Renal sodium handling and stimulation of medullary Na-K-ATPase during blockade of prostaglandin synthesis

The effect of suppression of prostaglandin synthesis on renal sodium handling and microsomal Na-K ATPase was studied in control and indomethacin treated intact rats maintained on a normal sodium diet (series A) and chronically salt loaded (series B). Indomethacin administration resulted in a decreased GFR and a significantly depressed urinary excretion and an increased fractional reabsorption of sodium in animals fed the normal sodium diet or chronically salt loaded. In rats maintained on a normal Na diet, the activity of the renal medullary Na-K ATPase after indomethacin was 206.3 +/- 6.4 ug Pi/mg protein, i.e. significantly higher as compared with the enzyme activity in the medullary renal fraction from control animals in which it averaged 148 +/- 7.79 ug Pi/mg protein (p less than 0.001). While after chronic salt load a similar increment in the activity of renal medullary Na-K ATPase was observed, no additional stimulation was elicited by subsequent indomethacin administration. The addition of exogenous PGE2, 0.1 mM to microsomal fractions obtained from kidneys of normal rats, was associated with a moderate suppression of the medullary Na-K-ATPase activity, from a basal level of 170 +/- 16 to 151.3 +/- 13 umol Pi/mg protein/hr (p less than 0.005). In isolated segments of medullary thick ascending limb of Henle's loop (MTAL) addition of PGE2 to the incubation medium resulted in a significant inhibition of Na-K ATPase from 37.2 +/- 2 to 21.25 +/- 1.17 x 10(-11) mol/mm/min (p less than 0.0001). These findings suggest that the increased renal Na reabsorption after inhibition of PG synthesis might be related, at least partly, to stimulation of medullary Na-K ATPase. In parallel, the reported natriuretic effect of prostaglandins might imply a direct inhibitory effect of these mediators on renal Na-K ATPase.     

Modulation of prostaglandin of the renal vascular action of arginne vasopressin

To determine whether the renal vascular effect of arginine vasopressin (AVP) is modulated by renal prostaglandin E₂ (PGE₂) were determined during the infusion of AVP in dogs during control conditions and after the administration of the inhibitor of prostaglandin synthesis, indomethacin. During control conditions, intrarenal administration for 10 min of a dose of AVP calculated to increase arterial renal plasma AVP concentration by 75 pg/ml produced a slight renal vasodilation (p<0.01) and an increase in renal venous plasma concentration of PGE₂. Renal venous PGE₂ concentration during control and AVP infusion averaged 33 ± 7 and 52 ± 12 pg/ml (p<0.05), respectively. After administration of indomethacin, the same dose of AVP induced renal vasoconstriction (p<0.05) and failed to enhance renal venous PGE₂ concentration (9 ± 1 to 8 ± 1 pg/ml). Intrarenal administration of 20 ng/kg. min of AVP for 10 min induced a marked renal vasoconstriction (p<0.01) and increased renal venous plasma PGE₂. Renal venous PGE₂ during control and AVP infusion averaged 31 ± 10 and 121 ± 31 pg/ml (p<0.01), respectively. Administration of the same dose of AVP following indomethacin produced a significantly greater and longer lasting renal vasoconstriction (p<0.01) and failed to increase renal venous plasma PGE₂ (10 ± 1 to 9 ± 1 pg/ml). These results indicate that a concentration of AVP comparable to that observed in several pathophysiological conditions induces a slight renal vasodilation which is mediated by renal prostaglandins. The results also indicate that higher doses of AVP induce renal vasoconstriction and that prostaglandin synthesis induced by AVP attenautes the renal vasoconstriction produced by this peptide.     

