Disrupted seasonal biology impacts health, food security and ecosystems.

The rhythm of life on earth is shaped by seasonal changes in the environment. Plants and animals show profound annual cycles in physiology, health, morphology, behaviour and demography in response to environmental cues. Seasonal biology impacts ecosystems and agriculture, with consequences for humans and biodiversity. Human populations show robust annual rhythms in health and well-being, and the birth month can have lasting effects that persist throughout life. This review emphasizes the need for a better understanding of seasonal biology against the backdrop of its rapidly progressing disruption through climate change, human lifestyles and other anthropogenic impact. Climate change is modifying annual rhythms to which numerous organisms have adapted, with potential consequences for industries relating to health, ecosystems and food security. Disconcertingly, human lifestyles under artificial conditions of eternal summer provide the most extreme example for disconnect from natural seasons, making humans vulnerable to increased morbidity and mortality. In this review, we introduce scenarios of seasonal disruption, highlight key aspects of seasonal biology and summarize from biomedical, anthropological, veterinary, agricultural and environmental perspectives the recent evidence for seasonal desynchronization between environmental factors and internal rhythms. Because annual rhythms are pervasive across biological systems, they provide a common framework for trans-disciplinary research.

Sex differences in emotionality in C3H/HeH mice, with hypogonadal mutant to distinguish activational effects of gonadal hormones.

The C3H/HeH mouse strain has a mutant hypogonadal (hpg) variant, providing an animal model to examine the activational effects of sex hormones because reproductive maturation is arrested at a neonatal stage. Thus in the adult mouse, the circulating concentrations of sex steroids are extremely low. The present study used a series of tests to distinguish sex differences in behaviour: open field, locomotor activity, hyponeophagia, and novel location recognition. The results showed some evidence for a role of sex hormones in emotionality underscoring the potential utility of the hpg model, to distinguish activational effects in the C3H/HeH strain. However, the direction that the sex differences took varied by task: whilst males showed the predicted sex difference of relatively greater anxiety in the open field, hyponeophagia tests suggested higher emotionality in females. The hpg mice of both sexes showed a reduction in anxiety measured as hyponeophagia. Overall it can be concluded that this set of experiments supports the potential of the hpg model to investigate hormonal influences on emotionality.

Human 2D (index) and 4D (ring) finger lengths and ratios: cross-sectional data on linear growth patterns, sexual dimorphism and lateral asymmetry from 4 to 60 years of age.

Human 2D : 4D ratios (measures of the relative lengths of index and ring fingers) attract considerable research interest because they exhibit sexual dimorphism and are associated with various morphological, physiological and behavioural traits as well as sporting abilities and medical conditions. In an attempt to identify potential confounding factors in such studies, we have examined how relative and absolute digit lengths vary with gender and tested whether they are influenced by age, right-left asymmetry and hand preference. Participants between 4 and 60 years of age were recruited from local educational sites. Hand photocopies and calliper measurement were used to obtain digit lengths. We employed linear regression analysis to examine the growth trajectories of individual digits, analyses of variance to isolate main and interaction effects of age, gender and hand preference, and paired t-tests to identify lateral asymmetries. Both digits exhibited biphasic growth with an early growth phase followed by a stable length phase. Digits in females attained their maximum length about 2.2 years (dextral subjects) or 5.1 years (sinistral subjects) earlier than those in males. Sexual dimorphism in 2D : 4D ratios was apparent by 4 years of age and age changes in ratios depended on gender, side and hand preference. Relative and absolute lengths displayed age, gender, hand-preference and age x gender interaction effects. Lengths tended to be greater in females in younger subjects and greater in males in older subjects. Ratios tended to be greater in sinistral subjects. In dextral subjects, significant lateral asymmetries in 2D lengths were seen at all ages but asymmetries in males and 4D lengths seemed to be age-dependent. We conclude that age, lateral asymmetry and hand preference are potential confounding factors and that future study designs should take account of these as well as other known confounders such as ethnicity, birth order, menstrual cycle phase and sexual preference.

