BASIC SCIENCE SECTION Autonomic nervous system regulation of baseline heart rate in the fetal lamb Akihiko Wakatsuki, MD, Yuji Murata, MD, Yuko Ninomiya, MD, Naoki Masaoka, MD, James G. Tyner, MD, and Krishna K. Kutty Orange, California OBJECTIVE: We examined 29 chronically instrumented fetal lambs from 125 to 143 days' gestation to investigate the effects of fetal behavioral states and autonomic nervous system maturation on baseline fetal heart rate. STUDY DESIGN: Behavioral states were defined from electrocorticographic analysis as low-voltage fast activity or high-voltage slow activity. Decrease and increase in baseline fetal heart rate subsequent to administration of propranolol and methylatropine represented l3-sympathetic and parasympathetic activity. RESULTS: Baseline fetal heart rate decreased with gestation in both states, with steeper regression in low-voltage fast activity (p < 0.001). Positive correlation was noted between gestational age and percent decrease baseline fetal heart rate in both states with steeper regression in high-voltage slow activity (p < 0.001), and between gestational age and percent increase baseline fetal heart rate with steeper regression in low-voltage fast activity (p < 0.001). Fetal heart rate l3-sympathetic and parasympathetic tones increased with age in both states, with elevation of l3-sympathetic tone in high-voltage slow activity and parasympathetic tone in low-voltage fast activity. CONCLUSION: Sympathetic and parasympathetic systems influence baseline fetal heart rate in these behavioral states and with age. (AM J OBSTET GYNECOL 1992;167:519-23.) Key words: Baseline fetal heart rate, tone, parasympathetic tone, fetal lamb, behavioral state, gestational age Baseline fetal heart rate (FHR) declines with gesta- tional age in the human fetus':" and fetal lamb," Schif- ferli and Caldeyro-Barcia' attribute this decline to ma- turing vagal parasympathetic functional control. Others report that baseline FHR is consistently higher in the high-voltage slow activity behavioral state than in the low-voltage fast activity state."? Using a-sympa- thetic and blockades, Zhu and Szeto" concluded that the changes in baseline FHR relative to behavioral states were determined uniquely by sym- pathetic activity. B The regulation mechanisms of these fetal cardiovascular changes, especially the interaction of fetal behavioral states and autonomic nervous mat- uration on baseline FHR, remains unidentified. Our study investigated these control mechanisms at differ- ent gestational ages and behavioral states. From the Division of Maternal-Fetal Medicine, Department of Ob- stetrics and Gynecology, University of California, Irvine. Received for publication July 9, 1991; revised March 9, 1992; ac- cepted March 18, 1992. Reprint requests: Yuji Murata, MD, Department of Obstetrics and Gynecology, University of California, Irvine, UCI Medical Center, P.O. Box 14091, Orange, CA 92613-1491. 6/1 /37965 Material and methods Surgical preparation. Twenty-nine chronically in- strumented fetal lambs between 125 and 143 days' ges- tation from time-mated, cross-bred Columbia-Ram- bouillet ewes were used in this study. Sheep gestation averages 145 days. All animals were maintained in a facility approved by the American Association for the Accreditation of Laboratory Animal Care and in ac- cordance with Guide for the Care and Use of Laboratory Animals and met the standards of care of the U.S. De- partment of Agriculture. Before implementation re- search protocols were approved by the Animal Re- search Committee at the University of California, Irvine. After 24 hours of food and water withdrawal anes- thesia was induced with xylazine (Rompun) 12 to 14 mg administered intramuscularly. Lidocaine 1% was used locally as needed. The uterus was exposed with a midline abdominal incision. The fetal head was deliv- ered through a uterine myometrial incision and cov- ered with a surgical glove filled with warm saline so- lution. Bilateral electrodes were sutured under the skin in the fetal chest wall for electrocardiography. Paired stainless steel electrodes (ethylcyanoacrilate, Loctite 519
Transcript
BASIC SCIENCE SECTION
Autonomic nervous system regulation of baseline heart rate inthe fetal lamb
Akihiko Wakatsuki, MD, Yuji Murata, MD, Yuko Ninomiya, MD, Naoki Masaoka, MD,James G. Tyner, MD, and Krishna K. Kutty
Orange, California
OBJECTIVE: We examined 29 chronically instrumented fetal lambs from 125 to 143 days' gestation to
investigate the effects of fetal behavioral states and autonomic nervous system maturation on baselinefetal heart rate.STUDY DESIGN: Behavioral states were defined from electrocorticographic analysis as low-voltage fast
activity or high-voltage slow activity. Decrease and increase in baseline fetal heart rate subsequent to
administration of propranolol and methylatropine represented l3-sympathetic and parasympathetic activity.RESULTS: Baseline fetal heart rate decreased with gestation in both states, with steeper regression in
low-voltage fast activity (p < 0.001). Positive correlation was noted between gestational age and percentdecrease baseline fetal heart rate in both states with steeper regression in high-voltage slow activity
(p < 0.001), and between gestational age and percent increase baseline fetal heart rate with steeperregression in low-voltage fast activity (p < 0.001). Fetal heart rate l3-sympathetic and parasympathetic
tones increased with age in both states, with elevation of l3-sympathetic tone in high-voltage slow activityand parasympathetic tone in low-voltage fast activity.
