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Women who return to work outside of the home while still breastfeeding must often store the expressed milk at less than optimal temperatures. Human milk provides digestive enzymes (amylase and lipase) that compensate in the newborn for immature pancreatic function.We have assessed the stability of amylase and bile salt-dependent lipase after storage for 1-24 h at 15, 25, and 38 degrees C.Both enzymes were stable at 15 and 25 degrees C for 24 h, whereas at 38 degrees C there was a 15 and 20% decrease in lipase and amylase activity, respectively. The stability of milk lipoprotein lipase was also tested. This very labile enzyme was more stable in milk than previously reported for blood and tissues, i.e., 20 and 50% decrease in activity after storage at 15 or 25 degrees C for 24 h, respectively. A two-unit drop in milk pH by 24 h of storage would not affect the activity of digestive enzymes, which are stable at pH3.5.We conclude that milk provides the same compensatory digestive activity after short-term storage, even at relatively high temperature, as when fed fresh to the infant.

The effect of diet, human milk or formula, on gastric function (lipase and pepsin activity, pH, and volume) and intragastric digestion of fat was assessed in 28 appropriate for gestational age preterm infants (gestational age, 28.9 +/- 1.4, 29.1 +/- 0.9, 29.5 +/- 0.6 wk; birth weight, 1.00 +/- 0.14 to 1.18 +/- 0.07 kg). The infants were fed either human milk (n = 11), SMA Super Preemie formula (n = 9), or Similac, Special Care formula (n = 8). Fasting and postprandial activity of digestive enzymes, pH, and gastric volume (measured before or during 50 min after gavage feeding) did not differ as a function of diet among the three groups of infants. Gastric lipase output, 23.1 +/- 5.1, 28.3 +/- 6.6, and 22.5 +/- 6.4 (U/kg of body weight) in human milk-, SMA SP-, or Similac SC-fed infants was comparable to the gastric lipase output of healthy adults fed a high fat diet (22.6 +/- 3.0). Pepsin output was, however, significantly lower (597 +/- 77, 743 +/- 97, and 639 +/- 142 U/kg of body weight) in human milk-, SMA SP-, and Similac SC-fed infants) than in healthy adults (3352 +/- 753 U/kg). The hydrolysis of dietary fat was 1.7-2.5-fold higher (p < 0.01) in human milk-fed infants than in infants fed either formula. We conclude that differences in type of feeding, i.e. different fatty acid profiles (long chain or medium chain triglycerides), different emulsions (natural or artificial), and different fat particle sizes do not affect the level of activity of gastric enzymes. However, the triglyceride within milk fat globules appears to be more accessible to gastric lipase than that within formula fat particles. We suggest that the contribution of gastric lipase to overall fat digestion might be greater in the newborn (a period of pancreatic insufficiency) than in the adult.

The onset of medium-chain fatty acid synthesis in the human mammary gland was investigated. Colostrum and serum were collected from 31 healthy women and the fatty acid composition of total lipid was analyzed by gas-liquid chromatography. Although colostrum/serum ratios for most fatty acids range from 0.7-2.4, very low levels of 10:0 and 12:0 were present in serum lipids as compared to much higher concentrations of these fatty acids in colostrum lipids (colostrum/serum ratio 16.23 and 17.11 for 10:0 and 12:0, respectively). We have previously found that medium-chain fatty acid levels are very low in prepartum mammary secretions (6-10 wk before term delivery) but are higher and similar in colostrum of women who deliver preterm (3-14 wk) or at full term. The data indicate that parturition, irrespective of length of pregnancy, is the trigger for medium-chain fatty acid synthesis in the human mammary gland.

Fat absorption from two different premature infant formulas and one full-term formula containing three different fat blends was investigated in two groups of premature infants. The first group of nine infants (gestational age, 29.1 +/- 0.88 weeks; postnatal age, 3.13 +/- 0.71 weeks) was fed alternately for 1 week each SMA preterm formula containing either high levels (50%) of medium-chain triglycerides (MCT) (6:0, 8:0, and 10:0) or high levels (86%) of long-chain triglycerides (LCT) (greater than or equal to C12). Except for fat blends, the formulas were otherwise identical. The second group of 11 infants (gestational age, 30.5 +/- 0.77 weeks, studied at a postnatal age of 4.33 +/- 0.91 weeks) was fed for 1 week a full-term infant formula, S-26, containing 98% LCT. Fat absorption (studied during a 3-day fat balance period) was similar irrespective of fat blend: 89.08 +/- 2.37% during feeding of preterm SMA, 50% MCT; 87.0 +/- 3.81% during feeding of preterm SMA, 86% LCT; and 83.00 +/- 2.89% during feeding of S-26, 98% LCT. Weight gain (grams per day) and increase in length (centimeters per day) were 23.2 +/- 1.7, 21.20 +/- 1.7, and 14.28 +/- 2.9, and 0.17 +/- 0.06, 0.16 +/- 0.04, and 0.22 +/- 0.07 during feeding of the three fat blends, respectively. Lipase activity levels in fasting gastric aspirates were higher during feeding of the LCT than the MCT formula. The possible stimulation of gastric lipase secretion secondary to long-chain fatty acid stimulation of cholecystokinin secretion might be related to the efficient digestion of formula fat, irrespective of triglyceride-fatty acid chain length.(ABSTRACT TRUNCATED AT 250 WORDS)

Plasma cholesterol and lecithin concentrations are regulated by the serum enzyme lecithin: cholesterol acyltransferase (LCAT). LCAT activity is low in cord blood of premature infants, suggesting that in these infants the hypercholesterolemia associated with Intralipid infusion might be due to low LCAT activity. The serum LCAT activity has not been quantitated in preterm infants receiving intravenous fat emulsions. We have therefore quantitated LCAT activity in eleven premature infants maintained on total parenteral nutrition (TPN). Ten infants were studied during the first 2 weeks after birth; they received daily infusions of Intralipid at a rate of 0.5-2.0 g/kg/day over 15 h. One infant received 3.8 g/kg/day during the second week. In addition to LCAT, serum apoprotein A1 (the cofactor of LCAT), cholesterol, triglycerides, and free fatty acids were quantitated. Blood specimens were taken before the start of the infusion and 15-45 min before its completion. The LCAT activity and apoprotein A1 concentrations remained, respectively, 21-24% and 30-35% of adult levels. However, serum cholesterol levels remained in the normal range during the fat infusion. It remains to be established whether low LCAT activity and apoprotein A1 levels are due to the administration of Intralipid (which lowers LCAT activity in rats), to the lack of enteral feedings, or to prematurity per se. Our data suggest that administration of Intralipid at a rate not exceeding 1-2 g/kg/day does not impair the clearing of Intralipid-lecithin and the metabolism of cholesterol.

