Gonadotropin-induced anomalies of the zona pellucida of the baboon ovum.

Vesiclulation and other anomalies were observed in the zona pellucida about the ovum of female baboons (Papio anubis) that had received treatment with Pergonal and human chorionic gonadotropin; and in somiie instances the zona pellucida was absent.

The effects of dexamethasone on the peripheral plasma concentrations of androstenedione, testosterone and cortisol in the pregnant rhesus monkey.

The concentrations of androstenedione, testosterone and cortisol have been measured in the maternal peripheral plasma of normal pregnant rhesus monkeys (Macaca mulatta), and in rhesus monkeys treated daily with dexamethasone during late pregnancy. During the last 30 days of pregnancy, the mean plasma concentrations of androstenedione, testosterone and cortisol were about 1400-2000 pg/m1375-425pg/ml and 300-400 ng/ml, respectively. None of these steroids increased significantly before parturition. As there was no rise in maternal androgens in late pregnancy when plasma estrogens increase sharply, it suggested that his elevation of plasma estrogens is related to an increase of fetal precursors Dexamethasone treatment resulted in 90% suppression of plasma cortisol and 40%-60% suppression of androstendione and testosterone. As the suppression of maternal androgens was of lesser magnitude than the decline of plasma estrogens, and as the androgens did not continue to fall with continuing dexamethasone treatment as did the estrogens, these findings also suggest that the decline in plasma estrogens is related in large part to suppression and atrophy of the fetal adrenals.

Absence of regular pulsatile gonadotropin secretion during implantation in the rhesus macaque.

The secretory response of the primate corpus luteum (CL) to CG after implantation suggests that gonadotropin receptors are not depleted despite increasing CG production and continuous elevated tropic stimulation. Such continuous stimulation is known to cause down-regulation of receptors in other tissues. To determine if CG secretion is intermittent during the initial stages of CL rescue, we assessed the secretory pattern of CG during the periimplantation period by collecting frequent (4/h) blood samples in two studies (for 4 h on 3 separate days between days 8-13, or for 2 separate 13-h sequences between days 10-15 postovulation) in 13 chair-adapted females. Day 0 of gestation was defined as the day of ovulation, as estimated by peak urinary estrone conjugate excretion in females mated on days 9, 11, and 13 of the menstrual cycle. Hormone concentrations were measured by either RIA [irFSH; estradiol and progesterone (P)] or Leydig cell bioassay (bioLH or bioCG). In the first study, 4 of 6 females conceived, and the mean for bioLH was not significantly elevated until days 12-13. In the second study, 5 of 7 females conceived, and the episodic secretory pattern of circulating pituitary bioLH typically observed in cycling females (2.7 +/- 0.3 peaks/13 h) was replaced by a relatively nonpulsatile, but steadily increasing profile during days 12-15 of gestation (1.5 +/- 0.4 peaks/13 h). Although occasional large fluctuations in bioLH/CG and P were noted, the bioLH/P peaks were less congruent than those in nonfertile cycles, and there was no diurnal pattern in the secretion of either hormone. In contrast, irFSH concentrations did not fluctuate and were similar in the two groups of females [2.93 +/- 0.28 vs. 2.34 +/- 1.7 ng/ml (mean +/- SEM)]. These data demonstrate that 1) a steady, gradually increasing secretory pattern of CG is associated with rescue of the CL; 2) the circulating profile of CG during the periimplantation interval is not consistently episodic and does not support the hypothesis that intermittent CG release prevents LH/CG receptor down-regulation in the CL during early pregnancy; 3) increased bioLH/CG levels during conceptive cycles in rhesus monkeys are not detected until days 12-13 after the midcycle bioLH peak; 4) irFSH patterns on pooled aliquots appear to be uninformative with regard to gonadotropin dynamics in early pregnancy; and 5) urinary estrone conjugate measurements provide a practical method for the precise timing of infrequent events, such as implantation, in the laboratory macaque.

Proliferation gradients in the inner ear of the monkey (Macaca mulatta) embryo.

