Cleavage of prolactin by its target organs and the possible significance of this process.

An enzymatically cleaved form of rat prolactin (rPRL) was first described in 1980. This cleavage produces a molecule that consists of two chains of amino acids linked by a disulfide bond between two Cys residues. Reduction of that bond produces two fragments of 6 and 16 Kd. A considerable amount of information has accrued in recent years about the cleaved molecule and its 16-Kd fragment. The enzyme that cleaves the molecule is present in target tissues of PRL in rodents (e.g., mammary gland and ventral prostate), and the activity of the enzyme changes with the functional state of the mammary gland. Rat mammary PRL-cleaving activity is specific for rodent PRL, and the enzyme is localized in the stroma of that gland. The enzyme that cleaves rPRL is probably cathepsin D, and the sites of cleavage on the molecule have been identified. The cleaved form of rPRL has a high degree of activity in various assays, but it has reduced activity in radioimmunoassays. The 16-Kd fragment retains a significant degree of bioactivity in in vitro mitogenic assays, and specific binding sites for the fragment have been identified. Novel bioactivities for the cleaved form of rPRL and its 16-Kd fragment have been reported, and these molecules may account for the fact that bioassay estimates of PRL in rat serum are generally higher than are RIA measurements. Although the 16-Kd fragment has significant bioactivity, it contains only six of the fourteen residues that are thought to participate in the coupling of the intact hormone to its receptor. Cleaved rPRL is present in rat serum (but not in milk), but whether the 16-Kd fragment is formed in vivo has not yet been determined.

A dissection of the chapter "Tools for Research" in Peter Singer's Animal Liberation.

The book Animal Liberation, by philosopher Peter Singer, is frequently referred to as the bible of the animal liberation/rights movement(ALARM). Thus, Singer is regarded as a major moral standard-bearer of the ALARM. Some have suggested that his book provides "intellectual rigor" to the moral arguments for animals' equality with humans, which had previously been based largely on emotionalism and sentimentality. We have analyzed the contents of the chapter "Tools for Research" which criticizes the use of animals in biomedical research as well as for drug and product-safety testing. In order to discredit these practices, Singer "documents" his arguments with 138 "notes", some of which are to the same reference and others of which contain multiple references. Of the 132 difference references, we attempted to verify the accuracy of 49 of them. Of these, 16 (33%) were inaccurate or we could not find. In addition, Singer mischaracterizes the cited studies in various ways. He quotes selectively and out of context from numerous research projects. He never mentions the objectives of these projects, except occasionally when, in our opinion, he distorts or trivializes them. Singer also cites supposedly damning "evidence" published by other antivivisectionists, even though this "evidence" has been refuted in the literature. Singer supposedly embraces utilitarianism, a philosophy which holds that the harm done by a practice should be balanced against the gain realized from it. However, he makes virtually no attempt to consider objectively the benefits that have been realized from animal-based medical research and he greatly exaggerates the costs. To him, animal research is "all pain and no gain." We believe that Singer's moral arguments for animal equality are not convincing. The lack of objectivity and the reliance upon distortion and selective quotation that characterize Singer's "scholarship" are surprising when one considers that he presents himself as an ethicist and moralist.

Regulation of insulin-like growth factor-binding proteins in rats with insulin-dependent diabetes mellitus.

