Study About Modulation Of Intrinsic Cardiac Nervous System On Heart Rate Variability And Its Mechanism In The Toads

IntroductionHeart rate variability (HRV) is an accepted term to describe the oscillation in the R-R interval between consecutive heart beats. The physiological foundation of HRV is the regulation of autonomic nervous system to the heart rate, which reflects the balance of the interaction between the neurohumor and the sinus node. HRV is affected by many peripheral and central factors in vivo, so it is complex to analyze. However, hardly anything is known about HRV of intrinsic sinus node activity under non-physiological conditions.It has been proposed that the intrinsic cardiac nervous system plays an active role in regulating cardiac function, but the impact of the intrinsic cardiac nervous system on HRV remains to be elucidated. The peripheral and central mechanism of HRV is not clear at the moment. Studies related to the intrinsic cardiac nervous system were few, especially about the effect of this system on HRV. The present study was designed as a basis for investigations into this problem in isolated perfused toad hearts. Electrocardiogram (ECG) was used to observe the conscious, resting toads, and electrocardiosignal was measured in isolated perfused toad hearts treated with atropine (an M-receptor agonist, Atr), phentolamine (an a-receptor agonist, Phe) and propranolol (αβ-receptor agonist, Pro). The above observation is aim at investigating the modulation of intrinsic cardiac nervous system on HRV in the toads, and exploring the possible mechanism involved in the effect. Materials and methods1. Animals and disposalMale healthy Zhoushan toads (35-50g) were held for up to a week at (24±2)℃and under a 12h light: 12h dark regime. The animals were raised by the manpower and divided into groups randomly. Three days before the experiment, each toad was placed individually in a plastic cage without restraint l-2h to adapt the experimental condition. All experimental trials were carried out in 8-12am.2. Experiment groups and protocol(1) In vivo group (n=10): ECG was collected in the conscious, resting toad.(2) In vitro group (n=10): isolated heart was purfused with Ringer's solution for 180min.(3) Atr group(n=10): isolated heart was purfused with Ringer's solution for 60min, then purfused with Atr(l.5μmol/L).(4) Phe+Pro group(n=11): isolated heart was purfused with Ringer's solution for 60min, then purfused with Phe(1μmol/L) and Pro(1μmol/L).(5) Atr+Phe+Pro group(n=10): isolated heart was purfused with Ringer's solution for 60min, then purfused with Atr(1.5μmol/L), Phe(1μmol/L) and Pro(1μmol/L).3. Electrocardiogram recordingPut the toad which was in the condition of conscious and resting into a plastic cage with electrodes, then collected the ECG of toad in vivo. Put the isolated heart into a small beaker with electrodes, and the electrocardiosignal was recorded.4. Observation on the histology of the intrinsic cardiac neuronsFour isolated toad hearts were perfused with Ringer's solution. Take a piece of 0.2cm×0.3cm tissue from venous sinus and posterior atrium respectively, put them into 2.5% Glutaral solution quickly, then sent them to the Electron Microscope Room of China Medical University. 5. Heart rate variability analysisSelect 10min continuous ECG or electro-cardio-signal records, analyzed all data with HRV software.6. Statistical analysisExperimental data were indicated as mean±SD, the differences between groups were assessed by student's test with SPSS software.Results1. As a result of histology, the intrinsic cardiac ganglion distributed under the epicardium or in the myocardium after toluidine blue (TB) dyeing.2. Compared with the heart in vivo, the isolated heart showed an obvious decrease in HRV.3. After medication with Atr, the change of high frequencies (HF) was not observed while low frequencies (LF) increased clearly; after medication with Phe+Pro, HF increased significantly while LF decreased; after medication with Atr+Phe+Pro, HF decreased while LF increased significantly.DiscussionCompared with the heart in vivo, all the time domain variables of HRV decreased obviously while HR increased, that is to say, HRV decreased after cardiac denervation. If the sympathetic and parasympathetic nerve controls the heart in the same degree in vivo and in vitro, HR should not change evidently. In our study, the increased HR indicate that the parasympathetic nerve system was predominant in vivo or the intracardiac sympathetic nerve enhanced its modulation in vitro.After treatment of Atr, the isolated heart was only controlled by the intracardiac sympathetic nerve. The changes of HR and HF were not observed while LF increased clearly. This indicates that the intracardiac parasympathetic nerve counterbalances a part of effect of the intracardiac sympathetic nerve before treatment. After treatment of Phe+Pro, the isolated heart was only controlled by the intracardiac parasympathetic nerve. HF increased significantly while HR and LF decreased. Compared with Atr group, the intracardiac sympathetic nerve seems to be predominant in the isolated heart before treatment. After treatment of Atr+Phe+Pro, the isolated heart lost the control of the intrinsic cardiac nervous system. There was no significant change in HR but HF decreased and LF increased significantly. This is the intrinsic heart rate and HRV of the heart. Compared Atr group to Atr+Phe+Pro group, we can see that after block of the intracardiac sympathetic nerve, the modulation of the intracardiac parasympathetic nerve was just appear. Above all, the experimental result suggests that the intracardiac sympathetic nerve seems to be predominant in the isolated heart before drug treatment.ConclusionIntrinsic cardiac nervous system may play an important role in the modulation on HR and HRV in the isolated toad heart, and the intracardiac sympathetic nerve seems to be predominant.