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This chapter discusses about prolactin, which is the specific hypophysiotropin that promotes lactation in the pregnant mammal. Prolactin is distinguished from other pituitary tropins by its inhibitory regulation originating in the central nervous system, that is, the hypothalamus, by lack of a reciprocal peripheral feedback mechanism; by its diverse effects among the vertebrates, including growth-promoting actions in several species; and by its close association with growth hormone. It is probably present in the hypophysis of all vertebrates, but it is more plentiful in some species and at certain stages of the life cycle than at others. It is not soluble in any fat solvent. Prolactin-like activity in urine has been demonstrated both by pigeon crop sac assay and by observation of the effect of urine on the hormonally-primed mammary gland of the immature hypophysectomized male rat. Present evidence indicates that mainly one anterior pituitary hormone prolactin—and perhaps also somatropin hormone (STH)—is involved in the development of the mammary gland. A most interesting development during the past few years is the growing evidence that prolactin and STH in combination can induce full mammary lobulo-alveolar growth in the apparent absence of ovarian hormones in the rat.

a) Introduction

Interest in the physiology of pituitary prolactin greatly increased in recent years, as indicated by the large number of investigations on this subject since 1960. Prolactin is distinguished from other pituitary tropins by its inhibitory regulation originating in the central nervous system, i.e. the hypothalamus, by lack of a reciprocal peripheral feedback mechanism, by its diverse effects among the vertebrates, including growth-promoting actions in several species, and by its close association with growth hormone.

b) Storage and Release

Prolactin is probably present in the hypophysis of all vertebrates (NICOLL, 1968), but it is more plentiful in some species and at certain stages of the life cycle than at others. Ox and sheep glands usually yield 30 to 40 IU per gram of fresh tissue, while pork glands contain much less. Measurements of the prolactin content of human pituitaries indicate that the hormone accounts for about 2 percent of the acetone-dried residue of the gland. More prolactin is contained in the adenohypophyses of goats, sheep and cows, and less in the glands of cats and rabbits. In most species the prolactin content of the pituitary increases steadily during pregnancy, with an increase sometimes occurring soon after parturition. This is also reflected in the MTI (Fig. 10). When nursing is permitted, the prolactin content remains at this higher level for a longer period than when nursing is prevented.

c) Properties

Prolactin has been prepared in a highly purified form from ox and sheep pituitaries. Isolation procedures have been described by LI (1961) in a review on the biochemistry of prolactin. The physical properties of sheep (198 aminoacids) and ox (205 aminoacids) prolactin are remarkably similar. The hormone from both species has a molecular weight of 24,000 to 26,000, an isoelectric point at pH 5.73, and a sedimentation constant of 2.19. DIXON and LI (1964) showed that the hormone molecule consists of a single peptide chain with one free amino acid group, but no free carboxy end-group. They suggested that the peptide chain has an intra-chain disulfide bridge forming a ring similar to that present in vasopressin and oxytocin. The results of their structural studies are summarized in the following schematic representation of prolactin:

In order to investigate aggregation and charge differences between prolactin components, a combination of gel filtration, ion-exchange chromatography, and zone electrophoresis was used by SLUYSER and LI (1964). As a result of these investigations, a method was developed for the isolation of monomer prolactin in highly purified form. The experiments presented in their paper show that the various peaks or bands which appear when prolactin is submitted to either ion exchange chromatography or zone electrophoresis are due to the presence of molecular aggregates in the preparations. The degree of aggregation differed from batch to batch. Some prolactin preparations contained about 20% monomer whereas others had a much lower content of unassociated material. In view of the reported finding that some aggregation of prolactin can occur when solutions are submitted to lyophilization, pervaporation, sucrose concentrates, or extremes of pH, it seems likely that the differences in degree of aggregation between various batches of lactogenic hormone arise at least partly during the preparation of the samples. In order to prevent aggregation, the hormone solution was concentrated by chromatography on small columns of DEAE-cellulose. A separation was achieved between fractions representing different states of polymerization of prolactin molecules. One of the fractions contained monomer prolactin, as revealed by exclusion chromatography and sedimentation equilibrium experiments. The molecular weight of monomer prolactin was estimated to be 23,000.

Prolactin is not soluble in any fat solvent. It is, however, soluble in water, except in the isoelectric region pH 5—6, and in strongly acid solution. It may be salted out with sodium or ammonium sulfate or sodium chloride, the required amount depending on the pH. It is also precipitated by basic salts of heavy metals at or near pH 7. It is rapdily inactivated by trypsin and pepsin. Salt-free solutions at pH 8 withstand boiling for one hour with only negligible loss; under other conditions, prolactin may be rapidly destroyed. Frozen preparations are not stable, once they have been thawed.

Despite the great similarities of bovine and ovine prolactin, certain differences have been observed. It seems that, proportionate to weight, ovine prolactin is more active than bovine prolactin. There is less tyrosine in ovine prolactin, and distribution of the two hormones in countercurrent analysis is different. However, BRYANT and GREENWOOD (1968) showed by radioimmunoassay of blood that ovine prolactin antigen can substitute for bovine prolactin. Human prolactin has yet to be characterized, but the fact that methods effective for isolating ox and sheep prolactin have completely failed when applied to human pituitaries suggests that primate prolactin may have peculiarities of structure which make it very similar to human growth hormone.

d) Prolactin in Body Fluids

Prolactin-like activity in urine has been demonstrated both by pigeon crop sac assay and by observation of the effect of urine on the hormonally-primed mammary gland of the immature hypophysectomized male rat. SEGALOFF and STEELMAN (1959) found daily up to 175 IU of prolactin in urine from menopausal women. In normal women the excretion was somewhat higher in the luteal phase of the cycle as compared with the follicular stage. Prolactin excretion also increased during the course of pregnancy. Prolactin-like activity of serum has been detected with both pigeon and mouse assays. The blood of lactating women has been found to contain prolactin activity (SIMKIN and GOODART, 1960). Somewhat smaller amounts of hormone were found in blood during the luteal phase of women with normal menstrual cycles. No prolactin activity was evident in the blood of women in the follicular phase of the menstrual cycle or in normal young men. These observations still require confirmation (cf. Chapter 26 —d).

e) Prolactin in Pituitary Tumors

Persistent lactation, with or without previous pregnancy, is encountered with considerable frequency in endocrine practice. The occurrence of this complaint in acromegalic women is well known. When lactorrhea and amenorrhea follow pregnancy (Chiari-Frommel syndrome), a functional disturbance in prolactin secretion may be present. General awareness that persistent lactation may also be a sign of pituitary tumor in the non-acromegalic patient followed the description in 1954 of fifteen such cases from Albright’s Clinic (Forbes-Albright syndrome—cf. Chapter 28). The problem has been studied in rats bearing pituitary “mammotropic” tumors by MIZUNO, TALWALKER and MEITES (1964).

f) Effect of Prolactin on Mammary Growth

Present evidence indicates that mainly one anterior pituitary hormone—prolactin, and perhaps also STH, are involved in the development of the mammary gland. The most convincing and impressive work was done by Lyons (1942), who injected small amounts of purified prolactin (according to standards of 1942) into the mammary ducts of spayed rabbits and obtained not only clear-cut mammary development and lactational response, but also histological evidence of cellular hyperplasia of the alveolar tissue. This “lactational” growth was characterized by the presence of...