C. Richard Barb

Central and peripheral administration of kisspeptin activates gonadotropin but not somatotropin secretion in prepubertal gilts

It is well established that kisspeptin signaling is necessary for the onset of puberty in laboratory animals. However, the role that kisspeptin may have in regulating puberty in large domestic animals is unknown. We tested the hypothesis that either central or peripheral infusion of kisspeptin would stimulate gonadotropin and GH secretion in prepubertal gilts. In experiment 1, prepubertal gilts were fitted with i.c.v. cannula and indwelling jugular catheters. Animals were randomly assigned to receive 0, 10, or 100 microg kisspeptin in saline. In experiment 2, prepubertal gilts, fitted with indwelling jugular catheters, randomly received 0, 1, 2.5, or 5 mg kisspeptin in saline intravenously. Serial blood samples were collected every 15 min for 3 h before and 5 h after infusions, and serum concentrations of LH, FSH, and GH were determined. Mean concentrations of LH and FSH remained at basal levels for control animals but were increased (P<0.001) for animals receiving i.c.v. infusion of kisspeptin. Area under the LH and FSH curves following i.c.v. infusion of kisspeptin increased (P<0.001) in a dose-dependent manner. Concentrations of GH were unaffected by i.c.v. treatment. Peripheral administration of kisspeptin increased (P<0.05) serum concentrations of LH but not FSH or GH. Thus, kisspeptin can activate gonadotropic but not somatotropic hormone secretion in prepubertal gilts. The present data support the concept that kisspeptin plays a role in the mechanism involved in initiating puberty in swine.

Gene expression profiling of the short-term adaptive response to acute caloric restriction in liver and adipose tissues of pigs differing in feed efficiency

Residual feed intake (RFI) is a measure of feed efficiency, in which low RFI denotes improved feed efficiency. Caloric restriction (CR) is associated with feed efficiency in livestock species and to human health benefits, such as longevity and cancer prevention. We have developed pig lines that differ in RFI, and we are interested in identifying the genes and pathways that underlie feed efficiency. Prepubertal Yorkshire gilts with low RFI (n = 10) or high RFI (n = 10) were fed ad libitum or fed at restricted intake of 80% of maintenance energy requirements for 8 days. We measured serum metabolites and hormones and generated transcriptional profiles of liver and subcutaneous adipose tissue on these animals. Overall, 6,114 genes in fat and 305 genes in liver were differentially expressed (DE) in response to CR, and 311 genes in fat and 147 genes in liver were DE due to RFI differences. Pathway analyses of CR-induced DE genes indicated a dramatic switch to a conservation mode of energy usage by down-regulating lipogenesis and steroidogenesis in both liver and fat. Interestingly, CR altered expression of genes in immune and cell cycle/apoptotic pathways in fat, which may explain part of the CR-driven lifespan enhancement. In silico analysis of transcription factors revealed ESR1 as a putative regulator of the adaptive response to CR, as several targets of ESR1 in our DE fat genes were annotated as cell cycle/apoptosis genes. The lipid metabolic pathway was overrepresented by down-regulated genes due to both CR and low RFI. We propose a common energy conservation mechanism, which may be controlled by PPARA, PPARG, and/or CREB in both CR and feed-efficient pigs.

Developmental Changes in the Long Form Leptin Receptor and Related Neuropeptide Gene Expression in the Pig Brain

Abstract The hypothalamus is the key site of central regulation of energy homeostasis, appetite, and reproduction. The long form leptin receptor (Ob-Rl) is localized within the hypothalamus along with several neuropeptides that are involved in regulation of the neuroendocrine axis. In the present study, developmental changes in gene expression of the Ob-Rl, preproorexin, proopiomelanocortin (POMC), corticotropin releasing factor (CRF), somatostatin, and GnRH in the hypothalamus was studied. Expression of Ob-Rl and neuropeptide mRNA was examined by semiquantitative reverse transcription-polymerase chain reaction in hypothalami collected from 106-day-old fetus (n = 3) and 7-day-old (n = 3), 3.5-mo-old (n = 3), and 6-mo-old (n = 2) gilts. In addition, leptin mRNA expression in the first three ages was examined in back fat. Leptin mRNA expression increased (P < 0.05) by 7 days postnatal, but Ob-Rl mRNA expression increased (P < 0.01) by 3.5 mo. Expression of preproorexin (P < 0.05), somatostatin, and GnRH (P < 0.01) mRNA peaked by 3.5 mo of age while POMC mRNA expression increased markedly (P < 0.01) by 6 mo of age. The CRF mRNA expression did not change across ages. These findings suggest a possible relationship among Ob-Rl and a number of hypothalamic and peripheral peptides in the development of the neuroendocrine axis. These peptides may serve as messengers that link mechanisms that regulate reproduction and energy balance.

