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Abnormalities of diaphragmatic muscle in neonates with ventilated lungs
Abnormalities of diaphragmatic muscle in neonates with ventilated lungs A. S. Knisely, MD, S u s a n a M. Leal, MD, a n d D o n B. Singer, MD From the Program .in Developmental Pathology and the Integrated Training Program in Pathology, Brown University, and the Department of Pathology and Laboratory Medicine, Women and Infants' Hospital of Rhode Island, Providence
Several infants and neonates who had received long-term ventilatory assistance had subnormal diaphragmatic muscle mass on gross necropsy examination. We conducted a retrospective study of our hospital infant necropsy files to determine whether prolonged ventilatory support was associated with diminution in myoflber cross-sectional area selectively affecting the diaphragm. We found that long-term ventilatory assistance may predispose diOphragmatic myoflbers to disuse atrophy or to failure of normal growth. This phenomenon may contribute to difficulties in weaning infants from ventilatory support. (J PEDIATR1988;113:1074-7)
On necropsy examination of several neonates and infants who had required long-term ventilatory support, we noted marked thinning of the muscular portions of the diaphragms. Diaphragmatic myofibers in these patients also appeared small; whereas voluhtary myofibers elsewhere appeared normal. Prolonged pharmacologic neuromuscular blockade has been associated with failure of muscular growth in infants with mechanically ventilated lungs, ~ but neuromuscular blocking agents could not be implicated in the findings in our patients. In healthy premature infants, the energy required for diaphragmatic work can represent as much as 10% of basal metabolic expenditure, a proportion that may be greater in infants with pulmonary disease. 2 It seemed possible to us that neonates respond to ventilatory support by conserving ventilatory effort, and that disuse of respiratory muscles leads to their relative atrophy; This report presents the i'esults of our retrospective evaluation of whether prolonged ventilatory support in severely ill neonates and infants was associated with decreases in muscle mass that disproportionately affected the diaphragm. Presented in part at the 76th annual meeting of the United States and Canadian Academy of Pathology, Chicago, Ill., March 8-13, 1987. Submitted for publication April 27, 1988; accepted June 16, 1988. Reprint requests: A. S. Knisel~r MD, Department of Pathology and Laboratory Medicine, Women and Infant's Hospital of Rhode Island, 101 Dudley St., Providence, RI 02905. 1074
METHODS
The study subjects were 13 neonates or infants who had received uninterrupte d ventilatory assistance for 12 days or more immediately before death. The comparison subjects D/S D/T
Diaphragm/strap muscle (ratio) Diaphragm/tongue (ratio)
]
were 26 neonates or infants, matched with the study subjects for postconceptional ages, who before death had received ventilatory support for 7 days or less. The intervals defining the two groups of subjects were arbitrary, but in selecting study and comparison subjects from necropsy files, we used the intervals prospectively. The intervals were chosen in the expectation that they would distinguish subjects who were severely ill and who required prolonged ventilatory support from subjects with severe illness but without chronic respiratory disease. Systemic neuromuscular disease was not identified in any subject. Both the study subjects and the comparison subjects were patients in the neonatal intensive care unit at Women and Infants' Hospital of Rhode Island from 1979 to 1986. Neuromuscular blocking agents were not administered to any subject. Chart review found that both sets of subjects were described as exhibiting 0nly slight resistance to ventilatory support. The study subjects as a group were characterized by minimal spontaneous respiratory effort and by a requirement for ventilatory assistance greater than that necessary for the comparison subjects. The
Volume 113 Number 6
Diaphragm abnormalities in neonates with ventilated lungs