Thyrotrophin-releasing hormone decreases feeding and increases body temperature, activity and oxygen consumption in Siberian hamsters.

Thyrotrophin-releasing hormone (TRH) is known to play an important role in the control of food intake and energy metabolism in addition to its actions on the pituitary-thyroid axis. We have previously shown that central administration of TRH decreases food intake in Siberian hamsters. This species is being increasingly used as a physiological rodent model in which to understand hypothalamic control of long-term changes in energy balance because it accumulates fat reserves in long summer photoperiods, and decreases food intake and body weight when exposed to short winter photoperiods. The objectives of our study in Siberian hamsters were: (i) to investigate whether peripheral administration of TRH would mimic the effects of central administration of TRH on food intake and whether these effects would differ dependent upon the ambient photoperiod; (ii) to determine whether TRH would have an effect on energy expenditure; and (iii) to investigate the potential sites of action of TRH. Both peripheral (5-50 mg/kg body weight; i.p.) and central (0.5 microg/ml; i.c.v.) administration of TRH decreased food intake, and increased locomotor activity, body temperature and oxygen consumption in the Siberian hamster, with a rapid onset and short duration of action. Systemic treatment with TRH was equally effective in suppressing feeding regardless of ambient photoperiod. The acute effects of TRH are likely to be centrally mediated and independent of its role in the control of the production of thyroid hormones. We conclude that TRH functions to promote a catabolic energetic state by co-ordinating acute central and chronic peripheral (thyroid-mediated) function.

Hypothalamic expression of human growth hormone induces post-pubertal hypergonadotrophism in male transgenic growth retarded rats.

Growth hormone (GH) is known to regulate peripheral components of the hypothalamo-pituitary gonadal (HPG) axis, but it remains unclear whether GH exerts a significant influence on the activity of the hypothalamo-pituitary components of the HPG axis. In this study, we investigated the development of HPG axis function in the male transgenic growth retarded (Tgr) rat, a model of moderate systemic GH deficiency caused by hypothalamic expression of human (h)GH. Impaired postnatal somatotroph expansion and moderate GH deficiency in male Tgr rats were accompanied by a two- to three-fold increase in pituitary gonadotrophin content, but without a significant change in the pituitary gonadotroph population. A three- to nine-fold elevation in basal circulating luteinising hormone concentration was seen in postpubertal Tgr rats, with a smaller increase in follicle-stimulating hormone. Despite this hypergonadotrophism, there was no corresponding increase in steroidogenic (circulating testosterone and seminal vesicle weights) or gametogenic (spermatozoa counts in seminiferous tubules) activity in the postpubertal Tgr testis. Following puberty, the plasma leptin concentration also became progressively elevated in Tgr males. Circulating gonadotrophin and leptin levels were normalised in Tgr rats by peripheral physiological replacement of rat GH, but plasma testosterone concentration was unaffected. These results confirm that hGH exerts a positive influence on the central control of gonadotrophin secretion in the Tgr rat, but the absence of a corresponding elevation in the steroidogenic or gametogenic function of the Tgr testis implies that the peripheral GH/insulin-like growth factor I axis may also exert a permissive influence on testicular function. The relative contribution of somatogenic and lactogenic mechanisms and the potential influence of elevated leptin and decreased sensitivity to androgen feedback to the development of postpubertal hypergonadotrophism in Tgr males remain to be determined.

The role of glutamate in the photic regulation of the suprachiasmatic nucleus.

