Despite the unquestioned galactopoiesis evoked in cows by thyroxine or dried thyroid gland [see Folley, 1940, for review] there is little unanimity about the effect of thyroidectomy on lactation [see Folley, 1938]. As regards the rat in particular there is a clear divergence of opinion on this point. Nelson & Tobin [1937] found that their rats weaned normal litters after thyroidectomy performed during pregnancy. Folley [1938], on the other hand, showed that thyroidectomy carried out early in lactation resulted in a prompt reduction in the growth of the pups, only a small proportion of which lived to be weaned and then only because the death of the majority of the young reduced the demands on the depleted milk supply. Nelson [1939] subsequently reaffirmed his original findings, though he now reported a slight retardation of the growth of litters of rats thyroidectomized during the current lactation. Results which may be considered

A very prolonged effect of certain hormones can be obtained by the subcutaneous implantation of a compressed tablet of undiluted, crystalline substance [Deanesly and Parkes, 1937]. This technique has proved to be specially useful with oestrone and testosterone, or testosterone propionate, and effects lasting many months can be obtained from a single administration. The great efficiency of the method seems to be due to slow and even absorption over a long period, so that the alternation of wasteful excess and sub-threshold concentrations, such as is often produced by the daily injection of solutions, is avoided. In considering whether such a technique could be usefully applied to other hormones it was reasonable to take account of adrenalin, the most rapidly acting and quickly destroyed of the hormones. Further, of the various physiological responses to adrenalin, that of the blood-sugar seemed the most promising for investigation of the effects of prolonged action.

Abstract

The aim of this study was to determine whether Type II phospholipase A₂ (PLA₂) is released from late pregnant human placental tissue. Placental explants were incubated in vitro and the release of immunoreactive (ir) Type II PLA₂ and PLA₂ enzymatic activity into the medium was determined. Both irType II PLA₂ and PLA₂ enzymatic activity accumulated in the incubation medium in a time-dependent manner (P<0·0001). This release was not associated with a loss of cell membrane integrity, as indicated by measurement of the intracellular enzyme, lactate dehydrogenase, in the incubation medium. The concentration of irType II PLA₂ and PLA₂ enzyme activity present in incubation medium were significantly correlated (P<0·01). Consistent with the hypothesis that Type II PLA₂ may be stored in secretory granules within human placental tissue, incubation in the presence of a membrane depolarising concentration of KCl (60 mm) caused the release of irType II PLA₂ 2·0-fold (P<0·001). PLA₂ enzyme activity released into the incubation medium displays biochemical characteristics consistent with those previously reported for secretory PLA₂ isozymes, that is, a requirement for millimolar concentrations of calcium for optimal enzyme activity, inhibited by reducing agents, such as dithiothreitol and insensitive to heat inactivation. The data obtained in this study establish that irType II PLA₂ is released from term placenta, when incubated in vitro. The release of this extracellularly-active PLA₂ isozyme may contribute to gestational and labour-associated increases in glycerophospholipid metabolism and prostaglandin formation.

Journal of Endocrinology (1997) 153, 151–157

ABSTRACT

The hypogonadal (hpg) mouse has a congenital deficiency in gonadotrophin-releasing hormone and the gonads consequently lack exposure to endogenous gonadotrophins during development. To determine the effect of FSH on Leydig cell function in these animals adult hpg mice were injected twice daily with FSH (2 μg injections) or LH (40 ng injections, the presumed LH contamination of FSH used). Following FSH treatment there was a clear stimulation of the seminiferous epithelium and in animals injected with FSH plus [³H]thymidine, the incorporation of label was largely confined to the germ cells with no apparent uptake by the Sertoli cells. In FSH-treated testes the Leydig cells contained numerous large lipid droplets, similar to the unstimulated hpg testis. There was no evidence of the interstitial hyperplasia which is observed following injection of high doses of LH (2 pg twice daily).

There was no change in basal androgen content of the testis in vivo following FSH treatment but injection of a maximal dose of human chorionic gonadotrophin (hCG), 1 h before death, markedly increased testicular androgen content only in the FSH-treated group. Testicular androgen production in vitro was significantly increased following FSH treatment both under basal conditions (FSH-treated, 17·4 pmol/testis; control, 1·46 pmol/testis) and during stimulation by hCG (FSH-treated, 940 pmol/testis; control, 81 pmol/testis). Associated with the increased androgen production following FSH treatment there were significant increases in the activities of three steroidogenic enzymes; cholesterol side-chain cleavage (186-fold increase over control), 17α-hydroxylase (103-fold increase) and 17-ketosteroid reductase (177-fold increase). The fourth enzyme involved in androgen synthesis, 3β-hydroxysteroid dehydrogenase, shows relatively high activity in the control hpg testis and was only increased by sixfold following FSH treatment. There was no effect of FSH on 5α-reductase activity.