comparison subjects were characterized by acute terminal illness such as meconium aspiration pneumonia or overwhelming sepsis. For each subject, sections of the costal diaphragm, the neck block (including portions of infrahyoid strap muscle ["strap muscle"]), and the posterior portion of the tongue were retrieved from hospital necropsy files. In three of the comparison subjects, no sections of the posterior portion of the tongue were available. Strap muscle and tongue were intended as internal controls for the systemic catabolic effects of illness? Muscles from these sites, rather than paraspinous or psoas muscles, were chosen because oropharyngeal motor control, essential to sucking, swallowing, and defense of the upper airway against aspiration, appears in infants to be coordinated with diaphragmatic function. 4,5 Bulbar muscle (posterior tongue), muscle innervated by C1-C2 high cervical motor neurons (infrahyoid strap muscle), and diaphragmatic muscle, which is innervated by C3-C5 motor neurons, might accordingly be expected to develop in parallel. Strap muscle and tongue thus were selected as less likely to be affected by maturation-related variation in myofiber size. s The tissues had been fixed in either Bouin fluid or 10% phosphate-buffered formalin, dehydrated, paraffin-embedded, sectioned at 4 ~m, and stained with hematoxylin and eosin in routine fashion. The perimeters of 25 contiguous myofibers in true cross section were traced at a magnification of x400, with the use of a camera lucida, for muscle from each site in each subject. An Apple Graphics digitizing tablet and Apple Graphics software with an Apple II computer (Apple Computer, Inc., Cupertino, Calif.) were used to determine the cross-sectional areas of individual rnyofibers ("areas") from the tracings. Average values were calculated for areas at each site in each subject. Ratios of diaphragm to strap muscle and diaphragm to tongue average areas were calculated for the study subjects and for the comparison subjects. Differences in values for D/S and D / T ratios between study and comparison subjects were compared for significance, after logarithmic transformation, by twotailed unpaired t testing. Statistical calculations were performed by means of version .99 of StatView 512+ (BrainPower, Inc., Calabasas, Calif.) with a Macintosh Plus computer (Apple Computer). RESULTS
Histopathologic findings. In the neonates and infants who had received long-term ventilatory support, extradiaphragmatic voluntary myofibers appeared normal, whereas diaphragmatic myofibers generally appeared small, with rounded outlines. Wohlfart type B fibers 6 in their diaphragms were more prominent than in the comparison subjects. These changes appeared to affect dia-
10 7 5
Fig, t. A, Diaphragmatic myofibers (average cross-sectional area: 97.4 iz~) of a male term infant, birth weight 3450 g, continuously ventilated from postnatal day 1 until death at 47 days postnatal age. Arrow indicates Wohlfart type B myofiber. (Hematoxylin-eosin stain; original magnification • bar = 30 ~m.) B, Diaphragmatic myofibers (average cross-sectional area: 315.9 ~m2) of a mate term infant, birth weight 3300 gin, never supported by ventilation before accidental death at 3 days postnatal age. Compare with A, above. (Hematoxylin-eosin stain; original magnification • bar = 30 am.)
phragmatic myofibers diffusely rather than focally. Photomicrographs of diaphragmatic myofibers from a study subject and a comparison subject are shown in Fig. 1. Statistical findings. The D / S ratios were significantly smaller in study subjects than in comparison subjects (p < 0.0009). The D / T ratios also were significantly smaller in study subjects than in comparison subjects (p < 0.0001). Graphs of the ranges of D / S and D / T ratios in the study and comparison subjects are shown in Figs. 2 and 3. DISCUSSION Normal spontaneous ventilation in infants is diaphragmdriven. When myofibers in the periphery of the diaphragm
107 6
Knisely, Leal, and Singer
The Journal of Pediatrics December 1988
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Fig. 2. Scattergraph of D/S ratios of myofiber cross-sectional areas in 13 study subjects (closed circles, left) and 26 comparison subjects (open circles, right). D/S ratios are generally higher in comparison subjects.
Fig. 3. Scattergraph of D/T ratios of myofiber cross-sectional areas in 13 study subjects (closed circles, left) and 23 comparison subjects (open circles, right). D/T ratios are generally higher in comparison subjects.
