Objective. To determine whether brainstem maturation as measured by brainstem auditory-evoked responses (BAERs) in preterm infants is a function of dietary intake.
Study Design. We obtained serial BAERs on infants 28 to 32 weeks' gestation at birth, cared for in the neonatal intensive care unit of a regional referral center in Upstate New York. Waveforms were analyzed for replicability and for the presence of waves III and V. Absolute and interwave latencies were measured. Baseline and follow-up BAER measurements were compared, and the rates of change were calculated. Patient charts were reviewed for type of enteral feeding during the interval between BAERs. Student's t test was used to analyze continuous variables and χ2 analysis was used to analyze categorical variables.
Results. Data from 37 study infants (17 fed breast milk and 20 fed commercial premature formula) revealed that there was no difference in absolute latencies of waves III and V at baseline; however, the rates of decrease of absolute latencies over the study interval were significantly greater in infants receiving human milk.
Conclusions. Infants fed breast milk have faster brainstem maturation, compared with infants fed formula, based on the rate of maturation of BAERs. This effect may be attributable to the constituent composition of breast milk, compared with synthetic formulas.
- LCPUFA =
- long-chain polyunsaturated fatty acid •
- LA =
- linoleic acid •
- LNA =
- linolenic acid •
- AA =
- arachidonic acid •
- DHA =
- docosahexaenoic acid •
- BAER =
- brainstem-evoked response •
- RBC =
- red blood cell
Human milk differs significantly from commercial infant formulas in fatty acid composition. Commercial formulas available in the United States today do not contain long-chain polyunsaturated fatty acids (LCPUFAs) with >18 carbons, whereas human milk contains not only linoleic acid (LA; 18:2ω6) and linolenic acid (LNA; 18:3ω3), but also their metabolites—arachidonic acid (AA; 20:4ω6) and docosahexaenoic acid (DHA; 22:6ω3), respectively. During fetal life, high levels of AA and DHA are maintained by selective placental transfer. After birth, long-chain metabolites of LA and LNA, including AA and DHA, are derived from human milk or by de novo synthesis from chain elongation–desaturation of LA and LNA. Thus, infants fed exclusively with formula must rely on endogenous synthesis of LCPUFAs. Experimental evidence suggests that premature infants may have limited capacity to elongate and desaturate LA and LNA.1–4
In the human brain, accelerated growth occurs during the last trimester of pregnancy. Long-chain metabolites of LA and LNA are actively incorporated into brain tissue during this critical period of brain development.5 These LCPUFAs play a major role in modulating the structure and function of neural membranes.6–8 Both neural synapses and photoreceptor plates of the retina are enriched in DHA. Numerous studies in animals have demonstrated the importance of DHA for brain and retinal development.9–11 Although there is considerable evidence for the importance of LCPUFAs for human brain development, the information available has not been sufficient to advocate routine supplementation of LCPUFAs in infant formulas. The importance of LCPUFAs on neurodevelopment in term infants was demonstrated by Agostini et al12 who found that term infants who received formula supplemented with LCPUFAs had better psychomotor performance at 4 months old, compared with term infants who received formula including LA and LNA but lacking LCPUFAs. Similarly, Horwood and Fergusson13 demonstrated that breastfed term infants had better cognitive skills and achieved higher academic scores when tested over the first 18 years of life, compared with formula-fed infants. In premature infants, Lucas et al14 showed the beneficial effect of breast milk during infancy on the neurodevelopment at 18 months old and at 8 years old. The factor associated with breast milk feedings responsible for this difference was postulated to be LCPUFAs. The importance of ω3 LCPUFAs for retinal and cortical function has been demonstrated for both term and preterm infants.15–19
Brainstem auditory-evoked response (BAER) has proven useful as a noninvasive electrophysiologic measure of brainstem function and maturation during early development.20 BAER has been used to evaluate the need for taurine supplementation for brain maturation in premature infants receiving formula devoid of taurine21and to evaluate brain maturation in infants with transient thyroxinemia.22 This test measures neural conduction both peripherally and centrally. The characteristic wave form in neonates is comprised of 3 waves (I, III, and V). Electrophysiologic data suggest that wave I is generated peripherally in the auditory nerve. Wave III reflects the firing of axons exiting the cochlear nuclear complex in the brainstem, while wave V reflects an action potential generated by axons from the lateral lemniscus at a more rostral brainstem location.23 Interwave intervals reflect axonal conduction and synaptic function.24 Axonal growth, synaptic function, and degree of myelination influence the wave form.25 BAER undergoes rapid changes that parallel the period of rapid brain development in preterm infants, particularly at 28 to 32 weeks' gestational age.20 With increasing age, both absolute and interwave latencies decrease suggesting maturation.
