Conformational change of human lens insoluble alpha-crystallin

BACKGROUND: It is useful to measure the luminal concentration of drugs which act in the gut. Dialysis of the rectum has not previously been used or validated for this purpose. AIM: To determine the precision of rectal dialysis for measuring rectal drug concentrations. METHODS: To establish the duration of dialysis required to approach equilibrium, the rate of methotrexate diffusion into dialysis bags was first determined in vitro. The precision of rectal dialysis for sampling the methotrexate concentration of colonic lumen extracellular fluid was determined in seven subjects who underwent two consecutive dialysis procedures. Subjects treated with subcutaneous methotrexate for refractory inflammatory bowel disease were studied. RESULTS: Methotrexate crossed the dialysis membrane by a first-order process, and after a 2 h in vitro dialysis, equilibration was 74 +/- 2% (mean +/- s.d.) complete. Rectal dialysis was well tolerated by all subjects. The mean +/- s.e. methotrexate concentration of 3.6 +/- 1.1 nmol/L in the first dialysate was not significantly different from 3.6 +/- 0.9 nmol/L in the second dialysate. P = 0.99 (paired two-tailed t-test). Similar precision was obtained for an endogenous molecule, potassium, secreted by the rectal mucosa. CONCLUSIONS: Dialysis of the rectum is a well tolerated and precise technique for sampling the colonic lumen extracellular fluid for quantitative analyses of exogenous and endogenous substances.     

Ontogeny of morphine withdrawal in the rat.

The present studies examined behavioral changes during precipitated morphine withdrawal in 7- to 42-day-old rat pups. One group of rats was injected with morphine sulfate (10.0 mg/kg) twice daily for 6.5 days. Another group of 7-day-old rats received a lower dose of morphine (3.0 mg/kg). Controls were saline injected or untreated litters (7-day-old pups only). On Day 7, a target pup was injected with saline or naltrexone (0.3–20.0 mg/kg). Preweaning pups were observed in a warm chamber with the litter. Forty-two-day-old rats were tested individually. Morphine-treated pups tested with naltrexone showed significant alterations in behavior that varied at different ages. For example, rolling, stretching, and head and paw moves were observed at the younger ages, whereas burrowing, diarrhea, jumps, teeth chatter, and wet dog shakes occurred in the older rats. These data indicate that morphine-abstinent rats demonstrate withdrawal signs that are within the developmental repertoire of the rat. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Ontogeny of thermoregulatory mechanisms in the rat.

The ability of rats to select a warm environment was studied as a function of postnatal age (birth to 13 days), using 100 Long-Evans hooded rats as Ss. Ss younger than 5 days demonstrated no choice response (movement to a warm compartment, 36-37 .C); however, they did demonstrate movement within the start compartment (23 .C). Increasing the motor capabilities of the pups, by injection of levodopa (50 mg/kg), elicited a choice response in 4-5 day-old Ss. Younger Ss demonstrated no choice of a warm environment even though they moved considerably. Also there was no difference between levodopa-treated and control Ss in the magnitude of temperature change in pups isolated from their mother for 1 hr. The evidence suggests development of behavioral thermoregulatory mechanisms prior to abilities for internal regulation. (22 ref) (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Immunohistochemical study of calpain-mediated alpha-crystallin proteolysis in the UPL rat hereditary cataract

The purpose of this study is to examine if microencapsulated PC12 cells may provide long term effects to the hemiparkinsonian rats. A modified technique was used to encapsulate PC12 cells into gelled microspheres. We found that the PC12 cells can survive in the modified microcapsules in vitro. Most of the PC12 cells formed cluster 3 weeks after incubation. The PC12 cell-loaded microcapsules were also examined in vitro. Adult Sprague-Dawley rats, anesthetized with chloral hydrate, were injected unilaterally with 6-hydroxydopamine into the medial forebrain bundle. The effectiveness of this lesion was tested by measuring apomorphine or methamphetamine-induced rotation one month after lesioning. The unilaterally lesioned rats were transplanted with microencapsulated PC12 cells. Results showed that apomorphine and methamphetamine-induced rotations were greatly suppressed after transplantation. One year after the grafting, the animals were anesthetized with urethane for the voltammetric study. Low dose of KCl was directly injected into the grafted striatum through pressure microejection. We found that KCl-induced DA release, as measured by voltammetric techniques, was regenerated in the striatum. The animals were later sacrificed for histological examination. We found that capsules were present in the lesioned striatum one year after grafting. Most of the capsules contained no PC12 cell. However, some capsules were filled entirely with PC12 cells. Taken together, our data suggested that PC12 cells can survive in the capsule in vitro and may provide long-term dopaminergic effects to the hemiparkinsonian rats.     

Ontogeny of meal patterns of the rat.

