1. Simultaneous changes of the width of the cloacal opening and plasma luteinising hormone (LH), follicle stimulating hormone (FSH), 17 beta-oestradiol, progesterone, prolactin, thyroxine (T-4) and triiodothyronine (T-3) during photo-induced ovarian growth and regression were measured in commercially bred Japanese quail from a heavy body weight line.2. Somatically mature female Japanese quail were transferred from short days (light: dark 8L:16D) at 10 degrees C to long days (16L:8D) at 20 degrees C, and sexually mature female Japanese quail were transferred from long to short days. All variables were measured at transfer and every five days thereafter (except for a measurement at 12 instead of 10 d) for 35 d.3. Transfer from short to long days caused significant increases in LH, FSH, 17 beta-oestradiol, ovary weights and oviduct weights after five days, and in the cloacal opening after 12 d. T-3 decreased after five days, whereas no significant changes were observed in T-4 concentrations. Progesterone and prolactin both decreased after 25 long days.4. The transfer of quail from long to short days caused significant decreases in LH, FSH, 17 beta-oestradiol, progesterone, prolactin, ovary and oviduct weights after 12 d and an increase in T-3. There was no significant change in T-4 concentrations. The cloacal opening decreased after 25 short days.5. These results are the first to show simultaneous changes in gonadotrophins, sex steroids, thyroid hormones and prolactin during photo-induced gonadal growth and regression in female Japanese quail.

Retrotransposons are mobile genetic elements that use a germline 'copy-and-paste' mechanism to spread throughout metazoan genomes. At least 50 per cent of the human genome is derived from retrotransposons, with three active families (L1, Alu and SVA) associated with insertional mutagenesis and disease. Epigenetic and post-transcriptional suppression block retrotransposition in somatic cells, excluding early embryo development and some malignancies. Recent reports of L1 expression and copy number variation in the human brain suggest that L1 mobilization may also occur during later development. However, the corresponding integration sites have not been mapped. Here we apply a high-throughput method to identify numerous L1, Alu and SVA germline mutations, as well as 7,743 putative somatic L1 insertions, in the hippocampus and caudate nucleus of three individuals. Surprisingly, we also found 13,692 somatic Alu insertions and 1,350 SVA insertions. Our results demonstrate that retrotransposons mobilize to protein-coding genes differentially expressed and active in the brain. Thus, somatic genome mosaicism driven by retrotransposition may reshape the genetic circuitry that underpins normal and abnormal neurobiological processes.

1. Simultaneous changes of cloacal gland area (CGA) and plasma luteinising hormone (LH), follicle stimulating hormone (FSH), testosterone (T), prolactin (PRL), thyroxine (T(4)) and triiodothyronine (T(3)) during photo-induced testicular growth and regression were measured in commercially bred Japanese quail from a heavy body weight line. 2. Somatically mature male Japanese quail were transferred from short days (light:dark 8L:16D) at 10°C, to long days (16L:8D) at 20°C; and sexually mature male Japanese quail were transferred from long to short days. All variables were measured at transfer and every 5 d thereafter for 35 d. 3. Transfer from short to long days caused significant increases in LH, FSH, T and testis weight (TW) after 5 d, and in CGA after 10 d. T(3) decreased after 5 d, whereas T(4) increased significantly after 25 long days and PRL did not undergo any consistent change. The testicular growth rate was k = 0·1146. 4. Transferring quail from long to short days caused significant decreases in LH and FSH after 5 d, and decreases in T, TW and CGA after 10 d. T(4) decreased after 5 d whilst T(3) increased significantly by day 15. PRL decreased significantly after 10 d then rose before declining again. The testicular regression rate was k = 0·0582. 5. The rates of photo-induced testicular development and regression in a strain of large Japanese quail did not differ from rates reported for other strains of quail. CGA was a better indicator of TW than plasma T concentrations during growth and regression. The role of PRL in photo-induced reproductive cycles in male Japanese quail remains to be determined. 6. The photoperiod-induced changes in gonad size and hormone concentrations, together provide valuable information that can be used in future studies of the endocrinology and neuroendocrinology of photoperiodism in birds.

