop the capacity to synthesize and accumulate acetylcholine and the catecholamines norepinephrine and dopamine. In contrast, detectable but relatively little synthesis and accumulation of 5-hydroxytryptamine gamma-aminobutyric acid, or octopamine from the appropriate radiolabeled precursors were observed. The capacity for synthesis and accumulation of radiolabeled acetylcholine and catecholamines is very low or absent at 2 days in vitro. Between 3 and 7 days in vitro, there is a marked rise in both catecholamine and acetylcholine accumulation in the cultures. These findings suggest that, under the particular conditions used in these experiments, the development of neurotransmitter biosynthesis in trunk neural crest cells ijs restricted and resembles, at least partially, the pattern observed in vivo. The development of this capacity to synthesize and store radiolabeled acetylcholine and catecholamines from the appropriate radioactive precursors coincides closely with the development of the activities of the synthetic enzymes choline acetyltransferase and dopamine beta-hydroxylase reported by others.

Acetylcholine↗
PubMed1981

Anatomical and neurochemical consequences of deafferentation in the development of the visual system of the moth Manduca sexta.

The primordium of the compound eye of the moth Manduca sexta was removed by surgery within 12 hours after the molt to the pupa. When the operated animals were examined as pharate adults just prior to emergence from the pupal cuticle, no eye tissue was present on the operated side. Histological examination of the brain at successive developmental stages showed that the volume of laminar neuropil on the operated side increased very little after deafferentation while the volume of the control-side laminar neuropil increased more than 20-fold. The impairment of development of the lamina was accompanied by a reduction in the synthesis and storage by the optic lobe of two neurotransmitter candidates, acetylcholine and 5-hydroxytryptamine. The endogenous content of 5-hydroxytryptamine was also reduced in these preparations. Excision of a section of the stemmatal nerve, which connects the primordium of the compound eye to the brain, also arrested the development of the lamina, but in this case the compound eye itself developed apparently normally without making contact with the brain. This finding supports the hypothesis that the centripetal growth of photoreceptor-cell axons normally is contact-guided. Results of neurochemical experiments on retinal tissue argue against the idea that acetylcholine, 5-hydroxytryptamine, or gamma-aminobutyric acid is a likely photoreceptor-cell neurotransmitter. From these and our previous studies of the metamorphosing brain in Manduca we conclude that different classes of neurons in the same central nervous system can exhibit widely different degrees of dependency on their normal innervation for their survival and differentiation.

Acetylcholine↗
PubMed1976

Substrate dependence of cell migration from explanted neural tubes in vitro.

Embryonic chick neural tubes containing neural crest cells were cultured in vitro on tissue culture plastic and collagen. Two parameters, the time of onset of cell migration from the neural tube and the rate of movement of the cell front away from the neural tube explant, were determined. On collagen, cell migration consistently began after four to six h in vitro, about five h prior to the onset of cell migration on tissue culture plastic. The identity of the migrating cells as neural crest cells is established by their eventual differentiation into melanocytes. Ablation experiments reveal that collagen also causes the early onset of migration of cells not of neural crest origin. These results provide in vitro support for the idea that extracellular materials may alter cell migratory behaviour in morphogenesis.

Animals↗