Onset Of Surge Research Articles

Bombesin acts in the suprachiasmatic nucleus to affect circadian changes in tuberoinfundibular dopaminergic neuron activity and prolactin secretion

Central administration of bombesin has been shown to block the afternoon PRL surge. A circadian change in the activity of tuberoinfundibular dopaminergic (TIDA) neurons that coincides with the onset of the PRL surge has also been found. Whether bombesin acts on the TIDA neuron to affect the PRL surge and where bombesin acts were the focuses of this study. Bombesin (0.75 microgram/3 microliters.rat) was first injected into the lateral cerebroventricle of estrogen-primed ovariectomized rats at 1200 h through preimplanted cannulas. Both the PRL surge and the circadian changes in TIDA neuron activity (using both 3,4-dihydroxyphenylacetic acid concentration and 3,4-dihydroxyphenylalanine accumulation in the median eminence as indexes) were blocked by the treatment. In suprachiasmatic nucleus-lesioned rats, however, both rhythms were absent, and the injection of bombesin exhibited no significant effect. Using microinjection of bombesin (50 ng/0.2 microliter) bilaterally into either the suprachiasmatic or arcuate nuclei at 1200 h, only the former injection effectively blocked both TIDA neuron activity and the PRL surge. We conclude that bombesin may act on the rhythm generation center, the suprachiasmatic nucleus, to exert its effect to disrupt TIDA neuron activity and the PRL surge.

Bombesin acts in the suprachiasmatic nucleus to affect circadian changes in tuberoinfundibular dopaminergic neuron activity and prolactin secretion.

Central administration of bombesin has been shown to block the afternoon PRL surge. A circadian change in the activity of tuberoinfundibular dopaminergic (TIDA) neurons that coincides with the onset of the PRL surge has also been found. Whether bombesin acts on the TIDA neuron to affect the PRL surge and where bombesin acts were the focuses of this study. Bombesin (0.75 microgram/3 microliters.rat) was first injected into the lateral cerebroventricle of estrogen-primed ovariectomized rats at 1200 h through preimplanted cannulas. Both the PRL surge and the circadian changes in TIDA neuron activity (using both 3,4-dihydroxyphenylacetic acid concentration and 3,4-dihydroxyphenylalanine accumulation in the median eminence as indexes) were blocked by the treatment. In suprachiasmatic nucleus-lesioned rats, however, both rhythms were absent, and the injection of bombesin exhibited no significant effect. Using microinjection of bombesin (50 ng/0.2 microliter) bilaterally into either the suprachiasmatic or arcuate nuclei at 1200 h, only the former injection effectively blocked both TIDA neuron activity and the PRL surge. We conclude that bombesin may act on the rhythm generation center, the suprachiasmatic nucleus, to exert its effect to disrupt TIDA neuron activity and the PRL surge.

Effect of restraint stress on the preovulatory luteinizing hormone profile and ovulation in the rat.

Plasma profiles of luteinizing hormone (LH) and follicle-stimulating hormone (FSH) were measured during restraint stress on the day of pro-oestrus; these profiles were considered in relation to ovulation rate on the next day. Rats bearing a permanent jugular vein cannula were subjected to restraint, which was started 0, 1 or 2 h before the presumed onset of the LH surge and ended just before the beginning of the dark period. Exposure to restraint resulted in a suppression of the secretion of both gonadotrophins on the day of pro-oestrus. Suppression of the LH surge was virtually complete (plasma LH < or = 0.2 ng/ml) in 15 out of 32 stressed rats, and the ovaries of these rats contained graafian follicles with oocytes in germinal vesicle stage. In these rats, the LH surge did not occur 24 h later. In the remaining 17 rats, restraint resulted in a considerable suppression of the LH surge. Of these rats, five had an ovulation rate of 100% and four ovulated partially. In unruptured follicles of the latter, the oocyte had not resumed meiosis and the follicle wall was not luteinized. In the remaining eight rats with a reduced LH surge, ovulations had not occurred and graafian follicles were unaffected. The results of this study indicate that during pro-oestrus restraint stress suppresses and does not delay the release of preovulatory gonadotrophins. Partial suppression of LH by restraint does not result in induction of meiotic resumption without subsequent ovulation or in luteinized unruptured follicles.