Results show that FSH causes a marked stimulation of the steroidogenic capacity of the hpg testis. Leydig cells do not contain FSH receptors and it is assumed that FSH acts through paracrine factors released by the Sertoli cells.

Journal of Endocrinology (1992) 135, 517–525

Short stature and osteoporosis are common in Duchenne muscular dystrophy (DMD) and its pathophysiology may include an abnormality of the growth hormone/insulin-like growth factor-1 (GH/IGF-1) axis, which is further exacerbated by long-term glucocorticoid (GC) treatment. Hence, an agent that has anabolic properties and may improve linear growth would be beneficial in this setting and therefore requires further exploration. A 5-week-old x-linked muscular dystrophy (mdx) mice were used as a model of DMD. They were treated with prednisolone ± GH + IGF-1 for 4 weeks and then compared to control mdx mice to allow the study of both growth and skeletal structure. GC reduced cortical bone area, bone fraction, tissue area and volume and cortical bone volume, as assessed by micro computed tomography (CT) In addition, GC caused somatic and skeletal growth retardation but improved grip strength. The addition of GH + IGF-1 therapy rescued the somatic growth retardation and induced additional improvements in grip strength (16.9% increase, P  < 0.05 compared to control). There was no improvement in bone microarchitecture (assessed by micro-CT and static histomorphometry) or biomechanical properties (assessed by three-point bending). Serum bone turnover markers (Serum procollagen 1 intact N-terminal propeptide (P1NP), alpha C-terminal telopeptide (αCTX)) also remained unaffected. Further work is needed to maximise these gains before proceeding to clinical trials in boys with DMD.

ABSTRACT

Hypogonadal (hpg) mice have a congenital deficiency of hypothalamic gonadotrophin-releasing hormone (GnRH) and the gonads consequently lack exposure to gonadotrophins during development. We injected male hpg mice with LH for 10 days to investigate whether LH alone can stimulate normal steroidogenesis in these animals. Control animals had an inactive interstitium and very few germ cells. Testicular content of androgens was undetectable by radioimmunoassay in control animals unless a single injection of LH was given 1 h before death, when androgens were just detectable. Control testes incubated in vitro with [³H]pregnenolone demonstrated that without gonadotrophin stimulation pregnenolone was metabolized only to progesterone in significant amounts. Assay for cholesterol side-chain cleavage cytochrome P450 (P450scc) mRNA showed basal expression in saline-treated hpg mouse testis. LH treatment induced hypertrophy and hyperplasia of Leydig cells and division of germ cells. Testicular androgen content increased significantly, with testosterone and androstenedione as the major androgens. LH-treated testes incubated with [³H]pregnenolone in vitro had a greater synthetic capacity for testosterone, suggesting an increase in 17α-hydroxylase/C_17–20-lyase activity. Basal and human chorionic gonadotrophinstimulated androgen production in vitro increased markedly following LH treatment to levels previously described in the normal adult animal. LH treatment caused a rapid and transient increase in the hybridization of P450scc mRNA which was sevenfold greater than that of saline-treated controls when the animals were killed 1 h after the last injection but fell to control levels within 24 h of cessation of treatment.

We conclude that LH alone can stimulate steroidogenesis in testes of hpg mice, and that previous exposure of the tissue to physiological concentrations of endogenous gonadotrophins is not required to obtain normal steroidogenic responsiveness.

Journal of Endocrinology (1990) 125, 131–138

ABSTRACT

Oxytocin secretion is inhibited by opioids, and oxytocin is important in parturition. The effects on parturition of morphine, a relatively selective μ-opioid receptor agonist, were studied in the rat. Morphine or vehicle with or without the opiate antagonist naloxone were administered immediately after the birth of the second pup and the subsequent course of parturition was recorded in a total of 80 rats. Both s.c. morphine (10 mg/kg) and intracerebroventricular (i.c.v.) morphine (18 μg through a previously implanted cannula) interrupted parturition, delaying the birth of the sixth pup after treatment to 187·3 ± 35·9 (s.e.m.) min and 195·4 ± 19·5 min respectively, compared with 46·4 ± 3·7 and 66·1 ± 17·5 min after vehicle alone. The dose of morphine given i.c.v. had no effect when given s.c. Naloxone given concurrently prevented the effects of morphine. Eventually the rate of parturition in the morphine-treated groups recovered. Perinatal pup mortality rate was not increased when morphine was given to the mothers, but it did inhibit the expression of normal intrapartum maternal behaviour.

