Photoperiodic requirement for the dissipation of scotorefractoriness in Japanese quail

Male Japanese quail were reared on short days (6L:18D) and at 15-20 weeks of age those which had become sexually mature (i.e., scotorefractory) were transferred to long days (18L:6D) for between 2 and 29 weeks. The birds were then returned to 6L:18D for 3 weeks to test for the dissipation of scotorefractoriness. This was assessed by a decrease in at least 3 of 4 indices of reproductive function: testicular weight, area of the cloacal gland, and levels of plasma LH and testosterone. There was great individual variation in the photoperiodic requirement for the dissipation of scotorefractoriness, ranging between 6 and 29 weeks of exposure to long days. Scotorefractoriness was dissipated in about 50% of the birds after exposure to long days for between 6 and 12 weeks. It is concluded that the photoperiodic requirement for the dissipation of scotorefractoriness in quail cannot be defined precisely.     

Episodic release of LH in gonadectomized male Japanese quail

Blood samples were taken every 15 min (for 4.75 h) from six castrated quail on three separate occasions. The birds were first bled while under short days (comprising 8 h light : 16 h darkness per 24 h; 8L : 16D) and, subsequently, after 1 and 20 days under long days of 16L : 8D. The photoperiodic alteration produced a marked increase in the mean plasma LH concentration of each bird and in four instances the rise was evident after only 1 long day. Pulsatile release patterns were not detected in the plasma LH profiles obtained during exposure to short days or after 1 long day but were pronounced in all of the birds after 20 long days. The peaks (n = 19) occurred on average every 83 +/- 14 (S.E.M.) min and had an amplitude of 12.3 +/- 1.2 micrograms/l. After 35 days under 16L : 8D three of the quail were bled more frequently (every 8 min for 2.5 h), allowing the LH pulses to be measured more precisely. Each pulse was composed of a sudden increase in secretion followed by a slower decrease which lasted for approximately 30 min. The development and functional significance of episodic LH release in the quail is discussed.     

Quantitative bioassay of luteinizing hormone releasing activity in the quail hypothalamus during photostimulated sexual development.

A quantitative bioassay using dispersed chicken pituitary cells was employed to measure luteinizing hormone releasing activity in pooled hypothalamic extracts from groups of quail during photostimulated sexual development. Levels of releasing activity with 95% confidence limits were expressed in terms of microlitres of a standard hypothalamic extract (CS).LH releasing activity was measurable in the hypothalamus of male and female quail reared under both short and long photoperiods. Using an approximate conversion factor from CS, levels ranged between 5.1 and 13.3 ng synthetic LH-RH/mg basal hypothalamic tissue.Dramatic increases in LH secretion seen after transfer to photostimulatory daylengths were not associated with any large changes in hypothalamic LH releasing activity. Several significant fluctuations in levels of releasing activity were observed but these could not readily be correlated with changes in plasma LH levels. Combined data from four experiments show that the mean concentration of releasing activity was just significantly higher in groups of birds receiving 7 or more long days than in short day quail and birds receiving fewer than 7 stimulatory photoperiods (P < 0.05). This difference represents a 30% increase in releasing activity in long-term photostimulated quail.Castration did not alter LH releasing activity during one 5-week experiment, although a group of castrated birds maintained on long days for 1 year showed the highest level of releasing activity found in any quail.     

Refractoriness in quail leads to a reduction in the photoperiodic drive on LH secretion

The hypothesis was advanced that the long-day castration response in quail should become less pronounced in birds made refractory by exposure to long photoperiods. This was tested by pretreating two groups of castrated quail with testosterone for 9 weeks to suppress LH secretion whilst exposing them either to short days or to long days. The castration response was then measured on long days by withdrawing the testosterone and following the subsequent rate of increase in LH secretion. In both groups LH concentrations increased steadily for 7 weeks after removing the testosterone but the rate of increase in the group previously exposed to long days was only 50% of that in the group previously held on short days (P less than 0.025). The long-day castration response in quail not pretreated with testosterone was not altered by retaining the birds for 9 weeks on short days before transfer to long days.     

