Marianne Garland

Prenatal nicotine-exposure alters fetal autonomic activity and medullary neurotransmitter receptors: implications for sudden infant death syndrome

During pregnancy, exposure to nicotine and other compounds in cigarette smoke increases the risk of the sudden infant death syndrome (SIDS) two- to fivefold. Serotonergic (5-HT) abnormalities are found, in infants who die of SIDS, in regions of the medulla oblongata known to modulate cardiorespiratory function. Using a baboon model, we tested the hypothesis that prenatal exposure to nicotine alters 5-HT receptor and/or transporter binding in the fetal medullary 5-HT system in association with cardiorespiratory dysfunction. At 87 (mean) days gestation (dg), mothers were continuously infused with saline (n = 5) or nicotine (n = 5) at 0.5 mg/h. Fetuses were surgically instrumented at 129 dg for cardiorespiratory monitoring. Cesarean section delivery and retrieval of fetal medulla were performed at 161 (mean) dg for autoradiographic analyses of nicotinic and 5-HT receptor and transporter binding. In nicotine-exposed fetuses, high-frequency heart rate variability was increased 55%, possibly reflecting increases in the parasympathetic control of heart rate. This effect was more pronounced with greater levels of fetal breathing and age. These changes in heart rate variability were associated with increased 5-HT(1A) receptor binding in the raphé obscurus (P = 0.04) and increased nicotinic receptor binding in the raphé obscurus and vagal complex (P < 0.05) in the nicotine-exposed animals compared with controls (n = 6). The shift in autonomic balance in the fetal primate toward parasympathetic predominance with chronic exposure to nicotine may be related, in part, to abnormal 5-HT-nicotine alterations in the raphé obscurus. Thus increased risk for SIDS due to maternal smoking may be partly related to the effects of nicotine on 5-HT and/or nicotinic receptors.

Family Nurture Intervention in the Neonatal Intensive Care Unit improves social-relatedness, attention, and neurodevelopment of preterm infants at 18 months in a randomized controlled trial

BACKGROUND Preterm infants are at high risk for adverse neurodevelopmental and behavioral outcomes. Family Nurture Intervention (FNI) in the Neonatal Intensive Care Unit (NICU) is designed to counteract adverse effects of separation of mothers and their preterm infants. Here, we evaluate effects of FNI on neurobehavioral outcomes. METHODS Data were collected at 18 months corrected age from preterm infants. Infants were assigned at birth to FNI or standard care (SC). Bayley Scales of Infant Development III (Bayley-III) were assessed for 76 infants (SC, n = 31; FNI, n = 45); the Child Behavior Checklist (CBCL) for 57 infants (SC, n = 31; FNI, n = 26); and the Modified Checklist for Autism in Toddlers (M-CHAT) was obtained for 59 infants (SC, n = 33; FNI, n = 26). RESULTS Family Nurture Intervention significantly improved Bayley-III cognitive (p = .039) and language (p = .008) scores for infants whose scores were greater than 85. FNI infants had fewer attention problems on the CBCL (p < .02). FNI improved total M-CHAT scores (p < .02). Seventy-six percent of SC infants failed at least one of the M-CHAT items, compared to 27% of FNI infants (p < .001). In addition, 36% of SC infants versus 0% of FNI infants failed at least one social-relatedness M-CHAT item (p < .001). CONCLUSIONS Family Nurture Intervention is the first NICU intervention to show significant improvements in preterm infants across multiple domains of neurodevelopment, social-relatedness, and attention problems. These gains suggest that an intervention that facilitates emotional interactions between mothers and infants in the NICU may be key to altering developmental trajectories of preterm infants.

