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Biomedical subjects

C L Weill

Publications and source records attributed to C L Weill.

At least 19 recordsLinked to original sources
PubMed1993

Regulation of NCAM by growth factors in serum-free myotube cultures.

Regulation of the neural cell adhesion molecule (NCAM) was examined in primary cultures of chick skeletal muscle grown in serum-free defined medium. Relative levels of NCAM (per microgram protein) increased 20-30% in myotubes grown on Matrigel, a reconstituted basement membrane preparation, compared to those grown on collagen; total NCAM levels on Matrigel were increased 40-55% due to the additional increase in total protein. A dose dependent increase in relative NCAM levels in myotubes grown on Matrigel in defined medium was observed with the addition of adsorbed horse serum, while relative NCAM levels in myotubes grown on collagen were unaffected by altering the serum concentration. Thus, extracellular matrix molecules and soluble factors exert trophic effects on myotube NCAM expression. Similar developmental changes in the expression of the different molecular size forms of NCAM occurred in myotubes grown on collagen and Matrigel: levels of 150K and 135K Mr forms decreased during development, while 125K remained prominent in older myotubes. Relative NCAM levels were specifically enhanced 11-26% by several factors: nerve growth factor, thyroxine, insulin-like growth factor II, dibutyryl cyclic AMP, veratridine (a sodium ion channel agonist), and nisoldipine (a calcium ion channel agonist). Total protein and overall myotube development in serum-free cultures were enhanced by fetuin, insulin-like growth factor II, acidic fibroblast growth factor, calcitonin gene-related peptide, dibutyryl cyclic AMP, and veratridine. Thus, changes in extracellular matrix, intracellular calcium, and sodium ions, as well as extracellular trophic factors, such as nerve growth factor, thyroxine, and insulin-like growth factor II, may regulate muscle NCAM expression during embryonic development.

Acetylcholinesterase↗
PubMed1992

Matrigel enhances myotube development in a serum-free defined medium.

Previously reported serum-free defined media for muscle cell culture require supplementation with hormones, purified growth factors or attachment factors. This report describes a culture system that enhances embryonic chick, skeletal muscle cell growth and differentiation in a serum-free defined medium, without added specialized trophic factors. Myoblasts adhered more to and proliferated more rapidly on a reconstituted basement membrane substrate, Matrigel, than on rat-tail collagen. Matrigel contains several basement membrane attachment molecules which apparently obviate the need for added purified attachment factors. Matrigel also appeared to play a trophic role in subsequent development by enabling the serum-free growth of myotubes which suggests that Matrigel mediates the cellular interaction of growth or attachment factors. Collagen, on the other hand, did not support serum-free myotube growth. Supplementation of defined medium with increasing levels of horse serum enhanced total protein in myotubes grown on both substrates; protein was higher in Matrigel cultures for each medium tested. The serum-free defined medium supported complete morphological differentiation of myotubes grown on Matrigel and maintained myotube cultures up to 22 days. Fibroblast proliferation was higher in cultures on collagen in defined medium with high serum levels, but was virtually eliminated in cultures on Matrigel in serum-free defined medium. The culture system described supports the differentiation of embryonic muscle cells in a simple, serum-free defined medium, thus providing an in vitro model of developing myotubes which should be particularly useful for studies of regulation mediated by extracellular factors.

Animals↗
PubMed1991

Somatostatin (SRIF) prevents natural motoneuron cell death in embryonic chick spinal cord.

Natural motoneuron cell death is a developmental process that effects the loss of about 50% of the cells in the lateral motor column of the spinal cord. The present study demonstrates that the systemic treatment of developing chick embryos with somatostatin (somatotropin release-inhibiting factor, SRIF) during the period of natural motoneuron cell death resulted in a 13-22% increase in the number of surviving motoneurons, suggesting that SRIF may be an endogenous contributor to motoneuron survival during normal development. It is hypothesized that SRIF may act, through its ability to reduce intraneuronal calcium, as an endogenous antagonist of neuronal death both during development and in the course of adult neurodegenerative diseases.

Animals↗
PubMed1990

Augmentation of GABAA receptor function by chronic exposure to GABA-neutral and GABA-negative benzodiazepine ligands in cultured cortical neurons.

