Thesis (M.S.)--Florida State University, 2005. / Advisor: J. Michael Overton, Florida State University, College of Human Sciences, Dept. of Nutrition, Food, and Exercise Sciences. Title and description from dissertation home page (viewed Jan. 26, 2006). Document formatted into pages; contains vii, 34 pages. Includes bibliographical references.

Thesis (M.S.)--West Virginia University, 2005. / Title from document title page. Document formatted into pages; contains vi, 84 p. : ill. Vita. Includes abstract. Includes bibliographical references (p. 76-83).

Thesis--Wisconsin. / Vita. Includes bibliographical references (leaves 44-48).

Two forms of gonadotropin-releasing hormone (GnRH) were extracted from brain-pituitary tissues of two species of Thai catfish, Clarias inacrocephalus and C. batrachus. The peptides were detected using high-performance liquid chromatography (HPLC) and radioimmunoassay (RIA), The amino acid sequences of both forms were determined using Edman degradation. One form of GnRH in the brain-pituitary tissues of the Thai catfish was novel, whereas the second form of GnRH was identical to chicken GnRH-XI. The presence of the N-terminal pGlu residue in both peptides was established by digestion with pyroglutamyl aminopeptidase. In addition, catfish GnRH-I was studied by mass spectrometry. The localization of these two peptides was determined to be in the discrete brain areas and in the pituitary of female and male catfish, C. macrocaphalus, using heterologous and homologous radioimmunoassays. Initially a heterologous RIA was used with mammalian GnRH as iodinated tracer and standard, and an antiserum made against salmon GnRH. Catfish GnRH-I (novel form) was found in most areas of the female and male brain with the highest content and concentration in the female pituitary and in the male hypothalamus,, Catfish GnRH-II (chicken GnRH-II) was found with the highest content in the female Cerebellum and highest concentration in the pituitary, catfish GnRH-II (chicken GnRH-II) was found with the highest content and concentration for males in the same area, hypothalamus. Additionally, a homologous RIA was used with catfish GnRH-II (chicken GnRH-II) as iodinated tracer and standard, and an antiserum prepared against chicken GnRH-II. Catfish GnRH-II was detected with the highest content and concentration in the cerebellum of both sexes. These values are higher than the results obtained in the heterologous assay. The location of catfish GnRH-I suggests that it plays a role in regulating the release of gonadotropin from the pituitary since the high content and concentration of this immunoreactive GnRH are detected in the hypothalamus and pituitary gland. In contrast, catfish GnRH-II may act as a neurotransmitter in the catfish brain, in particular in tha cerebellum where a high content and concentration of immunoreactive GnRH are detected. Physiological in vivo studies indicate that catfish GnRH-II is more effective than catfish GnRH-I and other forms of GnRH such as mammalian and dogfish GnRH for induction of ovulation in catfish, C. macrocephalus. Eight GnRH analogs had varying potencies for the induction of ovulation, but the most effective forms were two forms of catfish GnRH-II (chicken GnRH-II) modified in positions six and ten. In vitro studies found that catfish GnRH-I not only causes the release of gonadotropin but also the release of growth hormone in a dose-dependent manner. The primary structures of the two catfish GnRH peptides are important for understanding the evolution of this family peptide. The novel catfish GnRH shows that only positions 5, 7 and 8 vary in the GnRH molecule in jawed vertebrates, whereas catfish GnRH-II provides direct evidence that the structure of this GnRH is conserved in teleosts. / Graduate

