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Biomedical subjects

J R Sheppard

Publications and source records attributed to J R Sheppard.

At least 37 records · Page 2Linked to original sources
PubMed1981

Effects of caffeine withdrawal on isoproterenol-stimulated cyclic adenosine monophosphate.

The effect of short-term caffeine withdrawal on lymphocyte beta-adrenoceptor sensitivity was studied in seven men who had been drinking coffee daily for at least 3 mo before the study. The ratio of cyclic adenosine monophosphate production after in vitro incubation with isoproterenol and a blank was used to assess receptor sensitivity. There was a reduction in sensitivity to isoproterenol but not to prostaglandin E1 3 days after caffeine withdrawal. The mechanism of this effect is not clear, but may represent a biochemical correlate of caffeine-withdrawal syndrome.

Adrenal Medulla↗
PubMed1981

Down syndrome fibroblasts are hyperresponsive to beta-adrenergic stimulation.

The hormonal response to human skin fibroblasts after exposure to beta-adrenergic agonists, prostaglandin E1 (PGE1), and cholera toxin was monitored by intracellular cyclic AMP accumulation. Down syndrome (DS; trisomy 21) cells had an approximately 10-fold greater response to beta-adrenergic agonists than did either normal diploid skin fibroblasts or other aneusomic fibroblast strains (trisomy 13, 18, and 22). The altered response in DS fibroblasts was specific for beta-adrenergic agonists, because treatment of DS or control cells with PGE1 or cholera toxin resulted in the same degree of cyclic AMP accumulation. Experiments with 3-isobutyl-1-methylxanthine, a cyclic nucleotide phosphodiesterase inhibitor, indicated that the increased response of DS fibroblasts was not primarily a function of altered cyclic AMP degradation. Monosomy 21 cells responded less than normal diploid fibroblasts to stimulation by the beta-adrenergic agonist isoproterenol. These findings suggest that genetic information on chromosome 21 participates in regulating the beta-adrenergic response of human fibroblasts.

Adenylyl Cyclases↗
PubMed1981

Cell cycle changes in the adenylate cyclase of C6 glioma cells.

The adenylate cyclase of C6 glioma cell cultures was characterized for sensitivity to the beta-adrenergic agonist isoproterenol, as well as fluoride, and GTP as a function of the cell cycle. The mitotic phase of the cell cycle was emphasized because both the basal cellular cyclic AMP level and the intact C6 cell's capacity to accumulate cyclic AMP in response to isoproterenol decreased during mitosis. Basal and stimulated adenylate cyclase activities in mitotic cells were decreased relative to the enzyme activities in the G1, S, and G2 phases of the cell cycle. Analysis of the beta-adrenergic receptor using the radioligand(-)[3H]dihydroalprenolol showed that neither ligand affinity nor receptor density changed during the cell cycle, indicating that the reduced adenylate cyclase activity of the mitotic C6 cell was not caused by alterations in this hormone receptor. The reduction in the mitotic cell's basal adenylate cyclase activity was more prominent than the decrease in isoproterenol-, fluoride, or GTP-stimulated activities suggesting that the effectiveness of these enzymes activators (i.e., the efficiency of the coupling mechanism) was not attenuated during mitosis. These studies indicate that the intrinsic catalytic capacity (not the beta-adrenergic receptor or the coupling mechanism) of the C6 adenylate cyclase complex is reduced during mitosis and contributes to the mitotic cell's inability to accumulate and maintain the cyclic AMP concentration at the interphase level.

Adenylyl Cyclases↗
PubMed1981

Rapid and reversible reduction of junctional permeability in cells infected with a temperature-sensitive mutant of avian sarcoma virus.

The transformed or normal phenotype of cultured normal rat kidney cells infected with a temperature-sensitive mutant of avian sarcoma virus is conditional on the temperature at which the cells are grown. Using dye injection techniques, we show that junction-mediated dye transfer is also temperature-sensitive. The extent and rate of transfer between infected cells grown at the transformation-permissive temperature (35 degrees C) is significantly reduced when compared to infected cells grown at the nonpermissive temperature (40.5 degrees C) or uninfected cells grown at either temperature. Infected cells subjected to reciprocal temperature shifts express rapid and reversible alterations of dye transfer capacities, with responses evident by 15 min and completed by 60 min for temperature shifts in either direction. These results suggest that altered junctional capacities may be fundamental to the expression of the ASV-induced, transformed phenotype.