The effect of diet, usual (44 +/- 4% energy as fat), high-fat (49 +/- 4% energy as fat), and moderate-fat (33 +/- 2% energy as fat), on gastric function (lipase and pepsin activities, pH, emptying rate) and intragastric digestion of fat were assessed in six children with cystic fibrosis. Fasting and postprandial activity of digestive enzymes, gastric pH, and gastric volume measured before, during, and after 120 min of feeding did not differ significantly as a function of fat intake. Postprandial gastric lipase output (units per kilogram of body weight) during usual, moderate-fat, and high-fat diets was close to or higher than (38.8 +/- 7.2, 44.9 +/- 8.6, and 54.8 +/- 5.5 U/kg per 20 min) gastric lipase output of premature infants (22.5 +/- 6.4 to 28.3 +/- 6.6 U/kg per 20 min) or of healthy adults (5.4 +/- 0.4 U/kg per 15 min) fed a high-fat diet. Postprandial pepsin output was higher (4749 +/- 797, 6117 +/- 925, and 5444 +/- 819 U/kg per 20 min) than in premature infants (597 +/- 77 to 743 +/- 97 U/kg per 20 min) or healthy adults (781 +/- 56 U/kg per 15 min). Eighty minutes after feeding gastric lipolysis reached 20 to 36%. This study shows that gastric lipase activity is high in cystic fibrosis patients maintained on diets providing 32% to 49% energy as fat, and that gastric lipase level did not increase over the ranges of dietary fat intake tested.

Long-chain polyunsaturated fatty acids (LC-PUFA) accretion (essential for growth and neural development) was studied from late fetal throughout weaning age in the ferret, a species with maternal LC-PUFA sufficiency during pregnancy and lactation. The data show that a) accretion rate of LC-PUFA is rapid during early postnatal development, b) milk LC-PUFA decrease during lactation, c) adipose tissur, LC-PUFA level is directly related to milk LC-PUFA level, while accretion in brair and liver exceeds dietary intake, d) accretion of arachidonic acid occurs earlier than docosahexaenoic acid, suggesting earlier development of n6-fatty acid endogenous synthesis.

Gastric proteolysis is assumed to be low in the newborn (Britton & Koldovsky 1989). Postprandial pepsin output is significantly lower in preterm infants than adults, 589 vs. 3352U/kg, respectively (Armand et al. 1995, 1996). We now report on gastric proteolysis in preterm infants (gestation age, 29 weeks; postnatal age, 5-6 weeks) gavage-fed mother’s milk or preemie formula. The data show that a) the nonprotein component is higher in human milk than formula, b) net proteolysis amounts to 15% of protein, c) gastric proteolysis is lower than lipolysis and, contrary to the latter, is not enhanced by milk feeding (Armand et al. 1996). We suggest that stomach pH, enzyme output, and food structure are the reasons for differences in gastric digestion of protein and fat in infants.

Human milk contains many components that protect the newborn against infection at a time when the infant's own defense mechanisms are poorly developed. Fat is one of the major nutrients in human milk. The fat is contained within milk fat globules composed of a core of triglyceride and a membrane consisting of phospholipids, cholesterol, proteins, and glycoproteins. Both the membrane and the core components can provide protection against microorganisms. The major protective membrane glycoproteins, mucin, and lactadherin are resistant to conditions in the newborn's stomach and maintain their structure and function even at low pH and in the presence of the proteolytic enzyme pepsin. The core triglycerides upon hydrolysis by digestive lipases (especially gastric lipase, which is well developed in the newborn) produce free fatty acids and monoglycerides, amphiphylic substances able to lyse enveloped viruses, bacteria, and protozoa. Therefore, in addition to its nutritional value, the fat in human milk has a major protective function.

Human milk fat globule (HMFG) glycoproteins can prevent infections by microorganisms in breast-fed infants; the MUC-1 mucin inhibits binding of S-fimbriated Escherichia coli to buccal mucosa, and lactadherin may prevent symptomatic rotavirus infections. In this study, the survival of these HMFG glycoproteins in the stomach of human milk-fed preterm infants (gestational age = 27.5 +/- 0.4 wk) was assessed, and levels in their mothers' milk determined, using specific RIAs. Butyrophilin, a major component of HMFG membrane that has no demonstrated antimicrobial activity, was studied for comparison. The levels of mucin, lactadherin, and butyrophilin in 41 milk samples of 20 mothers were 729 +/- 75, 93 +/- 10, and 41 +/- 3 microg/mL, respectively. Mucin and lactadherin were significantly higher in early milk samples (15 d postpartum) than in later milk samples (15-90 d postpartum), whereas butyrophilin showed no such difference. Significant amounts of mucin and lactadherin were found in almost all gastric aspirates of human milk-fed infants, even 4 h after feeding (mucin, 270 +/- 30 microg/mL; lactadherin, 23.2 +/- 4.4 microg/mL), whereas butyrophilin was rapidly degraded in the majority of aspirates. Western blot analysis demonstrated that the immunoreactive mucin, lactadherin, and butyrophilin in the milk-fed gastric aspirates had the expected native molecular weights. Mucin and lactadherin survived at all gastric pH values, whereas butyrophilin was found only at pH4. Neither lactadherin nor butyrophilin were detected in gastric aspirates of formula-fed infants (gestational age = 27.8 +/- 0.5 wk), whereas the very low level of mucin (9.1 +/- 1.1 microg/mL) in this group is presumably cross-reacting gastric mucin. These results demonstrate that two HMFG glycoproteins implicated in prevention of infection, MUC-1 mucin and lactadherin, survive and maintain their integrity in the stomachs of human milk-fed preterm infants.

