Geographic distribution and regional impacts of Oxyops vitiosa (Coleoptera: Curculionidae) and Boreioglycaspis melaleucae (Hemiptera: Psyllidae), biological control agents of the invasive tree Melaleuca quinquenervia.

The invasive tree Melaleuca quinquenervia (Cav.) Blake is widely distributed throughout peninsular Florida and poses a significant threat to species diversity in the wetland systems of the Everglades. Mitigation of this threat includes the areawide release campaign of the biological control agents Oxyops vitiosa Pascoe and Boreioglycaspis melaleucae Moore. We summarize the results of this release effort and quantify the resulting geographic distribution of the herbivores as well as their regional impact on the target weed. A combined total of 3.3 million individual Melaleuca biological control agents have been redistributed to 407 locations and among 15 Florida counties. Surveys of the invaded area indicate that the geographic distribution of O. vitiosa encompasses 71% of the Melaleuca infestation. Although released 5 yr later, the distribution of B. melaleuca is slightly greater than its predecessor, with a range including 78% of the sampled Melaleuca stands. Melaleuca stands outside both biological control agents' distributions occurred primarily in the northern extremes of the tree's range. Strong positive association between herbivore species was observed, with the same density of both species occurring in 162 stands and no evidence of interspecific competition. Soil type also influenced the incidence of biological control agents and the distribution of their impacts. The odds of encountering O. vitiosa or B. melaleucae in cells dominated by sandy soils were 2.2 and 2.9 times more likely than those predominated by organically rich soils. As a result, a greater level of damage from both herbivores was observed for stands growing on sandy versus organic-rich soils.

Selection on herbivory resistance and growth rate in an invasive plant.

The evolution of increased competitive ability (EICA) hypothesis proposes that invasive species evolve decreased defense and increased competitive ability following natural enemy release. Previous tests of EICA examined the result of evolution by comparing individuals from home and introduced ranges, but no previous study of this hypothesis has examined the process of evolution by analyzing patterns of selection. On the basis of EICA, there should be selection for competitive ability without herbivores and selection for defense with herbivores. Selection on competitive ability should be stronger for genotypes accustomed to herbivores (home range genotypes), and selection on defense should be stronger for genotypes unaccustomed to herbivores (introduced range genotypes). Using a field experiment, we tested these hypotheses for the invasive plant Melaleuca quinquenervia. There was a negative genetic correlation between resistance and growth, indicating a trade-off. However, selection for stem elongation (an indicator of competitive ability) was always positive, and selection on resistance was always negative and did not depend on genotype source or the presence of herbivores. The patterns of selection found in this study contrast with predictions from EICA and accurately predict the lack of evolutionary change in growth and resistance following the introduction of this species from Australia to Florida.

Initial impacts and field validation of host range for Boreioglycaspis melaleucae Moore (Hemiptera: Psyllidae), a biological control agent of the invasive tree Melaleuca quinquenervia (Cav.) Blake (Myrtales: Myrtaceae: Leptospermoideae).

Invasion of south Florida wetlands by the Australian paperbark tree, Melaleuca quinquenervia (Cav.) S.T. Blake (melaleuca), has caused adverse economic and environmental impacts. The tree's biological attributes and favorable ambient biophysical conditions combine to complicate efforts to restore and maintain south Florida ecosystems. Management requires an integrated strategy that deploys multiple biological control agents to forestall reinvasion and to supplement other control methods, thereby lessening recruitment and regeneration after removal of existing trees. This biological control program began during 1997 when an Australian weevil, Oxyops vitiosa (Pascoe), was released. A second Australian insect, the melaleuca psyllid (Boreioglycaspis melaleucae Moore), first introduced during 2002, has also widely established. After inoculation of the psyllid in a field study, only 40% of seedlings survived herbivory treatments compared with 95% survival in controls. The resultant defoliation also reduced growth of the surviving seedlings. A weevil-induced decline at a site comprised mainly of coppicing stumps had slowed after a 70% reduction. Psyllids colonized the site, and 37% of the remaining coppices succumbed within 10 mo. The realized ecological host range of B. melaleucae was restricted to M. quinquenervia; 18 other nontarget plant species predicted to be suboptimal or nonhosts during laboratory host range testing were unaffected when interspersed with psyllid-infested melaleuca trees in a common garden study. Evaluations are ongoing, but B. melaleucae is clearly reducing seedling recruitment and stump regrowth without adversely impacting other plant species. Manifestation of impacts on mature trees will require more time, but initial indications suggest that the psyllid will be an effective supplement to the weevil.