Placental transfer and fetal metabolism of zidovudine in the baboon

Zidovudine (azidothymidine, AZT) is used in pregnancy to reduce mother to infant transmission of HIV. Understanding the disposition of AZT in the fetus is necessary to optimize therapeutic regimens directed toward the fetus. Recent studies in primates found similar steady-state levels of the glucuronide metabolite of AZT (AZT-glu) in the fetus to those in the mother, raising the question of whether metabolite was of fetal or maternal origin. The objective of this study was to determine whether glucuronidation occurred in the fetal compartment and to quantify the placental and fetal clearances of AZT using the two-compartment model at steady state. Steady-state concentrations were obtained after paired maternal and fetal infusions of AZT in chronically catheterized pregnant baboons. During maternal infusion, the mean (±SE) fetal to maternal ratio of AZT was <1 (0.84 ± 0.06, p < 0.02), suggesting clearance of AZT in the fetus. Mean total maternal clearance of AZT was 725 ± 49 mL/min and placental clearance was 36 ± 4 mL/min, or ∼5% of maternal clearance. Fetal clearance of AZT was estimated at ∼15% of placental clearance. This suggests fetal nonplacental clearance is minimal compared with that in the mother, but does not preclude the fetus from actively contributing to the metabolite in the fetal circulation. During infusion of AZT to the fetus, the concentration of AZT-glu in the fetus was 7.0 ± 0.8 times that in the mother. This is compelling evidence that glucuronide can be formed in the fetal compartment. Thus, fetal metabolism has an impact on the concentration of both AZT and AZT-glu in the fetal circulation.

Study of the development of fetal baboon brain using magnetic resonance imaging at 3 Tesla

Direct observational data on the development of the brains of human and nonhuman primates is on remarkably scant, and most of our understanding of primate brain development is extrapolated from findings in rodent models. Magnetic resonance imaging (MRI) is a promising tool for the noninvasive, longitudinal study of the developing primate brain. We devised a protocol to scan pregnant baboons serially at 3 T for up to 3 h per session. Seven baboons were scanned 1-6 times, beginning as early as 56 days post-conceptional age, and as late as 185 days (term approximately 185 days). Successful scanning of the fetal baboon required careful animal preparation and anesthesia, in addition to optimization of the scanning protocol. We successfully acquired maps of relaxation times (T(1) and T(2)) and high-resolution anatomical images of the brains of fetal baboons at multiple time points during the course of gestation. These images demonstrated the convergence of gray and white matter contrast near term, and furthermore demonstrated that the loss of contrast at that age is a consequence of the continuous change in relaxation times during fetal brain development. These data furthermore demonstrate that maps of relaxation times have clear advantages over the relaxation time weighted images for the tracking of the changes in brain structure during fetal development. This protocol for in utero MRI of fetal baboon brains will help to advance the use of nonhuman primate models to study fetal brain development longitudinally.

FETAL MORPHINE METABOLISM AND CLEARANCE ARE CONSTANT DURING LATE GESTATION

Fetal metabolism significantly contributes to the clearance of drugs from the fetus. To understand how the changes in fetal metabolism expected in late gestation alter fetal drug clearance, serial measurements of morphine metabolism were made in the fetal baboon over the latter third of gestation. Clearance and metabolism were evaluated in the context of fetal growth, onset of labor, and the administration of classical enzyme induction agents. Morphine, a probe substrate for the enzyme uridine diphosphate glucuronosyltransferase 2B7 (UGT2B7), was continuously infused to chronically catheterized fetal baboons while measuring morphine, morphine-3-β-glucuronide, and morphine-6-β-glucuronide concentrations. In some animals, intermittent infusions of the metabolites provided estimates of metabolite clearance and, hence, the rate of formation of metabolites and metabolic clearance. Overall, metabolic clearance of morphine from the fetus was 27 ± 9.0 ml · min–1 or 32% of total clearance. This is similar to the overall clearance in the adult baboon when standardized to weight. No change in any measure of metabolism or clearance of morphine or its glucuronide metabolites was found with gestational age, the presence of labor, or administration of UGT enzyme induction agents. Interpreting these findings using a physiologically based approach suggests that the intrinsic clearance of the fetal liver toward morphine is of sufficient magnitude that fetal hepatic clearance is flow-limited. The implication of a high intrinsic clearance is for significant placento-hepatic first-pass metabolism when drugs are administered to the mother. The previously held view of the “inadequacy of perinatal glucuronidation” needs to be reconsidered.

