Prostacyclin-independence in beta-adrenoceptor mediated renin release from dog renal cortical slices

There is some controversy regarding whether stimulation of renin release by the beta-adrenergic system is dependent on prostaglandin (PG) production. We have examined this problem in renal cortical slices of the dog and have obtained the following results: (1) Isoproterenol (4 X 10(-6) M) stimulated renin release, but had no effect on the formation of 6-keto PGF1 alpha, a stable metabolite of PGI2; (2) Indomethacin (2 X 10(-5) M) had no effect on isoproterenol stimulated renin release, but inhibited 6-keto PGF1 alpha formation; (3) Dibutyryl cyclic AMP (10(-3) M) stimulated both renin release, and 6-keto PGF1 alpha release. Indomethacin (2 X 10(-5) M) did not inhibit dibutyryl cyclic AMP-stimulated renin release, but did inhibit the production of 6 keto PGF1 alpha. These results indicate that the beta-adrenoceptor mediated renin release does not depend on the formation of PGI2, but renin release is dependent on cyclic AMP formation.     

Prostaglandins and renin release from submaxillary glands in vitro

The present studies were undertaken to investigate the effect of prostaglandins (PGs) on renin release from the submaxillary glands of mice. Pooled mouse submaxillary gland slices were incubated in Krebs-Henseleit buffer solution following a preincubation period, and renin release was measured by a radioimmunoassay for the direct measurement of submaxillary gland renin. Arachidonic acid (AA) significantly stimulated renin release at 10, 20, and 30 min of incubation. These increases of renin release were abolished by the presence of indomethacin. The synthetic prostaglandin endoperoxide analogue (EPA) strongly stimulated renin release at 10, 20, and 30 min of incubation. However, at a higher concentration the stimulating effect of EPA virtually disappeared. PGI2 caused the highest increase of renin release at 10 and 20 min of incubation. At higher concentrations the effect of PGI2 on renin release was drastically reduced, although it was still statistically significant. PGE2 and PGF2 alpha also exerted a significant increase in renin release; however, the extent of this effect was much less than that of EPA and PGI2. Other prostaglandins such as PGE1, PGA2, PGD2, PGF1 alpha, and 6-keto-PGF1 alpha were found to have no significant effect on renin release. These results suggest that the prostaglandin system directly affects renin release from submaxillary gland independent of systemic hemodynamic and neurogenic influences.     

Effect of vasodilator prostaglandins on the vascular renin-angiotensin system

The interaction of prostaglandin (PG) with the vascular renin-angiotensin (R-A) system was examined by studies on the effects of PGI2, PGE2 and the inhibitor of PG synthesis, indomethacin, on the release of angiotensin II (Ang II) from isolated rat mesenteric arteries. The Ang II released from the vasculature was measured after its concentration in a Sep-Pak C18 cartridge connected to the perfusion system. After perfusion with drugs, the specific vascular renin activity inhibited by anti-renin antibody was determined. The basal perfusion pressure was constant (19.6 +/- 1.1 mmHg) at a flow rate of 4.5 ml/min, and was not changed by any of these drugs. The basal levels of Ang II release and vascular renin activity were 44 +/- 5 pg/30 min and 113 +/- 8 pg Ang I/mg protein/hr, respectively. Infusion of PGI2 (10(-6) M) significantly decreased both Ang II release (p less than 0.01) and vascular renin activity (p less than 0.05) as compared with the control levels. Infusion of PGE2 (10(-6) M) decreased Ang II release significantly (p less than 0.05) and vascular renin activity slightly. Infusion of indomethacin (10(-6)M) increased vascular renin activity significantly (p less than 0.01). Pretreatment with indomethacin (10 mg/kg, ip) for 2 days also increased vascular renin activity (p less than 0.01). These results indicate that in contrast to their effects on the renal R-A system, PGs suppress the vascular R-A system and that these two local vasoactive factors interact to regulate vascular tone.     

Prostaglandins and renin release: I. Stimulation of renin release from rabbit renal cortical slices by PGI2.

