L- and T-type calcium channels in acutely isolated neonatal and adult cardiac myocytes.

We have compared transsarcolemmal Ca2+ currents in acutely isolated neonatal (1- to 5-d-old) and adult rabbit cardiac myocytes prepared using similar enzymatic techniques. Time- and voltage-dependent inward Ca2+ currents were measured using the whole-cell voltage clamp technique. In neonatal myocytes, peak Ca2+ currents measured 114 +/- 10 pA (mean +/- SEM, n = 18) as compared with 2014 +/- 403 pA in adult myocytes (n = 5, p less than 0.001). Although adult myocytes had a larger surface area (estimated from cell capacitance) than neonatal cells (113 +/- 15 x 10(-6) versus 28 +/- 2 x 10(-6) cm2, p less than 0.001), the calculated peak current density was also significantly larger in adult cells (17.9 +/- 2.5 compared to 4.3 +/- 0.4 microA/cm2 for neonatal cells, p less than 0.001). The voltage dependence of the peak Ca2+ current was similar in neonatal and adult myocytes. Early transient (T-type) Ca2+ currents were also studied by comparing the current induced by depolarization to -20 mV from holding potentials of -40 and -80 mV. T-type Ca2+ channels were present in 91% of the adult cells but were evident in only 39% of the neonatal cells. In summary, voltage-gated Ca2+ current amplitude, current density, and T-type Ca2+ channel prevalence all increase with maturation. These data suggest that neonatal myocytes may be relatively deficient in Ca2+ channel activity when compared to adult myocytes.     

Myocardial carbohydrate, ketone, and fatty acid uptake in conscious lambs with aortopulmonary shunts.

A left to right shunt increases myocardial work and is often accompanied by increased catecholamine levels. Because both increased myocardial work and increased catecholamine levels may induce increased fatty acid utilization, which could increase resting myocardial oxygen consumption and therefore unfavorably affect coronary reserve, we studied myocardial uptake of glucose, pyruvate, lactate, beta-OH-butyrate, acetoacetate, FFA, and triglycerides in 12 7-wk-old lambs with aortopulmonary left to right shunts (58 +/- 2% of left ventricular output, mean +/- SEM) and in 10 control lambs 2 wk after surgery. Despite the shunt, systemic blood flow in the shunt lambs was maintained at the same level as in the control lambs. This was accomplished by an increased heart rate and stroke volume. Furthermore, the shunt was accompanied by an increased myocardial oxygen consumption in the shunt lambs (834 +/- 70 versus 528 +/- 43 mumol O2.min-1 x 100 g-1; p less than 0.05). There were no significant differences in arterial substrate concentrations between the two groups. The same was true for arteriovenous differences across the myocardium, with the exception of lactate, which was substantially higher in shunt than in control lambs (72 +/- 25 versus 18 +/- 23 mumol/L; p less than 0.05). As a consequence, myocardial lactate uptake in the shunt lambs was increased 15-fold (18 +/- 6 versus 1 +/- 2 mumol.min-1 x 100 g-1; p less than 0.02), whereas uptake of the other substrates merely paralleled the increased myocardial blood flow.(ABSTRACT TRUNCATED AT 250 WORDS)     

Decreased fasting free fatty acids with L-carnitine in children with carnitine deficiency

At the time of acute presentation, children with carnitine deficiency may have increased free fatty acid concentrations and hypoglycemia. However, whether carnitine replacement affects the plasma concentration of these substrates remains to be determined. Therefore, to evaluate the effect of carnitine replacement on plasma substrate and hormone concentrations, five children with carnitine deficiency (two idiopathic, two secondary to long-chain acyl coenzyme A dehydrogenase deficiency, one secondary to isovaleric acidemia) were fasted overnight before and after treatment with oral carnitine (80 +/- 7 mg.kg-1.day-1). During carnitine supplementation, plasma total carnitine (19 +/- 4 versus 45 +/- 6 nmol/ml, pretreatment versus treatment, respectively) and free carnitine (11 +/- 3 versus 31 +/- 6 nmol/ml), as well as red blood cell total carnitine (0.057 +/- 0.019 versus 0.130 +/- 0.019 nmol/mg of hemoglobin) increased (p less than 0.05). Fasting plasma glucose (83 +/- 4 versus 85 +/- 3 mg/dl) and ketone body (0.54 +/- 0.18 and 0.56 +/- 0.20 mM) concentrations did not change with carnitine supplementation, but plasma free fatty acids (1.28 +/- 0.32 versus 0.77 +/- 0.07 mM) decreased (p less than 0.05). No differences in fasting insulin, growth hormone, or cortisol concentrations were observed. Urinary excretion of free carnitine (0.1 +/- 0.0 versus 2.4 +/- 0.7 mumol/mg creatinine), total carnitine (0.3 +/- 0.1 versus 3.4 +/- 0.9 mumol/mg creatinine) and acyl carnitine (0.2 +/- 0.1 versus 0.9 +/- 0.3 mumol/mg creatinine) increased (p less than 0.05) with carnitine supplementation. The decreased plasma free fatty acid concentrations with carnitine supplementation may be due to more efficient fatty acid oxidation and/or increased urinary excretion of fatty acids as acylcarnitines.     