CONCLUSION: Sympathetic and parasympathetic systems influence baseline fetal heart rate in thesebehavioral states and with age. (AMJ OBSTET GYNECOL 1992;167:519-23.)
Baseline fetal heart rate (FHR) declines with gestational age in the human fetus':" and fetal lamb," Schifferli and Caldeyro-Barcia' attribute this decline to maturing vagal parasympathetic functional control.Others report that baseline FHR is consistently higherin the high-voltage slow activity behavioral state thanin the low-voltage fast activity state."? Using a-sympathetic and ~-sympathetic blockades, Zhu and Szeto"concluded that the changes in baseline FHR relative tobehavioral states were determined uniquely by sympathetic activity.B The regulation mechanisms of thesefetal cardiovascular changes, especially the interactionof fetal behavioral states and autonomic nervous maturation on baseline FHR, remains unidentified. Ourstudy investigated these control mechanisms at different gestational ages and behavioral states.
From the Division of Maternal-Fetal Medicine, Department of Obstetrics and Gynecology, University of California, Irvine.Received for publication July 9, 1991; revised March 9, 1992; accepted March 18, 1992.Reprint requests: Yuji Murata, MD, Department of Obstetrics andGynecology, University of California, Irvine, UCI Medical Center,P.O. Box 14091, Orange, CA 92613-1491.6/1 /37965
Material and methods
Surgical preparation. Twenty-nine chronically instrumented fetal lambs between 125 and 143 days' gestation from time-mated, cross-bred Columbia-Rambouillet ewes were used in this study. Sheep gestationaverages 145 days. All animals were maintained in afacility approved by the American Association for theAccreditation of Laboratory Animal Care and in accordance with Guide for the Care and Use of LaboratoryAnimals and met the standards of care of the U.S. Department of Agriculture. Before implementation research protocols were approved by the Animal Research Committee at the University of California,Irvine.
After 24 hours of food and water withdrawal anesthesia was induced with xylazine (Rompun) 12 to 14mg administered intramuscularly. Lidocaine 1% wasused locally as needed. The uterus was exposed with amidline abdominal incision. The fetal head was delivered through a uterine myometrial incision and covered with a surgical glove filled with warm saline solution. Bilateral electrodes were sutured under the skinin the fetal chest wall for electrocardiography. Pairedstainless steel electrodes (ethylcyanoacrilate, Loctite
519
520 Wakatsuki at al.
Corp., Newington, Conn.) were placed and cementedbilaterally in the fetal parietal skull for e1ectrocorticography. The instantaneous FHR was obtained from theR-R interval of the electrocardiogram. Polyvinyl catheters were inserted into the fetal carotid artery,jugularvein, and trachea. The procedures were performedcarefully to avoid disturbing the fetal vagal nerve. Anopen-end catheter was retained in the amniotic cavity,and additional catheters were placed in the maternalfemoral artery and vein. All electrodes and catheterswere exteriorized to the flank of the ewe.
Experimental protocol. The animals were maintained in an air-conditioned room and given free accessto food and water. For 3 to 4 days after surgery themother and fetus received ampicillin (200 mg/kg/dayintravenously) and gentamicin (6 mg /kg/day intravenously), and ampicillin (1 gm/day) was administered tothe amniotic cavity. Maternal and fetal arterial pH,blood gases, and hematocrits were checked daily. ForpH and gas determinations approximately 0.6 ml arterial blood was collected anaerobically and measured(Corning pH/Blood Gas Analyzer, Model 168 BGA,Ciba Corning Diagnostic Corp., Medfield, Mass.) at37° C. Fetal well-being was determined by the followingcriteria: pH > 7.30. Po. > 16 mm Hg, hematocrit >30%.Fetal arterial blood pressure, FHR, electrocorticogram,tracheal pressure, intrauterine pressure, and maternalblood pressure were continuously monitored on an 8channel rectilinear recorder (Beckman Model R612,Sensor Medics Corp. , Anaheim, Calif.). Animals thatdemonstrated interpretable electrocorticogram wereselected for this study. Fetal behavioral states were analyzed from the electrocorticogram for low-voltage fastactivity and high-voltage slow activity. Experimentswere performed after allowing ~5 days of postoperative recovery. All experiments were carried out between9 AM and 5 PM with the ewe standing or lyingquietly.