contract, the membranotendinousdomes of the hemidiaphragms are drawn caudally. Intrathoracic volume increases, intrathoracic pressure decreases, and air enters the lungs. Assisted ventilation is externally driven. Although the domes of the hemidiaphragms are caudally displaced, diaphragmatic excursion is passive. Electromyography of respiratory muscle has shown that voluntary diaphragmatic activity in premature infants with mechanically ventilated lungs may be markedly decreased. 7 Decreased voluntary diaphragmatic activity is perhaps a valuable
adaptation to ventilatory support in that it conserves energy in the severely ill neonate, but it also may be expected to result in disuse atrophy of diaphragmatic muscle. Respiratory eleetromyograms were not obtained in our subjects, so the extent of voluntary diaphragmatic activity present is not specifically known. In this respect, it is of interest that the study subjects as a group were characterized by minimal spontaneous respiratory effort. Histologic findings in the diaphragms of the neonates
Volume 113 Number 6
Diaphragm abnormalities in neonates with ventilated lungs'
and infants supported by mechanical ventilation for 12 days or more were consistent with disuse atrophy, denervation atrophy, or failure of normal growth and maturation. If strap muscle, which may serve an accessory function in respiration, also underwent disuse atrophy, any resultant changes were not apparent by microscopy. This seeming discrepancy may reflect only the relative contributions of the diaphragm and of strap muscle to normal respiration. Changes resulting from ventilation-induceddisuse atrophy might be expected to be more pronounced in the diaphragm, which does more respiratory work. Loss of muscle mass because of the systemic catabolic effects of severe illness also might be expected to be relatively more pronounced in the bulky diaphragm than in tongue or strap muscle. Chart review did not identify apparent differences in nutritional status between study subjects and comparison subjects. We attempted to control for any effects of malnutrition on muscle bulk by studying ratios of myofiber cross-sectional areas from various sites rather than mean values for the areas themselves, but the possible confounding factor of differences in nutritional status could not be rigorously excluded in a retrospective study. Because no data were available on myofiber areas before ventilatory support was begun, we could not eliminate the possibility that abnormalities in growth or maturation contributed to our findings.8,9 If failure of normal growth occurred, it may be that passive ventilation, in turn, contributed to failure of diaphragmatic myofibers to grow. The prominence of Wohlfart type B myofibers is of interest in this regard; we could not determine whether they persisted abnormally or, as may occur, had returned to prominence in the setting of muscle atrophy. 6 The retrospective nature of our study did not permit histochemical evaluation of fiber type distribution, and neither peripheral nerves nor levels of brain stem and cervical spinal cord serving the muscles studied were specifically examined, but diaphragmatic muscle was selectively affected in a prominent manner, and strap and bulbar muscles were relatively spared. Accordingly, isolated phrenic-nerve injury or well-demarcated C3-C5 spinal-cord lesions must be invoked as common factors if the observed pattern of decrease in myofiber area is to be attributed to denervation atrophy. These explanations do not seem tenable. Weaning from ventilatory support represents a major
10 7 7
challenge to the severely ill patient, particularly in infancy, and the challenge may be greater if the capacity of the diaphragm to work is less than normal. Our results indicate that, in infants, prolonged ventilatory support is associated with a selective diminution in diaphragmatic muscle mass. This may in turn reduce infants' capacity for sustained respiratory work and complicate the process of weaning from ventilatory support. Serial respiratory-muscle electromyographic studies in infants receiving ventilatory support and prospective studies of patterns of diaphragmatic myofiber typing in such infants will be of interest in distinguishing whether this diminution in diaphragmatic muscle mass is due to abnormalities in myofiber growth and development, or to disuse atrophy of normally developing myofibers. They also m a y b e of value in selecting prophylactic and therapeutic measures. We thank S. H. Loring, MD, for a critical review of the manuscript, and S. R. Gaspar, MD, A. L. Mansetl, MD, and R. D. Plotz, MD, for helpful discussions.
REFERENCES
1. Rutledge ML, Hawkins EP, Langston C. Skeletal muscle growth failure induced in premature newborn infants by prolonged pancuronium treatment. J PEDIATR 1986;109: 883-6. 2. Guslits BG, Gaston SE, Bryan MH, England SJ, Bryan AC. Diaphragmatic work of breathing in premature human infants. J Appt Physiol 1987;62:1410-5. 3. Kelsen SG, Ference M, Kapoor S. Effects of prolonged undernutrition on structure and function of the diaphragm. J Appl Physiol 1985;58:1354-9. 4. Weber F, Woolridge MW, Baum JD. An ultrasonographic study of the organisation of sucking and swallowing by newborn infants. Dev Med Child Neurot 1986;28:19-24. 5. Selley WG, Ellis RE, Flack FC, Curtis H, Callon M. Ultrasonographic study of sucking and swallowing by newborn infants. Dev Med Child Neurol 1986;28:814-23. 6. Kakutas BA, Adams RD. Diseases of muscle: pathological foundations of clinical myology.4th ed. Philadelphia: Harper & Row, 1985:8-11, 16-7. 7. O'Brien MJ, Van Eykern LA, Oetomo SB, Van Vught HAJ. Transeutaneous respiratory electromyographic monitoring. Crit Care Med 1987;I5:294-9. 8. Keens TG, Bryan AC, LevisonH, lanuzzo CD. Developmental pattern of muscle fiber types in human ventilatory muscles. J Appl Physiol 1978;44:909-13. 9. Keens TG, Ianuzzo CD. Development of fatigue-resistant muscle fibers in human ventilatory muscles. Am Rev Respir Dis 1979;119:139-41.