Alpharetrovirus↗
PubMed1981

Fluorescence spectroscopic studies of Huntington fibroblast membranes.

Using fluorescence spectroscopic methods, we compared the membrane properties of intact fibroblasts from both normal subjects and patients with Huntington disease (HD). Cells were stained with various fluorophores, including 1-anilino-8-naphthalene sulfonic acid (ANS), 2-toluidinyl-6-naphthalene sulfonic acid (TNS), 1,6-diphenyl-1,3,5-hexatriene (DPH), and 6-lauroyl-2-(dimethylamino)-naphthalene (LAURDAN). Using these labeled cells, we measured fluorescence yields and emission maxima (ANS, TNS, and LAURDAN), polarizations (TNS, DPH, and LAURDAN), lifetimes (TNS), and differential polarized lifetimes (DPH). In each instance, comparisons were made between cells from normal and from HD individuals. These cultures were controlled for passage number in culture and for age of donor. We found no significant differences between the HD and the control fibroblasts in experiments using the above-mentioned probes and spectroscopic parameters.

Cell Membrane↗
PubMed1980

The induction and detection of diploid spermatozoa in Gallus domesticus.

Colcemid at the dose level of 0.37 mg/kg/day was injected intraperitoneally to 3 sexually active chicken males for 3 consecutive days. 10-12 days after the first colcemid injection, 14-25% of the sperm population in the semen samples from the treated males was found to be diploid in DNA content by flow microfluorometric analysis. Cytogeneic and developmental analyses on early embryos indicate that, during the process of spermatogenesis, the male germ cells are most susceptible to colcemid treatment 10-12 days prior to the maturation of the spermatozoa which is equivalent to the primary through secondary spermatocyte stages in chicken males. By the application of an extremely unequal chromosomal translocation as a cytological marker of parentage, it is confirmed that the diploid sperm induced are capable of uniting with a normal haploid or diploid egg to produce a triploid or tetraploid zygote.

Animals↗
PubMed1980

Comparative effects of vindesine, vinblastine, and vincristine on mitotic arrest and hormonal response of L1210 leukemia cells.

The Vinca alkaloids vinblastine, vincristine, and vindesine were compared for their capacity to arrest L1210 cells in mitosis and for their effect on the response of L1210 cells to beta-adrenergic and prostaglandin E1 hormones. Both microscopic and flow cytofluorimetric studies showed that, of the three drugs, vindesine was the most potent for inhibiting growth and arresting L1210 cells in mitosis. In the hormone response studies, vindesine was also found to be the most potent of the three drugs. Cells pretreated for 30 min with 10(-6) M concentrations of the drugs increased the initial hormone response after 5 min of L1210 cells to either hormone by 100%. After a 1-hr exposure to either isoproterenol or prostaglandin E1, however, the vindesine-treated cells had cyclic adenosine 3':5'-monophosphate levels several times higher than did controls. Vinblastine- and vincristine-treated cells were not different from the controls after 1 hr of hormone exposure. Dose-response studies show that the vindesine effect on enhanced cyclic adenosine 3':5'-monophosphate accumulation increases substantially with drug concentrations from 10(-8) to 10(-5) M. These and other studies indicate that the effect of the Vinca alkaloids on cyclic adenosine 3':5'-monophosphate metabolism occurs at the transduction mechanism which couples the hormone receptor to the catalytic component of the adenylate cyclase complex.

Animals↗
PubMed1979

Endogenous cyclic AMP does not modulate transport of hexoses, nucleosides, or nucleobases in Chinese hamster ovary cells.

