Homologous desensitization of substance-P-induced inositol polyphosphate formation in rat parotid acinar cells.

Maximal concentrations of substance P and methacholine induced a rapid increase in [3H]inositol trisphosphate ([3H]IP3) formation. After about 1 min, the [3H]IP3 in the substance-P-treated cells ceased to increase further, whereas in the methacholine-treated cells [3H]IP3 continued to increase. Addition of methacholine to the substance-P-treated cells caused a rapid increase in [3H]IP3, whereas a second addition of a 10-fold excess of substance P had no effect. Pretreatment of cells with substance P, followed by removal of the substance P by washing, resulted in a decreased response to a second application of substance P. A similar protocol involving pretreatment with methacholine had no effect on subsequent responsiveness to substance P. Analysis of [3H]substance P binding to substance-P-treated cells indicated that the number of receptors for substance P was decreased, but the affinity of the receptors for substance P was unaffected. After substance P pretreatment, a prolonged incubation (2 h) restored responsiveness of the cells to substance P, measured as [3H]IP3 formation, and restored the number of binding sites to control values. These findings indicate that, in the rat parotid gland, substance P induces a homologous desensitization of its receptor, which involves a slowly reversible down-regulation or sequestration of substance-P-binding sites.

Metabolism of inositol 1,4,5-trisphosphate in guinea-pig hepatocytes.

Metabolism of inositol 1,4,5-trisphosphate was investigated in permeabilized guinea-pig hepatocytes. The conversion of [3H]inositol 1,4,5-trisphosphate to a more polar 3H-labelled compound occurred rapidly and was detected as early as 5 s. This material co-eluted from h.p.l.c. with inositol 1,3,4,5 tetrakis[32P]phosphate and is presumably an inositol tetrakisphosphate. A significant increase in the 3H-labelled material co-eluting from h.p.l.c. with inositol 1,3,4-trisphosphate occurred only after a definite lag period. Incubation of permeabilized hepatocytes with inositol 1,3,4,5-tetrakis[32P]phosphate resulted in the formation of 32P-labelled material that co-eluted with inositol 1,3,4-trisphosphate; no inositol 1,4,5-tris[32P]phosphate was produced, suggesting the action of a 5-phosphomonoesterase. The half-time of hydrolysis of inositol 1,3,4,5-tetrakis[32P]phosphate of approx. 1 min was increased to 3 min by 2,3-bisphosphoglyceric acid. Similarly, the rate of production of material tentatively designed as inositol 1,3,4-tris[32P]phosphate from the tetrakisphosphate was reduced by 10 mM-2,3-bisphosphoglyceric acid. In the absence of ATP there was no conversion of [3H]inositol 1,4,5-trisphosphate to [3H]inositol tetrakisphosphate or to [3H]inositol 1,3,4-trisphosphate, which suggests that the 1,3,4 isomer does not result from isomerization of inositol 1,4,5-trisphosphate. The results of this study suggest that the origin of the 1,3,4 isomer of inositol trisphosphate in isolated hepatocytes is inositol 1,3,4,5-tetrakisphosphate and that inositol 1,4,5-trisphosphate is rapidly converted to this tetrakisphosphate. The ability of 2,3-bisphosphoglyceric acid, an inhibitor of 5-phosphomonoesterase of red blood cell membrane, to inhibit the breakdown of the tetrakisphosphate suggests that the enzyme which removes the 5-phosphate from inositol 1,4,5-trisphosphate may also act to convert the tetrakisphosphate to inositol 1,3,4-trisphosphate. It is not known if the role of inositol 1,4,5-trisphosphate kinase is to inactivate inositol 1,4,5-trisphosphate or whether the tetrakisphosphate product may have a messenger function in the cell.

ATP-induced calcium mobilization and inositol 1,4,5-triphosphate formation in H-35 hepatoma cells.

Addition of ATP (but not epinephrine, angiotensin II, vasopressin, or platelet-activating factor) to H-35 hepatoma cells whose cellular lipids have been pre-labelled with [3H]inositol, causes a rapid increase in [3H]inositol triphosphate. In H-35 cells pre-incubated in the presence of 45Ca2+, ATP causes a similarly rapid release of 45Ca2+. The concentration-effect relationships for inositol triphosphate formation and Ca2+ efflux are similar to those reported previously for differentiated hepatocytes. These results demonstrate that at least one of the Ca2+-mobilizing receptors normally found on hepatocytes is functionally retained in the H-35 hepatoma cell line and thus could provide a useful model for the study of these receptor mechanisms in liver.

Children's adrenocortical responses to classroom activities and tests in elementary school.

Levels of free-cortisol excreted by second-grade children during two morning hours on regular school days were compared with levels on days that achievement tests were administered. Cortisol excretion was significantly higher on test days than on normal school days but was not related to the children's self-reports of test anxiety. Children who were slightly above average in intelligence and children who were low achievers were found to have elevated cortisol levels. Sixty-eight percent of the variance in free-cortisol excretion was accounted for by the child's popularity with peers, hostility to the teacher, and on-task behaviors. The usefulness of measuring adrenocortisol responses to stress in the classroom was demonstrated.

