Role of frequency and amplitude of repetitive HCG stimulations for sustained progesterone secretion from the bovine corpus luteum in vitro.

This investigation aimed at evaluating a role for frequencies and amplitudes of repeated HCG stimulations for the optimal maintenance of progesterone (P4) secretion from the bovine corpus luteum in vitro. Slices (100-120 mg) of midluteal bovine corpora lutea were perifused with medium M199 (0.05% BSA, pH 7.2, 38.5 degrees C) and the perifusion effluent collected at 15 minute intervals for 20-29 hours. Unstimulated P4 release (n = 5) was distinctly pulsatile (by Pulsar pulse algorithm), with pulses occurring every 90 +/- 6 minutes (mean +/- SEM) and pulse amplitudes of 14.4 +/- 1.1 ng. Conversely, no pulses were detected in two control perifusions. Unstimulated P4 release increased during the first 5 perifusion hours (from 39.3 +/- 4.6 to 50.3 +/- 5.6 ng/15 min, p less than 0.01), but then appeared to decline (to 29.3 +/- 1.3 ng/15 min, p less than 0.05) towards the end of the perifusion periods. Hourly pulses of HCG (6.7 mM) did not change the P4 pulse amplitudes (16.6 +/- 2.0 ng), the pulse periodicities (105 +/- 15 min) and overall release rates (34.7 +/- 5.7 ng/15 min), nor did they prevent the decline in P4 secretion towards the end of perifusions (n = 5). In contrast, 2-hourly HCG stimulations maintained stable P4 release rates throughout the perifusion periods (34.7 +/- 6.8 ng/15 min), with P4 pulses of similar amplitudes (14.7 +/- 1.7 ng), but of lower periodicities (135 +/- 2 min, p less than 0.05) than during unstimulated conditions (n = 5).(ABSTRACT TRUNCATED AT 250 WORDS)     

Radioimmunoassay of 5-methoxy tryptophol in sheep plasma and pineal glands

No 5-methoxytryptophol (ML) could be detected in sheep plasma using a specific, sensitive radioimmunoassay developed for the purpose. Blood samples were collected from sheep during darkness and daylight and during various stages of the estrous cycle, but in no sample was the ML content above the detection limit of the method. Addition of 1 mM pargyline and neostigmine to blood immediately after collection to block metabolism did not result in a detectable ML concentration. The failure to detect ML was not due to degradation since added ML was not degraded by blood enzymes even after 16 hours incubation at 37 degrees C. Injection of 100 microgram and 1 mg ML sc in sheep resulted in a rapid rise of ML to 95-130pg/ml and 560-1000pg/ml respectively and disappearing with a half life of approximately 15-20 minutes. Sheep pineal glands collected during the light phase contained ML (51 +/- 5pg/mg tissue. X +/- SE, n = 7) which represents less than 6% of the melatonin content. It is concluded that if ML is present in sheep blood it is present at very low levels. It is thus unlikely to be a major circulating pineal hormone in this species, however, its role within the CNS as a local hormone cannot be excluded.     

Effects of ovarian input on GnRH and LH secretion immediately postovulation in pony mares

The potential involvement of ovarian factors in regulating GnRH and LH postovulation was studied in ovarian intact (Group 1; n=3) and ovariectomized (OVX; Group 2; n=3) mares (OVX within 12 hr of ovulation). Blood samples were collected every 10 min for 6 hr from jugular vein (JV) and intercavernous sinus (ICS) during estrus and on Day 8 postovulation for LH and GnRH analysis. Additionally, JV samples were collected twice daily (12-hr intervals) for 30 days for LH and progesterone (P4) analysis. A significant treatment x day effect (P<0.0001) describes declining plasma LH concentrations in intact mares, and regression analysis indicated that response curves were not parallel (P<0.001). Plasma LH concentrations remained elevated in OVX mares. LH increased further in OVX mares by Day 8 post-OVX (P<0.06), reflecting the increased (P<0.07) LH episode amplitude. GnRH decreased from estrus to Day 8 in both groups reflecting an effect of sampling period (P<0.03). GnRH episode amplitude declined (P<0.08) from estrus (62.8+/-3.1 pg/mL) to Day 8 (46.3+/-3.1 pg/mL) in OVX mares, but not in control mares (intact estrus, 36.5+/-6.4; intact Day 8, 37.5+/-7.3; OVX estrus, 62.8+/-3.1; OVX Day 8, 46.3+/-3.1 pg/mL). In conclusion, we propose that postovulatory LH decline requires ovarian feedback in mares, and that OVX alters GnRH secretory dynamics such that LH concentrations does not decline postovulation and, in fact, is further elevated with time after OVX.     