As previously reported, activation of the adrenocorticotropic hormone (ACTH)-adrenal cortical axis in rats with insulin-dependent diabetes mellitus (IDDM) reduces their growth and circulating insulin-like growth factor-I (IGF-I) levels and induces a resistance to growth hormone (GH) and IGF-I. The studies reported herein were conducted to determine whether the pituitary and/or adrenal gland influence the changes in basal and GH-stimulated serum concentrations of IGF-binding proteins (IGFBPs) in rats with IDDM. Male rats were made diabetic by injections of streptozotocin. Intact nondiabetic (NonDb), diabetic (Db), hypophysectomized diabetic (HxDb), and adrenalectomized diabetic (AxDb) rats were injected twice daily with 50 micrograms porcine (p) GH or with 0.9% saline for 2 weeks following the surgeries. Changes in serum IGFBP concentrations were determined by Western ligand- or immuno-blot analysis. Neither IGFBP-5 nor -6 was detected in any of the treatment groups. Induction of IDDM increased serum concentrations of IGFBP-1 and -2 and reduced those of IGFBP-3 and -4. Although serum IGFBP-1 and -2 concentrations remained elevated in the HxDb rats compared with the NonDb controls, IGFBP-1 levels were reduced compared with those in the Db controls. Serum IGFBP-3 and -4 were reduced to levels below those in Db controls. Although IGFBP-3 and -4 concentrations were elevated to normal in AxDb rats, the IGFBP-2 concentration was increased above those in both NonDb and Db rats and the IGFBP-1 concentration was reduced. Administration of pGH increased serum IGFBP-4 concentrations in all groups and IGFBP-3 concentrations in all groups except the Db. In addition, pGH reduced the concentration of IGFBP-1 in HxDb rats and nearly abolished it in AxDb rats, but had no effect on IGFBP-1 concentration in NonDb or Db rats. Administration of corticosterone (B; 25 micrograms/ml of 0.9% saline drinking water) to AxDb rats restored Db-like profiles of all IGFBPs. The refractoriness of Db rats to pGH is associated with a failure of the hormone to elevate IGFBP-2 and -3 titers and to reduce those of IGFBP-1. Adrenal B production appears to be responsible for this resistance to GH. However, the elevated IGFBP-2 concentration in Db rats does not appear to be due to B or any other pituitary-controlled or -derived factors. Impaired growth was associated with substantially reduced IGFBP-3 concentrations and elevated IGFBP-1, whereas growth restoration was associated with the opposite changes.

Corticosterone regulation of insulin-like growth factor I, IGF-binding proteins, and growth in streptozotocin-induced diabetic rats.

The experiments reported herein were conducted to determine how corticosterone regulates growth and plasma insulin-like growth factor (IGF) I and IGF-binding protein (IGFBP) concentrations in normal and streptozotocin (STZ)-induced diabetic rats. Males were bilaterally adrenalectomized (Ax) or sham Ax and given intravenous injections of 0, 30, or 65 mg STZ per kg body wt (0, 30, or 65 STZ) to induce varying degrees of insulin deficiency and implanted with 100-mg pellets containing 0, 40, or 80% corticosterone in cholesterol. Changes in plasma IGFBP concentrations were determined by Western ligand blotting or immunoblots. Neither IGFBP-5 nor IG-FBP-6 was detected in any of the treatment groups. Plasma IGFBP-2 was elevated and IGF-I was reduced in the nondiabetic Ax rats compared with sham Ax controls, but plasma IGFBP-3 and -4 were not significantly changed. Adrenalectomy had no affect on tibial growth or plasma IGFBP-1 in these animals. Plasma IGF-I, IGFBP-1 and -3, and tibial growth were equal among 0, 30, and 65 STZ Ax rats that did not receive corticosterone. Plasma IGFBP-4 was inversely related to the amount of STZ injected in these animals, and IGFBP-2 was elevated in those given the high dose of STZ. In the 0 STZ Ax rats, plasma IGF-I and IGFBP-3 increased in proportion to the corticosterone implant dose, but IGFBP-1 was unaffected. By contrast, IGF-I and IGFBP-3 were unaltered by corticosterone in the 30 STZ Ax rats, and IGFBP-1 increased in proportion with the dose of corticosterone.(ABSTRACT TRUNCATED AT 250 WORDS)

Comparison of the ability of normal mouse mammary tissues and mammary adenocarcinoma to cleave rat prolactin.

Previous studies have shown that target tissues cleave rat prolactin (rPRL) to form a two-chain derivative that yields approximately 16- and approximately 6-kDa fragments upon reduction. Both cleaved rPRL and the purified 16-kDa fragment have novel biological activities. Thus, cleavage may be of physiological significance. We determined whether normal mammary tissue or stroma (lacking mammary epithelial cells [MEC]) or mammary tumor tissue could cleave rPRL in vitro. Tumor explants were derived from a transplantable mammary tumor cell line (TPDTMT-4EP). The hormone was incubated with explants of those tissues from female BALB/c or DDD mice. Medium samples were processed by SDS-PAGE, and the relative abundance of intact and cleaved rPRL was determined by densitometric analysis of immunoblots using an antiserum to the 16-kDa fragment of rPRL. After 2-hr of incubation, normal DDD mammary explants cleaved approximately 25% of added rPRL, but tumor explants cleaved none. Explants of intact virgin BALB/c mouse mammary gland and of parenchyma-free "cleared" mammary fat pad cleaved rPRL equally well, but explants of abdominal adipose tissue had low cleaving activity. Homogenates of cleared mammary fat pads that were incubated with rPRL contained a high proportion of the cleaved form but none of the free 16-kDa fragment. These results indicate that cleavage of rPRL is highly specific to mammary stroma. Such cleavage may yield a form of the hormone that has novel activities within the mammary gland or elsewhere in the body.