Effect of chronic salt loading on renal Na-K-ATPase activity in the rat

The effect of chronic salt loading in rats fed regular chow diet on renal Na-K-ATPase was studied. The high salt intake was associated with increased filtered load of sodium (control: 126 +/- 3.9 mueq/min, salt loaded: 146 +/- 2.5, mueq/min, P less than 0.001), increased net reabsorption of sodium (control: 125.3 +/- 3.9 mueq/min, salt load: 134.8 +/- 2.4 mueq/min, P less than 0.05), increased urinary excretion of potassium (control: 2.4 +/- 0.09 mueq/min/min; salt loaded: 3.0 +/- 0.1 mueq/min, P less than 0.001) and increase in single kidney weight (control: 0.798 +/- 0.010 g, salt loaded: 0.937 +/- 0.015 g, P less than 0.001). The above mentioned changes were associated with significant increase in renal microsomal and whole homogenate medullary Na-K-ATPase activity in the salt loaded group (microsomes: control 74.1 +/- 4.9 mumole Pi/mg prot/hr, salt loaded 112.7 +/- 6.0 mumole Pi/mg prot/hr, P less than 0.001; whole homogenate: control 22.7 +/- 1.0 mumole Pi/mg prot/hr, salt load 29.4 +/- 1.6 mumole Pi/mg prot/hr, P less than 0.005), while cortical and papillary Na-K-ATPase activity remained unchanged. Taken together, these results show that increased filtered and reabsorbed load of sodium, which follows high salt intake, is associated with an increased renal Na-K-ATPase activity. The preferential rise in medullary enzymatic activity may be interpreted as suggesting that these changes may stem from increased delivery and reabsorption of sodium in the ascending limb of Henle's loop.     

Interaction between prostaglandin E2 and leukotriene D4 on the excretion of electrolytes by the dog kidney in vivo

Recent experiments indicate that prostaglandin E₂ potentiates the vasodilatory properties of leukotrienes in the skin microcirculation. The present experiments were undertaken to study the effect of leukotriene D₄ and prostaglandin E₂ on renal hemodynamics and urinary electrolytes in the dog. Experiments were performed in three groups of anesthetized Mongrel dogs: the first group was studied under hydropenia, whereas the two remaining groups were studied during water diuresis with (Group 3) or without indomethacin (Group 2). LTD₄ (100ng/min) and PGE₂ (3ug/min) were infused in the left renal artery to minimize systemic effects of these compounds. LTD₄ alone failed to influence urinary sodium excretion in all 3 groups. In Group 1, urinary sodium increased from 77 ± 6 to 393 ± 74uEq/min during PGE₂, and further increased to 511 ± 52uEq/min during LTD₄ + PGE₂. No change occured in the contralateral right kidney. In this group, glomerular filtration as well as renal plasma flow were not statistically influenced. In Group 2, the same phenomenon was observed for urinary sodium. The combined infusion of LTD₄ + PGE₂ increased urinary sodium without significant changes in glomerular filtration and renal plasma flow. Finally, in Group 3, indomethacin was shown to reduce the natriuretic effects of LTD₄ and PGE₂: during PGE₂ alone, urinary sodium increased from 90 ± 14 to 260 ± 66uEq/min, and only rose from 80 ± 10 to 175 ± 19uEq/min during the combined infusion of LTD₄ and PGE₂. In groups 2 and 3, free water clearance was utilized as an index of sodium chloride reabsorption in the thick ascending limb: this parameter increased from 2.35 ± 0.25 to 4.70 ± 0.30ml/min, while urinary volume was increasing from 3.55 ± 0.25 to 10.05 ± 0.65ml/min, during LTD₄ + PGE₂. Indomethacin, administered in Group 3, (3mg/kg/hr) again abolished the effect of combined PGE₂ + LTD₄. These results indicate a potentiating effect of leukotriene D₄ on the PGE₂-induced natriuresis in the anesthetized dog. These phenomena occured in the absence of significant changes in renal hemodynamics, therefore suggesting a direct tubular effect of these arachidonic acid metabolites. Finally, the water diuresis experiments suggest a proximal site of action of PGE₂ and LTD₄.     

Prostaglandin (PG) analysis in urine of humans and rats by different radioimmunoassays: Effect on PG-excretion by PG-synthetase inhibitors, laparotomy and furosemide

Thw radioimmunological (RIA) determination of prostaglandin (PG) E₂ and of PGF_2α in urine humans and rats is described in detail. After extraction and chromatography PGE₂ was determined by using a PGE specific antibody or by using either PGB or PGF_2α specific antibodies after the respective conversion procedures. The three different RIA procedures were compared to each other. PGF_2α was determined by a specific antibody to PGF_2α. Basal excretion of PGE₂ and of PGF_2α in healthy women on free diet was 9.3 ng/hour ± 0.96 and 18.3 ng/hour ± 2.5 respectively. Furosemide increased the excretion of PGE₂ and of PGF_2α in humans significantly, while PG-excretion rates decreased on indomethacin. In rat urine PGE₂ and PGE_2α increased markedly from 46.2 pg/min ± 9.3 and 27 ± 3.4 to 253.8 ± 43.3 and 108 ± 12.6 pg/min (per one kidney) in the anesthetized-laparotomized animal. This increase was abolished after giving two different PG synthetase inhibitors.     

