Potentiation of sympathetic neurotransmission in bovine isolated irides by isoprostanes.

Isoprostanes (IsoP) are formed by free radical catalyzed peroxidation of arachidonic acid independent of the cyclooxygenase enzyme. In the present study, we examined the effect of IsoP on norepinephrine (NE) release from the bovine isolated iris. Furthermore, we studied the role of IsoP's in hydrogen peroxide (H2O2)-induced enhancement of NE release from this tissue. Isolated bovine irides were prepared for studies of [3H]NE release using the superfusion method. Release of [3H]NE was induced via electrical field stimulation. Both 8-iso-prostaglandin E2 (E2-IsoP) and 8-iso-prostaglandin F2 alpha (F2-IsoP) produced a concentration-related enhancement of field-stimulated [3H]NE release from isolated bovine irides, an effect that was mimicked by the thromboxane (Tx) receptor agonist, U46619 and by H2O2. The Tx-receptor antagonist, SQ 29548 inhibited responses to E2-IsoP (10 microM) with an IC50 of 370 +/- 50 nM. SQ 29548 (10 microM) also blocked the enhancement of electrically-evoked [3H]NE release induced by U46619 (10 microM) but not that caused by H2O2 (300 microM). The Tx synthetase inhibitor, carboxyheptylimidazole (10 microM) prevented the stimulatory effect of E2-IsoP on evoked [3H]NE release without affecting responses induced by H2O2. We conclude that IsoP's can enhance sympathetic neurotransmission in the bovine isolated iris, an effect that can be blocked by a Tx-receptor antagonist. Furthermore, endogenously produced Tx's mediate the stimulatory effect of IsoP's on NE release. However, endogenously generated IsoP's or Tx's are not involved in H2O2-induced potentiation of sympathetic neurotransmission.     

Effect of Isoprostanes on Sympathetic Neurotransmission in the Human Isolated Iris-Ciliary Body

Isoprostanes (IsoP's) are prostaglandin-like compounds that are derived from free-radical catalyzed peroxidation of arachidonic acid independent of the cyclcooxygenase enzyme. In the present study, we investigated the effect of IsoP's on norepinephrine (NE) release from human isolated iris-ciliary bodies. Isolated human iris-ciliary bodies were prepared for studies of [³H]NE release using the superfusion method. Both 8-iso-prostaglandin F₂ (F₂-IsoP) and the thromboxane (Tx) receptor agonist, U46619 enhanced field-stimulated [³H]NE release from isolated, superfused human iris-ciliary bodies without affecting basal tritium efflux. On the other hand, an equimolar concentration (10 M) of 8-iso-prostaglandin E₂ (E₂-IsoP) inhibited evoked [³H]NE overflow. The Tx-receptor antagonist, SQ 29548 blocked the enhancements of electrically-evoked [³H]NE release induced by F₂-IsoP and U46619. However, the inhibitory responses elicited by E₂-IsoP was not antagonized by SQ 29548. We conclude that IsoP's can produce both excitatory and inhibitory effects on sympathetic neurotransmission in human isolated iris-ciliary bodies. The stimulatory effects of IsoP' on NE release may be mediated by Tx-receptors.     

Prejunctional α2-Adrenoceptors and Peroxide-Induced Potentiation of Norepinephrine Release from the Bovine Iris

Peroxides can enhance field-stimulated [³H]norepinephrine ([³H]NE) release in isolated irides from several mammalian species. In the present study, we investigated the role of prejunctional ₂-adrenoceptors in peroxide-induced potentiation of sympathetic neurotransmission in bovine isolated irides. Isolated hemi-irides were incubated in a Krebs buffered-solution containing [³H]NE and prepared for studies of neurotransmitter release using the superfusion method. ₂-Adrenoceptor agonists, oxymetazoline, UK-14304 and clonidine inhibited field-stimulated [³H]NE overflow without affecting basal tritium efflux. Pretreatment of tissues with H₂O₂ (300 M) had no effect on inhibition of evoked [³H]NE release caused by the ₂-adrenergic agonists. However, H₂O₂ (300 M) caused significant (P < 0.01) leftward shifts of excitatory concentration-response curves to yohimbine (10 nM–1 M). In contrast, yohimbine (1 M) did not prevent the enhancement of evoked [³H]NE overflow induced by H₂O₂ (300 M). In conclusion, excitatory effects of peroxides on sympathetic neurotransmission in bovine irides are not mediated by prejunctional ₂-adrenoceptors.     

