Distribution of hypothalamic vasoactive intestinal peptide immunoreactive neurons in the male native Thai chicken.

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.

Mesotocin and maternal care of chicks in native Thai hens (Gallus domesticus).

Oxytocin (OT) is known to induce and regulate maternal behaviors in mammals via the supraoptic nucleus and paraventricular nucleus (PVN), whereas the function of mesotocin (MT; the avian homolog of OT) is poorly understood in birds. To elucidate the association of MT and the regulation of maternal behaviors in birds, we studied changes in the number of MT-immunoreactive (ir) neurons in native Thai chickens using immunohistochemistry. We observed that MT-ir neurons and fibers appeared in discrete regions located close to the third ventricle from the level of the preoptic area through the anterior hypothalamus with an abundance observed in the nucleus supraopticus, pars ventralis (SOv), nucleus preopticus medialis (POM), and PVN. The number of MT-ir neurons was low in the SOv, POM, and PVN of non-laying hens, but it increased gradually when the hens entered the laying stage, and peaked in incubating and rearing hens. We compared the number of MT-ir neurons in the SOv, POM, and PVN of native Thai hens rearing chicks (R) with that of non-rearing chicks (NR). The number of MT-ir neurons was high in the R hens, but low in the NR hens in these nuclei. For the first time, these results indicate that the association between the MT neurons and the presence of chicks might, in part, play a role in the neuroendocrine reorganization to establish and maintain maternal behaviors in native Thai chickens. MTergic activity is likely related to the contribution of rearing behavior in this equatorial precocial species.

Gonadotropin releasing hormone and brooding behavior in the native Thai hen.

Changes in the number of hypothalamic gonadotropin releasing hormone-I (GnRH-I) neurons within the Nucleus commissurae pallii (nCPa) were associated with the reproductive cycle of native Thai chickens. In order to further understand the association of GnRH-I in the regulation of brooding behavior in this bird, the native Thai chickens were divided into two groups; chick-rearing (R) and non-chick-rearing (NR) hens. Numbers of visible of GnRH-I-immunoreactive (GnRH-I-ir) neurons in the hypothalamus of R and NR hens were compared utilizing immunohistochemistry. Numbers of visible GnRH-I-ir neurons within the Nucleus anterior medialis hypothalami, Nucleus suprachaiasmaticus, pars medialis, Nucleus septalis lateralis, Nucleus paraventricularis magnocellularis, and Regio lateralis hypothalami areas were observed in both groups, but no differences were seen between R and NR hens. The number of visible GnRH-I neurons in the nCPa was higher (P<0.05) in the NR than in R hens, and increased in NR hens by day 14 after chick removal. These findings suggest, for the first time, an association of the GnRH system with brooding behavior in continuously breeding birds. Furthermore, the expression of brooding behavior of native Thai chickens might be regulated, in part, by GnRH-I neurons in the nCPa.

Neuroendocrine regulation of rearing behavior in the native Thai hen.

Vasoactive intestinal peptide (VIP) is the avian prolactin releasing factor and changes in the concentrations of plasma prolactin (PRL) are found during the avian reproductive cycle. This study investigated the changes in the VIP/PRL system of native Thai hens rearing their young as compared to hens deprived of rearing their chicks. The number of VIP-immunoreactive (VIP-ir) neurons in the Nucleus inferioris hypothalami (IH) and Nucleus infundibuli hypothalami (IN) of hens rearing chicks (R) were compared with those of non-rearing chicks (NR). Plasma PRL levels were determined by enzyme-linked immunosorbent assay. The localization and number of VIP-ir neurons were determined by immunohistochemistry. The numbers of VIP-ir neurons in the IH-IN areas were high in the R hens, whereas the number of VIP-ir neurons decreased in the NR hens as compared to their respective R hens. During the rearing period, changes in the VIP-ir neurons within the IH-IN were correlated with plasma PRL levels. The results of the present study indicate for the first time that the VIP/PRL system plays a role in neuroendocrine reorganization to establish maternal behavior in native Thai chickens. The VIP/PRL system functions not only as a well established key regulator of incubation behavior, but is also involved in the regulation of rearing behavior. It is possible that VIP and the decline in the number of VIP-ir neurons and in turn VIPergic activity and the decrease in PRL levels are related to their contribution to rearing behavior of this non-seasonal breeding, equatorial precocial species.

Plasma leptin concentrations during the reproductive cycle in the native Thai chicken (Gallus domesticus).

