Distribution and anchoring of molecular forms of acetylcholinesterase.

Molecular forms of acetylcholinesterase exhibit tissue-specific distribution, and each form is anchored to the cell surface via a particular post-translational modification of the catalytic subunit. Nibaldo Inestrosa and Alejandra Perelman review evidence that heparan sulphate proteoglycans are the extracellular matrix receptors for the collagen-tailed enzyme, and that a glycolipid which contains phosphatidylinositol and a 20 kDa hydrophobic peptide participate in the anchoring of the hydrophobic globular forms of acetylcholinesterase to the cell surface.

Suppression of gonadotropin secretion by a long-acting gonadotropin-releasing hormone analog (leuprolide acetate, Lupron Depot) in children with precocious puberty.

The GnRH analog leuprolide acetate has been shown to be effective in the treatment of precocious puberty when given as a daily sc injection. We studied the effectiveness of a single im dose of a new depot form of leuprolide in suppressing estradiol and gonadotropin secretion in children with precocity. Five girls with previously untreated precocity showed significant decreases in basal serum estradiol and FSH levels and in peak LH levels (after GnRH testing) 30 days after a single im dose of leuprolide acetate for depot suspension. Mean peak FSH levels also fell greatly, but the difference was not significant. No adverse effects were noted during the first 4-6 months of monthly im injections. Depot im leuprolide appears to be effective in suppressing estradiol and gonadotropin secretion, and may be a useful method of treating children with central precocious puberty.

Intrauterine diagnosis and treatment of fetal goitrous hypothyroidism.

Newborn screening programs for the detection of congenital hypothyroidism have dramatically shortened the time before treatment is begun. However, concern still exists about central nervous system sequelae which may persist due to a period of untreated intrauterine hypothyroidism. Presence of polyhydramnios led to the ultrasound diagnosis of a fetal goiter. Hypothyroidism was confirmed at 34 weeks gestation by percutaneous fetal blood sampling, which revealed an elevated TSH (186 mU/L) and a low T4 (19.3 nmol/L). Intraamniotic fluid injections of 500 micrograms levothyroxine sodium (T4) every 10-14 days increased fetal serum T4 (59.2 nmol/L), decreased fetal serum TSH (14 mU/L), decreased amniotic fluid TSH, and decreased the size of the fetal goiter. The infant was born at term without perinatal complications. Thyroid function studies on cord blood were normal (T4, 109.4 nmol/L; TSH, 1.3 mU/L), and the infant was discharged on oral T4. Follow-up examination at age 6 weeks revealed that the infant was developmentally normal and clinically and chemically euthyroid. Intrauterine T4 therapy can suppress fetal TSH and treat fetal hypothyroidism despite hypothyroid levels of serum T3. Highly sensitive TSH assays may allow the use of amniotic fluid TSH as a marker for fetal hypothyroidism.

Plasma homocysteine levels in obstructive sleep apnea: association with cardiovascular morbidity.

OBJECTIVES: Obstructive sleep apnea (OSA) is associated with cardiovascular morbidity and mortality. Plasma levels of homocysteine are also associated with cardiovascular morbidity and mortality. We therefore investigated homocysteine and conventional cardiovascular risk factors in OSA patients with and without cardiovascular morbidity in comparison with normal control subjects and ischemic heart disease (IHD) patients without OSA. SETTING: Technion Sleep Medicine Center, Haifa, Israel. METHODS AND PARTICIPANTS: Levels of homocysteine, cholesterol, low-density lipoprotein, high-density lipoprotein, triglycerides, creatinine, vitamins B(12) and B(6), and folic acid were determined in 345 participants after overnight fasting. These included OSA patients with IHD (n = 49), with hypertension (n = 61), or without any cardiovascular disease (n = 127). Two control groups were employed: IHD patients without or with low likelihood for sleep apnea (n = 35), and healthy control subjects (n = 73). RESULTS: After adjustment for age, body mass index, creatinine, and existence of diabetes mellitus, OSA patients with IHD had significantly higher homocysteine levels (14.6 +/- 6.77 micromol/L) than all other groups including the IHD-only patients. Hypertensive OSA patients had comparable homocysteine levels to IHD patients (11.80 +/- 5.28 micromol/L and 11.92 +/- 5.7 micromol/L, respectively), while patients with OSA only had comparable levels to normal control subjects (9.85 +/- 2.99 micromol/L and 9.78 +/- 3.49 micromol/L, respectively). No differences in conventional cardiovascular risk factors or in vitamin levels were found between groups. CONCLUSIONS: Patients with the combination of IHD and OSA have elevated homocysteine levels. We hypothesize that these results may be explained by endothelial dysfunction combined with excess free-radical formation in OSA patients.