Effect of Inhibition of Cyclooxygenase on Pre- and Postjunctional Actions of Peroxides in the Iris-Ciliary Body

In the present study, we investigated the effect of inhibition of cyclooxygenase (COX) with flurbiprofen (FBF) on peroxide-induced enhancement of field-stimulated [³H]norepinephrine ([³H]NE) release from bovine isolated irides. Furthermore, the effect of FBF was examined on peroxide-induced attenuation of contractions evoked by carbachol on this tissue. Irides were prepared for studies of neurotransmitter release and for measurement of contractile tension in vitro. Pretreatment of tissues with FBF (10 M) caused significant (P < 0.001) rightward shifts of concentration-response curves to H₂O₂ and also decreased cumene hydroperoxide (cuOOH)-induced enhancement of evoked [³H]NE release. FBF (10 M) partially prevented the attenuation of carbachol-induced contractions induced by H₂O₂ (300 M) and cuOOH (300 M). We conclude that inhibition of the biosynthesis of prostanoids reduced both the prejunctional stimulatory effects of H₂O₂ and cuOOH on sympathetic neurotransmission and inhibitory effects of peroxides on carbachol-induced contractions the in the bovine isolated iris.     

Pharmacological Actions of Hydrogen Sulfide Donors on Sympathetic Neurotransmission in the Bovine Anterior Uvea,In Vitro

In the present study, we investigated the effect of three different sources of hydrogen sulfide (H₂S) on sympathetic neurotransmission from isolated superfused bovine iris-ciliary bodies. The three agents under consideration were: ACS67, a hybrid of latanoprost and a H₂S-donating moiety; l-cysteine, a substrate for endogenous production of H₂S and GYY 4137, a slow donor of H₂S. We also examined the contribution of prostaglandins to the pharmacological actions of the H₂S donors on release of [³H]-norepinephrine ([³H]NE) triggered by electrical field stimulation. ACS67, l-cysteine and GYY 4137 caused a concentration-dependent inhibition of electrically-evoked [³H]NE release from isolated bovine iris-ciliary bodies without affecting basal [³H]NE efflux. The cyclooxygenase inhibitor, flurbiprofen enhanced the inhibitory action of ACS67 and l-cysteine on stimulated [³H]NE release. Both aminooxyacetic acid, an inhibitor of cystathionine-β-synthase and glibenclamide, a K_ATP channel blocker reversed the inhibition of evoked NE release induced by the H₂S donors. We conclude that H₂S donors can inhibit sympathetic neurotransmission from isolated bovine iris-ciliary bodies, an effect partially dependent on the in situ production of H₂S and prostanoids, and is mediated by an action on K_ATP channels.     

Prejunctional Inhibitory Effects of Isoprostanes on Dopaminergic Neurotransmission in Bovine Retinae,In vitro

We investigated the effect of isoprostanes (IsoPs) on potassium (K⁺)-depolarization-evoked release of [³H]dopamine from isolated bovine retinae. Isolated retinae were preloaded with [³H]dopamine and then prepared for studies of [³H]dopamine release using the superfusion method. 8-iso(15R)PGF_2α, 8-isoPGE₂, 8-isoPGE₁ and 8-isoPGF_2α attenuated [³H]dopamine release from isolated bovine retinae. At a concentration of 1 μM, the rank order of activity displayed by IsoP agonists was: 8-iso(15R)PGF_2α > 8-isoPGE₂ > 8-isoPGE₁ > 8-isoPGF_2α. Inhibition of cyclooxygenase (COX) with flurbiprofen reversed the effects caused by 8-isoPGE₂ (10 nM and 10 μM), 8-iso(15R)PGF_2α (1 μM) and 8-isoPGE₁ (1 μM). Although the EP1/EP2 antagonist, AH 6809 (10 μM) had no significant effect on K⁺-induced [³H]dopamine release, it blocked the inhibitory effect of both 8-isoPGE₁ (10 μM) and 8-isoPGE₂ (10 μM). In conclusion, IsoPs attenuate K⁺-induced [³H]dopamine release in isolated bovine retinae, presumably via an indirect action on COX pathway leading to the production of prostanoids, which in turn, activates EP receptors.     