Background Gastric lipase has an important compensatory function in neonatal fat digestion. The activity level of pepsin and its role in protein digestion is less well understood. We have, therefore, studied the ontogeny of lipase and pepsin in the ferret, a species with a neonatal fat digestion pattern similar to that of humans. Methods Gastric lipase and pepsin activities were quantified from the late fetal period throughout lactation, and were compared with those of the adult. Results The data show earlier ontogeny and much more rapid rise of lipase activity than of pepsin. Lipase activity was present during the last week of fetal development, whereas pepsin was detected only postnatally. Lipase activity was 72.8% +/- 14.2% and 153% +/- 9.95% and pepsin activity was 11.6% +/- 1.3% and 30.1% +/- 1.3% of the adult level at 2 and 4 wk of age, respectively. Conclusions We conclude that lipase activity develops early and exceeds adult activity during the suckling period, when fat intake is very high. The low pepsin activity and high postprandial pH probably limit gastric proteolysis, thereby contributing to the structural and functional stability of milk proteins, many with protective or bioactive function in the gastrointestinal tract of the newborn.

The amount of mRNA hybridizing to bile salt-dependent lipase and to colipase-dependent lipase probes as well as their translation into active proteins were quantified in the adult and newborn pancreas and lactating mammary gland from the ferret, a species whose milk, similar to that of the human, has bile salt-dependent lipase. The concentration of colipase-dependent lipase mRNA correlated with the amount of activity found in the adult and newborn pancreas, whereas neither mRNA nor activity of this enzyme was detected in the kit pancreas or in the lactating mammary gland. These data indicate that colipase-dependent lipase is actually expressed in adult pancreas and might represent the main lipolytic system in the adult. mRNA hybridizing to the bile salt-dependent lipase probe used in this study were detected in adult and in newborn ferret pancreas as well as in lactating mammary gland. However, the bile salt-dependent lipase activity expressed in the newborn pancreas was very low when compared with the activity expressed either in the mammary gland or in the adult pancreas. These data argue for a compensatory role of milk bile salt-dependent lipase in lipid digestion in the newborn. The hydrolysis of dietary fat might be initiated by preduodenal lipase, the activity of which is only two times lower in the gastric mucosa of the newborn than in the adult ferret. The high concentration of mRNA hybridizing to the bile salt-dependent lipase probe associated with a very poor bile salt-dependent lipase activity and protein suggests either that these mRNA are very unstable or that they are poorly translated into an active pancreatic bile salt-dependent lipase.

The aim of this study was to determine whether the amount of dietary fat modulates the activity of gastric lipase in humans. Gastric juice was collected from six healthy subjects after 2-wk periods of either a high-fat (50% of energy as fat) or low-fat (25% of energy as fat) diet. The collection period lasted 2 h, the first hour under baseline conditions and the second hour after pentagastrin stimulation (6 μg/kg body wt). Gastric lipase and pepsin activities were quantitated at 15-min intervals and total enzyme outputs were calculated. Under baseline conditions there was a tendency for higher output of gastric lipase and pepsin after the high-fat diet than after the low-fat diet (gastric lipase : 745 compared with 446 U/h, pepsin : 107 677 compared with 78 505 U/h). The difference in output between diet groups was significant after pentagastrin stimulation (gastric lipase : 1323 compared with 875 U/h, pepsin : 191 751 compared with 128 961 U/h, for high-fat compared with low-fat diet, respectively, P < 0.05). This study is the first to report that a high-fat diet leads to an increase in the activity of gastric enzymes in humans. Am J Clin Nutr 1995 ; 62 :74-80.

In this chapter we will discuss gastric lipase and, briefly, other preduodenal lipases with similar function. The emphasis will be on the function of these lipases in the process of fat digestion in health and disease.

The aim of our study was to determine the role of excitatory amino acids in controlling cardiorespiratory activity. For this purpose we administered an antagonist of both N-methyl-D-aspartate (NMDA) and non-NMDA receptors (kynurenic acid), and an antagonist of the NMDA receptor complex (dizocilpine, more commonly known as MK-801) i.v. to chloralose-anesthetized cats while monitoring tracheal air flow, tidal volume, respiratory rate, inspiratory and expiratory durations, end tidal CO2, arterial blood pressure and heart rate. Administration of kynurenic acid in doses of 350 and 500 mg/kg produced respiratory depression as reflected by decreases in respiratory minute volume and increases in end tidal CO2. Inspiratory duration was increased with both doses and apnea (occurring during expiration) was observed with the high dose. Apnea was preceded by an apneustic pattern of breathing. Both doses resulted in an increase in blood pressure and, with the high dose, a later decrease in blood pressure was noted. Dizocilpine in doses ranging from 0.03 to 1 mg/kg produced dose-related decreases in respiratory minute volume, and increases in end tidal CO2. In addition, dizocilpine produced increases in inspiratory duration, an apneustic pattern of breathing and apnea (occurring during inspiration). Effects on blood pressure were similar to those observed with kynurenic acid. It is concluded that blockade of excitatory amino acid receptors results in pronounced effects on cardiorespiratoty activity.

LCPUFA are essential for growth and functional development. We have compared LCPUFA accretion from the late fetal throughout the nursing period in ferret kits with LCPUFA of maternal organs and milk. Table

Gastric Proteolysis in the Preterm Infant: Protein Digestion is Limited and Is Not Affected by Diet, Human Milk or Formula † 580

Most nutrients in human milk have multiple functions, among them the protection of the infant from invading micro-organisms (1). This symposium will address the topic of immune protection, i.e. specific antibody-mediated protection against micro-organisms and the much broader non specific protection which, contrary to the former, is independent of prior maternal exposure to micro-organisms. In this category there are milk components that act as analogs or homologues to host cell surface pathogen receptors(glycoproteins, gangliosides, proteins) have specific bactericidal and antiviral activity (lactoferrin) or are able to lyse micro-organisms(lysozyme, fatty acids and monoglycerides) (2). Lipid conjugates such as gangliosides in the milk fat globule membrane inhibit the action of the heat stable enterotoxins of V-cholerae and E coli. Milk lipids acquire the potential to lyse micro-organisms after their partial digestion to free fatty acids and monoglycerides (3). The fat contained in the core of milk fat globules is more accessible to gastric lipolysis than the fat in infant formula. Indeed hydrolysis of dietary fat is 1.7 to 2.5 fold higher in human milk-fed than in formula-fed infants (4). The free fatty acids and monoglycerides released cause rapid lysis of enveloped viruses, protozoa and certain bacteria. Highest lytic activity is associated with lauric and (C 12:0) and monolaurin and with long chain unsaturated fatty acids (C 18:2). Storage of milk, even at -20 C, releases large amounts of free fatty acids(5),that might lower viral and bacterial counts during frozen storage of human milk. The milk fat globule is an important protective factor through its core triglyceride (after lipolysis) and through the globule membrane gangliosides and glycoproteins (6).

