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G Shaw

Publications and source records attributed to G Shaw.

At least 91 records · Page 5Linked to original sources
PubMed1999

Virilization of the male pouch young of the tammar wallaby does not appear to be mediated by plasma testosterone or dihydrotestosterone.

Virilization of the male urogenital tract of all mammals, including marsupials, is mediated by androgenic hormones secreted by the testes. We have previously demonstrated profound sexual dimorphism in the concentrations of gonadal androgens in pouch young of the tammar wallaby Macropus eugenii during the interval when the urogenital sinus virilizes. To provide insight into the mechanisms by which androgens are transported from the testes to the target tissues, we measured testosterone and dihydrotestosterone in plasma pools from tammar pouch young from the day of birth to Day 150. Plasma testosterone levels were measurable (0.5-2 ng/ml) at all times studied, but there were no differences between males and females. These low concentrations of plasma testosterone appear to be derived from the adrenal glands and not the testes. Plasma dihydrotestosterone levels in plasma pools from these animals were also low and not sexually dimorphic. We conclude that virilization of the male urogenital tract cannot be explained by the usual transport of testosterone or dihydrotestosterone in plasma but may be mediated by the direct delivery of androgens to the urogenital tract via the Wolffian ducts. Alternatively, circulating prohormones may be converted to androgens in target tissues.

Adrenal Glands↗
PubMed1999

Apoptosis induced by infection of primary brain cultures with diverse human immunodeficiency virus type 1 isolates: evidence for a role of the envelope.

Apoptosis of neurons and astrocytes is induced by human immunodeficiency type 1 (HIV-1) infection in vitro and has been demonstrated in brain tissue from patients with AIDS. We analyzed a panel of diverse HIV-1 primary isolates for the ability to replicate and induce neuronal and astrocyte apoptosis in primary human brain cultures. Apoptosis was induced three- to eightfold by infection with the blood-derived HIV-1 isolates 89.6, SG3, and ADA. In contrast, the brain-derived HIV-1 isolates YU2, JRFL, DS-br, RC-br, and KJ-br did not induce significant levels of apoptosis. The ability of HIV-1 isolates to induce apoptosis was independent of their replication capacity. Studies of recombinant chimeras between the SG3 and YU2 viruses showed that replacement of the YU2 Env with the SG3 Env was sufficient to confer the ability to induce apoptosis to the YU2 virus. Replacement of the Env V3 regions alone largely conferred the phenotypes of the parental clones. The SG3 Env used CXCR4 and CCR3 as coreceptors for virus entry, whereas YU2 used CCR5 and CCR3. The V3 regions of SG3 and YU2 conferred the ability to use CXCR4 and CCR5, respectively. In contrast, the 3' region of Env, particularly the C3V4 region, was required in conjunction with the V3 region for efficient use of CCR3. These results provide evidence that Env is a major determinant of neurodegenerative mechanisms associated with HIV-1 infection in vitro and raise the possibility that blood-derived viruses which emerge during the late stages of disease may affect disease progression in the central nervous system.

Animals↗
PubMed1999

Reactivating tammar wallaby blastocysts oxidize fatty acids and amino acids.

The tammar wallaby, Macropus eugenii, has a ruminant-like digestive system which may make a significant concentration of amino acids and fatty acids available to the blastocyst via uterine fluids. Fluorescent and radioisotope analyses were performed to determine the rate of glutamine and palmitate use by blastocysts recovered on day 0, 3, 4, 5 and 10 after reactivation induced by removal of pouch young (RPY). Between day 0 and 4 glutamine uptake increased from 15.6 +/- 6.6 to 36.1 +/- 2.7 pmol per embryo h-1 (P < 0.01) and ammonium production increased from 8.2 +/- 4.3 to 26.6 +/- 3.0 pmol per embryo h-1 (P < 0.01). Glutamine oxidation did not increase until day 10 after RPY (P < 0.01), but the percentage of glutamine oxidized increased from 4.5 +/- 3.1% during diapause to 31.2 +/- 12.6% (P < 0.01) by day 5 after RPY and increased further to 51.0 +/- 15.8% (P < 0.01) by day 10 after RPY. Palmitate oxidation also increased from 0.3 +/- 0.1 by day 0 blastocysts to 3.8 +/- 1.7 pmol per embryo h-1 (P < 0.01) by day 4 blastocysts. This increase provides a greater potential for ATP production, possibly to supply increased demand due to the coincident resumption of mitoses. The ATP:ADP ratio within blastocysts had reduced by the time of the first measurement at day 3 (0.5 +/- 0.2 pmol per embryo h-1; P < 0.01) compared with day 0 blastocysts (1.4 +/- 0.3 pmol per embryo h-1). It is likely that metabolism of amino acids and fatty acids contributes to the energy supply during reactivation of tammar wallaby blastocysts after embryonic diapause.

