Energetic response to repeated restraint stress in rapidly growing mice.

There is a cost of stress that may result in the loss of normal biological function (e.g., growth). Repeated, and even single, applications of stressors have been shown to induce negative energy balance in rodents. However, here we addressed whether this energetic response changes during multiple stress exposure and whether there is complete recovery subsequent to the cessation of stress exposure. These questions were addressed in growing C57Bl/6 mice (31 day) by determining at different times the energetic and endocrine responses after the exposure to restraint (R) stress for 4 h applied once (R1), repeatedly over 3 days (R3), or repeatedly over 7 days (R7). Compared with control values, R elevated (P<0.05) plasma corticosterone and reduced plasma insulin-like growth factor I on all days of exposure to the stressor. Seven days, but not 1 or 3 days of R, decreased the net growth (126%, P<0.05) and deposition of fat (71%, P<0.05) and lean (60%, P<0.05) energy over the 7 days. Only R7 depressed the 7-day metabolizable energy intake (P<0.05), and R7, but not R1 or R3, increased the overall energy expenditure (10%, P<0.05). Our results demonstrate that repeated episodes of stress are energetically costly to the rapidly growing animal, but compensatory mechanisms mitigate this cost of repeated stress exposure and permit complete recovery of energy balance after the cessation of stress application.

Summation of behavioral and immunological stress: metabolic consequences to the growing mouse.

To address the hypothesis that multiple stressors can have cumulative effects on the individual, we determined the effects of restraint (R) stress (4 h/day for 7 days), immunological (L) stress [lipopolysaccharide (LPS) injection, 0.45 microg/g body wt on days 6 and 7], and R + L (RL) on the growth and energetics of C57Bl/6 male mice. R and L each repeatedly increased (P<0.05) circulating corticosterone (>8 times), but RL caused even greater (>250%, P<0. 05) concentrations of circulating corticosterone than did either stressor alone. Only L and RL increased (P<0.05) circulating interleukin-1beta. Although R, L, and RL impaired growth (>75% below controls, P<0.05), RL reduced growth to a greater extent. All stressors inhibited (P<0.05) lean (>33% below controls) and fat (>120% below controls) energy deposition, and like the effects on growth, combined RL stress inhibited lean and fat energy deposition to a greater extent than did either stressor acting alone. These results demonstrated that the summation of multiple stress results in a cumulative cost to the growing animal.

Effects of acute behavioral stress and LPS-induced cytokine release on growth and energetics in mice.

We examined the biological cost of a single episode of acute restraint stress on the growth and energetics of mice, and compared these effects with the effects of stress associated with LPS injection and the total cost of experiencing both stressors simultaneously. We monitored growth and energetics over a 24-h period in 31-day-old mice exposed to either 4 h of restraint stress (R), i.p. injection of 0.45 mg/g of lipopolysaccharide (L), or restraint plus L injection (RL). Compared to controls, either restraint or L significantly (p < 0.05) impaired growth, feed intake, and lean and fat energy deposition. The R and L treatments depressed (>100% lower than controls) body weight change. However, when applied in concert, the effect of restraint and LPS on growth, energy deposition, and feed intake did not summate. All stressors significantly increased circulating corticosterone (p < 0.05) at 1 and 4 h following the initiation of treatment, and there was no difference between the R, L, or RL treatments. Only L and RL increased (p < 0.05) circulating IL-1b at 1 and 4 h. LPS injection elevated (p < 0.05) IL-1b. Although a single episode of behavioral stress altered growth and energy partitioning, LPS-induced cytokine release inhibited energy deposition and growth to a greater extent than behavioral stress (p < 0.05), and no further suppression of energy deposition or growth occurred when the two stressors were combined over 24 h.

The effect of chronic testosterone administration on sturgeon gonadotropins in juvenile and pre-vitellogenic white sturgeon (Acipenser transmontanus).

The effects of chronic exogenous testosterone treatment on the synthesis and/or secretion of two sturgeon gonadotropins (stGTH I and stGTH II) were assessed in 2-year-old juvenile white sturgeon (Acipenser transmontanus) surgically implanted with silastic capsules filled with 75 mg of testosterone and in previtellogenic female white sturgeon females implanted with 150 mg of testosterone. In groups of juvenile white sturgeon sacrificed 30, 60, 90, or 442 days postimplantation, pituitary concentrations of stGTH I were significantly greater in testosterone-treated fish (P < 0.01) when compared to those of controls. Pituitary concentrations of stGTH II were significantly higher (P < 0.01) in juvenile fish treated 60, 90, or 442 days with testosterone when compared to those of controls. Exogenous testosterone had no effect on plasma concentrations of either stGTH. Additional testosterone-treated juvenile sturgeon which were injected intraperitoneally 90 or 442 days postimplantation with 10 microg/kg of the gonadotropin releasing hormone analog d-Ala6-des-Gly10-GnRH ethylamide (GnRHa) also showed no change in plasma concentrations of stGTHs. Similar results were obtained for previtellogenic white sturgeon, as pituitary concentrations of stGTH I and stGTH II were significantly greater (P < 0.01) after 60 days of testosterone treatment compared to those of controls. A second group of 60-day testosterone-treated previtellogenic females also failed to exhibit increases of plasma stGTHs when administered 10 microg/kg of GnRHa. These results indicate that long-term testosterone treatment stimulates the accumulation of pituitary stGTHs in both juvenile and previtellogenic white sturgeon but does not affect basal or GnRHa-induced stGTH secretion.

