Biomedical subjects

R G Downer

Publications and source records attributed to R G Downer.

At least 19 recordsLinked to original sources

PubMed2001

Testing for elevated disease rates using smoothed estimates.

A method of testing for elevated disease rates using smoothed estimates is introduced for counts aggregated to a specific administrative level. A test statistic is developed based on an approximation to the smoothed estimate from a penalized multinomial likelihood and an approximate critical value is derived. Empirical investigation reveals that smoothing can increase the ability to detect elevated rates. The methods are applied to data on gastric cancer in Nova Scotia, Canada.

Aged↗

PubMed1995

Isolation of nonsedimentable lipid-protein particles from insect intestine.

Nonsedimentable lipid-protein particles have been isolated from intestinal tissue of the American cockroach, Periplaneta americana. Most of the particles were within the range 30-50 nm in diameter and appear to originate from larger structures. Lipid analysis of the particles showed them to be enriched in neutral lipid components relative to microsomal membranes. Specifically, there is a decline in the amounts of phosphatidylcholine and phosphatidylethanolamine in the nonsedimentable particles compared with the microsomal membranes. Also, in contrast to microsomal membranes, the particles have a higher content of phosphatidic acid along with 1,2- and 1,3-diacyglycerols, free fatty acids and an unidentified lipid that co-migrates with sterol ester, wax ester and hydrocarbon standards in thin layer chromatograms. The cytosol, separated from the particles by ultrafiltration, contained phosphatidic acid, free fatty acids and the unidentified lipid. By contrast, the composition of neutral lipids in the cytosol resembles that of the particles. SDS-PAGE analysis of microsomal membranes, the particles and particle free cytosol shows an enrichment of low molecular weight proteins in the particles and cytosol. The particles and cytosol appear to possess proteolytic activity that is distinguishable from that of corresponding microsomal membranes since the incubation of these components with BSA resulted in the formation of distinct polypeptides. Many characteristics of these particles resemble those of the deteriosomes that have been isolated from plant tissue.

Animals↗

PubMed1994

Characterization of tyramine and octopamine receptors in the insect (Locusta migratoria migratorioides) brain.

The kinetic and pharmacological properties of [3H]tyramine and [3H]octopamine binding to membrane preparations of locust (Locusta migratoria migratorioides) brain were studied to characterize the tyramine and octopamine receptors. [3H]Tyramine and [3H]octopamine bind specifically and reversibly to the locust brain membrane with equilibrium achieved after 20 min. The dissociation of [3H]tyramine is monophasic while that of the [3H]octopamine shows a biphasic tendency. Scatchard analysis of the saturation curves reveals a single high affinity binding site for each of tyramine and octopamine. The mean (+/- S.E.M.) values of Kd and Bmax are 6.11 +/- 0.71 nM and 21.45 +/- 3.0 fmol/mg tissue for tyramine and 5.65 +/- 0.91 nM and 15.0 +/- 2.4 fmol/mg tissue for octopamine, respectively. Pharmacological analysis of the binding suggests the presence of both tyramine and octopamine receptors in the locust brain. alpha-Adrenergic agonists and antagonists have a high affinity to the octopamine but not the tyramine receptor whereas dopaminergic drugs have a higher affinity to the tyramine receptor than the octopamine receptor. No highly effective inhibitors of tyramine binding were identified. The serotonergic blockers, mianserin, LSD, BOL are effective blockers for both tyramine and octopamine receptors, whereas the serotonergic antagonist gramine is more active against the octopamine than the serotonin receptor. The results suggest that a G-protein binding mechanism is involved in the expression of both the tyramine and octopamine effects.

Adrenergic alpha-Agonists↗

PubMed1994

Octopamine enhances phagocytosis in cockroach hemocytes: involvement of inositol trisphosphate.

