| BackgroundBreathing is the precursor of all life activities in mammals,the basic rhythmic respiratory discharge activity(RRDA)of the respiratory center in medulla oblongata is the basis of respiratory movement.The RRDA of the medulla oblongata originates in the medial region of the nucleus retrofacialis(mNRF).Nicotinic acetylcholine receptors(nAChRs)are ion channel receptors and widely distributed in the central nervous system,such as attention,learning,awakening and cognitive functions.In recent years,most of the studies on the function of nAChRs have focused on the neuroprotective effect of injured central neurons,but the role of nAChRs in the respiratory system of medulla oblongata is relatively little known.With the occurrence of neonatal ischemic-hypoxic encephalopathy and depression of the respiratory center of the medulla oblongata caused by slow neonatal development increasing year by year,It is important to study the mechanism of nAChRs in the respiratory center of medulla oblongata under normal and hypoxic conditions.As an intermediate metabolite of ATP,adenosine indirectly increases ATP concentration by increasing adenosine concentration and regulates neuronal excitability through adenosine receptors.ATP is widely used in clinical treatment of ischemic and hypoxic diseases.In order to study the regulatory effect of nAChRs on RRDA in isolated medullary slices of neonatal rats under normal and hypoxic conditions and whether adenosine is involved in the regulation,we designed and completed this experiment.To provide new experimental basis and theoretical support for the research and treatment of central respiratory diseases.ObjectivesTo investigate the regulatory effect of nAChRs on RRDA in isolated medullary slices of neonatal rats under normal and hypoxic conditions and whether adenosine is involved in the regulation.To improve the function of the respiratory center of medulla oblongata in neonatal animals under hypoxic conditions,the experimental theoretical basis and therapeutic ideas were provided.Methods1.RRDA of isolated medullary slices.Prepared the In vitro medulla oblongata slices of newborn rats and recorded the RRDA of hypoglossal nerve roots.Brain slices were divided into 6 groups,each group consisting of 6 brain slices.The first group was blank control group,to compare the difference of the RRDA at 10 min ?20 min ? 30 min? 40 min?50 min under 95% oxygen concentration,to Test whether the experimental model is established and to provide blank control.The second group was the hypoxic group,artificial cerebrospinal fluid containing 95%,65%and 50% oxygen concentration was used to perfuse brain slices in turn to observe the effect of hypoxia on RRDA.The third group was nicotine group,the effect of nicotine on RRDA was observed at 95% oxygen concentration and the optimum nicotine concentration was selected.The third group was nicotine group.At 95% oxygen concentration,to observe the effects of different concentrations of nicotine on RRDA were and to select the optimal nicotine concentration.The fourth group was mecamylamine group,at 95% oxygen concentration,to observe the effects of different concentrations of methamphetamine on RRDA and to selected the optimum concentration of mecamylamine.The fifth group was hypoxic nicotine group.To observe the effects of 40?mol/L of nicotine on RRDA under 95%?65% and 50% oxygen concentration.The sixth group was hypoxic mecamylamine group.To observe the effects of10?mol/L of mecamylamine on RRDA at95%? 65% and 50% oxygen concentration.2.The adenosine concentration in brain slices was determined by high performance liquid chromatography(HPLC).In this experiment,36 newborn 2d rat medulla oblongata brain slices were prepared and randomly divided into 6 groups,namely:95% normoxic group,50%hypoxic group,95%nicotine group,95%megamine group,50%hypoxic nicotine group and 50%hypoxic megamine group.Brain slices of each group were incubated in artificial cerebrospinal fluid with 95% oxygen concentration,95% oxygen concentration,95% optimal nicotine concentration,95% oxygen concentration,50% oxygen concentration,50% optimal nicotine concentration,and 50% optimal megamine concentration for 20 min,and then the adenosine concentration of brain slices of each group was detected.Results1.In the blank control group,there was no statistical difference in RRDA between 10 min,20 min,30 min,40 min,50 min(P>0.05),which ensured the reliability of the experiment and the stability of the model.Hypoxia inhibited RRDA,with the decrease of oxygen concentration,RRDA gradually decreased,and there was significant difference between different oxygen concentrations(P<0.01).The RRDA was stimulated in nicotine group in the range of 10 ?mol/L to 80?mol/L,and 40?mol/L nicotine concentration is the optimum concentration(P<0.01).The RRDA was inhibited in the mecamylamine group in the range of 2.5?mol/L to 20?mol/L,and 10?mol/L mecamylamine concentration is the optimum concentration(P<0.01).nicotine enhanced RRDA at corresponding oxygen concentration(P<0.01),the effect of nicotine on RRDA decreased with the decrease of oxygen concentration.mecamylamine attenuate the RRDA at corresponding oxygen concentration(P<0.01),the effect of methamphetamine on RRDA increased with the decrease of oxygen concentration.2.Experimental results of HPLC detection showed that: Adenosine concentration at 95% oxygen concentration was 860.47% at 50% oxygen concentration.Nicotine increased adenosine concentration by 252.7% at 95% oxygen concentration,511.63% at 50% oxygen concentration,71.62% at 95% oxygen concentration and 290.69% at 50% oxygen concentration.nAChRs enhanced adenosine concentration during hypoxia,and adenosine participated in the excitatory effect of nAChRs on RRDA in isolated medullary slices of neonatal rats.ConclusionsnAChRs can excite RRDA,and hypoxia inhibits RRDA in isolated medullary slices of neonatal rats.Hypoxia inhibits the excitatory effect of nAChRs on RRDA;Adenosine is involved in the excitatory effect of RRDA on hypoxia by nAChRs... |
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