Sex differences in gastric mucosal protection after 16, 16-dimethyl PGE2 and lithium chloride

While the incidence of duodenal ulcer disease has been documented to be greater in men than in women, this observation has not been previously noted in animal studies of the upper gastrointestinal tract. In this study, we questioned whether the cytoprotective properties of 16, 16-dimethyl PGE₂ were sex-related by comparing the degree of ethanol-induced hemorrhagic gastritis in male and female rats pretreated with 16,16-dimethyl PGE₂ or lithium chloride. Animals receiving 16,16-dimethyl PGE₂ or lithium chloride had significantly less ethanol-induced hemorrhagic gastritis (1.17±0.15 and 1.24±0.13, respectively, p<0.001) when compared with controls (2.69±0.10). Female rats treated with 16,16-dimethyl PGE₂ had 59% less hemorrhagic gastritis than male rats treated similarly (0.76±0.14 vs 1.86±0.19 respectively, p<0.001). This sex-related difference in hemorrhagic gastritis was not noted in male and female rats receiving lithium chloride (1.24±0.15 vs 1.23±0.27, respectively). However, female rats treated with 16, 16-dimethyl PGE₂ had significantly less hemorrhagic gastritis when compared with female rats receiving lithium chloride (0.76±0.14 vs 1.24±0.15 respectively, p<0.05).These findings suggest that the protective properties of 16, 16-dimethyl PGE₂ are sex-related while those of lithium chloride are not.     

Bicarbonate-activated ATPase activity in renal cortex of chronically acidotic rats.

The effect of chronic NH4Cl-induced acidosis on the activity of a bicarbonate-activated component of ATPase was studied in homogenates of renal tissue from Wistar rats. This particular component of ATPase, which is maximally stimulated by 50 mM bicarbonate, and is insensitive to the action of ouabain, has been implicated in the active transport of bicarbonate in various tissues. The activity of this enzyme in cortical homogenates from an acidotic group of animals was 4.3 +/- 0.4 mumol Pi/mg protein per hour compared with 5.8 +/- 0.3 mumol Pi/mg protein per hour in a control group (p less than 0.02). No significant change in bicarbonate ATPase activity was observed in medullary homogenates, and NaK-ATPase activity remained the same in cortex and medulla of both groups. Subcellular fractionation of the cortical tissue homogenates revealed that bicarbonate ATPase activity in a microsomal fraction from acidotic animals was 6.5 +/- 1.1 mumol Pi/mg protein per hour compared with 9.4 +/- 1.2 mumol Pi/mg protein per hour in control animals (p less than 0.02). Bicarbonate ATPase activity in other subcellular fractions was not different in the two groups of animals. These findings are compatible with the hypothesis that a certain percentage of bicarbonate reabsorption in the nephron is mediated by a bicarbonate-activated component of ATPase.     

Prostaglandin stimulation of ornithine decarboxylase activity in mammary gland explants from mid-pregnant mice

The effects of various prostaglandins on ornithine decarboxylase (ODC) activity in mammary gland explants from mid-pregnant mice have been tested. PGE₁, E₂ and I₂ elicit a concentration-dependent stimulation of ODC activity. The minimally effective concentrations are 0.5 ug/ml for PGE₁ and E₂, and 50 ug/ml for PGF_2α and 6-keto-PGF_1α. The PGE₁ effect had a time course identical to that of prolactin. The prolactin action on ODC activity was attentuated by indomethacin, an inhibitor of prostaglandin biosynthesis. Arachidonic acid stimulation ODC activity and its effect was abolished by indomethacin. The phosphodiesterase inhibitor, 3-isobutyl-1-methylxanthine, potentiated the PGE₁ effect on ODC activity. The results suggest that the prostaglandins may modulate prolactin's action of ODC activity via a cAMP dependent mechanism.     