Several infants and neonates who had received long-term ventilatory assistance had subnormal diaphragmatic muscle mass on gross necropsy examination. We conducted a retrospective study of our hospital infant necropsy files to determine whether prolonged ventilatory support was associated with diminution in myoflber cross-sectional area selectively affecting the diaphragm. We found that long-term ventilatory assistance may predispose diOphragmatic myoflbers to disuse atrophy or to failure of normal growth. This phenomenon may contribute to difficulties in weaning infants from ventilatory support. (J PEDIATR1988;113:1074-7)
On necropsy examination of several neonates and infants who had required long-term ventilatory support, we noted marked thinning of the muscular portions of the diaphragms. Diaphragmatic myofibers in these patients also appeared small; whereas voluhtary myofibers elsewhere appeared normal. Prolonged pharmacologic neuromuscular blockade has been associated with failure of muscular growth in infants with mechanically ventilated lungs, ~ but neuromuscular blocking agents could not be implicated in the findings in our patients. In healthy premature infants, the energy required for diaphragmatic work can represent as much as 10% of basal metabolic expenditure, a proportion that may be greater in infants with pulmonary disease. 2 It seemed possible to us that neonates respond to ventilatory support by conserving ventilatory effort, and that disuse of respiratory muscles leads to their relative atrophy; This report presents the i'esults of our retrospective evaluation of whether prolonged ventilatory support in severely ill neonates and infants was associated with decreases in muscle mass that disproportionately affected the diaphragm. Presented in part at the 76th annual meeting of the United States and Canadian Academy of Pathology, Chicago, Ill., March 8-13, 1987. Submitted for publication April 27, 1988; accepted June 16, 1988. Reprint requests: A. S. Knisel~r MD, Department of Pathology and Laboratory Medicine, Women and Infant's Hospital of Rhode Island, 101 Dudley St., Providence, RI 02905. 1074
METHODS
The study subjects were 13 neonates or infants who had received uninterrupte d ventilatory assistance for 12 days or more immediately before death. The comparison subjects D/S D/T
Diaphragm/strap muscle (ratio) Diaphragm/tongue (ratio)
]
were 26 neonates or infants, matched with the study subjects for postconceptional ages, who before death had received ventilatory support for 7 days or less. The intervals defining the two groups of subjects were arbitrary, but in selecting study and comparison subjects from necropsy files, we used the intervals prospectively. The intervals were chosen in the expectation that they would distinguish subjects who were severely ill and who required prolonged ventilatory support from subjects with severe illness but without chronic respiratory disease. Systemic neuromuscular disease was not identified in any subject. Both the study subjects and the comparison subjects were patients in the neonatal intensive care unit at Women and Infants' Hospital of Rhode Island from 1979 to 1986. Neuromuscular blocking agents were not administered to any subject. Chart review found that both sets of subjects were described as exhibiting 0nly slight resistance to ventilatory support. The study subjects as a group were characterized by minimal spontaneous respiratory effort and by a requirement for ventilatory assistance greater than that necessary for the comparison subjects. The
Volume 113 Number 6
Diaphragm abnormalities in neonates with ventilated lungs