Regulation of [3H]D-Aspartate Release by the 5-F2t-Isoprostane and Its 5-Epimer in Isolated Bovine Retina

We have evidence that 15-F?-isoprostanes (15-F?-IsoPs) regulate excitatory neurotransmitter release in ocular tissues. Although 5-F?-IsoPs are abundantly produced in mammals, their pharmacological actions on neurotransmitter release remain unknown. In the present study, we compared the effect of the 5-F?-IsoP epimer pair, 5-F(2t)-IsoP (C5-OH in β-position) and 5-epi-5-F(2t)-IsoP (C5-OH in α-position), on K?-evoked [3H]D-aspartate release in isolated bovine retina. We further examined the role of prostanoid receptors on the inhibitory action of 5-epi-5-F(2t)-IsoP on [3H]D-aspartate overflow. Isolated bovine retina were prepared for studies of K?-evoked release of [3H]D-aspartate using the superfusion method. 5-epi-5-F(2t)-IsoP (0.01 nM to 1 μM), attenuated K?-evoked [3H]D-aspartate release in a concentration-dependent manner, with the inhibitory effect of 26.9% (P < 0.001; IC?? = 0.2 μM) being achieved at 1 μM concentration. Its 5-(S)-OH-epimer, 5-F(2t)-IsoP (0.1 nM-1 μM), exhibited an inhibitory biphasic action, yielding a maximal response of 35.7% (P < 0.001) at 10 nM concentration of the drug (IC?? value of 3 nM). Although the prostanoid-receptor antagonists, AH 6809 (10 μM; EP???/DP) and BAY-u3405 (10 μM; DP/Tx) exhibited no effect on 5-epi-5-F(2t)-IsoP (10 nM-1 μM)-mediated inhibition, SC-19220 (1 μM; EP?) completely reversed 5-epi-5-F(2t)-IsoP (0.1 μM and 1 μM)-induced attenuation of K?-evoked [3H]D-aspartate release. Similarly, both SC-51322 (10 μM; EP? and AH 23848 (1 μM; EP?) reversed the inhibitory action elicited by 5-epi-5-F(2t)-IsoP (0.1 μM) on the neurotransmitter release. We conclude that the 5-F?-IsoP epimer pair, 5-F(2t)-IsoP and 5-epi-5-F(2t)-IsoP, attenuate K?-induced [3H]D-aspartate release in isolated bovine retina presumably via prostanoid receptor dependent mechanisms. The trans-orientation of the allylic hydroxyl group at position C5 accounts for the apparent biphasic response exhibited by 5-F(2t)-IsoP on excitatory neurotransmitter release.     

Regulation of [<Superscript>3</Superscript>H] <Emphasis Type="SmallCaps">d</Emphasis>-Aspartate Release from Mammalian Isolated Retinae by Hydrogen Sulfide

Hydrogen sulfide (H₂S), can produce pharmacological effects on neural and non-neural tissues from several mammalian species. The present study investigates the pharmacological action of H₂S, (using sodium hydrosulfide, NaHS, and/or sodium sulfide, Na₂S as donors) on amino acid neurotransmission (using [³H] d-aspartate as a marker for glutamate) from isolated, superfused bovine and porcine retinae. Isolated neural retinae were incubated in Krebs solution containing [³H] d-aspartate at 37°C. Release of [³H] d-aspartate was elicited by high potassium (K⁺ 50 mM) pulse. Both NaHS and Na₂S donors caused an inhibition of K⁺-evoked [³H] d-aspartate release from isolated bovine retinae without affecting basal [³H] d-aspartate efflux yielding IC₅₀ values of 0.006 and 6 μm, respectively. Furthermore, NaHS inhibited depolarization-evoked release of [³H] d-aspartate from isolated porcine retinae with an IC₅₀ value of 8 μM. The inhibitory action of NaHS on [³H] d-aspartate release from porcine retinae was blocked by propargyglycine, a selective inhibitor of cystathionine γ-lyase (CSE). Our results indicate that H₂S donors can inhibit amino acid neurotransmission from both isolated bovine and porcine retinae, an effect that is dependent, at least in part, on intramural biosynthesis of H₂S.     