Chronic benzodiazepine agonist administration may lead to decreases in gamma-aminobutyric acidA (GABAA) receptor binding and function, but little information is available concerning chronic GABA-neutral or GABA-negative benzodiazepine exposure. We evaluated effects of chronic exposure to flumazenil (Ro15-1788) and FG 7142 (N-methyl-beta-carboline-3-carboxamide) on GABA-dependent chloride uptake in chick cerebral cortical neurons in primary culture. Acute flumazenil treatment (1 microM) had no effect on chloride uptake, but uptake was increased after 2 days of exposure. Similar increases were observed after 4 and 10 days. Flumazenil, 0.1 microM, had no effect after 10 days, and a 10 microM concentration had a similar effect as the 1 microM concentration. Acute FG 7142 (1 microM) decreased chloride uptake, but uptake was increased markedly after 2, 4, and 10 days of treatment. No effect was observed after treatment for 10 days with 0.1 microM, but a 10 microM concentration showed similar enhancement to the 1 microM concentration. Concurrent treatment with 0.3 microM flumazenil which did not affect chloride uptake and 1 microM FG 7142 for 10 days substantially attenuated the effects of FG 7142, suggesting that FG 7142 effects are mediated at the benzodiazepine site. Benzodiazepine receptor binding was increased in cultures treated for 10 days with 1 microM flumazenil or FG 7142, with an increase in receptor number in both cases but no change in apparent affinity. Neither flumazenil nor FG 7142 (1 microM for 10 days) altered GABA-independent chloride uptake, total cellular protein, protein synthesis or degradation, or neuronal survival. These results indicate that both chronic GABA-neutral and GABA-negative benzodiazepine exposures in cultured cortical neurons lead to increases in GABA-dependent chloride uptake and benzodiazepine binding. Effects of GABA-negative benzodiazepine exposure appear to be greater than those observed with GABA-neutral benzodiazepine exposure.

Animals↗
PubMed1990

The prevention of natural motoneuron cell death by dibutyryl-cyclic GMP in the spinal cord of White Leghorn chick embryos.

The effect of dibutyryl-cyclic guanosine monophosphate (db-cGMP) treatment of White Leghorn chick embryos on the extent of natural motoneuron cell death in the lumbar spinal cord was examined. A dose of 1.59 mumol of db-cGMP effected an increase in motoneuron survival of 26.7%. No effect on motoneuron survival was observed with equimolar doses of 8-bromo-cGMP, db-cAMP, GMP, guanosine, or butyric acid.

Animals↗
PubMed1990

Rapid activation by erythropoietin of protein kinase C in nuclei of erythroid progenitor cells.

The glycoprotein hormone erythropoietin (Ep) regulates the proliferation and differentiation of erythroid progenitor cells by a signal transduction system which is not well understood. It has recently been reported that prolactin, a mitogen and trophic hormone for liver, will activate a nuclear protein kinase C in hepatocytes. As similarities exist in the actions of Ep and prolactin in their target cells, we tested the hypothesis that Ep could activate protein kinase C in nuclei isolated from erythroid progenitor cells. In a pure population of such nuclei, Ep induced a rapid, time- and dose-dependent increase in phosphorylation of endogenous nuclear substrate which could be blocked by inhibitors of protein kinase C or by antibody to Ep. Other known activators of protein kinase C were also effective in this system. These findings show that Ep may exert its effects by a novel signalling pathway, the activation of a nuclear protein kinase C.

Animals↗
PubMed1990

The HIV protein, GP120, activates nuclear protein kinase C in nuclei from lymphocytes and brain.

Nuclear pool(s) of protein kinase C (PKC) may be a common target for hormones and growth factors which affect the trophic state of cells. The data presented demonstrate a time and dose-dependent activation of nuclear PKC by the HIV coat protein, gp120, in isolated nuclei from rat spleen and hippocampus. This gp120-stimulated PKC response was blocked by specific PKC inhibitors, a monoclonal antibody to PKC, and a monoclonal antibody directed against the murine T4 analog, L3T4. It is suggested that the gp120 interaction with the nuclear trophic factor-PKC system may impair normal gene expression, and thus result in the clinical symptoms associated in AIDS infection.

Alkaloids↗
PubMed1989

Persistent alterations in GABAA receptor binding and function after prenatal lorazepam administration in the chick.

Behavioral abnormalities have been reported in young and adult animals exposed to benzodiazepines prenatally. The presence of neurochemical alterations in the GABAergic system after prenatal benzodiazepine exposure was assessed in a chick model which avoids prenatal and postnatal maternal effects. The GABAA receptor complex, the presumed site of benzodiazepine action, was altered in adult chickens previously exposed to lorazepam for 10 days in ovo. Binding was decreased at the putative chloride channel site labeled by [35S]TBPS, coupling was decreased between this site and the GABA binding site, and function of the GABAA receptor in chloride uptake was diminished in animals exposed to prenatal lorazepam. Persistent neurochemical alterations in the GABAergic system accompany prenatal benzodiazepine exposure, and may influence subsequent behavior and development.

Animals↗
PubMed1989

Chronic clonazepam administration decreases gamma-aminobutyric acidA receptor function in cultured cortical neurons.

