Cardiac rhythms of late pre-pipped and pipped chick embryos exposed to altered oxygen environments

During the final stages of embryonic development in chickens, diffusive gas exchange through the chorioallantoic membrane (CAM) is progressively replaced by pulmonary respiration that begins with internal pipping (IP) of the CAM. Late chick embryos going through the transition from CAM respiration to pulmonary respiration were exposed to hyperoxic (100% O(2)) and hypoxic (10% O(2)/N(2)) environments for 2-h and the responses of baseline heart rate (HR), and HR fluctuation patterns were investigated. 16- and 18-day-old (referred to as 18-d) embryos and 20-d externally pipped (EP) embryos were examined as pre-pipped embryos and pipped embryos, respectively. 19-d embryos were divided into two groups: embryos that had not yet internally pipped (Pre-IP embryos) and embryos that had internally pipped (IP embryos). IP was identified by detecting the breathing signal with a condenser microphone attached hermetically on the eggshell (i.e. acoustorespirogram) on day 19 of incubation. In the hyperoxic environment, HR baseline of pre-pipped embryos remained unchanged and that of pipped embryos was depressed. In the hypoxic environment, HR baseline of 16-d pre-pipped embryos was depressed and that of pipped (IP and EP) embryos elevated. These different responses in pipped embryos might be partially attributed to increased cholinergic input from the vagus nerve in hyperoxia and increased adrenergic response in hypoxia. While hyperoxia did not induce marked modification of instantaneous heart rate (IHR) fluctuation patterns, hypoxia tended to augment transient decelerations of IHR in late pre-pipped embryos and markedly depressed HR fluctuations in pipped embryos.     

Pathogen-induced heart rate changes associated with cholinergic nervous system activation

The autonomic nervous system plays a central role in regulation of host defense and in physiological responses to sepsis, including changes in heart rate and heart rate variability. The cholinergic anti-inflammatory response, whereby infection triggers vagal efferent signals that dampen production of proinflammatory cytokines, would be predicted to result in increased vagal signaling to the heart and increased heart rate variability. In fact, decreased heart rate variability is widely described in humans with sepsis. Our studies elucidate this apparent paradox by showing that mice injected with pathogens demonstrate transient bradyarrhythmias of vagal origin in a background of decreased heart rate variability (HRV). Intraperitoneal injection of a large inoculum of Gram-positive or Gram-negative bacteria or Candida albicans rapidly induced bradyarrhythmias of sinus and AV nodal block, characteristic of cardiac vagal firing and dramatically increased short-term HRV. These pathogen-induced bradycardias were immediately terminated by atropine, an antagonist of muscarinic cholinergic receptors, demonstrating the role of vagal efferent signaling in this response. Vagal afferent signaling following pathogen injection was demonstrated by intense nuclear c-Fos activity in neurons of the vagal sensory ganglia and brain stem. Surprisingly, pathogen-induced bradycardia demonstrated rapid and prolonged desensitization and did not recur on repeat injection of the same organism 3 h or 3 days after the initial exposure. After recovery from the initial bradycardia, depressed heart rate variability developed in some mice and was correlated with elevated plasma cytokine levels and mortality. Our findings of decreased HRV and transient heart rate decelerations in infected mice are similar to heart rate changes described by our group in preterm neonates with sepsis. Pathogen sensing and signaling via the vagus nerve, and the desensitization of this response, may account for periods of both increased and decreased heart rate variability in sepsis.     

Increased vagal tone in adaptation to the effects of continuous stress and termination of cardiac arrhythmias

It was established in experiments on rats exposed to 5 day stress that 1 day stress resulted in a twofold decreased heart fibrillation threshold (HFT) and 5 day stress resulted in bradycardia and in the restoration of HFT to the control level. The restoration of the heart electric stability was due to an increased vagal tone because atropine eliminated the bradycardia and reduced HFT again. Adaptation to continuous 5 day stress increased 3-7-fold the heart resistance to ischemic and reperfusion arrhythmias. This protective effect was completely abolished with atropine. Thus adaptation to continuous mild stress has a potent antiarrhythmic effect which occurs due to the increased vagal tone.     

