| Light is an important environmental factor for animal growth and development.Photoreceptors such as the retina receive light stimulations,convert them into nerve impulses on the optic nerve,and then regulate the superior cervical ganglion to release norepinephrine(NE)through the suprachiasmatic nucleus(SCN).NE binds to adrenergic receptors in the pineal gland and regulates the rhythm of Melatonin(MLT)synthesis and secretion,which in turn affects the secretion of hormones such as insulin and cortisol,and ultimately impacts animal growth,development and behavior.Broilers are light sensitive animals,which can receive light signals from the external environment through photoreceptors such as the retina,pineal gland,and hypothalamus.Different light modes will have a significant impact on the growth,metabolism,immunity,reproduction,intestinal flora structure,meat quality and behavior of broilers.In intensive production of broilers,artificial light supplementation is commonly used to promote the growth of broilers and consequently to improve their production performance.However,the mechanism of light on growth of broilers has not been deeply interpreted yet.Therefore,research endeavor to clearly reveal the effect of light on the growth of broilers is of great significance to improve the benefit and welfare of large-scale broiler production.Melatonin is a well-known mediator of light to regulate physiological function of animals.Objectives of this study were to explore the effect of different light modes on melatonin synthesis,and then the impact of generated melatonin on intestinal flora and behavior,as well as its mechanism on the growth of broilers.In this study,Arbor Acre(AA)broilers were selected to test the role of melatonin,an important light signal factor in broiler production.Three different light modes were set up,namely short light group(1L:23D),long light group(23L:1D)and normal light group(16L:8D).By measuring the concentration of serum melatonin and other bioactive substances,effect of photoperiod on biological rhythm of serum melatonin and other bioactive substances was presented,and influence of light on growth and behavior of broilers was discussed.Different neurotransmitters were added to pineal gland cells cultured in vitro,and the effect of neurotransmitters on the synthesis of melatonin in pineal gland cells was tested.By comparing the changes in the transcription levels of key enzymes for the synthesis of melatonin and other substances in the pineal cells cultured in vitro and those in broilers growing under different light modes,the stimulation of the transcription of key enzymes for melatonin synthesis by light through epinephrine was explained.Variations of intestinal flora structure and behavior of broilers grown under different light conditions were analyzed to clarify the mechanism on how the light affects intestinal flora and behavior of broilers through the action of melatonin,and then broilers growth.Major findings of this paper are as follows:1.No significant effect on the body weight gain of broilers was found among the three different photoperiods,however feed consumption of the short photoperiod group was higher and its feed-to-meat ratio significantly increased.Compared with the normal light group,the results of serum melatonin and other bioactive substances showed that the rhythm period of melatonin,norepinephrine and epinephrine was changed from 24 h to 12 h for the group under long-term dark condition.The amplitude of melatonin synthesis increased to 390 pg/mL under long dark conditions with a peak value of 450 pg/mL,while under long light conditions,the amplitude was only 160 pg/mL with a peak value of approximately 200 pg/mL.Rhythm cycles of melatonin,norepinephrine and epinephrine were highly correlated.The rhythm of norepinephrine was opposite to melatonin,and the rhythm of epinephrine was consistent with melatonin.The circadian cycles of dopamine,antidiuretic hormone,cortisol and thyrotropin had no statistical relationship with photoperiod changes,which were all 24 h in the three photoperiod patterns.The contents of growth hormone,superoxide dismutase and immunoglobulin G in serum had no clear rhythm.2.Through in vitro culture,different neurotransmitters were added to pineal cells,in which the melatonin synthesis rhythm was lost.The results showed that the neurotransmitters,such as y-aminobutyric acid,glycine,glutamic acid,histamine and acetylcholine,could not restore the melatonin synthesis rhythm in pineal cells.Meanwhile,epinephrine and norepinephrine could restore the rhythm of melatonin synthesis in pineal cells cultured in vitro,and the lowest concentrations of epinephrine and norepinephrine were 10-13 mol/L and 10-11 mol/L,respectively.Real Time-PCR test suggested that epinephrine significantly increased the transcription level of Arylalkylamine-N-acetyltransferase(AANAT)gene in cultured pineal cells,and the abundance of aanat mRNA increased 6 times after 2 h,and it reached 12 times12 h later.While for the treatment of norepinephrine,the abundance of aanat mRNA increased only 3 times.Related hormones in serum of broilers under different photoperiod conditions showed that the concentration of epinephrine reached the peak 2 hours earlier than melatonin.Real Time-PCR test indicated that the changes of transcription levels of key enzymes in melatonin synthesis in bright and dark stages agreed with those of epinephrine synthase,but opposite to norepinephrine.The results of this study firstly confirmed that epinephrine can directly stimulate the transcription of aanat,suggesting synergistic action of epinephrine and norepinephrine affects melatonin synthesis level within a shorter time.3.The change of intestinal microbial community structure will affect the nutrient absorption and consequently the feed-to-meat ratio of broilers.In