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Biomedical subjects

G D Thorburn

Publications and source records attributed to G D Thorburn.

At least 73 records · Page 4Linked to original sources

Peripheral and brain tissue catecholamine content in intact and anti-NGF-treated fetal sheep.

Fetal lambs were treated with a single dose of anti-mouse nerve growth factor (anti-NGF) at 80 days gestational age. The catecholamine content of tissues was determined at 135 days gestational age. There was a reduction of either norepinephrine, epinephrine, or both, in the thymus, thyroid, atrium (but not ventricle), lung, liver, kidney, and jejunum when compared with age-matched control fetuses. The spleen, ileum, colon, and the adrenal glands were not affected by anti-NGF. In treated fetuses there was a reduction in catecholamine content of the thalamus, hypothalamus, hippocampus, medulla, cerebellum, and cervical spinal cord. These results show that some tissues are sensitive to, and some are refractory to, the action of anti-NGF at 80 days gestation. Also the results suggest that NGF may play a role in the development of catecholamine-containing neurons within the central nervous system.

Animals↗

Effect of hypophysectomy with and without thyroxine replacement on growth and circulating concentrations of insulin-like growth factors I and II in the fetal lamb.

The role of the pituitary gland in the regulation of skeletal growth and plasma insulin-like growth factor (IGF)-I and IGF-II concentrations in the late gestation sheep fetus has been studied. Singleton fetuses were either hypophysectomized (n = 14) or sham operated (n = 8) between days 110 and 125. Fetal and maternal blood samples were collected three times weekly through the remainder of gestation. In some hypophysectomized fetuses (HXT4 group, n = 4), T4 was administered (100 micrograms L-T4/day) to overcome the hypothyroidism caused by hypophysectomy. The other hypophysectomized fetuses received no replacement therapy (HXNR group, n = 10). Six HXNR fetuses were allowed to remain in utero post term and were killed at day 163 of gestation. All other animals were killed at day 147. All values are group means +/- SE. Hypophysectomized fetuses had significantly shorter limbs and long bones and delayed osseous maturation at term compared to sham controls. Plasma free T4 concentrations in HXT4 fetuses were not significantly different from those measured in sham fetuses (P greater than 0.05). Bone maturation at term was normal in HXT4 fetuses although there was no improvement in limb or bone elongation. Retention of hypophysectomized fetuses in utero until 16 days past term yielded fetuses which were heavier than controls but whose limb and bone lengths were no greater than hypophysectomized fetuses killed at term. Osseous maturation was appropriate for term in five of the six postterm hypophysectomized fetuses. The plasma IGF-I and IGF-II concentrations were not significantly affected by hypophysectomy, hypophysectomy with T4 replacement, gestational age, or prolonged gestation. The plasma IGF-I concentrations in the sham, HXNR, and HXT4 fetuses were 35.4 +/- 6.6, 28.2 +/- 3.0, and 34.4 +/- 1.7 ngeq human (h)IGF-I/ml, and the IGF-II concentrations were 656.3 +/- 59.2, 635.3 +/- 56.3, and 645.5 +/- 71.9 ngeq hIGF-II/ml, respectively, and remained within these ranges throughout the experiment. Fetal IGF-I concentrations were significantly lower than mean maternal IGF-I concentrations (88.0 +/- 6.8 ngeq hIGF-I/ml, P less than 0.05), and fetal IGF-II concentrations were significantly higher than mean maternal IGF-II concentrations (362.4 +/- 24.0 ngeq hIGF-II/ml, P less than 0.05). We conclude that in the late gestation fetal sheep, elongation of the appendicular skeleton is under some direct pituitary control whereas appendicular maturation exhibits some dependence on circulating T4 concentrations.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗

Fetal sheep serum contains a high molecular weight insulin-like growth factor (IGF) binding protein that is acid stable and specific for IGF-II.