Control of blood pressure mediated by baroreflex changes of heart rate in the chicken embryo (Gallus gallus)

Pharmacological manipulation of peripheral resistance via sodium nitroprusside and phenylephrine was used to study baroreflex function over the latter two-thirds of incubation in embryonic chickens. From day 9 to day 19 of incubation, there is a positive linear relation between heart rate and blood pressure, indicating the feedforward action of arterial pressure on heart rate. A reciprocal relationship between blood pressure and heart rate became pronounced during the last 3 days of incubation. For the purpose of the study, gain of the baroreflex was calculated as maximal gain (only those embryos that demonstrated the response) or average gain (all embryos). Maximal gain increased progressively from 13 +/- 7 beats. min(-1). kPa(-1) at 18 days to 105 +/- 83 beats. min(-1). kPa(-1) in 2-day-old hatchlings. The percentage of embryos older than 18 days with baroreflex responses increased from 33% on day 19 to 56% on day 21, indicating that baroreflex regulation begins late in incubation ( approximately 90% incubation time), and the gain of this reflex exhibits a maturation over the final 3 days of incubation.     

Chronic atropine administration diminishes the contribution of vasoactive intestinal polypeptide to heart rate regulation

Vasoactive intestinal polypeptide (VIP) is implicated in the modulation of vagal effects on the heart rate. In this study, the impact of acute and chronic atropine administration on VIP levels in rat heart atria was investigated in relation to heart rate in the course of vagus nerves stimulation. Anaesthetised control and atropinised (10 mg/kg/day for 10 days) rats pretreated with metipranolol and phentolamine that were either given or not a single dose of atropine were subjected to bilateral vagus nerve stimulation (30 min: 0.7 mA, 20 Hz, 0.2 ms). VIP concentrations in the atria were determined after each stimulation protocol. In control rats with or without single atropine administration, the heart rate upon vagal stimulation was higher than in atropinised animals with or without single atropine dose, respectively. VIP concentrations in the control atria were significantly decreased after the stimulation; the decrease was comparable both in the absence and presence of a single dose of atropine. Compared to controls, VIP levels were significantly decreased after chronic atropine treatment and they were not further reduced by vagal stimulation and single atropine administration. Administration of VIP antagonist completely abolished the differences in the heart rate upon vagal stimulation between control and atropinised groups. In conclusion, the data indicate that chronic atropine administration affects VIP synthesis in rat heart atria and consequently it modifies the heart rate regulation.     

Effects of diazepam on the isolated chick embryo heart.

Diazepam decreased the rate and amplitude of contraction in isolated embryonic chick hearts in a dose-dependent manner in both the noninnervated hearts obtained from 4-day-old embryos and the innervated hearts from 7-day-old embryos. The concentration of diazepam necessary to reduce the heart rate and contractile amplitude to 50% of the control values was about 1 X 10(-4) M. Concentrations less than 1.0 X 10(-5) M had no detectable depressant effects. Prior administration of atropine did not alter the depression induced by diazepam. Norepinephrine was able to stimulate the amplitude of contraction in the diazepam-depressed heart while atropine was without effect. The vehicle used in the clinical injectable preparation of diazepam had no depressant effects. The mechanism of action of the diazepam-induced depression on the isolated embryonic chick heart may be a direct depression of the myocardium.     

Ontogeny of heart rate in embryonic and nestling crows (Corvus corone and Corvus macrorhynchos)

The developmental patterns of mean heart rate (MHR) and instantaneous heart rate (IHR) were investigated in embryos and chicks of altricial Corvuscorone and Corvus macrorhynchos. The MHR of embryos increased linearly with time from 250 beats · min⁻¹ at mid-incubation to 290 beats · min⁻¹ in hatchlings. MHR during the pipping period was maximal, but only marginally higher than in hatchlings. MHR was stable at about 290–300 beats · min⁻¹ during the 1st week after hatching. Spontaneous heart rate (HR) decelerations and accelerations were found in embryos and chicks, disturbing the baseline HR with increasing frequency during development. However, the IHR accelerations developed later and were less frequent than in precocial species. IHR and body temperature decreased during mild cold exposure (23–25 °C) and IHR accelerations were reduced in nestlings during the 1st week. We suggest that the development of parasympathetic control of HR in crows occurs at 60% of incubation, similar to precocial embryos, but sympathetic control may be delayed and suppressed in contrast to precocial embryos. Accepted: 3 March 1999     