Nature and bioactivity of gonadotropin-releasing hormone (GnRH) secreted during the GnRH surge

Previous studies have demonstrated a neural action of estradiol in inducing a surge of GnRH in the ewe. However, although the GnRH and LH surges began concurrently, the GnRH surge consistently continued well beyond the surge of LH. Three experiments were conducted to test the hypothesis that the termination of the LH surge results from the secretion of a relatively inactive variant of GnRH during the later phases of the GnRH surge. In the first experiment, hypophyseal portal blood collected during an estrogen-induced LH surge was analyzed for GnRH immunoreactivity using two antibodies having specificity for the N- or C-terminal portion of the GnRH molecule. The duration, amplitude, and time course of the GnRH surge were found to be similar irrespective of the antisera used. In a second experiment, a competitive GnRH antagonist was administered at the beginning of the estrogen-induced GnRH/LH surge at a dose capable of blocking pituitary responsiveness for approximately half the duration of the GnRH surge. Antagonist treatment did not result in any change in the time of onset of the GnRH surge, but there was no increase in LH that naturally occurs coincident with onset of the GnRH surge. Rather, a persistent increase in LH secretion was observed during the latter stages of the GnRH surge, indicating that the GnRH molecules secreted at this time were biologically active. Finally, a sensitive and specific ovine pituitary cell bioassay was used to test bioactivity of GnRH in hypophyseal portal blood during different phases of the GnRH surge. GnRH bioactivity in samples collected early in the GnRH surge was greater than that before the onset of the GnRH surge but no greater than that collected during the descending limb of the surge. The results of all three experiments fail to support the hypothesis that the LH surge ends because of a change in the nature of the GnRH secreted. Rather they show that GnRH secreted throughout the surge is biologically active. Thus, the termination of the LH surge before that of the GnRH surge occurs for reasons other than lack of a bioactive GnRH signal.

Nature and bioactivity of gonadotropin-releasing hormone (GnRH) secreted during the GnRH surge.

Previous studies have demonstrated a neural action of estradiol in inducing a surge of GnRH in the ewe. However, although the GnRH and LH surges began concurrently, the GnRH surge consistently continued well beyond the surge of LH. Three experiments were conducted to test the hypothesis that the termination of the LH surge results from the secretion of a relatively inactive variant of GnRH during the later phases of the GnRH surge. In the first experiment, hypophyseal portal blood collected during an estrogen-induced LH surge was analyzed for GnRH immunoreactivity using two antibodies having specificity for the N- or C-terminal portion of the GnRH molecule. The duration, amplitude, and time course of the GnRH surge were found to be similar irrespective of the antisera used. In a second experiment, a competitive GnRH antagonist was administered at the beginning of the estrogen-induced GnRH/LH surge at a dose capable of blocking pituitary responsiveness for approximately half the duration of the GnRH surge. Antagonist treatment did not result in any change in the time of onset of the GnRH surge, but there was no increase in LH that naturally occurs coincident with onset of the GnRH surge. Rather, a persistent increase in LH secretion was observed during the latter stages of the GnRH surge, indicating that the GnRH molecules secreted at this time were biologically active. Finally, a sensitive and specific ovine pituitary cell bioassay was used to test bioactivity of GnRH in hypophyseal portal blood during different phases of the GnRH surge. GnRH bioactivity in samples collected early in the GnRH surge was greater than that before the onset of the GnRH surge but no greater than that collected during the descending limb of the surge. The results of all three experiments fail to support the hypothesis that the LH surge ends because of a change in the nature of the GnRH secreted. Rather they show that GnRH secreted throughout the surge is biologically active. Thus, the termination of the LH surge before that of the GnRH surge occurs for reasons other than lack of a bioactive GnRH signal.

Dietary-induced suppression of pre-ovulatory progesterone concentrations in superovulated ewes impairs the subsequent in vivo and in vitro development of their ova