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<0.05) compared to those of their corresponding incubating controls (0.77±0.08ng/ml). Similarly, plasma leptin concentration of chickens that were deprived of their chicks for 4 weeks was significantly lower (0.09±0.11ng/ml, P<0.05), when compared to those of chickens that rearing their chicks (0.71±0.18ng/ml). These findings taken together with the results that the low plasma leptin concentrations were observed in chickens having relatively greater ovary and oviduct weights led to the suggestion that circulating leptin concentrations are associated with the reproductive states of the birds, especially the ovarian activity (i.e. ovarian steroid hormone concentrations) in the native Thai chicken, a tropical and continuous breeding species.

Identification of cis elements necessary for glucocorticoid induction of growth hormone gene expression in chicken embryonic pituitary cells.

Glucocorticoid (GC) treatment of rat or chicken embryonic pituitary (CEP) cells induces premature production of growth hormone (GH). GC induction of the GH gene requires ongoing protein synthesis, and the GH genes lack a canonical GC response element (GRE). To characterize cis-acting elements and identify trans-acting proteins involved in this process, we characterized the regulation of a luciferase reporter containing a fragment of the chicken GH gene (-1727/+48) in embryonic day 11 CEP cells. Corticosterone (Cort) increased luciferase activity and mRNA expression, and mRNA induction was blocked by protein synthesis inhibition. Through deletion analysis, we identified a GC-responsive region (GCRR) at -1045 to -954. The GCRR includes an ETS-1 binding site and a degenerate GRE (dGRE) half site. Nuclear proteins, including ETS-1, bound to a GCRR probe in electrophoretic mobility shift assays, and Cort regulated protein binding. Using chromatin immunoprecipitation, we found that ETS-1 and GC receptor (GR) were associated with the GCRR in CEP cells, and Cort increased GR recruitment to the GCRR. Mutation of the ETS-1 site or dGRE site in the -1045/+48 GH reporter abolished Cort responsiveness. We conclude that GC regulation of the GH gene during development requires cis-acting elements in the GCRR and involves ETS-1 and GR binding to these elements. Similar ETS-1 elements/dGREs are located in the 5'-flanking regions of GH genes in mammals, including rodents and humans. This is the first study to demonstrate involvement of ETS-1 in GC regulation of the GH gene during embryonic development in any species, enhancing our understanding of GH regulation in vertebrates.

Distribution and variation in gonadotropin releasing hormone-I (GnRH-I) immunoreactive neurons in the brain of the native Thai chicken during the reproductive cycle.

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.

Presence of prolactin mRNA in extra-pituitary brain areas in the domestic turkey.

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.

Changes in vasoactive intestinal peptide and tyrosine hydroxylase immunoreactivity in the brain of nest-deprived native Thai hen.

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.

The dopaminergic system in the brain of the native Thai chicken, Gallus domesticus: localization and differential expression across the reproductive cycle.

Dopamine (DA) has a pivotal role in avian prolactin (PRL) secretion, acting centrally through D(1) DA receptors to stimulate PRL secretion by operating through vasoactive intestinal peptide (VIP). DA also inhibits PRL secretion by activating D(2) DA receptors at the pituitary level. This study was designed to investigate the distribution of DA neurons in the native Thai chicken, utilizing tyrosine hydroxylase (TH) as a marker for dopaminergic neurons. The differential expression of hypothalamic TH immunoreactive (TH-ir) neurons was also compared across the reproductive cycle. The results revealed that TH-ir neurons and fibers were found throughout the brain of the laying hen and were predominantly located within the diencephalon and mesencephalon. The observed distribution pattern of TH immunoreactivity was consistent with that reported previously in several avian species. However, changes in the number of TH-ir neurons in the nucleus intramedialis (nI) were observed across the reproductive cycle and correlated directly with variations in PRL levels. The population of TH-ir neurons in the nI increased significantly during the egg incubation period, where circulating PRL levels were the greatest. This study indicates, for the first time, that an association exists between DA neurons and the regulation of the reproductive system in the native Thai chicken. There is a paucity of information about the reproductive neuroendocrine regulation of tropical non-seasonally breeding avian species and it is suggested that the differential expression of DA neurons in the nI might play a role in the control of VIP secretion and subsequent PRL release in such birds.

Vasoactive intestinal peptide and its role in continuous and seasonal reproduction in birds.

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.