Change Of Clock Genes' Expression And Plasma Melatonin In HIBD Neonatal Rat Pineal

Background hypoxic-ischemic brain damage(HIBD) survivals often show disorders of psychomotor development and sleep-waking cycle. Circadian rhythm is known as the adaptation of the organism to the rhythmic environment, which ensures proper temporal organization of physiological processes and is related to the pathologic processes of many diseases. Comparing with the central master clock located in the suprachiasmatic nuleus(SCN), the pineal gland ,as a neuro-endocrine organ, not only acts like an central circadian oscillator, but also presents endocrine output effect by secreting mlatonin(MLT). The basic circadian structure is the intracellular circadian oscillator, which is made up of core clock genes. These genes and their expression products form interlocked feedback loops of transcription / translation, two significant genes among which are Clock mRNA and Per mRNA. However, Studies about circadian disturbance in HIBD are so few that the mechanism of circadian disorders in HIBD is indefinite.Objective To explore the effects of clock genes on Circadian disorder in hypoxic- ischemic brain damage(HIBD) by studying the expression of Clock mRNA, Per2 mRNA and CLOCK protein in neonatal rat pineal gland and plasma melatonin after HIBD.Methods 7-day-old Sprague-Dawley (SD) rats were randomly divided into 2 groups (36 pups each). HIBD models were set up according to modified Levine euthanized 0,2,12,24,36,48 hours afterwards. Semi-quantitative Reverse Transcriptase Polymerase Chain Reaction(RT-PCR)analysis was used to measure the expression profiles of Clock mRNA and Per2 mRNA , western blot for CLOCK protein in pineal gland and Radioimmunoassay(RIA) for plasma melatonin(MLT) concentrations, respectively.Results 1,Comparing with each corresponding sham-operated group,the expression level of Clock mRNA after HIBD was pretty much the same thing;The Clock mRNA expressions of all groups were very much alike,though a slight decrease of Clock mRNA in HIBD group can be observed(p>0.05). 2,There were no rhythmic variations of Per2 mRNA in sham-operated groups, but the amount of 24h after HIBD began to decline, and that of 36h and 48h displayed significantly lower than the sham-operated groups, respectively(p〈0.01). 3,the plasma melatonin concentrations began to decline 12h after HIBD, reached their minimi 24h after HIBD and returned to the former levels at 48h after HIBD. 4,The expression of CLOCK protein in pineal began to decrease at 2h after HIBD, back to the normal level in 24h after HIBD.Conclusion 1,There exists expression of Clock and Per2 mRNA and CLOCK protein in neonatal rat pineal. 2,Change of Clock mRNA expression seems hardly be observed under HIBD, however the down-regulation of Per2 mRNA and CLOCK protein expression in pineal gland and plasma melatonin concentrations under HIBD may indicate that clock genes may play a leading role in the mechanism of circadian disorder in HIBD.