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R W Dougherty

Publications and source records attributed to R W Dougherty.

At least 19 recordsLinked to original sources
PubMed1995

Modification of receptor selectivity and functional activity in cholecystokinin peptoid ligands.

Hybrid analogs of the cholecystokinin A (CCK-A) receptor selective tetrapeptide agonist Boc-Trp-Lys(Tac)-Asp-MePhe-NH2 (1,A-71623) and the CCK-B receptor selective antagonists PD-135118 (2) and CI-988 (3) were prepared. Incorporation of the Lys(Tac) side chain into 2 produced a moderately potent antagonist of CCK-8 in the isolated guinea pig gallbladder (GPGB). Incorporation of the Lys(Tac) side chain into 3 produced the novel agonist analog 7 (EC50 = 28 nM in the GPGB) with excellent affinity for both human CCK-A (IC50 = 12 nM) and CCK-B (IC50 = 17 nM) receptors. Analog 7 was a full agonist (EC50 = 3.5 nM) for calcium mobilization on CHO-K1 cells expressing hCCK-A receptors but a partial agonist on CHO-K1 cells expressing hCCK-B receptors, eliciting a weak agonist response (EC50 = 2800 nM) and antagonizing CCK-8-induced calcium mobilization (KB = 20 nM). Despite this unusual in vitro profile, analog 7 was a potent anorectic agent in rats (ED50 = 30 nmol/kg) following intraperitoneal administration.

Adamantane↗
PubMed1989

Dynamics of the diradylglycerol responses of stimulated phagocytes.

The generation of diradylglycerols (sn-1,2 diacylglycerols (DAG) and 1-O-alkyl-2-acylglycerols (AAG] was investigated in human polymorphonuclear leukocytes stimulated with fMet-Leu-Phe, phorbol myristate acetate (PMA), or A23187. With each stimulus, the elevations in the mass of DAG clearly preceded that of AAG. The levels of both lipids increased over time, peaked by 15-20 min (fMet-Leu-Phe) or 45-60 min (PMA or A23187) and returned slowly toward base line thereafter. The base-line levels of DAG were some 4-fold higher than levels of AAG. On stimulation, the relative increases in AAG (approximately 4-fold, fMet-Leu-Phe; approximately 20-fold, PMA and A23187) were much greater than the corresponding relative increases in the levels of DAG (approximately 2-fold fMet-Leu-Phe; approximately 5-fold, PMA and A23187). The diradylglycerol responses were dependent upon agonist concentration. Prior treatment with cytochalasin B augmented the fMet-Leu-Phe diradylglycerol responses but did not alter unstimulated or PMA- or A23187-stimulated diradylglycerol responses. Depletion of extracellular Ca2+ blocked responses to fMet-Leu-Phe, but not to PMA. Treatment with pertussis toxin: (a) completely blocked the responses to fMet-Leu-Phe, (b) slightly suppressed the AAG but not the DAG response to PMA, and (c) did not affect the responses to A23187. Gas chromatographic/mass spectral analyses indicated that the AAG generated during cell activation consists of a mixture of species differentiated by 1-O-alkyl chains of 16:0, 18:0, 18:1 and an additional species that remains uncharacterized. Since DAG and AAG are reportedly activators and inhibitors, respectively, of protein kinase C activities, the sequential generation of these lipid messengers may provide for a system to critically control the activation of protein kinase C.

Calcimycin↗
PubMed1988

Phorbol diesters and dioctanoylglycerol stimulate accumulation of both diacylglycerols and alkylacylglycerols in human neutrophils.

Human neutrophils treated with phorbol 12-myristate 13-acetate (PMA) or dioctanoylglycerol exhibited a large (10-fold), sustained accumulation of the mass of diradylglycerol, beginning 1 min after stimulation and continuing for 30 to 60 min. Phorbol dibutyrate was less potent than PMA in stimulating diradylglycerol accumulation, whereas the 4-alpha analogs of PMA and phorbol dibutyrate were inactive. Submaximal concentrations of PMA (0.5 to 2.5 nM) plus the calcium ionophore, ionomycin (15 to 60 nM), led to synergistic accumulation of diradylglycerols. Chlorpromazine and sphingosine, inhibitors of protein kinase C, blocked PMA-stimulated accumulation of diradylglycerol with IC50 concentrations of 32 and 9 microM, respectively, paralleling their inhibition of PMA-stimulated O2- production. These compounds also inhibited the ionomycin-stimulated accumulation of diradylglycerols. A third protein kinase C inhibitor, H-7, was less effective, inhibiting PMA-stimulated accumulation of diradylglycerol by 25% at 100 microM. Differential sensitivity to alkaline hydrolysis suggests that diradylglycerols that accumulate in response to PMA or ionomycin stimulation are composed of a mixture of two distinct diglyceride species, diacylglycerols and alkylacylglycerols. Whereas diacylglycerol may activate cellular protein kinase C, the importance of the production of alkylacylglycerols is uncertain.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗
PubMed1986