Acute normoxia increases fetal pulmonary artery endothelial cell cytosolic Ca2+ via Ca2+-induced Ca2+ release

To test the hypothesis that an acute increase in O(2) tension increases cytosolic calcium ([Ca(2+)](i)) in fetal pulmonary artery endothelial cells (PAECs) via entry of extracellular calcium and subsequent calcium-induced calcium release (CICR) and nitric oxide release, low-passage PAECs (<10 passages) were isolated from the intralobar pulmonary artery (PA) of fetal sheep and maintained under hypoxic conditions (Po(2), 25 Torr). Using the calcium-sensitive dye fura-2, we demonstrated that acute normoxia (Po(2) = 120 Torr) increased PAECs [Ca(2+)](i) by increasing the rate of entry of extracellular calcium. In the presence of either ryanodine or 2-aminoethoxy-diphenylborate (2APB), normoxia did not lead to a sustained increase in PAECs [Ca(2+)](i) Whole-cell patch clamp studies demonstrated that acute normoxia causes PAEC membrane depolarization. When loaded with the nitric oxide (NO)-sensitive dye, DAF - FM, acute normoxia increased PAEC fluorescence. In PAECs derived from fetal lambs with pulmonary hypertension, an acute increase in O(2) tension had no effect on either [Ca(2+)](i) or NO production. Hypoxia increases loading of acetylcholine-sensitive calcium stores, as hypoxia potentiated the response to acetylcholine We conclude that acute normoxia increases [Ca(2+)](i) and NO production in normotensive but not hypertensive fetal PAECs via extracellular calcium entry and calcium release from calcium-sensitive intracellular stores.     

Intestinal maturation: characterization of mitochondrial calcium transport in the rat

The mitochondria play a major role in the regulation of intracellular calcium. Despite the fact that the enterocytes receive the majority of absorbed calcium, the role of the intestinal mitochondria in calcium transport during maturation is not known. Therefore, the current studies were designed to characterize calcium pump activity of jejunal mitochondria of rats during maturation (suckling, weanling, and adolescent rats). The functional integrity of the intestinal mitochondria of suckling and adolescent rats was determined by oxygen consumption studies demonstrating respiratory control ratios of more than 3 when succinate was used as a test substrate. Ca++ uptake was significantly stimulated by the presence of 3 mM ATP at all age groups studied. Maximal Ca++ uptake in the presence of 3 mM ATP and 2 mM succinate was 31.1 +/- 0.4, 50.2 +/- 4.2, and 94.3 +/- 1.5 nmol/mg protein (mean +/- SE) in suckling, weanling, and adolescent rats, respectively. Rates of ATP hydrolysis were 15.5 +/- 1.5 and 2.9 +/- 0.3 nmol/ATP hydrolyzed/mg protein in adolescent and suckling rats, respectively (p less than 0.001). Ca++ uptake was completely inhibited by 0.25 microM ruthenium red, oligomycin (10 micrograms/mg protein), 0.5 mM dinitrophenol and 1 mM sodium azide at all age groups. Ca++ efflux in the presence of ruthenium red occurred by a Na+-dependent pathway, indicating a Ca++/Na+ exchange mechanism. Kinetic parameters for ATP stimulated Ca++ uptake at 10 s revealed a Km of 0.84 +/- 0.11, 0.65 +/- 0.17, and 0.57 +/- 0.03 microM and Vmax of 1.83 +/- 0.07, 3.62 +/- 0.26 and 14.15 +/- 0.21 nmol/mg protein/10 s in suckling, weanling, and adolescent rats, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)     

Adrenal function in thalassemia major following long-term treatment with multiple transfusions and chelation therapy. Evidence for dissociation of cortisol and adrenal androgen secretion

