Effect of bST administration on plasma concentrations of IGF-I and follicular dynamics and ovulation during the interovulatory cycle of sheep and goats.

This study used a comparative approach to gather clinical information to assess the effect of bovine somatotropin (bST) on follicular dynamics and ovulation in sheep and goats during an interovulatory cycle. The performance of general markers of ovarian function and specific features of follicular dynamics obtained by daily ultrasonography (US) were used to assess the hypothesis that bST, associated with supraphysiological levels of IGF-I, was able to disrupt the follicular dynamics and ovulation in Highlander ewes and Saanen goats. In Exp 1, 15 ewes and 14 goats were estrous-synchronized (P4-6 days + PGFα d-6) and then allocated to a bST-treated group (50 and 100 mg, Lactotropin®; n = 5 females each) and to an untreated control group (5 ewes and 4 goats) to assess the activity of bST through plasma IGF-I (RIA). In Exp 2, 12 animals from each species were synchronized. At day 6, they were divided into a bST-group (100 mg in sheep and 50 mg in goats, n = 6 each) and an untreated control group (n = 6 each). Starting at day 6 and up to 22 days after ovulation in sheep and 25 days in goats, each female was subjected to daily US (10 mHz probe) to assess follicular and luteal (CL) dynamics and ovulation. This included assessments of both general ovarian features and specific follicular wave features. Our results showed that bST increased plasma IGF-I by day 3 (p < 0.01) when compared to the control group. Moreover, these concentrations were maintained for at least 10 days in sheep and 10 days in goats before returning to pre-treatment concentrations. Increases in IGF-I after bST doses were similar in terms of a daily and total amount (P > 0.10). Results from Exp.2 indicate that in sheep, bST administration had a subtle inhibitory effect on follicular function. However, bST in goats had a stronger influence, extending the interovulatory cycle (P = 0,034), increasing the number of follicular waves during the period (P = 0.003), and reducing the functional potential of large follicles as measured by their lower follicular diameter (P = 0.02), duration of the follicle waves (P = 0.02), and persistence of follicles after reaching their maximum diameters (P = 0.04). In addition, untreated sheep and goats shared common patterns of terminal follicular development and ovulations characterized by overlapping between follicular waves and ovulations of follicles from different waves, features not seen in cattle.

Embryo splitting affects the transcriptome during elongation stage of in vitro-produced bovine blastocysts.

Embryo splitting has been used for the production of identical twins and to increase the pregnancy rate per available embryo. Split blastocysts can develop to term; however, little is known about the impact on gene expression of split embryos, especially at the whole transcriptome level. This work was aimed to evaluate the effect of blastocyst splitting on global gene expression profile at the elongation stage. For that, split and time-matched nonsplit (control group) bovine blastocysts were transferred to a bovine recipient and recovered at Day 17 of development. The number of collected embryos, their size, and global gene expression was compared between both groups. From 16 transferred split embryos, six (37.5%) were collected, whereas nine elongated were recovered from 17 nonsplit (52.9%). Neither the recovery rate nor the average length of the elongated embryos was significantly different between both groups. However more than 50% of embryos from the control group had a length surpassing 100 mm, whereas only 33% of the split embryos reached that size. Global gene expression was performed in individual elongated embryos from both groups using Two-Color Microarray-Based Gene Expression Analysis. From detected genes, 383 (1.31%) were differentially expressed between both groups, among them, 185 (0.63%) were downregulated and 198 (0.67%) genes were upregulated in split embryos. Bioinformatic analysis of differentially expressed genes revealed that embryo splitting affects transcriptomes of resulting elongated embryos, mainly downregulating genes involved in matrix remodelation, control of growth, detoxification, and transport of metabolites. These in turns might have a detrimental impact on the developmental potential of produced embryos.

Human baroreflex rhythms persist during handgrip and muscle ischaemia.