We have used radiolabeled ovine insulin-like growth factor II (oIGF-II) and human IGF-I (hIGF-I) to investigate the nature of the IGF binding proteins in fetal sheep serum. Incubation of fetal sheep serum with [125I]oIGF-II, followed by chromatography on Fractogel TSK HW55(S), revealed the presence of two major binding protein species, a lower molecular weight binding protein (apparent mol wt approximately 60,000) and a much higher molecular weight binding protein (apparent mol wt approximately 500,000). Only the lower molecular weight binding protein complex was seen in serum from adult nonpregnant sheep. The lower molecular weight binding protein in fetal and adult sheep serum bound both [125I]oIGF-II and [125I]hIGF-I, both of which could be displaced by unlabeled oIGF-II or hIGF-I. However, the very high molecular weight binding protein bound only [125I]oIGF-II, and this could only be displaced by unlabeled oIGF-II. The very high molecular weight binding protein appears to bind approximately 40% of the endogenous oIGF-II. We have purified the very high molecular weight binding protein from fetal sheep serum using anion exchange, Concanavalin A Sepharose, and hydrophobic interaction chromatography. The purified binding protein preparation did not contain any lower molecular weight binding protein and did not bind [125I]hIGF-I. In addition, this binding protein was stable at pH 3.2 for 1 h. Thus, fetal sheep serum contains a very high molecular weight IGF binding glycoprotein that is acid stable and specific for IGF-II.

Animals↗

Prostaglandin F2 alpha and oxytocin release during persistence of the corpus luteum in sheep.

Oxytocin (OT), progesterone and prostaglandin F2 alpha (PGF2 alpha) concentrations were measured in the utero-ovarian vein (UOV) of ewes which displayed persistence of the corpus luteum (CL). During the period of expected luteolysis, the frequency of OT and PGF2 alpha pulses in the UOV was significantly (P less than 0.005 for both) lower in ewes with persistent CLs, compared with ewes that underwent normal luteal regression. In contrast, the amplitude of both OT and PGF2 alpha pulses was similar in both groups of animals. It is suggested that persistence of the CL resulted from a decreased PGF2 alpha pulse frequency, which may have arisen from a decreased frequency of stimulation by OT. In two persistent CL ewes, however, it appeared that a failure at the level of the uterus may have contributed to the observed decrease in PGF2 alpha release. Although a PGF2 alpha analogue (Lutalyse) infusion into the uterine vein of two ewes with persistent CLs failed to induced luteolysis, it did stimulate a large release of OT into the UOV. This suggests that persistent CLs maybe more resistant to PGF2 alpha and, that at day 22 post-oestrus, these CLs are capable of releasing large quantities of OT into the UOV.

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Peripheral plasma concentrations of pregnenolone sulphate, pregnenolone, progesterone and 20 alpha-hydroxy-4-pregnen-3-one in ewes throughout the oestrous cycle.

Pregnenolone sulphate, pregnenolone, progesterone and 20 alpha-hydroxy-4-pregnen-3-one concentrations in peripheral plasma of normal cyclic ewes were measured by radioimmunoassay. The concentrations of these steroids were correlated with that of progesterone. The concentrations of all the steroids measured in peripheral plasma varied in a cyclic manner and showed a significant (P less than 0.05) positive correlation with the concentration of progesterone. Peripheral plasma concentrations of these steroids in ovariectomized and ovariectomized, dexamethasone-treated ewes were also determined. The plasma concentration of progesterone in ovariectomized ewes was undetectable but the concentrations of pregnenolone sulphate, pregnenolone and 20 alpha-hydroxy-4-pregnen-3-one remained similar to those observed at oestrus. Administration of dexamethasone to ovariectomized ewes had no effect on pregnenolone sulphate or pregnenolone concentrations but 20 alpha-hydroxy-4-pregnen-3-one concentrations, which were already very low, decreased further. It is proposed that the ovary, probably the corpus luteum, secretes pregnenolone sulphate, pregnenolone and 20 alpha-hydroxy-4-pregnen-3-one; however, pregnenolone sulphate and 20 alpha-hydroxy-4-pregnen-3-one may also arise from the metabolism of circulating pregnenolone and progesterone.

20-alpha-Dihydroprogesterone↗

Diurnal variations in plasma concentrations of cortisol, prolactin, growth hormone and glucose in the fetal sheep and pregnant ewe during late gestation.

