James Norman Dent

Ultrastructural observations on the developmental cytology of the pituitary gland in the spotted newt

Abstract Observations have been made on the ultrastructure of the pituitary gland in the eastern spotted newt during embryonic, larval, and red eft stages. Secretory granules were first seen in the presumptive pars intermedia prior to hatching at a developmental stage equivalent to embryonic stage 40 of the harrison series for Amblystoma. In recently hatched larvae the cells of the pars intermedia reached definitive form, being characterized by granules of varying electron opacity. At this stage axon endings become apparent in both the pars nervosa and the pars intermedia. Up to the hatching stage the cells of the pars distalis were without granules but soon thereafter minute granules were apparent in some cells. By the premetamorphic stage agranular cells with elongate processes called stellate cells and a cell type with granules in the 180–250 mμ range were distinguished. Three more cell types appeared; one with smaller granules late in metamorphosis, one with a double population of granules in the first year eft and one with larger granules in the second year eft. The possible functions of these cell types are discussed.

Cytological response of the newt pituitary gland to thyroidal depression.

Abstract 1. 1. Cytological examination was made of pituitary glands and ultimobranchial bodies from specimens of Diemyctylus (Triturus) viridescens killed at various times after surgical thyroidectomy. Pituitary and thyroid glands were studied after various periods of treatment with radioiodine, thiourea, and potassium perchlorate. 2. 2. Approximately two weeks after surgical removal of the thyroid, large chromophobic thyroidectomy cells began to appear in the pars distalis of the pituitary. They were abundant after the first month and persisted to at least the eighth month in some animals. In other animals sacrificed six or more months after thyroidectomy, none of these cells was present. The disappearance of the thyroidectomy cells is discussed. 3. 3. The formation of thyroidectomy cells was inhibited by keeping the thyroidectomized animals immersed in solutions of thyroxine. 4. 4. The source of the thyroidectomy cells seen is uncertain, but it is suggested that they arise from acidophils whereas it is generally agreed that in the mammal, thyroidectomy cells come from basophilic thyrotrophs. 5. 5. Following radiological damage to the thyroid, thyroidectomy cells developed apparently at the same rate as after surgical thyroidectomy. 6. 6. Continuous immersion in solutions of either thiourea (0.25%) or potassium perchlorate (0.005%) brought about the involution of the thyroid gland and, consequently, the development of thyroidectomy cells within 5 months (thiourea) or 6 months (potassium perchlorate). 7. 7. Molting was inhibited by surgical or radiological thyroidectomy and by immersion in a solution of potassium perchlorate but not by immersion in a solution of thiourea. Thiourea apparently has a direct stimulatory effect on the skin since inhibition of molting was not seen in animals injected with thiourea until thyroidal involution occurred.

Hormonal control of the tail fin and of the nuptial pads in the male red-spotted newt

Abstract Seasonal variation in tail fin height and in display of nuptial pads has been followed in the red-spotted newt ( Notophthalmus viridescens viridescens ), and various experiments have been conducted to elucidate the hormonal control of those structures. The height of the tail is constant in the female, but in the autumn that of the male increases to maximal proportions and black, keratinized nuptial pads form on the hind legs. At the end of the spring breeding season these nuptial pads are lost and the tail fin is markedly reduced in height. A similar loss of pads and decline in tail height takes place whenever pad-bearing males are brought into the laboratory and kept at room temperature with long day lengths. Loss of tail height was apparently unaffected by thyroidectomy or by the administration of testosterone on alternate days, but its decline was slowed by autografting the pituitary gland and was halted by the administration of prolactin on alternate days. Tail height of laboratory-conditioned male newts was restored over a period of 2–3 wk by the administration of prolactin. In hypophysectomized, conditioned males the response of the tail fin to prolactin was greatly reduced but was restored to a considerable extent by the additional administration of thyroid hormone or thyroid stimulating hormone. In intact animals the loss of nuptial pads was delayed by autografting of the pituitary gland, by thyroidectomy, and by the administration of prolactin. It was unaffected by testosterone, but was prevented by prolactin and testosterone given in combination. The combined dose also induced restoration of pads after their regression in laboratory-conditioned males. In hypophysectomized, laboratory-conditioned males thyroxine or thyroid stimulating hormone was required in addition to prolactin and testosterone for the restoration of nuptial pads. Luteinizing hormone, but not progesterone, was an effective substitute for testosterone.