comparison subjects were characterized by acute terminal illness such as meconium aspiration pneumonia or overwhelming sepsis. For each subject, sections of the costal diaphragm, the neck block (including portions of infrahyoid strap muscle ["strap muscle"]), and the posterior portion of the tongue were retrieved from hospital necropsy files. In three of the comparison subjects, no sections of the posterior portion of the tongue were available. Strap muscle and tongue were intended as internal controls for the systemic catabolic effects of illness? Muscles from these sites, rather than paraspinous or psoas muscles, were chosen because oropharyngeal motor control, essential to sucking, swallowing, and defense of the upper airway against aspiration, appears in infants to be coordinated with diaphragmatic function. 4,5 Bulbar muscle (posterior tongue), muscle innervated by C1-C2 high cervical motor neurons (infrahyoid strap muscle), and diaphragmatic muscle, which is innervated by C3-C5 motor neurons, might accordingly be expected to develop in parallel. Strap muscle and tongue thus were selected as less likely to be affected by maturation-related variation in myofiber size. s The tissues had been fixed in either Bouin fluid or 10% phosphate-buffered formalin, dehydrated, paraffin-embedded, sectioned at 4 ~m, and stained with hematoxylin and eosin in routine fashion. The perimeters of 25 contiguous myofibers in true cross section were traced at a magnification of x400, with the use of a camera lucida, for muscle from each site in each subject. An Apple Graphics digitizing tablet and Apple Graphics software with an Apple II computer (Apple Computer, Inc., Cupertino, Calif.) were used to determine the cross-sectional areas of individual rnyofibers ("areas") from the tracings. Average values were calculated for areas at each site in each subject. Ratios of diaphragm to strap muscle and diaphragm to tongue average areas were calculated for the study subjects and for the comparison subjects. Differences in values for D/S and D / T ratios between study and comparison subjects were compared for significance, after logarithmic transformation, by twotailed unpaired t testing. Statistical calculations were performed by means of version .99 of StatView 512+ (BrainPower, Inc., Calabasas, Calif.) with a Macintosh Plus computer (Apple Computer). RESULTS
Histopathologic findings. In the neonates and infants who had received long-term ventilatory support, extradiaphragmatic voluntary myofibers appeared normal, whereas diaphragmatic myofibers generally appeared small, with rounded outlines. Wohlfart type B fibers 6 in their diaphragms were more prominent than in the comparison subjects. These changes appeared to affect dia-
10 7 5
Fig, t. A, Diaphragmatic myofibers (average cross-sectional area: 97.4 iz~) of a male term infant, birth weight 3450 g, continuously ventilated from postnatal day 1 until death at 47 days postnatal age. Arrow indicates Wohlfart type B myofiber. (Hematoxylin-eosin stain; original magnification • bar = 30 ~m.) B, Diaphragmatic myofibers (average cross-sectional area: 315.9 ~m2) of a mate term infant, birth weight 3300 gin, never supported by ventilation before accidental death at 3 days postnatal age. Compare with A, above. (Hematoxylin-eosin stain; original magnification • bar = 30 am.)
phragmatic myofibers diffusely rather than focally. Photomicrographs of diaphragmatic myofibers from a study subject and a comparison subject are shown in Fig. 1. Statistical findings. The D / S ratios were significantly smaller in study subjects than in comparison subjects (p < 0.0009). The D / T ratios also were significantly smaller in study subjects than in comparison subjects (p < 0.0001). Graphs of the ranges of D / S and D / T ratios in the study and comparison subjects are shown in Figs. 2 and 3. DISCUSSION Normal spontaneous ventilation in infants is diaphragmdriven. When myofibers in the periphery of the diaphragm
107 6
Knisely, Leal, and Singer
The Journal of Pediatrics December 1988
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Fig. 2. Scattergraph of D/S ratios of myofiber cross-sectional areas in 13 study subjects (closed circles, left) and 26 comparison subjects (open circles, right). D/S ratios are generally higher in comparison subjects.
Fig. 3. Scattergraph of D/T ratios of myofiber cross-sectional areas in 13 study subjects (closed circles, left) and 23 comparison subjects (open circles, right). D/T ratios are generally higher in comparison subjects.