Corpus luteum function and embryonic mortality in buffaloes treated with a GnRH agonist, hCG and progesterone

The effect of treatment with a GnRH agonist, hCG or progesterone (P(4)) on corpus luteum function and embryonic mortality was investigated in buffaloes inseminated during mid-winter. Italian Mediterranean buffaloes (n=309) were synchronized using the Ovsynch with timed-AI program and mated by AI at 16 h (Day 0) and 40 h after the second injection of GnRH. On Day 5, buffaloes were randomly assigned to four groups: Control (no treatment, n=69), GnRH agonist (buserelin acetate, 12.6 microg, n=73), hCG (1500 IU, n=75) and P(4) (PRID without E(2) for 10 days, n=77). Progesterone (pg/ml) was determined in milk whey on Days 5, 10, 15 and 20 and pregnancy diagnosis was undertaken on Day 26 by ultrasound and Day 40 by rectal palpation. Treatment with buserelin and hCG increased (p<0.05) P(4) on Day 15 compared with controls (456+/-27, 451+/-24 and 346+/-28 pg/ml, respectively). Buffaloes treated with a PRID had intermediate P(4) concentrations (380+/-23 pg/ml). Embryonic mortality between Days 26 and 40 (22.9%) and pregnancies at Day 40 (48.9%) did not differ between treatments. A higher (p<0.01) P(4) concentration was found on Day 20 in pregnant animals compared with non-pregnant and embryonic mortality buffaloes, which did not differ. In summary, buserelin and hCG increased P(4) concentrations on Day 15 but this was not associated with a reduced incidence of embryonic mortality in buffaloes during mid-winter.     

Oxytocin plasma level in the lactating sows--effect of suckling

Oxytocin concentration in the peripheral blood was measured by RIA during suckling period in lactating sows (n = 8). Blood samples were taken from the jugular vein around the clock for every 2 h on day 5, 10, 15, 20, 25, 30 and on day 35 of lactation. Besides that blood samples were collected more frequently during suckling periods. Oxytocin plasma concentration was very low and in most cases it was on a border of sensitivity of our method (3 pg/ml). Marked but short-lasting rise of oxytocin was observed only during a period of initial massage of the udders by the piglets. This rise observed in all studied pigs was higher (p less than 0.01) compared to the values before the massage on the onset of lactation only, and was 14.6 +/- 4.2 pg/ml and 6.4 +/- 1.2 pg/ml on day 5 and day 10 of lactation, respectively. In all other studied days in a few cases only suckling stimulated the release of oxytocin over its basic concentration. Mean values (+/- SEM) of oxytocin in blood samples collected during massage of udder on day 15, 20, 25, 30 and day 35 were 3.7 +/- 0.5, 4.2 +/- 0.8, 4.9 +/- 1.1, 3.2 +/- 0.4 and 3.0 +/- 0.6 pg/ml plasma, respectively. There was no relationship between the size of the litters and neither basic level of oxytocin nor its blood concentration during suckling (r = 0.13).     

Rate of phosphate release after photoliberation of adenosine 5'-triphosphate in slow and fast skeletal muscle fibers.