Infection of sheep with the gastric nematode Teladorsagia circumcincta results in distinct Th2-type changes in the mucosa, including mucous neck cell and mast cell hyperplasia, eosinophilia, recruitment of IgA/IgE producing cells and neutrophils, altered T-cell subsets and mucosal hypertrophy. To address the protective mechanisms generated in animals on previous exposure to this parasite, gene expression profiling was carried out using samples of abomasal mucosa collected pre- and post- challenge from animals of differing immune status, using an experimental model of T. circumcincta infection. Recently developed ovine cDNA arrays were used to compare the abomasal responses of sheep immunised by trickle infection with worm-naïve sheep, following a single oral challenge of 50 000 T. circumcincta L3. Key changes were validated using qRT-PCR techniques. Immune animals demonstrated highly significant increases in levels of transcripts normally associated with cytotoxicity such as granulysin and granzymes A, B and H, as well as mucous-cell derived transcripts, predominantly calcium-activated chloride channel 1 (CLCA1). Challenge infection also induced up-regulation of transcripts potentially involved in initiating or modulating the immune response, such as heat shock proteins, complement factors and the chemokine CCL2. In contrast, there was marked infection-associated down-regulation of gene expression of members of the gastric lysozyme family. The changes in gene expression levels described here may reflect roles in direct anti-parasitic effects, immuno-modulation or tissue repair. (Funding; DEFRA/SHEFC (VT0102) and the BBSRC (BB/E01867X/1)).

In order to further advance the understanding of genes involved in avian photoperiodic signaling, a chicken hypothalamic cDNA microarray was made to identify changes in gene expression in the whole hypothalamus of juvenile male domestic chickens after 4 days' photostimulation. The most robust change was a depression in heat shock protein 90B1 (HSP90B1) expression. This observation was confirmed using quantitative PCR, and it was subsequently demonstrated that the depression in HSP90B1 expression first occurs in the anterior hypothalamus after 1 day's photostimulation, and was also depressed in the anterior and basal hypothalamus after 4 days' photostimulation. Four days after an intravenous injection of thyroxine (T4), an avian photomimetic, in short day birds, HSP90B1 expression was depressed in the anterior, but not in the basal hypothalamus. Depressed HSP901 expression after photostimulation or T4 treatment was associated with increased GnRH-I mRNA and plasma LH. HSP90B1 is abundant throughout the brain where it occurs in glial cells, and is involved in regulating white matter plasticity. It is suggested that photoperiodically depressed hypothalamic HSP90B1 may affect glial function in photoperiodic signaling pathways in the neuroendocrine system. This is the first report of a thyroid hormone-responsive gene involved in photoperiodic signaling.

Activation of the shell gland region of the avian oviduct is mediated by ovarian steroids. To understand more extensively how shell glands are maintained and function, we have compared gene expression in the shell glands from juvenile and laying hens using a chicken cDNA microarray. Average expression profiles of juvenile and sexually mature shell glands were compared resulting in the identification of 266 differentially regulated genes. Reverse transcription quantitative polymerase chain reaction confirmed expression differences. The differentially expressed genes included several with known involvement in shell gland function, including ion transport and shell matrix proteins. There were also many unpredicted differentially expressed genes, and for some we propose hypotheses for their functions. These include those encoding (a) osteoprotegerin, a decoy death receptor for receptor activator of nuclear factor NFkB ligand (RANKL) and tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), that in the shell gland, may prevent apoptosis and/or may have an endocrine effect by preventing RANKL's action on bone osteoclasts that mobilize stored calcium; (b) prostatic acid phosphatase (ACPP) and prostate stem cell antigen (PSCA) that could play a role in sperm physiology within the shell gland; (c) urea transporter (SLC14A2) that could provide a novel anti-microbial defence; (d) bactericidal/permeability-increasing protein-like 2 (BPIL2), and other potential anti-microbials that have not previously been documented in the chicken. These new hypotheses, if borne out experimentally, will lead to a greater understanding of shell gland function including the processes involved in eggshell formation and anti-microbial activity.

