Influence of altered photoperiods on serum melatonin and its receptors (MT1 and MT2) in the brain, retina, and ovary in carp Catla catla

Daily variation in melatonin receptor (MT1 and MT2) density in three specific tissues-brain, retina, and ovary-and its temporal relationship with serum melatonin were evaluated for the first time in a freshwater teleost, the carp Catla catla, under natural as well as altered photoperiods in different reproductive phases of the annual cycle. Cosinor analysis was used to determine rhythmic features of the serum melatonin and receptors (MT1 and MT2) in different tissues. In each photoperiodic group, irrespective of season, the daily minimum serum melatonin level was noted at midday. However, the daily peak value of melatonin varied in relation to both photo-schedules and reproductive phases. Under natural photoperiods (NPs; duration varied with seasons) and short photoperiods (SPs; light [L]:dark [D] 8:16), it occurred in the late dark phase during the preparatory phase, and at midnight in the remaining parts of the annual cycle. On the other hand, in each reproductive phase, compared to corresponding NP carp, the daily melatonin peak under long photoperiods (LPs; L:D 16:8) exhibited a phase delay of ～2-3?h (occurring during the late dark phase). The melatonin levels at each sampling point were highest during the postspawning phase and lowest during the spawning phase, irrespective of the photoperiodic history of the fish. In each tissue, Western blot analysis revealed a band at ～37?kDa and a band at ～36?kDa corresponding to the molecular weights of native MT1 and MT2 receptor proteins, respectively, with the band intensity of MT1 always being higher than that of a 36-kDa protein. The content of both melatonin receptor proteins varied significantly according to the studied tissue (being highest in the retina, intermediate in the brain, and lowest in the ovary), time in the daily cycle (peak at midnight and fall at midday), and reproductive phase in the annual cycle (highest in the spawning phase and lowest in the postspawning phase). Remarkably, no significant effects of altered photoperiod were detected on any rhythm parameters of either MT1 or MT2 in any of the studied tissues. Collectively, the results of the present study suggest a role of photoperiod in determining daily and seasonal profiles of serum melatonin, but not its receptor proteins, on the ovary or on any nongonad tissues in carp.     

The Endogenous Melatonin (MT) Signal Facilitates Reentrainment of the Circadian System to Light-Induced Phase Advances by Acting Upon MT2 Receptors

The indolamine melatonin is an important rhythmic endocrine signal in the circadian system. Exogenous melatonin can entrain circadian rhythms in physiology and behavior, but the role of endogenous melatonin and the two membrane-bound melatonin receptor types, MT1 and MT2, in reentrainment of daily rhythms to light-induced phase shifts is not understood. The present study analyzed locomotor activity rhythms and clock protein levels in the suprachiasmatic nuclei (SCN) of melatonin-deficient (C57BL/6J) and melatonin-proficient (C3H/HeN) mice, as well as in melatonin-proficient (C3H/HeN) mice with targeted deletion of the MT1, MT2, or both receptors, to determine effects associated with phase delays or phase advances of the light/dark (LD) cycle. In all mouse strains and genotypes, reentrainment of locomotor activity rhythms was significantly faster after a 6-h phase delay than a 6-h phase advance. Reentrainment after the phase advance was, however, significantly slower than in melatonin-deficient animals and in mice lacking functional MT2 receptors than melatonin-proficient animals with intact MT2 receptors. To investigate whether these behavioral differences coincide with differences in reentrainment of clock protein levels in the SCN, mPER1, mCRY1 immunoreactions were compared between control mice kept under the original LD cycle and killed at zeitgeber time 04 (ZT04) or at ZT10, respectively, and experimental mice subjected to a 6-h phase advance of the LD cycle and sacrificed at ZT10 on the third day after phase advance. This ZT corresponds to ZT04 of the original LD cycle. Under the original LD cycle, the numbers of mPER1- and mCRY1-immunoreactive cell nuclei were low at ZT04 and high at ZT10 in the SCN of all mouse strains and genotypes investigated. Notably, mouse strains with intact melatonin signaling and functional MT2 receptors showed a significant increase in the number of mPER1- and mCRY1-immunoreactive cell nuclei at the new ZT10 as compared to the former ZT04. These data suggest the endogenous melatonin signal facilitates reentrainment of the circadian system to phase advances on the level of the SCN molecular clockwork by acting upon MT2 receptors. (Author correspondence: M.Pfeffer@em.uni-frankfurt.de)     