Adenosine Diphosphate↗
PubMed1999

Effect of sodium cloprostenol and flunixin meglumine on luteolysis and the timing of birth in bitches.

At birth, the physiological role of prostaglandins in bitches is unclear. Bitches were treated before parturition with either saline, the prostaglandin analogue, sodium cloprostenol, or the prostaglandin synthetase inhibitor, flunixin meglumine. The animals were examined regularly to determine the onset of parturition and a series of blood samples were taken to define the hormonal profiles before, during and after birth. Animals treated with cloprostenol whelped earlier than did controls. In addition, the prostaglandin F2 alpha metabolite surge and decrease in plasma progesterone concentration and rectal temperature were earlier than in controls. Flunixin meglumine disrupted the normal 13,14-dihydro-15-keto prostaglandin F2 alpha profile but did not abolish prostaglandin synthesis completely or delay the onset of labour in treated animals. This study confirms that prostaglandins induce luteolysis and the onset of labour in the bitch. However, the partial inhibition of prostaglandin synthesis does not prevent parturition.

Analysis of Variance↗
PubMed1998

Mesotocin receptors during pregnancy, parturition and lactation in the tammar wallaby.

Mestocin receptor concentrations in membrane preparations from reproductive tissues of the tammar Macropus eugenii throughout gestation and lactation were assessed using [3H]-oxytocin as the ligand. There was a single binding site which bound both mesotocin and oxytocin with high and similar affinities. Mesotocin receptor concentrations in the myometrium were low (708 +/- 199 fmol mg-1 protein) in early and middle gestation but increased significantly on day 23 of pregnancy of the 26-day gestation period to 1921 +/- 552 fmol mg-1 protein. Myometrial receptors reached a peak of 2483 +/- 575 fmol mg-1 protein on days 25 and 26 of gestation, but returned to basal levels about an hour after birth. Receptor concentrations in the contralateral non-gravid uterus were much lower (605 +/- 75 fmol mg-1) and did not significantly increase throughout the period of gestation but dropped one day before birth. Mesotocin receptors were undetectable in the endometrium, the yolk sac placenta and the lateral, median and anterior vagina of all animals tested. In the lactating mammary gland after birth mesotocin receptors were initially high (588 +/- 38 fmol mg-1) but decreased after 200 days and by late lactation were 224 +/- 55 fmol mg-1 protein on day 240, close to the time of weaning. Mesotocin receptors in the ipsilateral non-lactating gland were also high in early lactation (430 +/- 153 fmol mg-1) and declined in late lactation (62 +/- 20 fmol mg-1). The changing concentrations of mesotocin receptors in pregnancy and lactation demonstrate that they are specifically regulated in tammar reproductive tissues. The increase in mesotocin receptors in gravid, but not in the non-gravid myometrium three days before birth may make the uterus responsive to the surge of mesotocin at birth. Since this rise is unilateral and only occurs in the gravid myometrium it must be due to local effects from the ipsilateral ovary or the feto-placental unit. Likewise, the down-regulation of mesotocin receptors in the contralateral, non-gravid myometrium may be due to its proximity to the developing follicle. The changing concentrations in the lactating and the adjacent, non-lactating mammary gland also reflect a differential regulation of mesotocin receptors, probably mediated via the sucking stimulus. Thus, local influences appear to be of primary importance in the regulation of mesotocin receptors during reproduction in this marsupial.