In the first of three experiments, eight ovariectomised Greyface ewes primed with exogenous progesterone were used to provide quantitative data on the effects of two contrasting feeding levels (0.3 vs. 1.4 × maintenance) on plasma progesterone concentrations. Over the 9 day study period, mean (± SEM) daily progesterone concentrations were 4.3 ± 0.13 and 3.3 ± 0.17 μg l −1 for the low and high feeding regimens, respectively ( P = 0.06), indicating that high feed intake suppressed circulating progesterone levels. The second experiment examined the effect in superovulated Finn-Dorset ewes of a diet supplying either 0.6 (Group L, n = 8) or 2.3 (Group H, n = 8) times their daily energy needs for maintenance, from 1 day before introduction of exogenous progesterone to the time of insemination, on plasma progesterone concentrations and the viability of ova recovered 4 days after insemination. Mean (± SEM) plasma progesterone concentrations were 4.5 ± 0.17 μg l −1 and 2.8 ± 0.16 μg l −1 for L and H ewes, respectively, during the 12 day priming period ( P < 0.001). Eight hours after progesterone withdrawal, levels had fallen to 0.9 ± 0.06 μg l −1 and 0.8 ± 0.07 μg l −1 , respectively, then rose to 17.8 ± 3.01 μg l −1 and 12.9 ± 2.50 μg l −1 ( P > 0.10) at ovum collection. Intervals (mean ± SEM) to oestrous onset (14.5 ± 0.38 h) and the luteinising hormone (LH) surge (27.1 ± 0.98 h) were unaffected by feed intake. Mean (± SEM) ovulation rates (8.1 ± 1.57 vs. 7.8 ± 1.10) and numbers of ova recovered (5.0 ± 1.39 vs. 4.8 ± 1.11) were also similar for each group. However, the proportions of ova considered viable (over 32 cells) at recovery were 0.53 and 0.22 for L and H groups, respectively ( P < 0.005). Following 72 h culture (Tissue Culture Medium-199 (M199) + 10% foetal calf serum (FCS)), 0.55 and 0.27, respectively, had developed to blastocysts ( P < 0.025). Of ova assessed as viable at recovery, similar proportions (0.86 vs. 0.75) from L and H treatments developed to blastocysts, with corresponding nuclei counts (mean ± SEM) of 55 ± 5.2 and 55 ± 13.2. The third experiment used 12 superovulated Greyface ewes, each offered a different feed level within the range 0.6–2.5 × maintenance, to determine the nature of the relationship between feeding level, pre-ovulatory progesterone concentrations and ovum development at Day 2 following insemination and subsequently during 7 day co-culture (M199 + FCS). Increases in feeding level were accompanied by linear decreases in plasma progesterone ( r 2 = 0.79, P < 0.001), the interval to oestrous onset ( r 2 = 0.52, P < 0.01) and timing of the LH surge ( r 2 = 0.32, P < 0.06). Although undetectable at ovum collection, and somewhat equivocal after 4 day culture, high feeding levels prior to ovulation reduced the proportion of ova (0.16 vs. 0.58) developing to or beyond the expanding blastocyst stage after 7 day culture. Quantitative indices of cell division and protein synthesis confirmed this. In conclusion, excessive feeding during follicular recruitment and oocyte maturation in superovulated ewes imparts a legacy of embryonic loss and developmental retardation.

Evidence that the switch from negative to positive feedback at the level of the pituitary gland is an important timing event for the onset of the preovulatory surge in LH in the ewe.

Experiments were performed to test the hypothesis that there is a negative feedback 'clamp' of ovarian hormones on the hypothalamus and pituitary gland during the follicular phase of the oestrous cycle that limits the secretion of GnRH and LH. GnRH secretion was monitored by sampling the hypophysial portal blood of ewes during the luteal phase of the oestrous cycle and either 24 h or 48 h after the induction of luteolysis by the injection of cloprostenol, a prostaglandin analogue. There was an increase in GnRH pulse frequency in the transition from the luteal to the follicular phase of the cycle. A reduction in the amplitude of GnRH pulses did not occur until 48 h after cloprostenol, suggestive of negative feedback at the level of the hypothalamus that is more profound in the latter part of the follicular phase. The responsivity of the pituitary gland to GnRH was monitored in ewes during the luteal phase of the oestrous cycle and 24 h or 48 h after cloprostenol. Injections of 250 ng or 1000 ng GnRH were given (i.v.) to ewes that had been anaesthetised to suppress endogenous secretion of GnRH and LH. Using the lower dose, the responses 48 h after cloprostenol were not significantly different from those in the luteal phase. With the higher dose of GnRH, a significant (P < 0.05) increase in mean responsivity was seen 48 h after cloprostenol. There was, however, a marked variation in response, with some ewes showing profound increases in LH secretion in response to GnRH and others showing responses that were similar to those obtained during the luteal phase of the cycle. These data are interpreted to mean that the secretion of LH is 'clamped' during the follicular phase of the oestrous cycle and the 'clamp' is only released near the time of the preovulatory LH surge. To test whether or not a rise in GnRH input to the pituitary gland could over-ride the 'clamp' on the pituitary secretion of LH in the late follicular phase of the cycle, sheep were treated 40 h after cloprostenol with either a bolus injection of 500 ng GnRH or four pulses of 125 ng GnRH given at 10-min intervals. These treatments caused small elevations in LH secretion but did not always cause preovulatory LH surges. In some cases, a small rise in LH secretion was induced by GnRH treatments and levels of LH in plasma returned to baseline with the preovulatory LH surge occurring a few hours later.(ABSTRACT TRUNCATED AT 400 WORDS)

