Selective stimulation of venous prostaglandin E 9-ketoreductase by bradykinin.

The effects of bradykin on prostaglandin metabolism in canine mesenteric vessels were examined. Bradykinin stimulated microsomal prostaglandin synthesis in both artery and vein; this stimulation was more pronounced when [14C] hosphatidylcholine rather than [14C] arachidonate was used as the substrate for prostaglandin synthetase. This suggested that bradykinin enhanced a membrane phospholipase. In addition, bradykinin selectively stimulated prostaglandin E 9-ketoreductase activity from veins but not arteries. This may explain the finding that bradykinin induces the release of prostaglandin E compounds from arteries but prostaglandin F compounds from veins.     

Endogenous acetylcholine release and labelled acetylcholine formation from [3H]choline in the myenteric plexus of the guinea-pig ileum.

The spontaneous release of acetylcholine (ACh) from the guinea-pig myenteric plexus - longitudinal muscle preparation superfused at a constant rate in the presence of physostigmine was 10 nmol-g-1-h-1. This release was decreased to one-third by tetradotoxin or by MnCl2 and increased 2.5 times by 0.1 Hz and 20 times by 16 Hz stimulation. The formation of [3H]ACh from [3H]choline increased from 3 to 33 nmol-g(-1)-h(-1) when the concentration of [3H]choline was increased from 1 muM to 50 muM. The rate of [3H]ACh formation was not affected by tetrodotoxin, MnCl2, or physostigmine in the absence of stimulation. It was increased by 50% by 0.1 Hz and by 100% by 16 Hz stimulation during the first 9 min of exposure to [3H]choline but not subsequently. The myenteric plexus - longitudinal muscle preparation contains 200 nmol/g choline. Results suggest that the apparent small [3H]ACh formation from low concentrations of [3H]choline is due to the dilution of [3H]choline by endogenous choline. The major part of [3H]ACh formation appears to be due to the intracellular turnover of ACh while the evoked release of [3H]ACh appears to originate from a small pool.     

Evidence for coupling of bradykinin receptors to a guanine-nucleotide binding protein to stimulate arachidonate liberation in the osteoblast-like cell line, MC3T3-E1.

The effect of bradykinin on the activation production of inositol 1,4,5-trisphosphate and prostaglandin E2 (PGE2) was examined in the murine osteoblastic cell line, MC3T3-E1. Bradykinin, at concentrations ranging from 1 to 1000 nM, stimulated the production of inositol 1,4,5-trisphosphate 2.5- to 3-fold within 10 s, and elevated cytosolic-free Ca2+, even in the absence of external Ca2+. This process is mediated through the activation of phospholipase C. Bradykinin at the same concentration also stimulated the production of PGE2 and caused a release of 3H radioactivity from the cells prelabeled with [3H]arachidonic acid, probably via the activation of phospholipase A2. Pretreatment of the cells with pertussis toxin inhibited the stimulation of PGE2 production and 3H radioactivity release, while the elevation in cytosolic Ca2+ and the production of inositol 1,4,5-trisphosphate were not altered by toxin-pretreatment. The addition of an unhydrolyzable analog of GTP, guanosine 5'-[gamma-thio]triphosphate (GTP gamma S) to the beta-escin-permeabilized cells prelabeled with [3H]arachidonic acid enhanced the release of 3H radioactivity. The simultaneous presence of bradykinin with GTP gamma S further activated the 3H radioactivity release in the beta-escin-permeabilized cells. These results provide evidence that receptors for bradykinin in the MC3T3-E1 couple stimulating arachidonate release, probably via the activation of phospholipase A2, through a guanine nucleotide binding protein sensitive to pertussis toxin.     

Alpha 1-adrenergic stimulation of arachidonic acid release and metabolism in a rat thyroid cell line. Mediation of cell replication by prostaglandin E2