The fetus in maternal hyperthyroidism.

A recent article in the New England Journal of Medicine reported the successful diagnosis and treatment of fetal goitrous hypothyroidism in a mother with Graves' disease. The fetus is being recognized as an important patient in its own right in terms of thyroid disease. The fetal thyroid system develops independently of the normal maternal thyroid axis. Presence of feedback suppression of TSH by T4 has been demonstrated in a 35-week fetus. Information learned from congenital hypothyroidism suggests that lack of fetal thyroid hormones may have a negative impact on the developing fetal brain with lack of normal myelination. It is uncertain at what gestational age the fetus and the developing central nervous system become adversely affected by thyroid hormone deficiency. Since congenital hypothyroidism is sporadic and since there is no current method for easily screening all pregnancies for hypothyroidism, the thrust in fetal diagnosis and therapy has been in those pregnancies suspected of having a hypothyroid fetus when a fetal goiter is detected by ultrasonography or in a hyperthyroid mother who may be on antithyroid therapy. Intraamniotic injections of L-thyroxine have proven successful for fetal therapy. Amniotic fluid TSH may prove useful in the diagnosis and treatment of a hypothyroid fetus. Previous studies have suggested that the period of thyroxine dependency of the fetal central nervous system is limited predominantly to the last 4-8 weeks of gestation. Fetal hyperthyroidism due to transplacental transmission of thyroid-stimulating immunoglobulins may occur in a mother with a history of hyperthyroidism due to Graves' disease.(ABSTRACT TRUNCATED AT 250 WORDS)

Infantile mammary duct ectasia: a cause of bloody nipple discharge.

Bloody nipple discharge in infancy has been rarely reported in the medical literature. Its cause is unknown. We report a three-year-old male infant and a five-month-old female infant with bloody nipple discharge. Because of persistent bloody discharge, a subcutaneous mastectomy was performed in the boy; the problem resolved in the girl after a period of observation. The specimen showed histologic changes identical to those seen in adult mammary duct ectasia. All the endocrinologic work-up was normal. We suspect that bloody nipple discharge in infancy is underreported. This is a benign condition with histologic changes similar to adult mammary duct ectasia and if persistent, should be properly investigated; biopsy or excision are not indicated.

The occult insulinoma operative localization by quick insulin radioimmunoassay.

Hypoglycemia secondary to organic hyperinsulinism in children can be caused by diffuse or localized pancreatic lesions. Differentiation between these two types of lesions is of utmost importance since the surgical approach will be different. Some tumors escape detection by all preoperative investigations including ultrasound, scintiscan, arteriography, and computerized tomography. We are reporting on a 12-year-old boy with organic hyperinsulinism in whom we were unable to localize a tumor preoperatively. Peroperative determination of insulin levels from a number of sites on the pancreas enabled us to localize an insulin-producing pancreatic adenoma. This technique can be done easily by catheterizing the splenic and portal vein through a branch of the splenic vein and by serial sampling at 2 cm intervals along the portal and splenic veins. Insulin levels at these sites were determined by quick double-antibody radioimmunoassay, which allows the determination of the insulin levels within 50 minutes after sampling. There was a perfect biochemic and anatomic correlation allowing us to perform a precise distal pancreatectomy. The technique can be used to localize pancreatic adenomas and to decide how much pancreas to remove in diffuse lesions avoiding "blind" pancreatectomies.

Studies of the fungistatic activity of antifungal compounds in mash and pelleted feeds.

The effect of calcium propionate (CP) and Agrosil (AG) as mold inhibitors in wetted mash and pelleted feed was studied using both commercial cattle and poultry rations. Number of fungal colonies isolated after pelleting was markedly reduced; however, wetting the pellets accelerated the build-up of inoculum and resulted in spoilage. The addition of CP (.3%) to the cattle ration before pelleting prevented mold proliferation during one month of storage while the number of fungal colonies counted in pellets treated with AG (.15%) markedly increased over that period. However, AG had a longer fungistatic effect than CP in preserving the mash diet. Both materials, applied at .2%, were ineffective in preserving wet pelleted poultry feed. After 17 days of storage, feed treated with either of the agents was visibly moldy. In all cases, an increase in mold population was concomitant with elevated carbon dioxide concentrations, which indicated the sensitivity of this parameter for measuring fungal activity. Fat content of the diets remained unchanged during the storage period in spite of increased fungal activity.

