Emberiza Melanocephala Research Articles

THE PHOTOPERIODIC ENTRAINMENT AND INDUCTION OF THE CIRCADIAN CLOCK REGULATING SEASONAL RESPONSES IN THE MIGRATORY BLACKHEADED BUNTING (EMBERIZA MELANOCEPHALA)

The properties of the circadian photoperiodic oscillator have been investigated in detail only in the Japanese quail. While the study of the quail is clearly very important, one cannot simply assume that other species, especially passerines that seem to have a different circadian organization than quail, function the same way. The current set of experiments was conducted to understand the entrainment and photoinduction of the circadian photoperiodic oscillator in a passerine species, the blackheaded bunting (Emberiza melanocephala). The experimental paradigm used skeleton photoperiods with two light periods, the first called the “entraining light pulse” (E-pulse) and the second called the “inducing light pulse” (I-pulse). Three experiments were performed on photosensitive male birds (N=6–8/group). Experiment 1 investigated the effects of the temporal relationship between E- and I-pulses on photoperiodic induction. Buntings entrained to 8h:16h L:D for 4 wk were released into constant dim light (LLdim, ∼1 lux). Beginning on subjective day 8, they received for 8 wk, E- and I-pulses only at alternate cycles. While I-pulse was 1 h and always began at zt 11.5, E-pulse varied in duration and timing (the 1h E-pulse beginning either at zt 0, zt 5, or zt 9, the 4h one beginning at zt 0 or zt 6, and the 10h one at zt 0; zeitgeber time 0=time of lights-on under 8h:16h L:D prior to release into LLdim). A photoperiodic response was induced only when the E-pulse began at zt 0, and thus the beginning of E- and I-pulses were separated by 11.5 h. Experiment 2 determined whether the duration of the E-pulse influences the position of the photoinducible phase (ϕi) of the circadian photoperiodic oscillator. Birds were entrained to 1h:23h L:D or 10h:14h L:D for 2 wk, and then exposed to 1h I-pulse at zt 11.5, zt 15, or zt 18.5 for another 8 wk. Photoperiodic induction occurred at all 3 zts in birds entrained to 10 h but only at zt 11.5 in birds entrained to 1 h, which infers the circadian rhythm of photoinducibility (CRP) in buntings was re-entrained when I-pulse fell at zt 15 and after. The last experiment examined the possibility of the re-entrainment of the CRP to light pulses falling at zt 15 and after. Birds received 1h I-pulse for 8 wk at zt 15 following 2 wk of 2.5h:21.5h L:D or 3.5h:20.5h L:D, or at zt 21.5 or zt 22.5 following 2 wk of 10h:14h LD. Photoperiodic induction was consistent with the hypothesis of the re-entrainment of the CRP under these light–dark cycles. The I-pulse appeared to be interpreted as a “new dawn”, and so the photoperiodic induction was determined by the coincidence of ϕi with the E-pulse. These results suggest a phase-dependent action of light on the circadian oscillator regulating photoperiodic responses in the blackheaded bunting. This could be a useful strategy for a photoperiodic species to regulate its seasonal responses in nature.

Effects of duration and time of food availability on photoperiodic responses in the migratory male blackheaded bunting (Emberiza melanocephala).

The effects of the duration and time of food availability on stimulation of the photoperiodic responses (fattening and gain in body mass, and growth and development of testes) were investigated in the migratory blackheaded bunting (Emberiza melanocephala). Two experiments were performed. Experiment I examined the effects of a reduction in the duration of food supply in buntings that were subjected to long day lengths (16h:8h L:D) and received food ad libitum (group I) or for restricted durations, coinciding with the end of the lights-on period, of 8h (group II) and 4h (group III). Buntings of group I gained in body mass, whereas there was a mixed response in group II (half the birds gained and half lost body mass), and all birds of group III lost body mass. There was no effect on testis growth in groups I and II, but testes grew more slowly in group III. Experiment 2 investigated the effects of both the duration and the time of food availability. Of five groups of birds, group I was exposed to an 8h:16h L:D photoperiod, and groups II-V were exposed to 16h:8h L:D. Whereas birds of groups I and II received food ad libitum, those of groups III-V were fed only for 5 h, at zt 0-5 (group III), zt 5.5-10.5 (group IV) or zt 11-16 (group V), where zt = zeitgeber time and zt 0 refers to the beginning of the lights-on period. Apart from duration, the timing of food availability also had an effect on photoperiodic stimulation under the 16h:8h L:D photoperiod. Birds that were fed ad libitum fattened and gained in body mass, whereas among restricted feeding groups, only birds in the group fed during the first 5 h (zt 0-5, group III) showed a significant increase in body mass (albeit considerably lower than in the ad libitum group). Birds fed during the middle 5h (zt 5.5-10.5, group IV) showed an intermediate response, and those fed during the last 5h (zt 11-16, group V) lost body mass. Testicular growth was suppressed in birds that were fed for 5 h in the evening, but not in those fed for the same period in the morning or in the middle of the long day. Taken together, these results show that the duration of food supply and/or the time of day at which food is available affect photoperiodic stimulation of fattening and gain in body mass as well as the growth and development of gonads in the blackheaded bunting.