The effect of suppression of prostaglandin synthesis on renal function in rats with intact and reduced renal mass

The present study was undertaken to assess the role of prostaglandin system in the compensatory response to reduced nephron population, respective to renal function and electrolyte excretion. Intact and nephrectomized rats were divided in 4 groups: 1) rats pretreated with indomethacin, 2) rats pretreated with the vehicle of indomethacin, 3) rats pretreated with sulindac, and 4) rats pretreated with the vehicle of sulindac.In normal rats, indomethacin administration resulted in a mild decrease in creatinine clearance and a significant reduction of the urinary Na excretion. In the rats with reduced renal mass treated with indomethacin, the creatinine clearance did not differ from that in the control group. The 24 h urinary sodium excretion and the fractional excretion of sodium, however, were significantly lower in the indomethacin treated animals than in the control rats. No change in the creatinine clearance or in the sodium excretion was observed in all groups pretreated with sulindac.The urinary PGE₂ and thromboxane excretion was significantly lower in the indomethacin treated intact rats and the rats with reduced renal mass. Sulindac induced a slight decrease in urinary excretion of PGE₂ in intact rats. No significant change in urinary excretion of PGE₂ or thromboxane was seen after sulindac in the rats with reduced renal mass.The antinatriuretic effect of indomethacin was dissociated from changes in urine flow in all groups of animals, suggesting that the increase in Na reabsorption tool place in a water impermeable segment of nephron.These results suggest that the compensatory increase in urinary Na excretion per nephron in rats with reduced nephron population at least partly depends on an intact prostaglandin synthesis.     

Salicylate nephropathy in the Gunn rat: Potential role of prostaglandins

We examined the potential role of prostaglandins in the development of analgesic nephropathy in the Gunn strain of rat. The homozygous Gunn rats have unconjugated hyperbilirubinemia due to the abscence of glucuronyl transferase, leading to marked bilirubin deposition in renal medulla and papilla. These rats are also highly susceptible to develop papillary necrosis with analgesic administration.We used homozygous (jj) and phenotypically normal heterozygous )jJ) animals. Four groups of rats (n = 7) were studied: jj and jJ rats treated either with aspirin 300 mg/kg every other day or sham-treated. After one week, slices of cortex, outer and inner medulla from one kidney wre incubated in buffer and prostaglandin synthesis was determined by radioimmunoassay. The other kidney was examined histologically.A marked corticomedullary gradient of prostaglandin synthesis was observed in all groups, PGE2 synthesis was significantly higher in outer medulla, but not cortex or inner medulla, of jj (38 ± 6 mg/mg prot) than jJ rats (15 ± 3) (p<0.01). Aspirin treatment reduced PGE2 synthesis in all regions, but outer medullary PGE2 remained higher in jj (18 ± 3) than jJ rats (9 ± 2) (p<0.05). PGE2α was also significantly higher in the outer medulla of jj rats with and without aspirin administration (p<0.05). The changes in renal prostaglandin synthesis were accompanied by evidence of renal damage in aspirin-treated jj but not jJ rats as evidenced by: increased incidence and severity of hematuria (p<0.01); increased serum creatinine (p<0.05); and increase in outer medullary histopathologic lesions (p<0.005 compared to either sham-treated jj or aspirin-treated jJ).These results suggest that enhanced protaglandin synthesis contributes to maintenance of renal function and morphological integrity, and that inhibition of protaglandin synthesis may lead to pathological renal medullary lesions and deterioration of renal function.     

Defibrotide, by enhancing prostacyclin generation, prevents endothelin-1 induced contraction in human saphenous veins

Spirals of human saphenous veins (HSV), mounted in a 5 ml organ bath containing Krebs-Henseleit solution (37°C), when kept in contact with defibrotide (100–200 ug/ml) for 15 min, enhance (2 and 3 fold) their own basal release of 6-keto-PGF_1α (61 ± 1.3 pg/mg w.t. n = 12). The phenomenon was long lasting upon repeated washing and sensitive to indomethacin (1 ug/ml). Endothelin-1 (ET-1, 20–40 ng) induced a sustained contraction of HSV and concomitantly released from the venous tissue a proportional amount of 6-keto-PGF_1α.Indomethacin (1 ug/ml), by inhibiting cyclo-oxygenase enzyme, potentiated the contractile activity of ET-1 in HSV whereas exogenous PGE₂ (20 ng/ml) considerably reduced the tension developed by the peptide on this venous tissue.Defibrotide (200 ug/ml), by releasing 6-keto-PGF_1α, and other vasoactive prostaglandins, antagonized the contractile effect ET-1 (20 ng) in HSV. This data indicates that the eicosanoid metabolism is involved in the modulation of the potent vasoconstrictor effect of ET-1 in HSV and that PGI₂-releaser, such as defibrptide, may have therapeutical value against immoderate changes of venous tone.     