We have measured fetal and maternal plasma concentrations of cortisol, prolactin, GH and glucose in samples collected during a 24-h period in 14 animals between 127 and 142 days of gestation. There was a significant increase in both the mean daily plasma cortisol concentration and mean daily coefficient of variation (C.V.) of plasma cortisol concentrations after 135 days of gestation. There was also a significant variation in the fetal plasma cortisol concentrations with a peak occurring at 19.00 h. There was a significant sinusoidal diurnal rhythm in the plasma prolactin concentrations in both the fetal sheep and pregnant ewe and the maximal prolactin concentrations occurred between 19.00 and 23.00 h (fetal) and 21.00 and 01.00 h (maternal). Although no significant diurnal variation was detected in fetal plasma GH concentrations, there was a significant sinusoidal diurnal rhythm in the plasma GH concentrations of the pregnant ewe and the maximal maternal GH concentrations occurred between 21.00 and 01.00 h. Both the fetal and maternal plasma glucose concentrations showed a significant sinusoidal diurnal rhythm. The maximal maternal and fetal glucose concentrations were measured between 21.00 and 01.00 h and between 23.00 and 03.00 h respectively. We have therefore established that diurnal variations in plasma cortisol and prolactin concentrations exist prenatally. Whether the presence of such hormonal rhythms reflects the activity of an endogenous fetal circadian pacemaker remains to be established.

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Ovine allantoic fluid inhibition of prostaglandin synthesis in cotyledonary microsomes.

Inhibition of microsomal prostaglandin (PG) biosynthesis by allantoic fluid, obtained from ewes at 80-120 days of gestation, was examined. Inhibition of cotyledonary microsomal PGE2 and PGF2 alpha biosynthesis by lyophilized allantoic fluid occurred in a dose-dependent manner. The concentration of allantoic fluid required to inhibit PGE2 and PGF2 alpha production by 50% averaged 17.9 +/- 3.2 (S.E.M.) mg dry weight/ml (n = 5). Microsomal PG biosynthesis was markedly enhanced by the addition of arachidonic acid (30 mumol/l). Synthesis of PGE2 and PGF2 alpha was increased to 245 +/- 65% and 184 +/- 14% of control (P less than 0.05, n = 5) respectively. Treatment of cotyledonary microsomes with porcine phospholipase A2 (PLA2; 0.125 units/ml) also stimulated PG synthesis, PGE2 increasing to 216 +/- 27% and PGF2 alpha to 172 +/- 14% of control (P less than 0.05, n = 5) respectively. Allantoic fluid (20 mg dry weight/ml) inhibited arachidonic acid-stimulated PG synthesis (PGE2 by 48.6 +/- 13.8% and PGF2 alpha by 44.2 +/- 7.7%) and PLA2-stimulated PG synthesis (PGE2 by 60.6 +/- 11.6% and PGF2 alpha by 74.8 +/- 8.5%). Allantoic fluid, however, did not affect PLA2-stimulated release of arachidonic acid from microsomes, thus negating the possibility that allantoic fluid suppresses PG synthesis by inhibiting PLA2 activity. These data indicate that allantoic fluid inhibits PG production at the level of PG synthase enzymes. Prostaglandin inhibitor(s) in allantoic fluid may play a role in maintaining uterine quiescence throughout gestation and its withdrawal, at term, may be involved in the initiation of labour.

Allantois↗

Passive immunization of pregnant goats against ovine LH.

Groups of three goats at 50, 90 and 130 days of gestation were passively immunized against ovine LH (oLH) by i.v. infusion of 8 ml serum equivalent of the immunoglobulin fraction of rabbit anti-oLH serum (LHAS). Goats at the same stages of gestation as above served as controls and received 8 ml serum equivalent of the immunoglobulin fraction of normal rabbit serum (NRS). Plasma concentrations of progesterone were determined by specific radioimmunoassay of blood collected at 20-min intervals from 6 h before infusion of LHAS or NRS to 12 h after infusion. Less frequent sampling was performed from 2 days before to 6 days after infusion. Plasma from all LHAS-immunized goats exhibited binding of oLH. Twelve hours after immunization, titres ranged from 1:135 to 1:215. All LHAS-treated goats had titres of less than 1:10 by 5 days after immunization, but a low level of oLH binding was still detectable. Treatment with LHAS or NRS did not shorten the length of gestation, with all goats delivering live offspring between 142 and 147 days after conception. Plasma concentrations of LH ranged from less than 0.15 micrograms/l to 4.8 micrograms/l and were greater than 0.15 micrograms/l in 181 of 255 samples (71%) for both the NRS-treated group, throughout the experiment, and the LHAS-treated groups before infusion of antiserum. Luteinizing hormone was not detectable in plasma samples obtained after LHAS infusion in goats at 50 or 130 days of pregnancy. Plasma concentrations of LH exceeded 0.15 micrograms/l in only five of 51 (10%) samples in 90-day-pregnant goats treated with LHAS, the maximum value reached being 0.80 micrograms/l.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