Radioautographic studies were conducted on early developmental stages of the inner ear in monkey (Macaca mulatta) embryos ranging in age from stage 11 to stage 20 (25 to 39 days' gestation) and labeled for one hour with [3H] thymidine. The most active period of proliferation occurred at stage 13, at which time all regions of the otocyst, including the endolymphatic duct, were heavily labeled. In subsequent stages the dorsal portion of the endolymphatic duct failed to incorporate label, whereas proliferation continued in the ventral portion of the duct near its communication with the medial wall of the otocyst. The duct thus appeared to grow upward as a result of cell division from below. Mitotic activity continued in the remainder of the otocyst although there was a progressive temporal decrease in the labeling index. Spatial gradients also occurred, with dorsal and lateral regions less active than ventral and medial ones. As the otocyst differentiated, the cells in various areas became attenuated. These flattened cells were still capable of undergoing cell division, as indicated by the presence of labeled nuclei in all regions of the utricle, saccule, semicircular ducts, and cochlear duct.

Studies of marginal zinc deprivation in rhesus monkeys. VIII. Effects in early adolescence.

Arrested adolescent growth and sexual maturation are striking symptoms of severe dietary zinc deprivation. More general implications of mild or marginal Zn deficiency during adolescence are not known. Five marginally Zn-deprived (ZD) male monkeys (4 mg Zn/kg diet) were compared with five controls pair fed a diet containing 100 mg Zn/kg during early adolescence. Mean plasma Zn levels were 38% lower in ZD group than in controls when evaluations began. During rapid growth plasma Zn decreased in controls but not ZD animals. ZD animals had delayed onset of accelerated weight gain and linear growth; loss of subcutaneous fat typical of early adolescence did not occur. ZD monkeys required two to three times more trials for both learning and reversal a visual discrimination task. Immune function was depressed 20-30% as reflected in early reduced proliferative response of peripheral lymphocytes and later lower immunoglobulin levels. Marginal dietary Zn deprivation affects growth and function in adolescence without producing frank developmental retardation.

Long-term marginal zinc deprivation in rhesus monkeys. II. Effects on maternal health and fetal growth at midgestation.

Adult nonpregnant female rhesus monkeys fed purified diets containing 100 or 4 ppm zinc for 1 yr were mated then studied through midgestation. At mating, zinc-deprived (ZD) monkeys showed maternal lymphocyte response to mitogens concanavalin A (Con A) and phytohemagglutinin (PHA), serum uric acid and carbon dioxide, and WBC lower than in control (C) monkeys. There was a significant positive association between plasma zinc and PHA response. At midgestation, discriminant analysis revealed that maternal lymphocyte response to Con A, fetal abdominal circumference (by ultrasound), plasma fibrinogen, serum IgM, and amniotic fluid iron level were discriminators for diet group, all lower in ZD than in C monkeys. Maternal plasma and RBC zinc at midgestation were positively associated with fetal growth and plasma uric acid. These observations suggest that immune function (ie, mitogen response and serum immunoglobulin level) is a strong discriminator of dietary zinc deprivation in rhesus monkeys, both before and during pregnancy.

Effect of phytate removal on zinc absorption from soy formula.

Low zinc bioavailability from soy formula may be the result of the formula's phytate content. We assessed the effect of phytate removal from soy formula on Zn absorption using infant rhesus monkeys and suckling rat pups as animal models. Zn absorption in monkeys, as determined by whole-body counting, was 65% from human milk, 54% from monkey milk, 60% from whey-predominant formula, 46% from casein-predominant formula, and only 27% from conventional soy formula (0.621 mmol phytate/L). In contrast, Zn absorption from dephytinized soy formula (0.067 mmol phytate/L) was 45%. In suckling rats, Zn absorption from conventional soy formula was only 16% vs 47% from dephytinized soy formula. Phytate concentration in a variety of experimental soy formulas was inversely correlated to Zn absorption. These results suggest that the low bioavailability of Zn from soy formula is a function of its phytate concentration and can be overcome by the removal of phytate.

Activity and attention in zinc-deprived adolescent monkeys.

Lethargy is characteristic of malnourished populations, but little is known about the biologic mechanism or consequences for cognitive performance. In the current experiment, 24-h activity patterns and performance of an attention task were studied in adolescent female monkeys (18-33 mo of age, n = 10/group) under conditions of moderate dietary zinc deprivation (2 micrograms Zn/g diet) and adequate dietary zinc (50 micrograms Zn/g). There were progressive decreases in daytime activity levels in the zinc-deprived group followed by slowing of growth around the time of the growth spurt. Attention performance was also impaired before the onset of growth retardation. Zinc-deprived monkeys failed to show the shift to later initiation of the rest phase of the diurnal cycle seen in controls in late adolescence. These data support previous findings that activity and attention can be affected during early stages of zinc deprivation before the onset of growth retardation.