Pup mortality was increased 48 h post partum by morphine given during parturition, and it reduced the proportion of rats with normal maternal behaviour 24 h post partum. Morphine did not affect spontaneous or oxytocin-stimulated contractile activity of the parturient uterus in vitro. The concentration of oxytocin in trunk blood plasma was decreased 40 min after i.c.v. morphine (24·3 ± 3·9 vs 39·3± 6·5 pmol/l in controls), as was vasopressin (7·2 ± 1·5 vs 19·7 ± 4·5 pmol/l in controls). Intravenous infusion of oxytocin (2–5 mU/min for 144·3 ± 8·2 min; total infused 448·5 ± 61·9 mU) after i.c.v. morphine re-started parturition; all pups were born to these rats (mean time to pup 6, 110·3 ± 12·7 min) before the i.v. vehicle-infused rats given i.c.v. morphine re-started (mean time to pup 6, 406·3±125·2 min).

It is concluded that morphine given during parturition acts centrally through opioid receptors to inhibit oxytocin secretion, and impairs the expression of maternal behaviour. Reversal of the effects of morphine on parturition by i.v. oxytocin demonstrates the important role of oxytocin in fetus ejection and expulsion.

Journal of Endocrinology (1989) 121, 521–536

It is unknown whether there is a gene signature in pancreas which is associated with type 1 diabetes (T1D). We performed partial pancreatectomies on 30-day preinsulitic, diabetes-prone BioBreeding (BBdp) rats to prospectively identify factors involved in early prediabetes. Microarrays of the biopsies revealed downregulation of endoplasmic reticulum (ER) stress, metabolism and apoptosis. Based on these results, additional investigations compared gene expression in control (BBc) and BBdp rats age ~8, 30 and 60 days using RT-qPCR. Neonates had increased ER stress gene expression in pancreas. This was associated with decreased insulin, cleaved caspase-3 and Ins1 whereas Gcg and Pcsk2 were increased. The increase in ER stress was not sustained at 30 days and decreased by 60 days. In parallel, the liver gene profile showed a similar signature in neonates but with an early decrease of the unfolded protein response (UPR) at 30 days. This suggested that changes in the liver precede those in the pancreas. Tnf and Il1b expression was increased in BBdp pancreas in association with increased caspase-1, cleaved caspase-3 and decreased proinsulin area. Glucagon area was increased in both 30-day and 60-day BBdp rats. Increased colocalization of BIP and proinsulin was observed at 60 days in the pancreas, suggesting insulin-related ER dysfunction. We propose that dysregulated metabolism leads to ER stress in neonatal rats long before insulitis, creating a microenvironment in both pancreas and liver that promotes autoimmunity.

Leucine-rich repeat-containing G-protein-coupled receptor 8 (LGR8, or RXFP2) is a member of the type C leucine-rich repeat-containing G protein-coupled receptor family, and its endogenous ligand is insulin-like peptide-3 (INSL3). Although LGR8 expression has been demonstrated in various human tissues, including testis, ovary, brain and kidney, the precise roles of this receptor in many of these tissues are unknown. In an effort to better understand INSL3–LGR8 systems in the rat, we cloned the full-length Lgr8 cDNA and investigated the presence and cellular localization of Lgr8 mRNA expression in adult and developing rat kidney. On the basis of these findings, we investigated the presence and distribution of renal ¹²⁵I-labelled human INSL3-binding sites and the nature of INSL3–LGR8 signalling in cultured renal cells. Thus, using in situ hybridization histochemistry, cells expressing Lgr8 mRNA were observed in glomeruli of renal cortex from adult rats and were tentatively identified as mesangial cells. Quantitative, real-time PCR analysis of the developmental profile of Lgr8 mRNA expression in kidney revealed highest relative levels at late stage gestation (embryonic day 18), with a sharp decrease after birth and lowest levels in the adult. During development, silver grains associated with Lgr8 mRNA hybridization were observed overlying putative mesangial cells in mature glomeruli, with little or no signal associated with less-mature glomeruli. In adult and developing kidney, specific ¹²⁵I-INSL3-binding sites were associated with glomeruli throughout the renal cortex. In primary cultures of glomerular cells, synthetic human INSL3 specifically and dose-dependently inhibited cell proliferation over a 48 h period, further suggesting the presence of functional LGR8 (receptors) on these cells (mesangial and others). These findings suggest INSL3–LGR8 signalling may be involved in the genesis and/or developmental maturation of renal glomeruli and possibly in regulating mesangial cell density in adult rat kidney.