Endogenous circadian rhythms govern most aspects of physiology and behaviour in mammals, including body temperature, autonomic and endocrine function, and sleep-wake cycles. Such rhythms are generated by the suprachiasmatic nucleus of the hypothalamus (SCN), but are synchronised to the environmental light-dark cycle by photic cues perceived by the retina and conveyed to the SCN via the retinohypothalamic tract (RHT). This review considers many lines of evidence from diverse experimental approaches indicating that the RHT employs glutamate (or a related excitatory amino acid) as a neurotransmitter. Ultrastructural studies demonstrate the presence of glutamate in presynaptic terminals within the SCN. In situ hybridisation and immunocytochemical studies reveal the presence of several NMDA (NMDAR1, NMDAR2C), non-NMDA (GluR1, GluR2, GluR4) and metabotropic (mGluR1) glutamate receptor subunits in the SCN. Messenger RNA encoding a glutamate transporter protein is also present. In behavioural tests, glutamate antagonists can block the effects of light in phase-shifting circadian rhythms. Such treatments also block the induction of c-fos within SCN cells by light, whereas a glutamate agonist (NMDA) induces c-fos expression. In hypothalamic slice preparations in vitro, electrical stimulation of the optic nerves induces release of glutamate and aspartate, and glutamate antagonists block field potentials in the SCN evoked by stimulation of the optic nerve. Circadian rhythms of electrical activity which persist in vitro are phase shifted by application of glutamate in a manner which mimics the phase shifting effects of light in vivo. This wide range of experimental findings provides strong support for the hypothesis that glutamate is the principal neurotransmitter within the RHT, and thus conveys photic cues to the circadian timing system in the SCN.

Involvement of 5-HT receptors in the regulation of food intake in Siberian hamsters.

The Siberian hamster provides a physiological model for understanding the hypothalamic control of energy metabolism as it undergoes annual photoperiod-regulated cycles of body weight (i.e. fattening in summer, and catabolism of fat stores in winter). As a first step to investigate whether enhanced serotonergic (5-HT) tone might underlie the catabolic processes in short days, we investigated whether serotonergic stimulation can produce catabolic actions in fat hamsters housed in long days. Acute treatment with the serotonin reuptake inhibitor (+/-) fenfluramine (8 mg/kg, i.p.) produced a prolonged, dose-dependent reduction in food intake in both photoperiods. Behavioural observations and radiotelemetry analyses revealed that this anorectic effect of fenfluramine was associated with short-term increases in locomotor activity and in core body temperature. In a subsequent series of studies, hamsters were pretreated with the 5-HT2C receptor antagonist SB242084 (4 mg/kg, i.p.). This 5-HT2C receptor antagonist completely blocked the anorectic actions of fenfluramine, but did not decrease the hyperthermia or hyperlocomotion induced by fenfluramine; thus, the anorectic actions of fenfluramine probably reflect actions via the 5-HT2C receptor. Consistent with these observations, treatment of hamsters with the 5-HT2C receptor agonist VER 3323 (10 mg/kg, i.p.) or the 5-HT1B/2C receptor agonist mCPP (3 mg/kg, i.p.) reduced food intake. The response to manipulation of serotonergic pathways was not affected by the ambient photoperiod in any of these studies. We conclude that the anorectic actions of fenfluramine are not an indirect consequence of serotonergic actions on arousal pathways, and that its actions on feeding in the Siberian hamster are most likely to be mediated by the 5-HT2C receptor.

Maternal entrainment of the developing circadian system in the Siberian hamster (Phodopus sungorus).

The aim of these studies was to investigate maternal entrainment of developing circadian locomotor activity rhythms in the Siberian hamster. In Experiment 1, mothers were transferred from a 16:8 LD cycle into constant dim red light (DD) from the day of parturition, and wheel-running activity of the mother and pups was individually monitored from the time of weaning. The phases of the individual pups' rhythms were found to be synchronized both to the phase of the mother and to the phase of lights off (ZT 12) of the photo cycle that the mother was exposed to until the day of parturition. To investigate whether this synchrony might reflect direct effects of light acting upon the fetal circadian system in late gestation, the experiment was repeated but with mothers placed into DD early in pregnancy (< or = day 7 of gestation). The results were similar to the first study, suggesting that the mother rather than the photo cycle during the latter part of gestation entrains the developing circadian system. The third experiment investigated whether this entrainment occurred during the postnatal period. Breeding pairs were maintained on alternative light-dark cycles, LD and DL, that were 12 h out of phase. Litters born to mothers on one light-dark cycle were exchanged on the day of birth with foster mothers from the reversed light-dark cycle, then raised in DD. Control litters exchanged between mothers from the same light-dark cycle had similar litter synchrony as shown by nonfostered litters of Experiment 1. However, pups cross-fostered with mothers on reversed LD cycles showed a very different distribution of pup phases. Pups were not synchronized to their natural mother but to their foster mother. Moreover, pups were more scattered over the 24-h period and were found to be significantly synchronized to the phase of the reversed LD cycle. These results demonstrate the occurrence of postnatal entrainment in the Siberian hamster. The increased scatter produced by the cross-fostering paradigm results from some litters being completely entrained to the phase of the foster mother, some with an intermediate distribution between the phase of the natural and foster mothers, and a minority being associated with the phase of the natural mother. These results suggest that Siberian hamster pups are initially synchronized either prenatally or at birth but that the mother continues to provide entrainment signals during the postnatal period.