Differential stimulation of the respiratory burst and lysosomal enzyme secretion in human polymorphonuclear leukocytes by synthetic diacylglycerols.

Binding of chemoattractants to receptors on human polymorphonuclear leukocytes (PMN) stimulates the phosphodiesteric cleavage of phosphatidylinositol 4,5-bisphosphate to produce inositol 1,4,5-trisphosphate and 1,2-diacylglycerols. To investigate the possible second messenger function of diacylglycerols in PMN activation, we tested the ability of a series of synthetic sn 1,2-diacylglycerols, known to stimulate protein kinase C in other systems, to promote superoxide anion release, oxygen consumption, lysosomal enzyme secretion, and chemotaxis. None of the diacylglycerols initiated the chemotactic migration of PMN. Several of the diacylglycerols however, were, active in stimulating superoxide anion release and lysozyme secretion, with dioctanoylglycerol (diC8) being the most potent. Unexpectedly, didecanoylglycerol (diC10) induced lysosomal enzyme secretion, but failed to stimulate superoxide production or oxygen consumption. All other biologically active diacylglycerols tested displayed similar EC50 for stimulating lysozyme secretion and superoxide production. The ability of the diacylglycerols to compete for phorbol dibutyrate (PDBu) binding in intact PMN suggested a mechanism for the divergent biological activity of diC10. Although the compounds that stimulated both superoxide production and lysosomal enzyme secretion competed for essentially all [3H]PDBu binding from its receptor, diC10, which only stimulated secretion, competed for 45% of the bound [3H]PDBu. Thus diacylglycerols can selectively activate certain functions of leukocyte chemoattractant receptor. The data suggest that a discrete pool of protein kinase C may mediate activation of the respiratory burst in PMN.

Binding, Competitive↗
PubMed1986

Cytosolic calcium regulates phorbol diester binding affinity in intact phagocytes.

The mobilization of internally sequestered stores of Ca2+ and activation of protein kinase C appear to be involved in neutrophil activation. We have examined the inter-relationship of these two pathways by investigating the effects of modulating Ca2+ activity on the binding of [3H]phorbol 12,13-dibutyrate (PDBU) to protein kinase C in intact phagocytes. Differentiated HL-60 cells were equilibrated with [3H]PDBU prior to stimulation with various agents known to alter Ca2+ homeostasis in cells. Agents that elevated cytosolic Ca2+, such as f-Met-Leu-Phe and A23187, up-regulated radioligand binding by increasing the affinity of the PDBU/protein kinase C interaction. These effects were time- and agonist concentration-dependent and temperature-sensitive. The kinetics of the up-regulation of binding by f-Met-Leu-Phe coincided with the kinetics of Ca2+ mobilization (by quin2 fluorescence measurements). The putative intracellular Ca2+ antagonist 8-(N,N-diethylamino)-octyl 3,4,5-trimethoxybenzoate alone down-regulated [3H]PDBU binding and inhibited the up-regulation of ligand binding by f-Met-Leu-Phe and A23187. Low concentrations of La3+ (0.1-10 microM) also inhibited up-regulation of radioligand binding to f-Met-Leu-Phe and A23187, whereas higher concentrations (0.1-1 mM) alone increased [3H] PDBU binding and supported further up-regulation of ligand binding by the Ca2+-mobilizing agents. These data suggest a role for Ca2+ in the regulation of phorbol diester binding to protein kinase C in intact cells.

Aminoquinolines↗
PubMed1986

Dominant lethal assay of chlordecone and its distribution in the male reproductive tissues of the rat.

