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The mesotocinergic (MTergic) and dopaminergic (DAergic) systems have been documented to play pivotal roles in maternal behaviors in native Thai chickens. In native Thai chickens, plasma prolactin (PRL) concentrations are associated with maternal behaviors, which are also controlled by the DAergic system. However, the role of MT in conjunction with the roles of DA and PRL on the neuroendocrine regulation of the transition from incubating to rearing behavior has never been studied. Therefore, the aim of this study was to investigate the association of MT, DA, and PRL during the transition from incubating to rearing behavior in native Thai hens. Using an immunohistochemistry technique, the numbers of MT-immunoreactive (-ir) and tyrosine hydroxylase-ir (TH-ir, a DA marker) neurons were compared between incubating hens (INC; n = 6) and hens for which the incubated eggs were replaced with 3 newly hatched chicks for 3 days after 6, 10, and 14 days of incubation (REC; n = 6). Plasma PRL concentrations were determined by enzyme-linked immunosorbent assay. The results revealed that the numbers of MT-ir neurons within the nucleus supraopticus, pars ventralis (SOv), nucleus preopticus medialis (POM), and nucleus paraventricularis magnocellularis (PVN) increased in the REC hens when compared with those of the INC hens at 3 different time points (at days 9, 13, and 17). On the other hand, the number of TH-ir neurons in the nucleus intramedialis (nI) decreased in the REC13 and REC17 hens when compared with those of the INC hens. However, the number of TH-ir neurons in the nucleus mamillaris lateralis (ML) only decreased in the REC13 hens when compared with the INC13 hens. The decrease in the numbers of TH-ir neurons within the nI and ML is associated with the decrease in the levels of plasma PRL. This study suggests that the presence of either eggs or chicks is the key factor regulating the MTergic system within the SOv, POM, and PVN and the DAergic system within the nI and ML during the transition from incubating to rearing behavior in native Thai chickens. The results further indicate that these two systems play pivotal roles in the transition from incubating to rearing behavior in this equatorial species.

It is well documented that vasoactive intestinal peptide (VIP) is an avian prolactin-releasing factor, regulating the reproductive cycle, that initiates and maintains incubation behavior in avian species. Native Thai chicken has strong maternal behaviors. Thus, it is an excellent animal model to study this phenomenon. Changes in the numbers of VIP neurons within the nucleus inferioris hypothalami (IH) and nucleus infundibuli hypothalami (IN) are associated with maternal behaviors in the native Thai chicken. The objective of this study was to elucidate the role of VIP during the transition from egg incubation to rearing behavior in this species. The distributions of VIP-immunoreactive (-ir) neurons and fibers within the nucleus septalis lateralis (SL), nucleus anterior medialis hypothalami (AM), regio lateralis hypothalami (LHy), nucleus commissurae pallii (nCPa), nucleus ventromedialis hypothalami (VMN), and IH-IN were compared between incubating (INC) hens and replaced-egg-with-chicks (REC) hens, in which eggs were replaced by 3-day-old chicks after 6, 10, and 14 d of incubation period for 3 days. Using an immunohistochemistry technique, the results revealed that VIP-ir neurons and fibers were found within the SL, AM, LHy, nCPa, VMN, and IH-IN, with the greatest expression observed in the IH-IN in both groups. The number of VIP-ir neurons in the IH-IN was markedly decreased (P 0.05) in the REC hens when compared with those of the INC hens at 3 different time points (at d 9, 13, and 17). However, changes in the number of VIP-ir neurons between the INC and REC hens were not observed within other hypothalamic areas tested. These findings indicate that the presence of eggs and chicks affects the VIPergic system, and VIP neurons in the IH-IN might play a role in the transition from incubating to rearing behavior in the native Thai chicken.