Effects of constant darkness and constant light on circadian organization and reproductive responses in the ram.

The relationship between circadian rhythms in the blood plasma concentrations of melatonin and rhythms in locomotor activity was studied in adult male sheep (Soay rams) exposed to 16-week periods of short days (8 hr of light and 16 hr of darkness; LD 8:16) or long days (LD 16:8) followed by 16-week periods of constant darkness (dim red light; DD) or constant light (LL). Under both LD 8:16 and LD 16:8, there was a clearly defined 24-hr rhythm in plasma concentrations of melatonin, with high levels throughout the dark phase. Periodogram analysis revealed a 24-hr rhythm in locomotor activity under LD 8:16 and LD 16:8. The main bouts of activity occurred during the light phase. A change from LD 8:16 to LD 16:8 resulted in a decrease in the duration of elevated melatonin secretion (melatonin peak) and an increase in the duration of activity corresponding to the changes in the ratio of light to darkness. In all rams, a significant circadian rhythm of activity persisted over the first 2 weeks following transfer from an entraining photoperiod to DD, with a mean period of 23.77 hr. However, the activity rhythms subsequently became disorganized, as did the 24-hr melatonin rhythms. The introduction of a 1-hr light pulse every 24 hr (LD 1:23) for 2 weeks after 8 weeks under DD reinduced a rhythm in both melatonin secretion and activity: the end of the 1-hr light period acted as the dusk signal, producing a normal temporal association of the two rhythms. Under LL, the 24-hr melatonin rhythms were disrupted, though several rams still showed periods of elevated melatonin secretion. Significant activity rhythms were either absent or a weak component occurred with a period of 24 hr. The introduction of a 1-hr dark period every 24 hr for 2 weeks after 8 weeks under LL (LD 23:1) failed to induce or entrain rhythms in either of the parameters. The occurrence of 24-hr activity rhythm in some rams under LL may indicate nonphotoperiodic entrainment signals in our experimental facility. Reproductive responses to the changes in photoperiod were also monitored. After pretreatment with LD 8:16, the rams were sexually active; exposure to LD 16:8, DD, or LL resulted in a decline in all measures of reproductive function. The decline was slower under DD than LD 16:8 or LL.(ABSTRACT TRUNCATED AT 400 WORDS)

Circadian and photoperiodic time measurement in male Syrian hamsters following lesions of the melatonin-binding sites of the paraventricular thalamus.