A comparison of the effects of electroimmobilisation and, or, shearing procedures on ovine plasma concentrations of beta-endorphin/beta-lipoprotein and cortisol.

The effects of electroimmobilisation (EIM) and shearing procedures on the plasma concentrations of beta-endorphin/beta-lipotrophin (beta-EP/beta-LPH) and cortisol were compared in conscious sheep. Serial blood samples were collected from five groups of animals subjected to either EIM during simulated shearing (EIM + SS), SS, EIM, shearing or control procedures. EIM and shearing each caused significant increases in the plasma concentrations of beta-EP/beta-LPH and cortisol. The responses to EIM and shearing were significantly greater than in the control animals but were not significantly different from each other. There were no significant differences in the mean plasma concentrations of cortisol following treatment between EIM and EIM + SS groups or between SS and control groups. However, the plasma cortisol concentrations after treatment in the EIM + SS group were significantly greater than in the SS group.

Animals↗

Lack of production of thromboxane A2 by the sheep heart following coronary occlusion.

Blood pressure, heart rate and ECG were monitored continuously and coronary venous blood samples assayed for thromboxane B2. Arterio-coronary venous lactic acid differences were estimated. In anaesthetized sheep, 60 sec occlusions of the left circumflex coronary artery caused hypotension with little change in heart rate. Five minute occlusions produced similar changes accompanied by cardiac arrhythmias and a larger fall in blood pressure. In conscious sheep heart rate rose and blood pressure was maintained. ECG and lactic acid changes indicated severe myocardial ischaemia but not change in thromboxane release was detected. The apparent lack of involvement of thromboxane may reflect a relative inability of sheep platelets to produce this substance or a failure to provide an adequate stimulus for synthesis.

Anesthesia↗

Myocardial ischaemia following electrical stimulation of the left circumflex coronary artery in sheep: a role for thromboxane A2?

The electrical stimulation model of thrombus formation was tested on rabbit carotid artery and adapted to sheep left circumflex coronary artery (LCCA). LCCA blood flow, mean arterial pressure (MAP), heart rate (HR) and ECG were monitored continuously and arterial and coronary venous blood samples were taken for radioimmunoassay of thromboxane B2. Stimulation of the LCCA mimicked acute myocardial infarction; reduction in LCCA blood flow preceded a fall in MAP and appearance of ECG abnormalities. Thromboxane B2 levels rose by 126% 35 min after stimulation. These findings support the proposal by other authors that thromboxane plays an important role in the pathogenesis of acute myocardial infarction.

Anesthesia↗

The effects of certain vasodilating prostaglandins on the coronary and hindlimb vascular beds of the conscious sheep.

Vasodilating prostaglandins were injected, in bolus doses, into the lower abdominal aorta or left circumflex coronary artery (LCCA) of conscious sheep. Local blood flow, mean arterial pressure (MAP), heart rate (HR) and ECG were monitored continuously. 6-Keto PGF1 alpha had no effect on either vascular bed in doses up to 100 micrograms. PGE2 was more potent than PGI2 in dilating hindlimb vasculature and PGE2 induced a more persistent hyperaemia whereas PGD2 elicited a biphasic response (constriction-dilation). PGE1, PGE2, PGD2 and PGI2 all produced dose-dependent vasodilation, the order of potency being PGD2 greater than PGI2 greater than PGE1 greater than or equal to PGE2. The effect of PGI2 was more transient and PGE1 and PGD2 caused small but consistent decreases in MAP and HR, respectively.

Animals↗

The effects of vasoconstricting prostanoids on the coronary and hindlimb vascular beds of the conscious sheep.