Octopamine and 5-hydroxytryptamine (5-HT) were previously shown to affect phagocytosis in cockroach hemocytes through unidentified receptor-mediated events. In the present study, we examined the ability of 5-HT and octopamine to enhance inositol trisphosphate (IP3) production using hemocyte membranes of the American cockroach, Periplaneta americana. Octopamine enhanced IP3 production with a maximal peak at 100 nM. Similarly, 5-HT enhanced IP3 production with a maximal effect at 10 nM. The effects of 5-HT and octopamine are not additive, suggesting that both are working through the same receptor. Phentolamine, a general octopamine antagonist, blocked the effects of octopamine and 5-HT, while a mammalian 5-HT2 antagonist that blocks 5-HT-sensitive receptors in insect peripheral tissue, ketanserin, did not. A pharmacological profile indicates that the receptor is similar to an octopamine1-type. Octopamine at 1 microM increased phagocytosis in cockroach hemocytes exposed to Staphylococcus aureus in vitro, and this effect was mimicked by IP3 (10 microM). The octopamine-treated hemocytes were shown to increase IP3 production in the latter stage of phagocytosis. Adult cockroaches exposed to an LD50 dose of S. aureus in conjunction with either 0.1 mM octopamine or the octopamine1 agonist, clonidine, had higher survival rates compared to saline-treated cockroaches. Correspondingly, the octopamine1 antagonist, chlorpromazine, partially blocked the octopamine-mediated increase in cockroach survival.

Analysis of Variance↗

PubMed1993

Partial characterization of protein kinase C from an insect cell line.

The characteristics of protein kinase C (EC 2.7.1.37) from an insect cell line (Choristoneura fumiferana) have been described. DEAE-cellulose chromatography produced a major peak of activity which eluted at 0.04-0.055 M NaCl. The enzyme was sensitive to phosphatidylserine in the presence of calcium. Phorbol 12-myristate 13-acetate (PMA) in nanomolar concentrations stimulated protein kinase C activity 8-fold over basal levels and reduced the enzymes requirement for Ca2+. The enzyme had a Ka of 10 nM for PMA. Diacylglycerols tested included diolein, dilinolein, diarachidonin, oleoyl-acetyl-glycerol, dioctonoyl-sn-glycerol, dipalmitin and distearin. A 2.5- to 3-fold activation was obtained in the presence of 26 microM diolein, 40 microM oleoyl-acetyl-glycerol and 46 microM dioctonoyl-sn-glycerol. The enzyme activity was sensitive to the inhibitor H-7 and 50% inhibition was achieved at a concentration of 52 microM H-7. Phosphatidylinositol enhanced enzyme activity in the absence of phosphatidylserine but phosphatidylethanolamine had no effect.

Animals↗

PubMed1993

Characterization of the tyraminergic system in the central nervous system of the locust, Locusta migratoria migratoides.

Tyramine occurs in the central nervous system (CNS) of the migratory locust, Locusta migratoria migratoides. The distribution of tyramine within the CNS does not parallel that of octopamine. Tyramine is synthesised from tyrosine in the presence of tyrosine decarboxylase. A second decarboxylase in the CNS is active against 5HTP and DOPA. The locust ganglia incorporate tyramine by high- and low-affinity uptake processes that appear to be independent of dopamine and octopamine. Depolarisation of the locust ganglia by high potassium concentration results in calcium-dependent release of incorporated [3H]tyramine.

Animals↗

PubMed1992

Comparative studies on the mode of action of proctolin and phorbol-12,13-dibutyrate in their ability to contract the locust mandibular closer muscle.