Regulation of Norepinephrine Release from Isolated Bovine Irides by Histamine

In the present study, we investigated the effect of histamine on sympathetic neurotransmission from isolated, superfused bovine irides. We also studied the pharmacology of prejunctional histamine receptors that regulate the release of norepinephrine (NE) from this tissue. The effect of exogenous histamine and various histamine receptor agonists was examined on the release of [³H]-norepinephrine ([³H]NE) triggered by electrical field stimulation using the Superfusion Method. Histamine receptor agonists caused a concentration-dependent inhibition of field-stimulated [³H]NE overflow with the following rank order of potency: imetit > histamine > R-α-methylhistamine. In all cases, the inhibitory action of histamine receptor agonists was attenuated at high concentrations of these compounds. The histamine receptor antagonists, clobenpropit (H₃-antagonist/H₄-agonist) and thioperamide (H₃-antagonist) blocked the inhibitory response elicited by R-α-methylhistamine and imetit, respectively. Inhibitory effects of R-α-methylhistamine and clonidine were not additive suggesting that prejunctional H₃- and α₂-adrenoceptors coexist at neurotransmitter release sites. We conclude that histamine produces an inhibitory action on sympathetic neurotransmission in the bovine iris, an effect mimicked by selective H₃-receptor agonists and blocked by H₃-antagonists.     

Modulation of GABA-augmented norepinephrine release in female rat brain slices by opioids and adenosine

GABA_A receptor activation augments electrically-stimulated release of norepinephrine (NE) from rat brain slices. Because this effect is not observed in synaptoneurosomes, GABA probably acts on inhibitory interneurons to disinhibit NE release. To determine whether opioids or adenosine influence GABA-augmented NE release, hypothalamic and cortical slices from female rats were superfused with GABA or vehicle in the presence and absence of 10 M morphine or 100 M adenosine. GABA augments [³H]NE release in the cortex and hypothalamus. Morphine alone has no effect on [³H]NE release, but attenuates GABA augmentation of [³H]NE release in both brain regions. Adenosine alone modestly inhibits [³H]NE release in the cortex, but not in the hypothalamus. Adenosine inhibits GABA-augmented [³H]NE release in both brain regions. The general protein kinase inhibitor H-7, augments [³H]NE release in both brain regions and may have additive effects with GABA in cortical slices. These results implicate opioid and adenosine interneurons and possibly protein kinases in regulating GABAergic influences on NE transmission.     

The effects of alpha1-adrenergic and muscarinic cholinergic stimulation on prostaglandin release by rabbit iris

We have investigated the effects of norepinephrine (NE) and acetylcholine (ACh) on prostaglandin (PGE₂ and 6 keto-PGF_1α) production by rabbit iris, measured by radioimmunoassay (RIA), and the type of phospholipase activated by NE in irides in which phosphatidylinositol (PI) was doubly prelabeled with [³H] myo-inositol and [1-¹⁴C] arachidonic acid (¹⁴C-AA), quantitated by radiometric and chromatographic methods. PGE₂ output in 60 min (3.6 μg/g tissue) was 2.6 times greater than 6 keto-PGF_1α. PG production is time-dependent and it is stimulated by NE and ACh in a dose-dependent manner. The Ne- and ACh-induced release of PGE₂, measured by RIA, is mediated through α₁-adrenergic and muscarinic cholinergic receptors, respectively, and it requeires Ca²⁺ for maximal stimulation. Studies on the mechanism of AA release PI in irides doubly prelabeled with ¹⁴C-AA and [³H] myo-inositol revelased the following: (a) Both Ne and ACh increased the breakdown of PI, and this was accompanied by a significant increase in the release of AA and consequently PGE₂. The stimulatory effects of NE and ACh are mediated through α₁-adrenergic and muscainic cholinergic receptors respectively. (b) The NE-induced formation of ³H-lyso PI and the NE-induced metabolism of ¹⁴C-1,2-diacyl-glycerol (DG) are time-dependent. Two pathways for AA release from PI are probably operaitve in the iris: (a) An indirect release by PI-specific phospholipase C which producers DG, followed by the actions of DG- and monoacylglycerol lipases on DG to release AA. (b) A direct release by phospholipase A₂. Whether lyso PI is a product of the polypholipids such as prosphatidylcholine and phosphatidylethanolamine could also serve as a source for AA in PG synthesis. In conclusion, the data presented provide evidence that in the iris the neurotransmitter-stimulated release of PG and AA, from phosphoinositides, for PG synthesis is coupled to the activation of α₁-adrenergic and muscarinic cholinergic receptors.     

