The effect of tyrosine-deficient total parenteral nutrition on the synthesis of dihydroxyphenylalanine in neural tissue and the activities of tyrosine and branched-chain aminotransferases.

The poor solubility of tyrosine (Tyr) limits the amount of this amino acid in total parenteral nutrition (TPN). In rats maintained on a standard pediatric TPN mixture, plasma and brain concentrations of Tyr are reduced to about 25% of the levels in chow-fed controls. To determine whether these low concentrations of Tyr affect the synthesis of catecholamines in neural tissue, the rate-limiting step (conversion of Tyr to dihydroxyphenylalanine [DOPA]) is studied by administering NSD-1015 to block the pyridoxal phosphate (PLP)-dependent decarboxylation of DOPA. However, in TPN rats, plasma concentrations of Tyr are increased by drug treatment. Because brain Tyr is also increased, these and other experiments using NSD-1015 clearly overestimate the rate of DOPA synthesis for drug-free rats on TPN. Nevertheless, in TPN rats, there is less DOPA in the brain in one experiment and less DOPA in the olfactory bulbs in another, versus control rats. Further examination of the metabolic effects of NSD-1015 reveals that the drug also elevates the concentration of branched-chain amino acids (BCAAs) in the plasma of TPN rats. These findings result from inhibition by NSD-1015 of the PLP-dependent aminotransferases that initiate catabolism of Tyr in the liver and BCAAs in the muscle. Despite the pronounced reduction in plasma Tyr, TPN rats showed a marked increase in the activity of hepatic Tyr aminotransferase compared with chow-fed controls. Conversely, although TPN elevates BCAA concentrations in plasma, the activity of branched-chain aminotransferase (BCAT) in the heart muscle of TPN rats is not different from control values. Different values but the same relationships are seen in drug-free rats.

Melatonin rhythms and pineal structure in a tropical bat, Anoura geoffroyi, that does not use photoperiod to regulate seasonal reproduction.

It has been hypothesized that pineal structure and function might differ between temperate zone and tropical species of mammals because of lower amplitudes of seasonal change in photoperiod and, in some areas, less seasonal climatic variation. Anoura geoffroyi produce a single offspring in November or December of each year on the Caribbean island of Trinidad, at 10 degrees N latitude in the deep tropics. Previous work has shown that this population lacks reproductive responses to photoperiod, and must be enforcing seasonal breeding using a non-photoperiodic cue. Anoura geoffroyi have a minute, thin, and rod-like pineal gland. Throughout much of its length, the pineal courses irregularly within the ventrolateral wall of the great cerebral vein. This intimate relationship may have functional implications. Despite having a very small pineal gland, this species produced a nocturnal rise in serum melatonin. Serum melatonin levels in most individuals were below or near undetectable levels during the light period and rose to a peak averaging 100 pg/ml in the last third of the dark period. Our results indicate that, although the pineal gland of A. geoffroyi is extremely small, serum melatonin levels are comparable to those of other mammals.

Melatonin rhythms and pineal structure in a tropical bat, Anoura geoffroyi, that does not use photoperiod to regulate seasonal reproduction

It has been hypothesized that pineal structure and function might differ between temperature zone and tropical species of mammals because of lower amplitudes of seasonal change in photoperiod and, in some areas, less seasonal climatic variation. Anoura geoffroyi produce a single offspring in November or December of each year on the Caribbean island of Trinidad, at 10 degrees N latitude in the deep tropics. Previous work has shown that this population lacks reproductive responses to photoperiod, and must be enforcing seasonal breeding using a non-photoperiodic cue. Anoura geoffroyi have a minute, thin, and rod-like pineal gland. Throughout much of its length, the pineal courses irregularly within the ventrolateral wall of the great cerebral vein. This intimate relationship may have functional implications. Despite having a very small pineal gland, this species produced a nocturnal rise in serum melatonin. Serum melatonin levels in most individuals were below or near undetectable levels during the light period and rose to a peak averaging 100 pg/ml in the last third of the dark period. Our results indicate that, although the pineal gland of A. geoffroyi is extremely small, serum melatonin levels are comparable to those of other mammals.(AU)

Intravenous gamma-glutamyl-tyrosine elevates brain tyrosine but not catecholamine concentrations in normal rats.