Hypoxic alligator embryos: chronic hypoxia, catecholamine levels and autonomic responses of in ovo alligators

Hypoxia is a naturally occurring environmental challenge for embryonic reptiles, and this is the first study to investigate the impact of chronic hypoxia on the in ovo development of autonomic cardiovascular regulation and circulating catecholamine levels in a reptile. We measured heart rate (f(H)) and chorioallantoic arterial blood pressure (MAP) in normoxic ('N21') and hypoxic-incubated ('H10'; 10% O(2)) American alligator embryos (Alligator mississippiensis) at 70, 80 and 90% of development. Embryonic alligator responses to adrenergic blockade with propranolol and phentolamine were very similar to previously reported responses of embryonic chicken, and demonstrated that embryonic alligator has α and β-adrenergic tone over the final third of development. However, adrenergic tone originates entirely from circulating catecholamines and is not altered by chronic hypoxic incubation, as neither cholinergic blockade with atropine nor ganglionic blockade with hexamethonium altered baseline cardiovascular variables in N21 or H10 embryos. In addition, both atropine and hexamethonium injection did not alter the generally depressive effects of acute hypoxia - bradycardia and hypotension. However, H10 embryos showed significantly higher levels of noradrenaline and adrenaline at 70% of development, as well as higher noradrenaline at 80% of development, suggesting that circulating catecholamines reach maximal levels earlier in incubation for H10 embryos, compared to N21 embryos. Chronically elevated levels of catecholamines may alter the normal balance between α and β-adrenoreceptors in H10 alligator embryos, causing chronic bradycardia and hypotension of H10 embryos measured in normoxia.     

INCREASED NUMBERS OF ANNULATE LAMELLAE IN MYOCARDIUM OF CHICK EMBRYOS INCUBATED AT ABNORMAL TEMPERATURES

Annulate lamellae have been observed in the myocardium of 18-day-old chick embryos maintained at the normal temperature of 100°F and at 90°F during the last week of incubation. An increased number of annulate lamellae was observed in heart muscle of embryos incubated at 90°F. This is probably caused by a persistent production of these organelles, since annulate lamellae are present in greater frequency than in 11-day-old embryos incubated at 100°F. In the hypertrophic hearts of 18-day-old embryos incubated at 90°F, the annulate lamellae were associated with a net increase of protein content and an elevated concentration of myocardial glycogen. It is suggested that the increased number of annulate lamellae is a sequela of reduced environmental temperature during incubation.     

The modulatory effects of noradrenaline on vagal control of heart rate in the dogfish,Squalus acanthias

The possible interactions between inhibitory vagal control of the heart and circulating levels of catecholamines in dogfish (Squalus acanthias) were studied using an in situ preparation of the heart, which retained intact its innervation from centrally cut vagus nerves. The response to peripheral vagal stimulation typically consisted of an initial cardiac arrest, followed by an escape beat, leading to renewed beating at a mean heart rate lower than the prestimulation rate (partial recovery). Cessation of vagal stimulation led to a transient increase in heart rate, above the prestimulation rate. This whole response was completely abolished by 10(-4) M atropine (a muscarinic cholinergic antagonist). The degree of vagal inhibition was evaluated in terms of both the initial, maximal cardiac interval and the mean heart rate during partial recovery, both expressed as a percentage of the prestimulation heart rate. The mean prestimulation heart rate of this preparation (36+/-4 beats min(-1)) was not affected by noradrenaline but was significantly reduced by 10(-4) M nadolol (a beta-adrenergic receptor antagonist), suggesting the existence of a resting adrenergic tone arising from endogenous catecholamines. The degree of vagal inhibition of heart rate varied with the rate of stimulation and was increased by the presence of 10(-8) M noradrenaline (the normal in vivo level in routinely active fish), while 10(-7) M noradrenaline (the in vivo level measured in disturbed or deeply hypoxic fish) reduced the cardiac response to vagal stimulation. In the presence of 10(-7) M noradrenaline, 10(-4) M nadolol further reduced the vagal response, while 10(-4) M nadolol + 10(-4) M phentolamine had no effect, indicating a complex interaction between adrenoreceptors, possibly involving presynaptic modulation of vagal inhibition.     