JC-1: alternative excitation wavelengths facilitate mitochondrial membrane potential cytometry.

Mitochondrial membrane potential provides a valuable indicator of cells' health and functional status. Cytometry- and microscopy-based analyses, in combination with fluorescent probes, are widely used to study mitochondrial behavior related to cellular pathways, most notably - apoptosis. The cyanine dye JC-1 (5,5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimi- dazolylcarbocyanine iodide) facilitates discrimination of energized and deenergized mitochondria because the normally green fluorescent dye forms red fluorescent aggregates when concentrated in energized mitochondria in response to their higher membrane potential. JC-1 fluorescence is usually excited by the 488 nm laser wavelength common in flow cytometers. In this study, we show that in practice this approach is not optimal for monitoring mitochondrial behavior. Investigation of fluorescence of JC-1 in solution and in cells using spectrofluorimetry, microscopy and flow cytometry reveals that excitation at 405 nm wavelength, now available on standard instruments, produces signals from aggregate fluorescence with considerably less spillover from dye monomer fluorescence than can be obtained using 488 nm excitation. The improved data are more accurate and eliminate the necessity for fluorescence compensation, making the use of the alternative excitation wavelengths beneficial for mitochondria-related biological and biomedial research.

Extinction and scattering by soft spheres.

On the basis of an approximate relation, the Mie series are replaced by new ones, which can be summed exactly with the aid of various modified and generalized forms of the addition theorem for cylindrical functions. The sums obtained simultaneously simplify the initial expressions for the scattering characteristics and preserve their analytical nature. The conventional approximations for the amplitude functions and the efficiency factors rigorously follow from the new approaches if the optical parameters are properly restricted. The acceptable domains of these approaches contain the long wavelength region and are extensive enough to study the various (including inverse) scattering problems for the real disperse systems, in which the particles are suspended in a medium with similar optical properties.

Scattering in spherically symmetric media.

The scattering problem within a multilayered spherically symmetric medium due to a source of perturbation located in the external region is considered. Assuming that the refractive index and its derivative vary continuously, with the exception of a finite number of jump discontinuities, the electromagnetic field vectors are represented in terms of various well-known potentials suitable for describing the azimuthal dependence of the incident wave. The linear dependence between the permittivity and the specific conductivity is proved to be equivalent to the vanishing of the electric charge density. Exact expressions for the field energy characteristics in the external region have been derived without any supplementary suppositions with respect to the wave zone. Scattering from a dielectric coated sphere, whose refractive index is a continuous function while its derivative has two jump discontinuity points (a modified Mie problem), has been studied. The generalized van de Hulst phase angle transformation is introduced and used to show the coincidence of the cross sections for scattering and extinction for the transparent spherical shell mentioned.

Scattering by particles with radially variable refractive indices.

The problem of scattering by a spherical particle whose refractive index arbitrarily depends on the distance from its center has been solved. A computational scheme for determining the scattering coefficients for the refractive-index profiles given by some piecewise smooth function is constructed, and the models to use it are presented. The simple algorithm for evaluating the scattered and internal field vectors is elaborated. The exact expression for the scattering cross section in terms of the generated Debye partial potentials that are dependent on the given refractive index has been obtained.

Different membrane-bound forms of acetylcholinesterase are present at the cell surface of hepatocytes.

In the present study we have determinated the acetylcholinesterase molecular forms present in rat liver hepatocytes; we have also studied the association of acetylcholinesterase with the cell surface of the hepatocytes. Subcellular fractionation indicated that rough endoplasmic reticulum and plasma-membrane-enriched fractions contains G4 and G2 acetylcholinesterase forms bound to membranes. Hepatocytes incubated with phosphatidylinositol-specific phospholipase C released about 70% of the surface acetylcholinesterase. Sedimentation analysis showed that all the solubilized acetylcholinesterase activity comes exclusively from a G2 dimer. The G4 hydrophobic form of acetylcholinesterase accounts for the additional cell-surface activity. The existence of these two forms of acetylcholinesterase on the surface of hepatocytes was further established by analyzing the phosphatidylinositol-specific phospholipase C sensitivity of the acetylcholinesterase molecular forms present in isolated rat liver plasma membranes.