We postulated that using serial BAERs, we could demonstrate a beneficial effect of human milk feedings, compared with formula on brainstem maturation in preterm infants. Testing during this period of development would allow the highest probability of detection of differences in maturation based on nutritional intake given the rapid changes that are normally occurring in the brain.
All infants 28 to 32 weeks' gestational age at birth admitted to the neonatal intensive care unit of the Children's Hospital at Strong from October 1996 to April 1998 were eligible for the study. Amin et al20 previously demonstrated that infants do not reliably demonstrate a mature wave form until after ∼28 weeks of corrected gestational age. Moreover, the BAER wave form showed the most marked maturation over the period from 28 to 32 weeks of corrected gestational age. For this reason, we concentrated on this gestational period for our study. Gestational age was assessed by obstetrical dating or, in those infants whose obstetric data were inadequate, by Ballard examination. After parental consent was obtained, sequential bilateral monaural BAERs were performed on 5 days during the first postnatal week by skilled audiologists blinded to type of feeding. BAERs were categorized into response types based on replicability and the presence of waves III and V (Fig 1). If a replicable wave form with both waves III and V (type 1) was not identifiable by the end of the first postnatal week, a BAER was repeated at weekly intervals until a type 1 response was obtained. Absolute and interwave latencies were measured. The baseline BAER was defined as a type 1 response with decreasing absolute latencies, compared with the previous BAER. The comparison BAER was performed at a mean interval of 2 weeks after the baseline BAER.
BAERs were recorded with a Bio-logic Navigator Evoked Response (Bio-logic Systems Corp, Mundelein, IL) with the infant lying supine in the isolette and with a skin temperature >35.5°C. Electrode sites were mastoid (reference), midline on high forehead or crown of the head (active), and shoulder (ground). Electrode gel was applied to silver/silver chloride electrodes. Monaural testing was performed on each ear using 80-dB nHL broadband click stimuli with supraaural earphones. The clicks were presented at a repetition rate of 39.9/second, and 3 runs of 2000 repetitions were recorded for each ear. These testing parameters were chosen to optimize BAER recordings in extremely premature infants. Testing took ∼20 minutes. The 2 most replicable runs for each ear were averaged and used for later analysis. The BAERs were analyzed by audiologists without knowledge of gestational age, previous BAER results, or dietary intake of the infant.
Neonates for whom serial BAERs demonstrated the presence of type 1 responses with decreasing latencies of waves III and V and for whom a follow-up BAER was obtained were analyzed for rate of brainstem maturation. The decrease in latencies of waves III and V and wave III/V interwave intervals were calculated over the interval between baseline and follow-up for each ear for each infant and expressed as milliseconds/week. The data from the ear with the more mature response were used for comparisons.
Patient charts were reviewed retrospectively to determine the type of enteral intake on each day during the interval between BAERs. The total caloric intake at baseline and the proportion of enteral intake (expressed as percent of total intake) were calculated at baseline and at follow-up. Infants who received both breast milk and formula were not included because of the wide variation in relative amounts and timing of breast milk and formula exposure. In addition, infants who did not receive any enteral feeds for >3 days during the study interval and infants with culture-proven sepsis were also excluded from analysis. Because repeated transfusions of adult packed red blood cells (RBCs) have been shown to increase infant plasma and RBC membrane content of polyenoic fatty acids and because this may affect the accretion of these fatty acids in the developing brain, the number and volume of transfusions received during the interval period also were recorded.