Inorganic phosphate (Pi) release was determined by means of a fluorescent Pi-probe in single permeabilized rabbit soleus and psoas muscle fibers. Measurements of Pi release followed photoliberation of approximately 1.5 mM ATP by flash photolysis of NPE-caged ATP in the absence and presence of Ca2+ at 15 degrees C. In the absence of Ca2+, Pi release occurred with a slow rate of 11 +/- 3 microM . s-1 (n = 3) in soleus fibers and 23 +/- 1 microM . s-1 (n = 10) in psoas fibers. At saturating Ca2+ concentrations (pCa 4.5), photoliberation of ATP was followed by rapid force development. The initial rate of Pi release was 0.57 +/- 0.05 mM . s-1 in soleus (n = 13) and 4.7 +/- 0.2 mM . s-1 in psoas (n = 23), corresponding to a rate of Pi release per myosin head of 3.8 s-1 in soleus and 31.5 s-1 in psoas. Pi release declined at a rate of 0.48 s-1 in soleus and of 5.2 s-1 in psoas. Pi release in soleus was slightly faster in the presence of an ATP regenerating system but slower when 0.5 mM ADP was added. The reduction in the rate of Pi release results from an initial redistribution of cross-bridges over different states and a subsequent ADP-sensitive slowing of cross-bridge detachment.     

Influence of gastric inhibitory polypeptide (GIP) and glucose on the regulation of glucagon secretion by pancreatic alpha cells

The effects of glucose and GIP on glucagon secretion were studied in perifused microdissected murine pancreatic islets. Glucagon levels were determined in effluent samples collected at 1-min intervals by radioimmunoassay using the glucagon-specific antibody, 30 K. There was no significant difference in the total amount (7740 +/- 212 pg vs 8630 +/- 36 pg, n = 10) of glucagon secreted over a 20 min period when the glucose concentration was alternately shifted between 5.5 mM and 11.1 mM, respectively. However, 22.2 mM glucose profoundly suppressed glucagon secretion. The suppressive effect of high glucose on glucagon release was partially, yet significantly, reversed by the presence of GIP, as glucagon secretion increased from a non-detectable level at 22.2 mM glucose alone to 10,175 +/- 145 pg, n = 10 (P less than 0.01). The glucagonotropic effect of GIP was dose-dependent in the range of 2 x 10(-9) - 2 x 10(-7) M, at 11.1 mM glucose. Our data show that GIP is able to substantially reverse the suppressive effect of a high glucose load on glucagon secretion.     

Hypothalamic proopiomelanocortin mRNA levels in suckled or nonsuckled beef cows: A preliminary study

This study was conducted to determine proopiomelanocortin (POMC) mRNA levels in the preoptic and hypothalamic brain regions of postpartum anestrous cows. An additional objective was to determine if calf suckling influences POMC mRNA concentration in these regions. Twenty cows were randomly assigned to suckled and nonsuckled treatment groups and slaughtered between 30 and 36 days postpartum. Serum luteinizing hormone (LH) concentrations were determined from blood collected every 15 minutes for 8 hours, starting 20 hours prior to slaughter. POMC mRNA levels in brain tissues were determined by dot blots. Serum LH concentrations between nonsuckled and suckled cows were 1.3 +/- 0.2 and 0.9 +/- 0.1 ng.ml(-1) (mean +/- SEM; P = 0.19), respectively. The POMC gene is expressed in the hypothalamus of postpartum anestrus cows with POMC mRNA levels higher (P<0.05) in the hypothalamus than in the preoptic region. Hypothalamic POMC mRNA levels tended (P = 0.12) to be lower in nonsuckled (14.9 +/- 3.8 ADU) than in suckled cows (23.5 +/- 3.6 ADU). Covariate analysis indicated (P = 0.10) that as mean serum LH concentrations increased, hypothalamic POMC mRNA levels decreased.     

Changes in somatostatin-like immunoreactivity in lungs from perinatal guinea pigs and the effects of somatostatin-14 on lung liquid production.