Endocervical mucus pH is inversely related to serum androgen levels and waist to hip ratio

To determine whether an inter-relationship exists between endocervical mucus pH, serum androgen levels, and waist to hip (W:H) ratio. Donor insemination clinic, University of Sheffield, Jessop Hospital, Sheffield, United Kingdom. One hundred patients receiving donor insemination. Waist, hip, height, and weight measurements were recorded for 100 patients receiving donor insemination. At the onset of the LH surge when patients attended for their first insemination, a sample of endocervical mucus and blood was taken. The Insler score of the mucus was recorded and the pH was measured with multirange pH paper. Of the 100 patients studied, 10 cases had an endocervical mucus pH < 6 despite Insler scores of > 10. Serum T, DHEAS, and androstenedione levels were measured in the 10 patients with pH < 6 and in 26 patients selected at random from the 90 patients with pH > or = 6. The W:H ratio was significantly higher in the patients with pH < 6 than pH > or = 6. No significant difference was seen in body mass index between the latter groups. All serum androgen levels were significantly higher in the patients with pH < 6 than pH > or = 7. A potential link (serum androgen levels) between a powerful predictive factor of fertility (W:H ratio) and a potential mechanism of subfertility (low endocervical mucus pH) has been demonstrated. This observation warrants further detailed evaluation particularly as it has been shown that a low endocervical mucus pH may be corrected by simple inexpensive treatment.

Evidence that hypothalamic neuropeptide Y gene expression increases before the onset of the preovulatory LH surge.

Neuropeptide Y (NPY), a 36 amino acid residue peptide, is involved in stimulation of LHRH and LH surges on proestrus and those induced by ovarian steroids in ovariectomized (ovx) rats. Recently, we observed that NPY gene expression in the medial basal hypothalamus (MBH) was increased before the onset of the LH surge in the ovarian steroid-primed ovx rats. Since the ovarian steroidal milieu during the estrous cycle is markedly different from that prevailing after ovarian steroid injections in ovx rats, we evaluated in cycling rats the temporal relationship between MBH preproNPY mRNA levels and the preovulatory LH surge on the day of proestrus and compared that with diestrus II, concomitant with basal LH levels. PreproNPY mRNA levels in the MBH were measured by solution hybridization/RNAse protection assay, using a cRNA probe. On the day of diestrus II, preproNPY mRNA levels changed little between 1000 and 1800 h. Quite unexpectedly, preproNPY mRNA levels at 1000 h on proestrus were similar to diestrus II levels, despite additional exposure to ovarian steroids during this interval. However, from these low levels at 1000 h, the preproNPY mRNA profile displayed a biphasic rise. During the first phase, preproNPY mRNA rose significantly at 1200 h and remained elevated at 1300 and 1400 h concomitant with basal serum LH levels. Thereafter, a second rise in preproNPY mRNA began at 1500 h, peaked rapidly at 1600 h and declined significantly at 1800 h.(ABSTRACT TRUNCATED AT 250 WORDS)

Regulation of gonadotropin-releasing hormone gene expression in the rat during the luteinizing hormone surge.