The rat thyroid cell line, FRTL-5, expresses an alpha 1-adrenergic receptor when exposed to thyrotropin. We have found that occupation of this alpha 1-adrenergic receptor by norepinephrine stimulated the release of [3H]arachidonic acid from prelabeled cells. Arachidonic acid was metabolized primarily to prostaglandin E2 and to much smaller amounts of 11-hydroxy-5,8,11,13-eicosatetraenoic acid, 15-hydroxy-5,8,11,13-eicosatetraenoic acid, prostaglandin D2, and thromboxane B2. Synthesis of all these metabolites was inhibited by the cyclooxygenase inhibitor indomethacin. When FRTL-5 cells were starved of thyrotropin for 24 h, norepinephrine nearly doubled [3H]thymidine uptake into DNA. Cyclooxygenase inhibitors inhibited norepinephrine-stimulated thymidine uptake by 60-70%. Of several arachidonic acid metabolites tested, none was able to stimulate thymidine uptake directly in the presence of indomethacin. Prostaglandin E2, however, was able to restore [3H]thymidine uptake when added together with norepinephrine in the presence of indomethacin. Thus, occupation of an alpha 1-adrenergic receptor in a functional rat thyroid cell line leads to arachidonic acid release. Subsequent metabolism of the arachidonic acid by the cyclooxygenase pathway leads to synthesis of prostaglandin E2, which mediates a norepinephrine-stimulated activity related to cell replication.     

Selective stimulation of venous prostaglandin E 9-ketoreductase by bradykinin

The effects of bradykin on prostaglandin metabolism in canine mesenteric vessels were examined. Bradykinin stimulated microsomal prostaglandin synthesis in both artery and vein; this stimulation was more pronounced when [¹⁴C]phosphatidylcholine rather than [¹⁴C]arachidonate was used as the substrate for prostaglandin synthetase. This suggested that bradykinin enhanced a membrane phospholipase. In addition, bradykinin selectively stimulated prostaglandin E 9-ketoreductase activity from veins but not arteries. This may explain the finding that bradykinin induces the release of prostaglandin E compounds from arteries but prostaglandin F compounds from veins.     

Dissociation of tumour-promoter-induced effects on prostaglandin release, polyamine synthesis and cell proliferation of 3T3 cells.

The phorbol ester 12-O-tetradecanoylphorbol 13-acetate induces tumour promotion, inflammation, cell proliferation and prostaglandin release. Recent reports suggest that the prostaglandins released by 12-O-tetradecanoylphorbol 13-acetate (TPA) initiate a cascade of events leading to polyamine synthesis and cell proliferation. In experiments designed to test this contention, it was found that addition of TPA (1 microM to 1 nM) to confluent mouse 3T3 fibroblasts successively caused the release of prostaglandins E2 and I2, induction of the enzyme ornithine decarboxylase (EC 4.1.1.17), stimulation of [3H]thymidine incorporation into DNA, and cell proliferation. Pretreatment of the cells with the anti-inflammatory steroid dexamethasone (1 microM) or the non-steroidal anti-inflammatory drug indomethacin (1 microM) inhibited TPA-induced prostaglandin release. However, dexamethasone enhanced the other effects of TPA, whereas indomethacin was ineffective. Addition of prostaglandin E2 to the cultures did not induce ornithine decarboxylase activity and cell proliferation. Pretreatment of the cells with 1,3-diaminopropane (1 mM) or alpha-methylornithine (5 mM), inhibitors of polyamine synthesis, decreased TPA-induced ornithine decarboxylase activity without affecting DNA synthesis. TPA stimulated [3H]thymidine incorporation into DNA, even when the ornithine decarboxylase activity was completely blocked. These data suggest that the proliferative effect of TPA on 3T3 cells is independent of prostaglandin release and polyamine synthesis.     

Decrease of Acetylcholine Release from Cortical Slices in Aged Rats: Investigations into Its Reversal by Phosphatidylserine

The release of total acetylcholine (ACh) and [3H]ACh was investigated in electrically stimulated cortical slices prepared from 4- and 18-month-old male Wistar rats. The slices were prelabeled with [3H]choline ([3H]Ch) and perfused with Krebs solution containing physostigmine. Total ACh was measured and the nature of the tritium efflux identified by HPLC. The total tritium content in the slices at the end of the incubation period was half as great in the old as in young rats. A linear relationship was found between stimulation frequencies (2, 5, and 10 Hz) and fractional [3H]ACh release in both young and old rats. In the latter the release was significantly smaller. At 10 Hz stimulation frequency the ratio between the two 2-min stimulation periods, S2/S1, was higher in the 18-month-old rats than in the young rats. Specific activity of the evoked ACh release was significantly smaller in S2 than in S1 in 4-month-old rats only. These findings indicate that the young synthetize ACh from endogenous unlabeled Ch more than older rats. In 18-month-old rats both the evoked total ACh and [3H]ACh release, expressed as picograms per minute, showed an approximately 50% decrease in both S1 and S2 stimulation periods, with no significant difference in specific activity. Phosphatidylserine (PtdSer) administration (15 mg/kg, i.p. daily) for 1 week to 18-month-old rats prevented the reduction in total evoked ACh release but not the reduction in evoked [3H]ACh release. The specific activity of ACh release was therefore significantly smaller than that of the young and untreated old rats.(ABSTRACT TRUNCATED AT 250 WORDS)     