Improvements to the spectral transparency method for determining particle-size distribution.

The previously developed spectral transparency method is improved in two ways: (a) A calculation scheme is developed that enables the consideration of light dispersion within a substance. We abandon the assumption that m(nu) = const and develop a method of calculating N(r) from gamma*(nu) provided that the function m(nu) is specified. (b) A new scheme of processing data on gamma*(nu)is proposed that significantly reduces the oscillation in the answer and requires less initial experimental data.

A simple assay to estimate the acetylcholinesterase molecular forms in crude extracts of rat skeletal muscle.

All the current methods available for analyzing the acetylcholinesterase (AChE) molecular forms are time consuming and require the use of expensive equipment. We have found that by using the differential inactivation of globular (G4 + G1) and asymmetric AChE forms by high Mg2+ concentration, we can set up a very easy and quick assay that allows us to determine the relative proportions of AChE molecular forms present in rat skeletal muscles. This assay will be of great help in estimating changes in the muscle AChE forms under experimental conditions that require several simultaneous determinations.

Efficacy of leuprolide therapy in children with central precocious puberty.

Eight girls with central precocious puberty were treated with the long-acting gonadotropin releasing hormone analogue leuprolide acetate (Lupron) for a period of six to 18 months. Suppression of gonadotropin and estradiol secretion and regression of secondary sexual characteristics and menses were observed while patients received a subcutaneous dose of 35 to 40 micrograms/kg/d. Growth velocity was slowed in all but one patient, and the rate of skeletal maturation was slowed even more, resulting in a stabilization or improvement in predicted adult height. There were no major side effects. Although the long-term effects of leuprolide therapy cannot be determined with this study, it appears to be efficacious in the treatment of central precocious puberty.

Neonatal adaptation: sympatho-adrenal response to umbilical cord cutting.

The present studies were designed to assess the individual effects of delivery and umbilical cord cutting on the stimulation of the sympatho-adrenal system during parturition. Pregnant ewes with time-dated singleton pregnancies were used in an acutely exteriorized fetal lamb model with an intact umbilical circulation. We observed a minimal, transient elevation in plasma catecholamines (CAT) coincident with the operative procedures and delivery. Subsequent cord clamping was observed to evoke a rapid and marked increase of both norepinephrine and epinephrine (E), maximal at 5 min and persisting over the 4-hr study period. Animals could be grouped on the basis of the observed CAT responses, severity of postpartum acidosis, the extent of free fatty acid (FFA) mobilization and degree of postpartum hypothermia. A blunted FFA response and slower correction of hypothermia were observed in the more acidotic animals despite higher CAT concentrations. One group of four animals had high peak CAT concentrations, 32,000 pg/ml norepinephrine and 35,000 pg/ml E, a deep nadir in pH of 6.88 +/- 0.09, a 2-hr delay in maximal FFA mobilization and slower correction of hypothermia. The other group of four animals had peak norepinephrine of 2800 pg/ml and E of 1100 pg/ml, a nadir in pH of 7.09 +/- 0.08, maximal plasma FFA concentration by 1 hr after cord cutting and a higher nadir in body temperature 35.7 versus 32.5 degrees C. The results demonstrate that umbilical cord cutting itself is a potent stimulus for fetal CAT release and FFA mobilization. Acidosis is capable of markedly augmenting E release in the mature fetus and obtunding chemical thermogenesis.

Differential association and distribution of acetyl- and butyrylcholinesterases within rat liver subcellular organelles.

Rat liver cholinesterases were found to share properties and characteristics with those expressed in cholinergic tissues. The distribution and presence of different molecular forms of cholinesterases in different subcellular organelles of rat liver were studied. The rough and smooth endoplasmic reticulum and Golgi apparatus were enriched in the G4 molecular form of acetylcholinesterase (AChE) (relative to the G2 molecular form), while the inverse was found in the plasma membrane. The interaction of these molecular forms of AChE with the Golgi membrane was studied in detail. Approximately one-half of the G4 form was free within the lumen while the remainder was an intrinsic membrane protein; all the G2 molecular form was anchored to the membrane via phosphatidylinositol. Only the G1 and G2 molecular forms of butyrylcholinesterase (BuChE) were found in the above subcellular organelles; both molecular forms were soluble within the lumen of Golgi vesicles. These results indicate that rat liver expresses several molecular forms of AChE which have multiple interactions with membranes and that liver is unlikely to be the source of the G4 form of BuChE present in high concentration in the plasma.