Receptor interactions in stimulation of hydrolysis of inositol phospholipids.

The amount of carbachol-induced hydrolysis of inositol phospholipid in the mouse exocrine pancreas is reduced when cholecystokinin octapeptide, an agonist which works through a separate class of receptors, is also present. This effect does not appear to be due to depletion of inositol phospholipid from the cell membranes. These results indicate that the hydrolysis of inositol phospholipid is not always dependent on the number of receptors occupied and that the different classes of receptors share at least one component in the response system.

The contribution of inositol exchange to agonist-stimulated breakdown of myo- [2-3H] inositol-labelled phosphatidylinositol in mouse exocrine pancreas.

Agonists stimulate the release of myo-inositol from phosphatidylinositol (PtdIns) labelled in vivo with myo-[2-3H] inositol. In the presence of lithium, which inhibits myo-inositol-1-phosphatase, the compound which accumulates following the breakdown of pre-labelled PtdIns is inositol-1-phosphate. This indicates that the agonist-stimulated release of the head group from this lipid is not the result of inositol exchange and is due to phosphodiesterase activity. The total amount of 3H-labelled compounds released from PtdIns in the presence and absence of lithium is the same, which indicates the labelled compounds which are released are not re-incorporated. Agonist-induced release of myo-[2-3H] inositol can be used as a reliable indication of PtdIns breakdown in the exocrine pancreas.

The role of calcium in agonist-stimulated hydrolysis of phosphatidylinositol in mouse pancreas.

The effects of Ca2+ on agonist-stimulated hydrolysis of myo-[2-3H]inositol-labelled phosphatidylinositol in mouse pancreas in vitro, were studied. The increase in cytosol Ca2+ concentration produced by the ionophore A23187 did not stimulate the breakdown of phosphatidylinositol. Cholecystokinin-octapeptide (CCK-8) stimulated the hydrolysis of phosphatidylinositol under conditions in which intracellular calcium stores were depleted. The breakdown of phosphatidylinositol was stimulated by bethanechol and CCK-8 in Ca2+ -free Krebs solution, and the addition of Ca2+ to the medium potentiated the effects of these agonists. Lanthanum significantly reduced bethanechol and CCK-8-stimulated hydrolysis of phosphatidylinositol in Krebs solution, but was without effect in Ca2+ -free Krebs solution. The results of this study support the proposal that hydrolysis does not occur as a result of Ca2+ mobilization and may be involved in Ca2+ gating in the pancreas.

Agonist-stimulated breakdown of myo-[2-3H] inositol-labelled phosphatidylinositol in mouse pancreas.

Agonist-stimulated hydrolysis of phosphatidylinositol (PI) in cell membranes has been proposed to lead to an increase of cytosol calcium concentration and activation of the cellular response in certain smooth muscles and glands. A method is described which allows the rapid, reproducible measurement of hydrolysis of phosphatidylinositol in mouse pancreas. The technique involves the in vivo labelling of the pancreas with myo-[2-3H] inositol. The majority of the label incorporated into phospholipids is in the form of PI, with only a small proportion of label in di- and tri-phosphoinositides. Tissue pieces of the labelled pancreata are incubated in vitro in the presence or absence of secretagogues, and the PI in homogenates of these pieces is precipitated with trichloroacetic acid. No PI remains in the acid-soluble supernatant. This technique does not require the time-consuming extraction and chromatographic separation of lipids which has been necessary in other assays of PI hydrolysis. Using this method, we have confirmed that PI breakdown is stimulated by carbachol and cholecystokininoctapeptide in the presence or absence of extracellular Ca2+. Agonist-stimulated hydrolysis of PI was potentiated by extracellular Ca2+, and was not dependent on agonist-activated Na+ influx. This technique will facilitate the investigation of the importance of PI breakdown in stimulus-response coupling.

Mepacrine inhibition of bradykinin-induced contractions of the rabbit ear vein.

Bradykinin induced concentration-dependent contractions in spiral strip preparations of the central vein of rabbit's ears. These contractions require the presence of Ca2+ in the extracellular medium. Mepacrine (2.1 to 42.3 mumol/l) significantly reduced these contractions. Since indomethacin (28 mumol/l) did not reduce the contractions produced by bradykinin, the effect of mepacrine is not explained by inhibition of prostaglandin synthesis. Procaine (10(-7) to 10(-4) mol/l) did not reduce the contractions produced by bradykinin and thus is does not seem that mepacrine acts through a non-specific, local anaesthetic effect. Mepacrine reduced the phosphatidate-mediated transfer of 45Ca from an aqueous phase to an organic phase. It has recently been proposed that phosphatidate may be the Ca2+ ionophore in smooth muscle. However, in the present experiments procaine was about as potent as mepacrine in inhibiting phosphatidate-mediated 45Ca transfer, and since it did not reduce the contractile responses these results do not support the concept that mepacrine inhibits contractions by reducing the Ca2+ influx via a phosphatidate ionophore.