In the present study we characterized GnRH gene expression in intact female rats across the estrous cycle and during a steroid-induced LH surge in ovariectomized (OVX) rats, using the quantitative ribonuclease protection assay. We measured cytoplasmic messenger RNA (mRNA) levels and nuclear primary transcript levels as an index of transcription. In Exp I, cycling rats were killed at 1100 or 1800 h on estrus, diestrus day 1, or diestrus day 2 (D2) or at 1100, 1500, 1800, or 2100 h on proestrus (P). In Exp II, proestrous rats were killed at the same time points or injected with pentobarbital (Pb) at the onset of the LH surge and killed on that day or the following day. In Exp III, a LH surge was induced in OVX rats treated with estradiol benzoate plus progesterone. Rats were killed at 1200, 1500, 1600, 1700, or 2100 h on the day of the surge. For all experiments, blood samples were collected and frozen for quantitation of LH by RIA. The preoptic area-anterior hypothalamus was dissected, and cytoplasmic and nuclear RNA were extracted and assayed separately by ribonuclease protection assay. In Exp I, cytoplasmic mRNA levels exhibited two significant peaks, one on D2 and another at 1500h on P. Primary transcript levels were significantly elevated only at 1500 h on P. In Exp II, proestrous rats and rats given Pb and killed the next day had a peak in cytoplasmic mRNA levels at 1500 h, which was blocked in rats given Pb and killed the same day. In Exp III with OVX rats, no significant changes in mRNA or primary transcript levels were observed between steroid or control groups. We hypothesize that the increase in cytoplasmic mRNA levels in cycling rats on D2 is probably due to a posttranscriptional mechanism, because it was not paralleled by changes in primary transcript levels, which would be expected if a transcriptional mechanism were involved. On P, both cytoplasmic mRNA and primary transcript levels changed, suggesting a transcriptional mechanism at this time.

Immunolocalization of Transforming Growth Factor a and Luteinizing Hormone Receptor in Healthy and Atretic Follicles of the Adult Rat Ovary1

At the onset of the LH surge in the rat, the proliferation of granulosa cells in preovulatory follicles is inhibited, aromatase activity is attenuated, and the cells enter a further phase of differentiation to form the CL. Whereas the LH surge induces these changes in the granulosa cells, it is possible that the actions of LH are mediated or modulated by paracrine and/or autocrine factors. A factor that may modulate the actions of LH is transforming growth factor alpha (TGF alpha). TGF alpha has the ability to inhibit granulosa cell growth and induce a change in the pattern of steroidogenesis from estrogen to progesterone synthesis, a change characteristic of early luteinization. To investigate whether there is a temporal relationship between the appearance of LH receptors and TGF alpha in the adult rat ovary, we localized LH receptors and TGF alpha during follicular development, atresia, and CL formation. In the interstitial compartment, the pattern of immunostaining for LH receptors and TGF alpha remained constant and was not affected by the stage of follicular development. As thecal cells differentiated from the stroma and formed several layers around the secondary follicle, the thecal cells stained positively for both LH receptors and TGF alpha. This immunostaining in thecal cells persisted in preantral, antral, and preovulatory follicles, even in follicles that became atretic. Granulosa cells did not contain detectable levels of LH receptor until the large antral stage of follicular development.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of transport stress and hydrocortisone on the oestrogen-induced luteinizing hormone surge in pigs

Pre-pubertal gilts were ovariectomized at 5–6 months age and 1 month later were challenged with oestradiol benzoate (OB; 20 μg kg−1 i.m.). In Experiment, 1 gilts were not transported (Treatment 1; n=6), or were transported for 24 h (Treatment 2; n=11) or 48 h (Treatment 3; n=6) after OB for a distance of 30 km. In Experiment 2, animals received saline (control; n=6) or an injection of hydrocortisone (HC; 500 mg per animal i.m.) at 24 h (HC 24; n=5) or 48 h (HC 48; n=5) after OB. In five of 11 gilts transported 24 h after OB (TR 24A), luteinizing hormone (LH) secretion was higher (P<0.05) during the negative feedback (6–48 h after OB) than in the remaining gilts (TR 24B) of the same treatment and the controls. LH concentrations during positive feedback (54–96 h after OB) did not differ between TR 24A and controls, but two of five gilts did not show LH surges. The TR 24A gilts had a higher rise in cortisol immediately after transportation than animals in the TR 24B group (P<0.05). During the positive feedback there was a significant increase of LH concentrations (ng ml−1) in gilts transported 48 h after OB when compared with the control and TR 24A groups (1.70±0.40 vs. 0.85±0.30 and 0.87±0.25, respectively; P<0.05). The LH surge parameters (times of emergence and greatest surge and amplitude) did not differ between the control, TR 24A, TR 24B and TR 48 groups. In Experiment 2, administration of HC caused significant elevation of cortisol in the blood of treated groups. An increase of LH level during ‘late’ negative feedback occurred in only one of five gilts given HC 24 h after OB. The positive feedback parameters were not affected in the two groups given HC with the exception of the earlier onset (h) of the LH surge in HC 24 group when compared with the control and HC 48 groups (52.8±1.2 vs. 62.0±2.0 and 61.2±2.2, respectively; P<0.05). These results demonstrate that transport can modify the profile of negative and positive feedback of LH secretion in ovariectomized pigs in different ways depending on the time of the gilts transportation after OB injection. Deviations of the positive and negative phases of feedback are not caused by cortisol.