Chronic benzodiazepine administration has been reported to decrease gamma-aminobutyric acidA (GABAA) receptor function in animals and may alter benzodiazepine binding in neuronal cultures. To assess GABAA receptor function in neuronal cultures exposed to chronic clonazepam, we measured muscimol-stimulated chloride uptake in chick cerebral cortical cultures treated acutely and for 2, 4, and 10 days. Acute clonazepam administration (1 microM) led to an increase in GABA-related chloride uptake at lower doses of muscimol. After chronic clonazepam (1 microM), maximal uptake was markedly decreased at day 10, but maximal uptake was unchanged after 2- and 4-day treatments. Benzodiazepine receptor binding was decreased by approximately 60% after 10 days due to a decrease in receptor number. Decreases in chloride uptake were also observed after 10 days of treatment with 0.1 and 10 microM clonazepam. Concomitant treatment with 0.1 microM Ro15-1788 abrogated the effect of 0.1 microM clonazepam on chloride uptake. Chronic clonazepam treatment (1 microM) did not alter total cellular protein, cellular protein synthesis or degradation or percentage of neuronal cells, as determined morphologically and by [3H]ouabain binding.

Animals↗
PubMed1989

Prenatal lorazepam administration is associated with GABAA receptor alterations in late embryonic and mature chicks.

Prenatal benzodiazepine administration has been associated with alterations in behavior in young and mature animals. Prior studies of neurochemical changes in animals treated in utero have produced conflicting results. We used a chick embryo system to assess the effects of chronic lorazepam administration on binding and function at the GABAA receptor in late embryos and in mature animals. Administration of lorazepam for 10 days of embryonic development (E8-E18) led to decreases in binding of the chloride channel ligand TBPS and in muscimol-stimulated chloride uptake in late embryos (E18). Similar alterations were observed in mature animals after the same regimen of prenatal lorazepam. Persistent alterations in GABAA receptor binding and function after prenatal lorazepam may be due to effects of neuronal differentiation or on receptor regulation. These neurochemical effects may underlie the behavioral abnormalities associated with prenatal benzodiazepine administration.

Animals↗
PubMed1988

Lorazepam administration during embryonic development alters GABAA receptor binding and function.

Prenatal exposure to diazepam has been reported to lead to behavioral alterations in young and mature animals, but the neurochemical basis of this effect is uncertain. To examine effects of embryonic benzodiazepine exposure on the gamma-aminobutyric acidergic (GABA)ergic) system, we assessed binding and function at the GABAA receptor complex in chick embryos treated with lorazepam, 2 mg/kg, for 2-10 days. There was no change in benzodiazepine receptor binding in cortex after lorazepam administration, but a decrease in the number of chloride channel sites was observed. Overall function at the GABAA receptor complex as assessed by chloride uptake into cortical synaptoneurosomes was decreased after 10 days of lorazepam, and possibly increased after 4 days of lorazepam. Similar results were obtained when embryos were treated for 2- and 4-day periods beginning on day 8. These results indicate that chronic lorazepam administration to embryos alters binding and function in the GABA system measured soon before hatching. These alterations, if persistent, may contribute to the behavioral changes seen in animals exposed prenatally.

Animals↗
PubMed1986

Expression of myelin proteolipid and basic protein mRNAs in cultured cells.

Studies were undertaken to investigate the regulation of myelin-specific mRNA expression in cultured cells. Three experimental systems were investigated: primary oligodendrocytes grown as enriched cell populations, primary oligodendrocytes grown in the presence of chick spinal cord neurons, and C6 cells. cDNA probes specific for the myelin proteolipid mRNA and the myelin basic protein mRNA were used to quantitate proteolipid and myelin basic protein mRNA levels in cells under different experimental conditions. C6 cells expressed less than 0.2% of the proteolipid mRNA that was expressed in primary oligodendrocytes. Primary oligodendrocytes expressed the myelin-specific mRNAs for at least 104 days in culture, and the level of these mRNAs in cultures was elevated fourfold by coculturing rat oligodendrocytes with chick spinal cord neurons.

Animals↗
PubMed1986

Characterization of androgen receptors in embryonic chick spinal cord.

While androgen binding sites have been localized to motoneurons of the lateral motor column (LMC) of 10-18-day chick embryo spinal cord, they have yet to be characterized biochemically. Studies were undertaken to characterize the binding of the androgen [3H]5a-dihydrotestosterone ([3H]DHT) to spinal cord cytosols. Saturable, high-affinity binding of [3H]DHT to cytosols prepared from both 6 and 10-day spinal cords was observed. The binding component was a macromolecular species as it displayed a sedimentation coefficient of 8S upon centrifugation in sucrose gradients; proteinaceous, as binding was eliminated by heating cytosols; and displayed steroid-specific binding, as other non-androgen steroid agonists did not significantly inhibit [3H]DHT binding. The number of binding sites increased 10-fold from embryonic day 6 to day 10. The availability of testosterone and presence of androgen receptors in spinal cord motoneurons as spontaneous limb motility begins suggests a possible role for androgen-receptor-dependent gene expression in the process of target-dependent differentiation of LMC motoneurons.