The role of proctolin has been further investigated in the locust (Locusta migratoria) mandibular closer muscles. Radioactive calcium uptake measurements were made using protease-dissociated muscle cells. Both the phorbol ester, phorbol-12,13-dibutyrate, and proctolin produce tonic contractions which are associated with the influx of extracellular calcium. The thresholds for proctolin and the phorbol ester to contract the muscle were 1-10 nM and 10-100nM, respectively, while their respective thresholds for evoking measurable calcium influx into the muscle cells were 0.1-1 nM for proctolin, and 0.1-1 pM for phorbol-12,13-dibutyrate. The effect of phorbol-12,13-dibutyrate is blocked by a number of protein kinase inhibitors (at a concentration of 0.1 mM), suggesting that an activation of a protein kinase can lead to calcium influx. These inhibitors, however, do not block the effect of proctolin, indicating that these two compounds work through different pathways, possibly converging on the same final target. In light of this finding, a number of other compounds have been tested to try to ascertain how proctolin mediates an increased calcium influx.

Amino Acid Sequence↗

PubMed1991

Pharmacological characterization of a 5-hydroxytryptamine-sensitive receptor/adenylate cyclase complex in the mandibular closer muscles of the cricket, Gryllus domestica.

The mandibular closer muscles of the cricket, Gryllus domestica, contain a 5-hydroxytryptamine (5-HT)-sensitive receptor that is coupled to adenylate cyclase. A structure-activity study of the 5-HT molecule indicates that the integrity of the ethylamine sidegroup and the presence of a negatively charged moiety at the 5 position (-OH, -OCH3) are essential for activity. A pharmacological profile is presented for this receptor. The receptor differs from any reported mammalian 5-HT receptor in that none of the mammalian agonists tested were effective. However, the mammalian antagonists for 5-HT receptors, spiperone, mianserin, and ketanserin as well as the anti-histaminic cyproheptadine were all effective antagonists in this preparation. Preliminary analysis of antagonism, particularly by spiperone, shows that these antagonists are probably acting non-competitively. On the basis of the pharmacological data, and comparisons with other insect systems, the 5-HT receptor present in the cricket mandibular muscles has been tentatively classified as 5-HT2-like.

Adenylyl Cyclases↗

PubMed1991

Immunofluorescent localization of dopamine-like and leucine-enkephalin-like neurons in the supraoesophageal ganglia of the American cockroach, Periplaneta americana.

Leucine-enkephalin- and dopamine-like nerve cells and fibers were localized in the supraoesophageal ganglia (brain) of the American cockroach, Periplaneta americana, using immunofluorescence. The presence of leucine-enkephalin-like material was confirmed using immunoperoxidase staining. Several cells containing leucine-enkephalin-like material were found in the pars lateralis, and nerve fibers belonging to these cells were traced through the brain. Dopamine-like material was detected in deutocerebral neurons as well as the nerve processes arising from these cells which lead into the area of the deutocerebral glomeruli. Specific immunofluorescence was also obtained in the alpha and beta lobes of the corpora pedunculata with both the leucine-enkephalin and dopamine antibodies. However, the fluorescent banding pattern observed in both lobes was distinctly different with the two antibodies. No specific fluorescence was observed in the stalk or peduncle of the corpora pedunculata with either the leucine-enkephalin or the dopamine antibody. The findings suggest a possible interaction of leucine-enkephalinergic and dopaminergic nerve fibers in the alpha and beta lobes of the cockroach corpora pedunculata.

Animals↗

PubMed1991

Characterization and pharmacological studies of an octopamine-sensitive adenylate cyclase from nerve cord of Locusta migratoria.

An octopamine-sensitive adenylate cyclase that is insensitive to stimulation by dopamine (DA) and 5-hydroxytryptamine (5-HT), was studied in a homogenized nerve cord preparation of the migratory locust, Locusta migratoria. The enzyme complex is similar to catecholamine-sensitive cyclases from mammals with respect to pH-optimum, requirement for Mg++ and GTP, and sensitivity to forskolin. The octopamine-mediated elevation of adenylate cyclase activity is antagonized by a variety of drugs with the following order of potency: mianserin greater than phentolamine greater than promethazine greater than gramine greater than cyproheptadine greater than cis-flupenthixol greater than chlorpromazine greater than metoclopramide.

Adenylyl Cyclases↗

PubMed1990

Proctolin in the innervation of the locust mandibular closer muscle modulates contractions through the elevation of inositol trisphosphate.