Extraocular control of seasonal reproduction in female tree sparrows (Spizella arborea)

The work reported here explored extraocular control of seasonal reproduction in a photoperiodic species. Photosensitive female tree sparrows (Spizella arborea) were subjected to bilateral or unilateral ocular enucleation or to sham operation and exposed to a photoregimen designed to stimulate luteinizing hormone (LH) secretion, to induce photorefractoriness, and to restore photosensitivity. As revealed by plasma LH profiles constructed from measurements taken over 181 days, neither bilateral nor unilateral ocular enucleation had a substantive effect on photoinduced LH release, on the development of photorefractoriness, or on the recovery of photosensitivity. In a related experiment, photorefractory sighted females were implanted epicranially with miniature self-powered lights (SPLs) or with non-emitting helium blanks (HBs) and transferred to short days. After 8 weeks, SPLs and HBs were removed, and birds were challenged with long days. As indicated by plasma LH and ovarian responses to the challenge, previously HB-implanted birds had regained photosensitivity, but previously SPL-implanted birds remained photorefractory. When interpreted within the context of the effects of blinding, these findings suggest that an extraocular encephalic mechanism maintains photorefractoriness. Collectively, the data support the hypothesis that photoinduced LH release and the biannual transitions between photosensitivity and photorefractoriness are controlled by an extraocular mechanism(s).     

Photoperiodic induction in vitro: the dynamics of gonadotropin-releasing hormone release from hypothalamic explants of the Japanese quail.

The Japanese quail is a photoperiodic animal that under certain experimental conditions can respond to a single long day with a wave of LH secretion. Such a system offers an opportunity to analyze the photoneuroendocrine changes as they occur in real time, especially as all of the neural machinery (photoreceptor, clock, and GnRH system) is believed to lie within the hypothalamus. The first detectable rise in LH occurs at about hour 23 of the long day, and this single inductive event leads to prolonged LH secretion lasting for up to 2 weeks and peaking 2-4 days after the dawn of the long day. The size of the quail's hypothalamus is such that the entire structure, including both the GnRH cell bodies and the median eminence, can be cultured for some hours, and the rates of GnRH release measured therefrom. The present experiments used hypothalamic explants from quail at different times throughout the photoperiodic response, superfused them for up to 7.5 h in vitro, and measured the dynamics of GnRH release. A significant step increase of 80% in GnRH release occurred between hours 22.5 and 23 in quail that had been exposed to a long day: an equivalent change was not found in hypothalami taken from quail maintained only under short day lengths. In explants taken from quail at the peak of LH secretion (53 h after dawn of the long day), the rates of GnRH release were double those found in control quail not exposed to the long day. Explants taken 14 days after the long day, when LH secretion had subsided fully, showed no difference in GnRH release between photo-stimulated and control quail. These results suggest that photoperiodic induction involves a timed increase in GnRH release, and the rise at hour 23 is believed to represent photoperiodic induction actually taking place within the brain in vitro. They also suggest that the wave of LH secretion triggered by the single long day is, at least in part, a neuroendocrine or neural phenomenon; this confirms earlier indirect evidence to this effect.     

Detection of neural connection to the infundibular complex by partial or complete hypothalamic deafferentation in male quail

To elucidate the stimulatory and inhibitory neural systems for photoperiodic control of avian reproduction, immature male Japanese quail were subjected to partial or complete hypothalamic deafferentation, followed by exposure to long and short photoperiods. The results indicated that when the encephalic photosensitive area (infundibular complex, INF) was preserved after hypothalamic deafferentation, birds were able to respond to long days and their gonads eventually recrudesced, and that testicular atrophy under short days was prevented by the semicircular cuts posterior to INF or by orbital enucleation. It is concluded that in male Japanese quail, INF plays the pivotal role in photoperiodic gonadostimulation and regulatory neurons in the retina and anterior hypothalamus may have neural connection to the posterior side of INF.     

