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Biomedical subjects

B I Posner

Publications and source records attributed to B I Posner.

At least 109 records · Page 6Linked to original sources
PubMed1986

Increase in the number of type II insulin-like growth factor receptors during propylthiouracil-induced hyperplasia in the rat thyroid.

We have observed that membranes isolated from rat thyroids contain receptors for the insulin-like growth factors (IGF). As IGFs are known to be important mediators of tissue growth, we conducted this study to determine whether modulation of thyroid IGF receptors might be involved in TSH-stimulated hyperplasia. A substantial increase in both the weight of the thyroid and its DNA content was observed within 2 days of exposing adult male rats to 0.1% propylthiouracil (PTU) in their drinking water. Serum T4 reached unmeasurable levels and serum TSH rose 3-fold over control by the tenth day of treatment. [125I]Iodo-human(h)IGF-II binding to membranes isolated from hyperplastic glands was significantly higher than control beginning at 2 days. A maximum was reached after 5 days (13.3 +/- 0.8%/25 micrograms protein vs. a control level of 6.7 +/- 0.7%, mean +/- SEM). The increase had disappeared by 15 days of PTU exposure, paralleling the drastic fall in the growth rate of the glands. This increase in binding was specific for the thyroid, as it was not seen in other organs. In both treated and control animals, the receptor involved was shown to be type II by preferential binding to IGF-II, lack of interaction with insulin, and molecular sizing. The observed increase in binding could be accounted for by an increase in receptor site number, the affinity remaining essentially the same. We conclude that the TSH-stimulated hyperplasia of the rat thyroid, induced by PTU, is associated with an increase in the binding sites of the type II IGF receptor. This observation raises the possibility that modulation of this receptor may play a role in the mediation of the mitogenic effect of TSH on the thyroid gland.

Animals↗
PubMed1986

Quantitative in vivo autoradiographic localization of [125I-Tyr11]somatostatin-14- and [Leu8,D-Trp22-125I-Tyr25]somatostatin-28-binding sites in rat brain.

Quantitative in vivo autoradiography was used to identify and compare the regional distribution of specific binding sites for blood-borne [125I-Tyr11]somatostatin-14 (S-14 section) and [Leu8,D-Trp22-125I-Tyr25]somatostatin-28 (S-28 section) in the rat brain. Rats were given intracardiac injections of 17 pmol S-14 section (with or without unlabeled S-14) or 17 pmol S-28 section (with or without unlabeled S-28). After whole body perfusion and fixation, brains were processed for light microscopic autoradiography of S-14 section- and S-28 section-binding sites. Of the peripheral tissues, the adrenal glands showed the highest uptake of S-14 section and S-28 section (determined by counting) and were subsequently processed for autoradiography for comparison with brain. Specific autoradiographic grains (ARG) associated with both radioligands were identified only in the circumventricular organs (CVOs). The highest ARG density associated with S-14 section was found in the area postrema, followed in decreasing order by the subfornical organ and the organum vasculosum lamina terminalis region. Median eminence (ME) contained virtually no specific S-14 section ARG. As with S-14 section, the highest ARG density of S-28 section-binding sites was also found in the area postrema, which labeled approximately equally with the two radioligands. This was followed by the ME, subfornical organ, and organum vasculosum lamina terminalis. The overall patterns of labeling of the CVOs with S-14 section and S-28 section showed significant differences, especially in the ME. Within the ME, labeled S-28 section was concentrated in a broad band throughout the external zone in a location identical to that of immunoreactive S-14. Analysis of dose-response curves obtained with 0.3-30 nmol unlabeled S-14 or S-28 revealed IC50 values for S-14 3- to 6-fold lower than those for S-28 for all labeled CVOs. With both S-14 section and S-28 section, the labelling density of the adrenal glands was double that of the area postrema. Adrenal binding of the radioligands was confined to the cells of the zona glomerulosa. We conclude: specific high affinity binding sites for S-14 section and S-28 section exist in the CVOs and the adrenal glomerulosa; and the 3- to 6-fold higher affinity of binding of S-14 to CVOs compared to S-28 together with the dissimilar patterns of labelling of the different CVOs by the two radioligands suggest the existence of separate populations of S-14 and S-28 receptors.

Animals↗
PubMed1986

Effect of insulin-like growth factors on human foetal, adult normal and tumour pituitary function in tissue culture.