A number of clinical situations may benefit from intravenous supplements of tyrosine (Tyr). In total parenteral nutrition (TPN), the supply of Tyr is limited by its poor solubility. In both rats and infants maintained on pediatric TPN, plasma Tyr levels are approximately 30% of normal, and in rat brains Tyr concentrations are similarly reduced. We reported previously that supplementing a TPN solution with the soluble peptide, gamma-glutamyl-Tyr [Glu(Tyr)], normalizes plasma Tyr and doubles brain Tyr in rats. To assess more fully the behavior of intravenous Glu(Tyr) in vivo, 20 mmol/L Glu(Tyr) was infused into the inferior vena cava of rats at rates increased every 2 hours over an 8-hour period (300 to 450 mumol Glu(Tyr)/kg body weight/h). The surgical procedure for catheterization is described. At the maximum rate of infusion, plasma Tyr and Glu(Tyr) concentrations reached mean plateau values of 326 and 252 mumol/L, respectively. Brain Tyr concentrations were 71 and 264 nmol/g wet weight in control rats infused with heparinized saline (SAL group) and rats infused with Glu(Tyr) (PEP group) respectively. No differences were found in concentrations of norepinephrine (NE), dopamine (DA), or homovanillic acid (HVA) in prefrontal cortex (PFC), striatum (STR), or remaining brain (RB) tissue in PEP and SAL rats. We did not detect undergraded Glu(Tyr) in the brain, and less than 0.5% of infused Glu(Tyr) appeared in the urine.

Observations on the pineal gland of the big brown bat, Eptesicus fucus: possible correlation of melanin intensification with constant darkness.

BACKGROUND: Our initial observation of the macroscopically pigmented pineal gland of the big brown bat, Eptesicus fuscus, led to this study. Information has been lacking on pigmentation in the pineal and its significance in mammals in general and bats in particular. This report begins to address this situation. METHODS: Bats were examined both in the wild and after exposure to various experimental conditions. The pineals were examined macroscopically as well as with light and electron microscopy. The pigment was identified as melanin by its color, the ultrastructure of its granules, and their reaction with hydrogen peroxide. RESULTS: Gross observations showed the pineals to be variably pigmented, which were subjectively scored from unpigmented to heavily pigmented. Pineals from bats exposed to a continuous 24 h light regimen or those from a summer population contained very little, if any, externally visible melanin. Such pineals are considered unpigmented in this study. In contrast, pineals from 74% of 156 animals taken together, either subjected to constant darkness or hibernation (simulated or natural), exhibited very heavily pigmented pineals. The pigment in these cases even extended to the juxtapineal meningeal covering. The pineal was pigmented even in a newborn Eptesicus. CONCLUSIONS: The pineal pigmentation in the big brown bat appears to intensify with constant darkness and may vary seasonally. The observation of macroscopically pigmented pineals in some other bats (Myotis lucifugus, Pipistrellus subflavus, and Lasiurus borealis) suggests that this phenomenon may be of taxonomic value for the family Vespertilionidae (Order Chiroptera).

Use of the soluble peptide gamma-L-glutamyl-L-tyrosine to provide tyrosine in total parenteral nutrition in rats.