Influence of sex hormones on prostaglandin dehydrogenase activity in the rat uterus

The present experiments report the effects of estradiol or of progesterone on the activity of 15-prostaglandin-dehydrogenase (PGDH) in the uterus of spayed rats. When the substrate was PGF_2α the treatment with progesterone (4 mg.day⁻¹, two days) or with estradiol-17-beta (0.5 ug + 1 ug) did not show any effect on the activity of the enzyme. On the contrary, uteri from ovariectomized rats injected with a higher dose of estradiol- 17-beta (0.5 ug + 50 ug) exhibited a significant increment. When the substrate was PGE₂, progesterone failed again to modify the enzyme activity, whereas estradiol, both at a low and at a high doses, enhanced significantly the uterine PGDH activity. The possibility of two different PGDHs for each PG and the role of estradiol in enhancing PGE₂ catabolism into 15-keto- PGE₂ as a mechanism subserving the effect of estrogens on the output of this PG in the rat uterus, are discussed.     

Role of renal prostaglandins in the fall in urine osmolality after papillary micropuncture

The effect of micropuncture of the renal papilla through an intact ureter on urinary concetrating ability of rats was examined. Micropuncture of the renal papilla caused a fall in urine osmolality in the punctured kidney from 1718 ± 106 to 1035 ± 79 mosmol/kg·H₂O. In order to investigate the role of renal prostaglandins in this process, PGE₂ excretion was measured and found to increase from 63.4 ± 14.0 to 205.5 ± 57.1 pg/min. Urine osmolality and PGE₂ excretion from the contralateral kidney were not significantly altered. In animals given meclofenamate (2 mg/kg·hr), renal PGE₂ excretion was reduced to 22.3 ± 5.1 pg/min prior to micropuncture and it remained low at 8.9 ± 1.8pg/min after papillary micropuncture. Meclofenamate also blocked the fall in urine osmolality caused by micropuncture of the renal papilla, with urine osmolality averaging 1940 ± 122 before and 1782 ± 96 mosmol/kg·H₂O after the micropuncture. These results indicated that papillary micropuncture through an intact ureter increased renal PGE₂ excretion and that a rise in renal production of PGE₂ or some other prostanoid is associated with a fall in urine concentrating ability.     

Prostaglandin synthesis by enterocyte microsomes of rabbit small intestine

We studied the prostaglandin (PG) synthetic capacity of microsomes of a relatively pure population of rabbit enterocytes and determined ideal conditions for product synthesis. The epithelial cells were freed from the basement membrane by a combination of calcium chelation and mechanical vibration, and 100,000 x g microsomes were prepared. These microsomes were found to synthesize PG from exogenously added arachidonic acid. The ideal conditions for the reaction were a microsomal protein concentration of 1.0 mg/ml, an arachidonic acid concentration of 33 μM, a reaction mixture pH of 8.0 − 9.5 and with epinephrine 1.5 mM added as a cofactor. The product yields increased linearly with time up to 30 min, of incubation and were inhibited by 100 μM indomethacin. Under the above ideal conditions enterocyte microsomes yielded the following products expressed as pmole/mg protein/20 min, incubation: PGF_2α 98±7, PGE₂ 48±9, PGD₂ 28±7, TxB₂ 40±5, 6 Keto PGF_1α 15 ± 6.     

The effect of adrenergic stimulation on urinary prostaglandin E2 and 6 keto PGF1α in man

