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Myometrium obtained from pregnant ewes (30-80 days gestation) contains a factor which inhibits phospholipase A2 (PLA2) activity. The activity of this moiety was assessed using an in vitro porcine pancreatic PLA2 assay system. Inhibitory activity was associated with a 35-45000 dalton molecular weight fraction, heat-labile, sensitive to protease degradation and did not partition into organic solvents. These data are indicative that PLA2-inhibitory activity resides in a protein moiety. Dixon-plot analysis of myometrial-inhibitory activity was indicative that the inhibition of PLA2 activity was of a non-competitive nature (Ki = 4.1 +/- 0.7 micrograms/ml, ca 118 nmol/l). Myometrial phospholipase-inhibitory protein(s) may be involved in the suppression of eicosanoid biosynthesis by the uterine tissues throughout gestation thus inhibiting uterine contractile activity.

The capacity of cotyledonary microsomes, prepared from pregnant ewes (20-145 days of gestation), to metabolize exogenous arachidonic acid was quantified using a radiolabel technique. During gestation, the capacity of microsomes to metabolize arachidonic acid increased 25-fold, from 0.36 +/- 0.06 mumol arachidonic acid/incubation (n = 8) at less than 100 days of gestation to 9.06 +/- 1.02 mumol arachidonic acid/incubation at 130-145 days of gestation (n = 5; P less than 0.05). Arachidonic acid was metabolized to prostaglandin E2 and F2 alpha, as determined by thin-layer chromatography and reverse-phase high performance liquid chromatography. The profile of prostaglandins synthesized by cotyledonary microsomes did not change throughout gestation. These data suggest that the increase in cotyledonary prostaglandin synthesis that occurs during late gestation and at term may reflect an increase in the tissue content of prostaglandin H2 synthase.

The authors examined the adhesion of seven human melanoma cell lines to cultured human umbilical vein endothelial cells (HEC) that were activated by cytokines or bacterial endotoxin. The adhesion of Hs 294T and MEL-24 cells was markedly increased (approximately 2 to 12-fold) after pretreatment of HEC monolayers for 6 hours with tumor necrosis factor, interleukin-1, or endotoxin. Smaller increases were noted with the cell lines RPMI 7951, HT 144, Malme-3M, MEL-2, and no significant increase was observed with MEL-5. Cytokine and endotoxin effects on melanoma-HEC adhesion were concentration- and time-dependent, with onset by 2 hours, peak at 6-8 hours and maintenance through 48 hours. Cytokine induction of increased HEC adhesiveness for melanoma cells was blocked by actinomycin-D or cycloheximide, suggesting the requirement for RNA and protein synthesis. Interaction of melanoma cells with subendothelial matrix did not appear to play a primary role because: 1) phase contrast and electron microscopy revealed direct contact between tumor cells and endothelial cells in standardized monolayer adhesion assays; 2) increased adhesion (rosette formation) of tumor cells to activated HEC was also observed after nonenzymatic resuspension of HEC, and 3) the matrix peptide GRGDSP partially blocked (approximately 45%) Hs 294T cell adhesion to subendothelial matrix, but had little or no effect on adhesion to activated HEC monolayers. Taken together, these data suggest that inducible HEC surface changes may mediate the adhesion of certain melanoma cells, thereby exerting an active influence over the metastatic process.

1. The effects, on parturition in the rat, of subacute and acute opioid administration were studied. Further experiments investigated the role of modulation of maternal plasma and pituitary oxytocin (OXY) and arginine vasopressin (AVP) levels in these effects. 2. Subacute opioid (M320, buprenorphine or bremazocine) administration prolonged the gestation of rats. This was accompanied by toxic effects on the offspring. Acute subcutaneous (s.c.) M320 (10 micrograms/kg) administration was accompanied by prolonged gestation without toxic effects. 3. Subacute M320 (10 micrograms/kg, s.c., twice daily) treatment was accompanied by increased interbirth intervals in parturient rats. 4. Maternal OXY but not AVP release, as assessed by measurement of plasma and pituitary immunoreactivity, was elevated during and up to 1 h after the completion of parturition. Subacute M320 treatment did not inhibit this elevated OXY release.