Autoradiographic studies using [125I]iodomelatonin in several species, including the Syrian hamster, have revealed that the rostral region of the anterior paraventricular nucleus of the thalamus (aPVT) contains a very high density of binding sites for melatonin. In two studies, small or large bilateral electrolytic lesions of the aPVT were made in adult male hamsters maintained on long days (LD 16:8). The hamsters were then transferred to short days (LD 8:16) to test whether testicular regression could occur in response to a decrease in photoperiod. Serum prolactin concentrations were measured as a second photoperiodic response. All unoperated control hamsters showed the typical short-day photoperiodic response: A decrease in serum luteinizing hormone (LH) and prolactin concentrations and testicular regression all occurred within 6 weeks in short days, followed by the development of scotorefractoriness. Lesions of the aPVT did not significantly affect the rate or the degree of the short-day-induced decline in serum levels of LH or prolactin, nor the pattern of testicular regression and the subsequent expression of refractoriness. To enable us to determine whether the aPVT might be involved in the entrainment or the expression of circadian rhythms, locomotor activity was monitored continuously in lesioned and control groups in Experiment 2, prior to and following the switch to short days. The reduction in photoperiod (involving an 8-hr advance in the time of lights-off and an 8-hr extension of the dark phase) caused a decompression of the nocturnal activity bout of control animals, so that after 2 weeks in short days, activity onset had also advanced to regain its phase relationship to the timing of lights-off. A similar pattern of reentrainment was observed in lesioned animals, and no differences were observed between treatment groups in the rate of entrainment and decompression. In addition, both intact controls and animals bearing large bilateral lesions of the aPVT exhibited robust free-running circadian rhythms of locomotor activity when held under constant dim red light. In summary, the integrity of the aPVT is not necessary for the seasonal response of the reproductive axis and prolactin secretion to photoperiod, nor for photic entrainment of activity rhythms, in the Syrian hamster.

Entrainment of the melatonin rhythms in early postnatal lambs and their mothers.

Although the developing sheep can produce an appropriately timed melatonin rhythm as early as 1 week after birth, it is not known whether the lamb is able to adjust its melatonin rhythm to a change in daylength. The ability of the young lamb to entrain its pattern of melatonin secretion to a new photoperiod was determined in the present study. Eight female lambs and their mothers were raised in long days (LD 16:8) beginning 2 weeks postpartum. At 7 weeks of age, the time of lights-off was advanced 8 hr, the short-day photoperiod then being LD 8:16; the time of lights-on remained unchanged. Concentrations of melatonin were measured in blood samples collected hourly on days - 1, 0, 2, 4, 6, and 13 relative to the light change. On day 0, all mothers and daughters had advanced the onset of melatonin secretion by at least 1 hr, and by day 13, 12 of 16 had completely entrained to the new photoperiod. The rate of entrainment among individuals varied; the mean rate for lambs and mothers did not differ. This study provides evidence that the melatonin-rhythm-generating system matures shortly after birth.

Hair.

The psychologic importance of hair to man is in inverse ratio to its physical function. Except for scalp hair and desultory areas of sexual hair, most of man's hair follicles are vestigial. Three problems of hair growth remain to be solved: (1) how the intermittent activity of hair follicles in both animals and man is controlled; (2) how the male hormone alters the hair cycle in human skin; and (3) why larger hairs are produced by testosterone in some areas of the body when in some individuals the hair follicles in the scalp regress. Studies in which skin grafts from rats of different ages were exchanged showed that hair follicles are innately programmed but can be slowly influenced by systemic factors. Steroid hormones, especially estrogens, slow down the moult cycle whereas thyroid hormones accelerate it. What establishes the innate rhythm remains problematical. The fact that plucking out the club hair initiates activity in resting follicles has been explained by the hypothesis that the mitotic inhibitor which accumulates during anagen is normally used up or dispersed during telogen or by wounding. However, contrary to this theory, follicular activity is not prolonged by epilation during anagen. Moreover, if rats are epilated within one or two days of eruption, only club hairs are removed since forceps cannot grasp the tips of the new hairs. Such epilation does not affect the anagen in progress, but remarkedly enough the subsequent resting phase is shortened. Both sexual hair and male-pattern baldness depend on androgenic hormones. Target organs of testosterone convert the hormone to active metabolites, chiefly 5alpha-dihydrotestosterone. In skin, however, 5alpha-dihydrotestosterone may not be the only active tissue androgen. The major metabolite of testosterone incubated with hair roots in androstenedione, and hirsute women without other obvious endocrine abnormality sometimes excrete high levels of androstanediol. Both steroids stimulated the sebaceous glands of hypophysectomized-castrated rats, which, however, showed only a limited response to testosterone. The androgenic steroids, the enzymes that convert them to their active metabolites, and the proteins that bind them are undoubtedly very important to the problems of the growth of sexual hair and male-pattern baldness.

19-Aldehydo-4-androstene-3,17-dione: an estrogen precursor that inhibits sebaceous secretion in a rat model.