Extracts of the locust (Locusta migratoria) mandibular closer muscle separated on reverse-phase HPLC and tested for bio-activity on the locust oviduct contain a bio-active substance that coelutes with authentic proctolin. Furthermore, the effect on oviduct contractions of this compound is indistinguishable from that of authentic proctolin. Antiserum to proctolin stains numerous axons with beaded endings that run along the fibres of the closer muscles and, in addition, the antiserum stains a number of cell bodies in the suboesophageal ganglion, some of which have axons in the mandibular nerve that innervates the mandibular musculature. The function of proctolin appears to be modulatory as its presence significantly increases the amplitude of neurally evoked contractions of the closer muscle. This effect can be mimicked by the addition of inositol 1,4,5-trisphosphate (IP3) to preparations in which the muscles have been permeabilized with dimethyl sulfoxide. The involvement of this second messenger is further implicated as we also show that proctolin produces a large, significant increase in the IP3 content of homogenized muscle.

Animals↗

PubMed1990

Serotoninergic innervation of the locust mandibular closer muscle modulates contractions through the elevation of cyclic adenosine monophosphate.

The mandibular closer muscles of the locust receive innervation that is immunoreactive for the putative transmitter 5-hydroxytryptamine (5-HT). Cobalt-labelling suggests that the origin of this innervation is a group of cells located anteriorly in the suboesophageal ganglion. Bath application of 5-HT while the muscles are active produces marked changes in the contractions, increasing their amplitude, rate of contraction, and rate of relaxation. Incubation of isolated muscles with 5-HT shows that this amine elevates the levels of the cyclic nucleotide cyclic adenosine monophosphate (cAMP). In addition compounds that artificially elevate the levels of cAMP in the muscle--3-isobutyl-1-methylxanthine (IBMX), forskolin, and the cAMP analogue 8-(4-chlorophenylthio) cAMP--mimic the actions of 5-HT, whereas a potent inhibitor of insect adenylate cyclase, adenosine, considerably delays the onset of the effects produced by 5-HT. The effects observed with 5-HT in the mandibular muscle are similar to those of octopamine in the locust extensor tibiae muscle, and it is possible that this is an analogous modulatory system.

Animals↗

PubMed1990

Alkaline phosphatase activity in the brain of the American cockroach Periplaneta americana L.

The supra- and suboesophageal ganglia of the American cockroach contain material which catalyses the alkaline hydrolysis (pH 9.5) of 5-bromo-4-chloro-3-indolyl phosphate in the presence of Nitro blue tetrazolium. Histochemical studies on unfixed cryostat sections indicate that this type of alkaline phosphatase is restricted to discrete regions in the cockroach brain. Highest enzyme activity is encountered in the mushroom bodies, central body, antennal glomeruli and specific parts of some distinct neural connections including the optic nerve, antennal nerve, circumoesophageal connectives and nerves leaving the suboesophageal ganglion. Tissue fixation by use of formaldehyde-type fixatives, as well as routine paraffin-embedding, completely destroy all histochemically detectable enzyme activity. Native polyacrylamide gradient electrophoresis suggests that the alkaline phosphatase activity is present as multiple isozymic forms, which show up in the 120-130 kD range of standard proteins. Enzyme activity becomes undetectable after fixation (trichloroacetic acid, formaldehyde containing fixatives) of electrophoretically separated native proteins, as well as after electrophoresis in denaturing conditions (SDS and beta-mercapto-ethanol, boiling). However, the enzyme activity remains virtually unaffected after storage of the sample for prolonged periods at -20 to -80 degrees C.

Alkaline Phosphatase↗

PubMed1990

Stimulation of carbohydrate metabolising enzymes by synthetic hypertrehalosemic peptides in thoracic musculature of the American cockroach, Periplaneta americana.

