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Cytochrome P450-epoxygenase metabolites play role in vasoconstriction of middle cerebral arteries in response to serotonin in healthy and diabetic rats that underwent hyperbaric oxygenation

INTRODUCTION: Our previous study demonstrated altered mechanisms of vasorelaxation in responses to acetylcholine in rats treated with hyperbaric oxygenation, involving arachidonic acid metabolites of CYP450-4A enzymes (1). The aim of the present study was to determine if vasoconstrictor response to serotonin (Ser), known to be mediated primarily with COX-originating vasoconstrictor metabolites is altered by diabetes mellitus and/or hyperbaric oxygenation.METHODS: Male Sprague-Dawley rats (N= 12-17/per group) were divided into 4 groups: control, diabetic (DM ; streptozocin 60mg/kg i.p.) and control+HBO2, and diabetic rats (DM-HBO2) that underwent hyperbaric oxygenation. HBO2 rats were treated in a hyperbaric chamber with 100% O2 (2 bar) 2 hours/day for 4 consecutive days. ON 5th day, prior to decapitation and vessel harvesting, rats were anesthetized with ketamin (75 mg/kg) and midazolam (2.5 mg/kg) i.p. Isolated middle cerebral arteries (MCA) s were mounted on glass pipettes of DMT 110P pressure myograph system for internal diameter measurements. Vasoconstriction responses to serotonin (10-6 M) were tested in the presence/absence of indomethacin (COX inhibitor ; 10-5M), DDMS (20-HETE inhibitor ; 10-5M) and combination of both inhibitors. Results are shown as mean ± SEM (microns). To compare differences among groups One way ANOVA was used ; p<00.5 was considered statistically significant. The Ethical Committee of Faculty of Medicine University of Osijek approved the study.RESULTS: Vasoconstriction in response to serotonin was preserved in all groups of rats (control=-23±9 (n=12), DM=-20±9 (n=12), Control+HBO2=-22±8 (n=17), DM+HBO2=-20±5 (n=17). Indomethacin completely blocked vasoconstriction in control group (0±0 (n=5)) and in DM group of rats (0±0 (n=5)). Vasoconstriction of MCA of control+HBO2 and DM+HBO2 rats was partially blocked by indomethacin (control+HBO2=-14±8 (n=11) ; DM+HBO2=-9±4 (n=10))) and almost completely blocked by DDMS+Indomethacin (control+HBO2= -4±4 (n=6) ; DM+HBO2 =-2±4 (n=6)).CONCLUSION: The presented results support the hypothesis that HBO2 alters mechanisms of vasoconstriction in response to serotonin in MCA of both, control and DM rats. There is a switch from vasoconstriction entirely mediated by COX- produced vasoconstrictor prostaglandins in HBO2 untreated rats to partial involvement of CYP450-4A metabolites, presumably 20-HETE in HBO2 treated rats, irrespectively of DM.Key words: 20-HETE, serotonin, diabetes mellitus, Sprague-Dawley rats, microcirculation

cytochrome P450-epoxygenase; middle cerebral artery; serotonin; diabetes mellitus; hyperbaric oxygenation

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