To determine the direct effects of insulin-like growth factors (IGFs) on hormone release by the human pituitary gland, human foetal, adult normal and tumour pituitary tissues were maintained in culture for 2 to 4 weeks and tested with acute (3 h) exposures to different preparations of IGF peptides. Adult normal pituitaries and adenomas were tested with a semipurified preparation of IGFs, free of immunoreactive insulin, containing IGF-I and IGF-II in a ratio of approximately 1:4. Human foetal pituitaries were tested with the semipurified IGFs as well as more purified preparations of IGF-I and IGF-II. Culture media were assayed for hGH, hPrl, hACTH and hLH using specific radioimmunoassays. Both foetal (n = 16 (No. of pituitaries), 33 (No. of observations] and normal adult (n = 3, 16) human pituitaries cultures responded to the semipurified IGFs (2-25 ngEq/ml for foetal and 2-4 ngEq/ml for adult pituitaries) with a significant decrease in hGH release compared to basal (P less than 0.01) whereas the GH-secreting pituitary tumours showed no effect when tested with from 2 to 25 ngEq/ml (n = 8, 129, NS). The effect of IGFs on human foetal somatotrope activity was dose-related for both the semipurified IGFs (2-25 ngEq/ml, n = 16, 33) and IGF-I or IGF-II (10-100 ng/ml; n = 3, 18).(ABSTRACT TRUNCATED AT 250 WORDS)

Adenoma↗
PubMed1985

Increase in the type 2 insulin-like growth factor receptors in the rat kidney during compensatory growth.

We have observed an increase in binding of IGF-I and IGF-II to microsomes obtained from rat kidneys undergoing compensatory growth following contralateral nephrectomy. This increase was evident by the 4th day and it preceded observable growth of the kidney. Binding returned to control levels just prior to the flattening of the growth curve of the kidney. The increase was due to an increase in the type 2 binding sites, the only type unequivocally present, from 95.3 +/- 2.7 to 117.2 +/- 4.1 pM/150 micrograms of microsome protein at its maximum at 4 days.

Animals↗
PubMed1985

Prolactin uptake into liver endocytic components. Reduced sensitivity to chloroquine.

Subfractionation of hepatic Golgi fractions on Percoll gradients revealed two populations of lactogen-receptor enriched components with one pool of rho = 1.040-1.050 and a high-density component of rho = 1.053-1.064. 125I-labelled prolactin uptake into Percoll gradient subfractions demonstrated rapid accumulation into low-density elements and slower accumulation in high-density structures. Electron microscope radioautography demonstrated that silver grains were largely associated with lipoprotein-filled structures. Radiolabel was highly concentrated in these components especially the very pure higher-density component where enrichment over the homogenate was 184-fold at the peak time after injection (10 min). Administered chloroquine accumulated in the higher-density component but not in low-density elements, suggesting that the former are at an acid intraluminal pH. In contrast to the marked effect of chloroquine on insulin uptake into endocytic structures found in Golgi fractions (Posner, B.I., Patel, B.A., Khan, M.N. and Bergeron, J.J.M. (1982) J. Biol. Chem. 257, 5789-5799), little effect of this pH disrupting agent was observed on prolactin uptake. The difference between the effect of chloroquine on 125I-labelled prolactin and 125I-labelled insulin uptake may reflect the greater stability of prolactin-receptor complexes in a low-pH environment.

Animals↗
PubMed1985

Internalization of insulin: structures involved and significance.

The binding of insulin to its receptor is followed by aggregation of hormone-receptor complexes and their internalization into the cell. Internalized hormone is concentrated in Golgi-enriched not lysosomal endocytotic structures which, in rat liver, contain lipoprotein particles and can be resolved by centrifugation techniques into three different entities. Recent work has shown that the bulk of endocytotic structures can be resolved from biochemically defined (i.e., galactosyltransferase-containing) Golgi elements. The endosomal apparatus or endosomes appear to function as a sorting center wherein internalized hormone-receptor complexes are concentrated and dissociated prior to directing hormone to lysosomes and receptor back to the cell surface for reutilization. Endosomes are heterogeneous and different functions might be subserved by different endosomal structures. Since an insulin stimulable receptor kinase activity can be identified in endosomes certain aspects of insulin action might be initiated herein.

Animals↗
PubMed1985

Separation of biological variant insulin molecules from different species by reversed-phase high-performance liquid chromatography (HPLC).