Animals↗
PubMed1986

Characterization of glucocorticoid receptors in whole and cellular subfractions of embryonic chick spinal cord.

Steroid-specific binding sites for tritiated corticosterone have been localized, via autoradiography, to motoneurons in the lateral motor columns of the adult rat spinal cord. Binding sites in adult rat spinal cord have been characterized biochemically and shown to possess the properties of a putative glucocorticoid receptor. The presence of receptors for glucocorticoids in embryonic chick spinal cord was determined and their characterization undertaken as a prelude to the study of the functions under regulation by glucocorticoids during development. Assay conditions were defined and binding of [3H]dexamethasone [( 3H]Dex) to cytosols of 6- and 10-day embryonic spinal cord and cellular subfractions of 6-day spinal cord determined. Saturable, high-affinity binding of [3H]Dex to cytosols prepared from both whole 6- and 10-day spinal cords and cells of all 3 cellular subfractions of 6-day spinal cords was observed. The binding component in 10-day cytosols was (1) proteinaceous, as binding was eliminated by heating cytosols, and (2) a macromolecular species, as it displayed a sedimentation coefficient of 8S upon centrifugation in sucrose gradients. The putative receptor displayed binding specific for glucocorticoids in a competition assay, with the exception that some inhibition of binding by the androgen ligand, methyltrienolone (R1881) was observed. The binding affinity decreased as the values for KD increased from 3.4 +/- 0.9 nM on day 6 to 15.7 +/- 1.8 nM on day 10, while the values for Bmax increased from 270 to 855 fmol/mg protein over the same period.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1986

Cyclic nucleotide content of ciliary and dorsal root ganglia during embryonic development in the chick.

The concentration of cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP) was measured in developing chick ciliary (CG) and dorsal root ganglia (DRG) as a function of embryonic age from day 8 through day 18 using radioimmunoassays. The concentration of cAMP and cGMP increased in both ganglia from day 8 through day 14. cAMP levels were nearly two-fold higher in DRG than in CG. Normalization of the data for ciliary ganglia to the number of cells per ganglion and calculation of their molar concentrations indicates a 48% increase in cGMP and a 3.2-fold increase in cAMP during the developmental process of natural neuronal cell death.

Animals↗
PubMed1986

Changes in glucose 6-phosphate dehydrogenase activity in developing embryonic chick skeletal muscle and spinal cord.

Glucose 6-phosphate dehydrogenase (G6PDH) activity was examined in the developing embryonic chick in brachial and lumbar spinal cord and pectoral and leg muscle. Enzyme activity was generally highest at the earliest stage examined, embryonic day 5. The developmental profiles for G6PDH activity in the two muscles were similar: a sharp initial decrease occurred between days 5 and 9, with relatively low levels present by day 18; peaks of G6PDH activity at days 12 and 16 were more prominent in leg muscle. Similar levels of G6PDH were also detected in spinal cord with the developmental profile in the brachial spinal cord resembling that seen in muscle. In lumbar spinal cord, initial G6PDH activity was lower than in brachial spinal cord; the developmental profile, however, resembled that seen in the brachial spinal cord, with an initial drop in enzyme activity seen between days 5 and 7. Neural regulation of G6PDH activity in mature muscle is believed to repress enzyme synthesis. The drop in G6PDH activity observed in embryonic spinal cord and muscle between days 5 and 9 coincides with the initiation of functional neuromuscular contacts. Hence, the normal regulation of G6PDH during embryonic development may involve the repression of G6PDH in spinal cord neurons and muscle, possibly effected by their interaction.

Animals↗
PubMed1985

Appearance of myelin proteins during development in the chick central nervous system.

The time course of the appearance of myelin-specific markers was studied in the developing chick central nervous system (CNS). Chick CNS tissue was studied for the presence of both proteolipid and myelin basic protein by electroblotting and for 2',3'-cyclic nucleotide 3'-phosphohydrolase (CNPase) by enzyme assay. Four regions of chick spinal cord (cervical, brachial, thoracic and lumbar), brain stem, cerebellum, optic nerve and cortex were studied. In general, myelin basic protein appeared approximately 1 day earlier than proteolipid. In spinal cord and brain stem, myelin basic protein appeared at 13 days incubation. In cerebellum and optic nerve, it appeared at 17 days incubation and in cortex at hatching. CNPase activity increased in most CNS regions between 16 days incubation and hatching. These results suggest that myelination occurs earlier in the chick than in the rat and that it occurs over a shorter time period.

2',3'-Cyclic-Nucleotide Phosphodiesterases↗