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.

The subcellular distribution and compartmentalization of choriomammotrophin (CM) and progesterone within ovine placentomes was investigated using differential and density gradient centrifugation techniques. Approximately 67% of placental CM and 45% of progesterone was associated with subcellular particles. The 10,000 g particulate fraction contained the highest specific activity of both CM and progesterone (19.1 +/- 3.8 (S.E.M.) micrograms/mg and 71.5 +/- 9.2 pmol/mg protein respectively). This fraction was also shown to contain electron-dense granules with morphology similar to that of hormone-containing secretory granules isolated from other endocrine tissues. Particle-associated CM sedimented to a density of 1.051-1.054 g/ml in colloidal silica gradients and displayed physicochemical characteristics consistent with its storage in secretory granules. During in-vitro incubations, particle-associated CM was stable for up to 90 min, but dissociated when incubated in hypoosmotic medium. Particulate progesterone, which was also present in the CM-rich fraction and was stable for up to 90 min of incubation, was not affected by decreasing the osmolality of the incubation medium. These data suggest that ovine CM (but not progesterone) is stored within a population of secretory granules located within placentomes.

This paper describes an exploratory multivariate analysis designed to determine the relative contributions of age, verbal ability, education, reading habits, and recall strategies to the explanation of variation in performance on prose recall tasks among adults. Four hundred twenty-two adults in three age groups--young (18 to 32), middle (40 to 54) and old (62 to 80)--read and recalled in writing two 388-word prose passages and answered questions about their background, reading habits, and recall strategies. Results indicate that a decrease in quantity of recall appears with increasing age, though verbal ability is a better predictor of recall than is age. In addition, a recall strategy factor representing a paragraph-by-paragraph retrieval strategy produces the highest simple correlations with total recall and contributes significantly to the other recall measures.

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.

The effects of organizational variables in prose on recall are examined for college-educated adults in three age groups. If older adults suffer a deficit in organizational processes [3], this may be manifest in lower in lower quantities of prose recall, difficulty in identifying and following the text's structure and main ideas, and diminished "levels effects" (information high in the hierarchical text structure recalled better than information low). The study reported tests these implications using Meyer's [15] prose analysis system to identify text structure. No age differences were found in total recall and recall of main ideas. However, young adults exhibited the typical " levels effects," while middle and old adults did not. This difference was attributed to the effects of current schooling practices on the youngest group, rater than organizational or reading comprehension deficits in the aged.

The involvement of arachidonic acid and arachidonic acid metabolites in the control of oxytocin secretion by ovine corpus luteum was investigated, using slices of luteal tissue incubated in vitro. Oxytocin was secreted at steady rates by luteal slices, during 60-min incubations (315.0 +/- 45.3 pg/mg.h). The secretion of oxytocin was stimulated by arachidonic acid, phospholipase A2 (PLA2), and phospholipase C (PLC) in a dose-dependent manner. The highest doses of arachidonic acid, PLA2, and PLC used stimulated oxytocin secretion by 145.8 +/- 23.0% (P less than 0.01; n = 6), 331.5 +/- 42.4% (P less than 0.02; n = 4), and 955.5 +/- 278.6% (P less than 0.01; n = 4), respectively. Oxytocin secretion by luteal slices was not affected by either prostaglandin F2 alpha (PGF2 alpha) or PGE2 over a concentration range from 3-3000 nM. Furthermore, inhibitors of the cyclo-oxygenase pathway of arachidonic acid metabolism did not consistently affect arachidonic acid and PLA2-stimulated oxytocin secretion. Nordihydroguaiaretic acid, which inhibits 5-lipoxygenase, however, totally abolished arachidonic acid- and reduced PLA2-stimulated oxytocin secretion. The presence of CoCl2 in the incubation medium also significantly reduced basal and PLA2- and PLC-stimulated oxytocin secretion [P less than 0.05 (n = 5), P less than 0.05 (n = 5), and P less than 0.01 (n = 6), respectively]. We have shown that oxytocin secretion from slices of ovine corpus luteum incubated in vitro is stimulated by exogenous and endogenously released arachidonic acid. The data show that PGF2 alpha and PGE2 do not have a role in luteal oxytocin secretion in vitro and PG formation does not appear to be involved in the stimulation of oxytocin secretion elicited by arachidonic acid or PLA2. Arachidonic acid may have its effect via the lipoxygenase pathway.