LONG CHAIN POLYUNSATURATED FATTY ACIDS (LC-PUFA) DURING EARLY DEVELOPMENT: RELATIONSHIP BETWEEN MATERNAL MILK AND NEONATAL ADIPOSE TISSUE AND BRAIN. † 1407

ONTOGENY OF GASTRIC DIGESTIVE FUNCTION: DIFFERENT PATTERN OF LIPASE AND PEPSIN DEVELOPMENT. † 703

Cholecystokinin (1–300 ng) was administered into the lateral brain ventricle of chloralose-anesthetized cats while monitoring tracheal airflow, arterial blood pressure, and heart rate. Dose-related increases in respiratory activity occurred in each animal tested, and were due to an increase in tidal volume. When 300–1000 ng of cholecystokinin was administered intravenously, no respiratory stimulant effect was observed. These results indicate that cholecystokinin acts in the brain to stimulate respiration.

Lipolytic activity was studied in gastric aspirates of 13 premature infants of birth weight 1,050 to 1,786 gm. All infants received a diet of infant formula fed by gastric tube. Gastric aspirates were collected after irrigating the stomach with 2 to 5 ml sterile saline before regular feeding. Lipolytic activity, tested with doubly labeled 3 H glyceryl- 14 C tripalmitin substrate, was 55.6±11.7 n mol/min/ml (range 4.2 to 140). The lipolytic activity had a pH optimum of 5.4 and produced partial glycerides (mono and diglycerides), glycerol, and free fatty acids. Lipolysis was inhibited by bile salts. Our findings show that in premature infants, as in adults, digestion of dietary fat starts in the stomach. Since bile salt concentrations are low in premature infants, the amphiphilic reaction products formed (monoglyceride and FFA) could play a significant role in the stabilization of lipid emulsions.

Dexamethasone was administered by continuous subcutaneous infusions (16 microgram/kg/h) to pregnant rats from day 16 of gestation. Administration of the hormone markedly affected maternal and fetal weight gain, fetal lung:body weight ratio and lipoprotein lipase activity of the lung. Cumulative maternal weight gain from days 15-21 of gestation was 80 +/- 4.0 g in control and 30 +/- 10 g in dexamethasone-treated rats. Fetal weight at 22 days of gestation and 1 day after birth was 5.5 +/- 0.39 and 8.6 +/- 0.30 in control and 4.65 +/- 0.26 and 5.9 +/- 0.34 in dexamethasone-treated rats. The ratio of lung weight to body weight was lower throughout the last 5 days of gestation in dexamethasone-treated than in control rats. Dexamethasone administration led to a 2- to 3-fold increase in lipoprotein lipase activity levels in fetal rat lung at 19 and 20 days' gestation and prevented the decline in enzyme activity shortly before birth. Stimulation of fetal lung lipoprotein lipase activity suggests that increased uptake of triglyceride-fatty acids by the lung could be a contributory factor to corticosteroid-enhanced surfactant synthesis.

Milk was collected from mothers of 18 very premature (26 to 30 wk gestation age), 28 premature (31 to 36 wk), and six term (37 + wk) infants on day 2 to 3 (colostrum), and at 1, 3, 6, and 12 wk postpartum. Fat content for 154 milk samples was 2.80 g/dl gravimetrically and 2.66 g/dl by quantitative thin-layer chromatography. Fat content increased during lactation, whereas phospholipids and cholesterol declined. Concentrations of medium-chain fatty acids increased from colostrum to mature milk and were highest in preterm milk. Compensatory decreases were observed in very premature and premature oleic acid. Long-chain polyunsaturated fatty acids were highest in colostrum and reduced in mature milk. Long-chain polyunsaturated fatty acids were also higher in very premature and premature milk than in term milk. These elevated levels of readily absorbed medium-chain fatty acids and long-chain polyunsaturated fatty acids in preterm milk may be of special benefit for the needs of premature infants.

The effect of smoking three cigarettes in rapid succession on maximal and partial expiratory flow rates was studied in ten healthy smokers. The mean decrease in maximal midexpiratory flow and partial midexpiratory and end-expiratory flow was statistically significant. The response was maximal after the first cigarette. These results suggest that the instantaneous midexpiratory flow after 50 percent of the forced vital capacity has been exhaled is a useful indicator of irritation of the airways. Reduction of partial expiratory flow rates was three times greater than reduction of maximal expiratory flow rates. We suggest the use of partial expiratory flow curves as a screening test for irritants of the airways.

We have used lung organ cultures to investigate the effects of prolactin on the synthesis of surfactant phospholipids in the fetal lung. Lung explants prepared from 18-day-old rat fetuses were maintained in culture for 48 h in serum-free medium with two different preparations of prolactin (2 μg/ml —NIH, ovine lot P-S-13 and Sigma, lot 19C-0396). In addition, lung explants were cultured for an initial 24 h with dexamethasone (10−6 M) followed by 24 h of culture with prolactin. Incorporation of surfactant precursors [methyl-3H]choline and [U-14C]glycerol, was measured after incubation for 4 h in the absence of hormones. Exposure of the explants to prolactin, NIH or Sigma, for 48 h led to a significant increase in the levels of total phospholipid −41 and 58% (P < 0.001), respectively. Culture of the lung expiants for the first 24 h with dexamethasone followed by an additional 24 h exposure to prolactin, NIH or Sigma, led to a further increase in the level of total phospholipid, 105 and 112% (P < 0.001) and disaturated PC, 260 and 300% (P < 0.001), respectively. Exposure of lung expiants to prolactin led to a significantly higher incorporation of [methyl-3H]choline into total phospholipid and disaturated PC, 32–35 and 42–45% (P < 0.001), respectively. [U-14C]Glycerol incorporation was significantly higher after culture in prolactin containing medium: 30 and 40% higher into total phospholipid (P < 0.001) and 77 and 105% (P < 0.001) higher into phosphatidylglycerol — the second largest component of lung surfactant. Culture for 24 h with dexamethasone, followed by an additional 24 h with prolactin, led to a further increase in the incorporation of labeled choline and glycerol into surfactant phospholipids: choline incorporation into phospholipid and disaturated PC was 63 and 67% (P < 0.001) and 71 and 75% (P < 0.001), respectively, higher than control; glycerol incorporation was 60 and 72% (P < 0.001) and 192 and 322% (P < 0.001) higher into total phospholipid and phosphatidylglycerol, respectively, than controls. We conclude that: 1, Prolactin acts directly on the fetal rat lung by stimulating the synthesis of surfactant phospholipids. 2, Enhancement of the prolactin effect by dexamethasone is suggested by a further increase in lung phospholipid and disaturated PC levels as well as in the incorporation of surfactant precursors in the presence of both hormones.

