Modulating effects of prostaglandins on parasympathetic-mediated secretory activities of rat salivary glands

Exogenously administered PGE1 or PGE2, like atropine, markedly decreased both the flow and calcium concentration of parasympathetically evoked rat parotid saliva; PGF2 alpha was less effective. Despite the fact that prostaglandins greatly reduced the Ca concentration of nerve-evoked saliva, they did not change the glandular Ca concentration of either control or parasympathetically stimulated parotid glands. Prostaglandins (20 micrograms/kg, i.a.) decreased the Na or K concentration of nerve-evoked parotid saliva, but at lower doses had no significant effect. PGE1, PGE2, PGF2 alpha or atropine markedly decreased flow rates of similarly evoked rat submandibular saliva. Prostaglandins and atropine, however, decreased the Na concentration and increased the K concentration of parasympathetically evoked submandibular saliva. PGF2 alpha, like atropine, increased the Ca concentration of such saliva. Drug vehicle, ethanol, slightly decreased the flow of both parotid and submandibular saliva but not the ion secretion, Endogenous prostaglandins themselves may not play a role in secretory activities during parasympathetic nerve stimulation of rat salivary glands, since administration of indomethacin, and inhibitor of prostaglandin biosynthesis, prior to or during nerve stimulation did not significantly alter nerve-evoked salivary secretion, The mechanisms by which prostaglandins modulate secretory responses of salivary glands during parasympathetic stimulation are not understood.     

Localization and in vitro synthesis of prostaglandins in components of rabbit preovulatory graafian follicles.

Graafian follicles obtained 9 hours after the injection of human chorionic gonadotropin (hCG) into mature rabbits were dissected into a follicular fluid component, a granulosa cell-oocyte component, and a residual wall component, (the latter containing mostly theca tissue with a small and variable amount of adhering granulosa cells). The amounts of PGE and PGF were determined for each component. The follicular fluid contained approximately 4-10 times more PGE and PGF than either the granulosa cell-oocyte component or the residual wall component. The latter two components contained approximately equal amounts of these prostaglandins. The in vitro biosynthesis of PGE and PGF was also studied and it was found that the granulosa cell-oocyte component had about 4 fold the capacity of the residual wall, and that the follicular fluid synthesized no prostaglandins. There was no significant effect of LH on either PGE or PGF synthesis in any of the components.     

Effects of indomethacin and prostaglandins on ovulation of goldfish.

A technique is described whereby elevated temperature and HCG injection yield a high percentage of ovulation in gravid goldfish. Indomethacin (10 mug/g; i.p. injection) completely inhibits ovulation if given within 6 hours following HCG (4 IU/g); the unovulated oocytes develop rapidly into corpora atretica. PGE1, PGE2, and PGF2alpha (5 mug/g; i.p. injection) induce ovulation in fish treated with indomethacin and HCG; PGE2 was most effective when given 11 hours after HCG. The results suggest that the ovulatory action of prostaglandins following HCG stimulation is at the level of the ovary and that it is restricted to a period between 7 and 12 hours after the gonadotropin injection.     

A specific radioimmunoassay for PGE2 using an antibody with high specificity and a sephadex LH-20 microcolumn for the separation of prostaglandins.

Antiserum against PGE2 was raised in rabbits following immunization with prostaglandin-hen-gamma-globulin conjugate. The antiserum exhibited 14% cross reactivity with PGE1 and far less cross-reaction with heterologous prostaglandins. A microcolumn of Sephadex LH-20 was used for a partial, but sufficient separation of PGE2 from PGE1 and a complete separation from heterologous prostaglandins to ensure a specific RIA for PGE2. The precision of the method in the rage 10-500 picograms showed a coefficient of variation varying between 4 and 13%. The detection limit was 10 picograms corresponding to 15 pg/ml of PGE2 in serum. In order to demonstrate the validity of the method values obtained for non-diuretic rat renal venous serum were compared with those obtained using the isotope derivative method of Bojesen & Buckhave (1972) on the same samples. The concentrations of PGE2 obtained were 239 +/- 25 pg/ml and 250 +/- 58 pg/ml, respectively.     