Zidovudine kinetics in the pregnant baboon

The devastating impact of human immunodeficiency virus (HIV) infection during pregnancy has made the pharmacologic evaluation of potentially therapeutic agents of high priority. The results presented here are the maternal pharmacokinetics from a series of experiments to delineate more clearly the complex maternal-fetal pharmacokinetics and the effects of AZT in the chronically instrumented maternal and fetal baboon during both steady state intravenous infusion and oral bolus dosage regimens. Two results of major clinical importance were found. First, during pregnancy, both the clearance and volume of distribution of AZT were increased. Plasma clearance in the pregnant animals was 51 +/- 10 ml/min/kg compared with 37 +/- 2 ml/min/kg in the nonpregnant animals, and steady state volume of distribution was 3.7 +/- 1.21/kg compared with 2.2 +/- 0.61/kg. Second, with continuous intravenous infusion plasma drug concentrations were easily maintained in the therapeutic range, whereas with oral administration plasma concentration fell below therapeutic levels within 2 h of the dose being given. Because maternal plasma concentrations are a major determinant of drug concentration achieved in the fetus, an understanding of drug kinetics in pregnancy is of vital importance when making recommendations regarding optimal drug therapy during pregnancy to maximize the beneficial effect--the prevention of HIV infection in children.

Implications of the kinetics of zidovudine in the pregnant baboon following oral administration.

Zidovudine (ZDV) therapy in pregnancy reduces mother-to-child transmission of HIV. The action of ZDV in the fetus is thought to be an important contributor to efficacy. Previous research in primates has demonstrated that continuous infusion of ZDV to the mother leads to sustained plasma concentrations in the fetus; however, it has not been determined what concentrations of ZDV are achieved in the fetus following oral administration. The pharmacokinetics of drug distribution to the fetus following oral administration of a 100-mg dose of ZDV to the mother are reported from 6 chronically catheterized baboons. The first order elimination half-life of ZDV from both the mother and fetus was approximately 1.2 hours. The area under the concentration-time curve for the fetus was 77% (r2 = 0.98; p < .001) that of the mother and the estimated peak drug levels in the fetus were 52% (r2 = 0.83; p < .01) those in the mother. The rapid transfer and short half-life of ZDV leads to a drug concentration-time profile that would not sustain levels in the fetus with dosing every 4 hours. After comparing these findings with existing data from pregnant and nonpregnant humans, it seems likely that current dose recommendations for ZDV in pregnancy would not maintain levels of the active intracellular metabolite of ZDV in all fetuses. This may explain in part the 8% failure rate of ZDV prophylaxis. The correlation between fetal and maternal plasma concentrations of ZDV would allow titration of dose based on maternal drug levels to achieve fetal levels within the therapeutic range.

Gestational age related changes in cardiac dynamics of the fetal baboon.

To provide insight into the maturation of neural mechanisms governing fetal heart rate and rate variability, seven chronically instrumented fetal baboons were monitored under steady state conditions between 120 and 165 days gestation (term 175 d). Forty records of 24 h duration (5-7 records/fetus) were evaluated. For each fetus, heart rate decreased with gestational age (mean+/-SD, r = -0.530+/-0.324, P <0.05). In contrast, there were increases with age in markers of various components of autonomic control of fetal R-wave to R-wave interval (RRi) variability as reflected in a positive correlation with age for all fetuses of SD RRi (r = 0.656+/-0.347, P < 0.01), root mean squared differences in RRi (r = 0.686+/-0.223, P <0.05), and power at low frequency in the RRi spectrum (r = 0.800+/-0.161 P < 0.01). In each of the seven fetuses, scatter plots of RRi as a function of the prior RRi (Poincare plots) had increased dispersion around the median with gestational age (0.605+/-0.371, P<0.05). Additional measures of variability evaluated changes in RRi from one interval to the next (deltaRRi). The incidence of sustained deltaRRi changes, either decelerations or accelerations, rose with gestation (r = 0.920+/-0.057, P < 0.001) while the incidence of no detected deltaRRi changes (<+/-1 ms) diminished (r = - 0.649+/-0.364, P <0.05). Sequential decreases in fetal heart rate, increases in RRi variability and increases in changes in RRi and deltaRRi with age imply an overall maturation in autonomic cardio-regulatory control processes. Increases with gestation in measures of high frequency components of variability are compatible with enhanced parasympathetic modulation of fetal heart rate.