The influence of light wavelength on reproductive photorefractoriness in migratory blackheaded bunting (Emberiza melanocephala).

There are two effects of long day length on reproductive responses in birds, one is the photoinduction of gonadal growth and maturation and the other is the induction of gonadal regression and photorefractoriness. Although it is likely that the same photoreceptors are involved in the photoinduction of gonadal growth and the onset and maintenance of photorefractoriness. and so the influence of wavelength should be similar, this has not been investigated. Therefore, we investigated the influence of light wavelength on reproductive photorefractoriness in the migratory male blackheaded bunting held under long photoperiods. In mid May, when photoperiod was approximately 14L:10D (14 hours light:10 hours darkness), eight groups of sexually mature birds were moved indoors on an artificial photoperiod of 14L:10D (L - 450 lux. D - 0 lux). Then after 3 weeks, for six groups, a 4-h light period in the morning (zt 0-4; zt 0 [zeitgeber time 0] refers to the beginning of lights-on period) or in the evening (zt 10-14) was substituted with green (428 nm), red (654 nm) or white light at 16 +/- 2 lux intensity. Of the remaining two groups, one was maintained on 14L: 10D and the other transferred to 10L:14D: these served as controls. At the end of 4 weeks, all birds were found to have undergone testicular regression, irrespective of LD cycle they were exposed to. When these gonadally regressed birds were subjected to 16L:8D for another 4 weeks, to test their responsiveness to the stimulatory effects of long day lengths, only those exposed to 10L:14D and 14L:10D with a 4-h green light period showed testicular regrowth. On the other hand, birds exposed to 14L:10D with a 4-h white or red light period remained fully regressed, similar to 14L:10D controls. Except for some individual difference, there was no difference in response between the groups that received a 4-h light period in the morning and that received it in the evening. These results suggest that the wavelengths of light influence induction of buntings from the photosensitive state into the photorefractory state. Whereas the short light wavelengths facilitated recovery from the photorefractoriness, the long light wavelengths were more effective in maintaining the photorefractoriness.

Photoperiodic induction of ovarian growth and plasma estradiol secretion in a migratory finch, Emberiza melanocephala: Involvement of circadian rhythm

To demonstrate the involvement of circadian rhythm in photoperiodic time measurement, photosensitive female blackheaded buntings were kept under different photoperiodic schedules consisting of 6 h of main photophase coupled with scotophases of various durations. Ovarian mass and circulating plasma estradiol concentration were found to be elevated in cycles of 6L:6D, 6L:36D, 6L:54D and in control 15L:9D groups. But cycles of 6L:18D, 6L:42D and 6L:66D did not stimulate ovarian growth or elevate circulating plasma estradiol concentration. These results are consistent with the Bünning hypothesis according to which a photoperiodic response is elicited as a result of the coincidence of light with the photoinducible phase of an endogenous circadian rhythm. The results thus indicate the involvement of a circadian rhythm of photoinducibility in ovarian growth and estradiol secretion.