Maternal care behaviors in birds include incubation and rearing behaviors. During incubating period, the hens stop laying and eating less due to food restriction as a natural fasting when compared with the rearing hens, resulting in low production of eggs and chicks. Neuropeptide Y (NPY), a neurotransmitter/neuromodulator, is very well known to be involved in food intake regulation in birds and mammals. The objective of this study is to elucidate the association between NPY and maternal behaviors in the female native Thai chicken. The distributions of NPY-immunoreactive (-ir) neurons and fibers in the brain of the incubating (INC), nest-deprived (ND), and replaced-egg-with-chicks (REC) hens at day 6 were determined utilizing immunohistochemistry technique. The results revealed that the distributions of NPY-ir neurons and fibers were observed within the septalis lateralis, nucleus rotundus, and nucleus dorsolateralis anterior thalami, with predominantly located within the the nucleus paraventricularis magnocellularis (PVN). NPY-ir fibers were located throughout the brain and the densest NPY-ir fibers were distributed in a discrete region lying close to the ventriculus tertius (third ventricle) through the hypothalamus. Changes in the number of NPY-ir neurons within the PVN of the INC, ND, and REC hens were compared at different time points (at days 6 and 14). Interestingly, the number of NPY-ir neurons within the PVN was significantly higher (P0.05) in the INC hens when compared with those of the ND and REC hens at day 14 but not day 6. In addition, the number of NPY-ir neurons within the PVN of the INC hens was significantly increased (P0.05) from day 6 to day 14 but not the ND and REC hens. These results indicated, for the first time, the asscociation between NPY and maternal behaviors in the femle native Thai chicken. Change in the number of NPY-ir neurons within the PVN during the transition from incubating to rearing behavior suggested the possible role of NPY in the regulation of the maternal behaviors in this equatorial species. In addition, the native Thai chicken might be an excellent animal model for the study of this phenomenon.

Avian prolactin (PRL) secretion is under stimulatory control by the PRL-releasing factor (PRF), vasoactive intestinal peptide (VIP). The neuroendocrine regulation of the avian reproductive system has been extensively studied in females. However, there are limited data in males. The aim of this study was to elucidate the VIPergic system and its relationship to PRL and testosterone (T) in the male native Thai chicken. The distributions of VIP-immunoreactive (-ir) neurons and fibers were determined by immunohistochemistry. Changes in VIP-ir neurons within the nucleus inferioris hypothalami (IH) and nucleus infundibuli hypothalami (IN) areas were compared across the reproductive stages. Plasma levels of PRL and T were determined by enzyme-linked immunosorbent assay and then compared across the reproductive stages. The results revealed that the highest accumulations of VIP-ir neurons were concentrated only within the IH-IN, and VIP-ir neurons were not detected within other hypothalamic nuclei. Within the IH-IN, VIP-ir neurons were low in premature and aging males and markedly increased in mature males. Changes in VIP-ir neurons within the IH-IN were directly mirrored with changes in PRL and T levels across the reproductive stages. These results suggested that VIP neurons in the IH-IN play a regulatory role in year-round reproductive activity in males. The present study also provides additional evidence that VIP is the PRF in non-seasonal, continuously breeding equatorial species.

Reproductive failure associated with aging is a well-known phenomenon. However, the mechanism by which this failure occurs in broiler breeder roosters is still unclear. A previous study conducted in our laboratory, comparing young and aging broiler breeder roosters, demonstrated an elevation in hypothalamic vasoactive intestinal peptide (VIP) and pituitary prolactin (PRL) gene expression accompanied by a deterioration of gonadal axis function. This resulted in a decrease in semen-quality variables as roosters aged. The objective of this study was to examine the involvement of the serotonergic axis in the age-associated reproductive failure in broiler breeder roosters. Cobb roosters aged 64 wk were divided into 3 groups (n = 20 each): parachlorophenylalanine (PCPA) administration, active immunization against chicken VIP, and controls. At 69 wk of age, each group was divided into 2 equal subgroups: 1 received ovine PRL and the other served as controls. Weekly semen volume, concentration and motility, and plasma testosterone, estradiol, and PRL concentrations were examined. At the end of the experiment, roosters were euthanized, testes were weighed, and hypothalamus and pituitary were removed to assay the expression of genes encoding hypothalamic GnRH-I, pituitary FSH, pituitary LH, hypothalamic VIP, and pituitary PRL. Both PCPA administration and active immunization against chicken VIP significantly increased testis weight, semen volume, sperm concentration, ejaculation grade, plasma testosterone level, and GnRH-I, FSH and LH gene expression compared with controls ( P ≤ 0.05). In addition, a decrease in plasma estradiol and PRL concentrations and VIP and PRL gene expression was observed in PCPA- and VIP-immunized birds compared with controls ( P ≤ 0.05). Administration of PRL in all groups decreased gonadal axis function and semen-quality variables ( P ≤ 0.05). Collectively, these results suggest that the increasing expression levels of the serotonergic axis in aging broiler breeder roosters inhibit proper gonadal function and reproductive performance. This article establishes for the first time the inhibitory role of serotonin on reproduction in aging roosters.