Student's t test was used to analyze continuous variables using Stata (Stata Corporation, College Station, TX). A χ2 analysis was used to analyze categorical variables. All tests were 2-sided and a P value <.05 was considered statistically significant.
Sixty-seven infants were enrolled in the study. Nine infants did not demonstrate a type 1 response by 32 weeks' corrected gestational age and were not included in the analysis. Of the remaining 58 infants, 21 were excluded from analysis: 10 infants who received both formula and human milk during the interval period, 10 infants who were not receiving any enteral feeds for >3 days during the interval, and 1 infant with culture-proven sepsis.
Demographic features of the 37 study infants are shown in Table 1. Of the study group, 17 infants received human milk and 20 infants received formula (Special Care Formula, Ross Products Division, Abbott Laboratories, Inc, Columbus, OH). There was no difference between the groups in gestational age at birth, corrected gestational age at baseline, weight at baseline, or duration of the interval between baseline and follow-up testing. The volume and caloric content of enteral intake at baseline and follow-up BAER were similar in the 2 groups. There was no difference in the number or total volume of blood transfusions. Three patients in the formula group and 4 patients in the breast milk group received gentamicin during the study. Gentamicin levels were monitored routinely and none were in the ototoxic range. No patients were receiving lasix during the study. Two patients, both in the breast milk group, were diagnosed with grade II intraventricular hemorrhages. There was a difference in the racial distribution in the 2 feeding groups. However, an analysis of BAERs performed during the first 24 hours after birth in ∼150 premature infants 28 to 32 weeks of gestational age showed that there was no difference in the frequency with which wave V could be detected (60/111 vs 17/38; P = .42) and that the wave V latency was no different between whites and blacks (9.53 milliseconds vs 9.64 milliseconds; P = .66).
There was no difference between the groups in the absolute latencies of waves III and V at baseline (Table 2). However, the rates of decrease in absolute latency of waves III and V over the interval period were significantly greater in infants receiving human milk, compared with those receiving formula (P = .04 and P = .02, respectively;Table 3). There was also a trend toward a greater rate of decrease in the III/V interwave interval in the infants receiving breast milk (P = .18; Table 3).
Our findings suggest that infants fed breast milk have faster brainstem maturation, compared with infants fed formula, based on the rate of maturation of BAERs. Using BAER measurements, we have demonstrated a short-term effect of breast milk on human brain development that may be attributable to the constituent composition of human milk compared with synthetic formulas.
The brain is 60% structural lipid and uses both AA and DHA as basic building blocks. During the third trimester of intrauterine development, marked accretion of LCPUFAs occurs in the human brain. Synaptogenesis and dendritic arborization increase significantly during the last trimester of intrauterine development explaining the rapid accretion of polyunsaturated fatty acids.26 LCPUFAs are essential for the structure and function of neuronal and glial membranes, creating a membrane with high fluidity, flexibility, and permeability.6–8 Their role as neuronal membrane components may involve modulation of the kinetics of carrier-mediated transport systems, properties of membrane receptors, and/or the activity of membrane-bound enzymes.11 In addition, some LCPUFAs (eg, adrenic acid; 22:4ω6) are important components of myelin.5 Myelination is associated with rapid conduction capability.25 There is experimental evidence from animal studies that both neural integrity and function can be permanently affected by deficiencies of these elongation–desaturation products of linoleic and linolenic acids.10,11
Human milk contains long-chain polyenoic derivatives of LA and LNA, primarily AA and DHA, while commercial formulas and intravenous lipids available currently in the United States do not contain either AA or DHA. Milk secreted by mothers of preterm infants has a higher content of LA and LNA and their long-chain polyenoiec derivatives.27 The amount of LCPUF present in preterm human milk seems to be sufficient to meet the requirements of neural tissue for fatty acids.28 In addition, animal experiments suggest that preformed LCPUFAs are incorporated into the developing brain with a 10-fold greater efficiency, compared with similar fatty acids synthesized de novo from LA and LNA.29
