Cocaine alters placental production of thromboxane and prostacyclin.

OBJECTIVE: The objective of this study was to test the hypothesis that cocaine alters placental prostaglandin production in vitro. STUDY DESIGN: Placentas were obtained from healthy women (n = 6) after normal vaginal delivery at term. Placental explants (300 mg) were incubated in duplicate at 37 degrees C in the presence of 0, 30, 300, or 3000 ng/ml cocaine. Thromboxane and prostacyclin production was measured by radioimmunoassay of their stable metabolites (thromboxane B2 and 6-keto-prostaglandin F1 alpha) at 0, 0.5, 1.0, 1.5, 2, 4, 8, and 12 hours. Analysis of variance with Newman-Keuls test was used for statistical analysis. RESULTS: Cocaine increased thromboxane production in a dose-dependent manner (p < 0.001) and decreased prostacyclin production (p < 0.05). Cocaine increased the ratio of thromboxane/prostacyclin production (p < 0.05). CONCLUSION: Cocaine alters the placental production of prostaglandins in vitro, favoring thromboxane production, which may cause vasoconstriction and decrease uteroplacental blood flow.

Streptozocin-induced diabetes mellitus in the pregnant ewe.

To evaluate the effects of streptozocin on maternal pancreatic beta-cell function, we administered the agent to 14 pregnant ewes at 85 to 90 days' gestation on two occasions, 4 days apart. Intravenous glucose tolerance tests were performed before the initial administration, before the second dose, and 4 weeks after the final dose of streptozocin. There was a significant elevation in maternal fasting blood glucose (82 +/- 8.1 mg/dl before streptozocin and 102.6 +/- 6.8 mg/dl after streptozocin, p less than 0.05). Five late-gestation ewes were used as controls, and a significant elevation in fasting plasma glucose levels was found in the streptozocin-treated animals (71.4 +/- 7.1 mg/dl control vs 102.6 +/- 6.8 mg/dl after streptozocin, p less than 0.05). The glucose tolerance test curves showed a significant elevation 4 weeks after streptozocin compared with before streptozocin (p less than 0.05). The maternal insulin response to streptozocin demonstrated a loss of the second-phase insulin response to the glucose load after one dose of streptozocin and loss of the first phase after two doses. The fetuses of the streptozocin-treated ewes showed a significant elevation in plasma glucose level compared with that of controls (13.3 +/- 0.8 mg/dl, n = 5) vs 42.1 +/- 8.1 mg/dl, n = 10; p less than 0.05, control vs streptozocin, respectively). There was a consistent trend to fetal hyperinsulinemia in the fetuses of the streptozocin-treated ewes, although this did not achieve statistical significance (3.3 +/- 0.8 microIU/ml, n = 5 vs 9.6 +/- 2.5 microIU/ml, n = 10; p = 0.06, control vs streptozocin, respectively). The fetal insulin/glucose ratio was preserved in the streptozocin-treated ewes. Comparison of fetal weights between the control and diabetic ewes showed a significant increase in fetal weight in the fetuses of diabetic ewes (3280 +/- 46 gm in control fetuses vs 3710 +/- 54 gm in diabetic fetuses, p less than 0.05). The alterations in the maternal glucose and insulin response resulting from streptozocin-induced pancreatic beta-cell destruction combined with elevations in fetal glucose, insulin, and weight provides a large animal model suitable for investigation of gestational diabetes in pregnancy.

The metabolic clearance rate of epinephrine in the fetus of the diabetic ewe.

Delayed organ maturation is a characteristic of the fetus of the diabetic mother with poor glucose control. We hypothesized that the ovine fetal catecholamine maturation sequence would be delayed in the fetus of the diabetic ewe. Twelve pregnant ewes were rendered glucose intolerant by the administration of streptozocin at 85 to 95 days' gestation. Maternal diabetic status was verified with fasting blood glucose assessments. The fetal metabolic clearance rate of epinephrine was determined at 126 to 140 days' gestation by the constant infusion of 0.1 micrograms epinephrine per kilogram estimated fetal weight per minute. Fetal arterial blood gas values, lactate, glucose, and insulin levels were measured before infusion; after 20, 30, and 40 minutes of epinephrine infusion; and 15, 30, and 60 minutes after cessation of infusion. Fetal plasma glucose rose significantly from a control level of 42.1 +/- 8.2 to 64.6 +/- 5.9 mg/dl during the epinephrine infusion (p less than 0.05). Fetal insulin levels increased from a baseline of 9.2 +/- 2.6 microIU/ml to 32.4 +/- 18.0 in the recovery period (p less than 0.05), and lactate levels similarly rose from 36.4 +/- 4.8 to 52.2 +/- 8.2 mg/dl (p less than 0.05). The plasma epinephrine production rates did not vary significantly between fetuses less than 135 and greater than 135 days' gestation (15.4 +/- 1.5 vs 13.9 +/- 2.0 ng/min, p greater than 0.5). The fetal metabolic clearance rate of epinephrine at early gestations (less than 135 days) was similar to that of the fetus of the nondiabetic ewe (35.3 +/- 3.4 vs 28.0 +/- 4.3 ml/min/kg, p greater than 0.2). However, at later gestations (greater than 135 days) the fetus of the diabetic ewe did not have the increase in the metabolic clearance rate previously published for the control fetus (32.0 +/- 4.6 vs 133.7 +/- 41.7 ml/min/kg, p less than 0.05). These data appear to indicate an absence in the maturation of the metabolic clearance rate of epinephrine in the fetus of the diabetic ewe. The observed alteration in the metabolic clearance rate of epinephrine of the fetus of the diabetic ewe could potentially impair the response to stress.