Hypophysectomy or adrenalectomy of rats with insulin-dependent diabetes mellitus partially restores their responsiveness to growth hormone.

The studies reported herein were conducted to confirm that the pituitary gland is involved in maintaining growth hormone (GH) resistance in rats with insulin-dependent diabetes mellitus (IDDM) and to determine whether the adrenocorticotropic hormone (ACTH)-adrenal cortical axis is responsible. The rats were made diabetic by injecting streptozotocin (85 mg/kg body wt) IP once daily on two consecutive days. They were then injected with 15 IU insulin SC twice daily on two consecutive days to enable them to survive hypophysectomy or adrenalectomy. Intact nondiabetic (NonDb), diabetic (Db), hypophysectomized diabetic (HxDb), and adrenalectomized diabetic (AxDb) rats were injected twice daily with 50 micrograms porcine (p) GH or with 0.9% saline for 2 weeks following the surgeries. Serum glucose levels of the saline-injected Db, HxDb, and AxDb rats were significantly greater than those of the NonDb rats by 106%, 65% and 49%, respectively. However, the levels in the HxDb and AxDb animals were significantly lower than those of the Db group by 20% and 28%, respectively. Injections of pGH into NonDb rats increased serum glucose concentrations by 38%, over their saline-treated controls, and by 29% in AxDb rats. This diabetogenic effect of GH was not seen in any other group. Administration of pGH to Db rats failed to increase body weight gain, tall growth, tibial epiphysial plate width, or serum IGF-I concentration over saline-injected controls. By contrast, HxDb and AxDb rats injected with pGH showed significant increases in all four growth parameters. Total serum IGF-I concentrations in AxDb rats injected with pGH equaled those in NonDb controls. To determine whether the lack of corticosterone (B) in the AxDb rats was responsible for the reduced hyperglycemia and restored responsiveness to pGH, AxDb rats were given B in their drinking water at 5 or 25 micrograms/ml. Administration of B reduced the beneficial effects of adrenalectomy by restoring hyperglycemia and growth impairment, and partially restored resistance to the pGH injections. These studies confirm that the pituitary contributes to diabetic growth impairment and show that the ACTH-adrenal cortical axis is primarily responsible for the GH-resistant state that develops in rats with IDDM.

Cleavage of rat prolactin occurs within rat mammary tissue, and the cleaved form is present in serum but not in milk.

The present study was undertaken to determine whether cleavage of rat prolactin (rPRL) occurs within rat mammary tissue and to investigate whether the cleaved form and its 16-kDa amino terminal fragment are present in rat serum and milk. The relative abundance of the different forms of rPRL was determined by SDS-PAGE and immunoblot analysis using an antiserum to the 16-kDa fragment. Explants of mammary glands from lactating rats were incubated with intact rPRL. The tissue was then washed extensively to remove external PRL forms. The ratios of cleaved to intact rPRL in mammary tissue homogenate, in mammary-conditioned medium, and in serum from lactating rats were 4.2, 0.44, and 0.16, respectively. Although intact rPRL was found in milk, the cleaved form was not detectable. The 16-kDa fragment of rPRL was not detected in concentrated extracts of either serum or milk. These results indicate that lactating rat mammary tissue can internalize and cleave rPRL, but the cleaved form does not appear in milk. The cleaved rPRL is present in rat serum, but no evidence was obtained for the presence of the 16-kDa fragment of rPRL in either serum or milk.

Pancreatic hormones differentially regulate insulin-like growth factor (IGF)-I and IGF-binding protein production by primary rat hepatocytes.

