Kisspeptin Stimulates the Pulsatile Secretion of Luteinizing Hormone (LH) during Postpartum Anestrus in Ewes Undergoing Continuous and Restricted Suckling

This study tested whether the intravenous application of kisspeptin can stimulate the pulsatile secretion of LH in suckling ewes during postpartum anestrus. Ten days after lambing, Pelibuey ewes were allocated among two groups: (1) continuous suckling (n = 8), where the lambs remained with their mothers; and (2) restricted suckling (n = 8), where the mothers suckled their lambs twice daily for 30 min. On Day 19 postpartum, the ewes were individually penned with ad libitum access to water and feed and given an indwelling catheter in each jugular vein. On Day 20, 4 mL of blood was sampled every 15 min from 08:00 to 20:00 h to determine LH pulse frequency. At 14:00 h, four ewes in each group received 120 μg of kisspeptin diluted in 3 mL of saline as a continuous infusion for 6 h; the remaining four ewes in each group received only saline. The interaction between kisspeptin and suckling type did not affect LH pulse frequency (p > 0.05). Before kisspeptin administration, pulse frequency was similar in all groups (1.50 ± 0.40 pulses per 6 h; p > 0.05). With the application of kisspeptin, pulse frequency increased to 3.50 ± 0.43 pulses per 6 h (p ≤ 0.014), so the concentration of LH (1.11 ± 0.14 ng mL−1) was greater in kisspeptin-treated ewes than in saline-treated ewes (0.724 ± 0.07 ng mL−1; p ≤ 0.040). The frequency of LH pulses was greater with restricted suckling than with continuous suckling (2.44 ± 0.29 versus 1.69 ± 0.29 pulses per 6 h; p ≤ 0.040). We conclude that intravenous application of kisspeptin increases the pulsatile secretion of LH in suckling ewes and that suckling might reduce kisspeptin neuronal activity, perhaps explaining the suppression of ovulation. Moreover, the effects of kisspeptin and suckling on pulsatile LH secretion appear to be independent, perhaps operating through different neural pathways.

SUN-LB57 Assessing the Requirement of Arcuate Nucleus Kisspeptin Neurons in Puberty Onset and Adult LH Secretion

Puberty is a critical developmental period marking the transition to adulthood and attainment of reproductive capability. A hallmark of puberty is increased pulsatile secretion of pituitary luteinizing hormone (LH) which is itself driven by increased gonadotropin-releasing hormone (GnRH) from the forebrain. The mechanisms governing GnRH neuron activation at puberty still remain unclear, but likely include enhanced stimulation from upstream reproductive neural circuits, including kisspeptin neurons. Kisspeptin is a potent stimulator of GnRH and is required for proper puberty onset. However, the specific brain site(s) from where kisspeptin signaling arises to trigger puberty remain unclarified. Kisspeptin is expressed in two primary nuclei in the hypothalamus, the arcuate nucleus (ARC) and anteroventral periventricular (AVPV) region. Studies suggest that, in adulthood, ARC Kiss1 neurons are involved in driving pulsatile secretion of GnRH (and hence, LH) in both sexes whereas AVPV Kiss1 neurons participate in the preovulatory GnRH/LH surge in females. However, the specific role of either kisspeptin neuron population in puberty onset still remains unknown. We previously showed that both kisspeptin populations show increased Kiss1 gene expression across the pubertal period, yet whether just one or both (or neither) population is needed for puberty to occur has not been determined. Here, we sought to tease out the role—if any—of ARC and AVPV Kiss1 neurons in the pubertal onset process. Since ARC Kiss1 neurons are abundant in both sexes and drive pulsatile GnRH secretion in adulthood, we hypothesized that ARC Kiss1 neurons are necessary for normal puberty onset and, conversely, that AVPV Kiss1 neurons are not sufficient on their own to induce normal puberty. To test this hypothesis, we used a Cre-specific diphtheria toxin approach to ablate just ARC Kiss1 neurons in juvenile mice (~ 2 weeks old) while leaving AVPV Kiss1 neurons intact. Preliminary data thus far indicates that site specific ablation of just ARC Kiss1 neurons during the juvenile period significantly delays puberty onset in both sexes, as measured by vaginal opening, first estrous, and preputial separation. In addition, selective ARC Kiss1 neuron ablation in juvenile life diminishes pulsatile LH secretion levels measured in adulthood, but does not alter LH surge generation in adult females. These preliminary findings empirically demonstrate that, in mice, ARC Kiss1 neurons are required for proper activation of the reproductive axis during puberty but not the LH surge in adulthood, and AVPV Kiss1 neurons are not sufficient to trigger normal pubertal onset.

374 Effects of maternal nutrition on secretion of leptin in the neonatal heifer and interaction of maternal and postnatal nutrition on age at puberty and postpubertal secretion of luteinizing hormone