BACKGROUND:The development of microarray resources for the chicken is an important step in being able to profile gene expression changes occurring in birds in response to different challenges and stimuli. The creation of an immune-related array is highly valuable in determining the host immune response in relation to infection with a wide variety of bacterial and viral diseases.RESULTS:Here we report the development of chicken immune-related cDNA libraries and the subsequent construction of a microarray containing 5190 elements (in duplicate). Clones on the array originate from tissues known to contain high levels of cells related to the immune system, namely Bursa, Peyers patch, thymus and spleen. Represented on the array are genes that are known to cluster with existing chicken ESTs as well as genes that are unique to our libraries. Some of these genes have no known homologies and represent novel genes in the chicken collection. A series of reference genes (ie. genes of known immune function) are also present on the array. Functional annotation data is also provided for as many of the genes on the array as is possible.CONCLUSION:Six new chicken immune cDNA libraries have been created and nearly 10,000 sequences submitted to GenBank [GenBank: AM063043-AM071350; AM071520-AM072286; AM075249-AM075607]. A 5 K immune-related array has been developed from these libraries. Individual clones and arrays are available from the ARK-Genomics resource centre.

Chicken gonadotropin‐releasing hormone‐I (GnRH‐I) and the avian prolactin‐releasing hormone, vasoactive intestinal polypeptide (VIP), were measured in the basal hypothalamus in male starlings during photo‐induced gonadal growth and the subsequent development and maintenance of reproductive photorefractoriness. Comparisons were made with thyroidectomized birds, which maintain breeding condition irrespective of changes in photoperiod. In intact birds, basal hypothalamic GnRH‐I increased four‐fold after photostimulation and then decreased 115‐fold over 12 weeks to values characteristic of long‐term photorefractoriness. Pituitary and plasma prolactin increased after photostimulation, reaching peak values when the testes were regressing, and returned to low values in long‐term photorefractory birds. Basal hypothalamic VIP did not change after photostimulation in intact birds. In photostimulated thyroidectomized birds, values for basal hypothalamic GnRH‐I and VIP, and for pituitary and plasma prolactin, remained no different to those of nonphotostimulated intact birds. These observations confirm that reproductive photorefractoriness is related to a decrease in hypothalamic GnRH‐I. However, photorefractoriness in terms of prolactin secretion is not similarly related to a decrease in basal hypothalamic VIP. The mechanisms responsible for the decrease in prolactin in long‐term photorefractory birds and for the total lack of photoperiodic responses in thyroidectomized birds remain unresolved.

The genetic control of incubation behavior was investigated in the domestic hen by analysis of the incidence of the behavior in reciprocal crosses between nonbroody White Leghorn (WL) and broody Bantam (B) lines and in a backcross of F-1 males (WL male x B female) and WL females. The hypothesis tested was that a sex-linked gene (or genes) plays a dominant role in the expression of incubation behavior. The incidence of incubation behavior was tested in hens held in floor pens with access to nests containing hard-boiled eggs during a 28-wk photoinduced laying cycle. The cycle was repeated if the behavior was not observed during the first cycle. The incidence of incubation behavior in B and WL hens was 78.6% (n = 28) and 0% (n = 28), respectively. Contrary to prediction, the incidence of incubation behavior in the WL male x B female and the B male x WL female crosses were not significantly different (61.6%, n = 73; and 56.8%, n = 37, respectively). The incidence of incubation behavior in the F-1 backcross was 5.8% (n = 103), which was significantly less (P < 0.001) than predicted (39.3%). It was concluded that incubation behavior was not controlled by major genes on the Z chromosome. It was hypothesized that at least two dominant autosomal genes are involved, one causing and the other inhibiting the behavior with equal influence.