Endocrine sensitivity to novelty in squirrel monkeys and titi monkeys: species differences in characteristic modes of responding to the environment.

The present study examined plasma cortisol and behavioral responses to environmental novelty in squirrel monkey and titi monkey male-female pairs. Overall, seemingly trivial increments in novelty evoked sustained plasma cortisol elevations. In individually tested animals, the minimal level of novelty sufficient to evoke a cortisol response was smaller, and the ability of the response to discriminate among levels of novelty was greater, in titis than in squirrel monkeys. When tested with the pairmate, the sensitivity of the response was reduced in titis but not in squirrel monkeys. Behavioral measures were not as sensitive to novelty as was the cortisol response. The results suggest that differential endocrine responsiveness to novelty is an important physiological concomitant to previously described differences between squirrel monkeys and titi monkeys in their characteristic modes of relating to the environment.

Social grouping tendencies and separation-induced distress in juvenile sheep and goats.

Social affinities in juvenile sheep and goats were compared by measuring grouping tendencies and separation-induced distress during experimental encounters with a person in either the presence or the absence of juvenile pen-mates. When tested with pen-mates, sheep spent more time near penmates than did goats. When separated from pen-mates, locomotor activity and plasma corticosteroid titers were higher in sheep, whereas vocal rates were higher in goats. Proximity to pen-mate scores were not correlated with either vocal rates, r = 0.20, or locomotion scores, r = 0.21, recorded when juveniles were tested alone. Proximity to pen-mate scores were correlated with posttest corticosteroid titers, r = 0.70; juvenile sheep spent more time near pen-mates and showed greater adrenocortical responses when temporarily separated from juvenile pen-mates. These findings support the possibility that interspecies differences in emotional reactivity contribute to the differences in grouping tendencies these ungulates display in natural or relatively unrestricted social groups.

Interaction between cortisol and arachidonic acid on the secretion of LH from ovine pituitary tissue.

In several species, glucocorticoids act directly on the pituitary gonadotroph to suppress the gonadotrophin-releasing hormone (GnRH)-induced secretion of the gonadotrophins, especially LH. A mechanism for this action of these adrenal steroids has not been established, but it appears that the glucocorticoids influence LH release by acting on one or more post-receptor sites. This study investigated whether glucocorticoids disrupt GnRH-induced LH release by altering the liberation of arachidonic acid from plasma membrane phospholipids, a component of GnRH-induced LH release. Using perifused ovine pituitary tissue, it was established that exposure of gonadotrophs to 1-1000 nmol cortisol/l for 4 h or longer significantly reduced GnRH-stimulated LH release with the maximal inhibitory effect being observed after 6 h of exposure to cortisol. This suppressive effect of cortisol could be reversed by administration of arachidonic acid, which in its own right could stimulate LH release from ovine pituitary tissue. Furthermore, the inhibitory effect of cortisol on GnRH-stimulated LH release could be directly correlated with decreased pituitary responsiveness to GnRH-stimulated arachidonic acid liberation, consistent with our hypothesis that glucocorticoids can suppress GnRH-induced secretion of LH by reducing the amount of arachidonic acid available for the exocytotic response of GnRH.

Disruption of estrous behavior in ewes by dexamethasone or management-related stress.

The effect of dexamethasone administration, isolation stress, or transportation stress on the ability of exogenous estradiol to induce estrus in progesterone-primed, ovariectomized ewes was evaluated in this study. Dexamethasone administered twice daily over a 6-d period, or a single injection at either 2 h before estradiol administration or 8 h after estradiol treatment, delayed or blocked the expression of estrous behavior. In those animals in which dexamethasone did not block the onset of estrus, the average length of estrus was reduced. Isolation stress, which induced significant increases in the plasma concentration of corticosteroids, failed to significantly alter the number of ewes expressing estrous behavior, although the expression of estrus was blocked in three of the nine treated animals. These nine animals all expressed estrus during the nonstress phase. In contrast to isolation stress, 8 h of transportation not only increased the average plasma concentration of corticosteroids, but also significantly (P less than .05) blocked the expression of estrus in five of eight ewes and delayed the expression of estrus in one other. During the nonstressed control phase, all eight expressed estrus. These findings indicate that management-related stress can block estrogen from inducing estrous behavior, however, the role of the adrenal axis response to stress in blocking estrus remains to be clarified.

