Active immunization with vasoactive intestinal peptide in turkey hens.

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.

Active immunization against vasoactive intestinal peptide prevents photo-induced prolactin secretion in turkeys.

Photostimulation initiates and maintains the rise in circulating prolactin (PRL) observed during the reproductive cycle of the female turkey. Vasoactive intestinal peptide (VIP) is the principal PRL-releasing factor. This study tested the hypothesis that gonadal stimulatory photoperiods stimulate PRL secretion by releasing hypothalamic VIP. Therefore, an experiment was designed to determine if VIP immunoneutralization altered photo-induced PRL secretion. Reproductively quiescent female turkeys were divided into two groups comprising turkeys actively immunized with synthetic VIP conjugated to keyhole limpet hemocyanin (VIP-KLH; immunized; n = 48) or KLH alone (control; n = 48). The first immunization was administered 6 weeks before photostimulation. Blood samples were collected at frequent intervals prior to and following photostimulation, and plasma PRL concentrations were determined. Vasoactive intestinal peptide antibody titer was estimated from the percentage of 125I-labeled VIP bound to plasma diluted 1:1000. At the onset of photostimulation (Day 0), plasma PRL levels were similar for immunized and control turkeys (9.1 +/- 0.3 versus 8.9 +/- 0.3 ng/ml, respectively). Plasma PRL of control birds increased (P < 0.05) by Day 16 of photostimulation, reaching a peak value of 724.9 +/- 90.1 ng/ml on Day 84. In contrast, plasma PRL remained essentially unchanged in immunized birds. Titer of anti-VIP antibodies expressed as 125I-VIP bound by plasma in immunized birds was 10.9 +/- 1.5% on the day of photostimulation. Incubation behavior was blocked in immunized birds, whereas 75% of controls exhibited incubation behavior. The control group laid 1.83 eggs/ week/hen compared to 3.40 eggs/week/hen in immunized hens. These findings suggest that photoperiodic modulation of PRL secretion in the turkey is influenced by hypothalamic VIP neuronal system.

Serotonergic stimulation of prolactin secretion is inhibited by vasoactive intestinal peptide immunoneutralization in the turkey.

The neuronal mechanisms that govern prolactin (PRL) secretion in the turkey appear to involve monoaminergic systems. Considerable evidence indicates that serotonin (5-HT), acting centrally, is a potent stimulator of PRL secretion. This study, using birds actively immunized against VIP, tests the hypothesis that 5-HT stimulates PRL secretion by releasing vasoactive intestinal peptide (VIP). Nonimmunized turkeys were injected ip with saline, quipazine (5-HT agonist; 5 mg/kg), methysergide (5-HT antagonist; 8 mg/kg), or methysergide plus quipazine, and VIP-immunized birds were injected with saline or quipazine. Quipazine increased plasma PRL levels from 26.8 +/- 7.1 ng/ml at Time 0 to a peak value of 148.1 +/- 31.4 ng/ml 2 hr after infection. Pretreatment with methysergide or VIP-immunoneutralization abolished the PRL response to quipazine. Intraventricular infusion of 5-HT (1 nmol/min) caused plasma PRL to rise from a baseline of 16.3 +/- 2.6 ng/ml to 85.2 +/- 14.3 ng/ml after 30 min in nonimmunized control birds. Serotonin infusion did not induce PRL secretion in the VIP-immunized birds. These findings suggest that serotonergic stimulation of PRL secretion in the female turkey requires a functional VIPergic system.

Tissue-specific alternative splicing of turkey preprovasoactive intestinal peptide messenger ribonucleic acid, its regulation, and correlation with prolactin secretion.