The objective of this study was to establish, for a short-day breeding bird, the male emu, whether the breeding season is principally controlled by changes in photoperiod, and to investigate the endocrine mechanisms involved. Two groups of adult males were subjected to three alternating periods of 150-185 days of 14 h light/day (LD) and 10 h light/day (SD) terminating in a 360-day period of LD or SD. Transfer from LD to SD led to increases in plasma concentrations of luteinizing hormone (LH) and testosterone, after 82 +/- 8 and 73 +/- 3 (SEM) days, and an increase in prolactin concentrations after 115 +/- 12 days. Concentrations of LH and testosterone began to decrease before transfer back to LD, at a time when prolactin concentrations were approaching peak values. Transfer from LD to 360 days of SD resulted in increases in LH and testosterone concentrations, and these terminated after an increase in prolactin concentrations. After transfer from SD to 360 days of LD, plasma concentrations of LH and testosterone began to increase, after delays of 222 +/- 24 and 225 +/- 13 days, and were high at the end of the study, while prolactin values remained depressed throughout. These observations clearly show that seasonal breeding in the emu is directly controlled by changes in photoperiod. The dynamics of the hormonal responses to change of photoperiod suggest that, despite being short-day breeders, the photoregulation of breeding in emus involves mechanisms that are currently accepted for birds, rather than mechanisms that have been proposed for short-day breeding mammals. The initiation of breeding in emus is due to dissipation of photorefractoriness by short days which leads to an increase in the secretion of gonadotrophins to levels that are sufficient to support full reproductive condition. The termination of breeding, while days are still short, is due to the antigonadotrophic action of prolactin which, unusually for birds, increases while the days are still short. In conclusion, breeding activity in male emus is strongly controlled by photoperiod. Emus are short-day breeders, but the central mechanisms that regulate the secretion of reproductive hormones seem to be similar to those previously proposed for long-day breeding birds. The pattern of prolactin secretion in emus suggests an important role for this hormone in the termination of the breeding cycle.

The aim of this study was to increase understanding of the occurrence and regulation of chicken gonadotropin releasing hormone I (cGnRH I) and chicken gonadotropin releasing hormone receptor (cGnRH-R) mRNA variants in the hypothalamic-pituitary-testicular axis (HPTA). The study was carried out in the cockerel. Fully processed cGnRH I mRNA (cGnRH Ia) and a variant transcript (cGnRH Ib) with a retained intron 1 were observed in the preoptic/anterior hypothalamus (POA), the basal hypothalamus, anterior pituitary gland, and testes. Fully processed cGnRH-R mRNA (cGnRH-Ra) and a variant transcript (cGnRH-Rb) with a deletion were detected in the same tissues. In juvenile cockerels, concentrations of cGnRH Ia and b in the POA increased after castration, and this was prevented by oestrogen treatment. In the anterior pituitary gland, the concentration of cGnRH-Ra increased after castration and this was reversed by oestrogen treatment. In intact adult cockerels, oestrogen treatment depressed plasma luteinizing hormone but did not affect concentrations of cGnRH I and cGnRH-R mRNAs in the POA, basal hypothalamus, and anterior pituitary gland, suggesting that locally produced oestrogen, by aromatization, may exert maximal suppression on cGnRH I and GnRH-R mRNAs. In intact adult cockerels, the concentrations of cGnRH Ia and b in the testis, but not cGnRH-Ra and b, were depressed by oestrogen treatment. It was concluded that fully processed and variant cGnRH I and cGnRH-R mRNAs occur in all components of the HPTA. Oestrogen appears to play a role in the regulation of cGnRH Ia and b in the POA and testes, and of cGnRH-Ra in the POA and anterior pituitary gland.