Studies of marginal zinc deprivation in rhesus monkeys. I. Influence on pregnant dams.

Rhesus monkeys were fed a diet marginally deficient in zinc (4 ppm zinc) throughout pregnancy and were monitored for changes in hematological, biochemical, and immunological parameters. This dietary zinc level was chosen because it did not produce an overt deficiency syndrome when fed for 10 wk to nonpregnant monkeys. Deprived animals were compared to control groups fed a zinc replete (100 ppm) diet ad libitum or on a food restricted (pair fed) basis. Beginning in the 3rd trimester zinc-deprived monkeys exhibited characteristic signs of deficiency including dermatitis, anorexia, and low levels of plasma zinc (less than 65 micrograms/100 ml) and of serum alkaline phosphatase activity. The extent of plasma zinc depression in deficient monkeys was dependent on total food intake; severely anorexic monkeys lost weight but maintained normal plasma zinc levels; monkeys that gained 20 to 30% of their body weight during pregnancy had severely depressed plasma zinc. Plasma vitamin A was reduced in the deprived group while copper, magnesium, and folate levels remained similar to controls. Hematological changes indicative of iron deficiency anemia (reduced packed cell volume, mean corpuscular volume, and Hb) were also seen in severely deficient monkeys. In addition, the peripheral lymphocyte mitogen response was reduced in deficient dams. We conclude that marginal deficiency of dietary zinc can produce significant abnormalities of nutritional status and has the potential for producing serious immunohematological dysfunction during pregnancy.

Studies of marginal zinc deprivation in rhesus monkeys. IV. Growth of infants in the first year.

Growth was evaluated in rhesus monkey infants that were the offspring of females given a marginally zinc-deficient diet (4 micrograms/g zinc) from the beginning of pregnancy and through 12 months of postnatal life. These zinc-deficient (ZD) infants were compared to controls whose mothers were fed a complete diet, either ad libitum or pair-fed to zinc-deficient dams, throughout gestation and lactation. Male ZD infants had evidence of growth retardation at birth. In contrast, growth retardation in female ZD infants was not observed until 1 month of age. From 3 to 9 months of age (late lactation and subsequent to weaning) ZD infants attained weights similar to those of the control group. However, analysis of crown-rump and femur length indicated that ZD infants' growth was less than optimal throughout the entire 1st yr of observation. In addition, skinfold thickness was markedly higher in ZD than in control infants in the postweaning period. In the juvenile period (9-12 months of age) both male and female ZD animals fell behind controls in body weight. ZD juveniles were also hypogeusic, as determined by a quinine acceptance test. Low weight ZD infants had reduced somatic growth as reflected in sitting height, long bone growth, head circumference, and limb circumference. Regression analysis indicated that impaired growth rates from 9 to 12 months were associated with both lower food intake and reduced food use efficiency. Plasma zinc concentration was, in general, inversely related to weight gain in both groups during the 1st yr.

Studies of marginal zinc deprivation in rhesus monkeys: infant behavior.

Rhesus monkey infants were marginally deprived of zinc (4 ppm diet) from conception and were compared to controls (100 ppm diet) during the first year of life in development of reflexes and motor patterns, mother-infant interaction, delayed response performance, discrimination learning and reversal, and open field behavior. Deficits in amount and variety of behavior were recorded in deprived infants; spontaneous locomotor activity was 50% below control levels in males at 1 mo of age; spontaneous activity was 7-10% lower in both males and females at 3 mo of age; response latencies were 50% lower than controls at 7-9 mo; failure to reach discrimination reversal criterion was seen in 71% of deprived infants as compared to 10% of controls at 10 mo of age; and abnormally low levels of climbing and exploration were seen in two of six deprived infants at 12 mo of age. No abnormalities in the rate of behavioral development or in emotional adaptability were observed. These and other results suggest that syndromes of lethargy, apathy, and hypoactivity are characteristic of behavioral effects of marginal zinc deprivation in primates.

Studies of marginal zinc deprivation in rhesus monkeys: II. Pregnancy outcome.