Changes in body weight, ovarian growth, and circulating plasma estradiol level in response to programmed photoperiods in blackheaded bunting, Emberiza melanocephala

Changes in body weight, ovarian growth, and circulating plasma estradiol level in response to programmed photoperiods in blackheaded bunting, Emberiza melanocephala

Changes in body weight, ovarian growth, and circulating plasma estradiol level in response to programmed photoperiods in blackheaded bunting,Emberiza melanocephala

Changes in body weight, ovarian growth, and circulating plasma estradiol level in response to programmed photoperiods in blackheaded bunting,Emberiza melanocephala

Involvement of Threshold Photoperiod in Control of Reproductive Rhythmicity in Migratory Blackheaded Bunting, Emberiza Melanocephala

To explore the need for minimum threshold photoperiodicity in regulation of metabolic and reproductive activities of a migratory finch, various programmed light-dark (LD) schedules, such as P1 (3L/21D), P2 (6L/18D), P3 (9L/15D), P4 (12L/12D), P5 (15L/9D), P6 (18L/6D), P7 (21L/3D), and P8 (24L/0D), have been used on photosensitive female blackheaded buntings for 42 days. Results indicate that the photoperiodic thresholds of 3h, 6h, and 9h completely failed to have any response on buntings, while threshold photoperiodicities of 12h, 15h, 18h, 21h, and 24h had significant effect (P <. 001) on body weight, ovarian weight, and circulating plasma estradiol concentration, suggesting the role of the photoperiod as a primary environmental source to regulate various metabolic and reproductive functions. Further, it has been suggested that the threshold photoperiod in this species appears to be of 12h duration.

Involvement of circadian rhythm in response to ultrashort photoperiods in blackheaded bunting, Emberiza melanocephala

Groups of photosensitive adult female blackheaded buntings were exposed to various ultrashort days photoperiodic regimes for 60 days in which a fixed 3 h photophase was coupled with dark phases in cycles of 22 to 32 hours duration. One group of buntings was kept in long days (15L:9D) as control. Significant increase in ovarian weight and circulating plasma estradiol concentration was marked in the cycles of 30 h (3L:27D), and 32 h (3L:29D) photoperiodic schedules as well as in control group (15L:9D), whereas there was no response in the cycles of 22 h (3L:19D) and 24 h (3L:21D). It seems that the response to ultrashort day cycles is due to a phase advance or delay in photosensitivity of the response system repeatedly shows coincidence of the external photophase (3 h) with the photoinducible phase of an endogenous circadian rhythm. Therefore, the present result appears to be consistent with Bünning hypothesis suggesting the involvement of an endogenous circadian component in the female blackheaded bunting.

Effects of wavelength and intensity of light in initiation of body fattening and gonadal growth in a migratory bunting under complete and skeleton photoperiods

A series of experiments investigated the effects of wavelength and intensity of light in initiation of body fattening (gain in body weight) and gonadal growth in migratory blackheaded bunting under complete and skeleton photoperiods. Using fluorescent light at an intensity ∼ 700 lx, the first experiment compared the inductiveness of a complete (13 h continuous light coupled with 11 h darkness; 13L:11D) and a skeleton (two light pulses of 6 and 1 h at 6 h apart; 6L:6D:1L:11D) photoperiod. Observations at the beginning and after 3, 8, 10, and 13 weeks of the treatment indicated that both photoperiods were fully inductive but that birds under 13L photoperiod fattened and lost body weight significantly earlier than birds under skeleton photoperiod. In the second experiment, bunting were subjected to 13L:11D ( L = 100 lx; D = 0 lx) of white, green (528 nm), and red (654 nm) light for a period of 5 weeks. Birds gained weight and testes grew in all groups except for an inconsistent fattening response in the white light group. The third experiment tested if the inductive effects of 1-h light pulse in a skeleton photoperiod were intensity dependent. Groups of bunting were exposed to 6L:6D:1L:11D (intensity of 1-h white light pulse = 2, 10, 50, or 100 lux) and examined at the beginning and after 3 and 8 weeks of the treatment. Photoinduction occurred at a slower rate and only at 50- and 100-lx intensities. The fourth experiment was similar to the third in design but it employed 1-h light pulse of two different wavelengths (green = 528 nm, and red = 654 nm) at 50- and 100-lx intensities. Birds fattened and testes grew only under red light. The last experiment varied the wavelength and intensity of the first (6 h) light pulse of the skeleton photoperiod (6L:6D:1L:11D): 6-h entraining light pulse of white, green (528 nm), or red (654 nm) colour at 10- or 50-lx intensity was used with 1-h inducing light pulse of white light at ∼ 700- lx intensity. Testes grew in all groups but significant fattening occurred only in birds entrained to 50-lx light intensity. These results indicate i) the dissociation of body weight and gonadal responses, in the sense that the timing of photostimulation and/or magnitude of photoperiod-induced body weight and testicular responses differed under various photoperiodic manipulations, and ii) the circadian processes involved in photoperiod-induced responses have differential spectral and intensity sensitivity.