Two experiments, one with 6 18-60 day old female Wistar rats and the other with 12 Wistar and Long-Evans rats, examined the development of adult rat meal patterns. Although diurnal differences in food intake were present as early as 18 days after birth, with significantly more food being consumed in the dark, these differences were much smaller than those seen in adults. Nocturnal feeding gradually increased, reaching adult levels at around 6 wks of age. In general, little or no direct correlation was observed between meal size and postmeal interval length until rats were 4-5 wks old. Thereafter, correlations of +.40 or higher were usually observed. On the other hand, correlations averaging around zero were observed between meal size and premeal interval at all ages studied. Results are discussed in terms of their implications for proposed physiological mechanisms determining meal onset. (25 ref) (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Ontogeny of cholinergic mediation of behaviors in the rat.

In 6 experiments with 159 immature Sprague-Dawley rats it was found that although scopolamine disrupted discriminative choice behavior in both 15- and 23-day-old pups, it increased latency to choice in 15-day-olds and decreased it in 23-day-olds. This disruption of discriminated choice behavior was not due to differential shock thresholds or differences in locomotor activity between drug-treated and control Ss, nor was it specific to a T-maze shock-escape discrimination task. Results suggest that central cholinergic mediation of different behaviors may mature at different rates. (26 ref) (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Ontogeny of thermal and olfactory determinants of huddling in the rat.

In Exp I, in standardized tests of huddling behavior, 64 5-, 10-, 15-, and 20-day-old Sprague-Dawley rat pups spent substantial and equivalent amounts of time with an immobile rat or a heated, fur-covered tube, which suggests that the conspecific and inanimate stimuli were equally attractive to the pups. In Exp II, with 32 Ss, 2-choice preference tests showed developmental differences in attraction. Younger Ss preferred to huddle with the warmer, inanimate target, whereas older Ss preferred the conspecific. The emergent conspecific preference appears mediated by attraction to species odors. In Exp III (64 Ss), the 5- and 10-day-olds huddled equally with an immobile rat and an immobile gerbil (stimuli with similar thermal and tactile properties), but older Ss preferred the conspecific. In Exp IV, with 16 15-day-old and 16 20-day-old Ss, intranasal zinc sulfate treatment eliminated preference for the conspecific but did not disrupt huddling per se. Results from all experiments show that thermal cues were sufficient to elicit huddling at all ages, but olfactory cues became a more salient influence before weaning. An ontogenetic transition from "physiological" to "filial" huddling is discussed in terms of changes in sensory control of early behavior. (22 ref) (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Ontogeny of estrogen receptor-beta expression in rat testis

The recently discovered estrogen receptor-beta (ERbeta) is expressed in rodent and human testes. To obtain insight in the physiological role of ERbeta we have investigated the cell type-specific expression pattern of ERbeta messenger RNA (mRNA) and protein in the testis of rats of various ages by in situ hybridization and immunohistochemistry. In fetal testes of rats 16 days postcoitum and testes of 4-day-old animals, fetal germ cells (gonocytes) reveal the ERbeta mRNA in their cytoplasm and the ERbeta protein in their nucleus. In testes of 11- and 15-day-old rats, ERbeta mRNA and protein were detected in Sertoli cells and type A spermatogonia. No signal was found in other types of germ cells. In the adult testes, expression of ERbeta mRNA as well as ERbeta protein was found in pachytene spermatocytes from epithelial stages VII-XIV and in round spermatids from stages I-VIII. Low ERbeta expression was observed in all type A spermatogonia, including undifferentiated A spermatogonia, whereas no expression was found in In and type B spermatogonia and early spermatocytes. At all ages, Sertoli cells showed a weak hybridization signal as well as weak immunoreactivity for ERbeta. In adult testes, no ERbeta mRNA or protein was detected in the interstitial tissue, indicating that Leydig cells and peritubular myoid cells do not express ERbeta. The expression of ERbeta in fetal and late male germ cells as well as in Sertoli cells suggests that estrogens directly affect germ cells during testicular development and spermatogenesis.     