We investigated the influence of and interactions among pancreatic hormones on the secretion of insulin-like growth factor-I (IGF-I) and IGF-binding proteins (IG-FBPs) by treating primary hepatocytes from young male Long-Evans rats with insulin or glucagon in combination with rat GH (rGH). The concentration of IGF-I secreted into the medium was estimated by radioimmunoassay after formic acid-acetone cryoextraction, and secreted IGFBPs were analysed by Western ligand blot and immunoblot; accumulation of IGF-I mRNA was analysed by Northern blot. Both insulin (0.1-100 nmol/l) and rGH (0.5, 5 and 50 pmol/l) produced a dose-dependent stimulation of IGF-I secretion over a 24-h incubation period. In contrast, glucagon (0.1-100 nmol/l) inhibited IGF-I production in a dose-related manner. Glucagon (10 nmol/l) also inhibited IGF-I secretion stimulated by rGH (5 pmol/l) and insulin (10 nmol/l). Northern blot analysis of total RNA isolated from rat hepatocytes revealed that rGH (5 pmol/l) elevated IGF-I mRNA levels, glucagon (10 nmol/l) alone had no effect on this parameter, but glucagon significantly reduced IGF-I transcript accumulation in response to rGH. IGFBPs secreted by rat hepatocytes run in two molecular weight ranges on SDS-PAGE: approximately 25 kDa (IGFBP-4) and approximately 29-31 kDa (IGFBP-1 and -2); the predominant hormonally regulated IGFBP was identified as IGFBP-1. Insulin produced a dose-dependent inhibition of production of IGFBP-1, while glucagon was stimulatory; when given together at an equivalent concentration (1 nmol/l), the effects of insulin were dominant to glucagon on IGFBP-1. These observations provide support for significant opposite roles for the pancreatic hormones, insulin and glucagon, in the regulation of liver IGF-I and IGFBP-1 production. As the production of pancreatic hormones is influenced by nutritional status, these polypeptides may mediate the effects of changing nutritional state on the hormonal control of protein anabolism and glucose homeostasis by directly influencing the circulating level of liver-derived IGF-I and its binding proteins.

Characterization of rat serum insulin-like growth factor-binding proteins by two-dimensional gel electrophoresis: identification of a potentially novel form.

The insulin-like growth factor-binding proteins (IGFBPs) in rat serum were analyzed by two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis followed by Western ligand or immunoblotting. A ligand blot of adult female rat serum revealed at least 20 proteins that bound labeled IGF-I specifically. They ranged in size from about 24-50K, and their pI values ranged from approximately 5.5-8.0. Immuno- and ligand blots showed that IGFBP-3 appeared as a broad band composed of numerous individual spots of about 40-45K, with pI values ranging from about 5.5-7.7. Immunoblots showed that IGFBP-4 resolved into at least three spots of approximately 29K, with pI values of about 6.3-6.7 and an approximately 24K nonglycosylated form that usually appeared as a single spot with a pI of about 7, but occasionally it resolved into a doublet. Immunoblots of neonatal rat serum using antiserum to IGFBP-2 resulted in immunoreactivity of two sets of proteins: approximately 33K forms composed of two proteins and approximately 30K forms comprising at least five different variants with pI values of about 7.0-7.5. However, ligand analysis showed that only the 30K forms had binding activity. IGFBP-1, -5, and -6 were not detectable by immunoblotting. However, a set of about 29K IGFBPs with pI values of approximately 5.9, which appears in significant amounts in the serum of pups and diabetic rats and in placental tissue-conditioned medium, has been tentatively identified as IGFBP-1 by ligand blotting. One finding of particular interest is an approximately 50K BP with a pI of about 6 that comigrates with IGFBP-3. However, unlike IGFBP-3, which is glycosylated and undergoes proteolysis during gestation, this approximately 50K IGFBP is not susceptible to endoglycosidase-F treatment and persists throughout pregnancy. Immunoblotting analysis revealed weak cross-reactivity between the approximately 50K IGFBP and antiserum to IGFBP-4. These results show that the rat serum IGFBPs are more heterogeneous than was previously realized. The two-dimensional system will allow changes in these different forms to be evaluated critically in different physiological and experimental states.

Mass spectrometric analysis of the fragments produced by cleavage and reduction of rat prolactin: evidence that the cleaving enzyme is cathepsin D.