Although vasoactive intestinal peptide (VIP) is a well characterized physiological PRL-releasing factor in avian species, its regulated expression is not fully understood. We cloned complementary DNAs encoding the prepro-turkey VIP (prepro-tVIP) molecule from an adult turkey hypothalamic complementary DNA library. When the amino acid sequence of the prepro-tVIP was compared to chicken and mammalian sequence, it was found that the isolated tVIP molecules lacked the 27-amino acid peptide histidine isoleucine (PHI) portion of the precursor protein. Several tissues showed an alternatively spliced tVIP transcript that lacked the PHI sequence. Only in the hypothalamus did tVIP-specific primer pairs and reverse transcription-polymerase chain reaction produce two alternatively spliced fragments. The larger hypothalamus-specific fragment was subjected to nucleotide sequence analysis and identified as containing the alternatively spliced PHI-containing exon, which encoded a 27-amino acid PHI peptide in addition to the 8 amino acids that flanked the peptide. Hypothalamic tVIP expression was shown to be up-regulated during the incubation phase of the reproductive cycle. The increased steady state level of tVIP messenger RNA appears to be regulated by nesting behavior, because nest deprivation dramatically suppressed its expression. Levels of the minor tVIP transcript containing both the PHI- and VIP-encoding exons did not significantly change between reproductive stages and were maintained at approximately 4-6% of the total tVIP transcript level. Our findings provide further evidence that VIP is the most important PRL-releasing factor in birds. Our study should serve as a useful model for determining whether PHI contributes in any way to the physiological role of PRL regulation. Revealing the tissue distribution of VIP and PHI gene expression and tissue-specific alternative splicing could contribute to an understanding of the physiological functions of the two peptides as well as their relative roles in PRL regulation.

Modulation of ovarian cytochrome P450 17 alpha-hydroxylase and cytochrome aromatase messenger ribonucleic acid by prolactin in the domestic turkey.

The effect of exogenous ovine prolactin (oPRL) on preovulatory follicle P450 17 alpha-hydroxylase (C17) and aromatase (ARO) mRNA abundance was investigated in turkeys. Ovine PRL (124 IU/hen per day) was injected i.m. into four sets (n = 8) of laying turkeys for 2, 4, 8, or 14 days. Vehicle was injected into control hens for 8 days (n = 8). Blood samples were collected and serum was assayed for LH, progesterone (P), testosterone (T), and estradiol (E). Theca layers from the largest (F1) and the third (F3), fifth (F5), and seventh (F7) largest preovulatory follicles and from small white follicles (SWF) were examined for C17 and ARO mRNA contents. The number of atretic follicles increased from 0 (vehicle-injected controls) to 9 (14-day-oPRL-injected hens). Serum E, T, and LH levels decreased, while P levels remained unchanged. There was a transient increase in theca C17 mRNA abundance of 2- and 4-day-oPRL-treated hen follicles. Cytochrome P450 ARO mRNA levels were reduced in SWF and F7 in response to oPRL. Thecal C17 and ARO mRNA content was reduced during follicular maturation in laying hens. ARO mRNA was not detectable in granulosa cells. The progressive decline in C17 and ARO mRNA content associated with follicular maturation as well as the absence of ARO mRNA in granulosa cells is consistent with the secretory activity of P, T, and E in preovulatory follicles. These findings suggest that reduced circulating E may be a consequence of suppressed ARO gene expression whereas the oPRL suppression of T secretion may not be coupled to C17 gene expression.

Sequence analysis of the turkey LH beta subunit and its regulation by gonadotrophin-releasing hormone and prolactin in cultured pituitary cells.

cDNAs encoding the precursor molecule of the turkey LH beta subunit (tLH beta) were cloned from a turkey pituitary cDNA library. The nucleotide sequence of the longest of two different tLH beta cDNA clones contained 592 bp, and included 23 bp of the 5' untranslated region (UTR) and 92 bp of the 3' UTR in addition to a 477 bp open reading frame that encoded a 39 amino acid leader polypeptide and a 120 amino acid mature apoprotein. Turkey and chicken LH beta sequences shared approximately 92 and 93% nucleotide and amino acid sequence similarities respectively. Northern blot analysis of total cellular anterior pituitary RNA showed that an approximate 800 base transcript hybridized to a 32P-labelled tLH beta cDNA probe. The gonadotrophin-releasing hormone (GnRH)- and prolactin (PRL)-regulated expression of LH and PRL in dispersed pituitary cells was determined by Northern blot analysis of tLH beta and PRL steady-state mRNA levels and by RIA analysis of secreted LH and PRL. GnRH-treated cells showed increased levels of both tLH beta mRNA and secreted LH, whereas mRNA and secreted levels of PRL did not change significantly. Cells treated with PRL showed lower levels of tLH beta and PRL mRNA as well as decreased release of LH and PRL. When cells were treated with both PRL and GnRH, increases in tLH beta mRNA and secreted levels of LH observed with GnRH alone were negated, whereas the decreases in mRNA and secreted levels of PRL observed with PRL alone were abrogated. These findings suggest that PRL can down-regulate tLH beta gene expression and spontaneous release of LH as well as autoregulate PRL gene expression and spontaneous release of PRL, while GnRH appears capable of modulating the effects of PRL-regulated LH and PRL gene expression and spontaneous release.

