Chronology of events in the first cell cycle of the polyspermic egg of the domestic fowl (Gallus domesticus).

The nuclear population in the polyspermic egg of the domestic hen was examined in whole-mount preparations of the germinal disc. The numbers of nuclei varied in groups of hens from averages of 5.9 to 26, depending on days from insemination. Changes in development from initial formation of pronuclei to the early mitoses of the zygote nucleus were staged according to the position of the egg in the oviduct. The findings substantiated earlier accounts on the timing of the apposition of the parental pronuclei towards the end of the first cell cycle. Additionally, analysis of the spatial distribution of accessory spermatozoal nuclei showed a slight, but significant, dispersal from a clustered arrangement at this time.

[14C]2-deoxyglucose uptake in the brain of the ring dove (Streptopelia risoria). I. Prolactin-induced uptake.

Quantitative [14C]2-deoxyglucose (2DG) autoradiography was used to identify areas of the ring dove brain involved in the expression of incubation behavior. Compared with non-breeding controls, 2DG uptake was increased in birds of both sexes during late incubation in all areas of the fore, mid- and hind-brain examined. This increase occurred irrespective of whether the birds were sitting on their eggs at the time of 2DG administration. A similar pattern of 2DG uptake into the brain was observed in non-breeding females treated with 30 I.U. ovine prolactin (i.p.) twice daily for 5 days. It is concluded that there is a generalised increase in neural activity in the brain of doves during late incubation which may be dependent on increased concentrations of plasma prolactin.

Projections of ankle joint afferents to the spinal cord and brainstem of the chicken (Gallus g. domesticus).

The projections of the ankle joint capsule afferents were studied by transganglionic transport of horseradish peroxidase injected directly into the ankle joint. The number and size of the labelled dorsal root ganglion cells were measured from synsacral nerves 2-9. In the dorsal root ganglia, all sizes of sensory neurones were labelled, and the largest number of labelled cells was in ganglia 5-7. The extensive sympathetic innervation of the ankle joint was identified by the large number of cell bodies labelled in the sympathetic ganglia of the paravertebral chain. Labelled afferent fibres projected to the spinal cord from the 2nd to the 8th synsacral nerves, with the rostral projection mainly via Lissauer's tract and the dorsal funiculus. Terminal labelling in the dorsal horn was identified in laminae I-III and VI, with a slight projection to V. Two areas of dense labelling, which did not correspond with the largest number of labelled dorsal root ganglion cells, were identified. A rostral area with the highest density of label was observed at the level of synsacral nerves 3-4 and a second slightly less dense area between synsacral nerves 7-8. In the caudal medulla, diffuse terminal labelling was observed in the nucleus gracilis et cuneatus, nucleus of the tractus solitarius, and the nucleus cuneatus externus. These results are discussed in a comparative context to identify similarities and differences between different primary afferent projections in birds and mammals and to highlight the possible functional significance of the avian articular afferent projection.

Pituitary prolactin messenger ribonucleic acid levels in incubating and laying hens: effects of manipulating plasma levels of vasoactive intestinal polypeptide.

Pituitary PRL messenger RNA levels in hens, measured by dot-blot hybridization, correlated directly with concentrations of plasma PRL, being 3-fold higher in incubating than in laying birds. Nest deprivation of incubating hens for 24 h caused a rapid decrease in both plasma PRL and pituitary PRL mRNA, which remained depressed thereafter. A single injection of vasoactive intestinal polypeptide (VIP) in laying hens resulted in an increase (P less than 0.05) in pituitary PRL mRNA whereas passive immunoneutralization of VIP in incubating hens resulted in a decrease (P less than 0.001) in pituitary PRL mRNA. The rapid decrease in pituitary PRL mRNA after nest deprivation or passive immunoneutralization of VIP was associated with a significant increase in pituitary PRL content, presumably a consequence of the decreased PRL secretion. In situ hybridization showed PRL mRNA to be localized in the cephalic lobe of the anterior pituitary gland in which most PRL cells, identified immunocytochemically, were found. Northern blotting studies showed that the pituitary gland contains a single 860 base(s) mature PRL mRNA transcript irrespective of physiological state or VIP manipulation. Both in situ and Northern hybridization studies confirmed that the amount of pituitary PRL mRNA was related directly to the concentration of plasma PRL. These observations are consistent with the view that in incubating hens hypothalamic VIP, in addition to acting as a PRL releasing hormone, also plays a major role in the regulation of the amount of PRL mRNA in the anterior pituitary gland.

The role of hypothalamic vasoactive intestinal polypeptide in the maintenance of prolactin secretion in incubating bantam hens: observations using passive immunization, radioimmunoassay and immunohistochemistry.