The photoperiodic clock in blackheaded buntings (Emberiza melanocephala) is mediated by a self-sustaining circadian system

Three experimental protocols were employed to clarify whether the circadian system is involved in photoperiodic time-measurement in the blackheaded bunting, Emberiza melanocephala. In a single-pulse paradigm, one 8-h light pulse was delivered at different times to groups of birds across three days of constant darkness (DD). Photoperiodic induction, as measured by a rise in plasma luteinizing hormone (LH), showed clear circadian rhythmicity. The second experiment examined the LH responses in birds exposed to lighting cycles using a Nanda-Hamner type of protocol and confirmed full photostimulation under 6L:30D. The third experiment measured the time of the first photoinduced rise in LH in birds subjected to 30 h of continuous light following entrainment under short days (6L:18D). This experiment aimed to identify the position of the photoinducible phase (phi i). LH first rose at hour 18 following dawn indicating that phi i lies in the middle of the day. Plasma concentrations of melatonin were also measured under 6L:18D and 6L:30D light cycles as another physiological marker of the circadian systems in buntings. The pattern of melatonin secretion with the driving oscillator being circadian in nature. It is concluded that the circadian pacemaker driving the photoinducible rhythm in blackheaded bunting is strongly self-sustaining and free-runs under constant conditions.

Entrainment of circadian system under variable photocycles (T‐photocycles) alters the critical daylength for photoperiodic induction in blackheaded buntings

AbstractThe present study was designed to investigate the photoperiodic responses of blackheaded buntings to various T‐photocycles [T =the period length of light:dark (LD) cycle]. Two series of experiments were performed. In series I, groups of buntings (n =8‐12) were exposed to LD cycles of 22 to 26 hr with photoperiods of 6, 10, or 11 hr (T22 =6L:16D, 10L:12D, 11L:11D; T24 =6L:18D, 11L:13D; T26 =6L:20D, 11L:15D). Among these groups, birds subjected to 11L:11D showed heavy deposition of fat, weight gain, and testicular recrudescence, a response that is characteristic of long day (≥ 12 hr photoperiod per day) photostimulation, indicating that a T‐photocycle of less than 24 hr (T =22 hr) was photoinductive in buntings when presented with a photophase long enough to interact with the photoinducible phase (ϕi), located in the middle of the photocycle. Experiments in series II included an interrupted‐night lighting protocol with a main photophase of 6 hr and 1‐hr light break in photocycles of T =22 hr, T =24 hr, and T =26 hr. The results revealed that a 1‐hr light pulse was inductive when it was introduced in the middle of the photocycles, and, thus, was coincident with the ϕi. It is suggested that the critical daylength for photoperiodic induction in buntings is altered as a result of entrainment to various T‐photocycles. © 1995 Wiley‐Liss, Inc.

Changes in food intake, body weight, gonads and plasma concentrations of thyroxine, luteinizing hormone and testosterone in captive male buntings exposed to natural daylengths at 29° N

We have investigated the seasonal changes in food intake, body weight, gonadal volume and plasma concentrations of thyroxine, luteinizing hormone and testosterone in male blackheaded bunting (Emberiza melanocephala) in captivity under natural daylengths at 29° N. The cycles in food intake, body weight and testis size in buntings appeared to be phase related. While the changes in body weight and testicular size were parallel to each other and correspond to the increasing daylengths of spring and early summer, cycle in food intake was almost antiphase to the cycles in body weight and testicular growth and development. Furthermore, buntings showed a distinct seasonal cycle in plasma concentrations of thyroxine, luteinizing hormone and testosterone. It is suggested that these seasonal cycles in buntings are endogenously programmed and their entrainment to the environmental photoperiod ensures the occurrence of different physiological functions at temporally fixed time of the year.