In a previous study we have demonstrated that neither extracellular nor intracellular cyclic adenosine monophosphate (AMP) levels directly affect the uptake of nucleosides, nucleobases, or hexoses by various types of cultured mammalian cells. Uptake of these nutrients into cells, however, involves two processes operating in tandem: facilitated transport across the membrane and intracellular phosphorylation; and uptake rates generally reflect the rates of substrate phosphorylation rather than of transport. In the present study we have examined the question of whether substrate transport per se is regulated by intracellular cyclic AMP. Initially various cell lines, grown both in suspension and monolayer culture, were screened for their cyclic AMP response to prostaglandin E1, isoproterenol, and inhibitors of cyclic AMP phosphodiesterase. Prostaglandin E1 treatment of Chinese hamster ovary cells was selected as the system giving the largest and most consistent (50-fold to 100-fold) elevation of cyclic AMP. Rapid kinetic techniques were used to measure the transport of 3-O-methylglucose, thymidine, adenosine, hypoxanthine, and adenine in wild-type cells and in mutant sublines incapable of phosphorylating these substrates. In no case was an increase in intracellular cyclic AMP accompanied by a significant change in the rate of transport of these substrates, although prostaglandin E1 slightly inhibited the transport of various substrates.

Animals↗
PubMed1978

Rise and fall of cyclic AMP required for onset of lymphocyte DNA synthesis.

The adenosine 3',5'-monophosphate (cyclic AMP) levels of mouse lymphocytes rose and fell sharply 10 hours after stimulation with concanavalin A. Treatment of the cells with indomethacin reversibly prevented the increase in cyclic AMP and the subsequent onset of DNA synthesis. When the heightened cyclic AMP before S phase was maintained by either inhibiting phosphodiesterase or by adding the 8-bromo derivative of cyclic AMP, DNA synthesis was also blocked. Both the increase and decrease in cyclic AMP appear to be required for progression of lymphocytes into the S phase of growth.

4-(3-Butoxy-4-methoxybenzyl)-2-imidazolidinone↗
PubMed1978

Brain reaggregate cultures: biochemical evidence for myelin membrane synthesis.

Myelin membrane synthesis was studied using mechanically dissociated fetal rodent CNS which formed spherical reaggregates while being maintained in rotating culture flasks. These reaggregate cultures exhibited myelinogenesis in vitro after precisely the same period of time needed for myelin synthesis to commence in vivo. The myelin membrane related enzymes, 2', 3' cyclic nucleotide phosphohydrolase (CNP) and cerebroside sulfotransferase (CST), appear similar in their specific activities and follow the same developmental patterns that these enzymes exhibit in vivo. In addition, phosphorylation of myelin basic protein occurs by the third week in vitro which agrees with previously published in vivo studies. These experiments indicate that this nerve-cell culture system may be a appropriate model for studying the biological regulation of myelinogenesis as well as a variety of other nervous-system functions.

2',3'-Cyclic-Nucleotide Phosphodiesterases↗
PubMed1978

Barbiturate dependence in mice: evidence for beta-adrenergic receptor proliferation in brain.

C57BL/6J and DBA/2J inbred mice were assayed for beta-adrenergic receptor density following seven days of chronic phenobarbital intoxication. These physically dependent mice showed a greater brain receptor density than did the control, while acute intoxication to phenobarbital produced no significant effects. C57BL/6J mice showed a higher average receptor density than did the DBA/2J mice under all conditions. These findings support the hypothesis that chronic barbiturate intoxication leads to a receptor supersensitivity which may play a role in the phenomena of functional tolerance and physical dependence.

Alprenolol↗
PubMed1977

G2+M arrest of cultured mammalian cells after incorporation of tritium-labeled nucleosides.

Novikoff rat hepatoma cells were propagated in suspension cultures containing 0.5 to 10 muC of 3H-methyl-thymidine, 3H-5-uridine, 3H-G-adenosine or 3H-8-adenine. The presence of the 3H-labeled precursors caused an inhibition of cell replication which was due to a delay or arrest of the cells in G2 and M. The degree of inhibition was proportional to the amount of radioactivity incorporated into nucleic acids. Almost immediate and complete inhibition resulted from incubation with 10 muC 3H-thymidine/ml. The presence of 0.5 muC 3H-thmidine/ml caused a significant increase in the relative proportion of cells in G2+M, even though the population doubling time of the culture appeared to be unaltered.

Adenine↗