Localization and dynamics of Mel1a melatonin receptor in the ovary of carp Catla catla in relation to serum melatonin levels

We studied the localization, sub-cellular distribution and daily rhythms of a 37 kDa melatonin receptor (Mel_1aR) in the ovary to assess its temporal relationship with the serum melatonin levels in four different reproductive phases in carp Catla catla. Our immunocytochemical study accompanied by Western blot analysis of Mel_1aR in the ovary revealed that the expression of this 37-kDa protein was greater in the membrane fraction than in the cytosol. Ovarian Mel_1aR protein peaked at midnight and fell at midday in each reproductive phase. Conversely, serum melatonin levels in the same fish demonstrated a minimum diurnal value at midday in all seasons, but a peak at midnight (during pre-spawning, spawning, and post-spawning phases) or at late dark phase (during preparatory phase). In an annual cycle, band intensity of Mel_1aR protein showed a maximum at night in the spawning phase and a minimum in the post-spawning phase, demonstrating an inverse relationship with the levels of serum melatonin. Our data provide first evidence of the presence of Mel_1a melatonin receptor in carp ovary and offer interesting perspectives especially for the study of the mechanisms of the control of its rhythmicity and its response to external factors.     

PHOTOPERIODIC REGULATION OF MT1 AND MT2 MELATONIN RECEPTOR EXPRESSION IN SPLEEN AND THYMUS OF A TROPICAL RODENT FUNAMBULUS PENNANTI DURING REPRODUCTIVELY ACTIVE AND INACTIVE PHASES

Photoperiodic regulation of melatonin receptor types on target tissues, such as lymphatic organs, has never been explored for any seasonal breeder. In the present study, we accessed the high affinity membrane melatonin receptors MT1 and MT2 expression dynamics in lymphoid organs (i.e., spleen and thymus) of a seasonally breeding rodent Funambulus pennanti during two major reproductive phases (i.e., active and inactive), when the internal hormonal (melatonin and gonadal steroid) as well as the ecological conditions were entirely different. Photoperiod regulates circulatory melatonin level; hence, we noted the effect of different photoperiodic regimes (long; 16L:8D and short; 10L:14D photoperiod) equivalent to summer and winter daylength on membrane melatonin receptor MT1 and MT2 expression in spleen and thymus. We have correlated the melatonin receptor expression with two major hormones varying seasonally (i.e., melatonin and testosterone) also being responsible for modulation of immunity of a seasonal breeder. Differential immunoreactivity of MT1 and MT2 receptor in spleen and thymus of F. pennanti suggests an involvement of both the receptor types in signal transduction of photoperiod for seasonal immunomodulation, because in the tropical zone, a slight difference (1:45–2?h) in daylength may change reproductive physiology and immunity of animals for adaptation. Our above suggestion receives strong support from the experiment of photoperiodic exposure on MT1 and MT2 expression at the translational level, where long daylength decreased the circulatory melatonin level and melatonin receptor expression in both lymphatic tissues. On the other hand, under short daylength, expression of MT1 and MT2 receptor increased in both spleen and thymus along with concomitant increase in circulatory melatonin level. Differential hormonal level of melatonin and gonadal hormones during reproductively active and inactive phase and its direct relation with melatonin receptor expression dynamics in lymphoid organs could be responsible for seasonal adjustment of immunity and reproduction. (Author correspondence: chaldar2001@yahoo.com)     

Diurnal and seasonal profiles of melatonin in the liver and their correlates of ovarian functions under natural photo-thermal conditions in adult carp Catla catla

Present study aimed to demonstrate daily rhythm features of hepatic melatonin concentrations in relation to ovarian functions during four reproductive phases of an annual cycle by measuring the levels of melatonin, 17-β estradiol (E2), vitellogenin (Vg) and maturation inducing hormone (MIH) in the liver and/or serum of adult carp Catla catla. Melatonin titres in liver, irrespective of reproductive phase, underwent daily variations with a peak in early dark phase and nadir at midday. However, the acrophase (Ø) of serum melatonin varied from late night in preparatory phase to midnight in the remaining parts of annual cycle. Their amplitude was highest during post-spawning phase and lowest during spawning phase. Hepatic E2 levels showed daily peak at midday and seasonal peak during pre-spawning phase. Though levels of serum Vg proteins and MIH did not exhibit daily variations, underwent seasonal changes with the highest and lowest values during spawning and post-spawning phases respectively. Hepatic melatonin titres always displayed significant negative correlation with the levels of both E2 and Vg. In essence, our study presented the first data on the daily and seasonal rhythm features of hepatic melatonin in carp and underlined their temporal relationship with the functions of ovary in any fish species.     

