| Background, objective and method—Autonomic nerve system including the sympathetic andparasympathetic (vagal) nerves plays an important role in the electrophysiology and pumpingfunction of mammalian heart. The disorder of autonomic innervation and function, especiallythe sympathetic nerve, may lead to arrhythmia. Therefore, the study of sympathetic efferentimpulse and transmitter release and the underlying mechanisms were essential for cardiology.Since the limit of technology, previous study always focused on the morphology ofsympathetic nerve instead of the transmitter release. In this study, we used the amperometryfor the first time to record NE release of cardiac sympathetic nerve in real time andinvestigated the mechanism of NE release both in neonatal rat heart slice and in adult rabbitin-situ heart.Results1. The reliability of the amperometry technique in recording the neurotransmitterrelease of cardiac sympathetic nerveIn rat ventricular slice, puffing high potassium (80mM) induced a bulk oxidation current of theneurotransmitter. The current decreased when the slice was incubated with reserpine for 15minutes. Puffingα2 adrenergic inhibitor yohimbine (2μM) without high potassium induced thesame bulk current as the KCl-induced current. The current induced by high potassium (80mM)and yohimbine (2μM) was significantly ( 8.75±3.3pA vs 17.29±8.50 pA) higher than highKCl-induced current. This phenomenon is consistant with the theory that presynapticα2adrenoceptor has a negative feedback on sympathetic neurotransmitter release. Herein, wepresume the bulk current is mainly due to the oxidation of NE, without excluding other traceoxidizable substances at this condition. In rabbit heart in vivo, asphyxia and hypoxia bynitrogen inhalation elicited the bulk current of NE release from cardiac sympathetic nerveterminal in left ventricle. Field electric stimulation of left stellate ganglion (LSG) induced thesame bulk current in left ventricle. We convinced that the amperometry technique is reliable indetecting sympathetic NE release in a real time fassion. 2. The gold fiber electrode can simultaneously record sympathetic neurotransmiterrelease and electrocardiographyIn the study of in situ heart, we used the polypropylene coated gold fiber electrode. Thiselectrode greatly improved the fragile problem of carbon fiber electrode. We found that thegold fiber electrode could record both the oxidation current signal and the cardiac electrogram.The data showed that the positive chronotropic effect of sympathetic nerve is lower than thenegative chronotropic effect of vagus nerve. We stimulated the left vagus nerve with a20Hz, 10mA pusle for 10 seconds, the heart rate fell to 115 beat/min from 263.5 beat/min.When a 20Hz, 30mA pusle for 10 seconds was given to the left stellate ganglion, the heart rateseemed unchanged (265.5 beat/min vs 263.5 beat/min), although the oxidation current of NErelease can be detected. The negative chronotropic effect of vagus nerves (left and right) isfrequency (5Hz-30Hz)-dependent. When the stimulation frequency was increased, thenegative chronotropic effect of vagus nerve became more obvious. Under low frequency(5～20Hz), the left and right vagus nerves show no asymmetry in the negative chronotropiceffect. At a higher frequency (30Hz), however, the negative chronotropic effect of right vagusnerve is significantly higher than the left vagus nerve (232 beat/min vs 183 beat/min, p＜0.05).3. The regulation of vagal nerve, glutamate and calcium on sympatheticneurotransmitter releasePre-stimulation of SG yielded a downward bulk current in the in-vivo left ventricle in responseto the stimulation of cervical vagus nerve, indicating that an increase in vagal nerve activitycan reduce NE release at a higher sympathetic tone. We also found that stimulating thecervical vagus nerve alone (without stimulating SG) induced an upward current, suggestingthat an increase in vagal nerve activity can enhance NE release at a lower sympathetic tone. Inthe heart slice study, we found that Mch and glutamate also elicited sympathetic NE release.In the presense of calcium (1mM), high potassium (80mM) induced a significantly higher bulkcurrent compared with the absence of calcium, indiating that calcium may play an partial rolein regulating NE release. Conclusion1. In this study, we demonstrate a novel technique, i.e., amperometry, in detecting cardiacsympathetic neurotransmitter release both in heart slice and in in-situ heart. This techniqueyields a reliable, stable and real time monitoring of NE release, with high time and spatialresolution.2. Extracellular high potassium, asphyxia, hypoxia and SG stimulation can elicit theneurotranmitter release of cardiac sympathetic nerve.3. Using amperometry in the heart, we again demonstrated the well-known theory thatpresynapticα2 adrenoceptor has a negative feedback on sympathetic neurotransmitter release.4. The modualtion of the vagus nerve system on cardiac sympathetic neurotranmitter releasewas bidirectional: under a higher sympathetic tone, stimulating vagus nerve can downregulatecardiac sympathetic neurotranmitter release; while under lower sympathetic tone, stimulatingvagus nerve can upregulate cardiac sympathetic neurotransmitter release. In heart slice,however, M receptor activation by Mch (from low to high concentrtion) all facilitated thecardiac sympathetic neurotransmitter release, suggesting that the interaction betweensympathetic and parasympathetic nerves at central or peripheral level may play an importantrole in the controlling of cardiac sympathetic neurotranmitter release.5. Glutamate likely modulates the sympathetic neurotranmitter release.6. Calcium plays a role in the regulation of NE release.7. The positive chronotropic effect of sympathetic nerve is lower than the negativechronotropic effect of vagus nerve.8. The negative chronotropic effect of vagus nerve is frequency (5Hz-30Hz)-dependent,Stimulation with 40Hz or over leads to cardiac arrest. Along with the increment of frequency,the negative chronotropic effect of vagal nerve becomes more obvious, both left and rightvagal nerves show this frequency-dependent manner.9. At a higher stimulation frequency (30Hz), the negative chronotropic effect of right vagusnerve is significantly higher than the left vagus nerve, indicating the functional asymmetry ofleft and right vagus nerves. |
PDF Full Text Request