Ten to 30% of dietary fat is hydrolyzed in the stomach by lingual lipase, an enzyme secreted from lingual serous glands. We investigated the substrate specificity of this enzyme as well as the potential of lingual lipase to act in the upper small intestine i.e., in the presence of bile salts and lecithin. The data presented show that partially purified preparations of rat lingual lipase and the lipase in gastric aspirates of newborn infants have identical substrate specificity: medium-chain triglycerides were hydrolyzed at rates 5-8-fold higher than long-chain triglycerides; the rat and human enzymes do not hydrolyze the ester bond of lecithin or cholesteryl-ester. In contrast to pancreatic lipase, the hydrolysis of triglycerides by lingual lipase is not inhibited by lecithin. But, similar to pancreatic lipase the activity of lingual lipase is inhibited by bile salts, the extent of inhibition varying with its nature and concentration. This inactivation is not prevented by colipase but is partially averted by lipids and protein, suggesting that lingual lipase can remain active in the duodenum. The pH optimum of the enzyme (2.2-6.5 in the rat and 3.5-6.0 in human gastric aspirates) is compatible with continued activity in the upper small intestine, especially during the neonatal period, when the luminal pH is under 6.5. The marked variation in lipase activity levels in gastric aspirates of newborn infants is probably due to individual variations in enzyme amounts. The characteristics of the lipase are however identical in infants with low, intermediate or high activity levels.(ABSTRACT TRUNCATED AT 250 WORDS)

The aim of this study was to determine the range of activity and the location of lipase in the human stomach. The range of lipase activity in gastric mucosa of surgical specimens from the fundic area of 22 subjects was 594 to 3350 mU [mean, 1598 +/- 144 mU tri[3H]olein, (1 mU-1 nmol [3H]oleic acid released from tri[3H]olein per minute per milligram protein)]. For localization of activity, pinch biopsy specimens of gastric mucosa from 6 subjects were taken from the greater and lesser curvatures within 2 cm of the gastroesophageal junction (upper greater curvature and upper lesser curvature) and within 2 cm of the pylorus (lower greater curvature and lower lesser curvature). Lipase activity was higher in the upper greater curvature (405 +/- 92 mU) than in the upper lesser curvature (32 +/- 13 mU) and lowest in the antral area (16 +/- 9 mU in the lower lesser curvature and 10 +/- 2 mU in the lower greater curvature). The data show that in the human, lipase activity is localized primarily in the fundic area of the stomach. Comparison of the lipase activity levels in the gastric mucosa with lingual lipase activity levels in specimens of lingual serous glands indicates that in humans, gastric lipase is the main lipase active in the stomach.

We have measured the hydrolysis of tri-, di- and monoacylglycerols and of phosphatidylcholine in the lung and have quantitated the incorporation of glyceride and phospholipid fatty acids into lung lipids. Lipolytic activity was studied in the isolated, ventilated, rat lung perfused for 100 min in a recycling system with Krebs-Ringer bicarbonate buffer containing 3% bovine serum albumin, 5.6 mM glucose, and the lipid emulsions. Triacylglycerols were hydrolyzed at significantly (P less than 0.05) lower rates than partial acylglycerols. Hydrolysis of di- and triacylglycerols was significantly lower in lungs of animals given i.v. heparin 60 min prior to death. The incorporation of fatty acids released from acylglycerols was greater into lung neutral fats than into lung phospholipids. Phosphatidylcholine was also hydrolyzed in the isolated perfused lung. The data show that (a) neutral glycerides and phospholipids are hydrolyzed in the isolated, ventilated, perfused rat lung; (b) the hydrolysis of tri- and diacylglycerols is markedly inhibited in lungs isolated from rats given heparin (100 U/kg) 1 h prior to death, whereas the hydrolysis of monoacylglycerol is unaffected by heparin administration, suggesting that tri- and diacylglycerols are hydrolyzed by endothelial lipoprotein lipase, whereas hydrolysis of monoacylglycerol is catalyzed by a separate monoacylglycerol lipase; (c) free fatty acids released by lipoprotein lipase are used by the lung for metabolic needs.

The aim of this study was to evaluate the cardiorespiratory effects of intravenously administered gamma-aminobutyric acid (GABA) alpha-(4,5,6,7-tetrahydroisoxazolo[5,4-c]pyridin-3-ol, THIP) and beta-(baclofen) receptor agonists and to locate the site of action of these drugs in the brain. THIP and baclofen were administered to alpha-chloralose-anesthetized cats while minute ventilation (VE), arterial blood pressure (AP), and heart rate were monitored. THIP, in doses of 0.5 to 2 mg/kg decreased VE, tidal volume (VT), and AP. No changes in respiratory rate (f) or inspiratory (TI) or expiratory (TE) duration were observed. Baclofen, in doses of 0.5 to 4 mg/kg, decreased VE, f, and AP. VT and TI increased and an “apneustic” breathing pattern was seen. THIP (9.5 micrograms), applied bilaterally to the glycine-sensitive area of the ventral medulla, reproduced the effects seen with intravenous administration. Application of 10 micrograms of bicuculline bilaterally to this area reversed the effects of intravenous THIP but not those of baclofen. Baclofen (5.6-56 micrograms), administered by the intracisternal route, produced the same respiratory effects seen with intravenous administration. We conclude that activation of GABA alpha- and beta-receptors produces cardiorespiratory depression. However, this is accomplished by different mechanisms and by actions exerted at different central nervous system sites.