Somatostatin-like immunoreactivity was measured by radioimmunoassay with a monoclonal antibody in lungs from perinatal guinea pigs (62 +/- 2 days of gestation). Fetuses delivered by Caesarean section and dissected before breathing showed 4748 +/- 758 pg/lung (n = 25). Fetuses allowed to breathe (neonates) showed marked increases in activity: 7629 +/- 1355 pg/lung (n = 12) after breathing 30 seconds, and 10729 +/- 1064 pg/lung (n = 6) after breathing 3 minutes (2.3-fold increase, P < 0.005). Values then declined (5203 +/- 1050 pg/lung (n = 9) at 30 minutes; 1458 +/- 105 pg/lung (n = 4) at 60 minutes). Changes were similar in pg/g wet tissue. HPLC characterized the immunoreactive peptides as somatostatin-14 (SS-14) and somatostatin-28 (SS-28) in both fetuses and neonates (n = 11). SS-28 made up only 13.7 +/- 1.7% of the activity; this percentage did not change with breathing. The effects of synthetic SS-14 on lung liquid production were investigated in in vitro lungs from 42 fetal guinea pigs. All 21 preparations immersed in 10(-5)-10(-7) M SS-14 during the middle hour of 3 h incubations reduced production, often approaching zero after treatment (rates, ml/kg body weight per h, succeeding hours: 10(-5) M (n = 9), 3.09 +/- 0.68, 0.93 +/- 0.39, -0.05 +/- 0.60 (fall significant during and after treatment, P < 0.025-0.005); 10(-6) M (n = 6), 3.06 +/- 0.68, 1.29 +/- 0.58, 0.36 +/- 0.38 (P < 0.05-0.005); 10(-7) M (n = 6), 1.96 +/- 0.66, 1.11 +/- 0.34, 0.64 +/- 0.28 (P < 0.05-0.025).(ABSTRACT TRUNCATED AT 250 WORDS)     

Thrombin-stimulated increases in cytosolic Ca2+ level and gonadotropin-releasing hormone release in GT1-7 neurons.

The effects of thrombin on cytosolic calcium levels ([Ca2+]cyt), and on gonadotropin-releasing hormone (GnRH) release, were characterized in cultured GT1-7 neurons. GnRH release from GT1-7 neurons was pulsatile with an average pulse amplitude of 14.3+/-5.8 pg x min x ml(-1) and an average pulse duration of 21.3+/-4.2 min. The [Ca2+]cyt response to 0.005 to 0.2 U/ml thrombin was saturable and concentration dependent (EC50 = 0.0268 U/ml). Ethyleneglycotetraacetic acid (EGTA) chelation of extracellular Ca2+ resulted in an approximately 70% attenuation of thrombin-stimulated increase in [Ca2+]cyt. By use of a special superfusion system, a 5-min exposure to 0.1 U/ml thrombin significantly increased the amplitude (193.2+/-67.8 pg x min x ml(-1); P = 0.001) but not the duration (22.5+/-2.4 min; P = 0.8) of GnRH release. These results suggest that thrombin increases [Ca2+]cyt and GnRH release from GT1-7 neurons via specific membrane-bound receptors.     

Prolonged pulsatile release of gonadotropin-releasing hormone from the guinea pig hypothalamus in vitro

The sexually mature mammal secretes luteinizing hormone in a pulsatile fashion. This is presumed to depend on the intermittent release of hypothalamic gonadotropin- releasing hormone (GnRH). The isolated guinea pig hypothalamus has been studied because, in this species, as in primates, the pulse generator appears to reside within the medial basal hypothalamus. The basal 2 mm of guinea pig hypothalami were rapidly removed and perifused at 37 degrees C with Krebs-Ringer solution containing 20 mM bacitracin gassed with 95% O2, 5% CO2. The eluates were sampled at 15 and 5 min intervals and pulsatile patterns of GnRH were consistently observed for periods up to 72 h. There was no difference in GnRH levels from hypothalami of intact and ovariectomized animals. Simultaneous measurement of TRH and somatostatin disclosed independent pulses of both neurohormones which did not coincide with GnRH, indicating that the peaks were secretory episodes not artefacts generated by varying perifusion rates. The hypothalami disclosed no histologic evidence of necrosis when examined after 20 h perifusion.     