The dopaminergic (DAergic) system plays a pivotal role in incubation behavior via the regulation of prolactin (PRL) secretion in birds, however the role of the DA/PRL system in rearing behavior is poorly understood. The objective of this study was to investigate the relationship between the DA/PRL system and rearing behavior in a gallinaceous bird, the native Thai chicken. Incubating native Thai hens were divided into two groups. In the first group, hens were allowed to care for their chicks (rearing hens; R). In the second group, hens were deprived of their chicks immediately after hatching (non-rearing hens; NR). In both groups, blood samples and brain sections were collected at different time points after the chicks hatched (days 4, 7, 10, 14, 17, 21, 24, and 28; 6 hens/time point/group). In this study, tyrosine hydroxylase (TH) was used as a marker for DAergic neurons. The numbers of TH-immunoreactive (-ir) neurons in the nucleus intramedialis (nI) and in the nucleus mamillaris lateralis (ML), which regulate the vasoactive intestinal peptide (VIP)/PRL system, were determined in R and NR hens utilizing immunohistochemical techniques. Plasma PRL levels were determined by enzyme-linked immunosorbent assays. The results revealed that both the number of TH-ir neurons in the nI and the plasma PRL levels were significantly higher in the R hens compared with the NR hens during the first 14 days of chick rearing (P

Fertility of domestic roosters decreases at ∼50 wk of age. In a previous study on aging white leghorn roosters, low fertility was accompanied by low levels of both hypothalamic vasoactive intestinal peptide (VIP) and pituitary prolactin (PRL) mRNA expression; however, their role in aging broiler breeder rooster reproduction is still unclear. In this study we compared reproductive activities of young (35-wk-old) and aging (73-wk-old) broiler breeder roosters. Weekly semen volume; concentration and ejaculation grade; and concentrations of plasma testosterone, estradiol, and PRL were examined. Every other week, 10 roosters from each group were euthanized, their testes weighed, and hypothalamus and pituitary removed to determine mRNA expression of hypothalamic GnRH-I, pituitary FSH, pituitary LH, hypothalamic VIP, and pituitary PRL. Aging roosters had significantly lower testis weight and semen volume, sperm concentration, ejaculation grade and plasma testosterone and low hypothalamic GnRH-I, pituitary FSH, and pituitary LH mRNA expression than young roosters (P ≤ 0.05). Aging roosters had higher concentrations of plasma estradiol and PRL and higher hypothalamic VIP and pituitary PRL mRNA expression than young roosters (P ≤ 0.05). We suggest that PRL, which is known to inhibit the gonadal axis, and its releasing factor, VIP, play an important role in the reproductive failure associated with age in broiler breeder roosters.

Plasma leptin concentrations were investigated during the reproductive cycle in the native Thai chicken. The plasma leptin concentration was high during non-laying (0.69±0.15ng/ml), lowered to a minimum concentration during egg laying (0.07±0.02ng/ml), and gradually increased during egg incubation and rearing of the chicks (0.53±0.22 and 0.74±0.29ng/ml, respectively). However, the differences were not significant. Incubating chickens that were deprived of their nests for 3 weeks showed a significant decrease in plasma leptin concentrations (0.29±0.04ng/ml, P

Gonadotropin releasing hormone-I (GnRH-I) is known to regulate the avian reproductive system. We investigated the roles of GnRH-I in the regulation of the reproductive system of the native Thai chicken. The distribution of GnRH-I neurons and changes in GnRH-I-immunoreactive (-ir) neurons throughout the reproductive stages and between incubating and nest-deprived hens were analyzed utilizing immunohistochemical techniques. The results revealed that GnRH-I-ir neurons were distributed in a discrete region lying close to the third ventricle from the level of preoptic area through the anterior hypothalamus, with the greatest abundance found within the nucleus commissurae pallii (nCPa). The number of GnRH-I-ir neurons in the nCPa was highest in laying hens when compared with that in the other reproductive stages. Nest deprivation caused an increase in the number of GnRH-I-ir neurons in the nCPa of nest-deprived hens when compared with incubating hens. These results indicate that GnRH-I expression is correlated with the reproductive state in the native Thai chicken and may be, in part, regulated by it. This study also confirms a pivotal role of GnRH-I in controlling avian reproduction of this non-seasonal breeding, equatorial species.