Growth hormone-releasing hormone (GHRH) is a neuropeptide which stimulates the synthesis and release of growth hormone (GH) from the pituitary gland. The primary structure of this peptide has been identified in 7 mammalian species while the gene has been isolated from only rat and human. GHRH is a member of the glucagon superfamily which includes vasoactive intestinal peptide (VIP), glucagon, secretin, peptide histidine methionine (PHM), gastric inhibitory peptide (GIP) and a recently identified peptide, pituitary adenylate cyclase activating polypeptide (PACAP). The evolutionary relationships of this superfamily are not well understood because the gene structure of these molecules has only been identified in mammals. This thesis presents immunological evidence of a GHRH-like molecule, and identifies a GHRH/PACAP precursor and gene that encode two peptides, a GHRH-like molecule structurally related to PACAP-related peptide (PRP) and PACAP, from sockeye salmon brain. An antiserum directed against a topologically assembled epitope of human GHRH 1-44 (NH2) was produced and used to develop a radioimmunoassay for detection of immunoreactive GHRH in brain extracts of salmon, guinea pig, mouse and alligator. An immunoreactive GHRH from salmon brain extracts with a retention time on reverse phase high-performance liquid chromatography (HPLC) distinct from human GHRH was present. In alligator, the same antiserum also detected a GHRH-like molecule. During attempts to purify alligator GHRH, alligator brain neuropeptide Y (NPY) was identified. Alligator NPY is the first non-mammalian vertebrate to have 100% sequence identity to human NPY. The sequence identity between alligator and human NPY suggests that this sequence is the same as the ancestral amniote NPY. Molecular biological techniques were used for the structural identification of the salmon GHRH-like molecule and another peptide. The salmon GHRH/PACAP precursor contains 173 amino acids and has dibasic and monobasic processing sites for cleavage of a 45 amino acid GHRH-like peptide with a free acid C-terminus and a 38 amino acid PACAP with an amidated C-terminus. The salmon GHRH-like peptide has 40% amino acid sequence identity with the human GHRH and 56% identity with human PACAP-related peptide (PRP). Salmon PACAP-38 is highly conserved (89%) with only 4 amino acid substitutions compared with the human, ovine and rat PACAP-38 peptides. Nucleotide sequencing and use of the polymerase chain reaction show the exon/intron organization of the salmon GHRH/PACAP gene to be similar to the human PACAP gene. Unlike the mammalian PACAP genes, the salmon gene produces two precursor forms by post-transcriptional processing. One form is similar to the mammalian PACAP precursors, while the second form is shorter due to the excision of exon 4. This deletion results in the loss of the first 32 amino acids of the GHRH-like peptide from the precursor. The high sequence identity and structural organization between the GHRH(PRP)/PACAP and PHM(PHI)/VIP genes suggest a duplication event occurred in an ancestral gene after the divergence from the other glucagon superfamily members. GHRH in mammals may have arisen by gene duplication after the divergence of the tetrapods from the other vertebrate lines. Thus, GH in fish may be controlled by the two molecules, GHRH-like peptide and PACAP, located on a single GHRH/PACAP gene. / Graduate

Observing cows in estrus and inseminating them at the optimal time are necessary steps for effective reproductive management of a dairy herd. However, increasing herd sizes can lead to reproductive inefficiency resulting in decreased profits on dairy herds. Synchronization of estrus, through pharmacological control, has been used to improve reproductive efficiency. Systematic breeding programs provide an organized approach for administering artificial insemination (AI) at first service. Moreover, reproductive management is based on a methodical approach for the entire herd rather than for the individual cow. Seven-hundred and thirty four Holstein cows from 16 commercial dairy herds were used to conduct this study evaluating three systematic breeding protocols; 14-d PGF2a, timed AI (TAI), and GnRH-PGF2α, in comparison with an untreated control group. Eight herds relied on visual observation as their primary method for detection of estrus, and 8 herds utilized the HeatWatch® (DDx, Inc., Denver, CO) electronic estrus detection system. The average days to first postpartum AI were longer for untreated control cows when compared to the other breeding protocols. First AI conception rates did not differ among control, 14-d PGF2a, or GnRH-PGF2a protocols, but were higher than the TAI protocol. However, first AI pregnancy rates were higher for untreated controls versus hormonally treated cows. Estrus characteristics associated with each protocol were also evaluated and no difference was detected across treatments. An economic analysis determining cost per pregnancy for each protocol when considering drug costs, and pregnancy rates, resulted in the highest cost per pregnancy for TAI followed by GnRH-PGF2a and 14-d PGF2a. These programs should be considered as tools for convenience and efficiency of estrus detection; however, reduced labor costs from less time spent on estrus detection may be offset by the cost of the drug protocols. Cost effectiveness must be calculated on an individual herd basis when deciding whether a systematic breeding program is the appropriate choice. / Master of Science