Relevance of 10-min delayed recall in dementia screening.

Within the context of early diagnosis of Alzheimer's disease (AD), there is a growing interest in neuropsychological screening tests. Amongst these tests, we focused on the largely used Memory Impairment Screen (MIS). The objective of the present work was to show that adding a 10-min delayed recall to the MIS, improves the test psychometric characteristics in order to detect dementia in the earliest stages. A prospective study was carried out on a cohort of 270 consecutive elderly ambulatory subjects attending the Broca Hospital Memory Clinic: normal controls (n = 67), mild cognitive impairment subjects (n = 98) and mildly demented patients [n = 105, Mini Mental State Examination (MMSE) = 23 +/- 4]. This study consisted in testing the advantage of the 10-min delayed recall entitled MIS-D compared with the MIS. At a cut-off score of 6, the MIS-D revealed satisfying psychometric characteristics with a sensitivity of 81% and a specificity of 91%, whilst the MIS alone indicated a sensitivity of 60% and a specificity of 88% in detecting dementia. In demented patients with MMSE score > or =26, MIS-D properties still remained satisfying (sensitivity: 75%, specificity: 92%). MIS-D is a more relevant screening test than MIS alone at very early stages of dementia.

Accidental MCI in healthy subjects: a prospective longitudinal study.

A study was realized on 130 healthy and autonomous volunteers (60-80 years old) who met specific medical and functional inclusion criteria. A comprehensive battery of neuropsychological tests was performed at baseline (M0), 6 and 12 months (M6, M12). At M0 the results indicated that 65% were cognitively normal on each of all the neuropsychological tests, whereas 35% presented a cognitive deficit on one or more tests. At M12, 52% of the subjects who had a cognitive deficit at M0 remained impaired, whereas 48% normalized their scores: they performed as well as the subjects classified normal at M0. The results also indicated that the subjects who remained impaired at M12, had at M0 low scores on three tests or more, whereas the ones who normalized their scores had one or two failed tests. This study focuses on the risk of false positive cases and shows that low scores can be accidental. The authors propose decision rules allowing to reduce the risk of false positive cases. The observation of accidental impairment invites to be cautious and makes this 1-year follow-up study particularly relevant, since a 1-year follow-up is generally needed to diagnose very mild dementia.

Anti-inflammatory mechanism of alminoprofen: action on the phospholipid metabolism pathway.

Alminoprofen is a nonsteroidal anti-inflammatory drug (NSAID) of the phenylpropionic acid class. It has anti-inflammatory properties different from the classical NSAID. Using both in vitro systems of cells in culture and in vivo models of inflammation, we report here that alminoprofen possesses both antiphospholipase A2 (PLA2) activity and anti-cycloxygenase (COX) activity. The PLA2 targeted by alminoprofen is likely the secretory phospholipase A2 (sPLA2) while the COX targeted is the COX-2.

Modulation of human myometrial PGE2 receptor by GTP characterization of receptor subtype.