Male rats received 3.6 or 11.4 mg/kg/day of chlordecone orally for 5 days. Some statistically significant events were seen in the reproductive data of females mated to males receiving chlordecone. However, these events did not follow a consistent pattern and do not suggest the conclusion that chlordecone causes dominant lethal effects. Male rats received a single oral dose (40 mg/kg) of chlordecone and were killed at 1, 2, 3, 5, 7, 14 or 21 days. Chlordecone was distributed throughout the reproductive tract. The descending order of concentration was seminal vesicular fluid greater than prostate greater than vas deferens greater than seminal vesicle greater than unwashed sperm greater than washed sperm. It is concluded that chlordecone is well distributed throughout the reproductive tract of the male rat, appears in the ejaculate, and does not appear to produce dominant lethal effects.

Animals↗
PubMed1985

Pertussis toxin inhibits chemotactic peptide-stimulated generation of inositol phosphates and lysosomal enzyme secretion in human leukemic (HL-60) cells.

The binding of the chemotactic peptide N-formyl-methionyl-leucyl-phenylalanine to its cell surface receptor rapidly elicits the hydrolysis of phosphatidylinositol 4,5-bisphosphate by phospholipase C to form the putative second messengers inositol 1,4,5-trisphosphate and sn-1,2-diacylglycerol. To investigate the possible role of a guanine nucleotide binding protein in transduction of this membrane signal, we examined the effects of pertussis toxin on chemotactic peptide-stimulated inositol phospholipid metabolism in differentiated HL-60 cells labeled with [3H]inositol. Pertussis toxin inhibited the chemotactic tripeptide-stimulated production of inositol mono-, bis-, and trisphosphates and secretion of N-acetyl-beta-D-glucosaminidase in a time- and concentration-dependent manner. Treatment with pertussis toxin did not alter the total incorporation or the distribution of [3H]inositol in inositol phospholipid. Chemotactic peptide receptor number was unchanged, although a slight decrease in binding affinity was observed. These findings suggest a role for a guanine nucleotide binding protein in coupling the chemotactic peptide receptor to phospholipase C.

Animals↗
PubMed1985

Reproductive toxicity and lack of dominant lethal effects of 2,4-dinitrotoluene in the male rat.

Adult male Sprague Dawley rats were gavaged with 2,4-dinitrotoluene (2,4-DNT) dissolved in corn oil at 0, 60, 180, or 240 mg/kg/day for five days. A single oral dose (0.5 mg/kg) of triethylenemelamine was used as a positive control. Induction of dominant lethal events was scored on the basis of early fetal deaths. At the two lower doses, no consistent changes were observed in the numbers of pre-implantation losses, implantation sites, or living or non-living fetuses. The highest dose of 2,4-DNT tested resulted in a marked decrease in the numbers of sperm-positive females (determined by microscopic examination of vaginal smears for sperm) and pregnant females. These two effects diminished in the latter weeks of mating. The low number of pregnant females at the highest dose made meaningful statistical evaluations difficult. The results indicate that 2,4-DNT does not cause dominant lethal mutations but does adversely affect reproductive performance.

Animals↗
PubMed1984

Secretagogue-induced phosphoinositide metabolism in human leucocytes.

The relationship between receptor binding of the formylated peptide chemoattractant formylmethionylleucylphenylalanine (fMet-Leu-Phe), lysosomal enzyme secretion and metabolism of membrane phospholipids was evaluated in both human polymorphonuclear leucocytes (PMN) and the dimethyl sulphoxide (Me2SO)-stimulated human myelomonocytic HL-60 leukaemic cell line. In both cell types, exposure to fMet-Leu-Phe (100 nM) induced rapid lysosomal enzyme secretion (maximal release less than 30 s) and marked changes in the 32P-labelling of the inositol lipids phosphatidylinositol (PtdIns), phosphatidylinositol 4-phosphate (PtdIns4P), phosphatidylinositol 4,5-bisphosphate [PtdIns(4,5)P2] as well as phosphatidic acid (PtdA). Specifically, levels of [32P]PtdIns and [32P]PtdIns(4,5)P2 decreased rapidly (peak decrease at 10-15s), with a subsequent increase at 30 s and later. PtdIns4P and PtdA showed only an increase. In Me2SO-differentiated HL-60 cells prelabelled with [3H]inositol for 20 h, fMet-Leu-Phe caused a net increase in the cellular content of [3H]inositol phosphates, including a rapid increase in [3H]inositol 1,4,5-trisphosphate, suggesting that PtdIns(4,5)P2 breakdown occurs by a phospholipase C mechanism. Both lysosomal enzyme secretion and changes in phospholipid metabolism occur over the same agonist concentration range with a similar time course. Binding of [3H]fMet-Leu-Phe, although occurring over the same concentration range, exhibited markedly slower kinetics. Although depletion of extracellular Ca2+ had no effect on ligand-induced polyphosphoinositide turnover, PtdIns turnover, PtdA labelling and lysosomal enzyme secretion were severely curtailed. These studies demonstrate a receptor-mediated enhancement of phospholipid turnover that correlates with a specific biological response to fMet-Leu-Phe. Further, the results are consistent with the idea that phospholipase C-mediated degradation of PtdIns(4,5)P2, which results in the formation of inositol trisphosphate, is an early step in the stimulus-secretion coupling pathway of the neutrophil. The lack of correlation between these two responses and the equilibrium-binding condition suggests that either these parameters are responsive to the rate of ligand-receptor interaction or only fractional occupation is required for a full biological response.