Postnatal changes of the tonic influence of the vagus nerves on the heart rate, and of the activity of choline acetyltransferase in the heart atria of rats

Postnatal changes in the resting heart rate and in its parasympathetic tonic inhibition have been measured in awake rats and compared with changes in the activity of choline acetyltransferase (ChAT) in the heart atria. The heart rate at rest increased from 372.min-1 on the 1st to 456 and 442.min-1 on the 15th and 24th day of life and then again decreased to 358 and 356.min-1 in 60-day-old and adult rats. Until the 15th day of postnatal life, the administration of atropine did not bring about an increase in the heart rate; the cardio-acceleratory effect of atropine (indicating the presence of tonic vagal inhibition of the heart) appeared only on the 18th day and increased steeply up to the 40th day of postnatal life. The activity of ChAT in the heart atria was measured as the difference between the synthesis of acetylcholine in atrial homogenates incubated in the absence and in the presence of bromoacetylcholine (BrACh), a specific inhibitor of ChAT; this procedure eliminated the contribution of carnitine acetyltransferase to the synthesis of acetylcholine. The activity of ChAT was found to increase steeply from the 1st to the 25th days of postnatal life; the steepest increase in the activity of the enzyme occurred between the 4th and the 15th days. Temporal correlation between the changes in the activity of ChAT, in the content of acetylcholine in the heart atria (Kuntscherová and Vlk 1979) and in the efficiency of transmural stimulation of sinoatrial region on the heart rate (Vlk 1979) indicate that the functional maturation of intracardiac cholinergic neurones, proceeding in rats during the first three weeks of their postnatal life, plays an important role in the onset and temporal development of the tonic parasympathetic inhibition of the heart rate.     

Antiarrhythmic drugs and the modulation of autonomic control of heart rate in rabbits

A model of the components of autonomic control of heart rate was developed and used for the evaluation of quantitative contribution of sympathetic and vagal tone to cardiac function. In conscious rabbits, sequential inhibition of muscarinic and beta receptors was produced and the relative contributions of vagal and sympathetic tone were characterized. Based on the model, the magnitude of presynaptic interaction between the vagal and sympathetic nerve endings was evaluated. From data in the literature, similar analysis of the control of heart rate was performed for the rat, dog, and human subject and compared with that of the rabbit. The results show that the resting rabbit heart is under less vagal tone than sympathetic tone as compared with other species. The effects of acute administration of amiodarone on the sympathetic and parasympathetic control of heart rate as well as intrinsic heart rate were investigated. Amiodarone decreased the heart rate, which resulted from a direct effect on the sinoatrial (SA) node. In addition, it attenuated the vagal as well as the sympathetic effects on the SA node. The effect on vagal component was greater. Further, the effects of other antiarrhythmic drugs on the electrocardiographic PP and PR intervals were studied. The usefulness of this model for the analysis of the effects of antiarrhythmic drugs is presented.     

A trial of evaluating static and dynamic loads in man by means of analysis of the variability of instantaneous heart rate

The instantaneous heart rate changes were examined in 12 forest workers during following tests: increasing (up to refusal) dynamic load on ergometer, rest in erect position, static load (about 75% of body weight) in erect position. The ECG signal, carrying information on heart rate, was amplified and processed by means of an especially designed heart rate analyzer. The dynamics of heart rate changes was evaluated through the analysis of differences of subsequent instantaneous heart rates and of distributions of heart rate accelerations and decelerations during the tests. Significantly different distributions of instantaneous heart rate changes as well as of accelerations and decelerations were observed for different loads.     