Comparative Effects of Hydrogen Sulfide-Releasing Compounds on [<Superscript>3</Superscript>H]D-Aspartate Release from Bovine Isolated Retinae

We investigated the pharmacological actions of a slow-releasing H₂S donor, GYY 4137; a substrate for the biosynthesis of H₂S, l-cysteine and its precursor, N-acetylcysteine on potassium (K⁺; 50 mM)-evoked [³H]D-aspartate release from bovine isolated retinae using the Superfusion Method. GYY 4137 (10 nM–10 µM), l-cysteine (100 nM–10 µM) and N-acetylcysteine (10 µM–1 mM) elicited a concentration-dependent decrease in K⁺-evoked [³H]D-aspartate release from isolated bovine retinae without affecting basal tritium efflux. At equimolar concentration of 10 µM, the rank order of activity was as follows: l-cysteine?>?GYY 4137?>?N-acetylcysteine. A dual inhibitor of the biosynthetic enzymes for H₂S, cystathionine β-synthase (CBS) and cystathionine γ-lyase (CSE), amino-oxyacetic acid (AOA; 3 mM) reversed the inhibitory responses caused by GYY 4137, l-cysteine and N-acetylcysteine on K⁺-evoked [³H]D-aspartate release. Glibenclamide (300 µM), an inhibitor of K_ATP channels blocked the inhibitory action of GYY 4137 and l-cysteine but not that elicited by N-acetylcysteine on K⁺-induced [³H]D-aspartate release. The inhibitory effect of GYY 4137 and l-cysteine on K⁺-evoked [³H]D-aspartate release was reversed by the non-specific inhibitor of nitric oxide synthase (NOS), l-NAME (300 µM). Furthermore, a specific inhibitor of inducible NOS (iNOS), aminoguanidine (10 µM) blocked the inhibitory action of l-cysteine on K⁺-evoked [³H]D-aspartate release. We conclude that both donors and substrates for H₂S production can inhibit amino acid neurotransmission in bovine isolated retinae, an effect that is dependent, at least in part, upon the intramural biosynthesis of this gas, and on the activity of K_ATP channels and NO synthase.     

Evidence that Inhibition of Nicotine-Mediated Catecholamine Secretion from Adrenal Chromaffin Cells by Enkephalin, β-Endorphin, Dynorphin (1-13), and Opiates is not Mediated via Specific Opiate Receptors

