We have evidence that 15-F2-isoprostanes (15-F2-IsoPs) regulate excitatory neurotransmitter release in ocular tissues. Although 5-F2-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-F2-IsoP epimer pair, 5-F2t-IsoP (C5–OH in β-position) and 5-epi-5-F2t-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-F2t-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-F2t-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; IC25 = 0.2 μM) being achieved at 1 μM concentration. Its 5-(S)-OH-epimer, 5-F2t-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 (IC25 value of 3 nM). Although the prostanoid-receptor antagonists, AH 6809 (10 μM; EP1–3/DP) and BAY-u3405 (10 μM; DP/Tx) exhibited no effect on 5-epi-5-F2t-IsoP (10 nM–1 μM)-mediated inhibition, SC-19220 (1 μM; EP1) completely reversed 5-epi-5-F2t-IsoP (0.1 μM and 1 μM)-induced attenuation of K+-evoked [3H]D-aspartate release. Similarly, both SC-51322 (10 μM; EP1) and AH 23848 (1 μM; EP4) reversed the inhibitory action elicited by 5-epi-5-F2t-IsoP (0.1 μM) on the neurotransmitter release. We conclude that the 5-F2-IsoP epimer pair, 5-F2t-IsoP and 5-epi-5-F2t-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-F2t-IsoP on excitatory neurotransmitter release.

Neuroprostanes (nPs) are a series of isoprostane (IsoP)‐like compounds that are spontaneously formed in vivo and in vitro by free‐radical catalyzed peroxidation of the polyunsaturated fatty acid (PUFA), docosahexanoic acid. Although nPs are elevated in neurodegenerative conditions and in animal models of neuronal oxidative stress (Musiek et al., Brain Pathol. 15:149,2005), it is unknown whether nPs, like IsoPs, are endogenously produced in mammalian retina. We sought to measure endogenous production of the PUFA metabolites, A4‐nPs and F2‐IsoPs in bovine retinae, in vitro. Freshly isolated bovine retinae were exposed to hydrogen peroxide (H2O2) and cumene hydroperoxide (cuOOH) for up to 6h, in vitro. PUFA metabolites were measured by the stable isotope dilution methods employing GS/negative ion chemical ionization MS. Basal endogenous levels of A4‐nPs and F2‐IsoPs were found to be 35.0± 2.2 and 1.1± 0.001 ng/g tissue, respectively. Exposure of retina to H2O2 (1 mM) for 1h and 6h stimulated production of A4‐nP levels by 15% and 56% while bound F2‐IsoPs were elevated by 44% and 68%, respectively. Similarly, cuOOH (1 mM) enhanced (p<0.001) A4‐nP levels by 408% and 420% and F2‐IsoPs by 831% and 918% after 1h and 6h periods of treatment, respectively. In conclusion, basal levels of nPs exceed that of IsoPs in bovine retina. Furthermore, endogenous production of PUFA metabolites is enhanced by oxidant stress in retina.