Prostaglandins have been shown to be involved in the mechanism of renin secretion in a variety of situations. Both arachidonic acid and prostaglandin endoperoxide have been shown to release renin from cortical slices and to be converted to PGI2 by cortical microsomes. In the present studies PGI2 was found to cause a time dependent increase in renin release from rabbit renal cortical slices, a system isolated from any indirect effects that result from the administration of prostaglandins in vivo. The stimulation was linear up to 30 minutes and effective over a range of concentrations from 10(7 M to 10(-5) M. At similar concentrations 6-keto-prostaglandin F1alpha was not active on these slices. Thus, it is proposed that PGI2 exerts a direct effect on the release of renin from cortical cells and may be the mediator of arachidonate or prostaglandin endoperoxide stimulated renin secretion.     

Epinephrine stimulates prostaglandin synthesis by bullfrog lung from warm-acclimated, but not cold-acclimated, animals

Exogenous prostaglandins (PGs) have been shown to have differing effects on frog lung contractility. In this study, prostaglandin synthesis was measured in lung tissues from warm-acclimated (WA, 22 degrees C) and cold-acclimated (CA, 5 degrees C) American bullfrogs, Rana catesbeiana, incubated for 30 min at 5 degrees or 22 degrees C. Media were assayed by radioimmunoassay for PGE2, PGF2 alpha, 6-keto PGF 1 alpha (the metabolite of PGI2), and thromboxane (TX)B2 (the metabolite of TXA2). PGE2 was produced in greatest quantity by tissues from WA and CA animals, at both incubation temperatures. Epinephrine stimulated PGE2, PGF2 alpha, and TXB2 synthesis at 22 degrees C but only stimulated PGE2 production at 5 degrees C. In tissues from CA frogs, epinephrine did not stimulate prostaglandin synthesis at either incubation temperature. Ibuprofen (10(-5) M) inhibited basal and epinephrine-stimulated prostaglandin synthesis in tissues from WA frogs incubated at 22 degrees C. The beta receptor antagonist propranolol (10(-6) M) blocked the epinephrine-stimulated synthesis of PGE2, PGF2 alpha and TXB2, suggesting epinephrine stimulates prostaglandin synthesis through beta receptor activation. The absence of stimulation by epinephrine in lung from CA animals, but not in 5 degrees C incubations of tissues from WA animals, suggests that a modification of beta receptors occurs during prolonged cold exposure.     

Dissociation of beta-adrenergic stimulation of renin secretion and prostacyclin synthesis in the rabbit kidney

The effect of inhibition of prostaglandin (PG) synthesis with indomethacin on basal and isoproterenol-stimulated renin secretion was examined in the isolated perfused rabbit kidney. 6-keto PGF1 alpha' the stable metabolite of prostacyclin, was measured in urine by radioimmunoassay using 125I labelled histamine coupled to 6-keto PGF1 alpha as ligand. The level in urine, prior to isolation and perfusion of the kidney, was 10.7 +/- 5.6 ng/min, and this was reduced to 0.32 +/- 0.25 ng/min (P less than 0.05) in rabbits treated with 2.0 mg/kg of indomethacin. Renin release was markedly stimulated by intrarenal infusion of isoproterenol (0.1 microgram/min) but urinary 6-keto PGF1 alpha did not change. These responses were not affected by indomethacin treatment. Renal perfusion pressure, perfusate flow rate and consequently renal vascular resistance, remained relatively constant during the course of perfusion and were unaltered by indomethacin treatment. These results therefore do not support a role for PGs, and in particular prostacyclin, in the renin response to beta-adrenergic stimulation with isoproterenol.     

A physiological role of E and F series prostaglandins in the regulation of TSH secretion