Diurnal rhythms of fetal and maternal heart rate in the baboon

To investigate the organization of diurnal rhythmicity during gestation, the relationship between daily cycles of maternal and fetal heart rate were measured in long-term studies of healthy chronically instrumented pregnant baboons. In each of six pregnancies, hourly mean values over a 168 h time series were obtained during a 7 to 10 day interval between 135 and 160 days of gestation. Data were modeled by a least squares fit to a cosine function with a period of 24 h. Hourly mean heart rate in the fetus ranged from 161 to 172 bpm (167.9+/-0.6 bpm), and the mother from 105 to 125 bpm (107.9+/-1.4 bpm). The amplitude of the daily fluctuations were 15 to 25 bpm for the fetuses and 25 to 60 bpm for the mothers. The relation between time series data and model estimates were significant (P < 0.001) in all cases with aggregate r2 = 0.747 for fetuses and 0.737 for the mothers. On average the time of day of the peak in fetal heart rate (15:05+/-0.3 h) was about 45 min after the maternal peak (14:21+/-0.4 h). This phase delay was significant (t = 2.63, P < 0.05). There was significant (P < 0.01) diurnal periodicity for each of six parameters used to assess different aspects of fetal heart rate variability with peak variability at night (23:00 to 2:00). Thus, during the latter third of pregnancy in both the maternal and fetal baboon 24 h periodicities of heart rate are present with peak rates in the midafternoon. The daily rhythms in fetal heart rate are linked with periodicities in maternal heart rate with a phase delay in the majority of cases. The synchrony of 24 h fluctuations in rate with parameters of rate variability is consistent with diurnal input into the fetal autonomic nervous system.

Placental Transfer and Fetal Elimination of Morphine-3-β-glucuronide in the Pregnant Baboon

The glucuronide metabolites of several widely used drugs are detected in fetal plasma after maternal drug administration. However, the disposition of these metabolites is poorly understood and clinical concerns have been raised about accumulation of active metabolites in the fetus. For this reason, morphine-3-β-glucuronide (M3G), an active metabolite of morphine, was studied to provide quantitative data on disposition. Maternal, fetal, and bidirectional placental clearances of M3G were measured in three pregnant baboons. During maternal infusion of M3G to steady-state, the glucuronide metabolite readily appeared in fetal plasma achieving a mean ± S.D. fetal-to-maternal concentration ratio of 0.79 ± 0.04. In paired maternal and fetal infusions, steady-state clearances were 53 ± 3 (maternal), 1.5 ± 0.5 (maternal-to-fetal), 2.6 ± 0.1 (fetal-to-maternal), and –0.70 ± 0.6 ml · min–1 (fetal). These clearance values support bidirectional transfer of M3G across the placenta and indicate negligible direct clearance from the fetus. The clearance of M3G across the placenta is more than 20-fold less than that of morphine. Despite this low index of permeability, placental transfer contributes significantly to the glucuronide pool in the fetus. Placental transfer emerges as the major clearance pathway for the glucuronide from the fetus and suggests a component of active efflux. What is more, the results do not support the concept of sequestration in the fetal intestine as a significant route of clearance. Together these results clarify the distribution and clearance of glucuronides in the pregnant primate and facilitate prediction of fetal exposure to active metabolites.