Vasoconstricting prostaglandins were injected, in bolus doses, into the lower abdominal aorta on the left circumflex coronary artery (LCCA) of conscious sheep. Local blood flow, mean arterial pressure (MAP), heart rate (HR) and ECG were continuously monitored. Thromboxane B2 had no effect on either vascular bed in doses up to 100 micrograms. PGF2 alpha produced mild vasoconstriction in both vascular beds with no systemic response. The endoperoxide analogues, U-44069 and U-46619, produced complex responses in both vascular beds. Initial vasodilation was followed rapidly by prolonged vasoconstriction. In the coronary circulation, vasoconstriction was temporarily masked by a hyperaemic phase. The U-compounds also affected MAP, possibly as a result of pulmonary and systemic vasoconstriction.

Animals↗

Secretion of oxytocin and progesterone by ovine corpora lutea in vitro.

The mechanisms involved in the control of oxytocin and progesterone secretion by the ovine corpus luteum have been investigated in vitro using luteal slice incubations. Oxytocin and progesterone were secreted at constant rates from luteal slices for 2 h of incubation (366 +/- 60 pg X mg X h and 18.9 +/- 0.18 ng X mg X h, respectively). Secretion of progesterone, but not of oxytocin, was significantly (p less than 0.02) stimulated in the presence of ovine luteinizing hormone. Incubation of luteal slices in medium containing 100 mM potassium, however, resulted in increased secretion of oxytocin and, to a lesser extent, of progesterone (294 +/- 59% and 142 +/- 15%, respectively, p less than 0.05). Basal oxytocin secretion was reduced during incubation in calcium-free medium, compared to secretion in the presence of calcium (70 +/- 15 and 175 +/- 25 pg X mg X 20 min, respectively, p less than 0.01), whereas progesterone secretion was not altered in the absence of calcium. Secretion of both hormones by luteal slices was stimulated by the addition of the calcium ionophore A23187 (p less than 0.05). Addition of prostaglandin F2 alpha (2.8 microM) had no effect on secretion of either oxytocin or progesterone. We have demonstrated that oxytocin and progesterone can be stimulated, independently, from corpus luteum slices incubated in vitro. The pattern of release is consistent with the proposal that oxytocin, but not progesterone, is associated with and actively released from luteal secretory granules. Our results also indicated that prostaglandin F2 alpha does not directly stimulate release of oxytocin or progesterone from luteal cells in vitro.

Animals↗

Effect of hypoxemia on plasma catecholamines in intact and immunosympathectomized fetal lambs.

Fetal lambs treated with sheep anti-mouse nerve growth factor antibodies (anti-NGF) at 80 days gestation subsequently showed a diminished cardiovascular response to intravenous infusion of tyramine (1 mg/min over 10 min) and no significant change in plasma norepinephrine concentrations as measured by reversed-phase high-pressure liquid chromatography and electrochemical detection. At autopsy at 135 days gestation, catecholamine content of the heart, thyroid, kidney, and ileum was reduced by greater than 70% compared with age-matched control fetuses. The anti-NGF-treated fetuses were thus judged to have impaired sympathetic function. When made hypoxemic (arterial PO2 12-14 Torr) for 1 h at 120 or 130 days gestation, anti-NGF-treated fetuses showed no significant change of arterial pressure, heart rate, or plasma catecholamines, whereas control fetuses showed bradycardia, a 28% increase in arterial pressure, and a 670% increase of plasma norepinephrine concentrations. These results suggest that the increase of arterial pressure that occurs during hypoxemia in fetal lambs between 120 and 135 days gestation is attributable to increased activity of the peripheral sympathetic nervous system. The adrenal medulla appears to contribute to the plasma catecholamine pool at rest, but the lack of increased plasma catecholamines during hypoxia after anti-NGF implies that the adrenal medulla was unable to release catecholamines. Possible reasons for this are discussed.

Animals↗

Oxytocin, oxytocin-associated neurophysin, and prostaglandin F2 alpha concentrations in the utero-ovarian vein of pregnant and nonpregnant sheep.