The site(s) at which mammary tissue enzymatically cleaves rat (r) PRL, and the possibility that the cleaving activity is cathepsin D, were investigated using mass spectrometry and enzyme inhibitors. Cleavage of intact rPRL [22,566 atomic mass units (amu)] by either mammary gland-conditioned medium or cathepsin D (both at pH 3) reduced the mass of the molecule by 397 amu. Subsequent reduction of the large rPRL fragment cleaved by either method generated two fragments of 16,364 and 5808 amu. The mass of the smaller fragment is consistent with the report of Vick et al. (Biochim Biophys Acta 931: 196-204, 1987) that its amino-terminal residue is Ser149. These results indicate that both enzyme preparations cleave rPRL by excision of the tripeptide Leu-Val-Trp (mass = 397 amu) between residues 145 and 149. The ability of both enzyme preparations to cleave rPRL at pH3 was inhibited by pepstatin A but not by phenylmethane sulfonyl fluoride, and both preparations were essentially inactive at pH7. Accordingly, the PRL-cleaving activity of rat mammary tissue is probably cathepsin D.

The primary structure of sturgeon prolactin: phylogenetic implication.

The complete amino acid sequence of prolactin (PRL) from a chondrostean species, the sturgeon (Acipenser gueldenstaedti), has been determined. Sturgeon PRL was isolated from the pituitary glands by gel filtration on a Sephadex G-25 column and high-performance liquid chromatography on a reverse-phase column following acid-acetone extraction. Sturgeon PRL was identified by immunoblot reactivity using antisera against salmon and ovine PRL. It consists of 204 amino acid residues, which is the largest among known PRLs, and contains three disulfide bonds corresponding to those of tetrapod PRLs. Sequence comparison with PRLs from other vertebrates revealed that sturgeon PRL has slightly higher sequence identities (35-46%) with teleost PRLs than with tetrapod PRLs (30-40%). These structural characteristics imply that an ancestor of the ray-finned fishes had PRL with three disulfide bonds and at some point after divergence of Chondrostei, the disulfide bond in the amino-terminus of PRL was lost.

Lungfish prolactin exhibits close tetrapod relationships.

This paper describes the isolation and the complete amino-acid sequence of prolactin (PRL) from the pituitary glands of African lungfish, Protoputerus aethiopicus. We purified the hormone from an alkaline extract of the pituitaries using a two-step chromatographic procedure by detecting specific immunoblot reactivity with rabbit antisera against salmon PRL. The lungfish PRL consists of 200 amino-acid residues. Sequence comparison revealed that the PRL shows 66% identities with amphibian, reptilian and bird PRLs, 57% with mammalian PRLs, and 38% with teleost (modern bony fish) PRLs. Moreover, the PRL contains three disulfide bonds homologous to those of tetrapod PRLs and differs from teleost PRLs which lack the amino-terminal disulfide bond. Thus, the structural features of lungfish PRL indicate a closer relationship to tetrapod PRLs than to teleost PRLs. All PRLs sequenced to date share 22 common amino acids, which may be important for the activities common to all PRLs.

Effects of prolactin, growth hormone, and triiodothyronine on prolactin receptors in larval and adult tiger salamanders (Ambystoma tigrinum).

The effects of porcine growth hormone (pGH) or ovine prolactin (oPRL) alone and in combination with triiodothyronine (T3) on renal PRL receptors were determined in both pre- and post-metamorphic tiger salamanders (Ambystoma tigrinum). The protein hormones were given at a dose of 1.0 micrograms/gm body weight/day and the T3 was given at 10.0 ng/gm body weight/day. The duration of treatment was 7 days. Effects on growth, and plasma thyroid hormone levels were also determined. Ovine PRL increased growth in both larvae and adults and reversed metamorphic changes. Administration of T3 increased the plasma T3 concentration, as measured by radioimmunoassay, and when given alone caused weight loss at both stages. The GH decreased plasma T4 and increased plasma T3 concentrations, indicating that it caused an increase in T4 deiodination. In adults the renal PRL receptor affinity of 2.9 +/- 0.7 x 10(10) L/mol and capacity of 160 +/- 22 fmol/mg protein were higher than the corresponding values of 1.8 +/- 0.3 x 10(10) L/mol and 29.2 +/- 3.8 fmol/mg in larvae. In adults only, there is an additional low-affinity, high capacity PRL binding site. The oPRL treatment decreased the binding capacity of 33.2 +/- 1.2 and 5.9 +/- 4.9 fmol/mg in adults and larvae, respectively. By contrast, pGH increased the capacities to 249 +/- 18 and 62.1 +/- 6.8 fmol/mg in adults and larvae, respectively. Treatment with T3 alone doubled the oPRL binding capacity to 58.3 +/- 4.7 fmol/mg in larvae, but there was no effect in adults. In both developmental stages the effects of oPRL and pGH on the receptors were not changed by the simultaneous T3 treatment.(ABSTRACT TRUNCATED AT 250 WORDS)

Experimental diabetes mellitus in a teleost fish. II. Roles of insulin, growth hormone (GH), insulin-like growth factor-I, and hepatic GH receptors in diabetic growth inhibition in the goby, Gillichthys mirabilis.