Animals↗
PubMed1998

The intermediate filament protein peripherin is a marker for cerebellar climbing fibres.

Immunocytochemical staining with antibodies to the class III intermediate filament protein peripherin reveals discrete subpopulations of neurons and nerve fibres throughout the rat central nervous system. Some of these fibres enter the cerebellar granular and molecular layers. Here we use light and electron microscopic immunocytochemistry and confocal fluorescence microscopy to identify the peripherin positive fibres in the molecular layer of the cerebella of various mammals. (1) The peripherin positive fibres in the molecular layer have morphological attributes of climbing fibres, and peripherin positive fibres are also detected in the olivo-cerebellar tract. Furthermore peripherin positive neurons can be seen in the inferior olive, from which climbing fibres originate. (2) The peripherin positive molecular layer fibres rapidly degenerate in rats treated with 3-acetylpyridine (3-AP), a reagent which destroys neurons in the inferior olive, and the time course of degeneration of these mirrors that previously described for 3-AP induced destruction of climbing fibres. (3) Cerebella of other mammal species tested (mouse, rabbit, pig, cow and human) revealed a similar peripherin staining pattern in the cerebellum, including fibres in the molecular layer with the morphology of climbing fibres. (4) We also noted peripherin positive spinocerebellar and vestibulocerebellar mossy fibres in the cerebellar granular layer of folia known to receive these inputs. (5) A subset of perivascular nerve fibres are also peripherin positive. These results show that peripherin is a useful marker for mammalian cerebellar climbing fibres, and that a subset of morphologically distinct cerebellar mossy fibres are also peripherin positive.

Animals↗
PubMed1998

Reactivating tammar wallaby blastocysts oxidize glucose.

Metabolic reactivation of the tammar blastocyst appears to be characterized by a change in the pathway of glucose metabolism rather than an absolute increase in substrate uptake. The switch in type of metabolism used was examined to gain information on the timing and physiology of blastocyst reactivation. Fluorescent and radioisotope techniques were used sequentially to determine the activity of pathways of glucose metabolism by individual wallaby blastocysts during diapause and 3, 4, 5, 6, 7, 8, and 10 days after removal of pouch young (RPY). Maternal endometrial and luteal cell metabolism and circulating hormone levels were measured and correlated with blastocyst activity. Observed differences between rates of blastocyst reactivation could be explained by variation in the maternal response between animals. While blastocysts recovered 4 days after RPY oxidized more glucose compared with Day 0 blastocysts (p < 0.05), rates of glycolysis did not change until Day 10. Blastocysts recovered between 4 and 10 days after RPY oxidized a significantly greater percentage of the glucose taken up (p < 0.01). The reduced ATP:ADP ratio within blastocysts recovered 3 days after RPY (p < 0.05) indicates that conditions are suitable for blastocysts to undergo a metabolic switch from glycolytic to oxidative metabolism of glucose on Day 4 after RPY. The increased oxidation results in greater ATP production, which plausibly fuels the increased energy requirements of wallaby blastocysts during the early stages of reactivation.

Adenosine Diphosphate↗
PubMed1998

Omeprazole may exert both a bacteriostatic and a bacteriocidal effect on the growth of Helicobacter pylori (NCTC 11637) in vitro by inhibiting bacterial urease activity.

AIMS: To assess the potential antibacterial effect of omeprazole, a benzimidazole proton pump inhibitor, on the growth of Helicobacter pylori in vitro and to evaluate the effect of this compound on bacterial urease activity. METHODS: The growth of H pylori was observed in liquid culture in the presence and absence of omeprazole (0.8 mg/ml). Urease activity was evaluated in aliquots removed from two hour cultures by monitoring the initial change in absorbency at 560 nm in the presence of 0.02% phenol red. RESULTS: The minimum inhibitory concentration of omeprazole against H pylori was 0.8 mg/ml. The concentration of omeprazole required to inhibit growth was dependent on inoculum density: omeprazole (0.8 mg/ml) prevented growth from a 1 x 10(6) cfu/ml inoculum, but not from the higher inocula of 10(7) or 10(8) cfu/ml. This is the first study to demonstrate that omeprazole exerts a bacteriocidal effect against low bacterial densities and a bacteriostatic effect when bacterial density is high. When used at the onset of growth, this concentration of omeprazole has a bacteriocidal effect after four hours, although it exerts a bacteriostatic effect when added to cultures after the exponential phase. Bacterial urease activity is competitively inhibited by omeprazole in a dose dependent manner. CONCLUSION: The results suggest that omeprazole exerts both a bacteriocidal and a bacteriostatic effect against H pylori and competitively inhibits bacterial extracellular urease activity.