Placental thromboxane and prostacyclin production in an ovine diabetic model.

We hypothesized that streptozocin-induced ovine diabetes would cause alterations in the placental production of thromboxane and prostacyclin. With a tissue incubation technique, we examined the placental production of thromboxane and prostacyclin in cotyledons from seven normal near-term ewes (127 +/- 3 days' gestation) and six streptozocin-induced diabetic ewes (125 +/- 3 days' gestation). Diabetic status was verified with serial fasting blood glucose assessments. Placental tissue was incubated in Dulbecco's modified Eagle's medium for 48 hours at 37 degrees C with 95% oxygen and 5% carbon dioxide. Samples were collected at 0, 1, 2, 4, 8, 20, 32, and 48 hours. Radioimmunoassay of the stable metabolites thromboxane B2 and 6-keto-prostaglandin F1 alpha were used to determine thromboxane and prostacyclin production, respectively. Placental thromboxane production was reduced in diabetic animals when compared with control animals (5.63 +/- 2.81 vs 7.32 +/- 1.37 pg/mg per hour, respectively; p less than 0.05). Prostacyclin production was also significantly reduced in the diabetic placentas compared with control placentas (11.44 +/- 4.06 vs 16.29 +/- 4.59 pg/mg per hour, respectively; p less than 0.05). We conclude that the ovine placenta produces thromboxane and prostacyclin. The ovine thromboxane production rate is comparable to that of the human placenta but the prostacyclin production rate is approximately two to three times higher. The observed decrease in the placental production of thromboxane and prostacyclin may reflect an adverse effect of hyperglycemia directly on eicosanoid production or indirectly through decreased placental cellular proliferation.

Characterization of a potent biotin-conjugated CRF analog and the response of anterior pituitary corticotropes.

A biotin-conjugated synthetic corticotropin releasing factor (B-CRF) was prepared and characterized. Its biological activity and binding affinity were compared with that of unlabeled synthetic CRF. Both forms of the releasing factor were equipotent in in vitro studies measuring the release of corticotropin (ACTH) (ED50 = 1 nM). The IC50 in the binding assays was 1.5 nM for CRF and 4 nM for B-CRF. Dual avidin-biotin peroxidase complex stains were then used in pituitary monolayer cultures to visualize receptivity to the releasing factor and to confirm opiocortin storage in the target cells. All corticotropes showed stain for B-CRF. The percentage of cells that were double-labeled for ACTH and CRF increased with the dose of B-CRF during a four hour incubation period. The CRF stain was abolished, however, when an excess of unlabeled CRF was added to compete with B-CRF. The distribution of the B-CRF and ACTH stains varied in the cells with the time of exposure to the analog. These studies show that biotin-conjugate CRF is a potent analog that can be demonstrated cytochemically on cells identified immunocytochemically as corticotropes. It can be used to follow important events associated with CRF stimulation including the rapid internalization of CRF coupled with the mobilization of corticotropin stores and the formation of cellular processes.

Somatostatin fibers and their relationship to specific cell types (GH and TSH) in the rat anterior pituitary.

Fibers immunocytochemically stained for somatostatin (growth hormone-release-inhibiting hormone) were localized in the anterior pituitaries of rats. Initial studies of fiber localization involved the use of thick frozen (30 micron) sections which allowed visualization of fibers as they coursed along the periphery of the anterior lobe in the sagittal plane and along blood vessels throughout the lobe. Fibers were observed most often at the rostral, caudal, and lateral poles. In thinner (1-3 micron) paraffin sections, stained somatostatin fibers could be localized in close proximity to cells that were stained for growth hormone or thyroid stimulating hormone in a double stain with a second peroxidase substrate. These and our previous light microscopic studies show that a few neuronal processes containing neurotransmitters extent beyond the level of the median eminence (or perhaps from a peripheral source), penetrate the anterior lobe in specific regions, and lie in close proximity to cells known to be controlled by the transmitter.