The ability of the synthetic hypertrehalosemic peptides, HT-I and HT-II, to influence the activities of glycogen phosphorylase, trehalase and hexokinase via elevation of Ca++ and cAMP levels was examined in thoracic musculature of the American cockroach, Periplaneta americana. The peptides effect dose- and time-dependent activation of phosphorylase, trehalase and hexokinase activities that occur concomitantly with elevated levels of intracellular calcium. In addition, HT-I increases the accumulation of cyclic AMP in muscle cells.

Amino Acid Sequence↗

PubMed1989

Purification and characterization of trehalase inhibitor from hemolymph of the American cockroach, Periplaneta americana.

An endogenous proteinaceous inhibitor of trehalase (alpha,alpha-trehalose-1-glucohydrolase: EC 3.2.1.28) has been isolated and purified from the serum of resting adult American cockroaches, Periplaneta americana. Purification procedures involved decreasing ionic strength, gel filtration, and reversed phase high performance liquid chromatography. Homogeneity was confirmed by polyacrylamide gel electrophoresis and end group analysis. The purified protein inhibited trehalase activity in a dose-dependent manner and was estimated to have a molecular weight of 86,000 and to contain sugar chains. An automated gas-phase sequencer was used to determine the following sequence for the N-terminal amino acid residues: H-Ala-Ilu-Pro-Thr-Pro-His-Val-Tyr-Lys-Val-X-Val-Pro-Asp-Gly-Ala-Le u-Asn-Asp.

Amino Acid Sequence↗

PubMed1989

Microassay for N-acetyltransferase activity using high-performance liquid chromatography with electrochemical detection.

A rapid, sensitive procedure has been developed for determination of N-acetyltransferase activity against octopamine, dopamine and 5-hydroxytryptamine. The assay, which is performed in a volume of 10 microliters, is based upon the separation and detection of monoamine substrates and their N-acetylated derivatives using high-performance liquid chromatography with electrochemical detection. The method has been used to measure N-acetyltransferase activity against octopamine, dopamine and 5-hydroxytryptamine in the cerebral ganglion of the American cockroach, Periplaneta americana and to study bi-substrate kinetics of the enzyme.

5-Hydroxytryptophan↗

PubMed1989

A substance resembling somatomedin C in the American cockroach.

Material antigenically resembling somatomedin C (type I insulin-like growth factor, IGF-I) is demonstrated in the American cockroach Periplaneta americana by means of a monoclonal antibody immunoperoxidase technique. It was localized histochemically in neuronal cell somata and axonal fibers (probably interneurons) of the central nervous/neuroendocrine system and in 'endocrine-type' cells lining the midgut epithelium. The IGF-I-like substance is different from vertebrate insulin and also distinct from materials immunostained by different insulin antibodies in the brain and neurohaemal complex of this insect species. These findings are viewed in the light of recent reports on the presence and action of insulin-like chemicals in insects, and with respect to the existence of an insect brain-midgut system similar to the mammalian brain-gastroenteropancreatic system.

Animals↗

PubMed1988

Modulation of octopamine-mediated production of cyclic AMP by phorbol-ester-sensitive protein kinase C in an insect cell line.

The presence of protein kinase C (EC 2.7.1.37) in an insect cell line has been demonstrated. Phorbol 12-myristate 13-acetate (PMA), in micromolar concentrations, activated protein kinase C with a translocation of the enzyme from the cytosol to the particulate fraction. Cyclic AMP production in the presence of PMA, octopamine and a combination of both increased in a dose-dependent and time-dependent fashion. The biologically inactive 4 alpha-phorbol 12,13-didecanoate had no effect on protein kinase C activity or on octopamine-mediated cyclic AMP production. Pretreatment of the cells with pertussis toxin had no effect on the response of cells to octopamine or PMA. However, pretreatment with cholera toxin resulted in increased cyclic AMP production which was further enhanced when both cholera toxin and PMA were used in combination. Our data indicate that the octopamine-mediated cyclic AMP production is modulated by protein kinase C.

Animals↗