Gonadotropin-releasing hormone secretion during the postpartum anestrous period of the ewe

An increase in episodic release of LH is putatively the initial event leading to the onset of postpartum ovarian cyclicity in ewes. This experiment was conducted to determine the relationship between hypothalamic release of GnRH and onset of pulsatile secretion of LH during postpartum anestrus. Control ewes (n = 7) were monitored during the postpartum period to determine when normal estrous cycles resumed. In controls, the mean interval from parturition to the first postpartum estrus as indicated by a rise in serum progesterone greater than 1 ng/mg was 25.8 +/- 0.6 days. Additional ewes (n = 4-5) at 3, 7, 14, and 21 days postpartum (+/- 1 day) were surgically fitted with cannula for collection of hypophyseal-portal blood. Hypophyseal-portal and jugular blood samples were collected over a 6- to 7-h period at 10-min intervals. The number of GnRH pulses/6 h increased (p less than 0.05) from Day 3 postpartum (2.2 +/- 0.5) to Days 7 and 14 (3.6 +/- 0.2 and 3.9 +/- 0.4, respectively). A further increase (p less than 0.05) in GnRH pulse frequency was observed at Day 21 postpartum (6.4 +/- 0.4 pulses/6 h). Changes in pulsatile LH release paralleled changes observed in pulsatile GnRH release over Days 3, 7, 14, and 21 postpartum (0.83 +/- 0.3, 2.8 +/- 0.4, 2.9 +/- 0.6, and 4.0 +/- 1.1 pulses/6 h, respectively). GnRH pulse amplitude was higher at Day 21 than at Days 3, 7, or 14 postpartum. These findings suggest that an increase in the frequency of GnRH release promotes the onset of pulsatile LH release during postpartum anestrus in ewes.     

Effect of perfusate potassium concentration on the contraction rate of the isolated rat heart during profound hypothermia

Effect of different concentration of K+ in perfusion fluid ([K+]) (5.9 mM, 3.6 mM, 2.38 mM) and the heart temperatures of 20 degrees C and below on the rat heart rate in the Langendorf preparations, were examined in conditions of retrograde perfusion with a modified Krebs-Henseleit buffer at constant perfusion volume. The lowering of [K+] diminished the temperature/heart rate ratio and depressed the heart standstill temperature from 12.3 +/- 0.6 degrees C at [K+] 5.9 mM (n = 12) to 6.7 +/- 0.6 degrees C at [K+] 3.6 mM (n = 5) and to 2.24 +/- 0.40 degrees C at [K+] 2.38 mM (n = 5). Temperature of the cold heart standstill had the liner relationship to Ig[K+]. Change the perfusion fluid with 5.9 mM K+ after heart cold standstill by the perfusion fluid with 3.6 mM K+ restored the heart beats to the rate of 40-50 min-1 in some experiments. The second heart standstill was at the mean temperature 3.6 degrees C lower than the first one.     

Size distribution and hormonal responsiveness of dispersed rabbit luteal cells during pseudopregnancy