The objective of these studies was to contribute to the understanding and assessment of thyroid function in horses. The first study evaluated methods of assessing thyroid function in horses, including validation of an enzyme immunoassay (EIA) for measuring equine thyroid hormones and development and assessment of a thyrotropin releasing hormone (TRH) response test. Our data indicated that EIA is an acceptable method for the measurement of total (T) and free (F) thyroxine (T4) and triiodothyronine (T3) in equine plasma. Its measurements are not equivalent to values obtained by radioimmunoassay (RIA), but they can be calibrated to predict corresponding RIA values. A protocol was developed for TRH response tests involving administration of 1 mg TRH intravenously, with blood sample collection immediately before, 2.5, 5.0, and 24 h after administration. Analysis of plasma TT4, FT4, TT3, and FT3 revealed that the magnitude of hormone response was best approximated by the area under the curve of hormone plotted against time and by the absolute change in thyroid hormone concentration. Baseline concentrations, peak concentrations, and percent of baseline values were not as well able to predict the magnitude of hormone response. The second study assessed the effects of exercise and feed composition on thyroid status. Thirteen mature Arabian geldings, adapted to either a high sugar and starch (SS) or high fat and fiber (FF) feed, underwent 15 wk of exercise training followed by a treadmill exercise test. The TRH response tests performed before training, after training, and the morning after the exercise test revealed that the exercise test decreased the TT4 and FT4 response, whereas feeding of high levels of sugars and starches increased the response of TT3 and FT3. During the first four weeks of training, increased TT4 and FT4 concentrations occurred simultaneously with increased nonesterified fatty acid concentrations, decreased triglyceride concentrations, and increased insulin sensitivity. The increase in TT4 and FT4 may have provided the cellular signaling necessary for increased lipolysis and insulin sensitivity. These metabolic changes facilitate the increases in lipid and carbohydrate metabolism that are needed to fulfill the additional energy requirements of regular exercise. The third study assessed thyroid status in ponies with different laminitic histories. Total T4, FT4, TT3, and FT3 were measured during March and May 2004 in 126 ponies that were categorized as either previously laminitic (PL; n = 54) or never laminitic (NL; n = 72) and evaluated for current laminitis in May (CL; n = 13). Decreased concentrations of TT4 and FT4 were found in PL ponies when compared to NL ponies in March (P = 0.018, 0.020) and May (P = 0.018, 0.001). However, TT4 and FT4 concentrations in CL ponies were not different than concentrations in NL ponies in May (P = 0.82, 0.72), and when retrospectively separated out in March, were not different than NL ponies (P = 0.90, 0.84). Therefore, basal thyroid hormone concentrations are not useful as a predictor or hormonal characteristic of pasture-associated laminitis. The decreased TT4 and FT4 in PL ponies may be an indication of a response or compensation to laminitis and may facilitate the metabolic changes necessary to cope with the disease. / Master of Science

Thesis (Ph. D.)--West Virginia University, 2001. / Title from document title page. Document formatted into pages; contains xii, 150 p. : ill. Vita. Includes abstract. Includes bibliographical references (p. 131-150).

One of the leading experts in the field of Neuroendocrinology and Neuroimmunmodulation, Samuel Mac Donald McCann, known by all his friends as ‘Don’, passed away in 2007. This article pays tribute to his outstanding scientific contribution and a glimpse on his fascinating personality. A member of the National Academy of Sciences of the United States and pioneer in the field of neuroendocrine regulation, he identified numerous hormones and peptides and set the stage for basic concepts in physiology and clinical medicine. / Dieser Beitrag ist mit Zustimmung des Rechteinhabers aufgrund einer (DFG-geförderten) Allianz- bzw. Nationallizenz frei zugänglich.

[Truncated abstract] It is becoming increasingly obvious that cell signalling pathways are more complicated than we originally perceived. Research is revealing that, not only is there a multitude of new proteins involved in signalling cascades, but also that previously identified proteins may have additional, alternate roles in intracellular trafficking. Gonadotropin-releasing hormone (GnRH) in conjunction with its receptor (GnRHR), the primary regulator of reproduction in all species, is no exception. In the past few years it has become readily accepted that the classic linear GnRHR-Gαq/11 signalling pathway is not universal and that this receptor is involved in a far greater range of cellular activities than was previously considered. In particular, it is widely accepted that continuous administration of GnRH analogs results in an inhibition of growth of a number of reproductive-derived tumours and that this may, in part, be mediated by direct activation of GnRHs expressed on these cells. However, it is not fully understood how the GnRHR mediates these growth effects or whether such effects are unique to reproductive-derived cancer cells. Research within this thesis aimed to determine how the presence or absence of this receptor in different cell types might affect the ability of GnRH to directly mediate growth effects. We demonstrate that continuous treatment with a GnRH agonist (GnRHA) induces an anti-proliferative effect in a gonadotropederived cell line (LβT2) and also in HEK293 cells stably expressing either the rat or human GnRHR. The anti-proliferative effect was time- and dose-dependent and was specifically mediated via the GnRHR, as co-treatment of the GnRHRexpressing cell lines with a GnRH antagonist blocked the growth suppressive effect induced by GnRHA treatment. Cell cycle analysis revealed that the GnRHA treated HEK/GnRHR cell lines induced an accumulation of cells in the G2/M phase while a G0/G1 arrest was observed in LβT2 cells. Previous identification by our group of a potential interaction between the GnRHR and the transcription factor E2F4, an integral cell cycle regulatory protein, prompted further investigation as to the nature of this interaction. Bioluminescence energy transfer (BRET) was utilised to demonstrate that the GnRHR also interacts with E2F5, another member of the E2F family of cell cycle proteins that shares a high level of homology to E2F4. In addition, it was determined that the interaction between human GnRHR and E2F4, detected using BRET, was influenced by cell density.