AIM: To determine whether physiological, rhythmic fluctuations of vagal baroreflex gain persist during exercise, post-exercise ischaemia and recovery. METHODS: We studied responses of six supine healthy men and one woman to a stereotyped protocol comprising rest, handgrip exercise at 40% maximum capacity to exhaustion, post-exercise forearm ischaemia and recovery. We measured electrocardiographic R-R intervals, photoplethysmographic finger arterial pressures and peroneal nerve muscle sympathetic activity. We derived vagal baroreflex gains from a sliding (25-s window moved by 2-s steps) systolic pressure-R-R interval transfer function at 0.04-0.15 Hz. RESULTS: Vagal baroreflex gain oscillated at low, nearly constant frequencies throughout the protocol (at approx. 0.06 Hz - a period of about 18 s); however, during exercise, most oscillations were at low-gain levels, and during ischaemia and recovery, most oscillations were at high-gain levels. CONCLUSIONS: Vagal baroreflex rhythms are not abolished by exercise, and they are not overwhelmed after exercise during ischaemia and recovery.

Follicular dynamics, interval to ovulation and fertility after AI in short-term progesterone and PGF2α oestrous synchronization protocol in sheep.

The study was aimed to assess the influence that short-term progesterone treatments have on follicular dynamics, oestrus and ovulation in sheep. The treatment was tested thereafter in a field trial to assess its fertility after AI with fresh semen. In a first experiment, 12 ewes without CL were grouped to receive a new (n = 6) or used CIDR (n = 6) for 7 days and blood samples were obtained to follow plasma progesterone profiles. In a second experiment, 39 cycling ewes were synchronized by a 7-day P4+PGF2α protocol using a new (n = 20) or a 7-day used CIDR (n = 19). Half of both groups received 400 IU eCG and half remained untreated as controls. Ultrasound ovarian examination and oestrous detection were used to compare follicular dynamics, oestrus and ovulation in both groups. In a third experiment, 288 ewes in 3 farms were synchronized by the short-term P4+PGF2α+eCG protocol and ewes were AI with fresh semen 24 h after oestrous detection. Lambing performance was used to test the fertility of the treatment. In Experiment 1, ewes with new inserts presented higher P4 concentration than ewes with used inserts throughout the sampling period (p < 0.05) and exhibited a P4 peak at days 1-2 of the treatment that was not observed in ewes with used inserts. In Experiment 2, ewes treated with new and used inserts show similar ovarian and behavioral traits (p > 0.10). However, ewes treated with eCG show shorter interval to oestrus (p = 0.004) and tend to have larger mature CL (p = 0.06). In Experiment 3, oestrous presentation and lambing performance after AI with fresh semen was considered normal compared to published results. Results suggest that the oestrous synchronization protocol based on P4+PGF2α allows little control of follicular dynamics without compromising fertility after AI with fresh semen provided that eCG is added at the end of the treatment.

MicroRNA expression profiling of elongated cloned and in vitro-fertilized bovine embryos.

The objective of this study was to identify microRNAs (miRNAs) expressed in bovine (Bos Taurus) cloned embryos at Day 17 of development (Day 0=day of nucleus transfer or in vitro fertilization) during elongation. Day 7 bovine expanded blastocysts produced by hand made cloning (HMC) or in vitro fertilization were bulk-transferred to synchronized recipient cattle (48 HMC embryos to 10 recipients and 28 in vitro-produced embryos to four recipients). Elongated embryos were retrieved at Day 17; miRNAs were isolated and subjected to microarray screening using custom composite slides spotted with human, mouse, and rat and in silico-predicted miRNAs. An initial profile of expressed miRNAs was determined in cloned embryos and somatic donor cells; this profile changed after somatic cell nucleus transfer, identifying differentially expressed miRNAs between cloned and in vitro-produced bovine embryos. Furthermore, microarray data were validated using a miRNA-specific quantitative reverse transcription-polymerase chain reaction (qRT-PCR) approach (miR-Q). There was an 83% correlation (P=0.01) between microarray and qPCR data. Based on qRT-PCR, correct reprogramming of some miRNAs from the donor cells was confirmed in cloned bovine embryos, whereas other somatic miRNAs were not appropriately reprogrammed. Some of the miRNAs that were equally reprogrammed clustered on the same chromosomal location in the bovine genome. In conclusion, reprogramming of miRNAs seemed to occur in cloned bovine embryos. This could have profound implications for elucidating nuclear reprogramming in somatic cloning, as well as for the role of miRNAs in preimplantation mammalian development.

CASA assessment of kinematic parameters of ram spermatozoa and their relationship to migration efficiency in ruminant cervical mucus.