Photoperiodic regulation of annual testicular events in the Indian major carp Catla catla

The importance of photoperiods in the regulation of annual testicular events in the carp Catla catla was evaluated by subjecting them to either long (16 h light : 8 h dark) or short (8 h light : 16 h dark) photoperiods for 30 days during the preparatory, prespawning, spawning and postspawning phases of an annual gonadal cycle. In each reproductive phase, testicular responsiveness to subjected photoperiods was determined by comparing the gonadal status in corresponding groups of control or natural photoperiodic fish. The values of testicular weight, gametogenic index, as well as testicular activity of two steroidogenic enzymes (Δ⁵3β‐, and 17β‐hydroxysteroid dehydrogenase), and the serum titre of testosterone were considered as the indices of functional status of the testis in the fish concerned. During the prespawning phase, exposure of fish to a daily long photoperiod schedule resulted in precocious maturation of testis, while retardation of testicular growth was noted under the influences of short photoperiod. However, none of the employed photo‐schedules could influence the gametogenic and steroidogenic functions of the testis in the remaining part of the gonadal cycle. Collectively, the present study provides evidence for the first time that in the case of a commercially important carp, Catla catla, artificial, long photoperiods may be used for advanced testicular maturation, while reductions in maturation‐associated growth and deterioration in flesh quality may be avoided by submitting the fish to shorter day lengths during the prespawning phase of the reproductive cycle.     

Neural Regulation of Dark-Induced Abundance of Arylalkylamine N-Acetyltransferase (AANAT) and Melatonin in the Carp (Catla catla) Pineal: An In Vitro Study

In all the vertebrates, synthesis of melatonin and its rhythm-generating enzyme arylalkylamine N-acetyltransferase (AANAT) reaches its peak in the pineal during the night in a daily light-dark cycle, but the role of different neuronal signals in their regulation were unknown for any fish. Hence, the authors used specific agonist and antagonists of receptors for different neuronal signals and regulators of intracellular calcium (Ca²⁺) and adenosine 3',5'-cyclic monophosphate (cAMP) in vitro to study their effects on the abundance of AANAT and titer of melatonin in the carp (Catla catla) pineal. Western blot analysis followed by quantitative analysis of respective immunoblot data for AANAT protein, radioimmunoassay of melatonin, and spectrophotometric analysis of Ca²⁺ in the pineal revealed stimulatory effects of both adrenergic (α₁ and β₁) and dopaminergic (D₁) agonists and cholinergic (both nicotinic and muscarinic) antagonists, inhibition by both adrenergic and dopaminergic antagonists and cholinergic agonists, but independent of the influence of any agonists or antagonists of α₂-adrenergic receptors. Band intensity of AANAT and concentration of melatonin in the pineal were also enhanced by the intracellular calcium-releasing agent, activators of both calcium channel and adenylate cyclase, and phophodiesterase inhibitor, but suppressed by inhibitor of calcium channel and adenylate cyclase as well as activator of phophodiesterase. Moreover, an inhibitory effect of light on the pineal AANAT and melatonin was blocked by both cAMP and proteasomal proteolysis inhibitor MG132. Collectively, these data suggest that dark-induced abundance of AANAT and melatonin synthesis in the carp pineal are a multineuronal function, in which both adrenergic (α₁ and β₁, but not α₂) and dopaminergic signals are stimulatory, whereas cholinergic signals are inhibitory. This study also provides indications, though arguably not conclusive evidence, that in either case the neuronal mechanisms follow a signal-transduction pathway in which Ca²⁺ and cAMP may act as the intracellular messengers. It also appears that proteasomal proteolysis is a conserved event in the regulation of AANAT activity in vertebrates. (Author correspondence: dgp_skmaitra@yahoo.co.in)     