Anti-Ulcer Agents↗
PubMed1998

Angiotensin II-induced nuclear targeting of the angiotensin type 1 (AT1) receptor in brain neurons.

Angiotensin II (Ang II) interaction with the neuronal AT1 receptor results in a chronic stimulation of neuromodulation that involves the expression of norepinephrine transporter (NET) and tyrosine hydroxylase (TH). In view of this unique property and the presence of putative nuclear localization signal (NLS) consensus sequence in the AT1 receptor, this study was conducted to investigate the hypothesis that Ang II would induce nuclear sequestration of this G protein-coupled receptor and that the sequestration may have implications on Ang II-induced expression of NET and TH genes. Incubation of neuronal cultures with Ang II caused a time- and dose-dependent increase in the levels of AT1 receptor immunoreactivity in the nucleus. A 6.7-fold increase was observed with 100 nM Ang II, in 15 min, that was blocked by losartan, an AT1 receptor-specific antagonist. Ang II-induced nuclear sequestration was specific for AT1 receptor, because Ang II failed to produce a similar effect on neuronal AT2 receptors. The presence of the putative NLS sequence in the cytoplasmic tail of the AT1 receptor seems to be the key in nuclear targeting because: 1) nuclear targeting was attenuated by a peptide of the AT1 receptor that contained the putative NLS sequence; and 2) Ang II failed to cause nuclear translocation of the AT2 receptor, which does not contain the putative NLS. Ang II also caused a time- and dose-dependent stimulation of P62 phosphorylation, a glycoprotein of the nuclear pore complex. A 6-fold stimulation of phosphorylation was observed with 100 nM Ang II, in 15 min, that was completely blocked by losartan and not by PD123,319, an AT2 receptor specific antagonist. Preloading of neurons with p62-pep (a peptide containing consenses of mitogen-activated protein kinase in p62) resulted in a loss of Ang II-induced p62 phosphorylation and stimulation of NET and TH messenger RNA levels. In conclusion, these data demonstrate that Ang II induces nuclear sequestration of AT1 receptor involving NLS in the AT1 receptor and p62 of the nuclear pore complex in brain neurons. A possible role of such a nuclear targeting of the AT1 receptor on chronic neuromodulatory actions of Ang II has been discussed.

Amino Acid Sequence↗
PubMed1998

Effects of cabergoline on reproduction in three families of Australian marsupials.

The effects of the prolactin inhibiting drug, cabergoline, on pregnant and lactating marsupials were investigated in four species from three diverse families: the tammar wallaby, Macropus eugenii, the quokka, Setonix brachyurus, the brushtail possum, Trichosurus vulpecula, and the fat-tailed dunnart, Sminthopsis crassicaudata. In tammar wallabies, 20 micrograms cabergoline kg-1 injected intramuscularly 1 day before expected birth did not alter the timing of parturition but neonates died within a day of birth, suggesting that the onset of lactation was compromised. During early lactation in tammars (56-69 days post partum), an intramuscular injection transiently retarded growth of the young, although they subsequently survived. This treatment induced reactivation of the quiescent corpus luteum and the blastocyst from diapause, so a new birth occurred 26-27 days later, despite the continued sucking of the young in the pouch. Intramuscular injection during late lactation (166-199 days post partum) apparently suppressed milk secretion since pouch young lost up to 20% of their bodyweight or died within 7 days of treatment. Oral administration of cabergoline had no effect on the growth of the young or on the quiescent corpus luteum and diapausing blastocyst. Quokkas showed similar responses to tammars after treatment in late lactation. Possums and dunnarts were less sensitive to injected cabergoline than the two macropodid species, and possums showed no response to oral administration. The lack of response of these marsupial species to oral cabergoline treatment suggests that accidental ingestion of baits, containing 20 micrograms cabergoline kg-1, used to control introduced eutherian pests such as the red fox, Vulpes vulpes, or the feral cat, Felis cattus, should not affect the reproduction of native marsupials.