Photorefractoriness in the black-headed buntingEmberiza melanocephala: Possible involvement of the thyroid glands

In the male black-headed bunting,Emberiza melanocephala, photostimulated testicular and/or body weight growth was followed by the regression. Transfer of photorefractory birds from 20L/4D to 23L/1D or from natural lighting (12–13 h) to 20L/4D failed to evoke testicular and/or body weight recrudescence. Thyroidectomy suppressed light-induced increase in the testes and resulted in early regression. Fully developed testes of breeding birds also regressed following thyroidectomy; an effect which was reversed by daily injections of 1 μg/bird of L-T4. Treatment with L-T4 at doses from 0·5-2·0 μ/bird/day/ 30 days had no effect on the testes of birds maintained on 12L/12D or following a shift from 12L/12D to 20L/4D. Photoinduced increase in body weight was inhibited by thyroidectomy; an effect which was reversed by treatment with L-T4 at dose level 1 μg/bird/day. The extent to which thyroidectomy decreased body weight of birds depended upon the lipid reserves at the time of operation. It is suggested that in the male black-headed bunting (i) breeding is terminated by development of absolute-gonadal and metabolic-photorefractoriness and (ii) thyroid hormones are necessary for sustaining light-induced increase in the gonads and/or body weight and for their maintenance, but not for the development of photorefractory state.

Plasma levels of luteinizing hormone in intact and castrated photosensitive blackheaded buntings (Emberiza melanocephala) exposed to stimulatory and nonstimulatory photoperiods.

Experiments were performed to investigate the effects of castration and/or photostimulation on plasma luteinizing hormone (LH) in photosensitive blackhead buntings. Castration evoked a significant rise in plasma LH in birds held under 8 h light: 16 h darkness (8L:16D). On exposure to 18L:6D for 24 d, both the intact birds and the castrates exhibited a significant rise in plasma LH, although the change in LH levels in the castrates was approximately 8 times greater than in the intact birds. When control birds under 8L:16D which showed no change in plasma LH up to d 13 were transferred to a 36-h resonance cycle (8L:28D), plasma LH rose significantly within 4 cycles of the treatment. Furthermore, the intact buntings showed rapid increase and decrease in the plasma LH levels during a 16-wk exposure to long (16L:8D) but not to short (8L:16D) photoperiods. These results suggest that: i) the photoperiodic drive on LH secretion is inhibited by the testes; ii) the blackheaded bunting has a strong photoresponsive system; and iii) a circadian rhythm of photosensitivity is involved in the photoperiodic time measurement in this species.

Thyroid involvement in photoperiodically induced testicular and body weight cycles of the black-headed bunting,Emberiza melanocephala

In the black-headed bunting, Emberiza melanocephala, thyroidectomy suppressed the amplitude of light-induced testicular growth and accelerated the onset of photorefractoriness. Extirpation of the thyroid glands during progressive phase inhibited further testicular growth and led to immediate testicular regression when performed during peak phase of a photostimulated cycle. The inhibitory influence of thyroidectomy on light-induced testicular growth was fully reversed by daily im injection of L-T4 at a dose level of 2 μg/bird/day. Neither thyroidectomy nor L-T4 had any effect on the regressing testes of thyroidectomized buntings. Body weight which was about 26 g increased to about 37 g within 15 days of photostimulation by 20L/4D. Removal of thyroid glands suppressed light-induced increase in body weight and resulted in premature decline. The inhibitory influence of thyroidectomy on body weight depended upon lipid reserves at the time of operation, an effect which could be reversed by daily im treatments of L-T4 at dose level 2 μg/bird/day. It is suggested that in the black-headed buntings thyroid hormones are required for sustaining light-induced increase in the testes and body weight and may not necessarily be involved in the development of photorefractory states—gonadal and metabolic. © 1992 Wiley-Liss, Inc.

Investigations of photoperiodically induced fattening in migratory blackheaded bunting (Emberiza melanocephala)(Ayes)

The body fattening and weight gain preceding vernal migration in birds is timed by a set of environmental factors of which daylength is predominant. However, the mechanism(s) by which these events is determined is poorly understood. Previous investigations on a photoperiodic migratory species, the blackheaded bunting (Emberiza melanocephala), indicate the involvement of a light‐sensitive circadian rhythm during initiation of fat deposition and body weight gain. This communication presents data from another set of experiments aimed to characterize further the mechanism(s) of fat deposition in the same species.Groups of photosensitive, unstimulated and stimulated birds were subjected to transfer and superimposition experiments for 30 days. While the former set included shifting of long‐day (LD) birds to DD, SD (short days), DD/LD and SD/LD, in the latter a 90‐minute bright light was superimposed at two different times of the day during the dim‐green lighted phase 15L:9D of varying intensity. Birds were weighed at the beginning and at the end of experiments. Those in transfer cycles were also weighed at 10‐day intervals. The results suggest that the premigratory body fattening and weight gain in blackheaded buntings is light dependent and timed by environmental daylength in accordance with the photosensitive endogenous circadian rhythm (ECR). They also show that the photoperiodic responses in birds in general are mediated by circadian rhythm(s).