Sperm motility is an indicator of male fertility because of its importance for sperm migration through the female genital tract and for gamete interaction at fertilization. This study analyses the relationship between computer assisted semen analysis (CASA) motility patterns and sperm migration of rams in ruminant cervical mucus. In experiment 1, spermatozoa extended with sperm analysis medium (SAM) and seminal plasma were compared in terms of motility. In experiment 2, 56 semen samples were collected either with artificial vagina (AV) or electroejaculator to be compared in terms of motility performance. In experiment 3, 104 ejaculates collected by AV from 26 males were analysed via the CASA system to characterize their motility patterns. In experiment 4, ejaculates from pairs of rams (20 rams in total) were simultaneously assessed for mucus migration (ovine, caprine, bovine) and motility patterns to evaluate the correlations between both parameters. Semen collected by AV and extended in SAM allows the most reliable assessment for sperm motility. Ram spermatozoa move fast and follow a linear trajectory compared with other ruminants. Continuous line velocity (VCL) and average path velocity (VAP) are the only sperm kinematic parameters that presented significant positive correlations with the ability to migrate in sheep cervical mucus (p < 0.05). Continuous line velocity, VAP, straight line velocity and linearity are highly significantly related with migration efficiency in goat cervical mucus (p < 0.01) and only lateral head displacement is negatively related to sperm migration in bovine cervical mucus (p < 0.05). These results suggest that specific kinematic parameters confer the ability of spermatozoa to colonize and migrate through epithelial mucus with different rheological properties.

In vitro fertilization and development of OPU derived goat and sheep oocytes.

The recent upgrade in IVP technology seen in cattle can be adapted to embryo production in small ruminants to overcome limitations exhibited by surgical procedures on preserving the reproductive potential of donors and the efficiency of embryo production. The aim of the present study was to assess the current procedures used in cattle for the production of IVP embryos in goats and sheep based on laparoscopic-aided ovum pick-up (LOPU) supplied oocytes. Sexually matured goat and sheep donors were treated during the breeding season with FSH and subjected to laparoscopic-guided follicular puncture under general anaesthesia. The collected cumulus-oocyte complexes were matured in medium 199 and fertilized by frozen-thawed spermatozoa using Talp medium supplemented with heparin and oestrus-sheep serum. Cleaved ova were either cultured in sheep in vitro fertilization medium plus amino acids or transferred to sheep oviducts. Blastocyst rate, hatching rate and development rate up to term were used as markers of embryo function. The results obtained for goat and sheep involving 30 and 35 donors respectively (10 and 9 LOPU sessions) were 81.2% and 85.2% of oocyte collection rate; 88.3% and 98.6% oocyte incubation rate; 85.6% and 76.0% fertilization rate; 82.4% and 93.4% of cleavage rate; 50.0% and 61.5% IVP blastocyst rate; 42.1% and 45.5% blastocyst rate in oviducts; 73.0% and 66.7% embryo survival up to term, respectively. The results are comparable to those obtained in small ruminants and in bovines suggesting that requirements for embryo production and development are similar.

Computer-assisted analysis of sperm motion in goats and its relationship with sperm migration in cervical mucus.

In vitro sperm migration in cervical mucus relates to sperm concentration at the utero-tubal junction and to in vivo fertilization performance in goats. The present study aimed to characterize, using Computer-Assisted Sperm Analysis (CASA), motility patterns depicted by buck sperm and their relation to the migration efficiency in homologous (goat) and heterologous (heifer) cervical mucus in vitro. Semen was collected from 23 sexually mature bucks from three breeds by artificial vagina and sperm were assessed for motility parameters with a Hobson Sperm analyzer following extension in Sperm Analysis Medium (SAM). To study the relationship between kinematics parameters and the ability of sperm to migrate in cervical mucus, in a first experiment, motility performance of buck sperm suspended in SAM was compared against seminal plasma. In a second experiment, kinematics parameters of sperm were characterized. In a third experiment, bucks with sperm that differed in specific motion parameters were compared for the ability of their sperm to migrate through goat and bovine cervical mucus collected at estrus. In a fourth experiment, ejaculates that were compared in their migration ability and were assessed simultaneously for their motility parameters. Overall, sperm suspended in SAM medium had better velocity and similar linearity and lateral head displacement than those suspended in seminal plasma; furthermore, caprine sperm swam relatively fast (relative to bovine and ovine sperm), following a very linear trajectory. Under the conditions used, velocity parameters, linearity and lateral head displacement seemed to be related to sperm migration efficiency in homologous mucus but not in bovine cervical mucus.