Eight patients with beta-thalassemia who were given long-term treatment with combined multiple transfusions and chelation therapy underwent adrenal testing. The six male and two female patients ranged in age from 7 to 19 years. Six of eight patients had delayed bone ages and height greater than 2.5 SDs below the mean. Of the six patients more than 13 years of age, two had clinical evidence of isolated adrenarche and only one had evidence of true puberty. Cortisol levels were similar in patients and controls at zero time (10.6 +/- 1.8 micrograms/dL [292 +/- 50 nmol/L] vs 10.8 +/- 1.4 micrograms/dL [298 +/- 39 nmol/L]) and at 60 minutes (26.6 +/- 2.5 micrograms/dL [734 +/- 69 nmol/L] vs 24.9 +/- 1.9 micrograms/dL [687 +/- 52 nmol/L]) after insulin hypoglycemia (all values are the mean +/- SE). During an eight-hour infusion of ACTH, cortisol responses in the patients with thalassemia were not significantly different from those of controls. Baseline levels of the adrenal androgens dehydroepiandrosterone (DHEA) and dehydroepiandrosterone sulfate (DHEA-S) were significantly lower in the subjects with thalassemia compared with controls of similar bone age and pubertal status. The prolonged ACTH infusion caused a significant increase in the DHEA level (79.2 +/- 14.7 ng/dL [2.74 +/- 0.51 nmol/L] vs 538.6 +/- 38.1 ng/dL [18.67 +/- 4.79 nmol/L]) and the DHEA-S level (37.5 +/- 10.8 micrograms/dL [1.02 +/- 0.29 mumol/L] vs 70.5 +/- 18.3 micrograms/dL [1.19 +/- 0.50 mumol/L]) in the patients. The patients' peak stimulated levels of DHEA-S were significantly lower than those of the controls, whereas peak levels of DHEA were similar in the patients and the controls. These results indicate that combined multiple transfusions and chelation therapy preserve the integrity of the ACTH-cortisol axis in patients with thalassemia. The reduced levels of adrenal androgens, short stature, and delayed puberty noted in our patients suggest, however, that alternative approaches to the therapy of thalassemia are needed.     

Ontogeny of calcium transport by intestinal Golgi in spontaneously hypertensive rats and genetically matched WKY rats

Our studies were designed to characterize calcium transport by intestinal Golgi vesicles in spontaneously hypertensive rats (SHR) and their genetically matched control, Wistar-Kyoto rats (WKY). The biochemical purity of the intestinal Golgi in SHR and WKY was validated by marker enzyme studies. Calcium uptake by Golgi vesicles represented transport into the intravesicular space as evidenced by temperature dependency and by calcium ionophore A23187-induced calcium efflux experiments. ATP-driven calcium uptake was stimulated several-fold compared with uptake in the absence of ATP and adenylyl-(beta-gamma-methylendiphosphate) (nonhydrolyzable ATP) in both SHR and WKY. ATP-dependent calcium uptake was significantly higher in WKY compared with SHR at early times points, 15 s-5 min (p less than 0.05-0.01). The initial rate of calcium uptake was linear up to 60 s. Kinetic parameters of calcium uptake at free calcium concentrations of 0.1 to 2.0 microM showed a Vmax of 1.64 +/- 0.06 and 1.2 +/- 0.06 nmol.mg protein-1.15 s-1 in WKY and SHR, respectively (p less than 0.01), and the Km values were 0.17 +/- 0.03 and 0.16 +/- 0.04 microM, respectively. Kinetic analysis of ATP-dependent calcium uptake in 3-wk-old rats showed a Vmax of 0.07 +/- 0.005 and 0.36 +/- 0.05 nmol/mg protein-1.15 s-1 (p less than 0.01) and a Km of 0.26 +/- 0.08 and 0.4 +/- 0.2 microM in SHR and WKY, respectively. These results suggest that intestinal Golgi vesicles in SHR and WKY demonstrate an ATP-driven calcium uptake. This ATP-dependent process is significantly decreased in the weanling and adult SHR compared with WKY. Such an abnormality in intracellular calcium regulation may have a role in the development of hypertension.     