We studied PGE2 specific binding sites in human myometrial microsomes prepared from uterine specimens obtained by hysterectomy (women between 38 and 55 years of age). Competition experiments showed that the potency order for various prostaglandins (PGs) was: PGE2 > or = PGE1 >> PGF2 alpha > Iloprost > or = Carbacyclin >> ZK 110841 (PDG2 analogue). These relative affinities indicated that the receptor was of the EP type. In kinetic experiments GTP, GppNHp and GTP gamma S increased the rate of PGE2 binding (steady state was reached more rapidly in the presence of nucleotides) but maximal specific binding was not significantly different. Complete dissociation could not be obtained, even in the presence of GTP. Only 50% of maximal binding was readily dissociable. The dissociation rate was 4.56.10(-4) sec-1 (half time of about 660 sec) and in the presence of GTP analogues it was slightly increased (k-1 = 7.16 10(-4) sec-1, half time 420 sec.). Scatchard analysis of saturation curves showed an increase in ligand receptor affinity in the presence of GTP or nucleotide analogues: the Kd shifted from 9.66 +/- 2.8.10(-9) M to 4.96 +/- 1.25.10(-9) M, but the number of binding sites did not change significantly (310 +/- 37 to 350 +/- 17 fmol/mgP). The effect of GTP was observed at a concentration of 5.10(-4)M. GppNHp and GTP gamma S were effective at 1.10(-5) M. Pretreatment of myometrial membranes with pertussis or cholera toxins had no effect on PGE2 binding to membrane sites. Our conclusion is that GTP induced conversion of a population of low affinity sites into a population of higher affinity sites. This effect of guanine nucleotides was described in adipocytes and kidney medulla. Competition studies with PGE2 analogues (sulprostone, 17-phenyl-omega-trinor PGE2, M&B 28,767, misoprostol, butaprost) showed that this receptor mediates a contractile response and is probably an EP3 subtype.

A rapid, simple method for calculating equilibrium constants and antibody site concentrations from dilution curves alone.

A rapid new practical method for calculating both the antibody-antigen equilibrium constant and the antibody concentration from antibody dilution curve data alone is described. This method is faster than the inhibition curve method for evaluating a humoral immune response. It is particularly suitable for monitoring the immune response of an immunization program. The response is assessed as an immunization index, Abi*Ka. This index is more exact than the antibody titer obtained from dilution curves and independent of the specific activity of the labelled molecule and total activity used in the assay. The method was used to monitor the production of a monoclonal antibody to the sulphide peptide leucotriene including immunization, cloning and purification.

Active metabolism of arachidonic acid by Kaposi sarcoma cells cultured from lung biopsies (KS-3); identification by HPLC and MS/MS of the predominant metabolite secreted as the 11,12-epoxy-eicosatrienoic acid.

The development of long-term culture of AIDS-KS cells has allowed us to investigate further a possible vascular origin of Kaposi sarcoma. Taking into account the relative specificity of arachidonic acid (AA) metabolism according to cell type, the AA 'cascade' was analyzed in cultured KS-3 cells established from lung biopsies and compared to human umbilical venous endothelial (H-UVE) cells and human myometrial smooth muscle (H-MSM) cells, under basal conditions and after stimulation with vasoactive agents such as histamine or thrombin. Considering strictly the 'prostaglandin' profile given by RIAs, the metabolism of AA was closer, whilst not identical, to H-UVE than to H-MSM cells. However, evaluation of all the eicosanoids released from [3H]AA labeled KS-3 cells revealed that the predominant metabolite was not prostacyclin (PGI2), as suggested from PG RIAs, but an epoxy-eicosatrienoic acid (EET), identified as the 11, 12 isomer by HPLC and MS/MS. The synthesis of this EET is probably cytochrome P-450 monooxygenase dependent. Its potential role in the development of the KS tumor cells is under investigation.

Prostaglandins of the E Series Stimulate Cyclic AMP Accumulation and N-Acetylation of Serotonin in Chick Pineal Cells.