Increased egg production by active immunization against vasoactive intestinal peptide in the turkey (Meleagris gallopavo).

Vasoactive intestinal peptide (VIP) is a hypothalamic prolactin (PRL)-releasing factor in the turkey. The hypothesis in the present study was that active immunization of turkeys with VIP would neutralize endogenous VIP, decrease circulating PRI, and consequently prevent the expression of incubation behavior. Large white female turkeys were divided into three experimental groups comprising untreated controls, control turkeys immunized with keyhole limpet hemocyanin (KLH), and turkeys immunized with synthetic chicken VIP conjugate (KLH-cVIP). Each turkey received four injections at 4-wk intervals, starting on the day of photostimulation. The immune response, measured by the percentage binding of monoiodinated chicken VIP (cVIP) to plasma at a dilution of 1:1000, averaged 11.8 +/- 2.5% during the reproductive life cycle. Immunization against KLH-cVIP prevented the normal increases of PRL that are associated with the photo-induced reproductive cycle. Over a 21-wk period beginning at photostimulation, KLH-cVIP-immunized birds exhibited a maximal plasma PRL level of 82.2 +/- 29.5 ng/ml, compared to 367.7 +/- 66.6 ng/ml and 227.5 +/- 51.7 ng/ml for non- and KLH-immunized turkeys, respectively. The total number of nest visits per hen during the 147-day experimental period decreased from 320.0 +/- 48.2 in the nonimmunized controls to 180.7 +/- 53.7 and 149.4 +/- 13.1 visits in KLH- and KLH-cVIP conjugate-immunized turkeys. Turkeys that showed an immune response to KLH-cVIP immunization did not exhibit incubation behavior, whereas 54% and 33% of non- and KLH-immunized hens incubated their eggs.(ABSTRACT TRUNCATED AT 250 WORDS)

Theca cell cytochrome P450 17-hydroxylase and aromatase messenger ribonucleic acid abundance and serum steroid levels during follicular atresia associated with incubation behavior in the domestic turkey hen.

This study was designed to examine changes in cytochrome P450 17 alpha-hydroxylase (C17) and aromatase (ARO) mRNA contents in the theca layer of preovulatory follicles (POF) as turkey hens transit from egg laying to incubation. Hens were grouped into the following categories: 1) laying hens--laid one egg per day and nested 1-2 times per day; 2) transitional hens--laid one egg per day and nested > 4 times per day; and 3) Day 1, Day 3, and Day 5 incubating hens--laid no eggs for 2, 4, or 6 days, respectively, and nested > 4 times per day. Small white follicles (SWF) and the theca layer from the largest (F1) and the third (F3), fifth (F5), and seventh (F7) largest POF were dispersed and challenged with testosterone (T) for 5 h. Relative levels of C17 and ARO mRNA were examined from the theca layers of F1, F3, F5, F7, and SWF. The number of atretic follicles increased from 0 (layers) to 8 (Day 5 incubating hens). Serum LH, progesterone (P), and estradiol (E), but not T, declined on Day 1 of incubation. Basal levels of P, T, and E from theca and SWF cells declined in incubating hens. Both basal and T-stimulated theca and SWF production of E decreased in incubating hens. C17 and ARO mRNA declined in SWF, F7, and F5 during follicular atresia. It is suggested that reduced gene expression of ovarian steroidogenic enzymes may be a partial determinant of reduced circulating sex steroid levels in incubating hens.(ABSTRACT TRUNCATED AT 250 WORDS)

Active immunization with vasoactive intestinal peptide prevents the secretion of prolactin induced by electrical stimulation of the turkey hypothalamus.

Immunoneutralization of endogenous vasoactive intestinal peptide (VIP) by active immunization with chicken VIP (cVIP) reduced both basal circulating prolactin (PRL) and steady-state levels of pituitary PRL mRNA in the turkey. This immunoneutralization severely curtailed the plasma PRL response induced by infusion of cVIP into the median eminence, and totally blocked the plasma PRL release effected by electrical stimulation of the medial preoptic nucleus. This is the first demonstration that a stimulated PRL secretion can be blocked by neutralizing VIP availability. These findings imply that among the neurochemicals released by electrical stimulation, only VIP directly stimulates PRL secretion. In addition to serving as a PRL releasing factor, VIP also appears to be involved in the regulation of pituitary PRL mRNA expression.

