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Biomedical subjects

R B Clark

Publications and source records attributed to R B Clark.

At least 181 records · Page 10Linked to original sources
PubMed1982

Generation of T cell colonies from responder strain 2 guinea pigs that recognize the copolymer L-glutamic acid, L-lysine in association with nonresponder strain 13 Ia antigens.

T cells from nonimmune responder strain 2 guinea pigs were primed in vitro to the copolymer GL in association with allogeneic, nonresponder strain 13 PEC. T cells that recognized GL in association with strain 13 Ia were separated from alloreactive T cells by cloning the population in soft agar following the priming in liquid culture. The existence of T cells of responder origin that recognize antigen in association with nonresponder macrophages is most consistent with clonal deletion models of Ir gene function.

Animals↗
PubMed1982

Ultrastructural effect of penicillin and cycloheximide on Chlamydia trachomatis strain HAR-13.

The effect of cycloheximide and penicillin on the ultrastructural morphology of C. trachomatis strain HAR-13 was examined by electron microscopy. HAR-13 infected McCoy cells were either treated with cycloheximide (1 microgram/ml) or cycloheximide (1 microgram/ml) plus penicillin G (100 U/ml). The studies revealed that cycloheximide alone induced no morphological alterations into the ultrastructure of HAR-13. Both HAR-13 developmental forms, the elementary body and reticulate body, were present inside the treated McCoy cells. The elementary bodies contained the central dense nucleoid and were about 0.3 microns in diameter, while the reticulate bodies were of typical gram negative bacterial morphology and were from 0.5-1.0 microns in diameter. Cycloheximide in combined treatment with pencillin produced giant, swollen reticulate bodies that were 2-4 microns in diameter and in some cases vacuolated. Elementary bodies were noticeably absent. These results indicate that cycloheximide does not alter the morphology of HAR-13. This system is a useful model for studying the ultrastructural morphology of C. trachomatis strain HAR-13.

Animals↗
PubMed1982

Influence of sodium and calcium ions and membrane potential on glutamate receptor desensitization.

1. The effects of Na, Ca and membrane potential on desensitization of postjunctional glutamate receptors on locust muscle were investigated. 2. The kinetics of desensitization were measured ionophoretically. 3. Replacement of Na by equimolar concentrations of the permeant cations Li, NH4 and guanidine and the impermeant cation choline accelerated desensitization onset, increased the steady-state leve of desensitization and reduced the rate of recovery from desensitization. 4. Desensitization onset rates and steady-state levels of desensitization were not significantly altered either by changing the extracellular Ca concentration or by changing the membrane potential of voltage clamped muscle fibres.

Ammonia↗
PubMed1982

Specific muscarinic-cholinergic desensitization in the neuroblastoma-glioma hybrid NG108-15.

The cholinergic agonist carbachol, epinephrine, and the opiate morphine all inhibit prostaglandin E1 (PGE1)-stimulated adenylate cyclase in homogenates from the neuroblastoma-glioma hybrid NG108-15. Pretreatment of the hybrid with 100 microM carbachol resulted in the rapid loss (desensitization) of the carbachol inhibition of adenylate cyclase (t1/2 less than 3 min). The desensitization of the carbachol inhibition was blocked by 0.1 microM atropine. Pretreatment with carbachol (1-24 h) did not significantly affect the inhibition of adenylate cyclase by either epinephrine or morphine, nor did it alter the PGE1-stimulated activity, that is, no supersensitization was observed. Cholate extracts of the particulate fraction from either carbachol-desensitized or of control NG108-15 were able to reconstitute adenylate cyclase activities of the coupling proteins (G/F)-deficient cyc- lymphoma cell membranes with equal efficacy. These results suggested that the coupling proteins of the adenylate cyclase were not altered by the carbachol pretreatment and that desensitization occurs at the receptor or at a receptor-associated level. However, the possibility remained that specific domains of the G/F, which interact only with muscarinic receptors, were altered.

Adenylyl Cyclases↗
PubMed1982

Localization of distinct surface antigens on Chlamydia trachomatis HAR-13 by immune electron microscopy with monoclonal antibodies.

Monoclonal antibodies were produced against the HAR-13 strain (A) of Chlamydia trachomatis. By an indirect immunofluorescence technique, three clones (1-7, 2-8, 9F) were found to produce antibody that reacted only with serotype A strains. Clone 3-5 produced antibody that cross-reacted with serotypes A, C, H, I, and J. Using an indirect immunoferritin technique, we examined the binding of these antibodies to strain HAR-13 by electron microscopy. Antibodies from all four clones were shown to bind to the outer membrane surface of both reticulate and elementary bodies. Two distinct binding patterns were demonstrated. Antibodies from clones 1-7 and 9F bound to the outer membrane surface in a homogeneous pattern, whereas antibodies 2-8 and 3-5 bound to the outer membrane surface in an irregular, patchy distribution. There was a direct correlation between the distribution of antigen on the outer membrane surface and neutralization of C. trachomatis HAR-13 infectivity in vitro. Neutralization of HAR-13 infectivity by antibodies 1-7 and 9F was complement dependent, whereas antibodies 2-8 and 3-5 did not neutralize under any conditions.

Antibodies, Monoclonal↗
PubMed1982

beta-Adrenergic receptors and cyclic AMP responses to epinephrine in cultured human fibroblasts at various population densities.