Abstract: The opioid peptides Met- and Leu-enkephalin, dynorphin (1-13), and β-endorphin and the narcotic analgesics, morphine, levorphanol, and dextrorphan all produced a dose-dependent inhibition of nicotine (5 × 10^?6m )-mediated release of [³H]norepinephrine ([³H]NE) from bovine adrenal chromaffin cells in culture. None of these agents affected [³H]NE release induced by high K⁺ (56 mm ). Although the above results suggest that the opioid peptides and narcotic analgesics inhibit catecholamine release from adrenal chromaffin cells in culture, we suggest that these effects are not mediated by specific opiate binding sites, since (1) the inhibition was only produced with high concentrations of the agents—the threshold concentrations were 10^?7 to 10^?5m and higher; (2) the inhibition produced by the narcotic analgesics did not display stereospecificity, because the (d-isomer, dextrorphan, was slightly more active than the l-isomer, levorphanol; (3) the narcotic antagonists naloxone, naltrexone, and levallorphan did not reverse the inhibition produced by either the narcotic analgesics (e.g., morphine) or the opioid peptides (e.g., dynorphin). These three antagonists themselves inhibited the nicotine-mediated release of [³H]NE from the adrenal chromaffin cells in culture. Finally (4), the I₂-Tyr¹ substituted analogues of β-endorphin and dynorphin that are biologically less active than the parent compounds produced an inhibition of the nicotine-mediated [³H]NE release similar to that of their parent compounds. These results do not support the idea that high-affinity stereospecific opiate binding sites are involved in the inhibitory modulation of nicotinic evoked catecholamine release from bovine adrenal chromaffin cells in culture.     

Succinate is the controller of O<Stack><Subscript>2</Subscript><Superscript>−</Superscript></Stack>/H<Subscript>2</Subscript>O<Subscript>2</Subscript> release at mitochondrial complex I : negative modulation by malate,positive by cyanide

Mitochondrial production of H₂O₂ is low with NAD substrates (glutamate/pyruvate, 3 and 2 mM) (G/P) and increases over ten times upon further addition of succinate, with the formation of a sigmoidal curve (semimaximal value at 290 μM, maximal H₂O₂ production at 600 μM succinate). Malate counteracts rapidly the succinate induced increased H₂O₂ release and moves the succinate dependent H₂O₂ production curve to the right. Nitric oxide (NO) and carbon monoxide (CO) are cytochrome c oxidase inhibitors which increase mitochondrial ROS production. Cyanide (CN⁻) was used to mimic NO and CO. In the presence of G/P and succinate (300 μM), CN⁻ progressively increased the H₂O₂ release rate, starting at 1.5 μM. The succinate dependent H₂O₂ production curve was moved to the left by 30 μM CN⁻. The V_max was little modified. We conclude that succinate is the controller of mitochondrial H₂O₂ production, modulated by malate and CN⁻. We propose that succinate promotes an interaction between Complex II and Complex I, which activates O₂⁻ production.     

Effect of Hydrogen Sulfide on Sympathetic Neurotransmission and Catecholamine Levels in Isolated Porcine Iris-Ciliary Body

In the present study, we investigated the pharmacological action of hydrogen sulfide (H₂S, using sodium hydrosulfide, NaHS, and/or sodium sulfide, Na₂S as donors) on sympathetic neurotransmission from isolated, superfused porcine iris-ciliary bodies. We also examined the effect of H₂S on norepinephrine (NE), dopamine and epinephrine concentrations in isolated porcine anterior uvea. Release of [³H]NE was triggered by electrical field stimulation and basal catecholamine concentrations was measured by high performance liquid chromatography (HPLC). Both NaHS and Na₂S caused a concentration-dependent inhibition of electrically evoked [³H]NE release from porcine iris-ciliary body without affecting basal [³H]NE efflux. The inhibitory action of H₂S donors on NE release was attenuated by aminooxyacetic acid (AOA) and propargyglycine (PAG), inhibitors of cystathionine β-synthase (CBS) and cystathionine γ-lyase (CSE), respectively. With the exception of dopamine, NaHS caused a concentration-dependent reduction in endogenous NE and epinephrine concentrations in isolated iris-ciliary bodies. We conclude that H₂S can inhibit sympathetic neurotransmission from isolated porcine anterior uvea, an effect that is dependent, at least in part, on intramural biosynthesis of this gas. Furthermore, the observed action of H₂S donors on sympathetic transmission may be due to a direct action of this gas on neurotransmitter pools.     