The regulation of TSH secretion by E1, E2, E1 alpha and F2 alpha prostaglandins was studied by means of a monolayer culture system of dispersed rat anterior pituitary cells which was appropriately responsive to TRH, T3 and SRIF. PGEs and Fs induced significant increases in basal TSH release of the order of 30% at 10(-9) or 10(-8) to 10(-5) or 10(-4) M. Only PGEs accentuated the TSH release induced by a half maximal dose of TRH (10(-9) M) of the order of 60% in a dose dependent manner (10(-9) to 10(-6) M of PGEs), whereas PGFs did not. SRIF (10(-8) or 10(-9) M) alone failed to alter basal TSH release but did completely inhibit the TSH response to TRH (10(-9) M). SRIF also significantly inhibited both the increase in basal TSH release and the accentuation of the TSH response to TRH induced by PGEs (10(-6) M) but did not diminish the enhancement of basal TSH release induced by PGFs (10(-6) M). 7-oxa-13-prostynoic acid (PY1), a prostaglandin antagonist, which can act as an agonist in some systems, itself exhibited agonistic properties of PGEs with respect to basal and TRH induced TSH release. PY1 failed to inhibit the TSH release induced by all PGs, but partially inhibited the accentuated TSH response to TRH induced by PGEs. Indomethacin, PG synthetase inhibitor, did not affect basal or TRH induced TSH release in our system. These data suggest that PGs of the E and F series probably modulate TSH release via different mechanisms and that the PGE effect on basal TSH release differs from its augmentation of TRH induced TSH response. It is speculated that these effects of PGs may have physiological significance.     

Comparative effects of prostaglandin E2 and prostaglandin E3 on water flow and cyclic AMP in the urinary bladder of the frog, Rana pipiens

Prostaglandins (PGs) modulate osmotic water flow in amphibian urinary bladders. Gas chromatographic analysis of prostaglandin precursors in bladders showed that arachidonic acid represented 13.0 +/- 0.6% and eicosapentaenoic acid 4.3 +/- 0.1% of the total fatty acid content. The effects of PGE2 and PGE3 on basal and arginine vasotocin (AVT) stimulated water flow were compared. Control water flow (1.1 +/- 0.2 mg/min) was increased to 4.6 +/- 0.3 mg/min with AVT (10(-6)M) present. PGE2 (10(-6)M) inhibited both basal and AVT stimulated water flow. In contrast, PGE3 (10(-6)M) stimulated basal water flow and further increased AVT stimulated water flow. Basal adenylate cyclase activity (ACA, 59 +/- 0.3 pmol cyclic AMP/mg protein/10 min) was stimulated by the addition of AVT in the absence or presence of exogenous guanosine 5' triphosphate (GTP, 10(-5)M). Both PGE2 and PGE3 stimulated basal ACA in the absence, but not in the presence of GTP. In the absence of exogenous GTP, PGE2 increased AVT stimulated ACA, whereas PGE3 decreased it. Both prostaglandins inhibited AVT stimulation when GTP was added. The effects of PGE2, PGE3 and AVT on tissue cyclic AMP levels in whole urinary bladders were similar to the effects seen on ACA in the absence of exogenous GTP. The contrasting effects of PGE2 and PGE3 on control water flow appear distinct from their similar effects on ACA. However, PGE2 and PGE3 may regulate AVT stimulation through mechanisms involving cyclic AMP.     

Role of phospholipase C in mediating oxytocin-induced release of prostaglandin F2 alpha from ovine endometrial tissue

Two experiments were conducted to determine if the ability of oxytocin to stimulate release of prostaglandin (PG)F2 alpha from ovine uterine tissue involved activation of phospholipase C (PLC). In the first experiment, 9 ewes were injected with progesterone for 11 d (12 mg/d, im). On days 11 and 12, ewes received an injection of estradiol (100 micrograms, im). Caruncular endometrial tissue was collected on d 13 and incubated in the presence or absence of oxytocin (10(-6) M). Concentrations of PGF2 alpha and its metabolite, 13,14-dihydro-15-keto-PGF2 alpha (PGFM), in culture media were determined by radioimmunoassay. PLC activity was determined by measuring the intracellular accumulation of 3H-inositol phosphates after preincubation with 3H-inositol. Concentrations of PGF2 alpha and total PGF (PGF2 alpha + PGFM) in culture media were greater for explants treated with oxytocin than for controls (p. less than .02, p less than .06, respectively). A similar effect of oxytocin on intracellular concentrations of 3H-inositol phosphates was observed (p less than .01). A second experiment was conducted to determine if agonists of second messengers, produced by activation of PLC, could stimulate release of PGF2 alpha from ovine endometrial tissue. Seven ewes were treated with progesterone and estradiol as in experiment 1. Explants of caruncular tissue from each ewe were incubated with 1) control medium, 2) A23187 (10(-5) M), 3) oxytocin (10(-6) M), 4) phorbol 12-myristate 13-acetate (PMA, 10(-7) M), 5) PMA + A23187 and 6) PMA + oxytocin. Significant stimulatory effects of oxytocin, PMA and A23187 on concentrations of PGF2 alpha and total PGF in culture media were observed (p. less than .05, p less than .1, p less than .1, respectively). In conclusion, oxytocin stimulated release of PGF2 alpha and activity of PLC in explants of ovine endometrial tissue in vitro. Second messengers associated with activation of PLC enhanced release of PGF2 alpha from ovine endometrial tissue.     