1. In this study, we examined the basal release of choriomammotropin (oCM) from monolayer cultures of cotyledonary cells obtained from ewes at different gestational ages. 2. oCM release increased with gestational age and displayed a similar profile to the concentration of oCM observed in maternal plasma. 3. Release of oCM was significantly (P less than 0.05; n = 9) increased in calcium-depleted medium, and by treatment with either phospholipase C (0.125 units/ml) or KCl (50 mM). 4. The calcium antagonist MgCl (12 mM) and the calcium channel-blocking agents verapamil (50 microM) and nefidipine (10 microM) all significantly stimulated oCM release. 5. These data are consistent with the suggestion that oCM release is inversely related to extracellular calcium concentration.

Hematogenous metastasis requires the arrest and extravasation of blood-borne tumor cells, possibly involving direct adhesive interactions with vascular endothelium. Cytokine activation of cultured human endothelium increases adhesion of melanoma and carcinoma cell lines. An inducible 110-kD endothelial cell surface glycoprotein, designated INCAM-110, appears to mediate adhesion of melanoma cells. In addition, an inducible endothelial receptor for neutrophils, ELAM-1, supports the adhesion of a human colon carcinoma cell line. Thus, activation of vascular endothelium in vivo that results in increased expression of INCAM-110 and ELAM-1 may promote tumor cell adhesion and affect the incidence and distribution of metastases.

1. The effects of mu- and K-opiate receptor agonists on plasma concentrations of immunoreactive (ir) oxytocin (OXY) and arginine vasopressin (AVP) in normally hydrated and water-deprived rats were studied. 2. Water-deprivation for 36 h elevated both plasma ir-OXY and ir-AVP concentrations of Long-Evans rats. These elevated levels were lowered in a dose-dependent manner after subcutaneous administration of bremazocine (3-10 micrograms/kg), M320 (10-100 micrograms/kg) or morphine (0.1-10 mg/kg). Comparable reductions of plasma concentrations of ir-AVP and ir-OXY were observed. 3. Plasma concentrations of ir-OXY and ir-AVP of normally hydrated Long-Evans rats were lower after subcutaneous administration of bremazocine (10 micrograms/kg), M320 (10 micrograms/kg) and morphine (1.0 mg/kg). 4. These data suggest that both mu- and K-opiate receptor agonists inhibit both OXY and AVP release from the neurohypophysis of rats.

Phospholipid metabolites have previously been implicated in receptor-mediated stimulation of protein hormone secretion. As the factors which regulate the release of choriomammotrophin remain to be elucidated, we investigated the potential involvement of phospholipase C-induced phospholipid metabolism in the release of this placental hormone. Phospholipase C (PLC) caused a dose-dependent release of choriomammotrophin from ovine placenta, incubated in vitro. At a concentration of 0.2 units/ml (0.25 microgram protein/ml), PLC caused the release of choriomammotrophin from placental tissue to approximately double that observed in control incubations (7.08 +/- 0.4 micrograms/50 mg/h and 3.26 +/- 0.3 micrograms/50 mg/h, respectively). PLC treatment did not significantly alter plasma membrane permeability, as indicated by the release of lactate dehydrogenase and protein. PLC-stimulated release of oCM was completely abolished by incubation in calcium-free medium or by preincubation with the inorganic calcium-channel blocking agents cobalt chloride (4 mM) and lanthanum chloride (1 mM). The effects of PLC treatment on ovine choriomammotrophin (oCM) release were also inhibited by preincubation of placental tissue with inhibitors of arachidonic acid metabolism: ibuprofen (10(-5) M), naproxen (10(-4) M) or nordihydroguaiaretic acid (NDGA) 5 X 10(-6) M). These results suggest that the effects of PLC on the release of choriomammotrophin are mediated via metabolites of arachidonic acid.