Three different isocratic systems for the separation by reversed-phase high-performance liquid chromatography (HPLC) of different species of insulin have been investigated. The effect of different solvent compositions and temperatures on elution time and resolution have been studied. These studies have been used to devise a method for reversed-phase liquid chromatographic separation of bovine, porcine, and human insulin, as well as the A and B chains of bovine insulin. The method can also be used for the separation of the various products of the iodination of porcine insulin. 125I-A14 tyrosine-labeled porcine insulin can be readily separated from nonlabeled porcine insulin and from other iodinated constituents of the mixture. A flow-though gamma-counting system that was designed for this work is described.

Animals↗
PubMed1985

Characterization of the prolactin receptor in cell fractions from rat liver.

[125I]Prolactin (PRL) was covalently cross-linked to its binding sites in subcellular fractions of female rat livers using NHSAB and UV irradiation. Analysis by non-reducing SDS-PAGE showed that all fractions with specifically bound radioactive hormone contained a major autoradiographic band (eliminated with unlabeled PRL) of similar electrophoretic mobility consistent with a MW of 36K for the receptor. In addition, microsomal membranes were treated with a zwitterionic detergent (CHAPS), solubilizing 30-60% of the specifically bound radioactivity. SDS-PAGE analysis of [125I]PRL cross-linked to CHAPS-soluble and CHAPS-insoluble material showed an autoradiographic band with a similar MW. These results suggest that prolactin receptors in different hepatocyte membranes are similar in MW and do not appear to be linked by disulfide bonds to other membrane proteins. Some of the binding sites appear to interact with membrane constituents in ways that affect their solubility by CHAPS.

Animals↗
PubMed1985

Concentration of native prolactin and prolactin binding sites in hepatic subcellular fractions from hyperprolactinemic rats.

Lactogen binding and prolactin content were measured in hepatic subcellular fractions from tumor-bearing rats (TBR; MtT/F4, MtT/W5, MtT/W10) with elevated prolactin and growth hormone levels and from control animals. Specific binding of 125I-oPRL to Golgi fractions from tumor-bearing animals was 2.5 to 7 fold greater than that from controls. Binding to plasmalemma was 6-fold greater in tumor-bearing rats. The specific binding of 125I-labelled bGH and insulin showed less marked differences between TBR and controls. Subcellular fractions were extracted with HCl to determine hormonal content. The content of prolactin and growth hormone in Golgi fractions from TBR was at least 20-fold that in fractions from controls. Rat prolactin extracted from Golgi heavy elements was 50% as effective as native material in binding to lactogen receptors as judged by radioreceptor assay. These studies demonstrate that the chronic elevation of prolactin was associated with an increase of receptors not only in the intracellular compartment but on the cell surface as well. Furthermore, they demonstrate that native prolactin is internalized and accumulated in rat liver Golgi fractions.

Animals↗
PubMed1985

NH2-terminal specificity and axonal localization of adrenocorticotropin binding sites in rat median eminence.

Adrenocorticotropin binding sites in the rat median eminence have been localized in vivo. These binding sites occur in the basalar zone, which is rich in axonal endings. Using competitive binding and quantitative light-microscope radioautography, we found that the median-eminence binding site, in contradistinction to the adrenal receptor, binds specifically the residue 4-10 region of the adrenocorticotropin molecule. Using quantitative electron-microscope radioautography and median-eminence deafferentation, we localized the binding sites to axon terminals in this region. In time-delayed uptake studies using light-microscope radioautography, we failed to observe concentration of radiolabel in neurons of the medial basal hypothalamus after the direct injection of radioiodinated adrenocorticotropin(1-24) into the median eminence.

Adrenal Glands↗
PubMed1985

Insulin and insulin receptor uptake into rat liver. Chloroquine action on receptor recycling.

In the present study, the effect of chloroquine on both insulin and receptor distribution was examined in vivo. Insulin injection (25 nmol/100 g body wt) caused a marked accumulation of both insulin and its receptor in purified hepatic Golgi fractions by 15 min postinjection. Percoll fractionation of parent Golgi fractions resolved two endocytic components of low (rho = 1.040-1.050) and high (rho = 1.053-1.064) density in which the relative distribution of insulin binding sites was unaltered by chloroquine. Chloroquine significantly accumulated in the high-density region of the Percoll gradient consistent with this being a low pH compartment. 125I-insulin accumulated first in the low-density (1 min) and subsequently in the high-density region (5-10 min) of Percoll-subfractionated Golgi fractions. Chloroquine treatment caused marked accumulation of 125I-insulin in the high-density compartment with substantial retention of radiolabel therein at 20 min postinjection. 125I-insulin extracted from the Percoll fractions was comparably intact in control and chloroquine-treated rats. These data suggest that the chloroquine-accumulating, high-density compartment of hepatic Golgi fractions is the site of dissociation of internalized insulin-receptor complexes before degradation of the ligand and receptor recycling.