Correlation of the biosynthesis of prostaglandin and cyclic AMP in monolayer cultures of rabbit articular chondrocytes

We have utilized ionophores to test whether stimulation of chondrocyte prostaglandin biosynthesis is accompanied by an increase in cyclic nucleotide levels in these cells. Radioimmunoassay of prostaglandin E2, 6-oxo-prostaglandin F1 alpha (the stable metabolite of prostaglandin I2) and prostaglandin F2 alpha showed that synthesis of each was stimulated by the divalent-cation ionophore, A23187 after short-term incubation (1-7 min) in serum-free medium. No stimulation of thromboxane B2 was detected. Two monovalent ionophores, lasalocid and monensin failed to stimulate prostaglandin biosynthesis after short-term incubation. Ionophore A23187-stimulated prostaglandin biosynthesis was variably and partially inhibited by sodium meclofenamate, indomethacin and aspirin, but not by sodium salicylate. Ionophore A23187-stimulated prostaglandin biosynthesis was accompanied by a 7.5-fold increase in cyclic AMP levels after 15 min. Sodium meclofenamate, indomethacin and aspirin which inhibited prostaglandin E2 biosynthesis also reduced cyclic AMP levels. Exogenous prostaglandin E2 (1 microgram/ml) stimulated cyclic AMP biosynthesis, which was not inhibited by aspirin. These results indicated that prostaglandins can be considered as one of the local effectors controlling cyclic AMP production in articular cartilage.     

Zero extracellular K+ and prostaglandin E release in the guinea-pig taenia coli

Prostaglandin E release rates from isolated strips of guinea-pig taenia coli increased during exposure to zero K+ bathing fluid, from control values of 0.78 +/- 0.11 ng/g per min to levels as high as 29.2 ng/per min. Release rates increased for 40-50 min and then remained constant or fell despite progressive increases in intracellular sodium [Nai+] or fall in intracellular potassium [Ki+]. Readmittance of K+ to the bathing solution resulted in rapid reversal of elevated prostaglandin E release rates. [Nai+] and [Ki+] were markedly more abnormal in strips exposed to zero K+ for 70-201 min compared to 30-min exposures. Upon the readdition of K+ after long zero K+ exposure, the rate of prostaglandin E release fell long before [Nai+] and [Ki+] returned to control levels. After K+ was readded to the bathing solution, the ion concentration of tissues exposed to zero K+ for 30 min returned to normal much more quickly than did those of tissues exposed for the longer time periods, yet the exponential rate constants for fall of prostaglandin E release rate after K+ was added were not significantly different after short or long zero K+ exposure. Thus there was a dissociation between the return of [Nai+] and [Ki+] and the fall of prostaglandin E release rate to control levels. Ouabain augmented prostaglandin E release under conditions where [Ki+] could not fall. Addition of known neurotransmitters present in this tissue to the bathing fluid did not augment prostaglandin E release. Guinea-pig taenia coli strips that had been incubated with [3H]arachidonic acid, constantly released [3H]arachidonic acid and [3H]prostaglandin E and a prostaglandin which cochromatographed with prostaglandin E but could not be converted to prostaglandin B by alkali and was shown to be 6-ketoprostaglandin F1 alpha. Release of [3H]arachidonic acid and [3H]prostaglandin E plus 6-[3H]ketoprostaglandin F1 alpha was increased when strips were exposed to zero K+. Data obtained in this study suggest the augmented prostaglandin E release seen during zero K+ or ouabain is related to increased availability of unbound arachidonic acid at the site of cyclooxygenase in the cell. Augmented prostaglandin E release is apparently not related to alterations in intracellular electrolyte concentrations or release of known neurotransmitters.     