The aim of the study was to establish, in contemporary broiler breeders, whether delayed photostimulation at 313 d results in a reproductive response similar to that after photostimulation at 134 d (standard practice). The standard lighting program was compared with a novel program in which daily hours of light were reduced to 3 h during rearing and kept at 3 h until photostimulation at 264 d (8 h) or at 313 d (16 h). This experiment was done with hens fed ad libitum or feed-restricted hens. In photostimulated and nonphotostimulated hens, feed restriction delayed the onset of egg production and enhanced the subsequent rate of laying. Standard photostimulation advanced the onset of lay and increased the subsequent rate of lay in hens fed ad libitum and feed-restricted hens. Delayed photostimulation of hens did not impair the photoinduced increase in the concentration of plasma luteinizing hormone (LH) or egg production. Delayed photostimulation in cockerels failed to stimulate LH secretion. Unexpectedly, for feed-restricted hens, transfer from 3 to 8 h light/d at 264 d resulted in an increased in plasma LH and increased egg production. A similar increase in plasma LH was observed for cockerels subjected to the same lighting treatment. We concluded that, in broiler breeder hens, the reproductive response to photostimulation is not impaired if photostimulation is delayed for up to 313 d. Cockerels may not respond well to delayed photostimulation.

Broodiness is a behavioural trait observed in most common breeds of domestic fowl with the exception of the White Leghorn. The genetics of broodiness has been investigated and has produced conflicting observations: some authors have produced evidence suggesting that the trait is polygenic with a major sex-linked effect (review Hutt, 1949; Saeki, 1957; Saeki and Inoue, 1979), while others found no evidence of sex-linked genes for broodiness in the Rhode Island Red (Hays, 1940). The aim of this study was to reassess the possibility that there is a major gene located on the Z chromosome implicated in the control of broodiness. The identification of the location of such a gene or its markers would facilitate marker-assisted selection against broodiness...

The DNA sequence of a chicken leptin gene that shares 95% nucleotide similarity with the mouse leptin sequence has been recently reported (Taouis et al., 1998, Gene 208, 239-242). Experiments have been performed independently in two laboratories to try to confirm this finding. Fourteen PCR primers based on the mouse leptin sequence were designed to amplify the avian leptin gene. Four of the primers were identical to the mouse and published chicken leptin sequences. PCR amplification was carried out on genomic DNA and reverse-transcribed mRNA from the fat, liver, and pancreas of several chicken strains and from the domestic turkey goose, and Japanese quail. No PCR products sharing close similarity to the mouse leptin sequence were generated from any avian templates. Amplification of mouse leptin sequence was consistently obtained when control mouse templates were used. Northern hybridization using a mouse leptin probe failed to produce a signal with poly(A)(+) RNA from chicken fat and liver and from the fat and liver of force-fed geese but a strong signal was obtained from control mouse fat total RNA. Southern hybridization under low stringency washing conditions revealed hybridization of a mouse leptin probe to chicken genomic DNA. Under higher stringency washing conditions, the chicken signal disappeared, while those from control mouse and sheep genomic DNA remained. This suggests that the putative chicken leptin sequence shares less than the 83% nucleotide sequence identity between the mouse and sheep genes. It is concluded that a chicken leptin gene sequence with close sequence similarity to mouse leptin is not present in the chicken genome. Furthermore, mRNA sharing high sequence identity with mouse leptin is not present in the fat or liver of the domestic chicken, turkey, goose, or Japanese quail. (C) 1999 Academic Press.