Due to the evidence for two distinct steroidogenic cell types in corpora lutea of large domestic animals, cells of the rabbit corpus luteum were characterized with respect to cell diameters, relative abundance, steroidogenic capacity and responsiveness to hormones. Pseudopregnancy was induced in New Zealand rabbits by injection of 30-160 IU pregnant mare's serum gonadotropin (PMSG) followed in 2-4 days by an i.m. injection of 20-35 micrograms gonadotropin-releasing hormone (GnRH). Corpora lutea were obtained 2, 5 and 9 days after injection of GnRH and dissociated into single cell suspensions. Suspended steroidogenic cells were incubated (2 h, 37 degrees C) in medium 199 alone or in medium containing ovine luteinizing hormone (oLH) (100 ng/ml), or isoproterenol (100 microM). Media were collected and assayed for progesterone content. Secretion of progesterone (means +/- SE, n = 4) was stimulated (p less than 0.05) by oLH on each day: Day 2 = 1.7 +/- 0.2-fold; Day 5 = 3.5 +/- 0.4-fold; and Day 9 = 3.1 +/- 0.6-fold stimulation above controls. Isoproterenol also stimulated (p less than 0.05) secretion of progesterone by suspended luteal cells on Days 2 and 9. Microscopic examination of cell suspensions stained for 3 beta-hydroxysteroid dehydrogenase (3 beta HSD) activity provided identification of cells with steroidogenic capacity. The diameters (means +/- SE) for steroidogenic cells increased (p less than 0.05) from Days 2 to 9 (Day 2 = 15.2 +/- 0.2 micron; Day 5 = 22.4 +/- 0.4 micron; Day 9 = 28.3 +/- 1.6 micron). The large cell to small cell ratio increased from 0.01 on Day 2 to 2.03 on Day 9.(ABSTRACT TRUNCATED AT 250 WORDS)     

Seasonal changes in the in-vivo activity of the luteinizing hormone-releasing hormone (LHRH) neural apparatus of male rabbits monitored with push-pull cannulae

Intact sexually mature New Zealand White male rabbits, raised under natural lighting and temperature conditions, were isolated and housed in air-conditioned quarters, in a 12-h light:12-h dark cycle. Push-pull cannulae were implanted towards the tuberal region of the hypothalamus, and animals were perfused with modified Krebs'-Ringer phosphate medium for an average period of 4 h. Most rabbits were repetitively perfused over an average period of 3.7 months. Perfusions were grouped into seasonal periods of about 40 or 80 days through the entire 1-year cycle: (A) 23 November to 31 December (winter solstice period: N = 6), (B) 1 January to 23 March (winter: N = 10), (C) 24 March to 13 June (spring: N = 9), (D) 14 June to 23 July (summer solstice period: N = 23), (E) 24 July to 13 October (late summer-early fall: N = 7) and (F) 14 October to 22 November (fall: N = 4). Maximal and minimal values of mean release, mean amplitude and mean frequency from every animal in each block were obtained. In the summer solstice group (D), maximal mean (+/- s.e.) LHRH release levels were significantly greater (14.42 +/- 6.62 pg/10 min) than for all groups (A, 0.76 +/- 0.27; B, 1.59 +/- 0.39; C, 1.34 +/- 0.22; E, 1.33 +/- 0.33; and F, 1.18 +/- 0.11) while during the winter solstice period (A), minimal mean LHRH release levels (0.48 +/- 0.04 pg/10 min) were significantly lower than in all other groups (B, 1.43 +/- 0.41; C, 0.96 +/- 0.11; D, 7.25 +/- 4.12; E, 1.18 +/- 0.37; and F, 1.18 +/- 0.11). Maximal values were highest during the summer solstice period while the minimal values were lowest during the winter solstice period. The amplitude and frequency of the LHRH pulses showed changes similar to those observed with the mean LHRH release. For 1 year, an estimated left testis weight was measured in 4-7 rabbits every 2-5 weeks. The percentage estimated testis weight peaked in early August and reached minimal levels during the winter months. These data demonstrate that the rabbit LHRH neural apparatus is very sensitive to seasonal influences although animals remained in a fixed photoperiod during the entire duration of the experiment.     