The pineal gland and the photoperiodic control of luteinizing hormone secretion in intact and castrated Japanese quail

The effects of pinealectomy on a range of photoperiodic responses were investigated in male Japanese quail by measuring plasma LH concentrations in intact, sham-operated and pinealectomized birds in the following four experiments: (1) transfer of sexually quiescent birds from a short photoperiod of 8 h light: 16 h darkness (8L:16D) to a photostimulatory daylength of 16L:8D; (2) transfer of sexually mature birds from 16L:8D to 8L:16D; (3) castration in 16L:8D and exposure to 13L:11D; (4) castration in 8L:16D and exposure to 13L:11D. There was no evidence of effects of the pineal gland on the photoperiodically induced changes in LH secretion, the quantitative relationship between LH secretion and photoperiod in intact and castrated birds, or the induction of relative photorefractoriness by prolonged exposure to 16L:8D. This suggests that there is no pineal influence on the photoperiodic clock or its effectors in this bird.     

Neuroendocrine regulation of gonadotrophin-releasing hormone secretion in the Japanese quail

Following the transfer of quail to long days, an increase in plasma and pituitary levels of LH and FSH is observed. The integrity of both the preoptic region (POR) and the posterodorsal part of the infundibular nuclear complex (PD-INC) appears to be essential for mediating this photoperiodic response. The INC may, however, operate autonomously in being able to regulate the low levels of LH-RH release of nonphotostimulated immature birds. By combining the techniques of electrical stimulation and electrolytic lesioning it can be demonstrated that stimulation of the POR can increase LH-RH secretion in quail where the photoinduced release has been blocked by a PD-INC lesion. Electrical stimulation of the preoptic region in immature birds similarly stimulates the secretion of LH-RH. These results suggest a direct link, perhaps neurosecretory, between the preoptic region and the median eminence.     

The stimulation of luteinizing hormone and follicle-stimulating hormone secretion in quail with complete and skeleton photoperiods

Photostimulation of quail by long daylengths stimulates LH and FSH release, the earliest increases in these hormones being detectable after 1 long day. Reproductive maturity is complete within 1 month. Steroid feedback becomes important during the second week of photostimulation and restricts LH and FSH secretion to a fraction of the concentrations observed in gonadectomized birds. The photoperiodic response is not all-or-none, and above a certain “critical daylength” the rate of gonadal growth, primarily controlled by the levels of FSH and testosterone, is proportional to daylength. Photoperiodic responses are qualitatively identical in gonadectomized and intact quail, arguing that stimulatory daylengths act directly on the hypothalamo-pituitary axis to alter LH and FSH secretion, and do not act by altering steroid feedback sensitivity and thereby gonadotrophin release. Any changes in feedback sensitivity are a consequence not a cause of the photoperiodic drive on the system. The use of “skeleton” photoperiods to mimic complete photoperiods is discussed. Whether or not one or two peaks of photoperiodic induction appear in asymmetric skeleton experiments depends both upon the duration of the first light period and of the night break itself. The rate of induction is less when quail are photostimulated with night breaks than with “complete” daylengths. The temporal position of the circadian rhythm(s) involved in photoperiodic time measurement in quail is controlled by both “dawn” and “dusk,” and so the position of the rhythm is phase-delayed as the days lengthen, occurring later and later in the night. This arrangement might have some bearing on the seasonal shift in critical daylength that occurs in quail exposed to natural photoperiods. The situation is compared with that in insects and plants.     

Photoinducible phase-specific light induction of Cry1 gene in the pars tuberalis of Japanese quail

Prolactin (PRL) secretion is regulated by photoperiod in mammals and birds. In mammals, the pars tuberalis (PT) in the pituitary is involved in the regulation of photoperiodic regulation of PRL secretion. In birds, however, hypothalamic vasoactive intestinal peptide is implicated in PRL secretion, and physiological roles of the avian PT remain unknown. In the present study, we show that PRL secretion increases under long days and short days with a night interruptive schedule, both of which also cause gonadal growth in Japanese quail. We have also found Cry1 gene expression in the PT of Japanese quail. Cry1 expression was rhythmic under long and short photoperiods in the PT, and the peak was phase delayed under a lengthened photoperiod. Moreover, expression of Cry1 gene was induced by a light pulse but only when given during the photoinducible phase. In our previous study, we have shown rhythmic Per2 gene expression with a peak in the PT during the early day under various photoperiods. When taken together with the results from the present study, different phase relationships between Per2 and Cry1 in the Japanese quail PT under different photoperiods may decode photoperiodic information and regulate photoperiodic PRL secretion in a manner similar to that of mammals.     