Bombesin-induced stimulation of cardiac parasympathetic innervation

The central nervous system effects of bombesin on cardiovascular function were examined in conscious, freely-moving rats. Intracerebroventricular administration of bombesin elevated mean arterial pressure, reduced heart rate and inhibited cold-induced tachycardia. Adrenalectomy prevented bombesin-induced elevations of mean arterial pressure. In contrast, bombesin-induced bradycardia was neither adrenal-dependent nor a baroreceptor-mediated reflex response to increased arterial pressure. Systemic atropine methyl nitrate treatment attenuated bombesin-induced bradycardia, suggesting that bombesin acts within the central nervous system to stimulate cardiac vagal activity.     

Ontogeny of cholinergic and adrenergic cardiovascular regulation in the domestic chicken (Gallus gallus)

Adrenergic and cholinergic tone on the cardiovascular system of embryonic chickens was determined during days 12, 15, 19, 20, and 21 of development. Administration of the muscarinic antagonist atropine (1 mg/kg) resulted in no significant change in heart rate or arterial pressure at any developmental age. In addition, the general cardiovascular depressive effects of hypoxia were unaltered by pretreatment with atropine. In addition, the ganglionic blocking agent hexamethonium (25 mg/kg) did not induce changes in heart rate. The beta-adrenergic antagonist propranolol (3 mg/kg) induced a bradycardia of similar magnitude on all days studied, with a transient hypertensive action on days 19-20, indicating the existence of an important cardiac and vascular beta-adrenergic tone. Injections of the alpha-adrenergic antagonists prazosin or phentolamine (1 mg/kg) reduced arterial pressure significantly on all days of incubation studied. Collectively, the data indicate that embryonic chickens rely primarily on adrenergic control of cardiovascular function, with no contribution from the parasympathetic nervous system.     

Dynamical systems analysis of arterial blood pressure signals in relation to heart rate fluctuations in chick embryos

We attempted a new approach based on a modern dynamical system theory to reconstruct the arterial blood pressure signals in relation to heart rate fluctuations of developing chick embryos. The dynamical systems approach in general is to model a phenomenon that is presented by a single time series record and approximate the dynamical property (e.g. heart rate fluctuations) of a system based only on information contained in a single-variable (arterial blood pressure) of the system. The time-series data of the arterial blood pressure was reconstructed in 3-dimensional space to draw characteristic orbits. Since the reconstructed orbits of the blood pressure should retain information contained in the pressure signals, we attempted to derive instantaneous heart rate (IHR) from the reconstructed orbits. The derived IHR presenting HR fluctuations coincided well with the IHR obtained conventionally from the peak-to-peak time intervals of the maximum blood pressure. Movements of the reconstructed orbits of the arterial blood pressure in 3-dimensional space reflected HR fluctuations (i.e. transient decelerations and accelerations).     

Cholinergic and adrenergic tone on the heart of the Antarctic dragonfish, Gymnodraco acuticeps, living at sub-zero temperatures

Heart rate and ventral aortic blood pressures were recorded from the Antarctic dragonfish,Gymnodraco acuticeps, a member of the family Bathydraconidae. At −1.0 °C, the resting heart rate was 17.4 beats per minute and the ventral aortic pressure was 3.4 kPa. Cholinergic and adrenergic tone on the heart was determined by administration of the muscarine and ß-adrenoreceptor antagonists, atropine and sotalol, respectively. Neither antagonist influenced ventral aorta blood pressure; however, injection of atropine resulted in a significant increase in heart rate, and sotalol a decrease in heart rate. The cholinergic tone accounted for 30% of intrinsic heart rate and the adrenergic tone 26% of intrinsic heart rate. Comparison of these cardiac data with those for other teleosts from a wide range of thermal environments revealed no significant correlation for either cholinergic or adrenergic tone with body temperature (i.e. thermal independence); however, the resting and intrinsic heart rate of teleosts were strongly correlated with temperature.     