Increased renal tubular reabsorption of calcium and magnesium by the offspring of diabetic rat pregnancy

Diabetic pregnancy has a marked influence on offspring calcium and magnesium homeostasis. Urinary excretion of calcium and magnesium is reduced, yet offspring of diabetic pregnancy exhibit hypomagnesemia and hypocalcemia. The aim of this study was to measure renal hemodynamic and tubular function in the offspring of diabetic (OD) and control, nondiabetic (OC) rats at 4 and 8 wk of age to determine the glomerular and tubular mechanisms through which renal calcium and magnesium handling are programmed in utero. The fraction of filtered calcium that was excreted was significantly lower in OD at both 4 and 8 wk of age [8 wk: OC (n = 6), 11.8 +/- 2.9 versus OD (n = 5), 4.3 +/- 0.6%; p < 0.05] and that of magnesium was lower at 8 wk of age [OC (n = 6), 42.4 +/- 7.5 versus OD (n = 5), 13.0 +/- 1.7%; p < 0.01]. This increased reabsorption occurred despite an elevated GFR in OD. These findings clearly indicate that tubular reabsorptive mechanisms for calcium and magnesium are increased markedly in OD. Serum PTH concentration was reduced in 8-wk-old OD [OC (n = 7), 539.4 +/- 142.1 versus OD (n = 9), 174.3 +/- 69.4 pg/ml; p < 0.05], consistent with previous reports in human infants. Taken together, these observations suggest that the basis for the altered renal magnesium and calcium handling in OD involves increased tubular transport activity and possibly increased sensitivity of these mechanisms to PTH.     

Changes in oxypurine concentrations in vitreous humor of pigs during hypoxemia and post-mortem

In the vitreous humor from three hypoxemic and one control group of pigs, hypoxanthine, xanthine, and uric acid concentrations were measured. The purine concentrations were measured before the hypoxemia, at the time of death, and 24 h post-mortem. During hypoxemia with arterial O2 tension between 2.1 and 3.0 kPa [fractional inspired oxygen (FiO2) = 0.08], hypoxanthine concentrations increased from a mean basal value of 11.7 +/- 5.6 mumol/L to 16.3 +/- 2.4 mumol/L at the time of death (NS). Xanthine concentrations changed from a basal value of 0.3 +/- 0.1 mumol/L to 0.6 +/- 0.2 mumol/L (p less than 0.02), and uric acid changed from 3.4 +/- 1.1 mumol/L to 5.0 +/- 4.5 mumol/L (NS). During hypoxemia with arterial O2 tension between 3.0 and 4.0 kPa (FiO2 = 0.11), hypoxanthine increased in the vitreous humor from a mean basal value of 9.1 mumol/L to 20.3 mumol/L at the time of death (p less than 0.02). Xanthine concentrations increased from 0.3 mumol/L to 1.3 mumol/L (p less than 0.05), whereas there was no change in uric acid concentration (basal 5.0 +/- 0.8 mumol/L and final 4.5 +/- 1.0 mumol/L). During milder hypoxemia with arterial O2 tension between 4.3 and 5.6 kPa (FiO2 = 0.14), or in the control group (FiO2 = 0.21), neither of the metabolites changed significantly. The vitreous humor was not stable post-mortem, inasmuch as the mean concentration of hypoxanthine increased from 18.2 +/- 7.7 mumol/L to 121.6 +/- 57.4 mumol/L 24 h post-mortem (p less than 0.01).(ABSTRACT TRUNCATED AT 250 WORDS)     

Effects of maternal protein-calorie malnutrition on the phospholipid composition of surfactant isolated from fetal and neonatal rat lungs. Compensation by inositol and lipid supplementation.

The effects of a maternal protein-calorie malnutrition during gestation and lactation were analyzed on fetal and postnatal lung growth and maturation, including a surfactant fraction isolated from lung tissue. There was a considerable reduction in body weight and in wet and dry lung weights of malnourished pups. Lung protein and DNA concentrations were similar in both groups except in late gestation (lung hyperplasia) and 2 and 15 d after delivery (hypocellularity). Lung glycogen breakdown was slowed down in malnourished newborns. Surfactant material was decreased the most perinatally and the reduction was more marked than for the nonsurfactant fraction of the lung. Disaturated phosphatidylcholine, the major surface active surfactant component, was decreased the most at birth (1.70 +/- 0.31 nmol/mg wet wt versus 3.68 +/- 0.17 nmol/mg in controls, n = 8) and on d 2 (5.04 +/- 0.53 nmol/mg versus 7.67 +/- 0.44 nmol/mg in controls, n = 8). There was an apparent recovery in the composition of surfactant in malnourished rats 5 d after delivery, due in fact to a decrease in controls, and an actual return to normal levels 15 to 20 d after birth. Postnatal lipid supplementation with Intralipid led to partial recovery on d 10. Inositol supplementation totally reverted the effects of malnutrition on surfactant phospholipids (8.36 +/- 0.94 nmol disaturated phosphatidylcholine/mg wet wt on d 2 versus 7.67 +/- 0.44 nmol/mg in controls and 5.55 +/- 0.62 nmol/mg in untreated malnourished rats, n = 10; 2.43 +/- 0.32 nmol disaturated phosphatidylcholine/mg wet wt on d 10 versus 3.26 +/- 0.32 nmol/mg in controls and 1.18 +/- 0.27 nmol/mg in untreated malnourished rats, n = 8).(ABSTRACT TRUNCATED AT 250 WORDS)     