Luteinized unruptured follicle: morphology, endocrine function and blood flow changes during the menstrual cycle

A case of spontaneous luteinized unruptured follicle syndrome is presented with full documentation of hormonal, morphological and haemodynamic changes. Changes in uterine blood flow were also noted. Growth of the leading follicle was slow during the follicular phase of the cycle. After the luteinizing hormone (LH) surge, growth of the follicle was more rapid. Concurrently, the follicle developed internal echogenicity with ultrasonic evidence of separation of the granulosa cell layer. The follicle was no longer visible 144 and 132 h after the LH rise and peak respectively. There was no primary progesterone rise associated with either the LH rise or peak, but a secondary progesterone rise occurred 42 h after the onset of the LH surge. Peri-follicular blood flow velocity was detected for the first time on cycle day 5 and appeared to rise after the onset of the LH surge. Peri-follicular blood flow velocity appeared to reduce after the LH surge to values associated with the follicular phase. These observations are consistent with an association of a primary granulosa cell defect with luteinized unruptured follicle syndrome which would account for the initial slow follicular growth, absent primary progesterone rise and reduction in blood flow in the wall of the follicle after the LH surge.

Control-Oriented High-Frequency Turbomachinery Modeling: Single-Stage Compression System One-Dimensional Model

In this paper, a one-dimensional unsteady compressible viscous flow model of a generic compression system previously developed by the authors is applied to a multistage axial compressor experimental rig configured for single-stage operation. The required model parameters and maps are identified from experimental data. The resulting model is an explicit system of nine first-order ODEs. The model inputs are compressor speed, nozzle area, compressor discharge bleed area, plenum bleed area, inlet total pressure and entropy, and nozzle and bleed exit static pressures. The model and experimental data are compared with respect to both open-loop uncontrolled and closed-loop controlled behaviors. These comparisons focus on (i) forced transients and (ii) global nonlinear dynamics and bifurcations. In all cases the agreement between the model and experimental data is excellent. Of particular interest is the ability of the model, which does not include any hysteretic maps, to predict experimentally observed hysteresis with respect to the onset and cessation of surge. This predictive capability of the model manifests itself as the coexistence of a stable equilibrium (rotating stall) and a stable periodic solution (surge) in the model at a single fixed set of system input values. Also of interest is the fact that the controllers used for closed-loop comparisons were designed directly from the model with no a posteriori tuning of controller parameters. Thus, the excellent closed-loop comparisons between the model and experimental data provide strong evidence in support of the validity of the model for use in direct model based controller design. The excellent agreement between the model and experimental data summarized above is attributed in large part to the use of effective lengths within the model, as functions of axial Mach number and nondimensional compressor rotational speed, as prescribed by the modeling technique. The use of these effective lengths proved to be far superior to the use of physical lengths. The use of these effective lengths also provided substantial improvement over the use of physical lengths coupled with fixed first-order empirical lags, as proposed by other authors for the modeling of observed compressor dynamic lag. The overall success of this model is believed to represent a positive first step toward a complete experimental validation of the approach to control-oriented high-frequency turbomachinery modeling being developed by the authors.

Coincident plasma ACTH and corticosterone time series: Comparisons between young and old rats

Senescence is accompanied by a reduced ability to respond to a variety of physical and behavioral stressors. A sizable literature has been devoted to the interplay between hypothalamic-pituitary-adrenocortical axis dysfunction and senescence; yet, the precise interactions remain an enigma. Adrenocorticotropic hormone (ACTH) is secreted in pulsatile bursts generating complex signals in the plasma compartment that must be “read” by adrenocortical cells in order to initiate appropriate secretory responses. We have previously demonstrated subtle differences between young and old rats in the pattern of fluctuations in plasma ACTH concentrations over time, despite no difference in mean levels. The present work addressed the physiological significance of these differences in the plasma ACTH signal by analyzing the corresponding plasma corticosterone concentration time series and the relationship between these two hormones over time. Time series of integrated 10-min ACTH and corticosterone concentrations were collected over 4 h at the time of diurnal activation and analyzed in the time and frequency domains. The time of onset of the diurnal surge occurred 20 min later in old rats, and the ratio of corticosterone to ACTH was less at the time of onset and peak of the diurnal surge. Corticosterone levels were lower in old rats and mean ACTH and corticosterone levels were correlated in young but not old rats, as were maximum levels of the two hormones. Cross-correlation of ACTH and corticosterone time series and comparison of spectra were consistent with smoother fluctuations in plasma corticosterone in old animals with less variability at time scales less than 55 min. We conclude that age may be associated with a delay in diurnal activation of the HPA axis, a loss of sensitivity of adrenal corticosterone secretion to plasma ACTH levels, and a relative loss of high frequency variability in the corticosterone signal, as seen in many physiological systems with age.