Insulin-dependent diabetes mellitus (IDDM), when untreated or poorly controlled in mammals, results in growth retardation. To determine whether the same relationship exists in an ectothermic vertebrate, IDDM-like symptoms were induced in a teleost fish, the goby Gillichthys mirabilis, by surgical removal of its pancreatic endocrine (islet) organ. Isletectomized (Ix) gobies lost body weight, their skeletal growth was retarded, as measured by changes in body length, and they exhibited a 50% reduction in cartilage 35SO4 incorporation in vitro, consistent with changes that occur in mammals with IDDM. Injections of bovine insulin into the Ix fish restored body growth parameters to control levels and stimulated cartilage 35SO4 incorporation in a dose-related manner. In contrast to mammals with IDDM, which are resistant to GH action, injection of teleost GH stimulated cartilage 35SO4 incorporation in the Ix fish. Furthermore, whereas cartilage from rats with IDDM is resistant to stimulation by insulin-like growth factor-I (IGF-I) in vitro, cartilage explants from the Ix fish were highly responsive to recombinant bovine IGF-I, exhibiting a dose-dependent stimulation of 35SO4 incorporation. As far as we are aware, these results represent the first demonstration of diabetic growth inhibition in an ectothermic vertebrate. This inhibition is similar to that which occurs in mammals with IDDM in some respects, but is different in others, as the diabetic fish did not develop resistance to growth stimulation by either GH or IGF-I. While these results support a role for insulin in maintaining the GH-IGF-I-growth axis in this ectothermic vertebrate, there may be important differences in the role of insulin in the promotion of anabolic processes.

Evidence for a role of the liver in the luteotropic action of prolactin in rats.

Recent studies indicate that the liver may participate in growth-promoting and lactogenic activity of prolactin (PRL). Accordingly, our study was designed to determine whether the liver participates in the luteotropic activity of the hormone in rats. Normally cycling Long-Evans rats received infusions of solvent or 50 micrograms ovine (o) PRL per day into either the external jugular vein (EJV) or the hepatic portal vein (HPV) via osmotic minipumps. The modes of delivery were continuous, 2 pulses of 8 h each per day (2-8), or 4 pulses of 2 h each per day (4-2). Observation of daily vaginal smears was used to classify rats as either cyclic or predominantly diestrus (PD). In a second study, rats fitted with osmotic minipumps received oPRL or vehicle as above, and serial blood samples were obtained for measurement of serum oPRL and rat (r) PRL concentrations. Estrous cycles of normal length were observed in 9 of 11 rats (82%) receiving solvent infusions. All 8 animals receiving continuous oPRL infusions were classified as PD, regardless of site of infusion. In addition, in 4 of 5 (80%) HPV-infused and 5 of 6 (83%) EJV-infused rats on the 2-8 pulse schedule, the smear patterns were classified as PD. However, when rats received 4-2 EJV, 9 of 13 animals (69%) were classified as cyclic, whereas 9 of 11 rats (82%) were classified as PD when oPRL was similarly pulsed into the HPV. Differences in serum concentrations of oPRL between oPRL-treated and vehicle-infused rats were significant only for continuously infused animals.(ABSTRACT TRUNCATED AT 250 WORDS)

An insulin-like growth factor I-resistant state in cartilage of diabetic rats is ameliorated by hypophysectomy. Possible role of metabolism.