Role of vasoactive intestinal peptide in the control of prolactin-induced turkey incubation behavior. I. Acute infusion of vasoactive intestinal peptide.

Vasoactive intestinal peptide (VIP) stimulates prolactin (PRL) secretion. Ovine PRL induces incubation behavior in avian species. This study was designed to determine whether VIP can elevate plasma PRL for up to 3 h. Saline or porcine VIP (pVIP; 30, 60, or 150 ng/min) was infused into the median eminence of laying turkeys for 1 h. The 60- and 150-ng doses of pVIP increased plasma PRL (p < 0.01), whereas the 30-ng dose was insignificant. Pituitary PRL content decreased in pVIP-treated turkeys. Two-hour infusion of 60 or 150 ng chicken VIP (cVIP)/min produced similar elevations of plasma PRL (p < 0.001), which declined within 80 min. Both treatments induced insignificant increases in pituitary PRL mRNA. Saline or cVIP (30, 60, or 60 [pulsed] ng/min) was infused into the median eminence for 3 h. Sixty ng cVIP/min induced the largest PRL release (p < 0.05). The pulsatile and low-cVIP treatments resulted in release of a significant amount of PRL in comparison to the saline treatment (p < 0.01). All cVIP treatments resulted in decreased pituitary PRL content (p < 0.05). The 60-ng dose increased PRL mRNA (p < 0.1). This study shows that 60 ng VIP/min causes the maximum PRL release in laying turkeys. However, pituitary PRL content is depleted and PRL synthesis cannot maintain PRL secretion at high levels.

Role of vasoactive intestinal peptide in the control of prolactin-induced turkey incubation behavior. II. Chronic infusion of vasoactive intestinal peptide.

Hyperprolactinemia is associated with incubation behavior in avian species. Increased nesting activity is a major indication of incubation behavior. Vasoactive intestinal peptide (VIP) stimulates prolactin (PRL) secretion from the anterior pituitary. The goal of this study was to induce incubation behavior by stimulating PRL through chronically infusing VIP into the third ventricle of turkey brains. In experiment 1, porcine VIP (pVIP) was infused into the median eminence at a rate of 60 ng/min for 7 days by means of osmotic pumps implanted s.c.. Plasma PRL increased significantly in the pVIP-treated turkeys (p < 0.001). Although egg laying was not affected by the pVIP infusion, the mean oviduct weight decreased (p < 0.057). In experiment 2, saline or pVIP (30 or 60 ng/min) was infused into the third ventricle of laying turkeys for 12 days. Both pVIP treatments increased plasma PRL for 9 days (p < 0.05). The 30-ng pVIP/min infusion decreased nesting activity, plasma LH, ovary and oviduct weight, hypothalamic GnRH I, and anterior pituitary VIP receptors (p < 0.1). However, ovine PRL infusion (20.8 ng/min) into the same turkey flock increased nesting activity (p < 0.01). In conclusion, pVIP does not induce incubation behavior in laying turkeys.

Hypothalamic and posterior pituitary content of vasoactive intestinal peptide and gonadotropin-releasing hormones I and II in the turkey hen.

An inverse relationship often exists between prolactin (Prl) and LH in avian species. Our study was designed to investigate the relationship between hypothalamic and posterior pituitary vasoactive intestinal peptide (VIP)--an endogenous Prl-releasing peptide--and chicken GnRH-I and GnRH-II. Hypothalamic VIP content was increased after photostimulation, reaching its highest levels in incubating and photorefractory birds. The highest hypothalamic GnRH-I content was in laying hens followed by that in photostimulated and incubating birds. The lowest levels were in the nonphotostimulated birds. Hypothalamic GnRH-II increased after photostimulation, then fell to nonphotostimulated levels during incubation and photorefractoriness. Posterior pituitary VIP content was elevated in response to photostimulation, reaching a peak value in the laying and incubating birds, then declining in the photorefractory hens. Posterior pituitary GnRH-I and GnRH-II content peaked in the incubating birds. Ovariectomy caused a significant reduction in hypothalamic GnRH-I and GnRH-II with no effect on VIP; no changes were found in the posterior pituitary due to ovariectomy. Reducing day length caused a significant decrease in the hypothalamic and the posterior pituitary content of VIP and GnRH-I, and GnRH-II. Ovine Prl (oPrl) administration to laying hens reduced the hypothalamic VIP and GnRH-I and GnRh-II content. Posterior pituitary GnRH-I content was also reduced. Although GnRH-II levels were reduced by Day 4 of injections, they rose to peak levels by Day 14 of oPrl administration. Posterior pituitary VIP content was not altered by oPrl.