We investigated the effects of prostaglandins on cyclic AMP (cAMP) levels and on the activity of the rate-limiting enzyme of melatonin biosynthesis, arylalkylamine-N-acetyltransferase (NAT). The study was performed on primary cultures of dispersed chick pineal cells. Prostaglandin E, (PGE,) increased cAMP levels 2-fold and this stimulation went up to 4-fold in the presence of a phosphodiesterase inhibitor. The PGE,- evoked increase in cAMP levels did not desensitize over 6 h. The potency order of a series of prostaglandins to increase cAMP levels (PGE(1) PGE(1) >PGA2>PGD(2) ≃PGF(2) α) agreed with the pharmacological profile of the adenylate cyclase-coupled prostaglandin receptor. Inhibition of endogenous prostaglandin synthesis by two cyclooxygenase inhibitors (indomethacin and aspirin) caused a 30% decrease in cAMP levels. This effect was completely reversed by the addition of exogenous PGE(1) or PGE(2) . Indomethacin and aspirin also caused a 50% decrease in NAT activity. Prostaglandins of the E series increased NAT activity up to 2-fold above basal level and restored NAT activity after inhibition by indomethacin or aspirin. These results are the first illustration of a role for prostaglandins in chick pineal cells. The correlations observed between cAMP levels and NAT activity suggest that the regulation of NAT activity by prostaglandins of the E series might be mediated by changes in cAMP concentration.

Basal and PAF-, interleukin 1-, ether stress-induced hypothalamic pituitary adrenal secretion of conscious rat: modulation by PAF antagonists.

We have previously shown that exogenous (1 to 5 nmol i.c.v.) PAF induces a rapid increase in plasma ACTH and beta endorphin followed by an increase in plasma corticosterone in conscious rats. The stimulatory action of PAF on the secretion of hypothalamic-pituitary-adrenal (HPA) axis products is mediated at least partly by stimulating hypothalamic CRF release. In addition rat hypothalamic membranes have two populations of specific PAF binding sites. In order to clarify the mode of PAF action on the stress-related hormones, we have now investigated the effect of two PAF antagonists, BN 50739 and RP 52770, on basal and PAF-induced ACTH and corticosterone secretion by conscious rats and on PAF specific binding to rat hypothalamic membranes. The role of PAF as a mediator of neuroendocrine secretion in response to acute stress was examined by determining the effect of PAF antagonists on ether-stress inducing HPA activity. We have also investigated their effect on IL 1-induced HPA activity. The ability of BN 50739 and RP 52770 to displace 3H PAF from its hypothalamic binding sites was correlated with their ability to alter basal hormone secretion and to counteract the PAF-stimulated secretion of HPA axis hormones in vivo (P less than 0.05 by ANOVA). Pretreatment with BN 50739. (50 nmol i.c.v.) did not alter ACTH response to a 1 min ether exposure or to IL1 beta injection (2 nmol i.c.v.). In contrast, RP 52770 (55 nmol i.c.v.) significantly inhibited the ether stress-induced ACTH and corticosterone production by 50% (P less than 0.05). In parallel, pretreatment with RP 52770 (55 nmol i.c.v.) caused a significant inhibition of IL1 beta-induced ACTH secretion. These results suggest that PAF acts, in vivo, on ACTH and corticosterone secretion, through a centrally mediated CRF dependent mechanism involving PAF receptor sites. Additionally, the data also indicate that PAF could have a central role in mediating basal and stress-induced ACTH secretion and that IL 1-induced HPA secretion may be mediated at least in part through the production of PAF.

A competitive receptor binding assay for platelet-activating factor (PAF): quantification of PAF in rat brain.