Maintenance of thyroid function in newts with transplanted pituitary glands

Following ablation of the hypophysis newts [ Triturus ( Diemictylus, Notophthalmus ) viridescens ] showed inhibition of molting, loss of appetite, and a high mortality rate. Animals with the pituitary gland transplanted to lie subcutaneously beneath the lower jaw, ate heartily and had a negligible mortality. Superficially they were indistinguishable from intact control newts except for a melanistic response indicating an elevated production of intermedin by the transplanted pars intermedia. From one group of eight animals that had been maintained in very good condition for 4 months after transplantation of the hypophysis, the gland was ablated from its transplanted position and within 3 weeks the animals gave the typical responses to hypophysectomy. Uptake and retention of 131 I by the thyroid was the same in newts with transplanted hypophyses as in control animals, but 131 I was lost more quickly from the thyroids of hypophysectomized animals. It appears, then, that in this form production of thyroid-stimulating hormone is either not controlled by the hypothalamus or that the hypothalamus discharges enough of a thyrotropic releasing factor to act through the general blood stream making the mediation of an hypothalamo-hypophyseal portal system unnecessary.

Hormonal regulation of cellular proliferation in the epidermis of the red-spotted newt

Abstract The effects of various hormonal conditions on epidermal proliferation were studied in the red-spotted newt. Prolactin was found to be a strong stimulant of epidermal mitotic activity. Hypophysectomy caused a rapid decline in the mean mitotic index, the thickness, and the number of cell layers in the epidermis. These parameters were increased by administration of ovine prolactin in both intact and hypophysectomized newts and by autografting of the pituitary gland. Injection of the prolactin inhibitor, ergocornine, into autografted newts decreased mitotic activity. Thyroid hormone increased the mean mitotic index of intact newts whereas thyroidectomy decreased the mitotic activity. The inhibitory effect of hypophysectomy was not prevented by exogenous thyroxine, indicating that thyroid hormone alone does not stimulate mitosis. Thyroid hormone apparently stimulates mitosis indirectly by increasing the rate at which sloughs are shed. Evidence for this view comes from the observation that the goitrogen thiourea, which mimics thyroidal effects on sloughing, also stimulates mitosis. Conversely, the accumulation of unshed sloughs may, in part, be responsible for the decrease in the mitotic activity in the thyroidectomized newts. Administration of adrenocorticotrophic hormone produced no detected effect on epidermal proliferation.

Hormonal effects on mitotic rhythm in the epidermis of the red-spotted newt.

Abstract The mitotic indices of epidermal cells in the red-spotted newt were measured at 3- or 4-hr intervals throughout 24-hr periods under a variety of hormonal conditions. In intact control newts maintained on a photoperiod of 16 hr of light/8 hr of darkness, a peak in mitotic activity was observed during the dark hours and a second smaller (but not statistically significant) peak was seen during the light hours. Inversion of the light cycle induced a new and more rapid rhythm. The basic rhythm was accentuated by autografting the pituitary gland to an ectopic site and also by the administration of thyroxine to intact animals. Hypophysectomy and administration of the prolactin-inhibiting drug, ergocornine, to auto-grafted animals greatly reduced the rate of epidermal proliferation and apparently reduced the peaks until they were no longer detectable. Injection of ovine prolactin into hypophysectomized or intact animals elevated the overall mitotic index and obscured or altered the mitotic rhythm. Removal of the rhyroid gland or administration of thiourea also altered the rhythm, but treatment with ACTH produced no detected effect. Apparently the basic rhythm of epidermal mitosis can be overridden or altered either by hormonal factors that increase or by ones that decrease the overall rate of cellular proliferation.