Termination of LH secretion in Japanese quail due to high- and low-temperature cycles and short daily photoperiods.

Mature male Japanese quail were transferred from 16L:8D (19 degrees) to one of the following combinations of daily light-dark and temperature cycles, 8L:16D (12 hr, 19 degrees:12 hr, 9 degrees), 12L:12D (12 hr, 19 degrees:12 hr, 9 degrees) and 12L:12D (16 hr, 19 degrees:8 hr, 9 degrees). The low temperature is for the middle of the dark period in each treatment. In the control groups, birds were transferred to the same photoperiodic conditions as the experimental groups, but without changes in ambient temperature. Blood samples were collected every other day for 30 days and circulating levels of plasma LH were estimated by radioimmunoassay. Both the change in conditions from 16L:8D to 8L:16D with the temperature lowered for 12 hr and that from 16L:8D to 12L:12D with temperatures lowered in one case for 12 hr and in the other for 8 hr caused a lowering in plasma LH levels in all the birds to reproductively quiescent levels. The cloacal protrusion of all these birds regressed completely. In control groups, however, most if not all the birds remained in active breeding states although the levels of circulating LH decreased to basal breeding levels of 1-2 ng/ml. The results indicated that in addition to a change from long to short days an alternation of high and low temperatures was sufficient supplementary information in causing termination of LH secretion and inducing regression of the gonads and the accessory sex organs in this species.     

Plasma follicle-stimulating hormone during photoperiodically induced sexual maturation in male Japanese quail.

Follicle-stimulating hormone (FSH) was measured in Japanese quail using a heterologous radioimmunoassay, the specificty of which was confirmed by its cross-reactions with purified chicken FSH and luteinizing hormone (LH). Plasma concentrations of FSH, LH and testosterone were determined in quail during the testicular growth and sexual maturation which follows their transfer from short to long daylengths. All three hormones could be detected in short-day birds but their concentrations were greatly increased following photostimulation. Plasma FSH increased 12-fold during the first 9 long days, remained at this level for a week, and then declined steadily so that by the time the birds were sexually mature the level of FSH had decreased to one-third of the maximum level. LH reached a high level (five times the short day level) after 4 long days. Thereafter two patterns of LH secretion could be distinguished. In one experiment the high level of LH was maintained unchanged throughout sexual maturation while in another experiment LH secretion decreased significantly between days 11 and 28 of photostimulation. A strong correlation existed between testicular growth and the plasma FSH concentration. It was maximal during the phase of rapid testicular growth and decreased as spermiogenesis began. The pituitary FSH content increased during photostimulation. Castration caused a 20-fold rise in plasma FSH compared with that in intact quail. The change in LH concentration after castration was about eightfold. The changes in hormone secretion were strikingly similar to those found during sexual development and puberty in the rat.     

Diurnal variation and the episodic release of plasma gonadotrophins in Japanese quail during a photoperiodically induced gonadal cycle.

Blood samples were taken every 3 h, over a 27 h period, from (1) a group of 12 intact male quail on short days (lights on 09.00-17.00 h) and during the 2nd, 15th and 36th day of photostimulation (lights on 09.00-05.00 h); (2) 12 castrated male quail on the 2nd, 20th and 43rd long day, and (3) 12 intact male quail on the 12th long day. Plasma LH was measured in all samples and FSH in the 43rd long day castrate and 12th long day intact male samples. Although there was considerable variation in the levels of LH and FSH, both between birds and between samples taken from the same bird, statistical analyses failed to reveal any diurnal (or circadian) rhythm at any time. There was a marked correlation between the LH and FSH levels in all samples. Possible episodic LH secretion was investigated by taking blood samples every 15 min for between 3 and 6 h from six intact male quail and six laying females on long days. Samples were obtained from each bird at three time-periods which were arranged so as to overlap and cover the first 12 h of the daily photoperiod. Statistical analysis suggested that episodes of secretion occurred 6-10 times/day in males, and 4-8 times/day in females. The pulses appeared to occur at random.     