The first step in fat digestion occurs in the stomach, where 10-30% of dietary triacylglycerols are hydrolyzed to partial (di- and mono-) acylglycerols and free fatty acids. Preduodenal fat digestion is an important compensatory mechanism in the newborn because of immature pancreatic (lipase) and hepatic (bile acid synthesis) function. Since hydrolysis of fat in the stomach can be catalyzed by enzymes of lingual (Hamosh, M. (1979) Pediatr. Res. 13, 615-622) and possibly gastric origin, we have studied the developmental pattern and quantitative contribution of these two enzymes to intragastric fat digestion by measuring lipase activity in homogenates of lingual glands and gastric mucosa of rats from birth until 60 days of age. Total lipolytic activity in rat gastric mucosa was only 2-10% of that in the lingual glands throughout the entire period studied. Lingual lipase activity increased steadily from birth until day 50, whereas the activity in the gastric mucosa reached peak levels at 17-20 days and declined sharply after weaning. Throughout the period of study--suckling, weaning, and young adulthood--lingual and gastric lipase had very similar characteristics: pH optimum in the range of 5.0-6.0 and 2.5-5.0-fold higher activity on medium-chain (tri[14C]octanoin) than long-chain (tri[3H]olein) triacylglycerols. In the lingual glands, lipase activity was higher during fasting, probably because of accumulation of enzyme (without depletion during meals), whereas in the gastric mucosa lipase levels were higher after feeding, suggesting adsorption of lingual lipase (which reaches the stomach with the ingested food) onto the gastric mucosa. From birth to weaning, there was rapid and extensive hydrolysis of triacylglycerol in the stomach (decrease from 98 mol% in rat milk to 33.6-48.9 mol% in the stomach contents half an hour after feeding). The intragastric lipolysis remained almost constant from birth until day 20, in spite of a marked increase in food consumption, probably because of the continued rise of lingual lipase levels. The direct relationship between high intragastric lipolysis and high lingual lipase activity suggests that lingual lipase is the major digestive enzyme in the newborn.

Phospholipids were determined in milk on postpartum day 3 (colostrum) and days 7, 21, 42, and 84 from mothers of 18 very premature (26 to 30 wk gestation age), 28 premature (31 to 36 wk), and 6 term (37 to 40 wk) infants. Lipids were analyzed by thin-layer and gas-liquid chromatography. Total fat content increased during lactation whereas phospholipids and cholesterol declined. Phospholipids were separated from neutral lipids by column chromatography and distributed by preparative thin-layer chromatography into classes, sphingomyelin, phosphatidyl choline, serine, inositol, and ethanolamine for fatty acid analysis. Phospholipids exhibited a remarkable constancy in class percentages in milks from mothers giving birth prematurely or at term. Changes were observed in fatty acid composition within each of the phospholipid classes as secretion progressed from colostrum (3d) to transitional (7d) to mature milk (21, 42, 84d). These changes in phospholipid fatty acid composition occurred only during the first 3 wk of lactation. Mature milk was found to be relatively constant in phospholipid composition.

The frequent inclusion of heparin in fluids used for total parenteral nutrition in infants, prompted an investigation of the ability of heparin to release lipoprotein lipase (LPL) and hepatic lipase (HL) from the endothelial surface into the circulation, and of the effect of heparin on tissue stores of lipase in the postnatal period. In rat pups, plasma postheparin lipolytic activity (PHLA) released by IP administration of heparin (0.5 unit/g body wt) was 15% of adult values at birth and increased rapidly to reach 60% on day 10. Repeated doses of heparin (in adult rats, given 0.1 unit/g IV) at 1 and 4 h after the initial dose did not affect the maximal response to heparin. In all age groups 80% of PHLA was inhibited by 0.5 M NaCl, suggesting a mostly nonhepatic origin for the released enzyme. Heart, lung, and liver lipase activities of rat pups were not significantly different from controls not given heparin. The pattern of change in tissue enzyme content was similar for heart and lung, but different from hepatic lipase. LPL activity in the former increased from 10 and 30% to 60 and 100% of adult values between birth and 10 days while in the latter enzyme activity exceeded adult levels at birth and decreased to 50% of adult values during the latter half of the suckling period (days 10-21). Our results demonstrate that heparin does not cause depletion of tissue lipases in the postnatal period. The parallel increases in LPL content of peripheral tissues and PHLA suggest that in all age groups heparin-induced release of LPL into the circulation is proportional to tissue lipolytic activity.

We previously demonstrated that GABA and muscimol administered either into the cisterna magna or the fourth ventricle to chloralose-anesthetized cats cause respiratory depression, hypotension, and bradycardia. Injection of these substances into the lateral and third ventricles had no effect. In order to localize the site of action, muscimol and GABA were applied by Perspex rings to the ventral surface of the medulla. Application of muscimol (0.25-2.66 micrograms) to Schlaefke's area in 6 cats reduced minute ventilation from 443 +/- 38 to 291 +/- 52 ml/min by reducing tidal volume from 31.8 +/- 2.3 to 17.6 +/- 1.4 ml, without changing respiratory rate and duration of inspiration. Hypotension and bradycardia were also observed. Application of GABA (0.14-4.86 mg) produced similar effects on respiratory activity and arterial blood pressure. No significant effects occurred when high doses of these agents were applied to Loeschcke's and Mitchell's areas. Application of bicuculline (5-25 micrograms) to Schlaefke's area had the opposite effect of muscimol and GABA on respiratory activity and blood pressure, and reversed the respiratory and cardiovascular depressant effects of both agents. We conclude that GABA receptors are present at Schlaefke's area, and that activation of these receptors results in respiratory depression, hypotension, and bradycardia. Our results suggest that GABA may be an important inhibitory neurotransmitter in the modulation of respiratory and cardiovascular control.