Acetylcholine Mobilization in a Sympathetic Ganglion in the Presence and Absence of 2-(4-Phenylpiperidino)Cyclohexanol (AH5183)

The present experiments measured the release of acetylcholine (ACh) by the cat superior cervical ganglia in the presence of, and after exposure to, 2-(4-phenylpiperidino)cyclohexanol (AH5183), a compound known to block the uptake of ACh by cholinergic synaptic vesicles. We confirmed that AH5183 blocks evoked ACh release during preganglionic nerve stimulation when approximately 13-14% of the initial ganglial ACh stores had been released; periods of rest in the presence of the drug did not promote recovery from the block, but ACh release recovered following the washout of AH5183. ACh was synthesized in AH5183-treated ganglia, as determined by the synthesis of [3H]ACh from [3H]choline, and this [3H]ACh could be released by stimulation following drug washout. The specific activity of the released ACh matched that of the tissue's ACh, and thus we conclude that ACh synthesized in the presence of AH5183 is a releasable as pre-existing ACh stores once the drug is removed. We tested the relative releasability of ACh synthesized during AH5183 exposure (perfusion with [3H]choline) and that synthesized during recovery from the drug's effects (perfusion with [14C]choline: the ratio of [3H]ACh to [14C]ACh released by stimulation was similar to the ratio in the tissue. These results suggest that the mobilization of ACh for release by ganglia during recovery from an AH5183-induced block is independent of the conditions under which the ACh was synthesized. Unlike nerve impulses, black widow spider venom (BWSV) induced the release of ACh from AH5183-blocked ganglia, even in the drug's continued presence. Venom-induced release of ACh from AH5183-treated ganglia was not less than the venom-induced release from tissues not exposed to AH5183. This effect of BWSV was attributed to the action of the protein, alpha-latrotoxin, because an anti-alpha-latrotoxin antiserum blocked the venom's action. ACh synthesized during AH5183 exposure was labelled from [3H]choline, and subsequent treatment with BWSV released [3H]ACh with the same temporal pattern as the release of total ACh. To exclude a nonexocytotic origin for the [3H]ACh released by BWSV, ganglia were preloaded with [3H]diethylhomocholine to form [3H]acetyldiethylhomocholine, an ACh analogue excluded from vesicles; the venom did not increase the rate of [3H]acetyldiethylhomocholine efflux. It is concluded that a vesicular ACh pool insensitive to the inhibitory action of AH5183 might exist and that this vesicular pool is not mobilized by electrical stimulation to exocytose in the presence of AH5183, but it is by BWSV.     

Storage and release of labelled acetylcholine in the myenteric plexus of the guinea-pig ileum.

Guinea-pig ileum myenteric plexus-longitudinal muscle preparation was superfused with [3H]choline for 15 min either without being stimulated or during field stimulation at 0.1 or 16 Hz; the preparation was then either removed immediately or after 75- or 135-min superfusion with hemicholinium-3 (HC-3) and the total acetylcholine (ACh) and [3H]ACh contents were determined. For measuring the release of [3H]ACh the preparation was stimulated for 60 min the second time at 0.1 or 16 HZ in the presence of hemicholinium. Exposure to [3H]choline without stimulation resulted in the formation of [3H]ACh stores which were maintained in the first 75 min but decreased therafter. Labelling during stimulation at 16 Hz produced the largest and best maintained [3H]ACh content. Following labelling during 0.1-Hz stimulation, more label could be released than following labelling in the absence of stimulation. Labelling during 16-Hz stimulation did not increase any further in fool of [3H]ACh accessible to release by 0.1-Hz stimulation, but caused a 2.5 times increase in the pool from which Hz stimulation released [3H]ACh. These results suggest that two populations of cholinergic neurons exist in the myenteric plexus, one activated only by high frequency stimulation, the other by both high and low frequency stimulation.     