Oxytocin, oxytocin-associated neurophysin (neurophysin), prostaglandin F2 alpha (PGF2 alpha), and progesterone concentrations were measured in the utero-ovarian vein (UOV) of sheep during the estrous cycle and early pregnancy. On days 13-16 of the cycle, large pulses of PGF2 alpha, oxytocin, and neurophysin were measured in samples collected at hourly intervals from the UOV draining a corpus luteum (UOV/CL). Most of the PGF2 alpha pulses (96.5%) coincided with a pulse of both oxytocin and neurophysin, whereas only 55.6% of oxytocin pulses coincided with a pulse of PGF2 alpha. Therefore, during luteolysis in sheep, uterine PGF2 alpha release is closely associated with ovarian oxytocin release, and oxytocin release is unlikely to be dependent upon a uterine PGF2 alpha stimulus. During frequent sampling, coincident oxytocin pulses were measured in 1) both UOVs when a CL was present in both ovaries and 2) the jugular vein, carotid artery, and UOV/CL, with a significantly higher oxytocin pulse concentration occurring in jugular venous compared with carotid arterial plasma. Pituitary and luteal release of oxytocin may, therefore, occur simultaneously and be controlled by a circulating factor in sheep. Compared to days 13-16 of the cycle, significantly (P less than 0.001) fewer pulses of PGF2 alpha, which were significantly (P less than 0.001) smaller in amplitude, were measured in UOV samples collected frequently during early pregnancy. The frequency of oxytocin pulses observed in the UOV/CL of pregnant sheep was not significantly (P greater than 0.1) different from that observed in cyclic ewes, although most (86.4%) oxytocin pulses occurred in the absence of a PGF2 alpha pulse. In contrast, when a pulse of PGF2 alpha was observed in the UOV/CL of pregnant ewes, it usually coincided with a pulse of oxytocin. The suppression of uterine PGF2 alpha release during early pregnancy is not considered to result from a lack of stimulation by oxytocin.

Animals↗

Cannulation of the utero-ovarian vein in intact ewes: hormone concentrations and blood gas levels during the oestrous cycle and early pregnancy.

A technique for implanting and maintaining catheters in both utero-ovarian veins (UOV) of intact, conscious sheep has been developed. Both UOV catheters remained patent for more than 2 weeks in 72.7% of animals. In non-pregnant ewes, large pulses of PGF2 alpha were observed in UOV plasma collected on days 13 to 16 of the cycle (frequency 1.3 +/- 0.2/12 h; pulse height range 1.0-26.9 ng/ml). In contrast, on days 13 to 16 of gestation, significantly (P less than 0.001) fewer pulses of PGF2 alpha (frequency 0.42 +/- 0.2/12 h) were observed in both UOVs of pregnant ewes, which were significantly (P less than 0.001) smaller in amplitude (pulse height range 1.4-4.9 ng/ml). The maintenance of the corpus luteum (CL) during early pregnancy may therefore directly result from a decreased uterine PGF2 alpha release. During the luteal phase (day 4-12), the PO2 of UOV blood draining an ovary containing a CL (UOV/CL) was significantly higher than blood in the contralateral UOV (P less than 0.001). By days 15 and 16 of the cycle, the PO2 of UOV/CL blood had decreased and was not significantly different to that observed in the contralateral UOV (P greater than 0.1). On day 17, the PO2 of blood in both UOVs was elevated significantly compared to levels observed on the two preceding and the two following days (P less than 0.001). We consider that the higher PO2 in UOV/CL blood resulted from a very high luteal blood flow, and that the decrease on days 15 and 16 of the cycle resulted from a decline in luteal blood flow during luteolysis.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

Comparison of particle-associated progesterone and oxytocin in the ovine corpus luteum.

The subcellular distribution of progesterone and oxytocin within the ovine corpus luteum was investigated using differential and density gradient centrifugation. Progesterone and oxytocin were associated with particles which sedimented to a density of 1.049-1.054 g/ml and 1.054-1.061 g/ml respectively. Particle-associated progesterone did not, however, display physical or biochemical characteristics consistent with its storage within secretory granules. When particle-associated progesterone was incubated in HEPES buffer at 37 degrees C, 70% of the total progesterone was recovered in the incubation medium. The remaining stable particle-associated progesterone was not affected by treatments which stimulated oxytocin release and which have been shown to cause the release of peptides and biogenic amines from secretory granules. These results suggest that particle-associated progesterone represents the intercalation of progesterone into cell membranes and they do not support the hypothesis that progesterone is stored, in a protein-bound form, in luteal secretory granules.

Animals↗