Premature infants fed commercial formulas containing LA and LNA, but not DHA and AA, have significantly lower plasma and erythrocyte membrane concentrations of the long-chain metabolites, compared with those receiving human milk. Similarly, lower hepatic content of DHA and AA were found in formula-fed preterm infants, compared with infants fed breast milk.3 This suggests a limited capacity of the enzyme system necessary to elongate and desaturate the 18-carbon precursors.2,17 Most infant formulas contain LA in excess of the infant's requirement, with a high LA:LNA ratio (>10:1). Despite high levels of the precursor LA in infant formulas, several studies have reported paradoxically reduced levels of AA in erythrocyte phospholipids.30 Because the 18-carbon precursors (LA and LNA) compete for the same elongase and desaturase enzyme systems, investigators have postulated that the high LA content of the formula may be responsible for decreased DHA levels in erythrocytes.30 However, despite decreasing the LA:LNA ratio in formula to 4:1, the DHA status of breastfed infants was not achieved in the formula-fed infants.30 Further decrease in the LA:LNA ratio affected AA status.30 Clark et al30 speculated that the presence of AA and DHA in human milk is biologically significant. Although there is an increase in erythrocyte membrane concentrations of both AA and DHA in breastfed infants, only DHA concentration is shown to be increased in the brain of breastfed infants, compared with infants fed formula.1,31
To date, there has been little information regarding the functional importance of the LCPUFAs in the developing brain of premature infants. The importance of DHA for retinal and cortical function has been demonstrated in premature infants.15,17 Our data are the first to suggest an effect of some component of human milk on brainstem maturation. Brainstem maturation is faster in premature infants fed human milk, compared with formula-fed infants. Evidence based on BAERs, especially the wave III and V absolute latencies and the interwave latencies, suggests more rapid axonal growth, synaptic function, and myelination in the breastfed neonates. Although breastfed infants were primarily white and formula-fed infants were primarily black, the fact that the latencies of waves III and V at study entry were similar argues against the differences in brainstem maturation being a function of racial differences. In addition, no racial differences were observed in the detectability of wave V or wave V latency at birth in a larger group of infants 28 to 32 weeks of gestational age at birth (data not shown).
The improvement in brainstem maturation may be clinically relevant because the degree of brainstem maturation may affect the frequency of other clinical entities such as apnea of prematurity.32Available literature reports that breastfed infants have a lower incidence of sudden infant death syndrome.33Although the underlying mechanism for the decreased incidences of these entities is unknown, more mature brainstem function (including control of breathing) in the breastfed infants could partially explain the observation. The rate of brainstem maturation also may reflect the rate of maturation of other parts of the brain. Our data are consistent with previous reports that showed improved IQ and visual and cortical function in premature infants fed human milk or formula supplemented with LCPUFAs, compared with those infants fed standard formula. Although we studied brainstem maturation in premature infants over a relatively short period of development, the possibility of a beneficial effect extending beyond term (40 weeks' gestational age) is very high because rapid accretion of structural lipids continues for several postterm months. We speculate that the difference in brainstem maturation is attributable at least in part to the presence of DHA in breast milk.
It remains to be shown that feeding premature neonates with commercial formulas supplemented with LCPUFAs will result in comparable rates of brainstem maturation as feeding with human milk. We believe that BAER, a noninvasive electrophysiologic measure of brainstem function, is an important and useful tool that may address this question.
We thank Todd M. Gibson, Teri D. Holt, Matthew MacDonald, Lynette McRae, Diane Puccia, and Catherine Papso Seeger for performing the brainstem auditory-evoked response testing on the infants.
We thank all of the neonatal intensive care unit nurses who assisted in positioning and monitoring the infants during testing.
We also thank Dr Ruth Lawrence for her critical review and guidance in preparing this manuscript.
- Received March 1, 1999.
- Accepted August 9, 1999.
Reprint requests to (R.G.) Department of Pediatrics, Division of Neonatology, Box 651, Children's Hospital at Strong, University of Rochester, 601 Elmwood Ave, Rochester, NY 14642. E-mail:
Dr Amin is currently affiliated with DC General Hospital, Washington, DC.
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- Copyright © 2000 American Academy of Pediatrics