Flow Physics Leading to System Instability in a Centrifugal Pump

The off-design performance in a centrifugal pump is investigated experimentally. The objective is to identify flow features that lead to the onset of surge as the fundamental pumping system instability. Results show that there are primarily two reasons for the onset of surge as the flow is reduced in the pump studied: (a) adverse flow in the tongue region, and (b) destabilizing effect of the pipe diffuser. The former is due to premature diffusion of the flow entering the tongue region, which is manifested by increased flow recirculation through the tongue/impeller gap and flow separation on the volute outer side-wall opposite the tongue. These effects in the tongue region flow coupled with the destabilizing behavior of the pipe diffuser lead to the eventual unstable pump operation.

Gene expression in a subpopulation of luteinizing hormone-releasing hormone (LHRH) neurons prior to the preovulatory gonadotropin surge

Gene expression in luteinizing hormone-releasing hormone (LHRH) neurons was analyzed during the periovulatory period to (1) characterize temporal patterns of LHRH gene expression and their relationship(s) to gonadotropin surges, and (2) determine if any such changes are uniform or dissimilar at different rostrocaudal levels of the basal forebrain. The number of neurons expressing mRNA for the decapeptide, and the relative degree of expression per cell were analyzed using in situ hybridization and quantitative image analysis. Rats were killed at 1800 hr on metestrus (Met), 0800 hr, 1200 hr, 1800 hr, and 2200 hr on proestrus (Pro), or 0200 hr, 0800 hr, and 1800 hr on estrus (E; n = 5-6 rats/group). All sections were processed for LHRH mRNA in a single in situ hybridization assay. Sections were atlas matched and divided into four rostrocaudal groups for analysis: vertical limb of the diagonal band of Broca (DBB), rostral preoptic area/organum vasculosum of the lamina terminalis (rPOA/OVLT), medial preoptic area (mPOA), and suprachiasmatic/anterior hypothalamic area (SCN/AHA). Plasma LH and FSH levels from all animals were analyzed by RIA. The labeling intensity per cell was similar among all time points at all four rostrocaudal levels. The number of cells expressing LHRH mRNA, however, varied as a function of time of death during the estrous cycle, and this temporal pattern varied among the four anatomical regions. At the level of the mPOA, the number of cells was highest at 1200 hr on Pro, and then declined and remained low throughout the morning of E. At the level of the rPOA/OVLT, the greatest number of LHRH neurons was noted later in Pro, at 1800 hr, dropping rapidly to lowest numbers at 2200 hr. No significant changes in LHRH cell number occurred at the DBB or SCN/AHA levels. At all anatomical levels, the secondary surge of FSH was unaccompanied by any change in the number of neurons expressing LHRH mRNA. These data demonstrate that (1) the number of detectable LHRH mRNA-expressing cells fluctuates during the periovulatory period and (2) peak numbers of LHRH-expressing cells are attained in the mPOA before the onset of the LH surge and before peak LHRH cell numbers are seen at more rostral levels. A model is proposed in which gene expression in this subpopulation of LHRH neurons may be activated by preovulatory estrogen secretion and acutely reduced following the proestrous surge of progesterone.

Steroidogenic cytochrome P450 enzyme messenger ribonucleic acids and follicular fluid steroids in individual follicles during preovulatory maturation in the pig.