The endogenous melatonin (MT) signal facilitates reentrainment of the circadian system to light-induced phase advances by acting upon MT2 receptors

The indolamine melatonin is an important rhythmic endocrine signal in the circadian system. Exogenous melatonin can entrain circadian rhythms in physiology and behavior, but the role of endogenous melatonin and the two membrane-bound melatonin receptor types, MT1 and MT2, in reentrainment of daily rhythms to light-induced phase shifts is not understood. The present study analyzed locomotor activity rhythms and clock protein levels in the suprachiasmatic nuclei (SCN) of melatonin-deficient (C57BL/6J) and melatonin-proficient (C3H/HeN) mice, as well as in melatonin-proficient (C3H/HeN) mice with targeted deletion of the MT1, MT2, or both receptors, to determine effects associated with phase delays or phase advances of the light/dark (LD) cycle. In all mouse strains and genotypes, reentrainment of locomotor activity rhythms was significantly faster after a 6-h phase delay than a 6-h phase advance. Reentrainment after the phase advance was, however, significantly slower than in melatonin-deficient animals and in mice lacking functional MT2 receptors than melatonin-proficient animals with intact MT2 receptors. To investigate whether these behavioral differences coincide with differences in reentrainment of clock protein levels in the SCN, mPER1, mCRY1 immunoreactions were compared between control mice kept under the original LD cycle and killed at zeitgeber time 04 (ZT04) or at ZT10, respectively, and experimental mice subjected to a 6-h phase advance of the LD cycle and sacrificed at ZT10 on the third day after phase advance. This ZT corresponds to ZT04 of the original LD cycle. Under the original LD cycle, the numbers of mPER1- and mCRY1-immunoreactive cell nuclei were low at ZT04 and high at ZT10 in the SCN of all mouse strains and genotypes investigated. Notably, mouse strains with intact melatonin signaling and functional MT2 receptors showed a significant increase in the number of mPER1- and mCRY1-immunoreactive cell nuclei at the new ZT10 as compared to the former ZT04. These data suggest the endogenous melatonin signal facilitates reentrainment of the circadian system to phase advances on the level of the SCN molecular clockwork by acting upon MT2 receptors.     

Importance of photoperiods in the regulation of ovarian activities in indian major carp Catla catla in an annual cycle

The present study attempted for the first time to explore the importance of photoperiod in the regulation of seasonal ovarian functions in any subtropical major carp. Adult Indian major carp Catla catla were transferred to a long photoperiod (LP; LD 16:8) or a short photoperiod (SP; LD 8:16) for 30 days on 4 dates corresponding to the beginnings of 4 reproductive phases in an annual cycle, and responsiveness of the ovary was evaluated by comparison with the gonadal weight (I(G)), relative number of developing oocytes, serum levels of vitellogenin, and the activity of 2 important steroidogenic enzymes, that is, Delta(5)3beta-hydroxysteroid dehydrogenase and 17.beta-hydroxysteroid dehydrogenase, in the ovary of fish in a natural photoperiod. Exposure of fish to LP during the preparatory phase (February-March) resulted in a significant (p < 0.001) increase in the values of vitellogenin and in the activity of both the steroidogenic enzymes but not in the ovarian weight and in the relative number of different stages of oocytes. A more stimulatory influence of LP was noted during the prespawning phase (April-May), when precocious maturation of ovary was evident from a significant (p < 0.001) rise in the values of each studied features of ovarian functions. However, no ovarian response was found when the fish were transferred to LP during the spawning (July-August) and the postspawning (September-October) phases. On the other hand, the SP was found to have an inhibitory influence on ovarian growth and maturation during the prespawning and the spawning phases or to have no influences on ovarian functions during the preparatory and the postspawning phases of an annual cycle. The results of our study provide the first evidence that photoperiod per se plays an important role in the seasonal maturation of ovary in a subtropical freshwater major carp.     

