I.R. McDonald

Stress and mortality in a small marsupial (Antechinus stuartii, Macleay)

An investigation was made into the role of increased adrenocortical activity in the causation of the total postmating mortality of male Antechinus stuartii—a small shrew-like marsupial. A progressive rise in plasma androgen concentration of the males up to the time of mating was associated with increasingly aggressive behaviour and a progressive fall in plasma CBG concentration. During the mating period, the rate of fall in plasma CBG concentration increased markedly and plasma total corticosteroid concentration rose above the MCBC, so that there was a sharp rise in plasma free corticosteroid concentration just before the males all died. Although the plasma CBG concentration of females also fell during the mating period, with a moderate increase in plasma total glucocorticoid concentration, there was only a slight increase in plasma-free corticosteroid concentration at that time and it did not change significantly during the postmating, lactation period. Castration of captive males caused a marked increase in their plasma CBG concentration, and injection of testosterone or ACTH depressed it. There was a marked reduction in immunological competence and serum immunoglobulin concentration of the males just before their death. Postmortem examination revealed evidence of increased invasiveness of parasites and microorganisms, as well as severe haemorrhagic ulceration of the digestive tract. These effects could all be reproduced in captive animals by injection of cortisol acetate. It is concluded that a state of stress, induced by aggressive interactions at the time of mating and exacerbated by the fall in plasma CBG, results in suppression of the immune and inflammatory systems of the males, causing their death from gastrointestinal haemorrhage as well as invasion by parasites and microorganisms.

Endocrine changes in dasyurid marsupials with differing mortality patterns.

A comparison was made between the breeding-related endocrine changes in two species of dasyurid marsupial (Antechinus swainsonii and A. flavipes), which exhibit a catastrophic male mortality after a brief mating period; and those of another dasyurid species (Sminthopsis crassicaudata), the males of which survive an extended breeding period. In the males of both Antechinus species, the mating period was preceded by a rapid rinse in plasma androgen concentration and accompanied by a marked fall in plasma corticosteroid-binding globulin (CBG) concentration, to less than the total glucocorticoid concentration, which itself rose. As a consequence plasma free glucocorticoid concentration rose approximately 10-fold just before the males disappeared from the populations. Severe hemorrhagic ulceration of the upper digestive tract was consistently found soon after capture near the time of mating. In females, CBG concentration was always well in excess of total glucocorticoid concentration and free glucocorticoid concentration remained low. There was a significant negative correlation between plasma androgen and CBG concentrations in male Antechinus species from the natural populations. In laboratory-held castrate male A. swainsonii, injection of either testosterone or dihydrotestosterone caused a dose-related fall in CBG. In contrast to the above, the CBG concentration was always well in excess of total glucocorticoid concentration in the plasma of male S. crassicaudata and there was no evidence of interaction between plasma androgen and CBG concentration. These findings support the hypothesis that, in those species with a breeding-related mortality of males, an androgen-dependent fall in plasma CBG concentration and a stressful mating period result in death from adrenocortical suppression of the immune and inflammatory responses.

Concentration of Free Glucocorticoids in Plasma and Mortality in the Australian Bush Rat (Rattus fuscipes Waterhouse)

We investigated the adrenocortical response to the stress of capture and anesthesia, and its association with seasonal mortality patterns in the Australian bush rat ( Rattus fuscipes ). In females, there were two marked seasonal peaks in the concentration of total corticosterone B of stressed plasma, one in the summer breeding period and the other in winter, a time of high mortality in females. The increase in summer was matched by a greater increase in maximum corticosterone-binding capacity of plasma so that free, biologically active, corticosterone B in plasma remained low. At the time of winter mortality, total corticosterone B exceeded maximum corticosterone-binding capacity and free corticosterone B in plasma was much higher than at other times. In males, total corticosterone B in plasma was much lower than in females during the summer period when mortality of males was high but maximum corticosterone-binding capacity was even lower, so that free corticosterone B in plasma was much higher than at other times. A similar increase in some males occurred at the time of winter mortality. The association of higher free corticosterone B in plasma and high mortality suggest an excessive adrenocortical response to stress causing increased susceptibility to disease, as in some dasyurid marsupials.