Effects of prostaglandins on intraocular pressure recovery rate in rabbits

The actions of a number of prostaglandins (PGs) were studied in unanesthetized rabbits using an intraocular pressure (IOP) recovery-rate method. In topical doses of 0.1 to 10 micrograms, these compounds accelerated the rate at which IOP returned to control levels after an infusion of hypertonic saline. In general, PGE1 appeared more potent than the other PGs at these doses. Arachidonic acid also increased the IOP recovery rate. The effect of arachidonic acid was completely blocked by the cyclooxygenase inhibitor indomethacin. Recovery rate responses to arachidonic acid were increased further after pretreatment with the lipoxygenase inhibitor phenidone. When administered alone, phenidone itself accelerated IOP recovery; this action was also blocked by indomethacin. The IOP recovery rate method appears to be a useful tool for studying ocular effects of PGs and other products or inhibitors of arachidonic acid metabolism.     

Interrelationships between prostaglandins, cyclic AMP and steroids in ovulation.

Ovulation is a complex phenomenon, involving a series of biochemical events within the ovary, leading to the rupture of the follicle. This paper summarizes recent studies in our laboratory of some of these biochemical changes using the rabbit as an experimental model. It has been shown in our laboratory that isolated Graafian follicles obtained from oestrous rabbits synthesize steroids and cyclic AMP when incubated in vitro. Luteinizing hormone added to the incubation medium increased steroidogenesis and cyclic AMP synthesis many fold. When follicles were isolated from rabbits at different times following the ovulatory stimulus (mating or HCG injection) it was found that the in vitro response to LH in terms of steroidogenesis and cylcic AMP synthesis was lost as ovulation approached. In contrast, when prostaglandins (PGF and PGE) were measured in rabbit Graafian follicles it was found that the PGF and PGE levels increased as ovulation approached. From these data and from reports in the literature, we have developed a hypothetical model for ovulation in the rabbit which may help in a better understanding of the ovulatory process.     

The stimulatory effects of prostaglandins on the melanophores of the lizard, Anolis carolinensis

The melanosome dispersing activity of prostaglandins PGE1, PGE2, PGF1 alpha, PGF2 alpha, PGI2 and 6 beta PGI, was tested on the melanophores of Anolis carolinensis. Only PGE2 and PGE1 were active and while PGE2 was the most potent and acted synergistically with alpha-MSH, PGE1 was additive with alpha-MSH. Arachidonic acid also stimulated melanosome dispersion but its effect was blocked by indomethacin suggesting an action through its conversion to PGE1 or PGE2. The effect of alpha-MSH, on the other hand, was unaltered by indomethacin which suggests that alpha-MSH stimulated melanosome dispersion does not depend upon prostaglandin synthesis. Thus, while some prostaglandins may interact with alpha-MSH to stimulate melanosome dispersion they are unlikely to mediate its action.     

Effects of prostaglandins E-1 and F-2ALPHA on serum luteinizing hormone concentration and on some sexual functions in male rabbits.

PGE-1(50 mu-g/animal) and PGF-2ALPHA (250 mu-g/animal) caused a transient increase in serum LH at 5 min after injection. PGE-1(250 mu-g/animal) had a biphasic effect on serum LH. A small peak was obtained at 5 min, and a second, larger peak at 60 min after injection. It is suggested that the first peak is a result of the stress associated with injection of the PGs, whereas the second peak represents a physiological effect of PGE. Subcutaneous injection of PGE-1(1 mg in arachis oil b.i.d.) for 10 days did not effect the concentration of LH in serum, the function of the accessory sexual glands or the sexual activity. PGF-2ALPHA, given at the same dose and in the same manner, increased the sexual activity but left all other variables unaffected. The pituitary responsiveness to LH-RH was unaltered by the treatment with PGE-1 and PGF-2ALPHA.     

Effect of prostaglandins on cyclic nucleotide levels in cultured keratinocytes.

Ginea pig ear epidermal cells (keratinocytes) were established in primary cultures using trypsin, and treated in their proliferative phase of growth with prostaglandins E1, D1, F1 alpha, E2, D2, or F2 alpha. This phase is induced by the addition of retinoic acid during cell plating. Intracellular content of cAMP and cGMP was measured by radioimmunoassay at various times after treatment. Maximum stimulation of cAMP levels was observed with PGD2, smaller increases with PGE2 and relatively transient rises with PGF2 alpha which were of low significance, but confirm earlier data. Similar results were observed with PGD1, PGE1, and PGF1 alpha with smaller increases. The effects of D and E PGs were biphasic. Significant increases in cGMP were immediately observed with PGD2 and PGE2. With PGF2 alpha, maximum cGMP levels were noted after some delay. All PGs tested showed some effect in elevating cyclic nucleotides in keratinocytes. The most striking result was the increase in cAMP on PGD2 treatment.     