Chronic hypoxic incubation blunts thermally dependent cholinergic tone on the cardiovascular system in embryonic American alligator (Alligator mississippiensis)

Environmental conditions play a major role in shaping reptilian embryonic development, but studies addressing the impact of interactions between chronic and acute environmental stressors on embryonic systems are lacking. In the present study, we investigated thermal dependence of cholinergic and adrenergic cardiovascular tone in embryonic American alligators (Alligator mississippiensis) and assessed possible phenotypic plasticity in a chronic hypoxic incubation treatment. We compared changes in heart rate (f _H) and mean arterial blood pressure (P _M) for chronically hypoxic and normoxic-incubated embryos after cholinergic and adrenergic blockade following three different acute temperature treatments: (1) 30 °C (control incubation temperature), (2) acute, progressive decrease 30–24 °C then held at 24 °C, and (3) acute, progressive increase 30–36 °C then held at 36 °C. f _H progressively fell in response to decreasing temperature and rose in response to increasing temperature. P _M did not significantly change with decreasing temperature, but was lowered significantly with increasing acute temperature in the normoxic group at 90 % of development only. Propranolol administration (β adrenergic antagonist) produced a significant f _H decrease at 24, 30, and 36 °C that was similar at all temperatures for all groups. For normoxic-incubated embryos at 90 % of development, atropine administration (cholinergic antagonist) significantly increased f _H in both 24 and 36 °C treatments, but not in the 30 °C control treatment. This atropine response at 24 and 36 °C demonstrated acute thermally dependent cholinergic tone on f _H late in development for normoxic-incubated, but not chronically hypoxic-incubated embryos. Collectively, data indicated that cardiovascular control mechanisms in embryonic alligators may be activated by thermal extremes, and the maturation of control mechanisms was delayed by chronic hypoxia.     

Cardiovascular effects produced by choline injected into the lateral cerebral ventricle of the unanesthetized rat

Intracerebroventricular (icv) injection of choline (50–150 μg) causes a transient increase in blood pressure and a more prolonged decrease in heart rate (HR) in conscious rats. The bradycardia results from a centrally mediated increase in vagal tone. The cardiovascular effects do not appear to involve endogenous brain acetylcholine since there is no significant difference in the responses induced by choline before and after icv injection of hemicholinium-3. Intracerebroventricular ventricular injection of atropine or mecamylamine, alone, failed to influence the choline effect. However, atropine and mecamylamine, given together, abolished the reduction of HR, but still failed to modify the pressor response. The changes in blood pressure and HR appear to be due to effects of choline on post-synaptic receptors in different brain regions.     

Heart rate recovery after exercise: a predictor of ventricular fibrillation susceptibility after myocardial infarction

Heart rate recovery after exercise, thought to be related to cardiac parasympathetic tone, has been shown to be a prognostic tool for all-cause mortality. However, the relationship between this variable and confirmed susceptibility to ventricular fibrillation (VF) has not been established. Therefore, myocardial ischemia was induced with a 2-min occlusion of the left circumflex artery during the last minute of exercise in mongrel dogs with myocardial infarction (n = 105 dogs). VF was induced in 66 animals (susceptible), whereas the remaining 39 dogs had no arrhythmias (resistant). On a previous day, ECG was recorded and a time-series analysis of heart rate variability was measured 30, 60, and 120 s after submaximal exercise (treadmill running). The heart rate recovery was significantly greater in resistant dogs than in susceptible dogs at all three times, with the most dramatic difference at the 30-s mark (change from maximum: 48.1 +/- 3.6 beats/min, resistant dogs; 31.0 +/- 2.2 beats/min, susceptible dogs). Correspondingly, indexes of parasympathetic tone increased to a significantly greater extent in resistant dogs at 30 and 60 s after exercise. These differences were eliminated by atropine pretreatment. When considered together, these data suggest that resistant animals exhibit a more rapid recovery of vagal activity after exercise than those susceptible to VF. As such, postexercise heart rate recovery may help identify patients with a high risk for VF following myocardial infarction.