Sustained-release terbutaline vs sustained-release theophylline in young patients with asthma

Twenty patients with asthma (mean age, 10.9 +/- 2 years) entered a six-week, randomized, double-blind, crossover comparison of sustained-release (S-R) terbutaline sulfate (Bricanyl Durules) vs S-R theophylline (Theo-Dur). In each two-week study period each patient received S-R theophylline twice daily in doses previously adjusted to give serum theophylline concentrations in the range of 10 to 20 mg/L (56 to 111 mumol/L); or S-R terbutaline sulfate, 5 mg twice daily; or S-R terbutaline sulfate, 7.5 mg twice daily. All treatment regimens produced significant improvement in one or more pulmonary function test values compared with prestudy values. The incidence of acute asthma episodes were similar during each treatment regimen. No clinically significant difference occurred between the regimens for daily symptom scores, peak expiratory flow rates, or use of a terbutaline metered-dose inhaler. At the end of the theophylline treatment period, the mean (+/- SD) theophylline level 12 to 14 hours after the last dose was 10.1 +/- 3.3 mg/L (56 +/- 18 mumol/L); at the end of the terbutaline treatment periods, the mean trough terbutaline levels were 2.22 micrograms/L (9.9 +/- 4.4 nmol/L) (S-R terbutaline sulfate, 5 mg twice daily) and 3.07 micrograms/L (13.7 +/- 5.4 nmol/L) (S-R terbutaline sulfate, 7.5 mg twice daily). Adverse effects, including tremor, occurred with similar frequency during all three drug regimens. Sustained-release formulations of theophylline and terbutaline, in the dosages studied, provided comparable control of asthma symptoms.     

Leucine kinetics during feeding in normal newborns

To examine how leucine and protein metabolism is affected by feeding, leucine kinetics were determined in 11 normal term newborns during feeding using a prime constant tracer infusion of 1-13C leucine combined with respiratory calorimetry. Fed newborns were compared with previously studied fasting newborns. Feeding and fasting newborns had similar rates of leucine oxidation (34 +/- 3 mumol/kg/h versus 31 +/- 4 mumol/kg/h) and leucine release from existing protein (156 +/- 16 mumol/kg/h versus 164 +/- 8 mumol/kg/h). In contrast, nonoxidative disposal rates of leucine (a reflection of protein synthesis) were significantly greater in feeding newborns (170 +/- 13 mumol/kg/h versus 129 +/- 9 mumol/kg/h). A significant positive correlation between birth weight and leucine flux was demonstrated in both feeding and fasting newborns. These results suggest that 1) newborns may accomplish protein accretion primarily by increases in protein synthesis rather than suppression of protein breakdown; 2) an estimate can be made of the minimal leucine intake required to replace irreversible leucine oxidative losses (816 mumol/kg/d, 107 mg/kg/d); and 3) the positive correlation between birth weight and leucine flux in both feeding and fasting newborns may be a result of differences in previous protein and energy supplies.     