It is well known that prolactin plays diverse roles in vertebrate reproduction. Besides expression in the pituitary, prolactin is also found in extra-pituitary tissues. In the present study, prolactin mRNA expression was studied utilizing in situ hybridization histochemistry. Prolactin mRNA, while found throughout the turkey brain, was predominantly localized within the pituitary, confirming a pivotal role of prolactin in turkey reproduction. The expression of prolactin mRNA was also observed within extra-pituitary brain areas including the cerebellum, nucleus accumbens, lateral septum, anterior hypothalamic nucleus, lateral hypothalamus, paraventricular nucleus, ventromedial nucleus, and infundibular nuclear complex. In the hypothalamus, an abundance of prolactin mRNA-expressing cells was observed in the anterior hypothalamic nucleus, lateral hypothalamus, and ventromedial nucleus. Cells expressing the least prolactin mRNA were found in the lateral septum, paraventricular nucleus, and the infundibular nuclear complex. This study reveals, for the first time, that prolactin mRNA was expressed in extra-pituitary brain areas in birds. In addition, the diverse expression of prolactin mRNA in the brain areas suggests that prolactin plays various physiological roles in birds.

Decreasing fertility in aging domestic roosters is a well-known phenomenon. Aging is manifested by a decrease in plasma testosterone level, testis function, and spermatogenesis, resulting in a low level of fertility. The roles of vasoactive intestinal peptide (VIP) and testicular inhibin in this aging process are not clear. The effects of active immunization against VIP, inhibin, or the combination of both hormones on the reproduction of aging White Leghorn (WL) roosters were assayed. In experiment 1a, 60 White Leghorn roosters (67 wk of age) were divided into 4 groups (n = 15/group). The first group was actively immunized against VIP, the second against inhibin, the third against VIP and inhibin, and the fourth served as a control. Active immunization against VIP decreased semen quality parameters, plasma steroid levels, and gene expression of gonadotropin-releasing hormone-I (GnRH-I), follicle-stimulating hormone (FSH), luteinizing hormone (LH), LH receptor, VIP, and prolactin (Prl). Immunization against inhibin increased some of the semen quality parameters and FSH mRNA gene expression but decreased inhibin gene expression. In experiment 1b, at 94 wk of age, we took the actively immunized against VIP group and the control group and divided them into 2 subgroups (n = 7 or 8): the first group was injected with 1 mg of ovine Prl (oPrl) daily for 7 d, and the second group served as a control. Administration of oPrl to previously VIP-immunized birds significantly elevated semen quality parameters. We suggest that VIP, Prl, and inhibin have an important effect on the reproductive axis in aging roosters. Active immunization against VIP-depressed reproductive activity and Prl administration restored their reproduction, indicating that both VIP and Prl are essential for reproduction in aging roosters. Immunization against inhibin improved FSH mRNA gene expression, suggesting a negative role of inhibin on FSH secretion in aging roosters. Not all semen quality parameters increased significantly after immunization against inhibin, even though FSH mRNA gene expression increased, suggesting interference in testicular function in aging roosters.

Hyperprolactinemia is associated with incubation behavior and ovarian regression in birds. To investigate the association of prolactin (PRL), vasoactive intestinal peptide (VIP), and dopamine (DA) with the neuroendocrine regulation of incubation behavior, changes in the number of visible VIP-immunoreactive (VIP-ir) neurons in the nucleus inferioris hypothalami (IH) and nucleus infundibuli hypothalami (IN) and tyrosine hydroxylase-immunoreactive (TH-ir) neurons in the nucleus intramedialis (nI) and nucleus mamillaris lateralis (ML) of incubating native Thai hens were compared with those of nest-deprived hens. TH was used as a marker for dopaminergic (DAergic) neurons. Blood samples were collected to determine PRL levels. The localization and the number of visible VIP-ir and TH-ir neurons were determined by immunohistochemistry. Disruption of incubation behavior was accompanied by a precipitous decline in plasma PRL levels. The number of visible VIP-ir neurons in the IH-IN and TH-ir neurons in the nI and ML were high during incubation and decreased when hens were deprived of their nests. This study indicated an association between VIP neurons in the IH-IN and DA neurons in the nI and ML with the degree of hyperprolactinemia, suggesting that the expression of incubation behavior in birds might be, in part, regulated by the DAergic input from the nI and ML to VIP neurons in the IH-IN and subsequent PRL release.