High affinity binding of adrenocortical and gonadal steroids by plasma proteins of Australian marsupials

Abstract High affinity binding of cortisol, progesterone, testosterone, and estradiol by plasma proteins was examined in 29 marsupial species by equilibrium dialysis and/or polyacrylamide gel electrophoresis. The association constant ( K a , 36°) and binding capacity ([Σ ϱ ]) for cortisol and testosterone were measured by microdialysis and found to be 3.2 to 9.6 × 10 7 M −1 and 0.3 to 3.7 × 10 −7 M respectively, for cortisol and 1.6 to 8.3 × 10 8 M −1 and 0.3 – 4.0 × 10 −7 M for testosterone. High-affinity binding of cortisol was detected in all 25 species studied by this method but testosterone was bound in 9 only. Steroid-binding proteins were further characterized by polyacrylamide gel electrophoresis of plasma equilibrated with radioactive steroid at 4°. Two types of non-albumin binding proteins were identified, one with a mobility like human corticosteroid-binding globulin (CBG) and an affinity for both cortisol and progesterone and a second protein with a mobility like human sex hormone-binding globulin (SHBG) and an affinity for testosterone. High affinity binding of estradiol was not detected, and a second CBG was found in four species.

Corticosteroid secretion in the Australian phalanger (Trichosurus vulpecula)

Abstract Corticosterone (B) as well as cortisol (F) is secreted by the adrenal gland of the marsupial Trichosurus vulpecula . The F:B ratio in controls varied from 3.5 to 12.8; and in all animals (including ACTH-treated) from 1.6–16.0. Both steroids were present in the erythrocytes as well as the plasma of adrenal venous blood. Their secretion was stimulated by intravenous ovine corticotrophin; and maximum cortisol secretion rate was 20.8–22.3 μg/hour per 100 mg adrenal, or 10–25 μg/hour per kg body weight. Cortisol concentration in peripheral blood plasma did not exceed 2.5 μg/100 ml. Aldosterone was sought, but not detected in excess of 0.1 μg in adrenal venous plasma, corresponding to a 2-hour collection period; or in a 48-hour urine sample. Urinary excretion of free cortisol, cortisone, their tetrahydro derivatives, and 17-ketosteroids was demonstrated; but their excretion rates were all very low, compared with eutherians.

Oestrogen receptors in the genital tract of the Australian marsupial Trichosurus vulpecula

Abstract Macromolecules with high binding affinity for oestrogens ( K a = 10 9 –10 10 M −1 at 0°) are detected in cytosols of vaginal and uterine endothelium of the marsupial brush-tailed possum Trichosurus vulpecula . There are two distinct classes, of high (8 S) and low (4 S) molecular weight, the proportions of which vary in relation to the reproductive state or hormone treatment. In the endometrium, the 8-S form predominates during all phases of the breeding cycle; but there is a relative increase in the 4-S form, up to 40% of the total, during lactation. In the vaginal endothelium, an 8-S form is predominant in cytosols from follicular and lactating animals, with little or no 4-S binding, but the reverse occurs in luteal or pregnant animals. Only the 4-S form is found in cytosols of endometrium or vaginal endothelium of 16-day castrate females. After treatment of castrates with oestradiol, testosterone, or progesterone, however, only the 8-S form is detected in the endometrium. In the vaginal endothelium, the high-affinity binding remains in the 4-S region in castrates treated with oestradiol or testosterone, but is in the 8-S form after treatment with progesterone. The concentration of high-affinity binding sites per milligram of protein is significantly increased in both endometrial and vaginal endothelial cytosols during the follicular phase of the cycle and in castrates treated with oestradiol. This increase is associated with a marked increase in size of the two organs. Treatment with progesterone or testosterone has no effect on the concentration of binding sites in castrates. These findings are discussed in relation to the unique method of reproduction in marsupials.

The effect of cortisol on the metabolic actions of insulin in the echidna, Tachyglossus aculeatus (Shaw)

Abstract The effects of insulin on plasma concentrations of glucose, free fatty acids (FFA), amino acids and urea were investigated in saline- and cortisol-treated echidnas ( Tachyglossus aculeatus ). Intravenous injection of 0.02 or 0.2 IU of regular insulin/kg in saline-injected echidnas was followed by dose-related decreases in plasma glucose, amino acid and urea concentrations. However, following a rapid decrease to 52 ± 10% of the control value, plasma FFA then increased slowly to a maximum of 254±35%, 8 hr after the injection of 0.2 IU of insulin/kg. An increase in plasma corticosteroid concentration was not observed. Pretreatment with five daily im injections of 0.2 mg of cortisol acetate/kg did not affect the early hypoglycemic and lipogenic actions of insulin but decreased its effect on plasma amino acid and urea concentrations. The later increase in plasma FFA concentration was abolished. It is concluded that insulin has potent actions on carbohydrate, protein, and fat metabolism in the echidna and that antagonistic effects of the glucocorticoids on these actions are minimal.