Microarray gene expression profiles of fasting induced changes in liver and adipose tissues of pigs expressing the melanocortin-4 receptor D298N variant

Transcriptional profiling coupled with blood metabolite analyses were used to identify porcine genes and pathways that respond to a fasting treatment or to a D298N missense mutation in the melanocortin-4 receptor (MC4R) gene. Gilts (12 homozygous for D298 and 12 homozygous for N298) were either fed ad libitum or fasted for 3 days. Fasting decreased body weight, backfat, and serum urea concentration and increased serum nonesterified fatty acid. In response to fasting, 7,029 genes in fat and 1,831 genes in liver were differentially expressed (DE). MC4R genotype did not significantly affect gene expression, body weight, backfat depth, or any measured serum metabolite concentration. Pathway analyses of fasting-induced DE genes indicated that lipid and steroid synthesis was downregulated in both liver and fat. Fasting increased expression of genes involved in glucose sparing pathways, such as oxidation of amino acids and fatty acids in liver, and in extracellular matrix pathways, such as cell adhesion and adherens junction in fat. Additionally, we identified DE transcription factors (TF) that regulate many DE genes. This confirms the involvement of TF, such as PPARG, SREBF1, and CEBPA, which are known to regulate the fasting response, and implicates additional TF, such as ESR1. Interestingly, ESR1 controls several fasting induced genes in fat that are involved in cell matrix morphogenesis. Our findings indicate a transcriptional response to fasting in two key metabolic tissues of pigs, which was corroborated by changes in blood metabolites, and the involvement of novel putative transcriptional regulators in the immediate adaptive response to fasting.

Prostaglandin-induced regression of porcine corpora lutea maintained by estrogen.

Corpora lutea were marked with suture in 24 crossbred gilts on day 7 to 9 of the estrous cycle (first day of estrus equals 0). All gilts were injected with 5 mg of estradiol benzoate (EB) daily from day 10 to 15 to extend the lifespan of corpora tutea, then the gilts were randomly assigned to two groups. On day 20, the 12 gilts of Group 1 were injected with 10 mg PGF-2ALPHA, and the 12 gilts of Group 2 were injected with saline. Ovaries were recovered 10 to 13 days after PGF-2ALPHA or saline injection. Ten gilts in Group 1 displayed estrus 5 plus or minus 0.7 days after PGF-2ALPHA injection, but only two gilts in Group 2 displayed estrus during the experimental period. In gilts that displayed estrus, all marked CL had regressed. Marked CL were still present in all 12 gilts that failed to exhibit estrus during the experimental period. These results show that in the pig, PGF-2ALPHA caused regression of CL that were maintained beyond the normal luteal phase of the estrous cycel by EB treatment.

N-Methyl-D, L-aspartate modulation of luteinizing hormone and growth hormone secretion from pig pituitary cells in culture

Administration of n-methyl-d, l-aspartate (NMA) to pigs in vivo increased GH and suppressed LH secretion. Cultures of anterior pituitary cells from pigs in the follicular phase (FOL; n = 3) and luteal phase (LUT; n = 3) of the estrous cycle, and ovariectomized (OVX; n = 10) pigs were treated with NMA (10(-4), 10(-6) or 10(-8) M) or the NMA antagonist, 2-amino-5-phosphonopentanoic acid (AP5; 10(-4), 10(-6) or 10(-8) M), to determine if NMA affects the pituitary directly. Secreted LH and GH were measured at 4 h after treatment. Basal LH and GH secretion (control; C) were 1.1 +/- 0.6, 4.4 +/- 2.1 and 5.6 +/- 1.3 ng/well and 5.2 +/- 1.2, 7.5 +/- 1.2 and 5.2 +/- 1.7 ng/well for FOL, LUT and OVX, respectively. Relative to C, 10(-4) M NMA increased (P < 0.001) LH secretion 2.4-, 2.2- and 5.1-fold in FOL, LUT and OVX cultures, respectively. The effect of 10(-4) M NMA was inhibited by 10(-4) M AP5 (P < 0.05) in FOL cultures, but not in OVX cultures. GnRH increased (P < 0.001) LH levels 3.1-, 2.3- and 3.8-fold in FOL, LUT and OVX cultures, respectively. Relative to C, 10(-4), 10(-6) and 10(-8) M NMA increased (P < 0.03) GH secretion 1.5-, 1.5- and 2.3-fold in LUT and 1.7-, 2.3- 2.0-fold in OVX cultures, respectively. AP5 alone or in combination with NMA failed to alter basal GH secretion.(ABSTRACT TRUNCATED AT 250 WORDS)

Prostaglandin F concentrations in utero-ovarian vein plasma of prepuberal and mature gilts.