Maternal alcohol consumption, birth weight, and minor physical anomalies.

The effect of maternal alcohol consumption during pregnancy upon neonatal outcome was examined in 278 mother-infant dyads. Accounts of the mother's drug use as well as information about confounding factors were obtained by personal interviews on two occasions during pregnancy and again after delivery. The infants were examined by a standardized procedure for the presence of minor physical anomalies. Alcohol, in moderate amounts, was found to be unrelated to infant birth weight, birth length, or head circumference. Minor physical anomalies associated with fetal alcohol syndrome were no more common in infants exposed to heavy amounts of alcohol in the first trimester than in nonexposed infants. A weak but significant negative correlation between alcohol consumption and gestational age was not accounted for by covariance with emotional stress, parity, socioeconomic status, mother's age, or use of other drugs.

Urinary cortisol excretion rates and anxiety in normal 1-year-old infants.

Urinary cortisol excretion rates were determined for each voiding during 8 hours on 3 days for 20 infants. The first day served as the control. On the second day stress was imposed by having the mother leave the infant for an hour. On a third day the child was stimulated by novel toys and socialization for an hour in the mother's presence. There was no significant difference between mean cortisol excretion rates on control and stress days. Variability in cortisol levels was significantly greater on the day of stress than on the other 2 days. Ratings of separation anxiety and levels of cortisol excreted 40 to 160 min after the imposed stress were positively correlated (r = 0.46, P less than 0.01). Cortisol levels also differentiated between anxious infants who were agitated from those who were withdrawn. No altered levels of cortisol were associated with excitement.

The early childhood development of 17 boys with sex chromosome anomalies: a prospective study.

Seventeen boys with sex chromosome anomalies were identified at birth and followed up developmentally for two to five years. The 17 included 12 boys with 47,XXY, four with 47,XYY, and one with 46,XY/XXY mosaicism. None had major disabilities in physical, intellectual, or emotional growth. Some of the boys with 47,XXY were characterized as tall, inactive, poorly organized in motor functions, placid, and delayed in language; two of the four with 47,XYY were impulsive at 1 year of age. These deviations from normal were relatively mild; the boys were not found to be at high risk for grossly abnormal development in the early years.

Cortisol excretion levels and daytime sleep in one-year-old infants.

Urine was collected from 20 one-year-old infants from the time of awakening in the moring for 8 consecutive h. Free cortisol was measured by competitive protein-binding analysis in each urine sample. The mean cortisol excretion rate for the entire collection period was .228 +/- .019 mug/h/kg. The mean excretion rate of sleep related cortisol (1.86 +/- .28 mug/h) was significantly different from the mean rate for awake periods (2.61 +/- .28 mug/h). The difference could not be attributed to time of day. The results suggest a direct relationship between cortisol production and daytime sleep-wake states in the one-year-old.

Aminocentesis: its impact on mothers and infants. A 1-year follow-up study.

In a 1-year follow-up study, the effects of amniocentesis for genetic diagnosis upon 22 infants and their mothers were assessed. The infants were tested at 1 year of age on the Bayley Infant Scale of Mental and Motor Development. No deviations from normal mental or motor development were found. The mothers were interviewed at the same time, in order to better understand the emotional aspects of the experience. Women between ages 35-39 with no previously affected child tended to accept amniocentesis easily, as an appropriate part of parnatal care. Mothers with a previous defective child tended to have more painful emotional reactions, including higher anxiety over the test and more conflict about a possible abortion. They needed more intensive and extended counseling. We suggest that the issue of abortion is important, even if manageable by most women, and that a chance to discuss it may be significant.

PIP: The effects of amniocentesis for genetic diagnosis were assessed in 22 mothers and their infants in a 1-year follow-up study. Among the infants, there lere no deleterious effects on mental and motor developme nt at 1 year of age. Women aged 35-39 years, whose previous children were normal, tended to accept the procedure easily, while those who already had a congenitally abnormal child were more deeply affected emotionally. These women were particularly anxious about the procedure and the possibility of abortion being indicated. This group required more intensive and longer counseling. It is suggested that the possibility of abortion be freely and openly discussed by the counselor with the amniocentesis patient.

A developmental study of girls with trisomy X.

Development of 11 girls (ages 2-10 years) with 47,XXX karyotype identified in a newborn survey is compared with eight girls having a mosaic sex chromatin pattern and with the normal siblings of each group. Delay in early motor development and speech, a mild intellectual deficit, and disturbance in interpersonal relationships occurred in one-third of the index cases, a higher frequency than in the comparison groups. two-thirds were considered normal and adequately adjusted. No consistent phenotype was found.