Inhibition of the Electrically Evoked Release of [3H]Acetylcholine in Rat Striatal Slices: An Experimental Model for Drugs that Enhance Dopaminergic Neurotransmission

The activation by endogenous dopamine of the inhibitory 3,4-dihydroxyphenylethylamine (dopamine) receptors modulating the electrically evoked release of [3H]acetylcholine [( 3H]ACh) and [3H]dopamine in rat striatal slices is a function of the concentration of dopamine accumulated in the synaptic cleft during electrical stimulation. When the release of 3H-neurotransmitters was elicited with a 2-min period of stimulation at a frequency of 1 Hz, neither dopamine autoreceptors nor dopamine receptors modulating [3H]ACh were activated by endogenously released dopamine. On the other hand, exposure to (S)-sulpiride facilitated the release of [3H]dopamine and [3H]ACh elicited when the 2-min stimulation was carried out at a frequency of 3 Hz but this effect was not observed at a lower frequency of stimulation (1 Hz). In the presence of amphetamine the dopamine receptors modulating the electrically evoked release of [3H]ACh can be activated by endogenous dopamine even at the lower frequency of stimulation (1 Hz). Similar effects can be obtained if the neuronal uptake of dopamine is inhibited by cocaine or nomifensine. The inhibition by amphetamine of the release of [3H]ACh elicited by electrical stimulation at 1 Hz involves dopamine receptors and can be fully antagonized by clozapine, haloperidol, chlorpromazine, or pimozide. The stereoselectivity of this antagonism can be demonstrated with the optical enantiomers of sulpiride and butaclamol. This inhibitory effect of amphetamine on cholinergic neurotransmission appears to be the result of the stimulation of dopamine receptors of the D2 subtype, as they were resistant to blockade by the preferential D1 receptor antagonist SCH 23390.(ABSTRACT TRUNCATED AT 250 WORDS)     

Diacylglycerol stimulates phospholipase A2 from Swiss 3T3 fibroblasts

We recently demonstrated that diacylglycerol induced arachidonate release and prostaglandin E2 synthesis in 3T3 fibroblasts, and greatly augmented prostaglandin E2 synthesis in response to submaximal and maximal concentrations of bradykinin. We have now partially purified a phospholipase A2 from the cells. When phosphatidyl[3H]choline was used as substrate, several diacylglycerols augmented phospholipase A2 activity. Diacylglycerol was effective at concentrations as low as 30 nM. Protein kinase C inhibition did not affect diacylglycerol's stimulation of phospholipase A2. Diacylglycerol did not alter the calcium requirement for phospholipase A2 or its pH optimum. The present study demonstrates that the effect of diacylglycerol to augment arachidonate metabolism is at the level of phospholipase A2, itself.     

Inhibition of prostaglandin E2 synthesis by a blocker of epithelial chloride channels

Arginine-vasopressin (AVP) elicits a variety of responses in cultured rat mesangial cells, among them stimulation of prostaglandin biosynthesis and activation of Cl- channels. AVP produced an 11-fold increase over basal levels in prostaglandin E2 release from cultured mesangial cells. This response was completely inhibited by 25 microM indomethacin and 82 +/- 5% inhibited by 25 microM 5-nitro-2-(3-phenylpropylamino)-benzoic acid (NPPB) which is a potent blocker of epithelial Cl- channels. The IC50 for NPPB inhibition of prostaglandin E2 release was 8 microM. Indomethacin and NPPB at 25 microM also inhibited AVP-stimulated cellular accumulation of prostaglandin E2 by 98% and 79 +/- 7% respectively. The inhibitory effect of NPPB was not due to interference with the cellular response to AVP since at 50 microM it did not block AVP-stimulated release of arachidonate metabolites from cells metabolically labeled with [3H]-arachidonic acid. It is suggested that NPPB inhibition of prostaglandin E2 synthesis is at the cyclooxygenase level on the basis of its structural similarity to the fenamic acid type of cyclooxygenase inhibitors.     