A five-year population study of persons involved in the mental health and local correctional systems: implications for service planning.

This study investigates the relationship between recipients of mental health services and incarceration within local jails. The analyses showed that male and female recipients of mental health services are incarcerated in local jails at a higher rate than the general community but that most are not long-term incarcerations. The study further showed that the rate of incarceration for recipients of mental health services decreases with age but the relative risk of incarceration, when compared with the general population, actually increases with age. In addition, across all age groups female recipients of mental health services were found to be at greater risk of incarceration than male recipients.

Utilization of local jails and general hospitals by state psychiatric center patients.

The idea that the deinstitutionalization of state psychiatric centers has resulted in increased utilization of general hospitals and correctional facilities by people with severe and persistent mental illness is widely held. This hypothesis of trans-institutionalization was tested by examining hospitalization and incarceration rates of people who had been or would be institutionalized in state psychiatric centers in 16 upstate New York counties. The results do not support the hypothesis of trans-institutionalization. Assumptions underlying the hypothesis are examined, potential explanations for the observed patterns are discussed, and areas for further research are suggested.

Nine months in space: effects on human autonomic cardiovascular regulation.

We studied three Russian cosmonauts to better understand how long-term exposure to microgravity affects autonomic cardiovascular control. We recorded the electrocardiogram, finger photoplethysmographic pressure, and respiratory flow before, during, and after two 9-mo missions to the Russian space station Mir. Measurements were made during four modes of breathing: 1) uncontrolled spontaneous breathing; 2) stepwise breathing at six different frequencies; 3) fixed-frequency breathing; and 4) random-frequency breathing. R wave-to-R wave (R-R) interval standard deviations decreased in all and respiratory frequency R-R interval spectral power decreased in two cosmonauts in space. Two weeks after the cosmonauts returned to Earth, R-R interval spectral power was decreased, and systolic pressure spectral power was increased in all. The transfer function between systolic pressures and R-R intervals was reduced in-flight, was reduced further the day after landing, and had not returned to preflight levels by 14 days after landing. Our results suggest that long-duration spaceflight reduces vagal-cardiac nerve traffic and decreases vagal baroreflex gain and that these changes may persist as long as 2 wk after return to Earth.

Controlled breathing protocols probe human autonomic cardiovascular rhythms.

The purpose of this study was to determine how breathing protocols requiring varying degrees of control affect cardiovascular dynamics. We measured inspiratory volume, end-tidal CO2, R-R interval, and arterial pressure spectral power in 10 volunteers who followed the following 5 breathing protocols: 1) uncontrolled breathing for 5 min; 2) stepwise frequency breathing (at 0.3, 0.25, 0.2, 0.15, 0.1, and 0.05 Hz for 2 min each); 3) stepwise frequency breathing as above, but with prescribed tidal volumes; 4) random-frequency breathing (approximately 0.5-0.05 Hz) for 6 min; and 5) fixed-frequency breathing (0.25 Hz) for 5 min. During stepwise breathing, R-R interval and arterial pressure spectral power increased as breathing frequency decreased. Control of inspired volume reduced R-R interval spectral power during 0.1 Hz breathing (P < 0.05). Stepwise and random-breathing protocols yielded comparable coherence and transfer functions between respiration and R-R intervals and systolic pressure and R-R intervals. Random- and fixed-frequency breathing reduced end-tidal CO2 modestly (P < 0.05). Our data suggest that stringent tidal volume control attenuates low-frequency R-R interval oscillations and that fixed- and random-rate breathing may decrease CO2 chemoreceptor stimulation. We conclude that autonomic rhythms measured during different breathing protocols have much in common but that a stepwise protocol without stringent control of inspired volume may allow for the most efficient assessment of short-term respiratory-mediated autonomic oscillations.

A solution to the problem of jail suicide.

