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Biomedical subjects

J H Clark

Publications and source records attributed to J H Clark.

At least 217 records · Page 12Linked to original sources
PubMed1980

Effects of salt extraction on the quantitation of nuclear estrogen receptors: interference by secondary estrogen binding sites.

The effects of salt-extraction on type I and type II estrogen binding sites were examined in uterine nuclei. Injection (10 ug) of estradiol or estriol in adult ovariectomized rate induced maximum numbers (80-100%, integral of 1 pmole/uterus) of 0.4 M KCL resistant type I estrogen complexes at 1 hour. Only estradiol, which sustained these levels for long periods of time (4-24 hours) stimulated true uterine growth. Likewise, a single injection of estradiol, but not estriol, also elevated nuclear type II sites throughout the entire uterine growth period (1 - 48 hours). However extraction of these nuclei from estradiol injected rats with 0.4 M KCL increased the numbers of type II sites from integral of 1 pmole/uterus (non-extracted nuclei) to integral of 8 pmoles/uterus (salt resistant plus salt-extractable fractions). Sixty percent of these sites were resistant to salt-extraction. Continuous exposure to either estradiol or estriol by beeswax implants stimulated nuclear type II sites which were highly resistant (80%) to KCL-extraction, and additional sites were not exposed by high salt. Thus chronic treatment with both estrogens "locked in" nuclear type II sites such that they were resistant to KCL-extraction. This resistance of type II sites to salt-extraction correlated with the ability of estradiol and estriol implants to stimulate true uterine growth. The procedures presented here for nuclear preparation and assay have reduced non-specific binding considerably in the uterine system, and may eliminate the need to perform exchange assays on salt-extracted nuclei in other systems.

Animals↗
PubMed1979

Nuclear binding of the estrogen receptor: heterogeneity of sites and uterotropic response.

Two kinds of estradiol binding sites are present in purified nuclei from the rat uterus following estradiol injection. One of these sites (type I) corresponds to the well-known estrogen receptor which undergoes translocation from the cytoplasm to the nucleus. The second site (type II) is not translocated from the cytoplasm to the nucleus, however, estradiol treatment does stimulate an increased number of these sites. Type II sites are observed in purified nuclei and chromatin isolated from the uterus but not from non-target tissues such as the spleen and diaphragm. Thus an elevation in the levels of type II sites appear to be a specific nuclear response of the rat uterus to estradiol. Saturation analysis over a wide range of [3H]-estradiol concentrations produces a binding curve for type II sites which is sigmoidal and hence no accurate estimation of the dissociation constant is possible. The binding of [3H]-estradiol to nuclear type II sites is inhibited by estradiol and diethylstilbestrol but not by progesterone, testosterone, or corticosterone. Extraction of nuclei isolated from estrogen treated rat uteri with KCl provides a complex picture. Direct labeling of nuclear estrogen receptors either by in vivo injection or in vitro incubation of intact uteri with [3H]-estradiol measures only a fraction of the specific estrogen binding sites associated with the nuclear pellet following 0.4 M KCl extraction. These sites are more accurately determined by performing saturation analysis over a wide range of [3H]-estradiol concentrations by exchange which measures specific estrogen binding sites, not [3H]-steroid. Saturation analysis of estradiol binding to KCl extracted nuclei when performed by exchange, with appropriate corrections for type II binding, reveals that approximately 1000--2000 receptors per nucleus are resistant to KCl extraction 1 hr after administration. The same numbers of type I sites display long-term nuclear retention. A single injection of estradiol results in long term (greater than 6 h) retention of type I sites, rapid and sustained elevations (1--72h) in type II sites and true uterine growth (uterine wet weight at 24--43 h). Estriol injections caused a rapid increase in nuclear type I sites which was not accompanied by an increase in type II sites and no true uterine growth occurred. Administration of estriol or estradiol as a pellet implant, which causes continuous occupancy of type I sites, increases the quantity of nuclear type II sites and stimulates true uterine growth. Therefore, we conclude that elevated levels of nuclear type II sites correlate with the long term uterotropic response to estrogenic hormones. Although we do not understand the function of this second class of binding sites it is possible that the type II sites represent a major component in the mechanism by which estrogens stimulate growth of the uterus.