The role of chicken vasoactive intestinal polypeptide (cVIP) as a prolactin-releasing factor was investigated in incubating bantam hens. Specific antibodies were raised against cVIP (anti-cVIP) for passive immunization studies, to develop a radioimmunoassay and to localize VIP neurones immunohistochemically in the hypothalamus. The concentration of plasma prolactin decreased after i.v. injection of anti-cVIP: this low concentration being maintained by daily injection of anti-cVIP. Incubating hens injected daily with anti-cVIP deserted their nests after 4.5 +/- 0.6 days and returned to lay after 20 +/- 1 days. This disruption of incubation behaviour with anti-cVIP was prevented by concomitant, twice daily, injections of 30 IU ovine prolactin. The concentration of plasma LH was not immediately affected after injection of anti-cVIP but increased when the hens deserted their nests. The amount of cVIP, measured by radioimmunoassay, was significantly higher in the median eminence (P less than 0.01) and medial basal hypothalamus (P = 0.05) in incubating than in laying hens. No differences were seen in the amounts of cVIP in the preoptic hypothalamus or in a part of the forebrain including the nucleus accumbens, between laying and incubating hens. Morphological observations were made on immunohistochemically identified cVIP cell bodies in the medial basal hypothalamus. These showed that cVIP cell number, cell area and density of immunoreactive product were significantly (P less than 0.05) greater in incubating than in laying hens. Further, the density of cVIP reaction product in the anterior median eminence was also significantly (P less than 0.01) greater in incubating than in laying hens.(ABSTRACT TRUNCATED AT 250 WORDS)

Relationships between prolactin, LH and broody behaviour in bantam hens.

The interactions between broody behaviour and changes in concentrations of plasma prolactin and LH were investigated in bantam hens. Adoption of newly hatched chicks caused incubating hens to leave their nests and prevented plasma prolactin decreasing as rapidly as in hens deprived of their nests and not given chicks. Further, the hens allowed to rear chicks came back into lay later (P less than 0.001) than the hens not allowed chicks. Plasma prolactin decreased and plasma LH increased in hens deprived of their nests: these changes were reversed when the hens re-nested. The changes in plasma LH and prolactin in nest-deprived and re-nesting birds were not always synchronous; this was particularly clear immediately after nest deprivation when the increase in plasma LH preceded the decrease in the plasma prolactin. Readiness to incubate disappeared between 48 and 72 h after nest deprivation and corresponded with the time when plasma prolactin decreased to baseline values. Administration of ovine prolactin depressed (P less than 0.01) the initial increase in plasma LH after nest deprivation, but repeated administration of prolactin for up to 72 h failed to suppress plasma LH to the values found in incubating hens. Repeated administration of ovine prolactin at 5- to 8-h intervals for 72 h maintained readiness to incubate in nest-deprived hens. It is concluded that the secretion of prolactin in broody hens is facilitated by the presence of chicks and that increased concentrations of plasma prolactin maintain incubation behaviour. In incubating hens the secretion of LH and prolactin may be partly regulated independently. In addition, LH secretion may also be inhibited by increased plasma prolactin.

Comparison of the fractionation and assay of domestic duck and fowl pituitary gonadotrophins.

1. Pituitary glycoproteins from domestic ducks and fowls were fractionated to separate luteinising hormone (LH) and follicle stimulating hormone (FSH) activities using the same chromatographic steps. 2. Fractions were bioassayed for LH using the release of progesterone from fowl granulosa cells and for thyroid stimulating hormone (TSH) by measuring the release of thyroxine in 3-d-old chicks. Follicle stimulating hormone activity was measured either in a cockerel-testes radioreceptor assay or by the release of oestrogen from cultured rat Sertoli cells. 3. Fractions containing predominantly FSH or LH activity were isolated from the fowl glycoproteins. Duck gonadotrophin did not occur in fractions corresponding to those containing fowl FSH. 4. Duck gonadotrophin was found in a fraction corresponding with the most highly purified fowl LH fraction. A duck LH fraction was found with little FSH activity for which there was no corresponding fowl LH fraction. 5. It is concluded that domestic fowl and duck gonadotrophins have different chromatographic properties. Further study is required to determine whether the purified duck gonadotrophin preparation comprises proteins with similar physico-chemical properties but with separate FSH and LH biological activities.

The distribution of nuclear progesterone receptor in the hypothalamus and forebrain of the domestic hen.