Changes in follicular concentrations of steroidogenic cytochrome P450 enzyme mRNAs were determined during preovulatory maturation. RNA was isolated from 59 individual follicles dissected from 18 pigs during altrenogest-synchronized preovulatory follicular maturation: at Day 1 (pre-follicular phase), Day 3 (early follicular phase), Day 5 (mid-follicular phase), and Day 7 (late follicular phase, 24-36 h after the onset of the LH surge). Follicular fluid was aspirated for steroid RIA. RNA was also isolated from pooled granulosa cells, theca tissue, and luteal tissue. RNA was analyzed by Northern and slot-blot procedures using cDNA probes to human aromatase cytochrome P450 (P450arom), porcine 17 alpha-hydroxylase cytochrome P450 (P450(17) alpha), and porcine cholesterol side-chain cleavage cytochrome P450 (P450scc). P450arom mRNA was expressed in both granulosa and theca interna cells but was not detectable in luteal tissue from Day 13 of the estrous cycle. Follicular P450arom mRNA concentration tended to increase between Days 1 and 5 and decreased (p < or = 0.05) by 92% between Days 5 and 7. Follicular fluid estradiol-17 beta concentration increased 17-fold between Days 1 and 5 and then decreased (p < or = 0.05) by 96% between Days 5 and 7. P450(17) alpha mRNA was present in theca interna but was not detected in granulosa cells or luteal tissue. Follicular P450(17) alpha mRNA concentrations did not differ significantly among days, but the content per follicle increased (p < 0.05) between Days 1 and 5 and decreased (p < 0.05) between Days 5 and 7.(ABSTRACT TRUNCATED AT 250 WORDS)

Transport stress delays the oestradiol-induced LH surge by a non-opioidergic mechanism in the early postpartum ewe.

The present study was designed to investigate whether transport, a mild environmental stressor, could affect the oestradiol-induced LH surge in postpartum ewes and, if so, the mechanism involved. Welsh Mountain ewes, with lambs removed at parturition (day 0) and hand-milked 12 and 48 h later, were given 50 micrograms oestradiol benzoate intramuscularly at various times postpartum. Blood samples were taken via an indwelling jugular venous catheter every 2 h from 8 to 24 h after oestradiol injection. All results are given as means +/- S.D. On day 1 oestradiol was unable to induce an LH surge in any ewe. Transport (10-14 h after oestradiol) delayed the onset of the oestradiol-induced LH surge on day 14 (17.5 +/- 1.7 vs 14.4 +/- 2.0 h, n = 5 each; P < 0.05), but not on day 28 (14.9 +/- 2.0 vs 14.0 +/- 2.4 h, n = 5 out of 7). Transport had no effect on the amplitude of the surge on either day. Naloxone treatment (1 mg/kg per 2 h) was unable to prevent the delay caused by transport (18.0 +/- 1.1 vs 17.5 +/- 1.7 h, n = 8 each), and did not affect the amplitude of the surge (28.4 +/- 5.3 vs 28.1 +/- 2.3 ng/ml, n = 8 each).(ABSTRACT TRUNCATED AT 250 WORDS)

Preferential induction of c-fos immunoreactivity in vasoactive intestinal polypeptide-innervated gonadotropin-releasing hormone neurons during a steroid-induced luteinizing hormone surge in the female rat

In small rodents, reproduction is critically dependent on the integrity of the circadian oscillator of the brain, the suprachiasmatic nucleus (SCN). Lesions of the SCN induce persistent estrus (anovulation) in intact female rats, whereas estrogen implantation in ovariectomized rats results in daily LH surges, which disappear after SCN lesions. Vasoactive intestinal polypeptide (VIP), a peptide synthesized in cell bodies of the SCN, has been implicated in the regulation of LH release. Recently, we have provided immunocytochemical evidence for a VIP-containing neuronal projection from the SCN to the GnRH system. This suggests that VIP from the SCN may modulate LH release via a direct influence on GnRH neurons. To investigate the involvement of VIP input on GnRH neurons and SCN neurons in the generation of a LH surge, we used immunoreactive c-fos as a marker for cell activation in ovariectomized mature rats and immature rats treated with steroids. VIP-containing fibers were observed in apposition to a substantial portion of the GnRH neurons containing c-fos. Expression of c-fos was more frequently observed in VIP-innervated GnRH neurons than in GnRH neurons in general. This difference in activation was most pronounced during the onset of the LH surge. In SCN neurons, steroid treatment did not induce c-fos immunoreactivity before or during the LH surge. The present results indicate that VIP-containing fibers, possibly originating in the SCN, are involved in the initiation of the LH surge. In view of the reported inhibitory effects of VIP on LH release, it is suggested that the role of VIP input in this respect is permissive.