A marginal state of zinc deficiency was induced in the pregnant nonhuman primate, Macaca mulatta, by feeding a diet containing 4 ppm zinc beginning at conception. Pregnancy outcome of marginally zinc-deficient monkeys (ZD) was compared to both pair-fed (PF) and ad libitum fed (AL) control animals (100 ppm zinc). Stillbirths, abortions, and delivery complications were more frequent in both ZD and PF dams than in AL controls; no malformations were detected (maternal plasma zinc was normal during organogenesis). Male ZD neonates weighed significantly less than same sex controls; also, in relation to colony norms, 7/8 ZD males, 2/8 ZD females, and 1/10 PF controls were of low birth weight. Further, plasma zinc and iron levels were lower in ZD neonates than in AL and PF controls. ZD neonates also had reduced muscle tonus. Birth weight and maternal plasma zinc concentration were negatively correlated in ZD group but positively correlated in AL and PF groups. Indeed, maternal plasma zinc concentration alone did not identify a state of zinc deficiency which impaired fetal growth in monkeys.

Extracellular superoxide dismutase activity is affected by dietary zinc intake in nonhuman primate and rodent models.

Assessment of zinc nutriture is often compromised by the lack of reliable biomarkers. In the present study the effect of dietary zinc deprivation on plasma extracellular superoxide dismutase (EC SOD) activity was investigated in rat and rhesus macaque models. This enzyme, which contains both zinc and copper, is distinct from cytosolic copper-zinc SOD. Young, growing rats fed zinc-deficient diets (1.5 nmol Zn/g diet) were characterized by low plasma zinc concentrations and plasma EC SOD activities (16% and 56% of controls, respectively). Adolescent rhesus macaques fed diets that contained a marginal amount of zinc (30.6nmol Zn/g diet) also had low plasma zinc concentrations and low EC SOD activities compared with controls fed diets containing 765 nmol Zn/g diet (75% and 40%, respectively). Enzyme activity was not affected after in vitro addition of zinc to plasma samples from control, restrict-fed, and zinc-deficient rats. Taken together, these data support the concept that plasma EC SOD activity can be a biomarker for zinc status.

Modulation of behavioral performance of prepubertal monkeys by moderate dietary zinc deprivation.

Young rhesus monkeys (Macaca mulatta, n = 14) 25-30 mo of age were fed a zinc-deficient (ZD) diet (2 micrograms Zn/g diet) (moderate zinc deprivation) over 15 week. The ZD diet period was compared with a zinc-adequate (ZA) diet period (50 micrograms Zn/g diet) of the same duration, which either preceded or followed it (crossover design). Plasma zinc was lower at the end of the ZD than after the ZA period. There were no overt signs of zinc deficiency or effects on growth rate. Spontaneous motor activity was lower and performance of a visual-attention task and short-term-memory task were poorer during the ZD period than during the ZA period. Behavioral effects were detected as changes from individual baseline values, and often represented a failure to improve. The results suggests that dietary zinc deprivation can modulate behavior in prepubertal monkeys without affecting growth.

Long-term marginal zinc deprivation in rhesus monkeys. I. Effects on adult female breeders before conception.

To assess long-term effects of marginal zinc deprivation on pregnancy, adult non-pregnant female rhesus monkeys were fed diets containing 100 or 4 ppm zinc for 1 yr. then mated; effects on pregnancy and its outcome are under study. During this year, food intake was not depressed in zinc-deprived (ZD) monkeys, and there were relatively few effects on biochemical or hematological indices. By the end of the year, plasma zinc concentration was somewhat lower in ZD monkeys than in controls. Several immune variables, including serum IgM and IgG levels and polymorphonuclear leukocyte (PMN) function, were depressed in the ZD group, changes closely reflecting circannual fluctuations in plasma zinc levels in both diet groups. Endotoxin-activated plasma from ZD monkeys had less potential to promote chemotaxis than that from control monkeys, suggesting that defective PMN function may relate to a plasma effect. Marginal zinc deprivation may thus influence immune function before other variables are affected.

Long-term marginal zinc deprivation in rhesus monkeys. IV. Effects on skeletal growth and mineralization.

Skeletal maturation was evaluated from ages 1 to 3 y in rhesus monkeys that had been subjected to a diet marginally deficient in zinc (4 micrograms/g Zn) from conception through age 3 y. Skeletal development was assessed at 18, 24, 30, and 36 mo of age and compared with that of controls fed ad libitum. Skeletal maturation was determined by the presence of epiphyseal ossification centers. To evaluate endochondral bone mineralization the appearance of the zone of provisional calcification on the metaphyseal side of the growth plate and the width of the growth plate were observed. Marginal Zn deprivation was associated with delayed skeletal maturation in monkeys up to age 3 y. Defective mineralization of bone was evident in these monkeys up to age 6 mo. Between ages 6 mo and 3 y bone mineralization increased in some of the marginal-Zn monkeys to values that were only slightly below those for control monkeys.