Summary: Lipoprotein lipase activity in lung and heart was studied in fetal (17-22 days of gestation) and newborn rats from the day of birth until 30 days of age. Enzyme activity in epididymal, omental, and parametrial adipose tissue was tested after 18 days of age. Blood triglyceride levels were measured at all ages from 17 days in utero until 30 days after birth. The developmental pattern of lipoprotein lipase differed markedly in lung and heart. Although lipoprotein lipase activity was 4 to 5 times higher hi adult rat heart than in lung (30-40 U/g in heart vs. 8-11 U/g in lung), the activity was almost completely absent from fetal heart and was very low during the first 3 days after birth. Lipoprotein lipase reached 60-70% of adult activity at 6 days and remained at that level until 19 days after birth; adult activity levels were reached at 24 days. In the lung, contrary to the heart, lipoprotein lipase activity was high in the fetus (84% of adult activity), decreased immediately after birth to 45% of adult activity at 2 days, and remained at that level up to 15 days after birth. Enzyme activity started to rise again at 15 days and reached adult levels at 21 days of age. Adipose tissue was present in trace amounts before the age of 2 weeks. In the three fat depots tested, lipoprotein lipase activity was 50% lower than in adults between 20 and 30 days after birth. Blood triglyceride levels increased 4- fold between 2 to 10 hr after birth and remained elevated during the first 3 days after birth. Speculation: The triglyceridemia that starts in the immediate postnatal period and lasts for several days in the rat probably results from the combination of high fat intake and low clearing ability of the extrahepatic tissues. In addition to its role in clearing circulating triglyceride, lipoprotein lipase may play a role in the growth and maturation of individual organs. The present study shows that lipoprotein lipase activity is high in fetal lung during the period of marked surfactant synthesis and suggests that circulating triglyceride-fatty acids are used by the fetal lung for surfactant synthesis.

Fatty acid composition of the major lipid classes in stomach contents of suckling rats at 1, 5, 10, 17 and 20 days of lactation was compared to that of milk lipids. In milk, 98% of fatty acids were in triacylglycerols at all lactation times. Medium-chain fatty acid concentrations increased from 8% in colostrum to 26% at day 5. Fatty acid composition of stomach acylglycerols at all lactation times was different from that of milk triacylglycerols, containing less medium-chain fatty acids, 8:0 and 10:0. This preferential hydrolysis was also shown by higher concentrations of medium-chain fatty acids in the free fatty acid fraction. The lipolysis of medium-chain fatty acids from triacylglycerols resulted in the appearance of di- and monoacylglycerols with 50–100% higher amounts of 14:0 and 16:0. The similar fatty acid composition of products suggests that considerable lipolysis occurred in stomachs of suckling rats even at 1 day of age. Although there was a 10-fold increase in milk consumption, the extent of lipolysis was similar throughout the suckling period because of a parallel rise in lingual lipase levels.

The source of the lipase(s) acting in the stomach was investigated in five animal species: rat, mouse (rodents), rabbit (lagomorphs), guinea pig (caviidae), baboon and human (primates). The activity of lingual and gastric lipases was quantitated in homogenates of lingual serous glands and of gastric mucosa, respectively, by the hydrolysis of tri[3H]oleylglycerol and is expressed in units/g (1 U = 1 mumol [3H]oleic acid released/min) per g tissue wet weight, mean +/- S.E. There were marked differences in the activity level of lingual and gastric lipases among species: mouse and rat had high levels of lingual lipase activity (250 +/- 20 and 824 +/- 224 U/g) and only traces of gastric lipase activity (4.5 +/- 0.9 and 0.04 U/g, respectively), whereas rabbit and guinea pig had no lingual lipase activity and only gastric lipase activity (78 +/- 48 and 27 +/- 7.4 U/g, respectively). In the baboon and human, gastric lipase was the predominant enzyme (109 +/- 20 U/g and 118 +/- 8.8 U/g, respectively), whereas lingual lipase activity was present in trace amounts only (0.04 U/g and 0.3 U/g, respectively). In addition to species differences in the origin of the preduodenal lipases, there were also species differences in the distribution of gastric lipase in the stomach. Thus, while in the rabbit, gastric lipase was localized exclusively in the cardia and body of the stomach, it was diffusely distributed in the entire stomach of the guinea pig and baboon. A comparison between the level of activity of lipase and pepsin (the two chief digestive enzymes secreted by the stomach), showed differences in their localization in the species studied. The difference in source (tongue vs. stomach) and site (cardia-body vs. entire stomach) of lipase secretion must be taken into account in future studies of these digestive enzymes. Although the exact contribution of lingual and gastric lipases individually to fat digestion in species which contain both enzymes cannot yet be evaluated, the markedly higher levels of gastric lipase activity in the baboon and human suggests that, in primates, gastric lipase is probably the major non-pancreatic digestive lipase.

We measured lipoprotein lipase activity in dried defatted preparations of rat lung using doubly labeled chylomicron triglyceride as substrate. The enzyme activity was linear for the first hour of incubation at 37° C, had a pH optimum of 8.1 and was completely inhibited by 0.5 M NaCl. Lungs from fed rats hydrolyzed chylomicron triglyceride at a rate of 13.00 μmoles/g per h; the activity rate was unchanged by fasting 8–72 h. Heparin infusion into isolated lungs caused immediate release of lipoprotein lipase to the venous effluent. The activity released was equivalent to about 10% of total lung lipoprotein lipase activity in both fed and fasted rats. Since the ability to remove blood triglyceride is directly related to the level of lipoprotein lipase activity, these findings indicate that the lung is one of the few tissues able to remove efficiently blood triglyceride during fasting.

We measured and compared the effect of smoking a filtered and a nonfiltered cigarette on instantaneous maximal expiratory flow rates ( and ) and airway resistance (Raw). We found a significant increase in Raw after both cigarettes, and a small decrease in after smoking the nonfiltered cigarette. The change in after smoking the filtered cigarette was not significant. We suggest, on the basis of these data, that filtered cigarette smoke reduces some of the constituents that cause bronchoconstriction in the large and central airways.