Native Thai chicken, an equatorial species breeds throughout the year, whereas turkeys are seasonal temperate zone breeder whose reproductive cycle is terminated by the onset of photorefractoriness. This study investigated VIPergic activity throughout a reproductive cycle in both species, hypothesizing that the differential expression of vasoactive intestinal peptide (VIP) would provide an insight into the differing reproductive strategies of the two species. Distribution of VIP neurons in the native Thai chicken and a comparison of VIPergic activity in the nucleus inferioris hypothalami (IH) and nucleus infundibuli hypothalami (IN) were investigated. VIP immunoreactivity was found throughout the native Thai chicken brain, predominantly located within the IH–IN. The pattern of VIP distribution in the native Thai chicken supports the findings reported in temperate zone species. Unlike the turkey, where there is a dissociation between VIPergic activity and prolactin levels during photorefractoriness, in the native Thai chicken, which do not express photorefractoriness, changes in VIP immunoreactive (VIP-ir) neurons within the IH–IN were directly correlated with prolactin throughout the reproductive cycle. VIPergic activity reached its lowest level after hatching of the chicks in the native Thai chicken, while in the turkey VIPergic activity was lowest only after exposure to a short day photoperiod and the acquisition of photosensitivity. This suggests that VIP neurons in the IH–IN may play a pivotal role in regulating the reproductive cycle and its differential expression following hatching of the young may, in part, account for the difference in reproductive mode between equatorial, continually breeding, non-photoperiodic birds and seasonally breeding, photoperiodic birds.

In mammals, prolactin (PRL) secretion is regulated by vasoactive intestinal peptide (VIP) and peptide histidine isoleucine (PHI). In birds, however, VIP is considered a PRL-releasing factor (PRF), while the role of PHI is unknown. The purpose of this study was to compare the effects of turkey PHI (tPHI) and turkey VIP (tVIP) on PRL secretion in vitro, and to study their physiological significance in vivo through active immunization against tPHI and tVIP. In vitro studies were conducted using pituitary cell cultures from female turkeys. In the in vivo study, female turkeys were immunized with keyhole limpet hemocyanin (KLH; control), synthetic tPHI conjugate (KLH-tPHI), or synthetic tVIP conjugate (KLH-tVIP). Both tVIP and tPHI stimulated PRL secretion from anterior pituitary cells in a dose response manner. However, tPHI was 100-fold less potent than tVIP in stimulating maximum PRL secretion in vitro. In addition, the highest dose (10(-4) M) of tPHI inhibited its own PRL-releasing activity as well as that of VIP-stimulated PRL release. Whereas, circulating PRL levels and nesting activity remained low and unchanged during the photo-induced reproductive cycle (i.e., experimental period) in tVIP-immunized birds, control and tPHI-immunized turkeys showed a significant increase in plasma PRL levels and in the incidence of incubation behavior over time following photostimulation. These findings, taken together with earlier results, indicate that VIP is the sole physiological PRF in the turkey (avian species).

Previous reports have suggested that green light enhances broiler growth at an early age, whereas blue light enhances growth at older ages. The aim of this study was to examine the effect of a switch in monochromatic light at 2 ages on growth and development of broilers. Male chicks (Anak, n = 640) were used. After hatch, chicks were weighed, wing-banded, and blocked into treatment groups. Chicks were grown in 1-m2 pens in 8 isolated light-proof rooms (20 birds/pen). The light treatments were (1) Control white (mini-incandescent lamps), 2) blue light-emitting diode (LED) lamps, 3) green LED lamps, 4) blue LED switching to green at 10 d of age, 5) blue LED switching to green at 20 d of age, 6) green LED switching to blue at 10 d of age, and 7) green LED switching to blue at 20 d of age. There were 8 pens for treatment 1, and 4 pens for each of the other treatments. The light schedule was 23L:1D, and intensity was 0.1 watts/m2. BW and feed consumption were recorded. Green light birds were significantly heavier at 4 d of age. Switching light at 10 d of age from green to blue caused a further increase in BW. This improved growth was maintained until the end of the experiment. Light switching from blue to green at 20 d of age also improved growth as compared with white light. Average feed efficiency and mortality rate did not differ between groups. No association was observed among light treatment, performance, and plasma triiodothyronine concentration. We suggest that green light stimulated growth of birds at early age, and shifting birds to a different light environment at 10 or 20 d of age might further stimulate growth.