Influence of GnRH infusion on endocrine parameters and duration of postpartum anestrus in beef cows

The effect of an intravenous infusion of gonadotrophin releasing hormone (GnRH) on the duration of postpartum anestrus in suckled beef cows was studied. Twenty-eight, mature, suckled beef cows were assigned in equal numbers to one of four treatment groups which were based on infusion with saline or GnRH (15ug/hour for 12 hours) and stage postpartum (pp) (20 or 35 days). Serum LH and progesterone were determined by radioimmunoassay for the period which began 5 days pre-infusion and ended at 55 days postpartum (ie: 35 or 20 days post-infusion). Serum LH remained below 5ng/ml during infusion in all control cows. Peak serum LH values, times of LH peaks, and duration of LH responses (means +/- SE) during infusion were 49 +/- 12 ng/ml, 162 +/- 42 minutes and 7.8 +/- 1.3 hours for the 20 day group and 44 +/- ng/ml, 144 +/- 6 minutes, and 8.2 +/- 1.1 hours for the 35 day group respectively. Serum progesterone levels indicated that the proportion of cows showing the onset of estrous cycles within 10 days of infusion was greater in the 20 day pp GnRH group (4/7) than the 20 day pp saline group (0/7) (p < .05) but was not significantly different between the 35 day pp GnRH (4/7) and 35 day pp saline (2/6) groups. The incidence of estrus was not affected by GnRH treatment and was 37% in all cows prior to 55 days pp. It was concluded that infusions of GnRH for 12 hours at a rate of 15 ug/hour could induce estrous cycles in suckled beef cows treated at 20 days postpartum.     

In vitro release of interleukin-15 by broncho-alveolar lavage cells and peripheral blood mononuclear cells from patients with different lung diseases

IL-15 shares several biological activities with IL-2 and uses the b and g chain of the IL-2 receptor. In addition to its T-cell stimulating capacity, IL-15 exhibits regulatory properties on macrophage proinflammatory cytokine release. IL-15 is released by non-lymphoid cells, e.g. muscle cells, fibroblasts and monocytes/macrophages. In many lung diseases alveolar macrophages (AM) are activated and release pro- inflammatory cytokines. We asked whether IL-15 is released ex vivo by AM and peripheral blood mononuclear cells (PBMC) from patients with inactive sarcoidosis (PSi), active sarcoidosis (PSa), tuberculosis (TB), hypersensitivity pneumonitis (HSP), cryptogenic fibrosing alveolitis (CFA) and pneumonia (PN). Additionally, we examined the kinetics of the IL-15 release of these cells. During 24 hours of culture, AM from controls (CO) released 3.8 +/- 1.9 pg/ml (mean +/- SD) of IL-15, which was significantly lower than in most of the patient groups (PSa: 8.7 +/- 3.9 pg/ml, TB: 8.4 +/- 1.9 pg/ml, CFA: 5.7 +/- 1.5 pg/ml, and PN: 7. 8 +/- 2.6 pg/ml) except PSi (4.0 +/- 2.6 pg/ml) and HSP (9.3 +/- 9.5 pg/ml). PBMC from patients with PSa released significantly more IL-15 than PBMC from CO (10.8 +/- 8.9 pg/ml versus 6.9 +/- 2.2 pg/ml) whereas PBMC IL-15 release of the other groups did not differ from CO (TB: 5.7 +/- 1.4 pg/ml; CFA: 4.6 +/- 1.6 pg/ml; HSP: 4.9 +/- 3.8 pg/ml). Kinetic studies revealed a minor peak after 5 hours and a major peak from 12 hours to 35 hours for AM and PBMC. In summary, AM from all patient groups but the PSi and the HSP group released increased levels of IL-15, although the total amount of this cytokine is very low.     

Comparison of pregnancy rates of repeat-breeder dairy cows given gonadotropin releasing hormone at or prior to the time of insemination