this study,V3+V4 regions in the 16S rDNA gene of the small intestine contents under three photoperiod modes were sequenced for broilers.The results revealed that the abundance of Barnesiella in the small intestine of broilers in short light group was significantly higher than that of the other two experimental groups,with the content up to about 17%,which was 3-6 times higher than that of the other two groups.Meanwhile the abundance of Bacteroides and Alistipes bacteria was lower,accounting for less than 5%of the total bacteria content,which was about half of that of the other two groups.KEGG cluster analysis stated that longer light time for broilers increased the abundance of bacterial genes with functions of glucose metabolism and membrane transport in intestinal microorganisms.Content of insulin and glucagon in serum showed that the change of photoperiod affects the synthesis of melatonin,and then the synthesis rhythm of insulin and glucagon,which ultimately affects the synthesis of fat and feed-to-meat ratio as well.The serum insulin concentration of broilers in the long light group was the highest in the light period,reaching about 32 pg/mL,which was about 20%higher than that in the short light group and the normal light group.The serum glucagon concentration of broilers in the short light group was the highest in the dark period,up to 90 pg/ml,which was about 12%higher than that in the other two groups.Therefore,the fat synthesis ability of the long light group is higher than that of the short light group.The results of this study suggested that short-term light could change the intestinal flora structure of broilers through the rhythm of melatonin,reduce the number of bacteria beneficial to the absorption and utilization of polysaccharides,and lower insulin synthesis and fat synthesis function,which finally increase feed-to-meat ratio of broilers.4.Light can affect broiler behaviors by changing the related hormones through the action of melatonin.Reasonable light is conducive to free expression of the biological behavior of chickens and to improving its welfare.In this study,the behavior of chickens under three different photoperiod modes was monitored.The results indicated that the maintenance time of broilers in short light group,long light group and normal light group were 91.87%,94.54%and 92.98%,respectively.However,effectiveness of different photoperiods on feeding,drinking,walking,standing and nesting were varied.Longer cumulative time of feeding and drinking behavior of broilers was observed as illumination time increased,while the walking and standing behavior lasted the longest in the normal light group,and lying behavior was not significantly affected by photoperiod.Short photoperiod was beneficial to the expression of grooming behavior of broilers,but reduced the expression of stretching behavior.The shorter was of the illumination time,the shorter the cumulative time of abnormal behaviors was observed,such as attacking,scratching and pecking of broilers.It should be noted that average frequency and duration of broilers under the condition of short photoperiod were both reduced,but after a continuous low frequency feeding for 12 h,a high frequency feeding behavior for 2-4 h typically occurred,during which the average feeding time reached 60%.Correlation analysis between the behavior and hormone content in serum of broilers showed that the contents of melatonin,antidiuretic hormone and cortisol were correlated with the behavior to varying degrees.Under long light and normal light feeding conditions,melatonin was significantly negatively correlated with broiler feeding behavior(-0.867,-0.753),and cortisol was positively correlated with feeding behavior(0.682,0.741).Under short light feeding conditions,cortisol was negatively correlated with feeding behavior(-0.650),while which was positively correlated with its lying behavior(0.715).Experimental results indicated that the photoperiod affected the endocrine of broilers,changed their biological rhythms,and impacted their behavior and growth.The behavior of concentrated feeding for a short time under the condition of short photoperiod leaded to a decrease in feed utilization rate and an increase in feed-to-meat ratio.Based on the above research in this paper,long-term dark condition increased the synthesis of adrenaline,stimulated the transcription of aanat,the encoding genes of key enzyme of melatonin synthesis,and increased the synthesis of melatonin.The increase in melatonin synthesis leaded to a decrease in the number of bacteria in the intestines of broilers that were beneficial to the absorption,transport and utilization of polysaccharides,reducing the utilization of sugar in food,while reducing the synthesis of insulin and reducing the function of fat synthesis.On the other hand,the increase of melatonin synthesis caused by short-term light could reduce the content of cortisol,and the stress response,while increased the disorder of feeding behavior,resulting in the decrease of feed conversion rate.The influence of light on intestinal flora and behavior increased the feed-to-meat ratio under short light conditions.In this paper,effect of photoperiod bioactive substances for biological rhythm such as melatonin was analyzed.It is firstly confirmed that epinephrine could directly stimulate aanat transcription,suggesting the synergistic effect of epinephrine and norepinephrine can significantly affect melatonin synthesis.It is revealed that light affected the intestinal flora structure,insulin synthesis and behavior of broilers through melatonin,and finally impacted the nutrient absorption and feed-to-meat ratio of broilers.This paper provides a theoretical support for the establishment of broiler welfare breeding mode based on photoperiod regulation. |
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