Retinal modulation of the hypothalamic sensitivity to testosterone feedback in photoperiodism of quail

This experiment was performed on two groups of male Japanese quail. One had been maintained in our laboratory as a closed colony (S-group), and the other had been obtained from a commercial source (R-group). Different responses of gonadal function were found between two groups following either testosterone treatment or exposure to short days. Immature birds of these groups responded to long days with rapid gonadal growth, but after sexual maturity, exposure to short days for 3 weeks induced testicular atrophy only in S-group. Involvement of the feedback effect of androgen in the photoperiodic response was then examined. Under long-day conditions, intraperitoneal placement of testosterone propionate (TP)-filled Silastic tube for 2 weeks decreased testicular weights in S-group but not in R-group. Apparently, sensitivity to short days is closely correlated with sensitivity to testosterone in the adult male. By bilateral enucleation, quail of S-group became less sensitive to both gonad inhibitory effect of short days and the negative feedback effect of TP. These results suggest that the photoperiodic mechanisms that are primarily mediated by the retinal system play a role in altering sensitivity to steroid feedback at the hypothalamus.     

Changes in gonadotrophin-releasing hormone and LH in Japanese quail during the first few days of photostimulation

Castrated Japanese quail responded to a single long day with a five- to eightfold increase in plasma LH levels. A rise in LH secretion appeared 19-24 h after dawn and LH levels were still increased 3 days later, despite the fact that the birds had been returned to a short daylength. Pituitary LH content decreased, reflecting these changes in secretion, although significant falls in content were only found 36-96 h after dawn, when LH secretion was maximal. Hypothalamic gonadotrophin-releasing hormone content was not altered. One interpretation of this is that increased synthesis of the peptide compensates fully for the increased secretion.     

The roles of day length and the testes in the regulation of plasma LH levels in photosensitive and photorefractory willow ptarmigan (Lagopus lagopus lagopus)

The interaction between day length and testicular hormones in the regulation of seasonal breeding in willow ptarmigan was investigated by measuring the changes in plasma LH levels after castration of photosensitive and photorefractory birds subsequently exposed to long or short days. Plasma LH levels increased after castration in photorefractory and photosensitive birds exposed to long days but this increase was greater (P < 0.001) in the photosensitive than in the photorefractory birds. Plasma LH levels increased immediately after castration in photosensitive but not in photorefractory birds exposed to short days. In the latter plasma LH levels increased steeply (P < 0.001) after exposure to short days for more than 5 weeks. This increase may have occurred at the time when the birds regained photosensitivity. The increase in plasma LH after castration in photosensitive birds was greater (P < 0.001) in those exposed to long days than in those kept on short days. We concluded that in the willow ptarmigan (1) there are seasonal changes in the sensitivity of the hypothalamo-hypophyseal unit to the inhibitory feedback action of steroids and (2) that the central nervous mechanism which controls seasonal breeding is stimulated more by long days than by short days in both photosensitive and photorefractory birds. These findings are consistent with the hypothesis that in the willow ptarmigan, seasonal breeding results from an interaction between a direct effect of day length on LH secretion and day length-induced changes in the sensitivity of the hypothalamus to the inhibitory feedback action of adrenal and testicular steroids.     

Involvement of catecholaminergic mechanisms in the photoperiodically induced rise in serum luteinizing hormone of Japanese quail (Coturnix coturnix japonica)