A total of 585 repeat-breeder dairy cows was used to study the effect of GnRH treatment, either at or prior to insemination, on the pregnancy rate. The cows were divided into 6 treatment groups. Cows in Group 1 (n = 142) were observed in estrus, and 11 +/- 0.42 hours (mean +/- SEM) later they were given 100 ug, i.m. gonadotropin releasing hormone (GnRH) and were inseminated. Cows in Group 2 (n = 139) were observed in estrus and were inseminated 11.4 +/- 0.43 hours later. Cows in Group 3 (n = 33) were monitored for estrus with an activated heatmount detector but were not observed in estrus; they were inseminated 1.5 +/- 0.87 hours later and were given 100 ug, i.m. GnRH. Cows in Group 4 (n = 35) were not observed in estrus, but they did activate the heatmount detector and were inseminated 2.2 +/- 0.87 hours later. Cows in Group 5 (n = 107) were observed in estrus, given 100 ug, i.m. GnRH 2.0 +/- 0.40 hours later, and were inseminated 9 +/- 0.60 hours after GnRH treatment. Cows in Group 6 (n = 129) were observed in estrus and were inseminated 10 +/- 0.50 hours later. Pregnancy rates were analyzed by Chi-square. Interactions between pregnancy rate, treatment and time of insemination were evaluated using ANOVA and LSM (P < 0.05). There was no effect on pregnancy rate when GnRH was given at or prior to insemination. Cows inseminated on the basis of observed estrus had a higher pregnancy rate (P < 0.05) than cows inseminated on the observation of an activated heatmount detector. From the results of this study, it is concluded that treatment with GnRH at or prior to insemination did not improve the pregnancy rate of repeat-breeder dairy cows.     

Rapid release of substance P and LH-RH from synaptosomes prepared from the medial basal hypothalamus and substantia nigra

We investigated Ca2+-dependent, depolarization-induced release of substance P (SP) and LH-RH from medial basal hypothalamic (MBH) and substantia nigra (SN) synaptosomes prepared from male rat brain. Depolarization of MBH synaptosomes evoked significant release of SP from 10.0 +/- 0.1 (5 mM K+) to 28.0 +/- 2.4 (75 mM K+) pg released/10 seconds. Fractional release was 1.0% and 2.7% respectively. In contrast, LH-RH was not released by depolarization of MBH synaptosomes: 11.6 +/- 0.9 (5 mM K+) to 11.0 +/- 0.7 (75 mM K+) pg released/10 seconds. Fractional release was 1.1 and 1.0% respectively. Depolarization-induced LH-RH release also did not occur in the presence of 10(-4) or 10(-6) M norepinephrine, 10(-7) M 12-O-tetradecanoylphorbol-13-acetate (TPA, PMA), 10(-5) M forskolin or in female rats. The inability of depolarizing concentrations of K+ to stimulate LH-RH release in physiological buffers remains an enigma. Significant depolarization-induced SP release was seen from MBH and SN synaptosomes at 20, 15, 10, 5 and only 1 second of release. Despite comparable basal release of SP from MBH and SN synaptosomes, the rate and magnitude of evoked release were much more pronounced in SN synaptosomes. The initial rate (0-1 second) of SP release was 4.5-fold greater from SN than from MBH synaptosomes [krel = 0.027(-1) (SN), krel = 0.006(-1) (MBH)]. The magnitude of SP release from SN synaptosomes was 2- to 3-fold greater at any given time interval compared with release from MBH synaptosomes.(ABSTRACT TRUNCATED AT 250 WORDS)     

The effect of zwitterion buffers on the freezability of Boer goat semen

Twenty semen samples with mass activity greater than +3 were collected from six healthy, mature Boer goat bucks. Each ejaculate was divided into four equal parts and extended at 37 degrees C in Tris, Test, Tes and Bes buffers containing egg yolk and glycerol. Semen was placed into medium size French strawsand after 2 hours of equilibration at 5 degrees C, frozen in the vapour phase and stored in liquid nitrogen for 7 days at -196 degrees C. Progressive motility, the number of live spermatozoa and glutamic oxaloacetic transaminase (GOT) release were studied after the initial extension, after equilibration and after 15 minutes and 7 days of freezing of semen. Semen samples when extended with Tris yolk glycerol showed significantly (P<0.01) higher progressive motility and live spermatozoa than when extended with the other zwitterion buffer-based extenders. The change of extenders did not influence the release of GOT at various stages of freezing of semen (P>0.05).