Cell nuclei containing progesterone receptor were identified immunohistochemically in the hypothalamus and forebrain of the domestic hen using an antiserum to the steroid binding "B" subunit (110 kDa) of chicken oviduct progesterone receptor and the avidin-biotin complex procedure. Cell nuclei containing progesterone receptor were widely distributed in the anterior, medial and basal hypothalamus with the highest density occurring in the lamina terminalis and the preoptic area. Abundant, though less intensely reacting progesterone receptor was present in cell nuclei in the tuberal infundibular area and in the internal zone of the median eminence. A large group of cell nuclei containing progesterone receptor occurred in the dorsal anterior hypothalamus between the anterior commissure and the lateral ventricle. This group of nuclei extended anteriorly into the telencephalon. A small number of cell nuclei containing progesterone receptor was also found in the ventral telencephalon in the region of the nucleus accumbens.

Evidence that vasoactive intestinal polypeptide is a physiological prolactin-releasing factor in the bantam hen.

Vasoactive intestinal polypeptide (VIP)-like material was localised immunohistochemically in the hypothalamus of the bantam hen. Abundant immunoreactive VIP terminals were seen in the external layer of the median eminence and most immunoreactive VIP cell bodies were located in the basal hypothalamus. A few immunoreactive VIP cell bodies and many fibres were found in the preoptic hypothalamus. Intravenous injections of synthetic porcine VIP over a dose range of 12.5 to 100 micrograms kg-1 body wt resulted in dose-related increase in concentration of plasma prolactin in incubating bantams deprived of their nests for 24 hr. These doses of VIP did not stimulate the release of growth hormone. Studies in vitro showed that synthetic VIP directly stimulated prolactin release from the anterior pituitary gland. The glands from incubating bantams were more responsive to the prolactin-releasing effects of VIP than were the glands from laying birds. Studies in vitro showed that the amount of prolactin released in response to an iv injection of 50 micrograms kg-1 VIP was greater in incubating birds deprived of their nests for 24 hr than in laying hens. Prolactin release was not stimulated in ovariectomized hens after an injection of 50 micrograms kg-1 VIP unless the birds were first treated with oestrogen or oestrogen and progesterone. It was concluded that a VIP-like material in the bantam hypothalamus may be a physiological prolactin-releasing factor acting at least in part at the level of the anterior pituitary gland.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of luteinising hormone releasing hormone and its analogues on plasma luteinising hormone concentrations in incubating bantam hens.

The ability of synthetic vertebrate luteinising hormone releasing hormones (LHRHs) and their long-acting analogues to maintain elevated plasma luteinising hormone (LH) concentrations and to stimulate ovarian growth was investigated in incubating bantam hens. Chicken LHRH-II (pGlu1-His2-Trp3-Ser4-His5-Gly6-Trp7-Tyr8-Pro9-G ly10-NH2) was more effective than chicken LHRH-I (pGlu1-His2-Trp3-Ser4-Tyr5-Gly6-Leu7-Gln8-Pro9-Gly10-N H2) or porcine LHRH (pGlu1-His2-Trp3-Ser4-Tyr5-Gly6-Leu7-Arg8-Pro9-Gly10-N H2) in stimulating the release of LH. Long-acting analogues of chicken LHRHs (chLHRHs) were created by substituting D-amino acids in position 6. An intravenous injection (10 micrograms/bird) of D-Arg6-chLHRH-II or of a long-acting mammalian analogue of LHRH (buserelin) resulted in a sustained release of LH for up to 8 h. Less sustained releases of LH were observed after the same doses of D-Ala6-chLHRH-I or of D-Trp6-chLHRH-I. Repeated subcutaneous injections of D-Arg6-chLHRH-II or buserelin at 7 to 9 h intervals for 9 d resulted in loss of pituitary gland responsiveness to these analogues. For this reason, the treatment failed to maintain elevated plasma LH concentrations and did not stimulate the growth of the ovary or oviduct.

Photoperiodic requirement for the dissipation of scotorefractoriness in Japanese quail.

Male Japanese quail were reared on short days (6L:18D) and at 15-20 weeks of age those which had become sexually mature (i.e., scotorefractory) were transferred to long days (18L:6D) for between 2 and 29 weeks. The birds were then returned to 6L:18D for 3 weeks to test for the dissipation of scotorefractoriness. This was assessed by a decrease in at least 3 of 4 indices of reproductive function: testicular weight, area of the cloacal gland, and levels of plasma LH and testosterone. There was great individual variation in the photoperiodic requirement for the dissipation of scotorefractoriness, ranging between 6 and 29 weeks of exposure to long days. Scotorefractoriness was dissipated in about 50% of the birds after exposure to long days for between 6 and 12 weeks. It is concluded that the photoperiodic requirement for the dissipation of scotorefractoriness in quail cannot be defined precisely.