Calcium metabolism and cystic fibrosis: mitochondrial abnormalities suggest a modification of the mitochondrial membrane

A disorder of calcium (Ca2+) metabolism may be central to the pathogenesis of cystic fibrosis (CF). Average cellular Ca2+ levels in fibroblasts derived from patients with CF (ages, 14-25 yr; n = 25) were 36-77% higher than in matched controls depending on age of cell culture (9.0-10.6 versus 5.1-7.8 nmol/mg cellular protein). Cellular Ca2+ was significantly elevated in CF, but was not a reliable criterion for identifying CF cells because of the high variability of results. Studies of Ca2+ fluxes in cell organelles showed that mitochondria isolated from CF fibroblasts accumulate 2-3 times more Ca2+ than controls [79.5 +/- 8.2 versus 33.7 +/- 4.7 nmols X mg mitochondrial protein-1 X 10 min-1 (+/- SD)], Ca2+ accumulation in mitochondrial reliably distinguished between CF and control or heterozygote cells (P less than 0.0005, n = 11). In vitro experiments showed that Ca2+ influx and efflux are increased in isolated CF mitochondria, resulting in net Ca2+ accumulation. Ca2+ uptake in mitochondria is energy-dependent; some inhibitors of mitochondrial energy metabolism (atractyloside, oligomycin) influenced Ca2+ intake significantly more in CF than in control mitochondria. Furthermore, the average activities of NADH oxidase, NADH- and succinate-cytochrome c reductase were 77, 58, and 48% higher in CF mitochondria, respectively. This indicates that many functions associated with energy metabolism and the mitochondrial membrane (electron transport, ATP transport, and ATP hydrolysis) are not operating properly in CF, thus possibly causing the derangement of Ca2+ metabolism found in CF mitochondria and cells.     

Increased cardiomyocyte intracellular calcium during endotoxin-induced cardiac dysfunction in guinea pigs

Septic shock is a complex pathophysiologic state characterized by circulatory insufficiency, multiple system organ dysfunction, and frequent mortality. Although profound cardiac dysfunction occurs during sepsis, the pathogenesis of this dysfunction remains poorly understood. To determine whether abnormalities in intramyocyte calcium accumulation might contribute to the development of cardiac dysfunction, we measured myocyte intracellular calcium during peak cardiac dysfunction after an endotoxin challenge. Intraperitoneal administration of Escherichia coli lipopolysaccharide 4 mg/kg to adult guinea pigs resulted in significantly impaired cardiac performance (Langendorff preparation) 18 h after challenge compared with control. This included diminished left ventricular pressure development (56 +/- 7 versus 95 +/- 4 mm Hg, p < 0.05), maximal rate of left ventricular pressure rise (998 +/- 171 versus 1784 +/- 94 mm Hg/s, p < 0.05) and left ventricular pressure fall (1014 +/- 189 versus 1621 +/- 138 mm Hg/s, p < 0.05). Assay of intracellular calcium in fura-2AM-loaded cardiac myocytes demonstrated increased intracellular calcium concentration in myocytes obtained from lipopolysaccharide-challenged animals compared with controls (234 +/- 18 versus 151 +/- 6 nM, p < 0.05). Inhibition of calcium-release channel (ryanodine receptor) opening by administration of dantrolene prevented the increase in intracytoplasmic calcium (159 +/- 8 versus 234 +/- 18 nM, p < 0.05) and partially ameliorated systolic and diastolic ventricular dysfunction. These data indicate that abnormalities of intracellular calcium contribute to the development of endotoxin-induced myocardial dysfunction.     

Mechanisms of increased susceptibility of immature and weaned pigs to Escherichia coli heat-stable enterotoxin.

Pigs demonstrate an increased sensitivity and susceptibility to Escherichia coli heat-stable enterotoxin (STa) in the 1st wk of life and immediately after weaning. To determine the possible mechanisms for this increased susceptibility, we compared STa binding, guanylate cyclase activation, and photoaffinity cross-linking to porcine jejunal brush border membranes prepared from immature (less than or equal to 1 wk of age) versus adult pigs as well as 3-wk-old weaned versus unweaned pigs. The STa binding capacity of immature pigs was nearly twice that of adult pigs (11.73 +/- 1.52 versus 6.00 +/- 0.96 x 10(-11) mol/L, p less than 0.001), and the STa binding capacity of weaned pigs was nearly three times greater than that of unweaned pigs (17.48 +/- 2.10 versus 4.86 +/- 1.02 x 10(-11) mol/L, p less than 0.001). Scatchard analysis suggested a single class of STa receptor, with an association of binding constant of approximately 10(9) L/mol at all ages. Maximum guanylate cyclase response (expressed as pmol cyclic GMP generated/mg brush border membrane protein/min) was greater in immature versus adult pigs (1312 +/- 831 versus 320 +/- 92, p less than 0.02). Weaned pigs had a greater maximum guanylate cyclase activation than unweaned pigs (1126 +/- 692 versus 624 +/- 298); however, this difference was not statistically significant. Autoradiograms demonstrated specific cross-linking of 125I-STa to a number of distinct radiolabeled bands (62, 66, 84, 92, 160, and 165 kD). There was a difference in the size and trypsin sensitivity of these radiolabeled bands as a function of age and weaning. Treatment with trypsin decreased the intensity of the 160 to 165-kD bands while increasing the intensity of the 62- to 66- and 84- to 92-kD bands. These differences in STa binding, guanylate cyclase activation, and STa receptor size may increase the susceptibility of pigs during the 1st wk of life and at weaning to STa-mediated diarrheal disease.     