This study investigated the effect of IDDM on cartilage anabolic activity in rats. Rats were injected with STZ to induce IDDM, were hypophysectomized, or were injected with STZ and hypophysectomized. After 14 days, control (intact and sham-Hx) and Hx rats were normoglycemic, whereas the rats with IDDM exhibited hyperglycemia and glycosuria. The HxDb rats, however, had normal blood glucose levels and no glycosuria. Both growth, serum levels of IGF-I, and basal cartilage 35SO4 incorporation measured in vitro were decreased in the Hx, IDDM, and HxDb groups. IGF-I added in vitro significantly stimulated 35SO4 incorporation by cartilage explants from control and Hx animals, whereas explants from the animals with IDDM were unresponsive. Explants from the HxDb rats, however, were stimulated by IGF-I in a dose-related manner. Because Hx corrected the glycemic status of the IDDM rats and restored cartilage responsiveness to IGF-I, a second set of experiments was undertaken to further investigate the relationship between cellular metabolism and anabolic activity in cartilage. Cartilage explants from rats fasted for 48 h showed significantly decreased basal 35SO4 incorporation, which was as low as that in explants from rats with severe IDDM. Whereas explants from the IDDM rats were completely unresponsive, those from the fasted rats (and fed rats) were significantly stimulated by the added IGF-I. However, incubation in the presence of 2-D-G, which causes intracellular glucopenia, or in the absence of glucose, completely blocked the anabolic response to IGF-I in otherwise responsive tissues. In conclusion, an important component of diabetic growth inhibition appears to be tissue resistance to the anabolic action of IGF-I, a condition that is correctable by Hx and that may be a result of metabolic impairment at the tissue level.

Restoration of lactation in bromocriptine-treated rats by prolactin replacement: comparison of constant versus pulsatile infusion and intrahepatic versus intrajugular routes of delivery.

The effectiveness of pulsed vs constant infusion of ovine(o) prolactin (PRL), given by different schedules, at restoring lactation in PRL-suppressed rats was compared, and the possibility that the liver participates in the restorative effects of the infused hormone was investigated. Lactating dams were given subcutaneous injections of bromocriptine (BC) between days 7 and 12 postpartum to suppress endogenous PRL secretion. Osmotic minipumps were used to infuse the oPRL into either the jugular vein or the hepatic portal vein. The latter route would expose the liver to higher concentrations of PRL than would intrajugular infusion. Constant infusion of oPRL in different doses was, overall, more effective at restoring lactation (i.e. litter weight gain) than was giving pulses, regardless of the site of delivery. Infusion of the PRL at 100 micrograms/rat/day in pulses of 1h duration was ineffective at frequencies of either 4 or 8/day, whereas pulses of 2h duration were effective at both of these frequencies. Infusing that dose of oPRL was equally effective whether it was given in 4 or 8 pulses/day of 2 h duration. Intrahepatic infusion of oPRL was not more effective than intrajugular delivery regardless of the schedule of administration. These results indicate that pulse duration is a more important determinant of the effectiveness of the galactopoietic action of PRL in the lactating rat than is pulse frequency. No evidence was obtained that the liver participates in the galactopoietic effects of PRL.

Human placental lactogen directly inhibits rat cartilage growth processes in vivo and in vitro.

We have recently reported that human placental lactogen inhibits the growth of young female rats without changing the serum levels of insulin like growth factor-I. Accordingly, experiments were conducted to determine whether human placental lactogen could directly inhibit cartilage growth processes in vivo and in vitro. Osmotic minipumps with attached polyethylene catheters were used to infuse the hormone for seven days into the left hindlimb of three-month-old female rats via the common iliac artery. The right hindlimb of each animal served as an internal control. Infusion of the placental lactogen at 10 micrograms/rat/day caused a slight (10%) but significant decrease in the width of the tibial epiphysial cartilage plate and a higher dose (100 micrograms/rat/day) caused a greater degree of inhibition (25%). However, the higher dose also inhibited the tibial cartilage plate of the contralateral (non-infused) limb. The possibility that human placental lactogen could directly inhibit cartilage anabolic activity in vitro was evaluated by measuring the incorporation of 35SO4 into costal cartilage explants from three to four-month-old female rats. The placental hormone inhibited the incorporation of 35SO4 in a dose-related manner at concentrations ranging from 1.0 to 100 micrograms/l. As a test of the specificity of this inhibition the effect of the hormone on the incorporation of 35SO4 into cartilage explants from Coho salmon was determined. The placental lactogen did not affect incorporation of the sulfate into the fish cartilage over a range of doses from 1.0 to 1000 micrograms/l.(ABSTRACT TRUNCATED AT 250 WORDS)