To evaluate the details of the adrenergic stimulation of urinary prostaglandins in man, ten normal volunteers were given various agonists and antagonists. The effect of 4 hour IV infusions of norepinephrine (NE), NE + phentolamine (PHT), NE + phenoxybenzamine (PHB), NE + prazosin (PZ), isoproterenol (ISO), and PHT alone on urinary PGE₂ and PGI₂ (6 keto PGF_1α) were determined. PGE₂ and 6 keto PGF_1α were measured by radioimmunoassay from 4 hour urine samples. NE stimulated both PGE₂ (196±40 to 370±84 ng/4 hrs/g creatinine and 6 keto PGF_1α(184±30 to 326±36), both p<0.01. In contrast, ISO had no effect on either PGE₂ or 6 keto PGF_1α excretion. Alpha blockade with PHT. PHB, or PZ inhibited the NE induced systemic pressor effect. However, the effect of the alpha blockers on the NE induced stimulation of PGE₂ and 6 keto PGF_1α varied. PHT did not alter the NE stimulated PGE₂ or 6 keto PGF_1α release (370±84 vs. 381±80) PGE₂ and (326±50 vs. 315±40) 6 keto PGF_1α, both p>0.2). PHT alone stimulated only 6 keto PGF_1α. PHB and the specific α₁ antagonist PZ similarly eliminated the NE induced prostaglandin release. These results suggest that adrenergically mediated urinary prostaglandin release in man is via an alpha receptor with α₁ characteristics.     

Metabolism of [C] arachidonic acid in the isolated rabbit spleen

Infusion of [¹⁴C] arachidonic acid (AA) into the isolated, Tyrode perfused rabbit spleen resulted in the release of a substance into the venous effluent with the musculotropic activity and chromatographic properties of prostaglandin (PG)E₂. Smaller amounts of radioactive materials with the chromatographic properties of PGF_2α, 6-keto-PGF_1α, and PGD₂ were also released. The radiolabeled material released in largest amounts from the spleen was identified as PGE₂ on the basis of: 1) Co-chromatography with PGE₂ in three solvent systems, 2) Conversion of the radioactive material and of authentic [³H] PGE₂ to similar products by treatment with sodium borohydride and with potassium hydroxide, and 3) Stability of the musculotropic activity in Tyrode solution at 37°C. Release of the major and minor radioactive products was inhibited by pretreatment of the spleen with either indomethacin or 5,8,11,14-eicosatetraynoic acid.     

Thromboxane receptor blockade improves cyclosporine nephrotoxicity in rats

Cyclosporine A (CyA) nephorotoxicity is associated with impaired renal hemodynamic funtion and increased production of the vasoconstrictor eicosanoid thromboxane A₂ (TxA₂). In CyA toxic rats, renal dysfunction cna be partially reversed by inhibitors of thromoboxane sysnthase. However, interpretation of these results is complicated since inhibitance of thromboxane synthase may cause accumulation of prostaglandin endoperoxides that can act as partial agonists at the TxA₂ receptor and may blunt the efficacy of treatment. Furthermore, these endoperoxides may be used as substrate for production of vasodilator prostaglandins causing beneficial effects on hemodynamics which are independent of thromboxane inhibition. To more specially examine the role of TxA₂ in CyA toxicity, we investigated the effects of the thromboxane receptor antagonist GR32191 on renal hemodynamics in a rat model of CyA nephrotoxicity. In this model, administration of CyA resulted in a significant decrease in glomerular filtration rate (GFR) 2.85±0.26 [CyA] vs 6.82±0.96 ml/min/kg [vehicle]; p<0.0005) and renal blood flow (RBF) (21.6±2.31 [CyA] vs 31.8±3.60 ml/min/kg [vehicle]; p<0.025). Renal vascular resistance (RVR) was significantly higher in rats given CyA compared to animals treated with CyA vehicle (5.32±0.55 [cyCyA] vs 3.54±0.24 mm Hg/min/ml/kg [vehicle]; p<0.05). These hemodynamic alterations were associated with a significant increase in urinary excretion of unmetabolized, “native” thromboxane B₂ (TxB₂ (103±18 [CyA] vs 60±16 pg/hour [vehicle]; p<0.05). Only minimal histomorphologic changes were apparent by light microscopic examination of kidneys from both CyA and vehicle treated animals. However, with immunoperoxidase staining, a significantly greater number of cells experssing the rat common leukocyte antigen was found in the renal interstitium of rats given CyA^*. There was no detectable increase in monocytes/macrophages in the kidneys of CyA toxic animals. In rats treated with CyA, intraarterial infusion of GR32191 at maximally tolerated doses significanlty increased GFR and RBD, and decreased RVR. Although both RBF and RVR were restored to levels not different from controls, GFR remained significantly reduced following administration of GR32191. These data suggest that the potent vasoconstrictor TxA₂ plays an important role in mediating renal dysfunction in CyA nephrotoxicity. However, other factors may be important in producing nephrotoxicity associated with CyA.     