Dynorphin modulates prolactin secretion in the turkey.

Big dynorphin (prodynorphin 209-240), dynorphin A (prodynorphin 209-225), dynorphin B (prodynorphin 228-240), beta-endorphin (beta-lipotrophin 61-90), or Met-enkephalin, each infused into the third ventricle, were tested for their effect on PRL release in the anesthetized turkey hen. Laying hens that received big dynorphin at the rate of 0.35 nmol/min showed a 5.1-fold increase in serum PRL at the end of a 30-min infusion period. In a second experiment, the big dynorphin-induced PRL increase was 2.6-fold. Nest-deprived, previously incubating hens that received big dynorphin displayed an 8.2-fold increase in serum PRL. Laying and nest-deprived incubating control birds infused with saline displayed no PRL increases. Laying hens that received dynorphin A (0.35 nmol/min) showed a 1.5-fold increase in serum PRL after 30 min of infusion; after 40 min of infusion, this increase rose to 2.7-fold. Infusions of beta-endorphin (0.35 nmol/min), or Met-enkephalin (0.35 nmol/min) failed to evoke PRL increases in either laying or nest-deprived incubating turkeys. Infusion of big dynorphin or dynorphin A for 120 min maintained an elevated PRL level across the period, a level equal to that evoked by electrical stimulation of the medial preoptic nucleus (ES/POM). Infusion of dynorphin B (0.48 nmol/min) or a reduced dose of dynorphin A (0.09 nmol/min) augmented the PRL response evoked by ES/POM. No augmentation was noted for beta-endorphin or Met-enkephalin, nor for saline-infused controls. The dynorphin-induced PRL response appeared to be dose-dependent.(ABSTRACT TRUNCATED AT 250 WORDS)

Ovarian involvement in the suppression of luteinizing hormone in the incubating turkey (Meleagris gallopavo).

A series of experiments were conducted to elucidate the role of the ovary in incubation behavior-associated luteinizing hormone (LH) suppression. Ovariectomy (Ovx) increased (p < 0.05) serum LH levels in nonphotostimulated, laying and photorefractory turkeys but not in incubating birds (p > 0.05). Ovx had no effect on intramuscularly injected mammalian luteinizing hormone-releasing hormone (4 micrograms/kg i.m.)-induced LH in incubating hens, but enhanced the LH response in laying hens. Serum LH and prolactin (Prl) were unaffected by Ovx, and nest deprivation (ND) decreased Prl levels (p < 0.05) with no effect on serum LH of incubating turkeys. However, serum LH increased (p < 0.05) and Prl decreased in Ovx-ND birds. Prl mRNA abundance (11.9 +/- 1.2 ng/microgram total RNA) decreased following Ovx (3.4 +/- 0.4 ng/microgram total RNA) or ND (3.6 +/- 0.5 ng/microgram total RNA). Nest deprivation increased LH beta mRNA (2.5-fold) which was further increased (4.8-fold) by Ovx. Hypothalamic GnRH-I and GnRH-II contents increased (p < 0.05) in Ovx-ND turkeys. We conclude that serum LH suppression during incubation behavior requires ovarian participation acting synergistically with elevated Prl and/or nesting stimulus on hypothalamic GnRH, and that the concentration fo LH beta mRNA may be a limiting factor in LH secretion.

Effect of ovine prolactin administration on hypothalamic vasoactive intestinal peptide (VIP), gonadotropin releasing hormone I and II content, and anterior pituitary VIP receptors in laying turkey hens.