Administration, Oral↗
PubMed1998

Effects of 5-hydroxytryptamine2 receptor antagonism on the behavioral activation and immediate early gene expression induced by dizocilpine.

The noncompetitive N-methyl-D-aspartate (NMDA) antagonists dizocilpine and phencyclidine cause behavioral changes in animals that can be blocked by antipsychotic agents, implicating NMDA receptors in the expression of schizophrenic symptoms. In the present study, we examined the effects of dizocilpine (0.1-3.0 mg/kg s.c.) on locomotor activity and on the expression of c-fos and hsp-70 immediate-early genes (IEGs) in mice. Results indicate that dizocilpine increases locomotor activity and selectively increases the expression of c-fos and hsp-70 in the posterior cingulate cortex. Haloperidol (0.01-0.1 mg/kg) and clozapine (0.6-1.25 mg/kg) block both the locomotor response and the increased IEG immunoreactivity induced by dizocilpine (0.6 mg/kg). The 5-HT2 antagonists ritanserin (0.06-0.25 mg/kg), ketanserin (0.03-0.12 mg/kg) and amesergide (0. 3-1.25 mg/kg) also significantly attenuated the locomotor response to dizocilpine. Haloperidol and clozapine suppressed the head weaving induced by dizocilpine, but ritanserin, as previously reported did not. Although some attenuation of the c-fos and hsp-70 immunoreactivity was seen with the 5-HT2 antagonists it was less pronounced than that induced by haloperidol or clozapine. In conclusion, 5-HT2 antagonists as well as antipsychotic compounds attenuate the locomotor response to dizocilpine in mice. Haloperidol and clozapine appear to be more effective, however, in attenuating the expression of c-fos and hsp-70 in the posterior cingulate gyrus than 5-HT2 antagonists ritanserin, ketanserin or amesergide. We thus have seen a dissociation in the capacity of compounds to alter the effects on behavior and IEG expression after dizocilpine administration.

Animals↗
PubMed1997

NMR studies of a viral protein that mimics the regulators of complement activation.

Vaccinia virus complement control protein (VCP) is a 243-residue protein that is similar in sequence to the regulators of complement activation; its role is to defend the virus against attack by the host complement system. A fragment of this protein spanning the two complement protein (CP)-modules (residues 126 to 243) which make up the C-terminal half of VCP has been expressed in Pichia pastoris. A 15N-labelled sample was purified for the purposes of structure determination and measurements of dynamics in solution using NMR. Structures were calculated on the basis of 1767 NMR-derived distance and angle restraints, with a longer than normal high-temperature simulated annealing (SA) protocol which improved convergence. The viral CP-modules are structurally very similar to the 15th and 16th CP-modules of human factor H (fH; average r.m.s.d., for invariant Trp and Cys, four pair-wise comparisons,=1.2 A) but less similar to the fifth CP-module of fH (average r.m.s.d.=2.2 A). In the VCP fragment, the orientation of one module with respect to the other is clearly defined by the experimental data, and T1 measurements are consistent with only limited flexibility at the module-module interface. The r.m.s.d. over all of the 118 residues (backbone atoms) is 0.73 A. The intermodular orientation is better defined than, and significantly different from, that observed in a CP-module pair from fH (re-calculated using the extended SA protocol). In VCP the long axis of the second module is tilted by 59(+/-4) degrees with respect to the first module (50(+/-13) degrees in the fH pair), and twisted with respect to the first module by 22(+/-6) degrees (223(+/-17) degrees in fH). The differences between the human and viral proteins may be rationalised in terms of the lack of hydrogen-bond stabilised secondary structure in the N-terminal portion of fH module 16, and the number and type of amino acid side-chains which make up the interface. A similar intermodular interface may be predicted between the third and fourth module of human C4 binding protein and, probably, between the third and fourth modules of the guinea pig acrosomal matrix protein 67; but the formulation of general rules for predicting the structure of interfaces between CP-modules awaits further experimental data.