In 10 fetal lambs between 125 and 143 da ys' gestationbaseline FHR was assessed under normal physiologicconditions in the low-voltage fast activity and high-voltage slow activity behavioral states. Baseline FHR wasmeasured after FHR was stable for at least 10 minutesin which uterine contractions were absent. A set of observations during adjacent low-voltage fast activity andhigh-voltage slow activity was obtained every day afterfetal well-being was confirmed. The average durationof observation per fetus was 6.3 da ys and the averagenumber of observations per animal was 12.6.
In 10 fetal lambs between 126 and 143 days ' gestationsaline solution was infused via the fetal jugular vein at4.0 ml per hour for a 2-hour control period. The fetusthen received an intravenous priming dose of propranolol (1.0 mg, 1 ml) followed by propranolol infusion (2.0 mg/hr, 4.0 ml/hr) for 2 hours to block ~
sympathetic control of the heart rate. Baseline FHR
August 1992Am J Obstet Gynecol
was obtained in low-voltage fast activity and high-voltage slow activity states during control and propranololperiods. The percent decrease in baseline FHR betweencontrol and propranolol periods was calculated as anindex of FHR ~-sympathetictone. Completeness of the~-blockadewas indicated by the absence of cardiac acceleration response to an injected ~-agonist (isoproterenol 2.0 IJ.g/kg estimated fetal weight) . The experimentwas repeated on the same fetuses in 2 to 5 da ys, confirming complete recovery from the blockade. The average number of experiments per fetus was 4.5.
In nine fetal lambs between 127 and 143 days' gestation a 2-hour control period was followed by methylatropine (200 IJ.g/kg estimated fetal weight intravenously) for blockage of the parasympathetic control ofthe heart rate. The methylatropine dose was determined in preliminary experiments. Baseline FHR wasmeasured in the low-voltage fast activit y and high-voltage slow activity states during the control period andsubsequent to methylatropine administration. Thepercent increase of baseline FHR from control tomethylatropine periods was taken as representing FHRparasympathetic tone. Completeness of methylatropineblockade was evidenced by the absence of decelerationresponse to norepinephrine (4 IJ.g/kg estimated fetalweight). The effect of the blocking dose in our studylasted for ~3 hours. The experiment was repeated onthe same fetuses in 2 to 5 days, confirming completerecovery from the blockade. The average number ofexperiments per fetus was 4.5 .
Data analysis. Regression lines were determined bythe least squares method. The regression slopes werecompared by t test regression analysis .to Statistical significance was accepted at p < 0.05.
Results
Figure I shows the normal physiologic relationshipof fetal gestational age and baseline FHR in low-voltagefast activity (LVFA) and high-voltage slow activity(HVSA). Baseline FHR in both behavioral states decreased with gestational age . The regression equationsfor baseline FHR (BFHR) versus gestational age (GA)are as follows:
The low-voltage fast activity slope (- 4.4 7) was significantly less (p < 0.001) than that of the high-voltageslow activity slope (- 2.44).
Figure 2 shows the influence of gestational age onbaseline FHR response to [3-sympathetic blockade withpropranolol infusion in the low-voltage fast activity andhigh-voltage slow activity states. A positive correlationwas found between gestational age and percent de-
The high-voltage slow activity slope (0.97) was significantly steeper than the low-voltage fast activity (0.54)(P < 0.001).
Figure 3 shows the relationship of gestational age andFHR response to parasympathetic blockade with methylatropine. The percent increase (% inc) in baselineFHR from control to methylatropine periods duringlow-voltage fast activity and high-voltage slow activityis illustrated. In both behavioral states the responseincreased with gestational age. The regression equations follow:
The low-voltage fast activity slope (1.94) was significantly steeper (p < 0.001) than the high-voltage slowactivity (1.01).
CommentBaseline FHR is regulated by the combined efforts
of both intrinsic and extrinsic factors. The extrinsic
factors are primarily the sympathetic and parasympathetic limbs of the autonomic nervous system.