Effects of bradykinin on DNA synthesis in resting NIL8 hamster cells and human fibroblasts

Bradykinin stimulates [3H]thymidine incorporation and DNA synthesis in resting, serum-deprived NIL8 hamster cells. The ED50 for this stimulation is 4.52 +/- 2.91 nM. Other kinin peptides including lys-bradykinin (kallidin) and met-lys-bradykinin also stimulate [3H]thymidine incorporation in the NIL8 cells, whereas desarg9-bradykinin is without effect, suggesting action of the kinin peptides through type B2 receptors. Bradykinin also stimulates DNA synthesis in IMR-90 human fibroblasts; however, this effect is observed only in the presence of indomethacin, which blocks prostaglandin synthesis. These results suggest that prostaglandins act as negative modulators of the growth-stimulatory effects of bradykinin in the fibroblasts. This conclusion is supported by the observation that exogenously added PGE1, PGE2, PGA1, PGA2, PGB1, and PGB2 strongly inhibit [3H]thymidine incorporation in the human fibroblasts. The direct effect of bradykinin observed in the NIL8 cells may be attributable to the relative resistance of these cells to growth inhibition by prostaglandins.     

Stimulation of anterior pituitary prostaglandin E content and somatotropin (growth hormone) synthesis by phospholipase A.

The prostaglandin E content of dispersed rat anterior pituitary glands was found to increase in the presence of phospholipase A or arachidonic acid. The increases were abolished by the addition of indomethacin. Similarly, the rate of somatotropin (growth hormone) synthesis was increased by these two agents, and the increases were again abolished by indomethacin. Phospholipase A also stimulated somatotropin release. The stimulation of prostaglandin E accumulation was a specific response to those fatty acids that are precursors for prostaglandin synthesis. One such precursor, [3H]arachidonic acid, was incorporated by rat anterior pituitary glands in vitro, and found to be associated mainly with phosphatidylethanolamine-like material. It is concluded that the intracellular concentration of prostaglandin E is limited by the availability of precursor fatty acids and that this can be increased by the addition of exogenous precursors or by the action of exogenous phospholipase A on the cellular phospholipid. Factors that increased prostaglandin E concentrations also increase the rate of synthesis of somatotropin, providing further evidence for the concept that prostaglandin E is involved in modulation of the rate of synthesis of this hormone.     

Biochemical evidence that acetylcholine release from cholinergic nerve terminals is mostly vesicular

The nature of the intraterminal compartments from which acetylcholine (ACh) is released following presynaptic stimulation was investigated. This was pursued by examining the effects of the anticholinergic drug 2-(4-phenylpiperidino)cyclohexanol (AH5183) on the release of newly synthesized [3H]ACh and of endogenous ACh from purified cholinergic nerve terminals (synaptosomes) which were isolated from the electric organs of Torpedo. Preincubation of the synaptosomes, with AH5183 (1-10 microM), does not affect either the intraterminal synthesis of [3H]ACh or the uptake of its precursors, but results in a marked inhibition (85%) of the release of the newly synthesized [3H]ACh. However, when AH5183 is added following the accumulation of [3H]ACh in the nerve terminals, it does not affect [3H]ACh release. AH5183 also has no effect on the release of preformed endogenous ACh. These findings, together with the previous in vitro demonstrations that AH5183 is a potent inhibitor of ACh uptake into isolated cholinergic vesicles, suggest that most of the synaptosomal ACh is secreted by a vesicular mechanism.     

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.     

Prostaglandin receptor-adenylate cyclase system in plasma membranes of rat liver and ascites hepatomas, and the effect of GTP upon it.

1. Adenylate cyclase in plasma membranes from rat liver was stimulated by prostaglandin E1, and to a lesser extent by prostaglandin E2. Prostaglandin F1alpha and A1 did not stimulate the cyclase. The prostaglandin E1-mediated activation was found to require GTP when the substrate ATP concentration was reduced from 3 mM to 0.3 mM in the reaction mixture. Adenylate cyclase of the plasma membranes from rat ascites hepatomas AH-130 and AH-7974 was not stimulated by prostaglandin E1 in the presence or the absence of GTP, although the basal activity of adenylate cyclase as well as its stimulation by GTP alone were similar to normal liver plasma membranes. 2. Liver plasma membranes were found to have two specific binders for [3H] prostaglandin E1 with dissociation constants of 17.6-10(-9) M and 13.6-10(8) M (37 degrees C) and one specific binder for [3H]prostaglandin F2alpha with a dissociation constant of 2.31-10(8) M (37 degrees C). The specific binders for prostaglandin E1 could not be detected in the hepatoma plasma membranes. 3. Binding of [3H] prostaglandin E1 to the liver plasma membranes was exchange by, GTP dGPT, GDP, ATP and GMP-P(N)P, but not by GMP, CGMP, DTTP, UTP or CTP. The increase in the binding of [3H] prostaglandin E1 was found to be due to the increased affinity of the specific binders to prostaglandin F2alpha was not affected by GTP. 4. GTP alone was found to increase V of adenylate cyclase of liver plasma membranes, while GTP plus prostaglandin E1 was found to decrease Km of adenylate cyclase in addition to the increase of V to a further extent.     