contract, the membranotendinousdomes of the hemidiaphragms are drawn caudally. Intrathoracic volume increases, intrathoracic pressure decreases, and air enters the lungs. Assisted ventilation is externally driven. Although the domes of the hemidiaphragms are caudally displaced, diaphragmatic excursion is passive. Electromyography of respiratory muscle has shown that voluntary diaphragmatic activity in premature infants with mechanically ventilated lungs may be markedly decreased. 7 Decreased voluntary diaphragmatic activity is perhaps a valuable
adaptation to ventilatory support in that it conserves energy in the severely ill neonate, but it also may be expected to result in disuse atrophy of diaphragmatic muscle. Respiratory eleetromyograms were not obtained in our subjects, so the extent of voluntary diaphragmatic activity present is not specifically known. In this respect, it is of interest that the study subjects as a group were characterized by minimal spontaneous respiratory effort. Histologic findings in the diaphragms of the neonates
Volume 113 Number 6
Diaphragm abnormalities in neonates with ventilated lungs'
and infants supported by mechanical ventilation for 12 days or more were consistent with disuse atrophy, denervation atrophy, or failure of normal growth and maturation. If strap muscle, which may serve an accessory function in respiration, also underwent disuse atrophy, any resultant changes were not apparent by microscopy. This seeming discrepancy may reflect only the relative contributions of the diaphragm and of strap muscle to normal respiration. Changes resulting from ventilation-induceddisuse atrophy might be expected to be more pronounced in the diaphragm, which does more respiratory work. Loss of muscle mass because of the systemic catabolic effects of severe illness also might be expected to be relatively more pronounced in the bulky diaphragm than in tongue or strap muscle. Chart review did not identify apparent differences in nutritional status between study subjects and comparison subjects. We attempted to control for any effects of malnutrition on muscle bulk by studying ratios of myofiber cross-sectional areas from various sites rather than mean values for the areas themselves, but the possible confounding factor of differences in nutritional status could not be rigorously excluded in a retrospective study. Because no data were available on myofiber areas before ventilatory support was begun, we could not eliminate the possibility that abnormalities in growth or maturation contributed to our findings.8,9 If failure of normal growth occurred, it may be that passive ventilation, in turn, contributed to failure of diaphragmatic myofibers to grow. The prominence of Wohlfart type B myofibers is of interest in this regard; we could not determine whether they persisted abnormally or, as may occur, had returned to prominence in the setting of muscle atrophy. 6 The retrospective nature of our study did not permit histochemical evaluation of fiber type distribution, and neither peripheral nerves nor levels of brain stem and cervical spinal cord serving the muscles studied were specifically examined, but diaphragmatic muscle was selectively affected in a prominent manner, and strap and bulbar muscles were relatively spared. Accordingly, isolated phrenic-nerve injury or well-demarcated C3-C5 spinal-cord lesions must be invoked as common factors if the observed pattern of decrease in myofiber area is to be attributed to denervation atrophy. These explanations do not seem tenable. Weaning from ventilatory support represents a major
10 7 7
challenge to the severely ill patient, particularly in infancy, and the challenge may be greater if the capacity of the diaphragm to work is less than normal. Our results indicate that, in infants, prolonged ventilatory support is associated with a selective diminution in diaphragmatic muscle mass. This may in turn reduce infants' capacity for sustained respiratory work and complicate the process of weaning from ventilatory support. Serial respiratory-muscle electromyographic studies in infants receiving ventilatory support and prospective studies of patterns of diaphragmatic myofiber typing in such infants will be of interest in distinguishing whether this diminution in diaphragmatic muscle mass is due to abnormalities in myofiber growth and development, or to disuse atrophy of normally developing myofibers. They also m a y b e of value in selecting prophylactic and therapeutic measures. We thank S. H. Loring, MD, for a critical review of the manuscript, and S. R. Gaspar, MD, A. L. Mansetl, MD, and R. D. Plotz, MD, for helpful discussions.
REFERENCES
1. Rutledge ML, Hawkins EP, Langston C. Skeletal muscle growth failure induced in premature newborn infants by prolonged pancuronium treatment. J PEDIATR 1986;109: 883-6. 2. Guslits BG, Gaston SE, Bryan MH, England SJ, Bryan AC. Diaphragmatic work of breathing in premature human infants. J Appt Physiol 1987;62:1410-5. 3. Kelsen SG, Ference M, Kapoor S. Effects of prolonged undernutrition on structure and function of the diaphragm. J Appl Physiol 1985;58:1354-9. 4. Weber F, Woolridge MW, Baum JD. An ultrasonographic study of the organisation of sucking and swallowing by newborn infants. Dev Med Child Neurot 1986;28:19-24. 5. Selley WG, Ellis RE, Flack FC, Curtis H, Callon M. Ultrasonographic study of sucking and swallowing by newborn infants. Dev Med Child Neurol 1986;28:814-23. 6. Kakutas BA, Adams RD. Diseases of muscle: pathological foundations of clinical myology.4th ed. Philadelphia: Harper & Row, 1985:8-11, 16-7. 7. O'Brien MJ, Van Eykern LA, Oetomo SB, Van Vught HAJ. Transeutaneous respiratory electromyographic monitoring. Crit Care Med 1987;I5:294-9. 8. Keens TG, Bryan AC, LevisonH, lanuzzo CD. Developmental pattern of muscle fiber types in human ventilatory muscles. J Appl Physiol 1978;44:909-13. 9. Keens TG, Ianuzzo CD. Development of fatigue-resistant muscle fibers in human ventilatory muscles. Am Rev Respir Dis 1979;119:139-41.