Isolation and characterization of erythrocyte membrane Ca2+-ATPase in cystic fibrosis

Ca2+-ATPase was purified from erythrocyte membranes prepared from cystic fibrosis (CF) blood samples (n = 10) and from age/sex-matched control blood samples (n = 10). The kinetics of calcium activation of the purified enzyme was investigated in the presence of asolectin phospholipids and found to be virtually identical for both CF and control preparations: VCa2+ = 3.01 +/- 0.24 mumol ATP hydrolyzed/mg pure enzyme/min (mean +/- SE) and 3.09 +/- 0.20 for CF and control Ca2+-ATPase, respectively; KCa2+ = 0.328 +/- 0.046 mu molar free calcium and 0.333 +/- 0.040 for CF and control enzyme, respectively. The preparative procedure used (one-step purification by calmodulin-affinity chromatography) allowed quantitative recovery of essentially 100% of the Ca2+-ATPase present in detergent-solubilized erythrocyte membranes, enabling expression of the yield of purified enzyme in terms of the quantity of starting membrane protein: 0.127% +/- 0.006% (w/w) and 0.140% +/- 0.007% for CF and control enzyme preparations, respectively. None of the parameters evaluated showed a statistically significant difference (p less than 0.05) between the CF and control groups. Furthermore, when CF and control purified Ca2+-ATPase samples were analyzed by high-resolution gradient SDS-polyacrylamide gel electrophoresis, no differences in mobility were observed (mol wt = 128 kdaltons). Thus, Ca2+-ATPase purified from CF erythrocyte membranes and assayed in the presence of asolectin appears to be quantitatively similar to control purified enzyme in amount, molecular weight, and kinetics of activation by calcium. These data suggest that Ca2+-ATPase may not be the defective gene product in CF.(ABSTRACT TRUNCATED AT 250 WORDS)     

Serum beta-carotene, retinol, and alpha-tocopherol levels during mineral oil therapy for constipation

Twenty-five children with chronic constipation underwent serial monitoring of serum beta-carotene, retinol (vitamin A1), and alpha-tocopherol (vitamin E) levels during mineral oil therapy. Mineral oil was administered between meals. Patients were monitored for up to four months of therapy. Mean serum beta-carotene levels fell from 1.0 +/- 0.5 mumol/L (55.7 +/- 26.0 micrograms/dL) to 0.7 +/- 0.4 mumol/L (35.9 +/- 22.1 micrograms/dL) after the first month of mineral oil therapy and remained depressed throughout the remainder of the study. Serum alpha-tocopherol levels remained unchanged throughout the observation period. There was a modest increase in serum retinol levels during the study, especially after three months (from 1.48 +/- 0.84 mumol/L [42.3 +/- 24.1 micrograms/dL] to 2.22 +/- 0.77 mumol/L [63.5 +/- 22.1 micrograms/dL]). We conclude that while a short course of mineral oil can induce a reduction in the serum level of beta-carotene, the treatment has no adverse effect on serum levels of retinol and alpha-tocopherol.     

Postnatal changes in calcium-regulating hormones in very-low-birth-weight infants. Effect of early neonatal hypocalcemia and intravenous calcium infusion on serum parathyroid hormone and calcitonin homeostasis