Effect of prostaglandins on hypothalamic-pituitary-testicular function in vitro.

The effect of prostaglandins (PG) A1, E1, E2 and F2 alpha in the concentration range of 10(-7)--10(-4) M were studied in vitro on a rat hypothalamic tissue, collagenase-digested isolated anterior pituitary cell and Leydig cell suspension system by measuring the testosterone production of incubated Leydig cells. PGs did not change the testosterone production and the hCG sensitivity of the Leydig cells, nor the LH secretion and the LHRH sensitivity of the anterior pituitary cells. PGE2 at concentrations of 10(-6), 10(-5) and 10(-4) M significantly increased the hypothalamic tissue-induced pituitary-testicular activation, and this stimulatory effect of PGE2 was dose dependent. PGA1, PGE1 and PGF2 alpha did not alter hypothalamic LHRH release measured in vitro. The results suggest that PGE2 has a direct stimulatory effect on hypothalamic LHRH release.     

Effect of prostaglandins on ornithine decarboxylase activity in the testis of immature rat

Intratesticular injection of prostaglandin E2 (PGE2) and F2 alpha (PGF2 alpha) caused stimulation of ornithine decarboxylase (ODC) activity in the testis of immature rats. PGE2 at a dose of 10 microgram per testis was maximally effective 2 hours after the injection. Dibutyryl cyclic AMP (cAMP) and 1 methyl, 3-isobutyl xanthine (MIX), a phosphodiesterase inhibitor, also stimulated ODC activity. Simultaneous injection of PGE2 and FSH or LH caused additional stimulation of ODC activity. Similarly injection of PGE2 in addition to cAMP or MIX also caused increased stimulation of ODC. Indomethacin (IM, 60 microgram/testis) inhibited LH, FSH or cAMP induced ODC activity. However, IM at the same dose inhibited the synthesis of total proteins. These results suggest that PGE2 and PGF2 alpha stimulate the activity of ODC. The action of prostaglandins may be independent of the action of gonadotropic hormones. cAMP appears to mediate the action of prostaglandins in the testis of rat.     

Synthesis and release of prostaglandins by rat brain synaptosomal fractions.

Abstract— Particulate fractions from rat brain homogenate containing the synaptosomes synthesize and release prostaglandins F and E on aerobic incubation. The prostaglandin of the F-typc released could be further identified as proslaglandin F_2α using specific radioimmunoassays for prostaglandins F_1α, and F_2α-. The metabolite 13,14-dihydro-15-keto-prostaglandin F_2α could not be detected. The amount of prostaglandins released is dependent on incubation time and temperature as well as pH and osmolarity of the incubation medium. Total brain homogenate released more prostaglandins than purified synaptosomes per mg protein, indicating that synaptosomes are probably not a main source of prostaglandins when compared with other subcellular brain fractions. While prostaglandin synthesis was only moderately increased by the addition of the precursor fatty acid arachidonic acid, anti-inflammatory drugs like indomethacin, high concentrations of some local anaesthetics and Δ¹-tetrahydrocannabinol inhibited prostaglandin release. The neurotransmitters noradrenaline, dopamine and 5-hydroxytryptamine did not influence prostaglandin release from the synaptosomal rat brain fractions.     

Effect of dietary calcium on renal prostaglandins.

The present study was designed to clarify the possible role of renal prostaglandins (PGs) on blood pressure (BP) regulation during calcium (Ca) restriction or supplementation. Twelve normotensive women with a mean age of 21.2 years participated in the study. After 1 week of normal Ca intake (mean +/- SE, 536 +/- 2 mg/day), a low-Ca diet (163 +/- 1 mg/day) was given for a further 1 week. Additional asparagine Ca (3 g as Ca/day) was also given to half of the subjects. BP, heart rate, and serum total and ionized Ca concentrations were measured at the end of each period. Levels of Ca, sodium, PGE2, 6-keto-PGF1 alpha and thromboxane (TX) B2 excreted into urine were also determined. The plasma level of ionized Ca was significantly increased without any change in total Ca in both groups. Low and high Ca intake decreased and increased urinary Ca excretion by 28% and 56%, respectively. BP was not altered after Ca deprivation or loading. However, urinary PGE2 excretion was significantly augmented from 668.9 +/- 68.1 to 959.7 +/- 183.1 ng/day by Ca loading, whereas Ca deprivation decreased PGE2 excretion (695.4 +/- 108.1 to 513.2 +/- 55.2 ng/day). No changes were observed in 6-keto-PGF1 alpha or TXB2 urinary excretion. These results suggest that renal PGE2 synthesis is stimulated or decreased by 1-week Ca loading or deprivation, indicating a possible antihypertensive role of renal PGE2 during high-Ca intake in hypertensives.     

