The Influences Of HIBD On Pineal Melatonin-Synthetic Function In Neonatal Rat

Objectives Hypoxic-ischemic encephalopathy is a common disease resulting in high fatality rate and disablement rate in neonatal department. There have been a lot of evidences that multiple factors participate in its complicated pathological course. But the exact pathological-physiological mechanism of hypoxic-ischemic brain damage (HIBD) has not been thoroughly clarified. By setting up rat HIBD model, we detected melatonin-synthetic enzymes - arylalkylamine-N-acetyltransferase (AANAT) and hydroxyindole-O-methyltransferase (HIOMT) mRNA expressions in pineal gland with semi-quantitative Reverse Transcriptase Polymerase Chain Reaction (RT-PCR) analysis. At the same time we determined plasma melatonin levels with Radioimmimoassay (RIA). The aims were to realize the influences of HIBD on pineal melatonin-synthetic function and to explore the possible significance of pineal function alterations in HIBD. Methods we did experiments with 60 seven-day-old Sprague-Dawley (SD) rats. Rat HIBD model was set up according to modified Levine procedure. Briefly, pup's left carotid artery was sectioned between double ligatures. The pups were allowed to recover for 1-2 h and then exposed to 2h of hypoxia with a mixture of humidified 8% oxygen balanced with nitrogen.60 pups were randomly divided into 10 groups (6 pups each), which were Test group l(0h after HIBD), Test group 2(6h after HIBD), Test group 3(12h after HIBD), Test group 4(24h after HIBD), Test group 5(48h after HIBD) and five corresponding sham-operated groups. We detected AANAT, HIOMT mRNA expressions in pineal gland and plasma melatonin levels of each group at different time. Results 1 .AANAT mRNA expressions in pineal gland: The AANAT mRNA amounts of 6h, 12h, 24h after HIBD were lower than that of corresponding sham-operated groups. The AANAT mRNA expressions of all sham-operated groups were very much alike; The amounts began to decline 6h after HIBD, reached their minimi 12h after HIBD, then rised 24h after HIBD, returned to their former levels 48h after HIBD.2.HIOMT mRNA expressions in pineal gland: Compared with each corresponding sham-operated group, the expression levels of HIOMT mRNA after HIBD were pretty much the same thing; The HIOMT mRNA expressions of all groups were very much alike.3. Plasma melatonin levels: The plasma melatonin levels of 12h, 24h after HIBD were lower than that of corresponding sham-operated groups. The melatoninlevels of all sham-operated groups were similar; The levels began to descend 12h after HIBD equal to that of 24h after HIBD, returned to their former levels 48h after HIBD. Conclusions l.HIBD may lead to down-regulate melatonin-synthetic key enzyme-AANAT gene expression in pineal gland. The AANAT mRNA amounts begin to decline 6h after HIBD, reach their minimi 12h after HIBD, then somewhat rise 24h after HIBD, return to their former levels 48h after HIBD.2. HIBD has no effect on melatonin-synthetic last enzyme-HIOMT gene expression. 3. HIBD can bring on diminution of plasma melatonin concentration. The levels begin to descend 12h after HIBD equal to that of 24h after HIBD, return to their former levels 48h after HIBD.4. Pineal melatonin-synthetic function is impaired on early HIBD stage. Down-regulation of AANAT gene expression is likely the "engine". 5. Knock-down of endogenous protective process of MLT maybe plays a part in early HIBD pathogenic mechanism.