Animals↗
PubMed1985

Acute reversal of the enhanced insulin action in trained athletes. Association with insulin receptor changes.

We studied the effect of aerobic training and detraining on insulin-stimulated glucose disposal and on erythrocyte insulin receptor binding. Seven endurance-trained athletes were studied at 12 h, 60 h, and 7 days after cessation of training and compared with three untrained, age- and weight-matched controls. The metabolic clearance rate of glucose as measured by the euglycemic clamp technique was 15.6 +/- 1.8 ml/kg/min (mean +/- SEM) in the trained subjects 12 h after the last bout of exercise compared with 7.8 +/- 1.2 ml/kg/min in the untrained control group. When the trained subjects refrained from physical training, the metabolic clearance rate decreased to 10.1 +/- 1.0 ml/kg/min at 60 h and further to 8.5 +/- 0.5 ml/kg/min after 7 days of detraining. The percentage of specific insulin binding to young erythrocytes (density 1.089-1.092), isolated by density gradient centrifugation, decreased from 10.4 +/- 0.9 at 12 h after the last exercise to 8.1 +/- 0.7%/3 X 10(9) cells after 60 h of detraining (P less than 0.001). The decrease in insulin binding to erythrocytes was almost entirely accounted for by a decrease in the number of insulin receptors. We conclude that the increase in peripheral insulin action seen in trained athletes is rapidly reversed, possibly by a mechanism separate from other phenomena associated with chronic training. The parallel findings of decreased in vivo insulin action and decreased insulin binding in young erythrocytes suggest that modulation of in vivo insulin response by detraining may be at least partially mediated by changes in insulin receptor number.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1984

In vivo uptake of insulin into hepatic Golgi fractions: application of the diaminobenzidine-shift protocol.

The hypothesis that insulin is internalized into the hepatic Golgi apparatus was tested by the diaminobenzidine-shift protocol of Courtoy et al. (1984, J. Cell Biol. 98, 870). Highly purified Golgi fractions were isolated after the coinjection of [125I]insulin and the synthetic ligand, galactose-bovine serum albumin-horseradish peroxidase. Golgi fractions were subsequently reacted in the presence or absence of diaminobenzidine, then subjected to Percoll gradient centrifugation. For incubations carried out in the absence of diaminobenzidine, [125I]insulin-containing components were found at a low density (peak density congruent to 1.042) identical to that of the Golgi marker enzyme galactosyltransferase. However after incubations carried out in the presence of diaminobenzidine, the majority of [125I]insulin-containing components was shifted to a higher density of greater than 1.06 while that of galactosyltransferase remained unchanged (peak congruent to 1.042). These observations indicate that the majority of internalized insulin is not located in galactosyltransferase-containing Golgi components.

3,3'-Diaminobenzidine↗
PubMed1984

Characterization of insulin-like growth factor receptors in rat anterior pituitary, hypothalamus, and brain.

Studies were undertaken to determine whether the insulin-like growth factors (IGF-I and -II), bind to specific membrane receptors in the pituitary and brain. Anterior pituitary glands, hypothalami, and brains (minus hypothalami) were obtained from adult male Sprague-Dawley rats (225-300 g) and 15,000 X g membranes prepared by differential centrifugation. Binding of 125I-IGF-I and 125I-IGF-II to all three membrane preparations was specific, time and temperature dependent, reversible, and increased in proportion to increasing concentrations of membrane protein or labeled ligand. Neither the pH of the assay buffer (6.5-8.5) nor the presence or absence of 1 mg/ml bacitracin had any significant effect on the levels of specific binding. In all three membrane preparations IGF-II specific binding was 3-5 times higher than that observed for IGF-I, and unlabeled IGF-II displaced either 125I-IGF-I or 125I-IGF-II better than comparable concentrations of IGF-I. All three membrane preparations showed similar low specific binding of 125I-insulin (1.3-2.2%) and negligible specific binding of 125I-rat GH (less than 0.5%). The presence of specific IGF and insulin receptors in rat anterior pituitary, hypothalamic, and brain tissue is additional evidence that IGFs and insulin are involved in modulating brain and pituitary function.

Animals↗
PubMed1984

Binding and uptake of [125I]iodoprolactin by epithelial cells of the rat choroid plexus: an in vivo autoradiographic analysis.