The involvement of brain catecholamines (CAs) in the neuroendocrine mechanisms that control the release of LH has been studied in somatically mature quail with regressed and with recrudescing testes and in sexually mature quail. In quail with recrudescing testes, serum LH was markedly increased 3 days after the onset of photostimulation. Administration of an inhibitor of CA synthesis, α-methyltyrosine (MT), started on the day of photostimulation and continued for 3 days thereafter, decreased brain dopamine (DA), norepinephrine (NE), and epinephrine (E) content and blocked the photoperiodically induced rise in serum LH. Treatment with bis(4-methyl-1-homopiperazinil-thiocarbonil) disulfide (FLA63), an inhibitor of DA-β-hydroxylase, caused a marked reduction of brain NE and E levels and a significant increase in brain DA content, and like MT, blocked the photoperiodically induced rise in serum LH. In sexually mature quail, serum LH and testicular regression were effected differently by MT and FLA63. MT partially reduced LH levels, but FLA63 suppressed them completely (i.e., to those of nonphotostimulated birds). Testicular weights were reduced by both MT and FLA63, but as with LH, the FLA63 was more effectual than MT. Treatment with l-dihydroxyphenylalanine, a direct precursor of DA, augmented DA content in the brain, caused a reduction of LH levels, and induced gonadal regression. The results are consistent with the hypothesis that NE-containing neurons facilitate central mechanisms which release LH in response to neural inputs involved in the photoperiodic stimulus, and that activation of DA-containing neurons is capable of inhibiting this release and inducing testicular regression.     

Seasonal change in luteinizing hormone subunit mRNA in Japanese quail and effects of short daylength and low temperature

Changes in pituitary mRNA levels of LHbeta-subunit (LHbeta) and glycoprotein hormone alpha-subunit (common alpha) were investigated in male Japanese quail under natural and laboratory conditions to clarify the mechanisms of seasonal regulation of luteinizing hormone (LH) secretion. In Experiment 1, birds were kept in outdoor cages under natural conditions from August for 12 months. Both LHbeta and common alpha mRNA levels decreased rapidly from August to September, and after a period of low levels from October through January, they began to increase in February and continued to increase until July. There were more pronounced seasonal changes in testicular weight and cloacal protrusion width with large decreases from August to September and increases from March to May. In Experiment 2, birds were kept on laboratory conditions and transferred from long to short daylengths at 20 or 9 degrees C and held for 14 days. Although common alpha mRNA levels, plasma LH concentrations, testicular weight, and cloacal protrusion area decreased on short days without low temperatures, levels of LHbeta mRNA did not change. Short daylengths combined with low temperatures induced testicular regression and caused decrease in all the parameters measured. Low temperatures under long days did not induce any change in the parameters significantly. These results suggest that (1) synthesis as well as secretion of LH is regulated seasonally, (2) short daylength does not suppress LH synthesis completely unless combined with low ambient temperature, and (3) the effect of photoperiod on the endocrine system regulating LH secretion is predominant over the effect of ambient temperature but ambient temperature acts as an environmental cue to terminate reproductive activities at late summer to early autumn in Japanese quail.     

Photoperiodic control of LH secretion in Japanese quail with special reference to the photoinducible phase

Repetitive blood sampling from Japanese quail showed that plasma LH concentration rose during the first dark period after the birds were transferred from 8L:16D to 16L:8D. In the second light period of long days, LH concentration decreased slightly (not significant) and maintained this lower level until the second dark period of long days. During the second dark period LH again increased. This suggests that at the first stage of LH release by long photoperiod, photoinduction can occur at least during 8 to 16 hr after lights are turned on. To test the hypothesis that LH secretion induced by long days is dependent on the photoinducible phase, 0.5-hr light pulses (0.5L) were given to quail kept under 8L:16D during the dark periods. Activity was also recorded for each bird. After 10 days of treatment, testicular growth was induced when 0.5L pulses were given at 13 or 19 hr after dawn. Testicular growth induced by 0.5L pulses given at 19 hr after dawn was less than that in birds given light 13 hr after dawn and had greater individual variation compared with other groups. The activity records of these birds revealed that some remained entrained to the primary light pulse (8 hr) while others phase shifted to the short pulse (0.5 hr). The latter were, in effect, then exposed to a 13-hr photoperiod while the former were exposed to 8L:11D: 0.5L:4.5D. LH increase by 0.5L given during the dark period was detected after 2 days of treatment when the pulses were delivered at 15 hr after dawn. These results suggest that LH release is induced when light impinges on the circadian photosensitive phase which is set by external lighting schedules.