Histamine alters prostaglandin output from diestrous rat uteri. Involvement of H2-receptors and 9-keto-reductase

The effects of exogenous histamine (H) on prostaglandin (PG) generation and release in uteri isolated from diestrous rats and the influences of H2-receptors blockers (cimetidine and metiamide) on the output of uterine PGs, were explored. Moreover, the action of H on the uterine 9-keto-reductase, was also studied. Histamine (10(-4) M) failed to alter the basal output of PGE1 but reduced significantly the generation and release of PGE2 and augmented the output of PGF2 alpha. On the other hand, cimetidine (10(-5) M) enhanced the basal release of PGE2 but had no action on the outputs of PGs E1 or F2 alpha. The enhancing effect of H on the production and release of PGF2 alpha was abolished in the presence of cimetidine. Also, the antagonist reversed the influence of H on the output of PGE2. Metiamide, another H2-receptor antagonist, did not alter the basal control generation and release of uterine PGs, but antagonized the augmenting influence of H on PGF2 alpha uterine output, as much as cimetidine did, and prevented the depressive action of H on the release of PGE2 from uteri. Histamine (10(-4) M) significantly stimulated uterine formation of cyclic-adenosine monophosphate, an action which was antagonized by the presence of cimetidine (10(-5) M), a blocker of H2 receptors. Also, histamine (10(-5) M) and dibutyrylcyclic-adenosine monophosphate (DB-cAMP) at 10(-3) M, enhanced significantly the formation 3H-PGF2 alpha from 3H-PGE2. Results presented herein demonstrate that H is able to diminish the generation of PGE2 in uteri from rats at diestrus augmenting the synthesis of PGF2 alpha, apparently via the activation of H2-receptors, enhancing adenylate-cyclase. These effects appear to increase uterine 9-keto-reductase activity which transforms PGE2 into PGF2 alpha. Relationships between the foregoing results and those evoked by estradiol, are also discussed.     

Endothelin inhibits renin release from isolated rat glomeruli

The effect of endothelin on renin release from isolated rat glomeruli was examined. Endothelin inhibited basal renin release in a dose-dependent manner with an IC50 of 1.0 x 10(-9) M. Endothelin also inhibited renin release stimulated by isoproterenol (10(-5) M). Nifedipine (10(-5) M), a calcium channel blocker, induced an increase in renin release. Endothelin did not affect this nifedipine-induced renin release. These results suggest that endothelin inhibits renin release via a calcium entry mechanism and increases intracellular calcium.     

Potential mediation of prostaglandin E2 release from isolated human parietal cells by protein kinase C

Parietal cells are a major source of gastric mucosal prostaglandins in various species. We examined cholinergic stimulation of prostaglandin E2 (PGE2) release from human parietal cells; using activators of the protein kinase C we attempted to get an indirect insight into cellular mechanisms which control PGE2 release. Gastric mucosal specimens were obtained at surgery and the cells were dispersed by collagenase and pronase E. Parietal cells were enriched to 65-80% by a Percoll gradient, and were incubated for 30 min. PGE2 release into the medium (radioimmunoassay) was 74-126 pg/10(6) cells/30 min under basal conditions and was 2.6-fold increased by carbachol (10(-5) and 10(-4) M). Similarly, PGE2 release was stimulated by phospholipase C (20-200 mU/ml, 364% above basal), 1-oleoyl-2-acetyl-sn-glycerol (10(-9)-10(-5) M, 229%), 12-O-tetradecanoylphorbol-13-acetate (TPA; 10(-9)-10(-5) M, 283%) and calcium ionophore A23187 (10(-7)-10(-5) M, 219%). Simultaneous presence of A23187 and TPA synergistically induced stimulation which was slightly higher than the sum of the individual responses. N-(6-aminohexyl)-5-chloro-1-naphthalene sulfonamide W-7, a putative calmodulin antagonist, inhibited TPA-induced PGE2 release at concentrations regarded specific for blocking calmodulin (IC50 = 1.5 X 0(-6) M). We conclude that in human parietal cells PGE2 is released upon cholinergic stimulation and that phospholipase C and protein kinase C are involved in the control of PGE2 release. We speculate that calmodulin might interact with a protein phosphorylated by protein kinase C to cause PGE2 release.     