19-aldehydo-4-androstene-3,17-dione (A; R = CHO), a biogenetic precursor of estrone in the body, has been found to suppress sebum secretion in the ovariectomized testosterone-treated Wistar rat at 1/2000 times the dose of cyproterone acetate required to produce an equivalent effect. The action of this steroid must therefore be analogous to that of an estrogen even though, in striking contrast to estradiol, it is without effect on uterine weight or vaginal cornification. It is postulated that 19-aldehydo-4-androstene-3,17-dione (A; R = CHO) is converted locally into estrone (B) by aromatase present in skin but only in low concentrations in vagina and uterus. The potential of biogenetic precursors (I) of estrogens for therapy of acne or alleviation of the worst effects of skin aging is worth investigation.

The local effects of topically applied estradiol, cyproterone acetate, and ethanol on sebaceous secretion in intact male rats.

Estradiol in ethanol was applied once daily to one flank of intact male rats and sebum production on both flanks was measured over periods of 18 h alternating with 6 h by absorbing the lipid on pads of cigarette paper held in place by a harness, starting on the 15th day. Sebum production was very significantly less on the treated flanks than on the contralateral flanks that received only vehicle, indicating an unequivocal local effect of the estradiol. At the same time, the values on the contralateral flanks were significantly below those of littermate rats which received vehicle only, indicating that the estrogen had been systemically absorbed to produce a distal action. The estradiol also significantly reduced plasma testosterone and the relative weights of the seminal vesicles, ventral prostate, and preputial glands, which demonstrated that part of the general action of the estrogen could have been by suppression of endogenous androgen production. In short-term experiments, in which measurements were started concurrently with treatment, the inhibitory action of estradiol, in contrast to that of cyproterone acetate, was not detected until the 4th day. Indeed, it appeared probable that the estrogen actually stimulated sebum secretion over the first 2 days, suggesting that it had a biphasic effect.

Local suppression of sebum secretion in rats by topical cyproterone acetate in ethanol.

Sebum production was measured on 2 symmetrically placed areas on the flanks of rats over alternating periods of 18 and 6 hr for 4 days, by absorbing the lipid on pads of cigarette paper held in place by a specially designed harness. Castrated rats receiving testosterone produced about twice as much sebum as untreated littermate controls. Once daily application of ethanol to one flank significantly reduced sebum production, in comparison with the other flank in testosterone-treated but not in untreated rats. Daily application of 5 mg cyproterone acetate in ethanol to one flank in a third group of rats significantly reduced sebum production in comparison with that of the other flank treated with vehicle only. The effects became significant within 24 hr. It seems likely that the sebaceous glands received the topically applied materials by way of the pilo-sebaceous orifices rather than by transepidermal absorption. The results demonstrate that one action of cyproterone acetate is at the target site and suggest that the anti-androgen may be effective when applied topically. The method could prove useful in assessing the local action on sebaceous activity of topically applied substances.

Beta-endorphin secretion in rams related to season and photoperiod.