The photoperiodic entrainment and induction of reproductive rhythms in male blackheaded buntings (Emberiza melanocephala).

Groups of photosensitive, unstimulated or stimulated, male blackheaded buntings were subjected to photoregimes of 15 hr of green light of three intensities and 9 hr of dark per day. In some groups green light was interrupted with 90 min of bright fluorescent light at different times in the subjective day. While gonads did not develop or regressed in some groups, birds in others behaved as if exposed to long daylengths. The results besides suggesting the involvement of endogenous circadian rhythm during initiation and maintenance of gonadal growth indicate that the reproductive rhythms are entrained and induced by environmental photoperiod.

Participation of the circadian component in the photoperiodic mechanism of the blackheaded bunting, Emberiza melanocephala

Groups of photosensitive male blackheaded buntings ( Emberiza melanocephala) were subjected to continuous darkness (DD), short days (8L:16D), alternating DD and long days (15L:(D), and alternating 8L:16D and 15L:9D after photostimulation (15L:9D) for either 15 or 30 days. The effect of transfer to any of these regimes on the size of the testes was found to be dependent on the size attained at the time of transfer. However, the rate of testis regression, after 15 days or 30 days of 15L:9D, after transfer to DD, once begun, was the same as in the group transferred to 8L:16D. (Testicular regression did not begin immediately upon transfer to DD or 8L:16D, but, development continued for about 10 days in the birds photostimulated for 15 days.) Similarly, alternating long days could not maintain the testis size when they had attained their maximal size. A significant growth occurred in birds transferred after 15 days, in contrast to those transferred after 30 days of photostimulation. These observations are compatible with an external coincidence model, but could also be explained by an internal coincidence model. Application of the Law of Parsimony may favour an explanation that combines an external coincidence model and the established “carry-over” function of the neuroendocrine—gonadal system in photoperiodic birds.

Circadian Rhythmicity and the Termination of Photorefractoriness in the Black-Headed Bunting

Groups of photorefractory male Black-headed Buntings (Emberiza melanocephala), previously maintained on long daylengths (15L:9D), were exposed to photoperiods in which a 6-h light period was coupled with dark periods of 6, 18, 30, 42, or 54 h; a sixth control group was kept on 15L:9D. Cycles with lengths of 24 (6L:18D) and 48 h (6L:42D) terminated photorefractoriness, but cycles of 12 (6L:6D), 36 (6L:30D), and 60 h (6L:54D) did not; refractoriness persisted in the birds on 15L:9D. The results demonstrate that the termination and maintenance of the refractory period in Black-headed Buntings, a palearctic migratory passerine, are mediated by an endogenous circadian system of photoperiodic time measurement similar to that involved in the initiation and maintenance of gonadal growth, as is the case for several nearctic migratory species.

Biochronometry of photoperiodically induced fat deposition in a migratory finch, the Blackheaded bunting (Emberiza melanocephala) (Aves)

This investigation attempts to identify the mechanism(s) involved in the fat deposition in a photoperiodic migratory species, the Blackheaded bunting (Emberiza melanocephala). Groups of photosensitive male buntings were exposed to resonance, interrupted night, and ultra‐short‐day light cycles for 35, 42 and 75 days, respectively. Birds were weighed at the beginning and at the end of the experiments. Those exposed to ultra‐short‐day light cycles were also weighed at critical intervals during the treatment period. Our results indicate that: (a) a light‐sensitive rhythm with a period of about 24 hr is involved in the photoperiodic induction of premigratory fattening and weight gain in Blackheaded buntings: (b) buntings possess a bimodal pattern 'of sensitivity to photoperiods that induce fattening, and (c) this endogenous circadian rhythm can be entrained by an ultrashort photophase of 3 h if the latter is coupled with scotophases of specific duration.