Stimulation of Phospholipase A2 and Secretion of Catecholamines from Brain Synaptosomes by Potassium and A23187

Abstract: Potassium depolarization of rat brain synaptosomes (containing incorporated l-acyl-2-[¹⁴C]arachidonyl-phosphatidylcholine) stimulated endogenous phospholipase A₁ (EC 3.1.1.32) and A₂ (EC 3.1.1.4), as determined by the formation of [¹⁴C]lysophosphatidylcholine, [¹⁴C]arachidonate, and [¹⁴C]prostaglandins, and also stimulated the secretion of [³H]catecholamines. The phospholipase A₂ stimulation, dependent on calcium, was elicited in resting synaptosomes by A23187 and was demonstrated with incorporated 1-acyl-2-[^l4C]oleoyl-phosphatidylcholine but not with incorporated [^I4C]phosphatidylethanolamine or [^l4C]phosphatidylserine. Inhibitors of phospholipase A₂ [p-bromophenacylbromide (10 μM), trifluoperazine (3 μM), and quinacrine (3 μM) reduced the potassium-stimulated [³H]catecholamine release from synaptosomes to 78, 39. and 55%, respectively, of depolarized controls. The addition of lysophosphatidylcholine increased the release of [³H]norepinephrine to levels observed with potassium depolarization, whereas lysophosphatidylethanolamine, lysophosphatidylserine, and sodium dodecyl sulfate were much less effective. Potassium stimulation of synaptosomes increased the endogenous levels of free arachidonic acid and prostaglandins E₂ and F_2α. Indomethacin and aspirin decreased the amounts of prostaglandins formed, allowed the accumulation of free arachidonic acid, and diminished the potassium-stimulated release of [³H]dopamine. p-Bromophenacylbromide inhibited the formation of prostaglandin F_2α. Addition of prostaglandin E₂ inhibited, whereas prostaglandin F_2α enhanced the release of [³H]norepinephrine. These results suggest that calcium influx induced by synaptosomal depolarization activates endogenous phospholipase A₂, with subsequent formation of lysophosphatidylcholine and prostaglandins, both of which may modulate neurosecretion.     

Effect of Oxygen-Derived Free Radicals and Oxidants on the Degradation in vitro of Membrane Phospholipids

The abilities of chemically generated hydroxyl radical (OH), superoxide anion (O^?) and hydrogen peroxide (H₂O₂) to degrade rat myocardial membrane phospholipids previously lableed with [1 -¹⁴C]arachidonic acid were studied. HO and H₂O₂ but not O₂^?_?, caused the degradation of phospha-tidylcholine (PC), phosphatidylethanolamine (PE), and phosphatidylinositol (PI). With OH' and H₂O₂, the loss of radiolable in PC was accompanied by an increase in the radiolabel of lysophosphatidylcholine (LPC), but not in that of free fatty acid (FFA). These results suggest the hydrolysis of l-oxygen ester bond of PC by HO' and that H₂O₂ and that HO' and H₂O₂, but not O^?, are detrimental to the structure and function of membrane phospholipids. However, since μM amounts of HO' and mM amounts of H₂O₂ were necessary to affect the membrane phospholipids, it is likely that in the reprefused myocardium only HO', but not H₂O₂, may directly cause the breakdown of membrane phospholipids.     

Selective synthesis of triangular cluster oxido-sulfidocomplexes of Mo and W: High yield preparations of [Mo3O2S2(H2O)9], [W3O2S2(H2O)9], [W2MoO2S2(H2O)9] and their derivatization

Reaction of [Mo₂O₂(μ-S)₂(H₂O)₆]²⁺ with Mo(CO)₆ or metallic Mo under hydrothermal conditions (140 °C, 4 M HCl) gives oxido-sulfido cluster aqua complex [Mo₃(μ₃-S)(μ-O)₂(μ-S)(H₂O)₉]⁴⁺ (1). Similarly, [W₃(μ₃-S)(μ-O)₂(μ-S)(H₂O)₉]⁴⁺ (2) is obtained from [W₂O₂(μ-S)₂(H₂O)₆]²⁺ and W(CO)₆. While reaction of [Mo₂O₂(μ-S)₂(H₂O)₆]²⁺ with W(CO)₆ mainly proceeds as simple reduction to give 1, [W₂O₂(μ-S)₂(H₂O)₆]²⁺ with Mo(CO)₆ produces new mixed-metal cluster [W₂Mo(μ₃-S)(μ-O)₂(μ-S)(H₂O)₉]⁴⁺ (3) as main product. From solutions of 1 in HCl supramolecular adduct with cucurbit[6]uril (CB[6]) {[Mo₃O₂S₂(H₂O)₆Cl₃]₂CB[6]}Cl₂⋅18H₂O (4) was isolated and structurally characterized. The aqua complexes were converted into acetylacetonates [M₃O₂S₂(acac)₃(py)₃]PF₆ (M₃ = Mo₃, W₃, W₂Mo; 5a-c), which were characterized by X-ray single crystal analysis, electrospray ionization mass spectrometry and ¹H NMR spectroscopy. Crystal structure of (H₅O₂)(Me₄N)₄[W₃(μ₃-S)(μ₂-S)(μ₂-O)₂(NCS)₉] (6), obtained from 2, is also reported.     