Amino Acid Sequence↗
PubMed1997

Phenotypic switch in a Spanish HIV type 1 isolate on serial passage on MT-4 cells.

A biological clone (F0) of a syncytium-inducing (SI) isolate (S61) was unable to produce syncytia in MT-4 cells. On serial passage on MT-4 cells this virus [F15(-3)] became capable of inducing syncytia (Sánchez-Palomino S, et al.: J Virol 1993;67:2938). After sequencing different regions of the env gene including V1-V2, V3, and the fusion domain of both viruses, we have found only an asparagine (N)-to-isoleucine (I) change in position 7 of the V3 loop. By mutagenesis and in vitro recombination, using infectious molecular clones, we have identified this amino acid change as the only one responsible for the syncytial phenotypic switch. However, this cytopathic change was not accompanied by a change in the replication rate, indicating that these two properties are not linked genotypic traits. Thus serial passaging of an HIV-1 isolate on MT-4 cells has produced a nonsyncytial-to-syncytial switch through a point mutation in position 7 of the V3 loop.

Amino Acid Sequence↗
PubMed1997

Membrane binding and enzymatic activation of a Dbl homology domain require the neighboring pleckstrin homology domain.

Dbl-homology (DH) domains are invariably located immediately N-terminal to a pleckstrin homology (PH) domain. To understand the functional relationship between these two domains we expressed the DH domain alone, the PH domain alone, and the DH-PH combination of the invasion inducing protein Tiam-1 fused to glutathione-S-transferase (GST) or green fluorescent protein (GFP). We found that the GST-DH-PH and the GST-PH constructs bind to preparations of brain membranes and to the beta gamma subunits of trimeric G proteins in vitro, while the GST-DH and GST control do not. The GFP-DH-PH and GFP-PH constructs are localized to peripheral membranes of COS-7 cells in vivo, while GFP and GFP-DH domain constructs are found diffusely in the cytoplasm. The DH-PH domain combination activates Jun N-terminal kinase (JNK) strongly, but the DH domain alone and the PH domain alone have little effect. We conclude that membrane localization and enzymatic activation of the DH domain require the adjacent PH domain.

Animals↗
PubMed1997

Valency dependent patterns of binding of human L-selectin toward sialyl and sulfated oligosaccharides of Le(a) and Le(x) types: relevance to anti-adhesion therapeutics.

The human L-selectin is known to bind to immobilized 3'-sialyl-Le(x) and -Le(a) oligosaccharides both under static and physiological flow conditions. Here the reactivities toward 3'-sulfated and 3'-sialyl-Le(a) and -Le(x) pentasaccharides are compared by in-vitro binding and inhibition assays using preparations of human L-selectin-IgG-Fc chimera in which the selectin is predominantly in di- and tetrameric form (paucivalent) or in the form of a complex with anti-IgG (multivalent). Affinity for the sulfated ligands is marginally greater than for the sialyl ligands, as judged by concentrations required to give 50% inhibition of the multivalent selectin binding to the immobilized sulfated and sialyl ligands. There is a striking difference, however, in the avidities of binding of the two L-selectin forms toward the sulfated and sialyl ligands when these are immobilized in the clustered state: the paucivalent selectin gives detectable binding only to the sulfated ligands when these are immobilized as neoglycolipids on plastic microwells (up to 100 pmol immobilized per well) whereas the multivalent L-selectin binds well to both classes of ligand. Moreover, binding of the paucivalent selectin form is effectively inhibited only by the sulfated ligand, although binding of the multivalent selectin is inhibitable by both the sulfated and sialyl ligands. Such striking valency-dependent differences in ligand binding avidity and inhibitability may be manifest in vivo with the membrane-bound L-selectin, as marked variations occur in its density of expression on leukocytes. Thus, for the purpose of selecting inhibitors for development of therapeutic anti-inflammatory compounds, experimental designs based on the paucivalent L-selectin would more clearly single out compounds with broad spectrum anti-adhesive activities toward the both the high- and low-avidity interactions of the cell adhesion protein.

Animals↗