Limited solubility restricts amounts of tyrosine (Tyr) in amino acid solutions used in total parenteral nutrition (TPN). Excess phenylalanine (Phe) is included in TPN for conversion to Tyr by liver Phe hydroxylase. However, this conversion is limited, especially in infants. We have confirmed that infants receiving TPN have low Tyr concentrations and high Phe/Tyr ratios in plasma compared with published values for enterally fed neonates. Tyr is important in the synthesis of proteins and other biomolecules, including catecholamines in the brain. We tested the soluble peptide gamma-glutamyl-tyrosine (Glu(Tyr)) as a possible precursor of Tyr in TPN. Groups of five rats were given infusions of TPN containing an amino acid mixture simulating a commercial formulation (group A), TPN in which Glu(Tyr) was substituted for half the Phe in the group A solution) (group B), or saline (group C). Control animals (group C) were fed rodent chow. Blood was sampled at 0 time and daily for 4 days. Brains were collected at 96 hours, and aromatic amino acids in plasma and brains were measured by high-performance liquid chromatography. Throughout the experiment, plasma of animals in group A had significantly elevated Phe and reduced Tyr concentrations compared with control values; plasma concentrations in groups B and C were similar. In groups A and B, brain Tyr levels were 31% and 63% of control values, respectively. In group B, Glu(Tyr) was not detected in brains. These data suggest that supplementing current TPN mixtures with Glu(Tyr), which is stable in solution, can produce normal plasma Tyr concentrations and Phe/Tyr ratios and improve the supply of Tyr to the brain.

Increased binding of epidermal growth factor at preimplantation sites in mouse uteri.

The presence of receptors for epidermal growth factor (EGF) in nonpregnant uteri and the elevation of EGF levels in blood during early pregnancy suggest that EGF and its receptor may play important roles in the early stages of pregnancy. We determined the distribution of EGF receptors in uteri of nonpregnant and pregnant mice during the late preimplantation period (days 4.5-5.0 of pregnancy) using radioautograph and quantitative binding techniques. Radioautography of [125I]EGF binding to cornua from nonpregnant mice showed low levels of specific binding evenly distributed throughout the cornua. In contrast, radioautographs of cornua from pregnant mice showed bands of elevated binding encircling the lumen at sites of preimplantation. Results from radioautography were supported by quantitative analysis of [125I]EGF binding to uterine homogenates from nonpregnant and pregnant mice. Binding of [125I]EGF to uterine membranes was highly specific and time dependent. The average level of specific EGF binding calculated from Scatchard plots of nonpregnant uteri (27 +/- 13 fmol/mg protein) was significantly (P less than 0.05) lower than that in pregnant superovulated uteri (106 +/- 67 fmol/mg protein). Furthermore, specific binding of EGF was significantly (P less than 0.05) higher in preimplantation sites than in the intervening nonimplantation regions from the same uteri (42 +/- 6 vs. 29 +/- 4 fmol/mg protein, respectively). Differences in EGF binding appear to be due to changes in the number of EGF receptors, since half-displacement values (1 nM) were similar in all samples. These results demonstrate that alterations of EGF receptor levels occur at sites where implantation will occur in mouse uteri and support the concept that the transforming growth factor-alpha/EGF receptor and its ligands are involved in implantation of concepti.

Effect of hypoxic hypoxia on taurine concentrations in ventricles of mouse hearts.

The effects of hypoxic hypoxia on the concentration of taurine in right ventricles was studied in the hearts of male CF1 mice caged individually and maintained for 16 hr per day in a hypobaric chamber evacuated to an air pressure of 307 mm Hg. After 23 days hearts were excised and right and left ventricles were separated and lyophilized. Hematocrits in chamber animals were 77-82%, compared to 45-49% for control mice. Mean weights of right ventricles of animals from the chamber were 11.2 +/- 0.9, compared to control values of 7.0 +/- 0.4, mg dry weight. The mean dry weights of left ventricles in both groups of animals were the same. There were no significant differences in the nmoles taurine per mg day tissue in either heart chamber, with mean values +/- S.E.M. of 124.0 +/- 4.6 and 135.0 +/- 4.5 in right ventricles and 128.0 +/- 4.3 and 110.9 +/- 15.3 in left ventricles of experimental and control animals respectively. Thus, hypertrophy which results from hypoxia is not accompanied by increased concentrations of taurine in right ventricles.

Intrauterine proximity to male fetuses predetermines level of epidermal growth factor in submandibular glands of adult female mice.