Serotonin Inhibits Acetylcholine Release from Rat Striatum Slices: Evidence for a Presynaptic Receptor-Mediated Effect

Rat brain striatum slices were incubated with [3H]choline, perfused with a physiological buffer, and stimulated by perfusion with a K+-enriched buffer for 2 min. The tritium overflow evoked by K+ was decreased by 5-hydroxytryptamine (serotonin, 5-HT) (maximal inhibition 10(-6) M). This effect of 5-HT was mimicked by several agonists (5-methoxytryptamine, N,N-dimethyl-tryptamine, bufotenin) and blocked by serotonergic antagonists (methiothepin, methysergide, cinanserin) but not by haloperidol; methiothepin and methysergide alone slightly increased the K+-evoked overflow of tritium (3H). Inhibition of the tritium release by 5-HT was not suppressed in the presence of tetrodotoxin (TTX) (10(-6) M). These results suggest that 5-HT tonically inhibits acetylcholine (ACh) release from striatal cholinergic neurons by acting on a presynaptic receptor localized on cholinergic terminals.     

Kinetic measurements of the release of prostaglandins from perfused rat lungs.

Prostaglandins released from isolated ventilated and perfused rat lungs were measured by a simple modification of the Vane technique using the rat stomach fundus as a continuous bioassay tissue. Exogenously supplied arachidonic acid was converted mainly to PGF2alpha which was determined by bioassay. A novel method for mixing a stream of inhibitors with the perfusate was used to determine PGF2alpha in the presence of substrate amounts of arachidonic acid. Using this system the apparent Km for PGF2alpha production with arachidonic acid as the substrate was found to be 1.90 X 10(-4)M, while the Ki for aspirin was found to 2.47 X 10(-4)M. These kinetic parameters are close to those reported for cell free systems and subcellular fractions suggesting that both substrate and inhibitor have ready access to the site of prostaglandin synthesis. The method appears to be generally useful to determine the effect of drugs and environment factors on the release of prostaglandins by the lung.     

Sex differences in arachidonate cyclo-oxygenase products in elicited rat peritoneal macrophages

Peritoneal macrophages were elicited by Freund's incomplete adjuvant from adult male and female Fisher 344 rats. The release of prostaglandin E2 and thromboxane B2 from these macrophages was determined by radioimmunoassay. The basal release of these products was the same for males and females. The macrophages of the female rats released, in a dose-dependent manner, significantly more prostaglandin E2 and thromboxane B2 than macrophages from the male, following challenge with either a particulate stimulus, zymosan (25-150 micrograms/ml) or a soluble stimulus, calcium ionophore A23187 (1 X 10(-7) -1 X 10(-6) M). These results may relate to gender differences in immune responses.     

Dissociation of hyperreninemia and renal prostaglandin synthesis in the adrenalectomized rat

The aim of this study was to determine whether hyperreninemia in the adrenalectomized (ADX) rat is dependent on renal prostaglandin synthesis, as has been suggested for two other hyperreninemic conditions, Bartter's syndrome and chronic liver disease. Plasma renin concentration (PRC) in anesthetized, ADX rats was significantly increased (delta +480%; p less than 0.001) compared to sham-operated controls. In vivo, indomethacin (10 mg/kg i.v.) significantly reduced PRC of anesthetized, ADX rats after both 45 min (delta -34%; p less than 0.05) and 90 min (delta -47%; p less than 0.05). In vitro renin release from renal cortical slices of ADX rats was also significantly greater (delta +130%; p less than 0.05) than from sham-operated control cortical slices. Renin release from cortical slices of ADX rats given dexamethasone (10 micrograms/kg/day) for 4 days prior to sacrifice did not differ from sham-operated control values. Prostaglandin E2 (PGE2) release from cortical slices of ADX rats did not differ significantly from controls. However, PGE2 synthesis in glomeruli microdissected from ADX rats was significantly increased (delta +110%; p less than 0.001) compared to controls. PGE2 synthesis in glomeruli of dexamethasone-treated ADX rats remained significantly elevated compared to controls. Ibuprofen (10(-6) M) decreased PGE2 synthesis in cortical slices by 80%. However, prostaglandin synthesis inhibition had no effect on renin release from either ADX or control renal cortical slices. These results suggest that despite increased glomerular synthesis, prostaglandins do not directly influence renin release in the ADX rat.     