Seasonal changes in testicular morphology and blood plasma concentrations of LH, testosterone, and prolactin are described for captive male emus in southwestern Australia. Testicular mass and testicular testosterone did not differ between the non-breeding (spring–summer) and the breeding (autumn–winter) seasons. Nevertheless, the testes obtained in the breeding season (May and August) were nearly two fold greater in mass than those collected in the non-breeding season (October and February). The highest testicular concentrations of testosterone were observed in February and lowest in October, while the values during the breeding season were intermediate. The patterns of histological changes in the testes also indicate that emus breed over the autumn–winter months. Tubule diameter was larger in the breeding season than in the non-breeding season, whereas the relative volume of the interstitium was larger in the non-breeding and smaller in the breeding season. Moreover, during the autumn and winter months, plasma LH and testosterone concentrations were high. Outside this period, in spring and summer, the concentrations of these hormones were low. Prolactin concentrations rose around the winter solstice, after the initial increases in plasma LH and testosterone. The end of the breeding season, in early spring, was marked by a gradual decrease in plasma LH concentrations but a rapid fall in testosterone concentrations. Prolactin concentrations continued to increase and peaked near the spring equinox, several weeks after the breeding season ended, and then decreased to reach baseline values by mid-summer. These testicular and endocrine changes are consistent with observations that the emu is a short-day breeder in southwestern Australia. Reproductive activity in the male begins soon after the summer solstice, well in advance of the development of suitable breeding conditions, and is then terminated in spring before food resources become limited by the onset of the dry season.

The peptide neurotransmitter vasoactive intestinal polypeptide (VIP) has several important functions in vertebrates, particularly, influencing the neuroendocrine and autonomic nervous systems both in developing and in adult animals. To document potential brain areas that might play significant functional roles, the distribution of VIP mRNA was examined throughout the entire chick brain by using in situ hybridization histochemistry (ISHH). In addition, a VIP binding-site study was completed that focused on the lateral septal organ (LSO), a circumventricular organ of potential significance in avian species. The areas where VIP message was found included the olfactory bulbs, posterior hippocampus, parahippocampal area, hyperstriatum, archistriatum/nucleus (n.) taenia (amygdala), medial part of the LSO, organum vasculosum of the lamina terminalis, medial preoptic region, bed n. of the pallial commissure, anterior hypothalamic (hypo.) n., lateral hypo. area (most extensive and dense message), periventricular hypo. n., lateral to the paraventricular n., ventromedial hypo. n., stratum cellulare externum, inferior hypo. n., infundibular hypo. n., median eminence, three layers within the stratum griseum et fibrosum superficiale, area ventralis of Tsai, n. tegmenti pedunculopontinus pars compacta (substantia nigra), intercollicular n., central gray, locus ceruleus, parabrachial n., ventrolateral medulla, reticular pontine area, in and about the n. vestibularis descendens. When compared with immunocytochemistry that detected the presence of the peptide product VIP, more areas of the brain were found to contain perikarya expressing VIP by using ISHH, particularly in the telencephalon and the mesencephalon. VIP binding sites were found in the lateral portion of the LSO where the blood-brain barrier is not fully developed. Hence, the LSO was found to contain neural elements that synthesize as well as bind VIP. VIP appears to be a useful peptide for defining major components of the visceral forebrain system in birds. (C) 1997 Wiley-Liss, Inc.

Transfer of intact, photosensitive starlings from short to long days causes an increase in plasma thyroxine and gonadal maturation and later induces photorefractoriness. Thyroidectomy of starlings prevents the induction of photorefractoriness. This study investigated whether the long-day-induced increase in plasma thyroxine is necessary for the induction of photorefractoriness. Photosensitive starlings were thyroidectomised, given thyroxine in their drinking water at concentrations that result in plasma thyroxine at short-day physiological concentrations or lower, and transferred to long days. Plasma thyroxine and prolactin, gonadal size, and moult were monitored. The group with short-day concentrations of plasma thyroxine became photorefractory at the same time as intact controls transferred to long days. The other groups, with lower plasma thyroxine, also became photorefractory, but the onset of photorefractoriness was delayed. The increase in plasma prolactin following photostimulation was proportional to plasma thyroxine concentrations. The onset of moult was also related to plasma thyroxine. We conclude that the long-day concentrations of plasma thyroxine observed in the plasma of intact starlings are not necessary for the induction of photorefractoriness. This suggests that thyroxine acts as a permissive factor rather than actively driving the photorefractory process.