Nutrition during gestation and early postnatally can program developmental changes in the offspring that persist until adult life. Here we tested the hypotheses that maternal nutritional during the second and third trimesters of gestation 1) affects neonatal secretion of leptin in heifer offspring, and 2) interacts with dietary energy intake during the juvenile period to affect age at puberty and postpubertal pulsatile secretion of luteinizing hormone (LH) in heifers. Bos indicus-influenced beef cows (n = 108) in 3 replicates bearing heifer fetuses were assigned randomly to receive low (L), moderate (M), or high energy (H) diets to achieve body condition scores (BCS) of 3–3.5 (thin; n = 36), 5.5–6 (moderate; n = 36), or 7.5–8 (obese; n = 36) by the second trimester of gestation until calving. Heifer offspring were weaned at 3–3.5 months of age and assigned randomly to either a high- (H; 1 kg/day) or low-gain (L; 0.5 kg/day) diet for 5 months. Blood samples from a subgroup of 30 heifers (Exp. 1) from cows in each maternal treatment (n = 10/treatment) were collected once every 2–3 days for 2 weeks after birth. In Exp. 2, 18 heifers representing 3 of the maternal × postnatal groups (HH, MH and LL) were ovariectomized and received estradiol replacement (OVX+E) after puberty. Pulsatile secretion of LH was evaluated for 5.5 hours. Maternal nutrition did not affect postnatal circulating leptin. Based on two replicates (n = 61), postnatal diets had the greatest effect on age at puberty (L > H; P < 0.001), with a lesser maternal diet effect (P < 0.10), although LL > HH by 99 days. None of the characteristics of pulsatile LH secretion differed among treatments in OVX + E heifers. Dietary effects on age at puberty were not associated with hormonal characteristics evaluated in these initial studies.

Electrical Excitability of the Endoplasmic Reticulum Membrane Drives Electrical Bursting and the Pulsatile Secretion of Insulin in a Pancreatic Beta Cell Model

ABSTRACTPancreatic β-cells secrete insulin, the hormone that controls glucose homeostasis in vertebrates. When activated by glucose, β-cells display a biphasic electrical response. An initial phase, in which the cell fires action potentials continuously, is followed by a phase with a characteristic firing pattern, known as electrical bursting, that consists on brief pulses of action potentials separated by intervals of rest. Electrical bursting is believed to mediate the pulsatile secretion of insulin. The electrical response of β-cells has been extensively studied at experimental and theoretical level. However, there is still no consensus on the cellular mechanisms that underlie each of the phases of the response. In this paper, I propose the hypothesis that the pattern of the plasma membrane (PM) response of stimulated β-cells is generated by the electrical activity of the endoplasmic reticulum (ER) membrane. In this hypothesis, the interaction of the two excitable membranes, PM and ER membrane, each operating at a different time scale, generates both, the initial continuous phase and the periodic bursting phase. A mathematical model based on the hypothesis is presented. The behavior of the model β-cell replicates the main features of the physiological response of pancreatic β-cells to nutrients and to neuro-endocrine regulatory factors. The model cell displays a biphasic response to the simulated elevation of glucose. It generates electrical bursting with frequencies comparable to those observed in live cells. The simulation of the action of regulatory factors mimics the actual effect of the factors on the frequency of bursting. Finally, the model shows that a cell with a defective ER response behaves like a dysfunctional β-cell from individuals with type 2 diabetes mellitus, a result that suggests that the electrical malfunction of the ER membrane may represent one of the primary causes of type 2 diabetes. Dynamic analysis of the ER behavior has revealed that, depending on the transport rates of Ca2+ in and out of the ER, the system has three possible dynamic states. They consist on the hyperpolarization of the ER membrane, periodic oscillations of the electric potential across the membrane, and the depolarization of the membrane. Each of these states determines a different functional program in the cell. The hyperpolarized state maintains the cell at rest, in a non-secreting state. Periodic oscillations of the ER membrane cause electrical bursting in the PM and the consequent pulsatile secretion of insulin. Finally, the depolarized state causes continuous firing and an acute secretory activity, the hyperactive conditions of the initial phase of the β-cell response to glucose. The dynamic states of the ER are also associated with different long-term effects. So, conditions that induce the hyperactive depolarized state in β-cells also potentiate apoptosis. The induction of the oscillatory state by glucose and neuro-endocrine factors seems to activate also cell proliferation. In extreme conditions though, such as the chronic treatment of T2DM with incretin analogs, the activation of the oscillatory state may lead to the appearance of cancer. The mathematical model presented here is an illustration of how, even in a extremely simplified system, the nonlinearity or excitability of the ER membrane can produce a repertoire of dynamic states that are able to generate a complex response comparable to the response observed experimentally in pancreatic β-cells. In actual cells, with a much higher number of parameters susceptible to be modified by environmental and genetic factors, the ER membrane is likely to have a significantly bigger set of dynamic states each capable to direct the cell in a particular functional or developmental direction. The potential role of the electrical activity of the ER membrane in cellular processes such as fertilization, cell proliferation and differentiation, and cell death, as well as in the development of diverse pathological conditions is analyzed in the discussion.

Temporary undernutrition during early gestation, corpora lutea morphometrics, ovarian progesterone secretion and embryo survival in gilts

The present study reports effects of severe undernutrition on luteal function and pregnancy in pigs. Gilts were inseminated and either fasted on Day 10 and 11 after conception (n = 11) or fully fed throughout (n = 10). Fasting did not affect LH or progesterone pulsatile secretion pattern on Day 11 in samples taken from blood vessels draining an ovary. Ultrasonographic measurements of the size of the corpora lutea did not show any effect of fasting either. However, fasted gilts had 10 to 30 % lower systemic progesterone from Day 12 through Day 15 after conception (P < 0.05). All gilts farrowed, but fasted gilts had fewer born piglets than fully fed gilts (8.8 ± 0.8 vs 10.9 ± 0.5 respectively; P < 0.05). In conclusion, fasting during embryo elongation can compromise embryonic survival by affecting ovarian function in the days after fasting, without having an immediate effect on LH secretion and progesterone output by the ovaries.