A radioreceptor assay (RRA) was developed using rabbit platelet membrane preparations to quantify platelet-activating factor (PAF) and lyso-PAF, the deacylated derivative of PAF, in a variety of tissues and biological fluids. We examined PAF and lyso-PAF levels in different rat brain areas with regard to the many proven and postulated actions of PAF in brain functions. Human saliva was selected to check the validity of this RRA. The samples were extracted with methanol/chloroform/water and purified by high-performance liquid chromatography on a 5-microns Nucleosil Si column (overall recovery: 78%). Sample extracts were acetylated before chromatography to assay lyso-PAF. PAF itself was assayed in non-aceylated samples. A competitive binding assay was performed using aliquots of platelet membrane preparation and tritiated PAF. The minimum detectable amount of PAF was 144 pg per tube and the receptor was highly specific for PAF. In human saliva, we confirm the presence of PAF and lyso-PAF within the range expected. Moreover there was a good correlation between the RRA and the aggregation assay (r = 0.976). A defined cocktail of protease inhibitors allowed storage of platelet membrane preparations for at least 3 months at -20 degrees C with no change in binding properties. In the brain we observed the prevalent presence of lyso-PAF and large variations in PAF and lyso-PAF concentrations between the different brain areas analyzed. PAF was undetectable in the hypothalamus but the lyso-PAF concentration was 2.5 micrograms/g wet tissue. The PAF concentration in the cortex varied from 0 to 16 ng/g wet tissue while that of lyso-PAF was 0.7 micrograms/g wet tissue. Moreover the amount of lyso-PAF varied between the different brain areas analyzed. The hippocampus contained the highest amount (7 micrograms/g wet tissue), and relatively high levels were found in the hypothalamus, medulla oblongata and corpus striatum. The cerebellum and cortex contained the lowest levels of lyso-PAF. These findings show that PAF is present in the central nervous system mainly in its inactive form, lyso-PAF, and suggest that its effects as a modulator of brain function may be dependent on deacetylation, rather than synthesis.

Prostaglandin E2 and leukotriene C4-induced luteinizing hormone-releasing hormone release from immature and adult male rat median eminences in vitro: eicosanoid formation and binding parameters.

The amounts of prostaglandin E2 formed in vitro by the median eminences of adult male rats were greater than those produced by the median eminences of immature, 22 day-old rats. However, the amount of leukotriene C4 produced by the adult rat median eminences was lower than that produced by the immature rat median eminences. Analysis of the prostaglandin E2 binding parameters of hypothalamic P2 membrane fractions indicates that there are two binding components, one high affinity (RH) and one low affinity (RL) in both adult and immature rats. The maximal binding capacity of RH from adult rat membranes was significantly lower than that of immature rat membranes, correlating with greater prostaglandin E2 production by the adult rat median eminence. Only one leukotriene C4 binding site was detected in both adult and immature rat membranes. Exogenous prostaglandin E2 and leukotriene C4 both stimulated, the release of luteinizing hormone-releasing hormone to the same extent from both the adult and immature median eminences.

Elimination of undesired cross-reactants by using mixtures of antibodies: experimental and theoretical evaluations of hapten radioimmunoassays.

This study shows that the specificity of radioimmunoassays can be improved by including a second antibody raised against an undesired cross-reactant. In a radioimmunoassay of prostaglandin E2 (PGE2) involving a monoclonal antibody, the cross-reactivity with 6-keto-prostaglandin E1 (6kPGE1) was decreased from 20% to 2% by including a high concentration of a polyclonal anti-6kPGE1. A similar increase in specificity was obtained in the assay of a larger hapten, luliberin (luteinizing hormone releasing hormone); the cross-reactivity of a luliberin analog was decreased 20-fold. Equations derived from the Law of Mass Action were used for the mathematical analysis and for the computer simulation of changes in assay affinity and specificity according to the quantity and quality of the mixed antibodies. The model gave values that agreed well with experimental data; it promises to be quite useful in designing specific radioimmunoassays.

Practical method for optimizing radioimmunoassay detection and precision limits.

A model of the competitive radioimmunoassay standard curve, based on the Law of Mass Action, has been developed and used in conjunction with experimental and counting errors to predict the assay detection limit and precision profiles. We verified the model with hapten radioimmunoassays performed in our laboratory. The resulting computer program can be used to determine the optimum antiserum concentration--depending on its affinity--and labeled-antigen concentration--according to its specific activity and nonspecific binding. The graphical representation of this model provides radioimmunologists with a practical tool for assay optimization.

Comparative study of biosynthetic human growth hormone immunogenicity in growth hormone deficient children.