The effect of ovine prolactin (oPRL) administration to laying and incubating turkey hens was studied. In experiment one, five treatment groups (n = 6) of laying hens received injections of vehicle or oPRL (4 mg/bird/day) for 2, 4, 8, or 14 days. Hypothalamic vasoactive intestinal peptide (VIP) content decreased to its lowest level (p < 0.05) by Day 2 of oPRL injection. Serum turkey PRL decreased by Day 8 of injection, reaching nadir by Day 14. Similar to the decline in hypothalamic VIP content, the maximal decline in the number of anterior pituitary VIP binding sites was achieved by Day 2 of oPRL administration. Hypothalamic GnRH-I content decreased after 2 days of oPRL injection and remained low through Day 14 of the experiment. GnRH-II levels declined with time, reaching significantly lower levels after 8 days of injections, and remained low through Day 14. Plasma LH levels also declined (p < 0.05) after Day 14 of oPRL administration. In experiment 2, two groups (n = 6) of incubating birds were used: controls receiving injections of vehicle only and birds receiving injections of oPRL (4 mg/bird/day) for 10 days. Exogenous oPRL had no effect on hypothalamic VIP, GnRH-I or II, anterior pituitary VIP binding sites, or plasma turkey PRL or LH. The findings suggest that in laying hens, PRL inhibits its own secretion by acting on both the hypothalamus and the anterior pituitary; this phenomenon does not occur in incubating birds. Furthermore, we provide evidence that pRL acts centrally to reduce LH levels by reducing GnRH levels in the hypothalamus.

Effect of reproductive condition on luteinizing hormone and prolactin release induced by electrical stimulation of the turkey hypothalamus.

In the turkey, the onset of incubation behavior is associated with decreased luteinizing hormone (LH) and increased prolactin (PRL). This study was designed to clarify the contribution of the hypothalamus and the anterior pituitary to the changes in plasma LH during the reproductive cycle of the turkey. Plasma LH and PRL were measured in anesthetized turkey before, during, and after electrical stimulation in the median eminence. In one experiment, luteinizing hormone releasing hormone (LHRH; 4 micrograms/kg) was injected intramuscularly 30 min after termination of electrical stimulation, and blood samples were obtained 5, 10, 20, and 30 min after injection. Electrical stimulation in the median eminence significantly increased (P < 0.05) plasma LH of laying (LAY), nest-deprived, previously incubating (NEST DEP), and photorefractory (REFRAC) hens, but not of photosensitive short-day (SHORT DAY) birds (P > 0.05). Plasma LH of LAY hens peaked at 4.06 +/- 0.78 ng/ml from a prestimulation baseline of 2.30 +/- 0.21 ng/ml and that in NEST DEP birds increased from 1.08 +/- 0.18 ng/ml to 2.57 +/- 0.53 ng/ml. Administration of LHRH increased plasma LH levels in SHORT DAY, LAY, and NEST DEP hens with the increase being 2.0-, 2.5-, and 6.1-fold, respectively. Electrical stimulation in the median eminence increased plasma PRL (P < 0.05) in all the reproductive groups tested, with peak response being greatest for NEST DEP birds (661 +/- 126 ng/ml) followed by LAY (317 +/- 26 ng/ml), REFRAC (50 +/- 7 ng/ml), and SHORT DAY (39 +/- 12 ng/ml) hens.(ABSTRACT TRUNCATED AT 250 WORDS)

Complementary DNA cloning and expression of Pit-1/GHF-1 from the domestic turkey.

The transition from egg laying to incubation activity in birds is associated with a dramatic rise in serum prolactin levels. To further our understanding of the regulation of prolactin gene expression in birds, a cDNA clone encoding turkey Pit-1/GHF-1 was isolated. The turkey cDNA, designated tPit-1/GHF-1, was 1,123 nucleotides in length and encoded a protein of 327 amino acids, including a conserved 80-amino-acid POU-specific domain and a 60-amino-acid POU homeodomain. tPit-1/GHF-1 POU-specific domain and POU-homeodomain showed 94-95% amino acid identity with the corresponding rat Pit-1/GHF-1 domains. At its amino terminus, tPit-1/GHF-1 contained a 26-amino-acid insertion comparable to that found in the rat variant isoform, Pit-1 beta. Two other insertions of 38 and 7 amino acids were present and were not found in the mammalian protein. Levels of tPit-1/GHF-1 mRNA in pituitary tissue were examined at different phases of the turkey reproductive cycle by Northern blotting. tPit-1/GHF-1 mRNA was expressed as a 3.5-kb transcript, whose abundance remained relatively constant throughout the reproductive cycle. Thus, the dramatic rise in prolactin mRNA, observed during hyperprolactinemia in incubating turkey hens, was not associated with a concomitant increase in tPit-1/GHF-1 gene expression.