Effect of endogenous prostaglandins on acetylcholine release from dog trachealis muscle

We used a radioenzymatic technique to measure effects of the prostaglandin synthesis inhibitor indomethacin and of exogenous prostaglandin E2 (PGE2) and prostaglandin I2 (PGI2) on acetylcholine (ACh) efflux from canine tracheal smooth muscle (TSM) during sustained electrical field stimulation (EFS; 2 Hz, 2 ms pulse duration, 50 V for 15 min). ACh efflux from indomethacin (INDO, 10(-6) M)-pretreated and control TSM increased with consecutive stimulations. However, efflux of ACh was greater in INDO-treated than control muscles. INDO increased the tension produced by TSM in response to EFS. Neither PGE2 (10(-8) M) nor PGI2 (10(-6) M) had any effect on ACh efflux from INDO-pretreated TSM during the first of three periods of EFS. However, PGI2 and PGE2 prevented the progressive increase in ACh efflux observed on subsequent stimulations. PGE2 but not PGI2 decreased contractions of TSM caused by EFS. Our results demonstrate that endogenous prostaglandins, probably PGE2, do inhibit EFS-evoked ACh release from canine TSM in vitro, but suggest that these prostaglandins modulate EFS-evoked contractions predominantly by postsynaptic mechanisms.     

Stimulatory effect of prostaglandin E2 on 1 alpha,25-dihydroxyvitamin D3 synthesis in rats.

The effect of prostaglandin E2 on accumulation in plasma of 1 alpha,25-dihydroxy[3H]vitamin D3 from 25-hydroxy[3H]vitamin D3 was studied in vivo using vitamin D-deficient thyroparathyroidectomized rats. Intra-arterial infusion of 10-50 micrograms of prostaglandin E2/h caused a significant stimulation of 1 alpha,25-dihydroxy[3H]vitamin D3 production. No significant changes in plasma Ca2+ and Pi concentrations or urinary cyclic AMP excretion were observed after prostaglandin E2 infusion. Theophylline did not enhance the effect of a submaximal dose of prostaglandin E2 on 1 alpha,25-dihydroxy[3H]vitamin D3 production. These data indicate that prostaglandin E2 stimulates plasma accumulation of 1 alpha,25-dihydroxy[3H]vitamin D3 independent of the adenylate cyclase/cyclic AMP system, and suggest that prostaglandin E2 has a modulatory role in the regulation of 25-hydroxyvitamin D3 1 alpha-hydroxylase in the kidney.     

Prostaglandin generation in rabbit kidney. Hormone-activated selective lipolysis coupled to prostaglandin biosynthesis.

The endogenous release of prostaglandins and free fatty acids from the isolated perfused rabbit kidney in the absence or presence of stimulation by bradykinin or angiotensin-II was investigated. Basal (nonstimulated) release of prostaglandin-precursor arachidonic acid was 15-20-fold higher than that of prostaglandin E2 indicating a low conversion of released arachidonate to prostaglandins. Addition of bovine serum albumin to the perfusion medium caused a substantial (50-250%) increase in the release of all fatty acids except myristic and arachidonic acids, and no significant change in prostaglandin E2 generation. In contrast, administration of bradykinin (0.5 microgram) or angiotensin-II (1 microgram) caused a 10-15-fold increase in prostaglandin E2 release, and with albumin present, also a 2-3-fold selective increase in arachidonic acid release. Thus, unlike what was observed under basal conditions, arachidonic acid released following hormone stimulation is efficiently converted to prostaglandin E2. We conclude that administration of bradykinin or angiotensin-II into the perfused kidney activates a lipase which selectively releases arachidonic acid, probably from a unique lipid entity. This lipase reaction is tightly coupled to a prostaglandin generating system so that the released arachidonate is first made available to the prostaglandin cyclooxygenase, resulting in its substantial conversion to prostaglandins.