Calcium↗
PubMed1975

Intestinal infusion of endotoxin in adrenalectomized calves.

Four adrenalectomized calves, 3 to 4 months old, were allowed to develop adrenocorticosteroid insufficiency and then were given 50 mg of Escherichia coli endotoxin by infusion via cannula into the duodenum. Several physiologic variables were studied to detect changes indicative of endotoxemia. It is concluded that adrenocorticosteroid insufficiency in calves is not an independent factor allowing an intestinal source of endotoxin to cause the development of clinical signs and physiologic changes characteristic of endotoxemia.

Adrenal Cortex Hormones↗
PubMed1975

Preliminary study of appearance of endotoxin in circulatiory system of sheep and cattle after induced grain engorgement.

Edotoxin was detected, using the limulus amebocyte lysate (LAL) test, in the blood of 3 sheep and 1 steer which had been experimentally "overfed" (induced grain engorgement) with a mixture of corn and oats (2:1). The 1st postfeeding blood samples were collected 24 hours after overfeeding. In 2 sheep and 1 steer, the 24-hours blood samples were test positive. In 1 sheep which died, the 48-hour blood sample was the 1st test-positive sample. In all cases, pre-overfeeding blood samples were taken just before overfeeding.

Animal Feed↗
PubMed1975

Grain overload in cattle and sheep: changes in microbial populations in the cecum and rumen.

Samples from the rumen and cecum of cattle and sheep were cultured to determine changes in microbial populations resulting from overfeeding with grain. Before the animals were overfed, the predominant organisms from both sides were those that grew anaerobically on a relatively nonselective ruminal fluid medium and would not grow on selective mediums designed to culture lactobacilli, streptococci, coliforms, or Clostridium perfringens. By 24 hours after overfeeding, lactic acid bacteria had increased in numbers so that they were the most numerous organisms in both the rumen and the cecum. The concentrations of coliforms and C perfringens also increased after overfeeding and were generally higher in the cecum than in the rumen.

Animal Feed↗
PubMed1975

Changes in motility and ph in the digestive tract of experimentally overfed sheep.

Motility of the digestive tract of 4 sheep was studied with radiotelemetric equipment. After base line records were made, each sheep was overfed with 70 g of grain per kilogram of body weight. The ruminoreticulum did not become static until the ingesta pH was less than 5. The cecum had the same patterns of motility and pH as did the ruminoreticulum, but these returned to normal more quickly in surviving sheep. The motility patterns of the abomasum and the small intestine were more erratic. Results indicated that considerable quantities of grain (substrate for microbial growth) reached the cecum before ruminoreticular motility was inhibited.

Abomasum↗
PubMed1975

Physiologic studies of experimentally grain-engorged cattle and sheep.

Seven sheep weighing 34 to 41 kg, each, 3 steers and 1 heifer weighing 230 to 460 kg each, were experimentally, "overfed" (induced grain engorgement). The most significant changes occurred in ruminal ingesta pH, blood pH, packed cell volume (PCV), hemoglobin (Hb), carbon dioxide pressure (PCO2, total CO2 (volume %), blood D-lactic acid, blood HCO3, and base excess. There was no common denominator that was especially pathognomonic.

Abomasum↗