| Objective:1. To study on changes of atrial effective refractory period (AERP) and monophasic action potential (MAP) and investigate atrial electrical remodeling by rapid atrial pacing in rabbits with intracardiac electrophysiology technique. To study on field action potential duration (fAPD) of atrium and pulmonary veins (PV), and explore calcium and potassium ion channels remodeling with Microelectrode arrays (MEA) technique by RAP in right atrium of rabbits.2. The experiment study on histological changes and remoding of autonomic nervous of pulmonary veins (PV) myocardial sleeves in rapid atrial pacing (RAP) right atrium of rabbits and dissolve mechanisms of the onset and maintenance of AF.3. The experiment study on changes of discharge activity and remoding of autonomic nervous in RAP right atrium of rabbits. Methods:1. Twenty New Zealand white rabbits of either sex (weight 2.5 to 3.0 kg, n=20) were were randomly divided into 2 groups:sham operated group (n=10) and model group (n=10). All animals were esthetized with pentobarbital (30mg/kg followed by 5mg/kg/h, i.v.). AERP was measured by programmed electrical stimulation before pacing and from 0 to 24 hours after the onset of the pacing.20 rabbits were randomely divided into 2 groups:sham operated group (n=10), model group (n=10). The hearts were quickly removed. right atrial appendage (RAA) and PV were sliced (slice thickness 500μm). each slice was perfused with Tyrode's solution and contiuous stimulated about 30min. Sham operated groups received Tyrode's solution superfusion 10 min, blocker groups and amiodarone groups received blocker and amiodarone superfusion 10 min, respectively.We recorded RAA and PV field action potential duration (fAPD) and field action potential morphology with MEA technique.2. Twenty New Zealand white rabbits of either sex (weight 2.5 to 3.0 kg, n=20) were were randomly divided into 2 groups:sham operated group (n=10) and model group (n=10). All animals were esthetized with pentobarbital (30mg/kg followed by 5 mg/kg/h, i.v.). The hearts were quickly removed.RAA and PV sample were collected. RAA and pulmonary vein sleeves was observed under light microscope:the structure of PV was observed by HE staining, myocardial fibrosis was displayed by Masson staining, myocardial abundant glycogen was displayed by PAS staining. Ultrastructural observation showed under electron microscope.3. Twenty New Zealand white rabbits of either sex (weight 2.5 to 3.0 kg, n=20) were were randomly divided into 2 groups:sham operated group (n=10) and model group (n=10). All animals were esthetized with pentobarbital (30 mg/kg followed by 5 mg/kg/h, i.v.). We recorded discharge activity of autonomic nervous and amplitude of integrated discharges (AID), time course of discharges (TCD) and time course of discharge interval (TCDI) was indexes of discharge activity. Right atrial appendage and PV samples were obtained and the experssions of some nerve makers protein were studyed in the immunofluorescence technique. Results:1. The AERP and MAP was shortened and the rate adapation of AERP was lost through the pacing process compared with those before pacing (P<0.05). a) In sham operated groups, RAA fAPD was (188.33±18.29) ms after Tyrode's solution superfusion, and fAPD was (173.91±6.83) ms after RAP. In model groups, CdCl2, TEA and BaCl2 superfusion prolonged atrial field action potential (fAPD) (control vs blocker:176.67±8.66 ms vs 196.11±10.76 ms, 182.22±12.87 ms vs 191.11±13.09 ms with TEA and BaCl2 superfusion, respectively, P< 0.05).4-AP superfusion prolonged significantly FAPD (control vs 4-AP:169.38±10.56 ms vs 188.56±13.82 ms, P<0.01). In pace/Amiodarone groups,4-AP superfusion extended Obviously fAPD (control vs Amiodarone:167.38±13.67 ms 185±15.14 ms, P<0.01). b) In sham operated groups, PV fAPD was (187.67±16.33) ms and amplitude of field potential (FP) was (-189.44±17.04)μV after Tyrode's solution superfusion, and fAPD was (170.83±10.65) ms and amplitude of FP was (-53.75±8.76)μV (n=10) in model groups.Compared with sham operated groups, fAPD shortened significantly (P<0.01) in model groups.2. RAA:Histological examination revealed remarkable myocardial hypertrophy, irregular array of cardiocyte, Central part of myocytes is free of sarcomeres and contains abundant glycogen (PAS-positivestaining) and myocardium interstitial fibrosis. PV:In model groups. HE staining revealed interstitial edema and myocardial sleeves (MS) fiber lining up wavily, contains abundant glycogen (PAS-positivestaining) and myocardium interstitial fibrosis after RAP 24h.vacuolization in some endochylema, intermuscular transverse striation were notclear.Ultrastructural observation showed that MS fibers were well arranged in sham operated groups, mitochondria were morphologically normal, nuclear membrane was smooth.After 24h of rapid pacing, mitochondria were deformed significantly, showing even more apparent vacuolization and glucogen accumulation as well as ruptures of cristal membrane, both of which were randomly distributed using electronmicroscopy in model groups.3. Compared with sham operated groups, vagus nerve AID prolonged significantly (sham operated groups vs model groups:32.4±0.78μV·ms vs 50.47±3.14μV·ms, P<0.01, n=10), but TCD and TCDI were not change (P>0.05) in model groups. Compared with sham operated groups, Sympathetic nerve AID shortened significantly (sham operated groups vs model groups: 58.93±2.99μV·ms vs 46.98±1.65μV·ms, P<0.01,n=10) in model groups. Compared with sham operated groups, sympathetic nerve TCD prolonged and TCDI shorted in model groups. The density of TH, ChAT and GAP43 positive nerve fibers were significantly higher in model groups than sham operated groups in RAA. GAP43 positive nerve fibers were significantly higher in model groups than sham operated groups in PV.Conclusions:1) The AERP and MAP was shortened and the rate adapation of AERP was lost by rapid atrial pacing in rabbits. a) fAPD of RAA and PV were shortened with MEA technique. b) After superfusing calcium and potassium ion channel blockers, fAPD have different extent prolongation. This results suggest that Ito, IKur and IKl have remodeling and mediate RAP-induced atrial electrical remodeling. Amiodarone can effect on potassium ion channels (Ito, IKur, IKl and IKs), prevent fAPD shortening.The findings of study indicate a potential antiarrhythmic effect of vAmiodarone during AF, because it reduces electrical remodeling of the atria. c) MEA was a sensitive and stable and tissue action potential multiple-channel recording mapping system in animal heart slices. fAPD electrophysical properties was similar to action potential duration (APD) and AERP.2) After 24h RAP, myocardial hypertrophy, irregular array of cardiocyte, Central part of myocytes is free of sarcomeres and contains abundant glycogen (PAS-positivestaining) and myocardium interstitial fibrosis in RAA and PV, which is ready for the onset and maintenance of AF.3) RAP induced cardiac nerve degenerating, regenerating and remodeling. The finding suggested that autonomic nerve remodeling could maintain developing of atrial fibrillation.
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