Melatonin rhythms in the pineal and non-pineal tissues and their physiological implications in subtropical fish

Abstract

Melatonin (N-acetyl-5-methoxy tryptamine), following discovery from the extracts of bovine pineal gland, has been detected in the pineal as well as several extra-pineal tissues/organs of different vertebrates including fish. The unique feature of melatonin in the pineal gland is its rhythmic biosynthesis and release in blood in synchronization with the environmental light-–dark cycle. Accordingly, melatonin produced in the pineal of an animal living in a changing environment is implicated to the regulation of seasonal reproduction by acting as a hormone at one or more levels of hypothalamo-hypophyseal-gonadal axis. Additionally, melatonin is known to act as a potent free-radical scavenger or antioxidant to influence maturation of oocytes. However, possible relationship between extra-pineal melatonin and seasonality of reproduction in any animal remains enigmatic. Perhaps, carp is the only known animal in which temporal patterns of melatonin levels in the serum as well as in the extracts of pineal, retina, ovary, gut, and liver have been studied in relation to the reproductive events in an annual cycle. The purpose of current review is to bring those fascinating, and arguably most important data together to underline their significance in the control of seasonal reproduction in subtropical fish in general and in carp in particular.     

Effects of melatonin and antagonists of MT1 and MT2 receptors on somatic pain induced within the fixed circadian rhythm

We studied behavioral pain-related reactions (PRRs) induced in mice by subcutaneous injections of 5% formalin within different phases of the fixed circadian illumination rhythm under conditions of administration of exogenous melatonin and of blocking of MT1 and MT2 melatonin receptors. It was demonstrated that modulation of experimentally induced somatic pain depends considerably on the phase of the preset circadian rhythm. In the norm, the duration of PRRs in the middle of the dark phase was 30% smaller than that in the middle of the light phase. Administration of exogenous melatonin in the middle of the light phase decreased the duration of episodes of noxious behavior by 43%, on average. Injections of melatonin within the dark phase resulted in no significant changes in the duration of PRRs. In the dark phase, the blockade of MT1 receptors by luzindole led to an increase in the duration of PRRs by 45%, as compared with the norm, while in the light phase we observed no significant alterations of this duration under conditions of blocking of the above-mentioned receptors. The blockade of MT2 receptors by prazocine in the middle of dark and light phases increased the durations of PRRs by 92 and 28%, respectively. Our data indicate that the analgesic effect of melatonin depends significantly on the level of this hormone in the organism; in turn, such a level is determined by the illumination conditions. The antinoxious effect of melatonin is mediated by MT receptors, in particular by MT2 receptors. Neirofiziologiya/Neurophysiology, Vol. 39, No. 3, pp. 255–259, May–June, 2007.     

The expression of MT1 and MT2 melatonin receptor mRNA in several rat tissues

The mechanisms that mediate the various effects of melatonin in mammalian tissues are not always known. Therefore, the aim of this study was to investigate whether MT(1) and MT(2) melatonin receptors are expressed in certain tissues of the rat. The expression of MT(1) and MT(2) melatonin receptor mRNA was determined using a real-time quantitative RT-PCR method. In addition, we examined whether mRNA for either subtype of receptor shows any difference in the expression between midnight and noon, similar to the changes in melatonin concentrations in plasma and tissue samples. MT(1) and MT(2) melatonin receptor mRNAs were found in the rat hypothalamus, retina and small intestine. We also showed a low expression of MT(2) mRNA in the rat liver and heart SA node. In the heart apex and the Harderian gland, no appearance of either of the receptor mRNAs was detectable. A significant difference in the expression of MT(1) mRNA between day and night was found in the hypothalamus. In conclusion, our findings suggest that at least some effects of melatonin are mediated through membrane MT(1) and MT(2) receptors in the hypothalamus, the retina and the small intestine. Down-regulation of receptors might be one reason for the difference in the hypothalamic MT(1) melatonin receptor mRNA expression between midnight and noon. In the liver and the heart SA node, the physiological significance of possible MT(2) receptors remains unclear. According to our negative midnight and noon results in the Harderian gland and heart apex melatonin may exert its effect on these tissues by a non-receptor mechanism.     

Do thyroid and testis modulate the effects of pineal and melatonin on haemopoietic variables inClarias batrachus?