Abstract Prostaglandin F (PGF) and progestins in utero-ovarian vein (UOV) plasma during the late luteal phase of the estrous cycle in unbred mature gilts and following induced ovulation in unbred prepuberal gilts were determined. Prepuberal gilts (120 to 130 days of age) were induced to ovulate with Pregnant Mare Serum Gonadotropin and Human Chorionic Gonadotropin (HCG). The day following HCG was designated as Day 0. Mature gilts which had displayed two or more estrous cycles of 18 to 22 days (onset of estrus = Day 0) were used. Polyvinyl catheters were inserted into the UOV of all gilts and blood was collected at 15 min intervals from 0800 to 1045 hr on Days 10 through 20 or Days 12 through 18. Plasma PGF concentrations in the mature gilts were elevated on Days 13, 14, 15, 16 and 17, whereas, plasma PGF concentrations in the prepuberal gilts were elevated only on Days 15, 16 and 17 resulting in a reproductive age (mature vs prepuberal) by day interaction (P

Gene expression in hypothalamus, liver, and adipose tissues and food intake response to melanocortin-4 receptor agonist in pigs expressing melanocortin-4 receptor mutations.

Transcriptional profiling was used to identify genes and pathways that responded to intracerebroventricular injection of melanocortin-4 receptor (MC4R) agonist [Nle(4), d-Phe(7)]-α-melanocyte stimulating hormone (NDP-MSH) in pigs homozygous for the missense mutation in the MC4R, D298 allele (n = 12), N298 allele (n = 12), or heterozygous (n = 12). Food intake (FI) was measured at 12 and 24 h after treatment. All pigs were killed at 24 h after treatment, and hypothalamus, liver, and back-fat tissue was collected. NDP-MSH suppressed (P < 0.004) FI at 12 and 24 h in all animals after treatment. In response to NDP-MSH, 278 genes in hypothalamus (q ≤ 0.07, P ≤ 0.001), 249 genes in liver (q ≤ 0.07, P ≤ 0.001), and 5,066 genes in fat (q ≤ 0.07, P ≤ 0.015) were differentially expressed. Pathway analysis of NDP-MSH-induced differentially expressed genes indicated that genes involved in cell communication, nucleotide metabolism, and signal transduction were prominently downregulated in the hypothalamus. In both liver and adipose tissue, energy-intensive biosynthetic and catabolic processes were downregulated in response to NDP-MSH. This included genes encoding for biosynthetic pathways such as steroid and lipid biosynthesis, fatty acid synthesis, and amino acid synthesis. Genes involved in direct energy-generating processes, such as oxidative phosphorylation, electron transport, and ATP synthesis, were upregulated, whereas TCA-associated genes were prominently downregulated in NDP-MSH-treated pigs. Our data also indicate a metabolic switch toward energy conservation since genes involved in energy-intensive biosynthetic and catabolic processes were downregulated in NDP-MSH-treated pigs.

Luteinizing Hormone Secretion in Hypophysial Stalk-Transected Pigs Given Progesterone and Pulsatile Gonadotropin-Releasing Hormone

Abstract This study was conducted to determine whether progesterone inhibits luteinizing hormone (LH) secretion in female pigs by a direct action on the pituitary gland. Eight ovariectomized, hypophysial stalk-transected gilts were given 1-μg pulses of gonadotropin-releasing hormone iv every 45 min from Day 0 to 12. On Days 5–12, each of four gilts received either progesterone or oil vehicle im at 12-hr intervals. Serum progesterone concentrations in steroid-treated gilts reached 70 ± 6.8 ng/ml (mean ± SE) by Day 8 and remained elevated thereafter, whereas serum progesterone concentrations in oil-treated controls were less than 1 ng/ml for the entire study. Daily serum LH concentrations were not different between gilts treated with progesterone or oil. The 1-μg pulses of gonadotropin-releasing hormone reliably evoked pulses of LH in both treatment groups. The LH pulse frequency and amplitude, assessed from samples collected every 15 min for 6 hr on Day 12, were similar for progesterone- and oil-treated gilts. These results provide evidence that progesterone does not act at the pituitary gland to alter LH secretion in pigs.

Role of Adipose Secreted Factors and Kisspeptin in the Metabolic Control of Gonadotropin Secretion and Puberty

© 2013 Lents et al., licensee InTech. This is an open access chapter distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Role of Adipose Secreted Factors and Kisspeptin in the Metabolic Control of Gonadotropin Secretion and Puberty