Ontogeny of iodothyronine deiodinases in human liver

The role of the deiodinases D1, D2, and D3 in the tissue-specific and time-dependent regulation of thyroid hormone bioactivity during fetal development has been investigated in animals but little is known about the ontogeny of these enzymes in humans. We analyzed D1, D2, and D3 activities in liver microsomes from 10 fetuses of 15-20 weeks gestation and from 8 apparently healthy adult tissue transplant donors, and in liver homogenates from 2 fetuses (20 weeks gestation), 5 preterm infants (27-32 weeks gestation), and 13 term infants who survived up to 39 weeks postnatally. D1 activity was determined using 1 microM [3',5'-125I]rT3 as substrate and 10 mM dithiothreitol (DTT) as cofactor, D2 activity using 1 nM [3',5'-125I]T4 and 25 mM DTT in the presence of 1 mM 6-propyl-2-thiouracil (to block D1 activity) and 1 microM T3 (to block D3 activity), and D3 activity using 10 nM [3,5-125I]T3 and 50 mM DTT, by quantitation of the release of 125I. The assays were validated by high performance liquid chromatography of the products, and kinetic analysis [Michaelis-Menten constant (Km) of rT3 for D1: 0.5 microM; Km of T3 for D3: 2 nM]. In liver homogenates, D1 activity was not correlated with age, whereas D3 activity showed a strong negative correlation with age (r -0.84), with high D3 activities in preterm infants and (except in 1 infant of 35 weeks) absent D3 activity in full-term infants. In microsomes, D1 activities amounted to 4.3-60 pmol/min/mg protein in fetal livers and to 170-313 pmol/min/mg protein in adult livers, whereas microsomal D3 activities were 0.15-1.45 pmol/min/mg protein in fetuses and <0.1 pmol/min/mg protein in all but one adult. In the latter sample, D3 activity amounted to 0.36 pmol/min/mg protein. D2 activity was negligible in both fetal and adult livers. These findings indicate high D1 and D3 activities in fetal human liver, and high D1 and mostly absent D3 activities in adult human liver. Therefore, the low serum T3 levels in the human fetus appear to be caused by high hepatic (and placental) D3 activity rather than caused by low hepatic D1 activity. The occasional expression of D3 in adult human liver is intriguing and deserves further investigation.     

Ontogeny of serotonergic inhibition of behavioral arousal in the rat.

Tested the effects of para-chlorophenylalanine (P-CPA), which depletes brain serotonin, in 3 experiments with 10-25 day old albino Sprague-Dawley rats. Experimental Ss were tested for locomotor effects 0, 24, 48, and 72 hrs after P-CPA injection. Depletion of serotonin increased behavior arousal in 15-, 20-, and 25-day-old Ss, but not in those 10 days old. P-CPA potentiated the locomotor activity induced by amphetamine, but again 10-day-old Ss did not show the effect. These results are interpreted as evidence for the delayed maturation of a serotonergic inhibitory system that modulates behavioral arousal. (53 ref) (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Ontogeny of the ionic specificity of sodium appetite in the rat pup

Sodium-deficient adult rats prefer NaCl to other monochloride salts (e.g., Denton, 1991; Schulkin, 1991). However, it is not known when or how this specificity develops. Our experiments charted the development of the ionic specificity of sodium appetite aroused by sodium depletion or intracerebroventricular injection of renin. We compared intake of 3% NaCl to three other monochlorides, potassium (K), ammonium (NH4), and lithium (Li), and calcium chloride (CaCl2) at various ages between 72 hr postnatal and weaning. This revealed a biphasic developmental scheme: The adult pattern of discrimination between the salts emerges between 3 and 18 days of age. Subsequently, the preference for Na over the other salts increases into adulthood.     

pH-dependent secondary conformation of bovine lens alpha-crystallin: ATR infrared spectroscopic study with second-derivative analysis

Of 37 Yersinia isolates from various aquatic environments, seven were Y. enterocolitica and 30 Y. intermedia. These isolates were biotyped, serotyped and tested for their susceptibility to 20 antibiotics. All Y. enterocolitica isolates were of biovar 1; those of Y. intermedia were distributed amongst four biovars (1, 2, 4 and 6). On the basis of combined biotyping and serotyping results, Y. enterocolitica isolates were distributed in five and Y. intermedia in 17 groups. With the exception of one Y. enterocolitica isolate which was resistant to tetracycline and streptomycin, the isolates were sensitive to the non-beta-lactam antibiotics. In contrast, various patterns of beta-lactam insensitivity were detected, including ampicillin and ticarcillin (35 isolates), cephalothin (33 isolates), carbenicillin (32 isolates), amoxycillin/clavulanate (23 isolates) and cefoxitin (22 isolates). No correlation between biotype or serotype and the susceptibility pattern of the isolates was apparent. Both inducible cephalosporinase activity against third-generation cephalosporins and inhibition of resistance to penicillins were detected in all Y. enterocolitica and Y. intermedia isolates by double-disk tests.     