Light and electron microscopic (EM) autoradiography were used to determine the subcellular fate of [125I] iodo-PRL that specifically binds to epithelial cells of the rat choroid plexus. Experimental animals (total binding) received an external jugular vein injection of ovine [125I]iodo-PRL, whereas control animals (nonspecific binding) received an identical dose of [125I]iodo-PRL plus a 500-fold excess of unlabeled PRL. Experimental and control animals were killed by vascular perfusion 3, 20, 40, 60, and 240 min after hormone injection. Choroid plexuses were removed, processed to Epon, and prepared for light and EM autoradiography. Quantification of silver grains at the light microscopic level revealed an intense autoradiographic reaction over the epithelial cells of experimental animals 3, 20, 40, and 60 min after hormone injection. The co-injection of unlabeled PRL resulted in a statistically significant reduction in the autoradiographic reaction throughout the initial 60 min of the test period, reflecting specific binding of PRL during this interval. Analysis of EM autoradiographs revealed that the initial binding of [125I]iodo-PRL occurred primarily at the basal and lateral plasmalemma of the epithelial cells. With time there was a dramatic reduction in the amount of radiolabel localized to the cell membrane and a concomitant increase in silver grains localized over the interior of the cells. Internalized [125I]iodo-PRL exhibited a preferential localization to small cytoplasmic vesicles, short tubules, multivesicular bodies, and dense bodies. Combined acid phosphatase cytochemistry and EM autoradiography revealed that only 11.2% of the internalized silver grains were localized over acid phosphatase-positive structures 40 min after injection. The results indicate that subsequent to the initial binding of [125I]iodo-PRL to the cell membrane, internalization and translocation occurs primarily to nonlysosomal cytoplasmic organelles. The study also demonstrates at least a partial or ultimate entrance of internalized PRL to the lysosomal system.

Acid Phosphatase↗
PubMed1984

Effects of insulin-like growth factors on adult male rat pituitary function in tissue culture.

UNLABELLED: To determine the direct effects of insulin-like growth factors (IGFs) on pituitary secretion of GH, PRL, and ACTH, adult male rat pituitary explant cultures were tested with acute (3-4 h) or chronic (24 h) exposure to a semipurified preparation of IGF peptides, free of immunoreactive insulin, containing IGF-I and IGF-II in a ratio of approximately 1:4. To examine the effect of serum binding proteins on IGF bioactivity, certain experiments were run in parallel using culture medium supplemented with 10% fetal bovine serum or 1% purified BSA. To compare IGF effects with those of known regulators of pituitary function, cultures were also tested with SRIF, TRH, human pancreatic GH-releasing factor, insulin, and human GH (hGH). IGFs, at 10-100 ngeq/ml, were able to inhibit significantly both basal and (1 mM) theophylline-stimulated rat GH (rGH) and rat PRL (rPRL) release during acute (3-4 h) exposure. Only the higher concentration (100 ngeq/ml) was consistently effective in inhibiting rGH and rPRL output after 24 h in culture, due to gradual metabolism of IGF peptides by the cells. Parallel experiments carried out in medium containing 10% fetal bovine serum or 1% BSA gave similar results, demonstrating that IGF serum binding proteins did not interfere with IGF bioactivity in this test system. Chronic 5-day exposure to IGFs, at 100 ngeq/ml, resulted in a significant inhibition of rGH release for the entire 5-day period and rPRL release for the first 3 days. IGFs (10-100 ngeq/ml) had no acute or chronic effect on basal or theophylline-stimulated ACTH release. Purified IGF-I (50 ng/ml) and IGF-II (50 ng/ml) gave approximately equivalent effects on basal rGH and rPRL release during an acute (3 h) exposure suggesting that both IGFs can exert inhibitory influence on pituitary function. Ten thousand nanograms per ml insulin and 10(-9) M SRIF had acute inhibitory effects on rGH and rPRL release similar to what were observed for 100 ngeq/ml semipurified IGFs. hGH (200 and 1000 ng/ml) had no effect on rGH, rPRL, or ACTH release when administered either acutely (3-4 h) or chronically (24 h). CONCLUSIONS: These studies demonstrate that IGFs, administered acutely or chronically, directly inhibit basal as well as theophylline-stimulated rGH and rPRL output by the rat pituitary; ACTH release remains unaltered. Insulin, at high concentrations, can mimic these effects, whereas hGH has no effect either acutely or chronically.

Adrenocorticotropic Hormone↗