Involvement of pineal and its major hormone, melatonin, in the process of erythropoiesis in a freshwater catfish,Clarias batrachus has been investigated. The study was conducted during four phases, namely preparatory phase, spawning phase, postspawning phase and late postspawning phase of its annual reproductive cycle. During each phase a fish received either melatonin injections or subjected to pinealectomy. In addition, each fish in all the groups, received either iopanoic acid or cyproterone acetate or vehicle in the morning or late afternoon. Results clearly indicate that melatonin stimulates the rate of erythropoiesis inClarias batrachus. It appears that the extent of stimulation depends upon the phase of the annual reproductive cycle. However, in general, the pineal-or melatonin-induced modulation of blood variables is gonad dependent and thyroid seems to play a time of the day dependent subtle role     

Pineal and cortical melatonin receptors MT1 and MT2 are decreased in Alzheimer's disease

The pineal hormone melatonin is involved in physiological transduction of temporal information from the light dark cycle to circadian and seasonal behavioural rhythms, as well as possessing neuroprotective properties. Melatonin and its receptors MT1 and MT2, which belong to the family of G protein-coupled receptors, are impaired in Alzheimer's disease (AD) with severe consequences to neuropathology and clinical symptoms. The present data provides the first immunohistochemical evidence for the cellular localization of the both melatonin receptors in the human pineal gland and occipital cortex, and demonstrates their alterations in AD. We localized MT1 and MT2 in the pineal gland and occipital cortex of 7 elderly controls and 11 AD patients using immunohistochemistry with peroxidase-staining. In the pineal gland both MT1 and MT2 were localized to pinealocytes, whereas in the cortex both receptors were expressed in some pyramidal and non-pyramidal cells. In patients with AD, parallel to degenerative tissue changes, there was an overall decrease in the intensity of receptors in both brain regions. In line with our previous findings, melatonin receptor expression in AD is impaired in two additional brain areas, and may contribute to disease pathology.     

Activation of MT(2) melatonin receptors in rat suprachiasmatic nucleus phase advances the circadian clock

The aim of this study was to identify the melatonin receptor type(s) (MT(1) or MT(2)) mediating circadian clock resetting by melatonin in the mammalian suprachiasmatic nucleus (SCN). Quantitative receptor autoradiography with 2-[(125)I]iodomelatonin and in situ hybridization histochemistry, with either (33)P- or digoxigenin-labeled antisense MT(1) and MT(2) melatonin receptor mRNA oligonucleotide probes, revealed specific expression of both melatonin receptor types in the SCN of inbred Long-Evans rats. The melatonin receptor type mediating phase advances of the circadian rhythm of neuronal firing rate in the SCN slice was assessed using competitive melatonin receptor antagonists, the MT(1)/MT(2) nonselective luzindole and the MT(2)-selective 4-phenyl-2-propionamidotetraline (4P-PDOT). Luzindole and 4P-PDOT (1 nM-1 microM) did not affect circadian phase on their own; however, they blocked both the phase advances (approximately 4 h) in the neuronal firing rate induced by melatonin (3 pM) at temporally distinct times of day [i.e., subjective dusk, circadian time (CT) 10; and dawn, CT 23], as well as the associated increases in protein kinase C activity. We conclude that melatonin mediates phase advances of the SCN circadian clock at both dusk and dawn via activation of MT(2) melatonin receptor signaling.     

Photoperiod, melatonin secretion, and sexual maturation in a tropical rodent

Descendants of a sample of cane mice (Zygodontomys brevicauda) trapped at 8 degrees latitude in Venezuela were tested for reproductive photoresponsiveness. This species breeds continuously, year around, despite living in a seasonally harsh habitat. At 50 days of age there were no differences in the weights of the testes or seminal vesicles or in sperm counts of males born and reared on 16L:8D, 13L:11D, 11L:13D, or 8L:16D photoperiods, although there were small differences in body weight. Females born and reared on 16L:8D vs. 8L:16D cycles became pregnant at the same rates and ages when paired with males at 21 or 31 days of age. The daily duration of melatonin secretion depended on the length of the dark phase of the cycle in both sexes. Circulating levels of melatonin were elevated for 8 h on a 16L:8D cycle and for between 9 and 16 h on an 8L:16D cycle. In this tropical species, the neuroendocrine pathway that links photoperiod to reproduction apparently is disconnected somewhere between melatonin and gonadotropin secretion, causing cane mice to be reproductively unresponsive to variation in photoperiod.     