A newly established RIA was used to measure changes in the concentration of beta-endorphin in peripheral blood and pituitary tissue from adult Soay rams living outside under natural conditions and housed indoors under artificial photoperiods. A pronounced seasonal cycle in plasma beta-endorphin immunoreactivity occurred in the outdoor animals, with low levels in spring and early summer (February-May; less than 200 pg/ml plasma) and maximal levels 10-20 times higher in late summer and autumn (July-October). Seasonal changes in plasma levels of PRL, FSH, and cortisol, testis size, and body weight were also monitored; the seasonal cycle in the levels of immunoreactive beta-endorphin occurred in parallel with the cycle in plasma FSH and body weight. There were no significant seasonal changes in plasma cortisol concentrations. Marked changes in the plasma levels of beta-endorphin were also seen in rams kept under the artificial photoperiod regimen of alternating 12- to 16-week periods of long days (16 h of light and 8 h of darkness; 16L:8D) and short days (8L:16D). Transfer from long days to short days led to a greater than 20-fold increase in the levels of beta-endorphin, reaching a maximum after 4-8 weeks; the reverse switch in photoperiod led to a rapid decrease in the levels. There was no diurnal rhythm in the plasma levels of beta-endorphin based on hourly samples collected for 24 h under long and short days. The total content of immunoreactive beta-endorphin in the pituitary gland was lower in rams under short days than under long days, converse to the pattern in the blood. Sephadex chromatography of the plasma samples revealed that most of the beta-endorphin immunoreactivity coeluted with synthetic beta-endorphin-(1-31), and a small amount of activity eluted with beta-lipotropin. The seasonal and photoperiod-induced changes were largely due to changes in the levels of beta-endorphin. Extracts of pituitary tissue revealed a large proportion of beta-lipotropin to beta-endorphin compared to plasma, with no consistent change in ratio related to the photoperiod. The overall results illustrate that there are pronounced seasonal and photoperiod-induced changes in immunoreactive plasma beta-endorphin levels in the ram. Under artificial photoperiods, long days inhibit and short days stimulate beta-endorphin secretion. Under natural conditions, the development of refractoriness to both the inhibitory effects of long days and the stimulatory effects of short days may explain the timing of the annual cycle of beta-endorphin secretion.

Endogenous opioid regulation of pulsatile luteinizing hormone secretion during sexual maturation in the female sheep.

The developing female sheep, which attains puberty after 25 weeks of age, was used as an experimental model to investigate the role of endogenous opioid peptides in the control of pulsatile LH secretion during sexual maturation. Treatment of ovary-intact prepubertal sheep at 12 weeks of age with the opiate antagonist naloxone resulted in a dose-dependent increase in LH secretion. Subsequent studies used ovariectomized (OVX) lambs implanted with capsules containing 17 beta-estradiol to provide a constant, ovarian steroid feedback signal throughout development. Naloxone treatment (hourly iv injections of 1 mg/kg BW for 4 h) produced an increase in the frequency of episodic LH secretion at all prepubertal ages, when lambs were highly sensitive to the estradiol negative feedback. However, increases in LH pulse frequency were also induced by naloxone treatment at a postpubertal age in estradiol-treated OVX sheep, indicating that opioid inhibition is still present at a time when sensitivity to the feedback effects of ovarian steroids is markedly reduced and endogenous LH secretion is increased. These observations in ovary-intact and estradiol-treated OVX lambs suggest that opioid mechanisms inhibit pulsatile tonic LH secretion during both the prepubertal and postpubertal periods. Endogenous opioid inhibition of LH secretion is not dependent on the presence of ovarian steroids, as evidenced by the response to naloxone 3 weeks after removal of an estradiol implant from OVX lambs, when LH pulse frequency was already high. Naloxone treatment increased LH pulse frequency further, at both a prepubertal age (18 weeks) and a postpubertal age (38 weeks). Naloxone also increased LH pulse frequency in OVX lambs in which LH secretion was inhibited chronically by progesterone rather than by estradiol. The response to naloxone was similar in postpubertal P-treated OVX lambs and age-matched prepubertal P-treated OVX controls in which puberty had been delayed by means of an inhibitory seasonal photoperiod. In addition, after removal of steroid implants to allow LH secretion to increase, the degree of inhibition of LH secretion by the opiate agonist morphine was similar between age-matched postpubertal sheep and those with photoperiodically delayed puberty. We conclude that endogenous opioid mechanisms are an important inhibitory mechanism controlling pulsatile LH secretion in the developing sheep. However, changes in opioid inhibition are unlikely to underlie the decrease in sensitivity to steroid negative feedback and increase in pulsatile LH secretion that occur at puberty.

Distribution of estrogen receptor-immunoreactive cells in the sheep brain.