The effects of the autonomic nervous system on baseline FHR in different behavioral states and gestationalages were assessed. Most previous studies used atropinesulfate to evaluate parasympathetic tone on FHR I I
-I" ;
however, atropine sulfate has been reported to altersleep behavior. Animals that received atropine sulfatedid not show desynchronized sleep for several hours,and the desynchronized sleep that ultimately appearedwas markedly modified. 14 For our study methylatropinewas used as a parasympathetic blockade. This substanceis much less lipid soluble and does not appear to actcentrally. 15, 16 According to Guazzi et aI.,16 methylatropine (1 mg/kg) could be administered intravenouslywithout any apparent sleep disturbance. 17
In our study methylatropine 200 I-lg/kg estimatedfetal weight was determined by prior experiments tobe the appropriate dose. Complete parasympatheticblock was obtained for :::0:3 hours with no effect uponlow-voltage fast activity- and high-voltage slow activity-cycling behavior. For the evaluation of autonomicnervous system effects on FHR, the sympathetic andparasympathetic blocking agents were not studied together. Twenty-four hours were allowed between tests
for recovery of homeostasis. The percent decrease inbaseline FHR subsequent to propranolol infusion andthe percent increase in baseline FHR subsequent tomethylatropine injection were calculated as representing FHR l3-sympathetic and parasympathetic tone, re-
522 Wakatsuki et al. August 1992Am J Obstet Gynecol
Fig. 2. Relationship between percent decrease in baseline FHR (BFHR) and in low-voltagefast activity(LVFA) and high-voltage slow activity (HVSA) states after propranolol.
Fig. 3. Relationship between percent increase in baseline FHR (BFHR) and gestational age in lowvoltage fast activity (LVFA) and high-voltage slow activity (HVSA) states after atropine.
spectively.":" The percent decrease in baseline FHR
increased with gestational age, and the slope of theregression line was steeper in the high-voltage slow ac
tivity state. These data indicate that FHR l3-sympathetictone increases with gestational age and is greater in thehigh-voltage slow activity state. The difference be-
tween low-voltage fast activity and high-voltage slowactivity sympathetic tone also increased with gestationalage.
The percent increase in baseline FHR rose with gestational age, and the regression line slope was steeperduring the low-voltage fast activity state. These results
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indicate that FHR parasympathetic tone increases withgestational age. FHR parasympathetic tone was higherin low-voltage fast activity, and the difference betweenstates increased with gestational age.
A number of studies have reported conflicting resultsabout the autonomic system influence on FHR. Theexistence of adrenergic cardiovascular function in thefetal lamb as early as 85 days' gestation was demonstrated by bradycardia response to propranolol. 13 Parasympathetic cardiac function in the fetal lamb at 60days' gestation was shown by vagally driven heart rateslowing." Nuwayhid et al." reported that parasympathetic tone on resting heart rate in the fetal lamb wasfeeble at <130 days and sympathetic tone was constantthroughout gestation. Walker et al." found a progressiveincrease in parasympathetic tone and decrease in sympathetic tone toward term in the fetal lamb.' Vapaavouri et al." reported that parasympathetic tone isfully developed by 120 days' gestation in the fetal lamband that sympathetic tone increased from 101 days on.Other studies have reported that sympathetic influenceupon FHR is significant" or minor." In our study gestational age correlated positively with increased sympathetic and parasympathetic tone from 85% of gestational period to term. These results demonstrate thatautonomic effects on FHR develop markedly duringthe last 15% of gestation in the fetal lamb. Assali et al."concluded that sympathetic tone becomes active earlierthan parasympathetic tone, and Walker et al." showeda decrease of sympathetic influence toward term thatresulted from a strong increase in parasympathetic activity. They did not, however, consider differences associated with behavioral states. In our study FHR sympathetic tone was more notable during high-voltageslow activity and parasympathetic tone more pronounced during low-voltage fast activity. Thus each behavioral state has unique autonomic effects upon FHR.
It would appear that FHR in the absence of autonomic effector action declines progressively with gestational age and that this may be a function of increasedcardiac muscle mass and cardiac work. In our study weconclude that with behavioral state changes shifts inrelative magnitudes of the simultaneous parasympathetic inhibitory and sympathetic acceleratory influences modulate the basal or intrinsic heart rate. Duringthe low-voltage fast activity state increases in parasympathetic activity and decreases in sympathetic activityresult in lower baseline FHR. On the other hand, dur
ing high-voltage slow activity state sympathetic activityincreases and parasympathetic activity decreases so thatbaseline FHR rises.
We thank Lanie M. Adamson, MS, for editorial assistance in the preparation of the manuscript.
Baseline heart rate in fetal lamb 523
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