Regulation by prostaglandin E2 of interleukin release by T lymphocytes in mucosa

Regulation of immune cell activation in lymphocyte-bearing human tissues is a pivotal host function, and metabolites of arachidonic acid (prostaglandin E₂ in particular) have been reported to serve this function at non-mucosal sites. However, it is unknown whether prostaglandin E₂ is immunoregulatory for the large lymphocyte population in the lamina propria of intestine; whether low (nM) concentrations of prostaglandin E₂ modulate immune responses occurring there; and whether adjacent inflammation per se abrogates prostaglandin E₂'s regulatory effects. To address these issues, intestine-derived lymphocytes and T hybridoma cells were assessed, T cell activation was monitored by release of independently quantitated lymphokines, and dose-response studies were performed over an 8-log prostaglandin E₂concentration range. IL-3 release by normal intestinal lamina propria mononuclear cells was reduced (up to 78%) in a dose-dependent manner by prostaglandin E₂, when present in as low a concentration as 10⁻¹⁰M. PGE₂ also inhibited(by ≥ 60%) mucosal T lymphocytes' ability to destabilize the barrier function of human epithelial monolayers. Further, with an intestine-derived T lymphocyte hybridoma cell line, a prostaglandin E₂ dose-dependent reduction in IL-3 and IL-2 (90 and 95%, respectively) was found; this was true for both mitogen- and antigen-driven T cell lymphokine release. Concomitant [³H] thymidine uptake studies suggested this was not due to a prostaglandin E²-induced reduction in T cell proliferation or viability. In contrast, cells from chronically inflamed intestinal mucosa were substantially less sensitive to prostaglandin E², e.g., high concentrations (10⁻⁶ M) of prostaglandin E² inhibited IL-3 release by only 41%. We conclude that prostaglandin E² in nM concentrations is an important modulator of cytokine release from T lymphocytes derived from the gastrointestinal tract, and it may play a central role in regulation of lamina propria immunocyte populations residing there. © 1996 Wiley-Liss, Inc.     

A passive apparatus for controlled-flux delivery of biocides: hydrogen peroxide as an example

A new test method has been developed to estimate the required release rate of hydrogen peroxide (H₂O₂) to prevent marine biofouling. The technique exploits a well-defined concentration gradient of biocide across a cellulose acetate membrane. A controlled flux of H₂O₂, an environmentally friendly biocide, was obtained. Larvae of the barnacle, Balanus improvisus, were subjected to known release rates of H₂O₂ from a surface, under laboratory conditions. It was found that the distribution of settled larvae was not significantly different from the controls when H₂O₂ fluxes of 5–8 μg cm^?2 day^?1 were applied. However, release rates of 40 μg cm^?2 day^?1 significantly displaced the distribution of settled larvae towards the area of the chamber farthest away from the membrane. Membrane tests in seawater (Jyllinge Harbour, Denmark) for over 16 weeks showed that release rates of H₂O₂ of approximately 2800 μg cm^?2 day^?1 deterred biofouling efficiently. A H₂O₂ release rate of about 224 μg cm^?2 day^?1 resulted in some slime formation, but it was less than that on the H₂O₂-free control. It appears that to obtain efficient resistance to biofouling in natural seawater requires much higher membrane release rates of H₂O₂ (factor of between 5 and 50) than laboratory membrane exposure assays using barnacle larvae.