Active immunization of turkey hens against vasoactive intestinal peptide (VIP) has been shown to inhibit incubation behavior and to increase egg production in second-cycle hens. The objective of this study was to compare the effect of VIP immunization on first- and second-cycle turkey hens during a 27-wk production period. First- (25-wk-old) and second- (54-wk-old) cycle hens were intermixed, distributed among 16 pens, and subjected to a photoperiod of 6 h of light and 18 h of darkness for 10 wk. The first-cycle hens were divided into two groups: keyhole limpet hemocyanin (KLH)-immunized controls (n = 16) and VIP-immunized (n = 18). Second-cycle hens were divided into four groups: 1) unimmunized controls (n = 19), 2) KLH-immunized controls (n = 18), 3) VIP-immunized (n = 19), and 4) VIP-preimmunized (immunized during first cycle; n = 16). Each hen received four antigen injections beginning the day of photostimulation (4-wk intervals), except for the preimmunized hens, which received three injections beginning 4 wk after photostimulation. The maximum titer of VIP antibodies in first-cycle, second-cycle, and preimmunized hens was 17.2+/-2.2, 20.9+/-2.9, and 21.7+/-3.2%, respectively. After photostimulation, plasma prolactin of first- and second-cycle control hens peaked between 484 +/-105 and 630+/-118 ng/mL. In contrast, prolactin changed very little in VIP-immunized turkeys. The average number of daily nest visits was less in first- and second-cycle VIP-immunized hens (1.68+/-0.23 and 1.09+/-0.15 visits per hen per day, respectively) than in their respective KLH-immunized controls (2.47+/-0.36 and 2.65+/-0.45 visits per hen per day). Expression of incubation behavior was 50.0 and 52.6% in first- and second-cycle control hens, respectively, upon termination of the study. In contrast, only 11.1% first-cycle and 5.2% second-cycle VIP-immunized turkeys exhibited the hormonal and behavioral characteristics of incubating hens. Average weekly egg production of first- and second-cycle VIP-immunized turkeys was similar (3.58+/-0.19 vs. 3.63+/-0.14 eggs per hen per wk). First- and second-cycle control hens laid 2.63+/-0.25 and 2.41+/-0.20 eggs per hen per wk, respectively. The present results show that comparable egg production was attained in first- and second-cycle hens by active immunization with VIP.

In order to ascertain if prolactin (PRL) secretion stimulated by serotonin (5-HT) was mediated by the dopaminergic system, both dopamine (DA) and 5-HT were infused into the third ventricle of anesthetized laying turkeys in conjunction with the D1 DA receptor antagonist R(+)-SCH-23390 HCl or the 5-HT receptor antagonist Mianserin HCl. When infused alone at the rate of 10 nmol/min, both DA and 5-HT increased circulating PRL levels significantly (P0. 05) after 20 min (111.0 +/- 19.0 to 334.0 +/- 115.1 ng/ml and 93.3 +/- 13.2 to 173.8 +/- 33.3 ng/ml, respectively). When infused in conjunction with the D1 DA antagonist, the stimulatory effect of both DA and 5-HT on PRL release was significantly suppressed. The 5-HT antagonist completely blocked 5-HT-stimulated PRL release when infused at the rate of 10 nmol/min. When the 5-HT antagonist was co-infused with DA, PRL levels significantly increased (33.5 +/- 3.3 to 112.2 +/- 19.7 ng/ml) after 30 min, an increase that did not differ significantly from that caused by DA alone (63.2 +/- 16.4 to 144.5 +/- 44.1 ng/ml). These data suggest that 5-HT stimulation of PRL secretion requires an intact dopaminergic system and that 5-HT synapses are located further upstream than DA synapses along the PRL-releasing pathway within the avian hypothalamus.