Aspects of the neuroendocrine control of ovulation and broodiness in the domestic hen.

The neuroendocrine control of ovulation and broodiness in the domestic hen involves complex interactions between hypothalamic neuropeptides, neurotransmitters, and ovarian steroids which regulate the secretion of luteinizing hormone (LH) and prolactin. Nuclear progesterone receptor is localized in many neurons throughout the hypothalamus but is absent from LHRH neurons. Hence, the positive feedback action of progesterone on LH release is not mediated by a genomic mechanism within the LHRH neuron. Precursors of 5-hydroxytryptamine (5HT) and dopamine (DA) inhibit the preovulatory release of LH, while the turnover rates of these neurotransmitters in the anterior hypothalamus decrease when preovulatory levels of LH are at their highest. Further, a population of receptors for 5HT which occurs in the anterior hypothalamus in laying birds is absent in nonlaying, incubating hens. Taken together, these observations suggest that the preovulatory surge of LH is mediated by a transitory decrease in the inhibitory action of 5HT and possibly DA, on the secretion of LHRH. Neurons containing 5HT may play a role in the regulation of prolactin release and, more specifically, in the control of broodiness. Drugs which enhance the function of 5HT neurons stimulate prolactin release while increased prolactin secretion in incubating hens is associated with an increase in the turnover of 5HT in the anterior hypothalamus. No receptors for 5HT were demonstrable in the anterior pituitary gland, showing that the prolactin-releasing activity of 5HT must be mediated by a prolactin-releasing factor (PRF). A candidate for a physiological PRF is vasoactive intestinal polypeptide (VIP).(ABSTRACT TRUNCATED AT 250 WORDS)

A comparison of the luteinizing hormone-releasing activities of synthetic chicken luteinizing hormone-releasing hormone (LH-RH), synthetic porcine LH-RH, and buserelin, an LH-RH analogue, in the domestic fowl.

The luteinizing hormone-releasing activities of synthetic chicken luteinizing hormone-releasing hormone (chLH-RH), synthetic porcine LH-RH (pLH-RH), and an analogue of LH-RH (buserelin, D-Ser-(But)6-des-Gly10-LH-RH ethylamide) were compared in the domestic fowl. In adult cockerels, intravenous injections of 0.5 or 1 microgram chLH-RH/kg released the same amount of LH as the same doses of pLH-RH; subcutaneous injections of 0.5 or 1 microgram buserelin/kg were about twice as effective as the same doses of pLH-RH. In laying hens, injections of 1, 10, 20, and 50 micrograms buserelin induced more sustained releases of LH than the corresponding doses of pLH-RH. Daily injections of 1 or 10 micrograms buserelin/bird or of 10 micrograms pLH-RH/bird for 12 days synchronized the timing of most ovipositions showing that the injections of releasing hormone could induce preovulatory surges of LH. In contrast with mammals, daily injections of buserelin in laying hens did not reduce pituitary responsiveness to the analogue. It is concluded that the structural difference between mammalian and chicken LH-RH does not affect their LH-releasing activities in the domestic fowl. Although the LH-releasing activity of buserelin in the hen is greater than that of pLH-RH, the difference in activity is not as great as that observed in most mammals. This view is strengthened by the finding that chronic treatment with buserelin, which exerts an antagonistic effect on ovulation in mammals, does not do so in the domestic hen.

Absence of nuclear progesterone receptor in LH releasing hormone in laying hens.

Using a double immunohistochemical technique, LH releasing hormone (LHRH) neurones and 110kDa nuclear progesterone receptor were localized in the hypothalamus of the laying hen. Nuclear progesterone receptor was widely distributed throughout the hypothalamus, occurring in the preoptic, septal, anterior and basal areas. The region where progesterone receptor was revealed in nuclei of neurones overlapped that containing LHRH neurones. However, LHRH cell bodies did not contain progesterone nuclear receptor. It is concluded that the positive feedback action of progesterone on LH release is not mediated by a genomic mechanism within the LHRH neurone.

Plasma concentrations of luteinising hormone, follicle stimulating hormone, androgen, growth hormone, prolactin, thyroxine and triiodothyronine during growth and sexual development in the cockerel.