Inhibition of ovulation in the gonadotropin-primed immature rat by exogenous prostaglandin E2

In the past two decades there have been innumerable reports that prostaglandins (PGs) are essential for mammalian ovulation. However, we have recently found that a relatively low dose of 0.03 mg indomethacin (INDO) sc to PMSG/hCG-primed immature Wistar rats can significantly reduce ovarian PG levels without inhibiting the control ovulation rate of 60+ ova/rat (1–3). In view of this information, the present study was an effort to duplicate the earlier reports that PGs can reverse the “inhibitory” effect of INDO on ovulation. In control animals, which received PMSG and hCG only, the ovulation rate was 63.8 ± 4.5 ova/rat. This rate was reduced to 4.1 ± 1.1 ova/rat when the animals were injected with 1.0 mg INDO at 3 h after hCG. In no instance was this inhibition reversed when the animals were treated with 1.0 mg of PGE₂ or PGF_2α, or a combination of both prostanoids in either a single dose at 3 h after hCG, or in 4× doses at 2-h intervals beginning at 3 h after hCG. Furthermore, in animals that did not receive INDO, the ovulation rate in PGE₂-treated animals was reduced to 20.0 ± 6.7 ova/rat, and in animals treated with PGE₂ and PGF_2α (combined) it was reduced to 19.4 ± 6.5 ova/rat. In summary, not only did the PGs fail to reverse the anti-ovulatory effect of INDO, PGE₂ actually suppressed the ovulation rate.     

The effects of a diet rich in fish oil on human neutrophils: Identification of leukotriene B5 as a metabolite

Diets that are enriched with fish oil have been shown to alter arachidonic acid metabolism via the cyclooxygenase pathway. Recently it has been shown that one of the major component fatty acids of fish oil, eicosapentaenoate (EPA), is a substrate for the leukotriene B (LTB) pathway when added exogenously to human neutrophils . We fed a diet that contained 8–10 gm/day of EPA to four human subjects for three weeks and compared the arachidonate metabolism of their neutrophils to the same functions while the subjects were on their usual diet. The fish oil-supplementation increased neutrophil EPA content from undetectable levels to 7.4 ± 2.4% (p<0.01, expressed as % of total fatty acid), and decreased arachidonate from 15.4 ± 2.3% to 12.8 ± 2.3% (p<0.05). Leukotriene B₅ was identified as a metabolite during the fish oil-diet by its chromatographic profile and mass spectrum. During the experimental diet LTB₄ decreased from 160 ± 37 ng/10⁷ neutrophils to 120 ± 12 (p<0.05), and LTB₅ increased from 0 to 39 ± 9 ng/10⁷ neutrophils (p<0.005). The diet had no effect on neutrophil aggregation or adherence to nylon fibers.     

The in vitro effect of indomethacin on basal bone resorption, on prostaglandin production and on the response to added prostaglandins

Mouse calvaria were maintained in organ culture without serum additives. Basal active resorption, as measured by ⁴⁵Ca and hydroxyproline release, was significantly inhibited to 74% control levels by indomethacin (1.4 × 10⁻⁷ M). Prostaglandin F and prostaglandin E₂ production, determined by radioimmunoassay, were both significantly lowered by this concentration of indomethacin. DNA, protein and hydroxyproline synthesis, as indices of cell toxicity, were unaffected by low concentrations of indomethacin, while concentrations of 1.4 × 10⁻⁶M inhibited protein synthesis (p<0.005). In the presence of indomethacin (1.4 × 10⁻⁷M) both PGE₂ and PGF_2α stimulated resorption in a dose-dependent manner, with PGE₂ being the more potent. Neither prostaglandin affected hydroxyproline synthesis at low concentrations, but PGE₂ had a marked inhibitory action at a higher concentration (10⁻⁶M). In combination, the effects of PGE₂ and PGF_2α showed no evidence of synergism or any antagonistic action. The study shows that in vitro calcium and hydroxyproline resorption in the unstimulated mouse calvaria are inhibited by indomethacin at concentrations measured in serum during human therapy. The decreased PGF and PGE₂ production associated with this decreased bone resorption in the presence of non-toxic concentrations of indomethacin would suggest a role for these prostaglandins in maintaining the basal resorption of cultured bone.