The beta-adrenergic receptors of the intact human lung diploid fibroblast line Wl-38 and an SV-40 transformed clone of Wl-38, Wl-38-VA-13-2RA (VA13), were estimated in experiments utilizing the beta-adrenergic ligand, 125l-hydroxybenzylpindolol (125IHYP). When specific 125IHYP binding was measured in cells grown to relatively low population densities (0.15x10(6)cells/35mm dish), both Wl-38 and VA13 cells had approximately 40,000 beta-adrenergic receptors per cell. Wl-38 cells, when cultured to a high population density (0.5x10(6) cells/35/mm dish) had clearly diminished numbers of beta-adrenergic receptors and greatly decreased cAMP responses to epinephrine stimulation. On the other hand, in VA13 cells, neither the receptor number nor the beta-adrenergic response was affected by cell population density. In Wl-38 cells, the diminished cAMP response to epinephrine paralleled the decrease in number of beta-adrenergic receptors. Prostaglandin E1 (PGE1) stimulation of cAMP levels was unaffected by cell population density in either Wl-38 or VA13 cells. Thus, increased cell population density, perhaps related to density dependent inhibition of growth, caused a specific diminution in 125IHYP binding concomitant with decreased cAMP responses to epinephrine.

Cell Count↗
PubMed1982

Differences in the forskolin activation of adenylate cyclases in wild-type and variant lymphoma cells.

The ability of the diterpene forskolin to stimulate cyclic AMP accumulation in intact cell and membrane preparations of wild-type S49 lymphoma cells (WT) and a number of variants has been confirmed. Additionally, a number of salient new findings have emerged: (a) A time delay in forskolin stimulation of cyclic AMP accumulation and adenylate cyclase (t 1/2 approximately equal to 1.5 min) occurred in all hormone-sensitive WT and variant cell and membrane preparations tested. (b) The time delay was missing in the adenylate cyclase-deficient variant (cyc-) of the S49 lymphoma cell, which also lacks functional adenylate cyclase-coupling proteins. (c) The simultaneous addition of epinephrine and forskolin to WT cells or to membrane preparations eliminated the time delay. (d) Forskolin stimulation of intact WT cells did not appear to desensitize adenylate cyclase. (e) The activation of WT adenylate cyclase by forskolin was biphasic with respect to concentration, with both high- and low-affinity components being apparent. In cyc-, only the low-affinity component was detected.

Adenylyl Cyclases↗
PubMed1982

Direct evidence for the role of the coupling proteins in forskolin activation of adenylate cyclase.

Forskolin stimulation of adenylate cyclase in wild type (WT) S49 lymphoma membrane preparations exhibited a lag and biphasic kinetics as a function of the forskolin concentration (i.e., both a high and low affinity component were observed). In contrast to WT, forskolin stimulation of cyc- adenylate cyclase in membranes demonstrated no observable lag and only the low affinity component. Both the lag and the high affinity component characteristic of forskolin activation of WT were observed in cyc- reconstituted with cholate extracts of WT which contained the G/F protein. The potency of forskolin stimulation of reconstituted adenylate cyclase was increased still further if the reconstitution was carried out with epinephrine and Gpp(NH)p. The Vmax of the forskolin stimulation of adenylate cyclase was approximately the same in the reconstituted and unreconstituted cyc-. In addition to the experiments with reconstituted cyc-, we have demonstrated that forskolin increased the apparent affinity of epinephrine for activation of adenylate cyclase in WT, and reciprocally, epinephrine increased the apparent affinity of forskolin for activation. We conclude that the lag, biphasic kinetics of forskolin activation and the synergism of hormone and forskolin activation of WT are attributable to functional G/F and are consistent with the forskolin stabilization of the activated catalytic unit of adenylate cyclase.

Adenylyl Cyclases↗
PubMed1981

Adenylate cyclase coupling proteins are not essential for agonist-specific desensitization of lymphoma cells.

Epinephrine or prostaglandin E1 pretreatment of intact S-49 lymphoma cells specifically desensitized adenylate cyclase activity. Cholate extracts of membranes from these desensitized cells showed no difference from controls in their reconstitution of hormone-stimulated adenylate cyclase activity in the adenylate cyclase-deficient variant (cyc-) of the lymphoma cell line. Adenylate cyclase in the cyc- variant was also specifically desensitized by epinephrine in prostaglandin E1. The desensitization was assessed by reconstituting the cyc- adenylate cyclase with cholate extracts of the wild type lymphoma cells. Since the cyc- has been shown to lack the coupling or G/F proteins of the adenylate cyclase complex, and these proteins are restored by cholate extracts from wild type cells, we conclude that the G/F proteins are not altered or necessary for specific desensitization of the lymphoma cell lines.

Adenylyl Cyclases↗
PubMed1981

Influence of agonists on desensitization of glutamate receptors on locust muscle.

1. The desensitization, by ionophoretically and bath-applied L-glutamate and agonists, of excitatory post-junctional receptor populations on locust extensor tibiae muscle was investigated. 2. The kinetics of onset of and recovery from desensitization were determined ionophoretically either with trains of glutamate/agonist pulses of different frequencies ('conditioning trains') followed by a single 'test' pulse at different intervals after cessation of the conditioning train or with trains of constant frequency drug pulses superimposed upon steady 'conditioning' doses. Both methods gave quantitatively similar results. 3. For approximately equipotent doses, L-glutamate and agonists produced different rates of desensitization onset. Qualitatively similar results were obtained when changes in input conductance of single muscle fibres were measured during bath application of the amino acids. 4. The time courses of recovery from desensitization for receptor populations activated by glutamate and agonists could be described adequately by single exponentials and were independent of the concentration of amino acid except when desensitization exceeded ca. 90%. 5. The rate of recovery from desensitization was different for each agonist.

Animals↗