Effects of insulin and prior exercise on prostaglandin release from perfused rat muscle. Evidence that prostaglandins do not mediate changes in glucose uptake.

Prostaglandin generation and its inter-relation to the metabolic effects of insulin and prior exercise were examined in perfused muscle of fed rats. During a 60 min perfusion of the rat hindquarter, a substantial release of the prostaglandins PGF2 alpha, PGE2 and 6-oxoPGF1 alpha was observed. Blood cells present in the perfusate released these substances in negligible amounts indicating the prostaglandins were produced by the hindquarter. Addition of insulin to the perfusate increased both glucose uptake and the generation of PGE2 and 6-oxoPGF1 alpha. At 30 min after intense treadmill exercise, glucose and alpha-aminoisobutyric acid (AIB) uptake by the hindquarter were increased in the absence of added insulin, but prostaglandin release was not increased. Insulin further increased glucose and AIB uptake; however, in contrast with its effects in non-exercised rats, insulin no longer stimulated prostaglandin generation. Indomethacin (10 microM) added to the perfusate inhibited the release of PGF2 alpha and PGE2 by 90% and the release of 6-oxoPGF1 alpha by 54%. It had no effect on the stimulation of glucose uptake by either insulin or prior exercise. The data indicate that insulin increases prostaglandin synthesis by perfused rat muscle, and that prior exercise blocks this effect. They suggest that under the conditions studied prostaglandins do not mediate the effects of insulin or prior exercise on glucose uptake.     

Effects of infusion of some prostaglandins in essential fatty acid-deficient and normal rats

Infusion of 1 mg/kg per day of prostaglandin E(1) (PGE(1)) for 2 and 7 wk failed to correct the dermal signs of essential fatty acid (EFA) deficiency in rats despite the known conversion of EFA to certain prostaglandins. PGE(1) caused no significant changes in serum cholesterol, triglycerides, or phospholipids or in liver neutral lipids in EFA-deficient or normal rats. In normal rats epinephrine-induced lipolysis was greater in fat pads from infused than from untreated rats. The effect on epinephrine-induced lipolysis was greater after the 7 wk infusion than after the 2 wk infusion. The 7 wk infusion also lowered plasma free fatty acid (FFA) concentrations. Infusion of PGE(2) and PGF(2alpha) in combination for 4 wk had no significant effect on either dermal signs of EFA deficiency, lipolysis, or plasma FFA concentrations.     

Biosynthesis of prostaglandins and thromboxane B2 by fetal lung homogenates

The conversion of arachidonic acid to prostaglandins (PG's) and thromboxane B2 (TXB2) was investigated in homogenates from fetal and adult bovine and rabbit lungs. Adult bovine lungs were very active in converting arachidonic acid (100 microgram/g tissue) to both PGE2 (10.7 microgram/g tissue) and TXB2 (6.2 microgram/g tissue. Smaller amounts of PGF2alpha (0.9 microgram/g) and 6-oxoPGF1alpha were formed. Homogenates from fetal calf lungs during the third trimester of pregnancy were quite active in converting arachidonic acid to PGE2, but formed very little TXB2, PGF2alpha or 6-oxoPGF1alpha. Homogenates from rabbit lungs converted arachidonic acid (100 microgram/g) mainly to PGE2, both before and after birth. The amount of PGE2 formed increased during gestation to a maximum of about 6 microgram/g tissue at 28 days of gestation. It then decreased to a minimum (1.5 microgram/g) which was observed 8 days after birth, followed by an increase to about 4 microgram/g in older rabbits.     