Effects of parathyroidectomy and of parathyroid extract on levels of calcium and phosphate in the blood and urine of the red-spotted newt☆

Abstract A single pair of parathyroid glands is typical of the red-spotted newt but an auxilary pair is sometimes present. The gland is composed of a single type of epithelial cell arranged in cords. The structure of the gland is stable throughout the year and shows no sexual dimorphism. Parathyroidectomy causes the calcium levels of both the blood and the urine to fall but control values are reestablished within 2 weeks. Administration of mammalian parathyroid extract (PTE) produces a rise in the concentration of calcium in the urine but no change in the calcium of the blood. Plasma phosphate rises after parathyroidectomy but the phosphate level of the urine remains unchanged. Within 24 hr of the injection of PTE a transitory peak in the phosphate appears in both blood and urine. It is suggested that in the red-spotted newt the parathyroid hormone mobilizes calcium from bone but, perhaps, may not act on the kidney.

Observations on the interaction of prolactin and thyroxine in the tail of the bullfrog tadpole.

Explants of tail fin from tadpoles of Rana catesbeiana maintained in tissue culture underwent resorption when exposed to a medium containing 150 nM thyroxine (T4). Increases in the specific activity of hexosaminidase (Hex) and of acid phosphatase (AP) were associated with this response. Ovine prolactin inhibited resorption of explants in a dose-responsive manner; however, prolactin had no influence on the specific activity of either Hex or AP cultured explants. This result contrasted with the effect of prolactin in vivo, where it inhibited both the resorption of the tail and the increase in specific activity of hydrolytic enzymes induced by T4. The actions of both hormones, when applied in vivo, were more pronounced in the tail fin than in the muscular core of the tail at the stage of regression tested, indicating that the failure of prolactin to influence enzyme induction in vitro did not result from an incapacity of the fin to respond to prolactin in this fashion. It is suggested that the inhibition of T4-induced enzymic activity by prolactin which is observed in the intact animal may come about in response to some related but indirect systemic action of prolactin that is lacking when explants are cultured in vitro. It is concluded that inhibition of the induction of hydrolytic enzymes is not an essential factor in the inhibitory effect of prolactin on the resorption of the tail fin.

Responses to thiourea and to surgical thyroidectomy by the autotransplanted pituitary gland in the red spotted newt

Abstract Two tests of thyrotropic function were conducted on autotransplanted pituitary glands in red spotted newts, Notophthalmus (Diemictylus, Triturus) viridescens viridescens , Rafinesque. In the first of these, the engrafted newts together with intact control animals were maintained in aqueous solutions of thiourea for 150 days. A marked goitrogenic response was seen in half the intact animals and in slightly more than a third of the animals with autotransplanted pituitary glands. In the second test, other newts with autotransplants and newts with intact pituitary glands were kept in thiourea and were also thyroidectomized surgically. Representative groups were killed at 15, 30 and 60 days after thyroidectomy. Thyroidectomy cells appeared in most of the intact and autotransplanted pituitary glands of newts killed at 30 and 60 days after thyroidectomy but not at 15 days. In similar tests mammalian glands have failed to support either goitrogenesis or the formation of thyroidectomy cells. It follows, then, that the transplanted pituitary gland of the newt is more active thyrotropically than those of the mammals thus far examined. Reasons for this difference are discussed.

The developmental cytology of the nuptial pad in the red-spotted newt☆

Abstract The ultrastructure of developing, mature and regressing nuptial pads has been examined and interpreted in the red-spotted newt. The development of the pad begins with a thickening of the dermis. Mitotic activity then increases the cell layers of the epidermis from about four to approximately eight. Simultaneously, keratinocytic synthetic activity shifts to produce more tonofilaments and fewer mucous granules. In the upper cell layers, the shift is followed by an increase in cytoplasmic volume with bundles of tonofilaments accumulating on the anterior side of each cell, displacing the nucleus posteriorly. After this rearrangement, the enlarged cells become grouped into ascending columns that tilt posteriorly from the basal epidermal layer at an angle of about 45°. Also the flattened cells of the monolayered stratum corneum become superficially roughened and with successive molts are replaced by orderly rows of cornified conical structures possessing cusps that are directed posteriorly. Each cone then lies at the top of one of the germinative columns. In rudimentary pads induced on female newts, the epidermis attains a height of only five or six layers and columns are not evident, but other developmental features of the male are present. During regression, mitosis is slowed and the developmental sequence is reversed.