The subcellular localization of oxytocin within the corpus luteum of sheep was investigated using differential and density gradient centrifugation. Oxytocin was associated with a particulate fraction which sedimented to a density of 1.054-1.061 g/mL. The exclusion of [3H]oxytocin from this particulate fraction is indicative that particulate oxytocin represents endogenous compartmentalization. Particulate oxytocin, incubated in buffered medium at 37 degrees C, was stable for up to 1 h and the release of oxytocin was not affected by the pH of the incubation medium, over the range 5.5-8.5. Oxytocin release, however, was stimulated by incubating particle-bound oxytocin in buffered medium of low osmolality (less than 200 mosmol). These data are similar to the physicochemical properties reported for peptide-containing neurohypophysial secretory granules. Ultrastructural analysis of oxytocin-containing fractions revealed the presence of electron-dense granules (diameter, 200-250 nm). These data are suggestive that oxytocin, in the corpus luteum of sheep, is contained within a population of secretory granules which occur in high numbers during the midluteal phase of the oestrous cycle.

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.

1. Administration of 10 micrograms kg-1 of the long lasting potent kappa- and weaker mu-opioid agonist N-(cyclopropylmethyl)-19-isopentylnororvinol (M320) twice daily from day 20 of gestation prolonged the internal gestation period of the rat and retarded the development of the offspring in the perinatal period. 2. The capacities of myometrial, placental and cervical tissues to produce prostaglandin E2 (PGE2) were not affected by M320 treatment. 3. During the period in which parturition normally occurred in saline-treated rats, foetal pituitary levels of immunoreactive oxytocin (ir-OXY) but not immunoreactive arginine-vasopressin (ir-AVP) were greater in M320-compared to saline-treated animals. Following the completion of parturition, foetal pituitary ir-OXY and ir-AVP levels continued to rise in saline-treated rats, but fell dramatically in rats treated subacutely with M320. 4. These data indicate that subacute treatment with M320 may inhibit foetal oxytocin release at term. This foetal OXY release may be a stimulus for the initiation of labour.

The osmolality and concentrations of Na, K, Cl and the hormones arginine vasotocin (AVT), prolactin, aldosterone and corticosterone were measured in plasma as functions of time in relation to oviposition, changing NaCl content of the diet, and feeding-inanition. AVT was significantly increased immediately after oviposition (but not during the hour before) with a calculated average value of 38.0 +/- 4.1 pg/ml at oviposition. A moderate increase in concentrations of prolactin and corticosterone were observed immediately after oviposition. Oviposition was not associated with detectable changes in plasma osmolality (and electrolyte concentrations) nor with the concentration of aldosterone. After a sudden change from a high NaCl diet to a low NaCl diet the plasma osmolality and concentrations of NaCl, AVT and prolactin reached new stable levels in 24 hr, whereas the plasma aldosterone concentration required more than 4 days to reach a steady level. After resalination plasma aldosterone was suppressed in less than 8 hr. Both osmolality and concentrations of AVT and prolactin showed transient overshoots during the first 24 hr. NaCl depletion resulted in a transient increase of corticosterone.

Luteal oxytocin-containing secretory granules have been isolated and characterized in terms of their physicochemical parameters. The isopynic sedimentation density (1.03 +/- 0.003 g/ml) and sedimentation coefficient (1670 S, 0.32 mol sucrose/l, 4 degrees C) of these granules have been estimated. Based upon these estimates, the average vesicle diameter (258 +/- 17 nm) and vesicle weight (9.92 +/- 0.67 fg/vesicle) were calculated. The exchangeable water content (58.2%) of these granules was determined using density gradients prepared with deuterium oxide. Luteal oxytocin-containing granules displayed similar physicochemical characteristics to those reported for neurohypophysial peptide-containing granules, with the exception of particle size. Luteal granules were 1.3 times greater in diameter than neurohypophysial granules.