The purpose of our study was to identify central nervous system sites involved in the respiratory depressant effect of drugs that stimulate opioid receptors. Diacetylmorphine (heroin) was administered into several cerebroventricular regions of chloralose-anesthetized cats, while monitoring pulmonary ventilation with a Fleisch pneumotachograph. Administration of heroin (17, 50, 150, and 450 micrograms) into the forebrain ventricles, which was restricted to these ventricles, resulted in no significant respiratory effects. In contrast, administration of heroin into either the fourth ventricle or the cisterna magna resulted in a significant (P less than 0.05) decrease in respiratory minute volume (VE). In the fourth ventricle this was because of a decrease in frequency (f) and in the cisterna magna, to a decrease in tidal volume (VT). Intravenous administration of heroin in the same dose-range produced a decrease in VE, which was primarily due to a decrease in f. Bilateral application of heroin (70 micrograms/side) to each of three ventral medullary surface sites (Mitchell's, Schlaefke's, and Loeschcke's areas) known to influence respiration elicited a decrease in VE only at Mitchell's area. This decrease was due to decreases in f and VT. The role of this site in the action of intravenously administered heroin was tested by topical application of naloxone to this area in animals with respiratory depression evoked by intravenous heroin. Bilateral application of naloxone (15 micrograms/side) to Mitchell's area restored breathing to normal. These results lead us to suggest that the site of heroin-induced respiratory depression is a specific area (Mitchell's area) on the ventral surface of the medulla.

Human milk samples were divided at collection and stored at -70 degrees C or -20 degrees C, or extracted immediately with organic solvent, to compare lipid class composition. Storage at -20 degrees C was not satisfactory for maintaining milk lipid composition, for it resulted in hydrolysis of triglycerides and the appearance of free fatty acids.

Summary Plasma lipolytic activity (lipoprotcin lipase and hepatic lipase). free fatty acids (FFA). triglyceridcs. cholesterol, and glucose levels were measured in 21 premature infants [gestational age 26–37 weeks (mean ± SEM 30.4 ± 0.63 weeks), aged 1–8 days (mean ± SEM 3.00 ± 0.35 days)]. All infants were maintained on total parenteral nutrition with heparin (1 U/ml) and were given Intralipid, 1. 2, and 3 g/kg/day. over 15 h on days 1, 2, and 3. respectively. Blood samples were drawn before and at the end of Intralipid administration. Baseline plasma lipolytic activity, before the start of lipid infusion, was 1.54 ± 0.24 U/ml (1 U = I μmol [3H]oleic acid released from tri[3H]olein/h). Lipolytic activity increased after lipid infusion to 4.04 ± 0.96, 4.32 ± 0.63. and 6.09 ± 1.00 U/ml on days 1, 2, and 3 of the study. Hepatic lipase amounted to 38–47% of total lipolytic activity. During the 3 days of lipid infusion, there were dose-dependent increases in plasma FFA, triglyceride, and cholesterol. Whereas FFA and triglyceride concentrations returned to prelipid infusion levels 9 h after stopping the infusion of Intralipid, 1, 2, or 3 g/kg, there was a cumulative increase in plasma cholesterol and glucose concentrations. The close correlation between FFA concentrations and plasma lipolytic activity (r = 0.655, p < 0.001) suggests considerable in-travascular lipolysis. The positive correlation between plasma FFA and triglycerides (r = 0.632, p < 0.001) and FFA and cholesterol (r = 0.582. p < 0.001) indicate, however, that intravascular lipolysis does not prevent the lipemia associated with Intralipid infusion to low birth weight infants. This study suggests that the infusion of a mixture of small amounts of heparin and Intralipid leads to detachment of lipoprotcin lipase and hepatic lipase from the capillary endothelium of low birth weight infants. The resulting intravascular lipolysis probably exceeds the FFA disposal capacity of premature infants. The rise in plasma FFA concentration to 2.0 μmol/ml warrants caution in the combined use of Intralipid (at rates exceeding 2 g/kg/day) and low levels of heparin for the clinical management of premature infants maintained on total parenteral nutrition.

The presence of a triacylglycerol lipase in human gastric juice was described in previous studies. Its source and role in intragastric lipolysis was, however, uncertain. Our study presents definitive evidence for gastric origin of a lipase and its release by secretagogues. Both carbachol and cholecystokinin-8 stimulate release of this enzyme for dispersed human gastric glands. While the two secretagogues had similar efficacies, with nearly a 3-fold stimulation over basal rates, cholecystokinin-8 was about four orders of magnitude more potent in releasing lipolytic activity than carbachol (maximum stimulation at concentrations of 1 X 10(-9) and 1 X 10(-5) M, respectively). Lipolytic activity measured against triolein (18:1), tricaprylin (8:0) and tributyrin (4:0) emulsions was 1.18 +/- 0.12, 4.48 +/- 0.64, and 12.17 +/- 0.88 units (1 unit = 1 mumol free fatty acid released/min per mg protein), respectively. Characterization of the pH optimum for each substrate showed maximum lipolysis at 4.5 for tributyrin, and at 5.5 for tricaprylin and triolein. These results indicate that a lipase which hydrolyzes long-, medium- and short-chain triacylglycerols is secreted by human gastric mucosa. At pH 6.0, the pH of the duodenum, there is appreciable lipolytic activity in the presence of bile salts. This suggests that gastric lipase, in addition to initiating lipolysis in the stomach, might also aid in the digestion of lipids in the duodenum. It remains to be determined whether gastric lipase is distinct from lingual lipase, or is the same enzyme secreted by the lingual serous glands and the gastric mucosa.

The effect of heparin (10 U/kg) on serum lipolytic activity, triglyceride and FFA levels, during four hours infusion of 0.5 gm/kg Intralipid was measured in 18 AGA infants, 25 to 32 week's gestational age. PHLA, TG, and FFA were measured at 0, 10, 30, 120, and 240 minutes of infusion of Intralipid, before and following a bolus of 10 U/kg heparin iv. Lipolytic activity, measured by hydrolysis of activated tri- 3 H-oleate and expressed in μmol FFA released per milliliter serum per hour, was not detected in serum before heparin administration. Ten minutes after heparin administration peak PHLA was significantly higher in infants of 27 to 32 weeks' gestation than in infants of 25 to 26 weeks' gestation. There was no significant difference in peak PHLA between infants of 27 to 28 and 29 to 32 weeks' gestation. PHLA returned to baseline (zero) two hours after heparin administration in all infants. Infants of 25 to 26 weeks' gestational age had significantly higher concentrations of serum triglycerides before and during Intralipid infusion than in infants of 27 to 32 weeks' gestational age. Although there was a transient rise in FFA 10 and 30 minutes after heparin administration, the levels of FFA and triglycerides were not different at the end of infusion with or without heparin in either group, suggesting that a single bolus of heparin has only a transient effect on Intralipid clearance.