How behavioral stress disrupts the endocrine control of reproduction in domestic animals.

Behavioral stress can prevent animals from achieving normal reproductive success. Stressors associated with intensive livestock management may be responsible for reduced reproductive efficiency. However, before appropriate management decisions can be made to alleviate the effects of behavioral stress on reproduction, it is necessary to identify the mechanisms by which stress disrupts normal reproduction. The neuroendocrine regulation of follicular development and ovulation requires a complex and delicate interplay between the pituitary gonadotropins and the feedback actions of the major follicular steroid, estradiol. Because of this complexity, the regulation of the follicular stage of the estrous cycle and ovulation is especially vulnerable to the effects of stress. Although the pathophysiological mechanisms by which stress disrupts reproduction are not fully understood, the stress-induced secretion of adrenal glucocorticoids seems to be of special significance because these steroids can effect both the synthesis and secretion of gonadotropins. Of additional importance may be the role of corticotropin-releasing hormone and adrenocorticotropin on the regulation of the hypothalamic-pituitary-gonadal axis.

Influence of the hypothalamic-pituitary-adrenal axis on the menstrual cycle and the pituitary responsiveness to estradiol in the female rhesus monkey (Macaca mulatta).

In order to examine the effect of glucocorticoids on the menstrual cycle of rhesus monkeys, cortisol was injected twice daily during the follicular phase. This cortisol treatment did not alter basal gonadotropin secretion but blocked the normal follicular rise of estrogens, the gonadotropin surge and the luteal rise of progesterone, and delayed the onset of the next cycle. In a second study, estradiol benzoate (E2B) was injected on the sixth day following the start of menstrual bleeding either with or without concurrent adrenocorticotropic hormone (ACTH) treatment. E2B injection was able to stimulate surges of luteinizing hormone (LH) and follicle-stimulating hormone (FSH) whether or not the animals had been treated with ACTH. These data suggest that, the action of cortisol, the final mediating step in the hypothalamic-pituitary-adrenal axis, occurs at the level of the gonads versus the pituitary in the rhesus monkey. While the pituitary response to endogenous gonadotropin-releasing hormone or exogenous E2B stimulation appears to remain unaffected, normal folliculogenesis is disrupted, preventing the follicular secretion of estrogens and the subsequent gonadotropin surges. The effects of corticosteroids are temporary, with normal cycling returning when plasma corticosteroids return to basal concentrations, albeit after a delay.

Interspecific contrasts in responses of macaques to transport cage training.

Corticosteroid values in response to brief confinement in a transport cage were compared between rhesus, bonnet, and crabeating macaques before and after they were trained to enter the cage. Behavioral data were collected to assess performance during training. Species differences were found both in training measures and in corticosteroid response to confinement in the transport cage after training. Bonnets took longer to train than rhesus or crabeaters. Rhesus showed the smallest adrenocortical response to cage confinement after training and crabeaters the greatest, suggesting that this group habituated more slowly to confinement than the other two groups. The results have implications for choice of experimental subject species and for management and husbandry of laboratory primates.

Social modulation of pituitary-adrenal responsiveness and individual differences in behavior of young domestic goats.

The effects of three variables known to influence pituitary-adrenal activity were examined in kid goats who were accompanied by adult goats in standard goat-human encounters. Serum corticosteroid concentrations in bold kids who characteristically spent above average amounts of time closely interacting with the person were never significantly different from control values. Corticosteroids in timid kids who spent little time near the person depended on the adult goat's behavior toward the person and the nature of the social relationship between the kid and the adult goat. When timid kids were accompanied by adult goats (familiar pen-mates or strangers) who behaved as if they were fearful of the person, their corticosteroids were significantly greater than control values. In contrast, bold pen-mates or bold strangers effectively reduced pituitary-adrenal responsiveness in timid kids. Only mothers unconditionally reduced kid pituitary-adrenal responsiveness and systematically affected kid behavior toward the person. The functioning of behavioral and pituitary-adrenal systems in young goats was influenced by consistent individual differences in responsiveness and the differing capacities of social companions to modulate this responsivity.

A model for assessing the impact of behavioral stress on domestic animals.