Inductive effects of prostaglandins on alkaline phosphatase in osteoblastic cells, clone MC3T3-E1

The effects of prostaglandins (PGs) on the induction of alkaline phosphatase (ALP) were investigated in osteoblastic clone MC3T3-E1 cells cultured in serum-free medium. Prostaglandin E2 (PGE2) stimulated ALP activity in the cells in a dose-dependent fashion with a maximal effect which was about twice that in the control cells at concentrations of 100-500 ng/ml. Actinomycin D and cycloheximide inhibited the stimulative effect of PGE2 on ALP activity in the cells. PGE2-induced and native ALPs in the cells were of the same type as that in adult mouse calvaria, being heat-labile, L-homoarginine- and levamisole-sensitive, and L-phenylalanine-insensitive. Isobutyl methylxanthine (IBMX), a cAMP phosphodiesterase inhibitor, stimulated the inductive effect of PGE2 on ALP activity at 0.1 mM, at which concentration IBMX alone had little effect on the activity. PGE2 also increased the intracellular cAMP content in a dose-dependent fashion with a maximal effect at 100 ng/ml. PGE1, PGF1 alpha, and PGF2 alpha (primary PGs like PGE2) increased the activity. Our present results suggest that PGs stimulate the differentiation of osteoblasts and are involved in bone formation in vivo, as well as in bone resorption.     

Identity and synthesis of prostaglandins in the lone star tick, Amblyomma americanum (L.), as assessed by radio-immunoassay and gas chromatography/mass spectrometry.

High concentrations of PGE(2) and PGF(2alpha) were identified by radio-immunoassay (RIA) and/or gas chromatography/mass spectrometry (GC/MS) in the hemolymph, salivary glands and saliva of the lone star tick Amblyomma americanum (L.). Binding studies indicated that PGE(2) was free and not bound to any proteins in the hemolymph. A small amount of 6-keto-PGF(1alpha) (breakdown product of PGI(2); prostacyclin) was also found in the salivary glands but not in the hemolymph or saliva. Neither PGD(2) nor PGA(2)/B(2) was detected in any tick material investigated. Although PGE(2) was found in the gut contents, only small amounts of label crossed the gut into the hemolymph during artificial feeding with labeled PGE(2), indicating that the high amounts of PGE(2) in hemolymph and salivary glands are not sequestered from the host blood meal. Isolated salivary glands and salivary gland homogenates demonstrated robust synthesis of PGE(2) at high concentrations of exogenous arachidonic acid. Synthesis by the salivary glands was monitored by measuring increasing PGE(2) with increasing arachidonic acid by RIA, GC/MS and labeled PGE(2) in the presence of labeled arachidonic acid. Synthesis was inhibited in a dose-dependent manner by indomethacin indicating that the cyclooxygenase synthesizing prostaglandins in ticks shares similarities to the enzyme found in mammals.     

Role of prostaglandins in acute phase proteins in inflammation

Prostaglandin E1, a mediator of inflammation, was investigated for its effects on serum acute phase proteins, alpha 2 macroglobulin (alpha 2M). Induction of carrageenin inflammation in rats caused an elevation of alpha 2M to a maximum level (100%) at 1 day. Similarly, administration of PGE1 (1 mg/kg) was found to increase serum alpha 2M levels in normal rats. On the other hand, sc injection of PGE1 into inflamed rats significantly reduced the alpha 2M in serum as well as edema. In vitro studies with liver slices showed increasing rates of incorporation of [14C]leucine into alpha 2M with the addition of PGE1 to the medium. It was followed by the secretion of alpha 2M-bound radioactivity into media. But addition of higher doses (greater than 100 ng/ml) of PGE1 resulted in the suppression of incorporation and secretion of alpha 2M-bound radioactivity. Incubation of inflamed liver slices with PGE1, however, showed only decreased incorporation and secretion of alpha 2M-bound radioactivity. These results indicate that (a) primary prostaglandins, like PGE1, generated during inflammation may be responsible for the increase of alpha 2M in serum, and (b) PGE1 enhanced the synthesis of alpha 2M in liver and its secretion into the medium, so the anti-inflammatory drugs which decrease levels of PGs are likely to alter alpha 2M levels.     

Localization and in vitro synthesis of prostaglandins in components of rabbit preovulatory Graafian follicles

Graafian follicles obtained 9 hours after the injection of human chorionic gonadotropin (hCG) into mature rabbits were dissected into a follicular fluid component, a granulosa cell-oocyte component, and a residual wall component, (the latter containing mostly theca tissue with a small and variable amount of adhering granulosa cells). The amounts of PGE and PGF were determined for each component. The follicular fluid contained approximately 4–10 times more PGE and PGF than either the granulosa cell-oocyte component or the residual wall component. The latter two components contained approximately equal amounts of these prostaglandins. The $\text{in vitro}$ biosynthesis of PGE and PGF was also studied and it was found that the granulosa cell-oocyte component had about 4 fold the capacity of the residual wall, and that the follicular fluid synthesized no prostaglandins. There was no significant effect of LH on either PGE or PGF synthesis in any of the components.