Testing the "redirection hypothesis" of prostaglandin metabolism in the kidney

Furosemide increases the synthesis of two major renal eicosanoids, prostacyclin (PGI2) and thromboxane A2 (TXA2), by stimulating the release of arachidonic acid which in turn is metabolized to PGG2/PGH2, then to PGI2 and TXA2. PGI2 may mediate, in part, the early increment in plasma renin activity (PRA) after furosemide. We hypothesized that thromboxane synthetase inhibition should direct prostaglandin endoperoxide metabolism toward PGI2, thereby enhancing the effects of furosemide on renin release. Furosemide (2.0 mg . kg-1 i.v.) was injected into Sprague-Dawley rats pretreated either with vehicle or with U-63,557A (a thromboxane synthetase inhibitor, 2 mg/kg-1 followed by 2 mg/kg-1 X hr-1). Urinary 6ketoPGF1 alpha and thromboxane B2 (TXB2), reflecting renal synthesis of PGI2 and TXA2, as well as PRA and serum TXB2, were measured. Serum TXB2 was reduced by 96% after U-63,557A. U-63,557A did not affect the basal PRA. Furosemide increased PRA in both vehicle and U63,557A treated rats. However, the PRA-increment at 10, 20 and 40 min following furosemide administration was greater in U-63,557A-treated rats than in vehicle-treated rats and urine 6ketoPGF1 alpha excretion rates were increased. These effects of thromboxane synthesis inhibition are consistent with a redirection of renal PG synthesis toward PGI2 and further suggest that such redirection can be physiologically relevant.     

Differential actions of corticosterone on luteinizing hormone and follicle-stimulating hormone biosynthesis and release in cultured rat anterior pituitary cells: interactions with estradiol

Previous in vivo studies from our laboratory suggested that glucocorticoids antagonize estrogen-dependent actions on LH secretion. This study investigated whether corticosterone (B) may have similar actions on gonadotropin biosynthesis and secretion in vitro. Enzymatically dispersed anterior pituitary cells from adult female rats were cultured for 48 h in alpha-modified Eagle's medium containing 10% steroid-free horse serum with or without 0.5 nM estradiol (E2). The cells were then cultured for 24 h with or without B in the presence or absence of E2. To evaluate hormone release, 5 x 10(5) cells were incubated with varying doses of GnRH (0, 10(-11)-10(-7) M) or pulsatile GnRH (10(-9) M; 20 min/h) for 4 h. Cell and medium LH and FSH were measured by RIA. To evaluate LH biosynthesis, 5 x 10(6) cells were incubated for an additional 24 h with 10(-10) M GnRH, 60 microCi 3H-glucosamine (3H-Gln), 20 microCi 35S-methionine (35S-Met), and the appropriate steroid hormones. Radiolabeled precursor incorporation into LH subunits was determined by immunoprecipitation, followed by SDS-PAGE. Continuous exposure to GnRH stimulated LH release in a dose-dependent manner, and this response was enhanced by E2. B by itself had no effect on LH release, but inhibited LH secretion in E2-primed cells at low concentrations of GnRH (10(-10) M or less). Total LH content was not altered by GnRH or steroid treatment. Similar effects of B were observed in cells that were given a pulsatile GnRH stimulus. In contrast to LH, E2 or B enhanced GnRH-stimulated FSH release at the higher doses of GnRH, while the combination of E2 and B increased basal and further augmented GnRH-stimulated release. Total FSH content was also increased in the presence of B, but not E2 alone, and was further augmented in cells treated with both steroids. There were no effects of the steroids on the magnitude of FSH release in response to GnRH pulses, but the cumulative release of FSH was greater in the E2 + B group compared to controls, indicating an increased basal release. Independent of E2, B suppressed the incorporation of 3H-Gln into LH by more than 50% of control, with only subtle effects on the incorporation of 35S-Met.(ABSTRACT TRUNCATED AT 400 WORDS)