Ultrasound evaluation of fetuses of zinc-deprived monkeys (Macaca mulatta).

Fetal body movements were studied in three groups of gravid rhesus macaques fed different amounts of dietary zinc (100 micrograms Zn/g diet, control, n = 12; 4 micrograms Zn/g diet, marginal deprivation, n = 7; 2 micrograms Zn/g diet, moderate deprivation, n = 11). Sonographic examinations were conducted during the third trimester in awake chair-restrained dams. Movement categories, derived from the human biophysical profile, were motor activity (trunk and limb movements), startle, and breathing movements. Moderately deprived fetuses were more active than controls on gestational day (GD) 115; maternal plasma zinc concentrations were significantly correlated with fetal activity at this time. In addition moderately deprived fetuses exhibited fewer breathing episodes on GDs 115-135. Biometrics measures indicated growth retardation in one moderately deprived fetus. These data suggest that moderate, but not marginal, dietary zinc deprivation influences fetal status as evaluated by sonography.

Adolescent growth and maturation in zinc-deprived rhesus monkeys [see comment].

Growth retardation has been associated with zinc deficiency in adolescent human populations, but animal models were not available previously to explore this syndrome. Moderate dietary zinc deprivation (2 micrograms Zn/g diet) was introduced in female rhesus monkeys (Macaca mulatta; n = 10) from the beginning of puberty through menarche. Subgroups of animals (n = 4) continued to be fed the zinc-deficient diet through 45 mo of age (sexual maturity). Reduced weight gain and linear growth and lower plasma zinc concentrations (11.8 +/- 0.6 and 9.2 +/- 0.8 mumol/L in control and zinc-deficient monkeys, respectively) were evident during the premenarcheal growth spurt. Slower skeletal growth, maturation, and mineralization were recorded in the postmenarcheal period and some indicators of sexual maturation were delayed. Food intake was slightly higher in the zinc-deficient group than in controls. These data confirm that adolescent growth and maturation are vulnerable to disruption by moderate dietary zinc deprivation in nonhuman primates.

Studies of marginal zinc deprivation in rhesus monkeys. V. Fetal and infant skeletal effects.

Skeletal maturation was evaluated in newborn and infant rhesus monkeys that had been subjected to a marginally zinc-deficient diet (4 ppm zinc) from conception through 12 months of postnatal life. Serial radiographic assessment of skeletal development was performed and compared to both ad libitum and pair-fed controls. Radiographs were obtained at birth and at 1, 3, 9, and 12 months of age. In each age group a maturation indicator was selected to identify individuals with abnormal skeletal maturation defined on the basis of presence of epiphyseal ossification centers. Animals were compared only within a given sex group. Additionally, to evaluate endochondral bone mineralization, the appearance of the zone of provisional calcification on the metaphyseal side of the growth plate and the width of the growth plate were assessed. A marginal level of zinc deprivation during gestation and during the 1st yr of life was found to be associated with significantly delayed skeletal maturation and defective mineralization. This abnormality of bone mineralization has many features similar to human rachitic syndromes and suggests that zinc plays an important role in endochondral bone formation.

Studies of marginal zinc deprivation in rhesus monkeys: VI. Influence on the immunohematology of infants in the first year.

Rhesus monkey infants fed a marginally zinc-deficient diet (4 ppm) from conception through 12 mo of postnatal life were monitored for changes in hematological, biochemical, and immunological parameters. These zinc-deprived (ZD) infants were compared to control infants whose mothers were fed a zinc-replete (100 ppm) diet either ad libitum (AL) or pair-fed (PF) throughout gestation and lactation. Blast transformation of peripheral blood lymphocytes to phytohemagglutinin (PHA-P), concanavalin A (Con A), and pokeweed mitogen (PWM), was dramatically depressed in the zinc-deficient (ZD) group. Similarly, ZD infants had reduced polymorphonuclear leukocyte function as measured by chemotaxis to endotoxin-activated plasma and phagocytosis of Candida albicans. Levels of serum IgM were significantly altered in zinc-deficient infants compared to controls. Serum concentrations of IgG and IgA were similar in zinc-deficient and control infants. ZD infants also manifested a hypochromic microcytic anemia at one month of age, reduced activity of the zinc metalloenzyme alkaline phosphatase, and lower activity of SGPT.