The immunogenicities of six recombinant human growth hormone (rhGH) preparations, from KABI (A rhGH191 and B rhGH192), Eli Lilly (C), Nordisk (D), Sanofi (E) and Serono (F), used to treat 260 GH-deficient children, have been compared using a common specific and sensitive procedure for antibody determination. For this purpose we developed two immunoassays: a competitive liquid radioimmunoassay using 125I-rhGH, and an immunometric solid enzymoimmunoassay in which the rhGHs were immobilized. Blood samples were collected from the GH-deficient children before treatment and after 3, 6, 9, 12, 18 and 24 months of therapy. Human GH antibodies were detected in children treated with 3 of the 6 rhGH preparations. Seven percent of the patients treated with hormone A, 14% with hormone B and 22% with hormone C formed antibodies against the respective rhGH. Differences in capacity and affinity of the hGH antibodies were observed between these anti-GH-positive groups. They could be divided into 2 groups according to their immunopotency. One group (7, 14 and 6% of the patients treated with hormones A, B and C, respectively) developed anti-hGH antibodies with very low binding capacities (30-100 fmol/ml). The other group (16% of the patients treated with hormone C) developed IgG-type antibodies to hGH with higher binding capacities (200-1,200 fmol/ml) and a measurable binding affinity (Ka = 10(8) M-1). These hGH antibodies partially inhibited the binding of labeled GH to its specific liver membrane receptor. However, because of their low titer, they did not inhibit growth in the treated children.(ABSTRACT TRUNCATED AT 250 WORDS)

[Prostaglandins and reproduction. I. Physiological aspects]. / Prostaglandines et reproduction. I. Aspects physiologiques.

1) Eicosanoids are a family of polyunsaturated 20-carbon fatty acids and their metabolites. The metabolites are produced by three enzymatic pathways: the cyclooxygenase pathway, giving prostaglandins (PGs), the lipoxygenases and the epoxygenases pathways. Arachidonic acid (C20:4) is the most common fatty acid precursor in mammalian cells, where it is incorporated, as an ester, into the membrane lipid complex. 2) The eicosanoids have a variety of effects on several cell activities, including secretion, muscle contraction, cell growth and differentiation. The type of effect--stimulation or inhibition--depends on the metabolite, its concentration, the metabolic activity of the cell and the involvement of other humoral factors. 3) The message may be transmitted via a specific membrane receptor to a specific transduction system: the adenyl or guanyl cyclase system and mobilization of free cytosolic Ca2+, or via the participation of membrane ion channels. Depending on which is involved, the eicosanoid message applies to the cell in which it was synthesized or to neighboring cells (autocrine or paracrine action). 4) The eicosanoids, especially the PGs, take part in many reproductive processes; in the hypothalamic-pituitary axis, particularly through the synaptic modulation by PGE2 (stimulation of LHRH secretion and inhibition of noradrenaline secretion); in the ovary: follicle maturation and luteolysis; in the oviducts: gamete migration; in the uterus: ovum implantation and parturition. 5) PGs seem to have a variety of species-dependent effects on the normal onset of labor. In sheep there is an increase in fetal cortisol, a drop in the progesterone/estradiol ratio and increased PG synthesis. In women, there is an increase of phospholipase A2 activity in amnios and uterus with an increase of PGE2 in the first tissue and of PGF2 alpha in the second one. 6) The PGs from the seminal fluid have several actions. They effect fertility by acting on the female genital tract or on the spermatozoa. PGE1 and PGE2 influence the fertilization capacity. PGs also effect the process of ejaculation (inhibition of the stimulatory effect of noradrenaline). Finally, they effect the immune responses: PGEs and 19 hydroxy PGEs immuno-suppressive characteristics.

Effects of N-acetyl-aspartyl glutamic acid and sodium cromoglycate on leukotriene B4 secretion by human leukocytes.

Peripheral leukocytes from allergic subjects were treated for 30 min with sodium cromoglycate (SCG) or with N-acetyl-aspartyl glutamic acid (NAAGA) and challenged for leukotriene B4 (LTB4) production with calcium ionophore A 23187. NAAGA significantly inhibits LTB4 release at concentrations of 10(-2) M (-86%), 5 x 10(-3) M (-49%) and 10(-3) M (-34%), while SCG was not able to block LTB4 production within the range of 10(-2)-10(-4) M. In spite of the fact that SCG and NAAGA are chemically unrelated and that both show antiallergic properties, only NAAGA is able in this model to block production of LTB4, a chemical mediator strongly involved in inflammatory and hypersensitivity reactions.

