Comparative Effects of Hydrogen Sulfide-Releasing Compounds on [3H]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 ß-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 KATP 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 KATP channels and NO synthase.

Effect of Hydrogen Sulfide Donors on Intraocular Pressure in Rabbits.

PURPOSE: In this study, we investigated the effect of a slow-releasing hydrogen sulfide (H2S) donor, GYY 4137, on intraocular pressure (IOP) in normotensive rabbits. Furthermore, we compared the IOP-lowering action of GYY 4137 with those elicited by other H2S-producing compounds, l-cysteine and ACS67 (a hybrid compound of latanoprost with an H2S-releasing moiety). METHODS: IOP was measured in New Zealand normotensive male albino rabbits using a pneumatonometer (model 30 classic; Reichert Ophthalmic Instruments, Depew, NY). At 0 h, 50 µL of test compounds were applied topically to 1 eye of each animal, while the contralateral eye received the same quantity of vehicle (saline). IOP was measured hourly until baseline IOP readings were attained and animal eyes monitored for potential side effects (i.e., tearing, hyperemia). RESULTS: GYY 4137 (0.1%-2%) produced a dose-dependent decrease in IOP reaching a maximum of 27.8% ± 3.14% (n = 5) after 6 h. Interestingly, a significant contralateral effect was observed in vehicle-treated controls eyes at all doses tested. l-cysteine (5%) and ACS67 (0.005%) also elicited a significant (P < 0.01) decrease in IOP that achieved a maximum of 28.84% ± 1.53% (n = 5) and 23.27% ± 0.51% (n = 5), respectively, after 3 h. All 3 H2S-producing compounds also caused a significant contralateral effect in vehicle-treated control eyes. CONCLUSION: We conclude that GYY 4137 and other H2S-producing donors can reduce IOP in normotensive rabbits. However, the profile of IOP-lowering action of GYY 4137 was different from the other H2S donors affirming its ability to act as a slow-releasing gas donor.

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 [(3)H]-norepinephrine ([(3)H]NE) triggered by electrical field stimulation. ACS67, L-cysteine and GYY 4137 caused a concentration-dependent inhibition of electrically-evoked [(3)H]NE release from isolated bovine iris-ciliary bodies without affecting basal [(3)H]NE efflux. The cyclooxygenase inhibitor, flurbiprofen enhanced the inhibitory action of ACS67 and L-cysteine on stimulated [(3)H]NE release. Both aminooxyacetic acid, an inhibitor of cystathionine-ß-synthase and glibenclamide, a KATP 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 KATP channels.

Role of the non-enzymatic metabolite of eicosapentaenoic acid, 5-epi-5-F3t-isoprostane in the regulation of [ (3)H]D-aspartate release in isolated bovine retina.

We have evidence that F2-isoprostanes (F2-IsoPs) regulate the release of excitatory neurotransmitters in isolated bovine retina. Although 5-F3-IsoPs are generated in mammals, in vivo, their pharmacological actions on neurotransmitter release remain unknown. In this study, we investigated the effect of 5-epi-5-F3t-IsoP on K(+)-evoked [(3)H]D-aspartate release in isolated bovine retina using the superfusion method. Furthermore, we examined the role of arachidonic acid metabolites in the regulation of the neurotransmitter release by this novel IsoP. In the concentration range, 0.01 nM-0.1 µM, 5-epi-5-F3t-IsoP inhibited K(+)-evoked [(3)H]D-aspartate release in a concentration-dependent manner, achieving a maximum inhibition of 46.9 % at 0.1 µM (IC30 = 1 nM). The prostanoid receptor antagonists, AH 6809 (EP1-3/DP; 10 µM), SC 51322 (EP1; 10 µM) and SC 19220 (EP1; 1 µM) partially reversed 5-epi-5-F3t-IsoP-mediated inhibition of K(+)-induced [(3)H]D-aspartate release. Pretreatment of retinal tissues with the cyclooxygenase (COX) inhibitor, flurbiprofen (3 µM) unmasked a biphasic action of 5-epi-5-F3t-IsoP that was inhibitory at lower (0.1-10 pM) and stimulatory at higher concentrations (≥0.1 nM). The prostanoid pathway antagonists, BAY-u3405 (10 µM; TP/DP-receptors), SQ 29548 (10 µM; TP-receptor) and ozagrel (10 µM; Tx-synthase inhibitor) abolished the stimulatory action of the 5-epi-5-F3t-IsoP (0.1 µM) on neurotransmitter release. In conclusion, 5-epi-5-F3t-IsoP attenuates K(+)-induced [(3)H]D-aspartate release in a concentration-dependent manner by mechanisms that are partially dependent on activation of pre-junctional prostanoid EP1-receptors. Moreover, blockade of the COX-pathway unmasks a biphasic action for 5-epi-5-F3t-IsoP that is inhibitory at low concentrations and stimulatory at higher concentrations. Products of the thromboxane synthase pathway may partially account for the stimulatory action of this F3-IsoP on isolated bovine retina.