The Avian Pineal Gland

The pineal gland plays a key role in the control of the daily and seasonal rhythms in most vertebrate species. In mammals, rhythmic melatonin (MT) release from the pineal gland is controlled by the suprachiasmatic nucleus via the sympathetic nervous system. In most non‐mammalian species, including birds, the pineal gland contains a self‐sustained circadian oscillator and several input channels to synchronize the clock, including direct light sensitivity. Avian pineal glands maintain rhythmic activity for days under in vitro conditions. Several physical (light, temperature, and magnetic field) and biochemical (Vasoactive intestinal polypeptide (VIP), norepinephrine, PACAP, etc.) input channels, influencing release of melatonin are also functional in vitro, rendering the explanted avian pineal an excellent model to study the circadian biological clock. Using a perifusion system, we here report that the phase of the circadian melatonin rhythm of the explanted chicken pineal gland can be entrained easily to photoperiods whose length approximates 24 h, even if the light period is extremely short, i.e., 3L:21D. When the length of the photoperiod significantly differs from 24 h, the endogenous MT rhythm becomes distorted and does not follow the light‐dark cycle. When explanted chicken pineal fragments were exposed to various drugs targeting specific components of intracellular signal transduction cascades, only those affecting the cAMP‐protein kinase‐A system modified the MT release temporarily without phase‐shifting the rhythm in MT release. The potential role of cGMP remains to be investigated.     

Melatonin in the Regulation of Annual Testicular Events in Carp Catla catla: Evidence from the Studies on the Effects of Exogenous Melatonin,Continuous Light,and Continuous Darkness

The physiological significance of melatonin in the regulation of annual testicular events in a major carp Catla catla was evaluated through studies on the effects of graded dose (25, 50, or 100 µg/100 g body wt.) of melatonin exogenously administered for different durations (1, 15, or 30 days) and manipulation of the endogenous melatonin system by exposing the fish to constant darkness (DD) or constant light (LL) for 30 days. An identical experimental schedule was followed during the preparatory (February–March), pre‐spawning (April–May), spawning (July–August), and post‐spawning (September–October) phases of the annual cycle. Irrespective of the reproductive status of the carp, LL suppressed while DD increased the mid‐day and mid‐night values of melatonin compared to respective controls. Influences of exogenous melatonin varied in relation to the dose and duration of treatment and the reproductive status of the carp. However, testicular response to exogenous melatonin (at 100 µg, for 30 days) and DD in each reproductive phase was almost identical. Notably, precocious testicular maturation occurred in both DD and melatonin‐injected fish during the preparatory phase and in LL carps during the pre‐spawning phase. In contrast, testicular functions in both the melatonin‐treated and DD fish were inhibited during the pre–spawning and spawning phases, while the testes did not respond to any treatment during the post‐spawning phase. In conclusion, this study provided the first experimental evidence that melatonin plays a significant role in the regulation of annual testicular events in a sub‐tropical surface‐dwelling carp Catla catla, but the influence of this pineal hormone on the seasonal activity of testis varies in relation to the reproductive status of the concerned fish.     

Melatonin Does Not Modulate Testicular Responsiveness to Altered Photoperiods. A Study During Different Phases of the Annual Gonadal Cycle in Roseringed Parakeets (Psittacula krameri)

The roseringed parakeet has been shown to exhibit a variable testicular responsiveness to both altered photoperiodic regimens and to treatment with melatonin during different phases of the annual gonadal cycle. Adult male roseringed parakeets were held under either natural photoperiods (NP), or long photoperiods (LP; 16L 8D), or short photoperiods (SP; 8L 16D) for a total period of 90 days. From day 46 onward, half of the total birds in each group were administered with the vehicle of melatonin, and the other birds were injected daily in the afternoon with melatonin (25 µg/ 100 g body wt.) till the end of the experiment. An identical experimental schedule was followed during the four different (preparatory, progressive, pre-breeding, and breeding) phases of the annual testicular cycle. The testicular activities in various bird groups were evaluated by volumetric, gravimetric, histometric and karyometric measurements, and by quantitative histological studies. The findings revealed that exogenous melatonin may exert either a suppressive influence or none at all on the testicular functions in relation to the photoperiodic schedule as well as to the reproductive phase of the concerned bird, but in no case modulates gonadal responsiveness to artificially altered photoperiods.