Animals↗
PubMed1979

Mechanism of action of estrogen agonists and antagonists.

The relationship between estrogen agonists and antagonists and the nuclear binding of the estrogen receptor was investigated in the immature rat uterus. Estriol (and other short acting estrogens) are antagonistic when administered as a single injection due to the inability of the receptor-estriol complex to be retained by uterine nuclei for a critical period necessary to stimulate true uterine growth. This failure of the receptor-estriol complex to be retained appears to be due to rapid clearance of estriol from uterine tissue following a single injection of the hormone. When estriol is present in a chronic fashion, it acts as an agonist because receptor estriol complex is retained at nuclear sites for long periods of time. Nonsteroid estrogen antagonists, such as Nafoxidine, also cause long term nuclear retention of the estrogen receptor and act as estrogen agonists after a single injection. However, after multiple injections these drugs are antagonistic and this effect is correlated with decrease availability of cytoplasmic estrogen receptors. In addition, these compounds appear to specifically stimulate the growth of uterine epithelial cells while minimally affecting other cell types which results in hyperestrogenization of the epithelium.

Animals↗
PubMed1979

Effect of intraruminal urea administration on glucose metabolism in dairy steers.

Isotope dilution techniques were used to study steady-state glucose kinetics in four rumen-fistulated Holstein steers and to study the effect of rapid absorption of ammonia from the rumen on glucose metabolism. Steers were fed a high-concentrate diet at hourly intervals from automatic feeders. Plasma glucose specific activity curves following single intravenous injection of [2-3H]glucose were used to construct a two-compartment model of the glucose pool with inflow and outflow from compartment one. Primed continuous infusion of [2-3H]glucose was used to determine the steady-state turnover rate of glucose and to monitor changes in the rates of inflow and outflow of glucose from the glucose pool following a single dosage of urea (0.4 g/kg body weight) into the rumen. Compartment sizes of the glucose pool were 65.6 and 33.5 g for compartments 1 and 2, respectively. Glucose turnover rate during steady-state was 15.4 mg/minute/kg body weight 0.75 and transfer rate of glucose between compartments was 17.9 mg/minute/kg body weight 0.75. Concentrations of rumen ammonia-nitrogen, plasma ammonia-nitrogen and plasma urea-nitrogen were 6.1, 0.5 and 4.0 mg/100 ml, respectively, before urea dosage. Rumen ammonia-nitrogen, plasma ammonia-nitrogen increased after urea dosage and reached peak concentrations, 170.0 and 1.2 mg/100 ml, respectively, approximately 120 minutes after urea dosage. Plasma urea-nitrogen increased linearly throughout the 4-hour sampling period and reached 12.0 mg/100 ml at end of the experiment. Concentration of glucose in plasma increased from 98.2 mg/100 ml before urea dosage to 114.6 mg/100 ml at 100 minutes after urea dosage. Estimates of glucose production and utilization indicated that the increased concentration of glucose in plasma in all steers was due, at least partially, to a decrease in the rate of glucose utilization. A rapid rate of glycogenolysis which resulted in a marked increase in the plasma glucose concentration also was evident in one steer.

Ammonia↗
PubMed1979

Muscle and liver glycogen content: diurnal variation and endurance.

The purpose of this study was to determine the influence of diurnal variations of muscle and liver glycogen stores on exercise endurance in male albino rats. Animals were swum to exhaustion at either 0700 or 1900 h, after which samples of soleus, white vastus lateralis, and red vastus lateralis muscles as well as liver were excised and subsequently analyzed for glycogen content. Glycogen content of all tissues from nonexercising control animals was higher in the morning than in the evening. Consequently, animals at 0700 h swam 60% longer than those at 1900 h (209 +/- 20 min vs. 130 +/- 23 min, respectively). However, because the skeletal tissues of the exhausted animals were not totally depleted of glycogen, it was concluded that fatigue under the swim protocol was the result of hypoglycemia secondary to hepatic glycogen depletion. The results of this study demonstrate the physiological consequences of diurnal glycogen fluctuation and establish experimental support for the importance of controlling this variable in rodent exercise investigations.

Animals↗