Intracerebroventricular injection of platelet-activating factor induces secretion of adrenocorticotropin, beta-endorphin and corticosterone in conscious rats: a possible link between the immune and nervous systems.

To investigate whether platelet-activating factor (PAF) exerts an indirect action on immune cells by altering the secretion of hypothalamic-pituitary-adrenal (HPA) axis products, the effects of intracerebroventricular (i.c.v.) PAF on adrenocorticotropic hormone (ACTH), beta-endorphin and corticosterone blood levels were examined in adult male rats. Hormones were radioimmunoassayed on blood samples from conscious or ether-anesthetized rats after i.c.v. injection of PAF or vehicle into the left lateral ventricle. PAF induced significant increases in these stress-related hormones under both, basal and ether-induced stress conditions. The analysis of the time course response to PAF of hormone release into the blood of unrestrained rats revealed that: i.c.v. injection of 5.4 nmol PAF resulted in rapid increases in ACTH and beta-endorphin, at the latest within 15 min after the onset of injection. The maximal response of both hormones was reached within 45 min after the onset of injection and was followed by an elevation of plasma corticosterone. Hormone release is related to the PAF dose infused, the lowest effective PAF concentration was 1 nmol. The stimulatory effect of PAF on ACTH and beta-endorphin secretion was strongly decreased in rats previously treated with purified anti-rat corticotropin-releasing factor (CRF) antibody. These results, associated with the in vitro demonstration that PAF increases CRF release from incubated rat median eminence, strongly support the hypothesis that the stimulatory action of PAF on the secretion of HPA axis products is mediated at least partly, by stimulating hypothalamic CRF release.(ABSTRACT TRUNCATED AT 250 WORDS)

Epoxygenase products of arachidonic acid are endogenous constituents of the hypothalamus involved in D2 receptor-mediated, dopamine-induced release of somatostatin.

The epoxyeicosatrienoic acids (EETs) were discovered as products of a cyclooxygenase/lipoxygenase-independent, cytochrome P-450 catalyzed metabolism of arachidonic acid (AA) termed the "epoxygenase" pathway. The rat hypothalamus is able to synthesize EETs from exogenous AA, and 5,6-EET has been found to release the neuropeptide somatostatin (SRIF) from hypothalamic nerve terminals of the median eminence (ME). In the present study, hypothalami from male rats were examined for the presence of endogenous EETs, using chemical, chromatographic, and mass spectral analysis procedures. The samples were initially separated in a C18 Sepralyte column, fractionated on TLC plates, and purified by reverse phase HPLC. Thereafter, they were esterified (pentafluorobenzyl esters) and subjected to negative ion chemical ionization/gas chromatography (GC)/mass spectral (MS) analysis. The GC retention time and the MS fragmentation patterns revealed the presence of a mixture of 8,9-, 11,12- and 14,15-EETs; instability of 5,6-EET during the isolation protocol precluded its identification. Total hypothalamic EET concentration was estimated to be 120 ng/g wet tissue. The 8,9-regiosomer released SRIF from ME nerve terminals with an ED50 of 5 x 10(-12) M; Dopamine (DA) and the D2 receptor agonist PPHT, but not the D1 receptor agonist SKF-38393, induced SRIF release from the ME. This effect was blocked by clotrimazole and ketoconazole, two inhibitors of microsomal cytochrome P-450 function and AA epoxygenase in particular. In contrast, the inhibitors failed to affect the increase in SRIF release induced by 8,9-EET. These results indicate that: 1) in addition to cyclooxygenase and lipoxygenase products, epoxygenase metabolites of AA are endogenous compounds of the hypothalamus, and 2) EETs may mediate the increase in SRIF release from hypothalamic neurons induced by the interaction of DA with D2 receptors.