Ontogeny of innervation of rat and ovine fetal adrenals

The formation of adrenocortical zonation occurs in rats during late gestation. Since adult cortical function is modulated by neural mediators, it is possible that the development of differentiated function is dependent on cortical innervation. The goal of this study was to compare the pattern and timing of rodent and ovine adrenal innervation during late organogenesis by staining with antibodies directed against the neuropeptides vasoactive intestinal peptide (VIP), calcitonin gene-related peptide (CGRP) and neuropeptide tyrosine (NPY) and the catecholamine biosynthetic enzyme, tyrosine hydroxylase (TOH). Rat adrenals were collected from fetal days 17-21 (term=21 days) and ovine adrenals from fetal days 101-136 (term=145 days). Adrenals were fixed, cryosectioned at 100 microns and immunostained using Cy3-conjugated secondary antibodies. In both species, staining of VIP, CGRP, NPY and TOH fibers was observed in the capsule and subcapsular layers of the cortex during gestation. In late gestation, VIP- and NPY-positive ganglions cells were observed near the medulla extending processes toward the outer cortex; in ovine adrenals, fibers from ganglion cells appeared to surround nests of outer cortical (presumably, zona glomerulosa) cells. These data show that phenotypically distinct neural elements appear at different stages of adrenocortical development. The presence of neural elements in contact with adrenal cortical cells supports the possibility for neural control of adrenocortical development.     

Ontogeny of FAD-linked glycerophosphate dehydrogenase in rat pancreatic islets

The activities of the mitochondrial FAD-linked glycerophosphate dehydrogenase (m-GDH), glutamate dehydrogenase, alpha-ketoglutarate dehydrogenase, glutamate-pyruvate transaminase (GPT) and glutamate-oxaloacetate transaminase were measured in islet and liver homogenates from fetal, neonatal, adult male, adult female, pregnant and lactating rats. Either parallel or dissociated ontogenic changes were observed in islet and liver homogenates. The activity of islet m-GDH was slightly, albeit not significantly, lower in neonates than in adult rats, comparable in male and female adult animals, unaffected by pregnancy, and increased during lactation. It was much higher in fetal or adult islets cultured for 7 days than in freshly isolated islets from adult rats. In cultured islets from adult rats, the increase in m-GDH activity coincided with a dramatic decrease of GPT activity, a situation the mirror image of that found in several animal models of non-insulin-dependent diabetes mellitus. The intrinsic properties of m-GDH, as judged by comparison of measurements made by either a radioisotopic or a colorimetric procedure, were not identical in islet and liver homogenates and differed between fetal and adult islets, suggesting the existence of distinct iso-enzymes. These findings illustrate adaptive changes of islet enzymes, with exclusive or partial mitochondrial location, in ontogenic situations characterized by a remodelling of fuel homeostasis.     

Ontogeny of calretinin expression in rat dorsal root ganglia

The relationship between the expression of calretinin and the maturation level of peripheral sensory neurons was investigated by means of immunohistochemistry and immunoblot analysis. Our immunohistochemical results show that calretinin is expressed during two different developmental phases in rat dorsal root ganglia. The early phase lasts between embryonic days 11 and 14, when calretinin is detectable in the majority (75%) of the cells. A second phase starts at embryonic day 17 and lasts throughout the whole postnatal life, when calretinin is expressed only in a small proportion of the neurons (less than 8%). Between these two periods no calretinin is found in the ganglia. These changes in calretinin expression during embryonic development were confirmed by Western blot analysis. The early expression of calretinin in dorsal root ganglion cells suggests that calretinin may act as a calcium regulator until neurotrophins take over the precise tuning of intracellular calcium concentration.     

Epithelial cell migration in the normal rat lens

The purpose of the present study was to estimate the rate of the rat lens epithelium displacement. Twenty-four adult male rats were injected intraperitoneally with 18.5 kBq/g [3H] thymidine. Rats in groups of three were killed at the following times: after one hour, 3, 6, 9, 12, 15, 18 and 21 days. The enucleated eyes were fixed in formalin, embedded in paraffin, cut into 5 microns-thick sections, and subjected to autoradiography. The number of cells was counted, and the x-coordinate of each nucleus with three grains or more, and its grain content were recorded. Statistical analysis included analysis of variance and linear regression. The average lens diameter was 68 fields of 75 microns each (5.1 mm). Cell displacement was measured in two regions: Region A, the equatorial region, extending from field 0-20, and region B extending from field 20 to the lens center (field 34). Cells in region A are divided into two kinetic compartments, progenitor (P-cells) occupying fields 9-20, and non-proliferating (Q) cells occupying fields 0-8. Cells advanced from the P into the Q-compartment at a velocity of 21.8 microns/day. Cells did not move between field 20 and the lens center. These findings resemble kinetically renewing tissues, e.g. epidermis and crypt-villus in the gastrointestinal mucosa.     

Co-culture with assisted hatching of human embryos using Buffalo rat liver cells

Two-dimensional nutation pure NQR experiments on 35Cl have been carried out on a single crystal of NaClO3. The 2D nutation experiment separates out different orientations of each chemically equivalent site in a unit cell as a separate frequency in the omega1 domain. The squares of the observed frequencies lie on a straight line with respect to the squares of the offsets, confirming the expected offset dependence quantitatively. The intercepts at zero offset yield the relative orientations of the efg tensors with respect to the axis of the radiofrequency coil.