In very-low-birth-weight (VLBW) infants, we studied the hypotheses that in early neonatal hypocalcemia the serum parathyroid hormone (PTH) concentration would rise; the serum calcitonin (CT) concentration would decline; and, in response to intravenous (IV) calcium (Ca) infusion, the serum PTH concentration would be lowered; and the serum CT concentration would rise. Fifteen infants appropriate for gestational age (age, less than 32 weeks; birth weight, less than 1,500 g) were enrolled in the study. In eight infants in whom the serum Ca level declined to less than 6.0 mg/dL, changes in serum magnesium, phosphorus, PTH, CT, and whole blood ionized calcium (iCa) were evaluated on entry into the study, when serum Ca declined to less than 6.0 mg/dL, immediately after infusion of 18 mg/kg of elemental calcium as calcium gluconate, and at eight hours post-Ca infusion (+ 8 hr). The serum Ca concentration declined from 7.9 +/- 0.6 baseline (mean +/- SE) to 5.2 +/- 0.2 mg/dL pre-Ca infusion and rose to 9.17 +/- 0.74 mg/dL post-Ca infusion and 7.1 +/- 0.5 mg/dL at +8 hr post-Ca infusion. Whole blood iCa declined from 4.82 +/- 0.24 to 3.72 +/- 0.19 mg/dL pre-Ca infusion, rose to 6.68 +/- 0.32 mg/dL post-Ca infusion, and was 4.12 +/- 0.21 mg/dL at + 8 hr post-Ca infusion. The serum P concentration did not change significantly. The serum PTH concentration rose from 116 +/- 17 to 204 +/- 34 pmole/L pre-Ca infusion, declined to 149 +/- 22 pmole/L post-Ca infusion, and was 187 +/- 28 pmole/L at + 8 hr post-Ca infusion. The serum CT concentration was elevated and did not change significantly. Thus, in infants less than 32 weeks' gestation, the serum PTH level rises in early neonatal hypocalcemia and is suppressed by IV Ca infusion; the serum CT level is markedly elevated and is not altered in early neonatal hypocalcemia and does not rise further in response to IV Ca infusion in VLBW infants. We suggest that hypercalcitoninemia occurs in VLBW infants and that serum CT concentrations are unresponsive to changes in serum Ca.     

Assessment of cardiac function by M-mode echocardiography in selenium-deficient phenylketonuric children

Selenium (Se) deficiency is associated with cardiac and skeletal muscle dysfunction. Twenty well children aged 2-16 years (10 male) attending the Phenylketonuria (PKU) Clinic at the Royal Children's Hospital, Brisbane, had low Se levels (mean 0.29 +/- 0.02 s.e.m. mumol/L; normal range 0.56-1.16 mumol/L). Their myocardial function was assessed at rest and after exercise provocation by M-mode echocardiography in order to exclude occult left ventricular dysfunction. At rest, fractional shortening (FS) was normal (mean 38.1 +/- 1.1 s.e.m. %, n = 20). After exercise, FS increased significantly (P less than 0.001) from 37.6 +/- 1.4% to 44.3 +/- 1.2%, n = 12). This was associated with a significant rise (P less than 0.001) in heart rate (HR) from 77.3 +/- 3.1 beats/min to 125.8 +/- 5.2 beats/min (n = 12). The normal resting FS and normal increase in FS and HR with exercise is evidence against significant cardiac impairment in this group of Se-deficient children.     

Effects of a protein-rich diet during convalescence from shigellosis on catch-up growth, serum proteins, and insulin-like growth factor-I.

Shigellosis in children can cause growth retardation, worsening of malnutrition, and hypoproteinemia. To assess the effects of ingestion of a protein-rich diet during convalescence, 22 children aged 2 to 4 y with culture-proven shigellosis were randomly assigned after 5 d of antibiotic treatment to 21-d feeding regimens of either a 150 kcal/kg/d high-protein diet with 15% of calories as protein or an isocaloric control diet with 6% of calories as protein. At the start and end of dietary treatment, weight, height, mid-arm circumference, skinfold thickness, serum protein concentrations, and serum IGF-I were measured. Means of weight gain and increases in mid-arm circumference were greater in children fed high-protein diets than those fed control diets (1.23 versus 0.76 kg; 1.40 versus 0.96 cm; p < 0.05). Mean increase in height in children fed high-protein diets (0.83 cm) was not significantly greater than with control diets (0.74 cm). Mean increases in serum concentrations of total protein, prealbumin, and retinol-binding protein were greater in the high-protein group than in controls (p < 0.05). Mean serum concentrations of IGF-I were low in both groups before treatment [4.2 +/- 2.6 nmol/L (31.9 +/- 19.6 ng/mL) in controls; 3.1 +/- 3.4 nmol/L (24.0 +/- 26.3 ng/mL) in the high-protein group] but increased more in the high-protein group [39.0 +/- 16.2 nmol/L (298 +/- 124 ng/mL)] than in the control group [16.7 +/- 9.2 nmol/L (128 +/- 70 ng/mL), p < 0.01].(ABSTRACT TRUNCATED AT 250 WORDS)