Pituitary luteinizing hormone and prolactin messenger ribonucleic acid levels are inversely related in laying and incubating turkey hens.

The relationships of prolactin (PRL) and LH messenger (m) RNA to serum and pituitary content were determined for turkey hens at different phases of the reproductive cycle. In the nonphotostimulated, reproductively inactive hen, serum and pituitary PRL content and pituitary PRL mRNA levels were low. All three PRL values rose after photostimulation and peaked during the incubation phase. Relative to nonphotostimulated hens, hyperprolactinemic incubating hens showed 220-, 11-, and 57-fold increases in serum PRL, pituitary PRL content, and pituitary PRL mRNA levels, respectively. These peak levels declined 80-, 3-, and 6-fold, respectively, in photorefractory hens. In contrast to PRL levels, serum LH, pituitary LH, and pituitary LH beta-subunit mRNA levels did not change as dramatically. Serum LH showed no significant changes for the different reproductive phases. Pituitary LH peaked after photostimulation and declined to its lowest level in incubating hens. Pituitary LH-beta mRNA abundance was highest in photostimulated and laying hens and lowest in incubating and photorefractory hens. These results demonstrate that the abundance of LH-beta and PRL mRNA shows an inverse relationship in photostimulated/laying and incubating turkey hens.

Evidence of a role for the turkey posterior pituitary in prolactin release.

The aims of this study were: (1) to examine whether the posterior pituitary contains prolactin releasing factor (PRF) activity, (2) to determine to what extent known neurohypophyseal peptides contribute to this activity, and (3) to compare posterior pituitary PRF activities of hens in different reproductive stages. Anterior pituitary cells derived from juvenile female turkeys were incubated with posterior pituitary extracts or test substances for 3 hr. Posterior pituitary extracts (0.1-0.8 equivalent) contained a potent substance(s) which stimulated PRL release in a concentration-dependent manner (2.4 +/- 0.08 to 6.5 +/- 0.23 micrograms/500 k cells). Arginine vasotocin (AVT) and vasoactive intestinal peptide (VIP) antisera (1:500) completely abolished the PRL-releasing activities of their respective peptides but partially reduced (P less than 0.05) the PRF activity of the posterior pituitary (AVT, 19.9%; VIP, 55.1%). Mesotocin antiserum did not alter (P greater than 0.05) PRL release induced by posterior pituitary extract. Posterior pituitary extract (0.01-0.5 equivalent) from hens in each of the various stages of the reproductive cycle induced a concentration dependent PRL release. The 0.5 posterior pituitary equivalent dose from reproductively quiescent (nonphotostimulated), laying, photorefractory, and incubating hens increased PRL release 2.4-, 2.9-, 3.8-, and 11.1-fold, respectively. The turkey posterior pituitary contains a potent PRF activity, partially accounted for by VIP and AVT, at the assayed concentrations, which varies with the reproductive cycle.

Involvement of dopamine in prolactin release induced by electrical stimulation of the hypothalamus of the female turkey (Meleagris gallopavo).

Controversy exists regarding the role of dopamine (DA) in the regulation of avian prolactin (PRL) secretion. Consequently, we injected apomorphine, a DA agonist, and pimozide, a DA receptor blocker, into laying and nest-deprived incubating turkeys and studied their effect on PRL secretion before (-20, -10, 0 min), during (5, 10, 20, 30 min), and after (5, 15, 30 min) electrical stimulation in the ventromedial nucleus of the hypothalamus. Apomorphine (10 mg/kg, ip) completely abolished the electrical stimulation-induced PRL increase in both laying and nest-deprived incubating hens. Pimozide (2 mg/kg, ip) potentiated electrical stimulation-induced PRL secretion in laying hens. In the two pimozide experiments, peak responses were 10.9-fold for the pimozide-treated group vs 2.9-fold for the control group, and 5.4-fold for the pimozide-treated group vs 2.6-fold for the control group. In nest-deprived incubating hens, PRL response to electrical stimulation was unaffected by pimozide treatment. These data support the concept that DA is inhibitory to the neuroendocrine system which stimulates PRL secretion in laying hens. In incubating hens, the dopaminergic inhibition is diminished, allowing for the increased PRL level observed during incubation.