Animal scientists need a reliable measure of behavioral stress in domestic animals if they are going to be able to assess the stress of various management practices and to answer public concern about the well-being of animals used in agriculture. Popular measures of stress, alterations in behavior or changes in hormone secretion, are not adequate because of a failure to establish any direct correlation between changes in these characteristics with adverse effects on animal well-being. Further complicating the use of these indicators of stress is the variation in their pattern of response to different kinds of stressors. Even the same stressor can elicit divergent responses in different animals because of inter-animal variation in the stress response. To address these problems, a model of animal stress is discussed and tested. From this model it is proposed that the best indicator of an animal suffering from stress is the development of a pre-pathological state; i.e., a stress-related change in biological function that threatens the animal's well-being. Examples of such pre-pathological states would be a suppression of the immune system, the loss of reproductive events critical for normal reproduction, or the development of behaviors that would lead to such undesirable acts as tail-biting or excessive fighting. Although determining the existence of such pre-pathological states is not convenient, their existence is currently the only defensible indicator of an animal suffering from behavioral stress.

Effects of epinephrine, norepinephrine, and dopamine on gonadotropin-releasing hormone-induced secretion of luteinizing hormone in vitro.

Recent evidence indicates that catecholamines may directly alter anterior pituitary function. In the present study, an in vitro perifusion system was used to investigate whether catecholamines affect the gonadotrope. Pituitary tissue from castrated ram lambs was incubated in the presence of epinephrine (EPI), norepinephrine (NE), or dopamine (DA). During a 2-h treatment period, neither DA (10(-8) M), NE (10(-7) M), nor EPI (10(-7) - 10(-9) M) significantly affected basal LH secretion. In contrast, the LH response to a subsequent 10(-10) M GnRH challenge was significantly potentiated by NE and EPI. NE increased the amount of LH secreted in response to GnRH 14 +/- 1.1% (P less than 0.01). Likewise, 10(-7), 10(-8), and 10(-9) M EPI resulted in 22 +/- 1.4% (P less than 0.001), 13 +/- 1.2% (P less than 0.001), and 6 +/- 1.3% (P less than 0.03) increases, respectively. The stimulatory effect of 10(-7) M EPI was blocked by pretreatment with propranolol (a beta-adrenergic blocker), but not with phentolamine (an alpha-adrenergic blocker). The beta-adrenergic agonist isoproterenol enhanced GnRH-induced LH secretion 46 +/- 1.5% (P less than 0.001), but had no effect on basal LH release. DA had no effect on LH secretion; however, it inhibited PRL release 24 +/- 0.9% (P less than 0.001). Neither NE, EPI, nor isoproterenol had any effect on PRL secretion. These results suggest that EPI, acting by a beta 2-adrenergic receptor, modulates the pituitary gonadotrope's response to GnRH.

Adrenocorticotropin-induced changes in ovine pituitary gonadotropin secretion in vitro.

In vitro pituitary perifusion experiments were conducted to examine the effect of ACTH and related peptides on basal and GnRH-stimulated gonadotropin release. Treatments of 5 X 10(-7) M ACTH-(1-39), ACTH-(1-24), or ACTH-(18-39) were examined for their ability to influence basal gonadotropin secretion and the subsequent response to a 10(-9)- or 10(-8) M GnRH challenge. Administration of the 1-39 or 18-39 peptide sequences of ACTH similarly stimulated the release of LH and FSH (P less than 0.01). ACTH-(1-24) had no effect on basal gonadotropin secretion. Pretreatment with ACTH-(1-39) inhibited the LH and FSH responses to 10(-9) and 10(-8) M GnRH (P less than 0.05). Suppression of the LH response to 10(-8) M GnRH (P less than 0.05) and the FSH response to 10(-9) M GnRH (P less than 0.05) was observed after ACTH-(1-24) treatment. The administration of ACTH-(18-39) had no significant effect on GnRH-induced gonadotropin release. PRL concentrations were not affected by any of the ACTH peptides. Exposure to 10(-10) M GnRH or 5 X 10(-7) M synthetic ACTH-(1-39) produced an equivalent stimulation of LH secretion. GnRH pretreatment enhanced (P less than 0.05), while ACTH-(1-39) diminished (P less than 0.05), the subsequent response to GnRH. The GnRH receptor antagonist [D-pGlu1, D-Phe2, D-Trp3,6]GnRH attenuated the LH and FSH responses to GnRH and ACTH-(1-39) (P less than 0.05). The results obtained in this study indicate that certain portions of the ACTH molecule may affect gonadotropin secretion, perhaps by interacting with the GnRH receptor.

Inability to predict sexual and aggressive behaviors by plasma concentrations of testosterone and luteinizing hormone in Hereford bulls.

The relationships between sexual and aggressive behaviors and levels of plasma testosterone and luteinizing hormone (LH) were examined in Hereford bulls at 18 and 24 mo of age. Concentrations of hormones in bulls during periods of sexual rest (13 blood samples collected from each animal during a 24-h period) were compared with individual differences in sexual and aggressive behaviors when exposed to restrained females, either individually or in groups. Nearly all correlation coefficients were low and nonsignificant. It was concluded that individual differences in sexual performance and aggressive behavior cannot be predicted based on circulating levels of testosterone and LH in bulls during periods of sexual rest.