Changes in concentrations of plasma luteinising hormone (LH), follicle stimulating hormone (FSH), androgen, growth hormone (GH), prolactin (Prl), thyroxine (T4) and triiodothyronine (T3) were measured during growth and sexual maturation in broiler cockerels reared in continuous light to 7 weeks and 14 h light/d thereafter. Concentrations of LH and FSH began to increase between 13 and 15 weeks, while those of androgens increased between 16 and 17 weeks. FSH concentration increased faster than that of LH. Concentrations of GH and Prl were high at 3 weeks; that of GH decreasing progressively between 3 and 14 weeks of age and thereafter remaining low, while that of Prl was low between 5 and 9 weeks, relatively high between 10 and 13 weeks, and then temporarily decreasing before increasing progressively during sexual maturation. Concentrations of T3 and T4 were higher in juvenile than in adult birds.

Concentrations of triiodothyronine, growth hormone, and luteinizing hormone in the plasma of thyroidectomised fowl (Gallus domesticus).

Surgical thyroidectomy increased (P less than 0.05) the basal concentrations of growth hormone (GH) and luteinizing hormone (LH) in the plasma of 10- to 12-week-old domestic fowl. The administration of thyrotrophin releasing hormone (TRH) (100 micrograms, sc) increased (P less than 0.01) the GH concentration in both intact and thyroidectomised birds. The magnitude of the TRH-induced increase in GH level was greater (P less than 0.01) in thyroidectomised birds than in intact controls. Although TRH had no effect on LH secretion in the controls, it induced a small (P less than 0.05) rise in the plasma LH level in thyroidectomised birds. In both the intact and thyroidectomised birds the LH concentration was enhanced (P less than 0.05) following the administration of LH-releasing hormone (LH-RH) (20 micrograms, sc). The increase in the LH level by LH-RH in the thyroidectomised birds was greater (P less than 0.001) than that in the intact controls. Plasma GH concentrations were unaffected by LH-RH treatment. These results suggest that thyroid hormones inhibit the secretion of LH and GH in birds. In thyroidectomised birds low levels of immunoreactive triiodothyronine (T3)-like material were measurable in the circulation, despite the absence of regenerated thyroid tissue. The administration of TRH (100 micrograms, sc) did not enhance the plasma level of this material in thyroidectomised birds, whereas plasma T3 concentrations were enhanced in intact birds following TRH treatment. These results suggest that the T3 immunoreactive substance in thyroidectomised birds is extrathyroidal in origin.

The localisation of LH-RH neurones in the diencephalon of the domestic hen.

Nerve fibers and perikarya containing LH-RH-like material were identified immunohistochemically in the diencephalon of the domestic hen using the peroxidase-anti-peroxidase technique. Perikarya were thinly scattered in bilateral bands close to the third ventricle extending from the nucleus praeopticus paraventricularis magnocellularis, passing in front of the anterior commissure into the septal area. In this latter area, the perikarya tended to spread out laterally. A few perikarya were seen in the anterior portion of the nucleus paraventricularis magnocellularis but were not found in the infundibular nuclear complex. Fibre tracts were seen running dorso-ventrally in the preoptic area apparently associated with the lamina terminalis. Fibres, possibly nerve terminals, were found in the lamina terminalis and in the external layers of the anterior and posterior divisions of the median eminence. A large number of fibres was seen distributed throughout the infundibular nuclear complex; scattered fibres were found close to the third ventricle in the anterior hypothalamus. Extrahypothalamic fibers were also observed to project from the septal area into other parts of the telencephalon.

Diminution of thyrotrophin releasing hormone-induced growth hormone secretion in adult domestic fowl (Gallus domesticus).

Age-related changes in the response of GH to administration of thyrotrophin releasing hormone (TRH) have been investigated in the domestic fowl. In two strains of chicken the i.v. administration of TRH (10 microgram/kg) to 4-week-old male and female birds markedly increased (greater than 100 ng/ml) the plasma GH concentration within 10 min of treatment and the concentration remained higher than the pretreatment level for at least a further 20 min. Saline (0.9%) administration had no effect on GH secretion in comparable groups of control birds. The same dose of TRH had no effect on plasma GH concentrations in adult (greater than 24-week-old) laying hens or cockerels. The administration of TRH at doses of 0.1-100 microgram/kg (i.v.) or 0.39-50 microgram/bird (s.c.) also had very little, if any, effect on GH secretion in laying hens. In laying hens slight increases (10-20 ng/ml, P less than 0.05) inthe plasma concentrations of GH were observed in one experiment 60 min after the s.c. injection of 100 microgram TRH, and in another 60, 90 and 120 min after the serial s.c. injection of TRH (100 microgram/bird) every 30 min over a 150 min period. The poor GH response to TRH of the adults to TRH stimulation was not due to high circulating concentrations of endogenous gonadal steroids, as surgical gonadectomy had no effect on the GH response to TRH. These results suggest maturational differences in the control of GH secretion in the fowl.