| Glaucoma is one of the most common irreversible blinding eye diseases worldwide.The pathogenesis of primary open-angle glaucoma(POAG)is very complicated.It is currently recognized that the pathological increase and abnormal fluctuation of intraocular pressure(IOP)is the most important risk factor for the progression of the disease.And the abnormal fluctuation of IOP has a greater impact on the disease development than the increase of a single intraocular pressure value.Researches have shown that the IOPs of human and rats both have the characteristic of circadian rhythmic fluctuation.Change of the circadian environment can cause the change of rats' activity pattern and IOP.However,whether the fluctuation rhythm of IOP is related to the activity habitual nature of individuals? It still requires further exploration.The stability of IOP depends on the dynamic balance of the production and the outflow of aqueous humor(AH).In patients with glaucoma,increasing resistance of the AH outflow channel is the main reason for the increase of IOP.Schlemm's canal(SC)and trabecular meshwork are the vital parts causing the resistance to the outflow of AH.Autonomic nervous system can cause fluctuation of IOP by regulating the diameter of Schlemm's canal(SC).And the excitement of sympathetic nervous system(such as exercise)and adrenaline(eye-dropping)can reduce intraocular pressure by expanding Schlemm's canal.In addition,the expression of sympathetic nerve marker dopamine-?-hydroxylase(D?H)is observed at Schlemm's canal and trabecular meshwork,which suggests that there are sympathetic nerve fibers.However,the mechanism by which sympathetic nerve fibers regulate the morphology of SC and IOP is still unclear.This research is divided into the following two parts,to explore the source of sympathetic nerve fibers at SC and whether the fibers can regulate IOP by regulating the morphology of SC,and to explore whether the circadian rhythmic fluctuation of IOP is related to the different degree of sympathetic nerve excitement,which is caused by the different circadian activity habitual nature of individuals.Part 1 Effects of Different Circadian Rhythms and Electrical Stimulation of the Superior Cervical Sympathetic Ganglion on Intraocular Pressure in RatsPurpose: To investigate the relationship between circadian rhythm and intraocular pressure,and to explore the effect of electrical stimulation of the superior cervical sympathetic ganglion(SCG)on IOP and fluctuation of IOP.Methods: Sprague Dawley rats weighting 220 ± 20 g were housed under normal(N-Normal),constant dark(N-Dark),and constant light(N-Light)rhythms.After 24 hours and during the following 48-hour period,Icare PRO tonometer was used to measure the intraocular pressure,and IOP was measured at 16:00,20:00,00:00,04:00,08:00,12:00.Then the eyes were collected.Rats were raised under different circadian rhythms.After 72 hours,electrical stimulation(parameters: intermittent wave,20 Hz,2 m A,10 minutes)was given to the right SCGs,and then,they were raised under the corresponding circadian rhythm environment,and after 24 hours,Icare PRO tonometer was used to monitor the intraocular pressure of the right eyes for 48 hours.The time points of IOP measurement were 16:00,20:00,00:00,04:00,08:00,12:00.After finishing the measurement,the right eyes were collected.Two-tailed unpaired t-test was used to analyze the mean value,peak value,nadir value and 24-hour fluctuation range of IOP in each group.Results: Compared with the N-Normal group,the intraocular pressure of rats increased in the N-Dark group,and the difference in the fluctuation amplitude of intraocular pressure between N-Normal and N-Dark groups had no statistical significance.Under normal circadian rhythm,electrical stimulation of SCG increased IOP and decreased IOP fluctuation.Under constant light rhythm,electrical stimulation of SCG also increased IOP and decreased IOP fluctuation.Under constant dark rhythm,electrical stimulation of SCG had no significant effect on IOP and IOP fluctuation range of rats,compared with the control group.Conclusions: Under constant dark rhythm,intraocular pressure of rats increased,but the short-term alteration of circadian environment could not cause significant change in fluctuation amplitude of IOP.Under normal and constant light rhythms,electrical stimulation of SCG increased intraocular pressure and decreased IOP fluctuation of rats.Part 2 Effects of Different Circadian Rhythms and Electrical Stimulation of the Superior Cervical Sympathetic Ganglion on the Morphology of Schlemm's Canal and Sympathetic Neurotransmitter around Schlemm's Canal in RatsPurpose: To explore whether alteration of IOP caused by different circadian rhythms and electrical stimulation of SCG is related to the expression of sympathetic neurotransmitter around SC and the morphology of SC.Methods: In the eyes from rats' models in the first part of the research,hematoxylin-eosin(HE)staining was used to observe the morphology of SC,and Image J was used to measure the cross-sectional area and circumference of SC.And after immunofluorescence staining,Image J was used to evaluate the expression of sympathetic nerve marker dopamine-?-hydroxylase(D?H)in Schlemm's canal endothelial cells(SCEs).Sprague Dawley rats weighting 220 ± 20 g were housed under normal(N-Normal),constant dark(N-Dark),and constant light(N-Light)rhythms,and Atropine was applied to the right eyes for 6 days(3 times/day).After 72 hours,electrical stimulation(parameters: intermittent wave,20 Hz,2 m A,10 minutes)was given to the right SCGs,and then,they were raised under the corresponding circadian rhythm environment.After 72 hours,the right eyes were collected,then HE staining was used to evaluate the cross-sectional area and circumference of SC.Dil fluorescent tracer was injected into the right SCGs of rats.After 5 days,the right eyes were collected to observe the Dil fluorescent around the SC.After electrical stimulation(parameters: intermittent wave,20 Hz,2 m A,10 minutes)was given to the right SCGs,immediately,the ipsilateral eyes were collected to evaluate the expression of D?H in SCEs with immunofluorescence staining.Two-tailed unpaired t-test was used to analyze the data of these groups.Results: In the N-Dark group,the cross-sectional area of SC decreased,and the expression of D?H in SCEs increased.Nerve projections between SC and SCG were detected,and electrical stimulation of SCG up-regulated D?H expression in SCEs.Under normal and constant light rhythms,electrical stimulation of SCG increased the expression of D?H,and decreased the cross-sectional area and circumference of SC.After paralyzing the ciliary muscles,electrical stimulation of SCG decreased the cross-sectional area and circumference of SC under normal and constant light rhythms.Conclusions: Under constant dark rhythm,the expression of sympathetic neurotransmitter around SC increased,and the cross-sectional area of SC decreased.There are noradrenergic nerve fibers between SC and SCG.Under normal and constant light rhythms,electrical stimulation of SCG increased sympathetic neurotransmitter in SCEs,and reduced the cross-sectional area and circumference of SC.SCG can regulate the morphology of SC without entirely relying on the traction of the ciliary muscles.Combining the results of part one and part two,it is speculated that SCG may regulate intraocular pressure by regulating the expression of sympathetic neurotransmitter around SC and the morphology of SC. |
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