Morphological, behavioral, and physiological masculinization of adult female mice that developed in utero between two male fetuses (2M females) has been previously attributed to the significantly higher concentration of testosterone in their fetal blood and amniotic fluid than that in female mice which had not been contiguous to males in utero (0M females). Serum testosterone levels of adult 2M and 0M females are not significantly different. To determine whether exposure of fetuses to different levels of testosterone during prenatal development alters adult biochemical parameters of a system responsive to testosterone, the level of epidermal growth factor (EGF) was measured by radioreceptor assay in the submandibular glands of adult CF-1 mice of known intrauterine position. The concentration of EGF was significantly higher (P less than 0.05) in the glands of 2M females (mean +/- SEM, 0.36 +/- 0.14 nmol/mg dry wt of tissue; n = 6) than that in 0M females (0.05 +/- 0.00 nmol/mg dry wt; n = 8). In contrast, EGF concentration did not differ significantly between the glands of 2M and 0M males (0.51 +/- 0.01 and 1.10 +/- 0.42 nmol/mg dry wt, respectively). EGF levels were also determined in the submandibular glands from adult animals of unknown intrauterine position which were gonadectomized and then treated with testosterone and estradiol. The concentrations of EGF in the glands of gonadectomized males and females were similar (0.13 +/- 0.01 and 0.23 +/- 0.09 nmol/mg dry wt, respectively). However, there was a significant difference in response to hormonal administration between males and females. The response of females exceeded that of males at 400 and 800 micrograms testosterone/day. These results suggest that the hormonal environment of a fetus, specifically modification of the fetal environment by the production of hormones from adjacent fetuses, is a major factor in the adult expression of testosterone-responsive proteins such as EGF.

Taurine in hearts and bodies of embryonic through early postpartum CF1 mice.

The hearts and remaining bodies of embryonic and fetal mice of known gestational age and of neonatal mice up to the age of 8.5 days were freeze-dried, weighed, and analyzed for the amino acid, taurine, by high performance liquid chromatography. Although cardiac taurine is only a small fraction of the taurine in the rest of the body in all animals studied, the concentration of taurine in the heart is similar to that in the rest of the body (40-45 nmole/mg freeze-dried wt) in embryos through Day 14.5 of gestation. Cardiac taurine concentration then begins to exceed that of the remainder of the body which gradually declines throughout the period studied. A doubling of cardiac taurine concentration is seen at birth (Day 19.5) when the cardiac to body taurine ratio rises markedly and is maintained at 2-4 throughout the period of observation. A maximum concentration of cardiac taurine (110 nmole/mg freeze-dried wt) is recorded 2.5 days after birth. The dramatic increase in cardiac taurine concentration at the time of birth follows the reported appearance in neonatal mouse hearts of adult levels of beta-adrenergic receptors and the increased work load of the heart.

Further evidence for behaviorly induced hypoxic conditions in subordinate mice.

When two male mice fight to establish social rank, the subordinate exhibits marked splenomegaly, reticulocytosis and decreased hematocrit. Individuals were isolated at weaning, paired 3 weeks later for two 30-min encounter periods daily for up to 3 weeks. After 2 days and 1 week encounter periods, subordinates had significantly decreased levels of hemoglobin in the kidney compared to dominants and controls. Subordinates had increased ATP levels in blood after 1, 2 and 3 weeks of encounter, bone marrow erythroid hyperplasia, as well as a substantial increase in the erythropoietin titre and Fe99 incorporation in spleen and blood.

Changes in concentrations of taurine in murine Harderian glands and retinas during postnatal development.

Quantitative measurements of taurine in the Harderian glands of male and female CF-1 mice were made by standard methods of amino acid analysis. Values, in nmoles per mg dry weight of gland, were 60 to 70 in the first 4 postnatal days with a gradual decrease to 31.1 +/- 3.5 at 12 days, just prior to eye opening. This was similar to the value of 27 +/- 1 observed in sexually mature males, 53 days old. Two to 11 animals were used to establish each value. Other free amino acids in the glands are present in much lower concentrations, difficult to quantitate. Comparisons of the nmolar concentrations of taurine in dry retinas and Harderian glands from the same animals revealed two to three fold higher concentrations in the retina until the time of eye opening, after which concentrations increased markedly to greater than 160 nmoles per mg dry retina at 16 days and older.