The distribution of immunoreactive (IR) estrogen receptor (ER)-containing cells was studied in the brains of adult Suffolk ewes using a rat monoclonal antibody (H222) which recognizes the human estrogen receptor. IR cells were characterized by dense nuclear reaction product, and in some instances, cytoplasmic immunostaining which filled dendrite-like processes. The greatest densities of ER-IR cells were found in the medial preoptic area, the mediobasal hypothalamus, and in a number of limbic system structures (amygdala, bed nucleus of the stria terminalis, lateral septum). ER-IR cells were found at lower densities in several other subregions of the hypothalamus and limbic system, and in the periaqueductal gray of the caudal midbrain. Cytoplasmic ER immunoreactivity was most prominent among ER-IR cells in the ventrolateral-ventromedial nucleus, the bed nucleus of the stria terminalis, the midbrain periaqueductal gray, and some ER-IR cells in the substantia innominata. The distribution of ER-containing cells in the sheep brain closely parallels that seen in other mammals. ER-IR cells are found in sites such as the medial preoptic area and ventrolateral-ventromedial hypothalamus which have been implicated as targets in this species and others for the influence of estradiol on sexual behavior and reproductive neuroendocrine function.

Prenatal photoperiod influences neonatal prolactin secretion in the sheep.

This study tested the hypothesis that the fetal sheep can respond to photoperiod cues. Pregnant Suffolk ewes were maintained in artificial photoperiod of either long days [16 h of light, 8 h of dark (16L:8D)] or short days (8L:16D) from approximately 100 days of gestation until term at approximately 147 days. On the day of birth, all lambs and their mothers were transferred to an intermediate photoperiod of 12L:12D; both groups were housed together. To provide an index of response to photoperiod, serum PRL concentrations were measured in blood samples collected daily 3-4 h after lights on. In lambs (n = 8 male; n = 7 female) born to mothers on long days, serum PRL concentrations were high (greater than 200 ng/ml) for the first few days after birth, but fell rapidly to low levels (less than 50 ng/ml) within 14 days postnatally in 12L:12D. Conversely, lambs (n = 8 male; n = 7 female) born to mothers on short days initially had low PRL concentrations, but these gradually increased in the postnatal 12L:12D photoperiod to 150 ng/ml by 32 days of age. Thus, serum PRL concentrations in lambs at birth reflect the photoperiodic treatment of their mother, and the subsequent PRL response to an intermediate photoperiod of 12L:12D depends on the photoperiodic history received in utero. We infer from these findings that the fetal lamb receives and responds to information about day length in utero and begins developing a seasonal photoperiod history before birth.

Cessation of long day melatonin rhythms time puberty in a short day breeder.

This study tested the hypothesis that in the female sheep, a short day (SD) breeder, puberty can occur normally in the absence of ambient short days. More specifically, the photoperiod cue timing the transition into adulthood is exposure to and then termination of a long day melatonin rhythm. Control lambs born in the spring were exposed to 5 weeks of long days (LD; 16 h of light, 8 h of darkness; 18-23 weeks of age) and were raised in SD (8 h of light, 16 h of darkness) at other times. As expected from previous studies, this alternating photoperiod sequence (SD-LD-SD) induced puberty at the normal age in autumn [33 +/- 2 weeks (mean +/- SEM); n = 6]. The other three groups were exposed only to LD from birth; the superior cervical ganglia were removed bilaterally at different ages to denervate the pineal gland in order to block transduction of subsequent LD cues. Puberty occurred normally (31 +/- 1 weeks; n = 7) after ganglionectomy at 23 weeks of age, indicating that ambient short days are not required to initiate reproductive cycles. LD are necessary, as evidenced by the results for the other two groups ganglionectomized neonatally at 4 weeks of age. With no further treatment, puberty was either delayed (n = 1) or did not occur during the first year of life (n = 5), after which the study ended. This delay was prevented in the other group of ganglionectomized lambs by a 5-week (18-23 weeks of age) exposure to LD melatonin patterns by means of 8-h melatonin infusions nightly; 12 weeks after melatonin replacement therapy, puberty occurred at the normal time (34 +/- 1 weeks; n = 6). The inference is that for puberty to occur in the female lamb the animal must be exposed to relatively limited periods of LD, followed by the blockade or absence of further LD cues (pineal denervation, termination of LD melatonin infusion, or presence of SD). This supports the concept that the LD of summer, followed by their disappearance in autumn, time puberty in the female sheep.