Suicide in local jail facilities can be reduced through interagency cooperation and the implementation of core services. In 1985, the State of New York implemented a comprehensive suicide prevention program within its upstate local jail facilities. The program utilized key coordination strategies and risk-management service components. It addressed not only the immediate needs of inmates with high-risk profiles, but also focused on the impact of the stressful jail experience on this already vulnerable population. Despite a nearly 100% increase in the jail population, there has been more than a 150% decrease in jail suicides since program implementation.

Linear and nonlinear contributions to step responses in cat retinal ganglion cells.

We measured excitatory and inhibitory step responses of cat retinal ganglion cells to square wave contrast reversal of stationary sinusoidal gratings. In most Y-cells the initial increase in firing rate (early peak) of the excitatory responses was followed by a distinct second increase in firing rate (late peak). Analysis of the spatial frequency and spatial phase dependence of the two peaks indicated that the early peak appears to be produced by the spatially linear center mechanism, while the late peak appears to be produced by the rectifying subunits described by Hochstein and Shapley (1976) Journal of Physiology, London, 262, 237-264, 265-284. These results indicate that the presence of two peaks in ganglion cell step responses is the result of two excitatory inputs with different time courses, and that inhibitory inputs are not required to explain the appearance of these responses.

Vernier acuity in amblyopic and nonamblyopic children.

We measured vernier acuity in normal and amblyopic children using a procedure that resembles a video game and is suitable for testing most children older than 4 years old. In this procedure, subjects align bars using two keys of a computer keypad. Following binocular practice trials, monocular vernier acuity was measured in 38 control children, 5 to 15 years old, and in 18 children with histories of amblyopia. Vernier acuity was defined as the standard deviation of adjusted position across a block of six trials. Vernier acuity improved as a function of age in the control subjects, indicating developmental improvement. Amblyopic subjects with strabismus (n = 5) and with both anisometropia and strabismus (n = 8) showed markedly impaired vernier acuity in their amblyopic eyes, with vernier acuitys four to five times larger than those of age-matched controls. However, the amblyopic subjects who were anisometropic (n = 5), without any history of strabismus, were not significantly different from control subjects in either eye. Testing with bars and gratings gave similar results. To our knowledge, this is the first study showing that vernier acuity deficits seen in adult amblyopes are also seen in child amblyopes.

Multiple estimation of concentrations in immunoassay using logistic models.

Immunoassay techniques yield estimates of concentrations of analytes based on comparison to known concentrations of a reference solution. The use of the nonlinear logistic model makes the error estimates and confidence levels approximate. When the goal of such a study is estimation of several unknowns, methods in common usage do not account for 'simultaneous' inference, i.e. the repeated use of the standard curve for estimating several concentrations. Alternative methods are described which take multiple use of the reference curve into account. Simulations using normally distributed data with variance proportional to a power of the mean compare different methods of obtaining calibration intervals and illustrate the approximate nature of all such techniques. Calibration intervals based on simple, commonly used methods do not provide the coverage promised, even for one-at-a-time estimation, and are not suited for multiple estimation and comparison.

Dose-dependent effect of heparin on fertilizing ability of goat spermatozoa.

Intact bovine oocytes were used to study the effect of heparin on goat IVF. Oocytes were matured in Medium 199 plus estrous sheep serum. Fresh semen was incubated for 4 h at room temperature, and spermatozoa were then resuspended in medium Talp plus serum and incubated further for 1 h at 39 degrees C in 5% CO(2) in air. Later, spermatozoa were resuspended in Talp plus serum and heparin and were then incubated in microdrops until the oocytes were matured. In Experiment 1, the effect of heparin on spermatozoa from individual males was studied by a dose-response curve. In Experiment 2, the timing of sperm penetration in matured oocytes was studied to assess the stage at which the action of heparin could be expressed in the fertilization process. In Experiment 3, heparin from the same source but at different grades of bioactivity was adjusted for bioactivity and its effect on spermatozoa was compared in terms of penetration rates in order to identify heparin-dependent variations on goat IVF. In Experiment 4, the influence of calcium on the effect of heparin at different levels of bioactivity on the fertilizing ability spermatozoa was assessed as in Experiment 3. In Experiment 5, different batches of heparin from the same source and grade of bioactivity were compared as above. The results suggest that 1) heparin stimulates fertilization rates following a comparable pattern between males; 2) the most probable site of action is at the stage of sperm capacitation; and 3) provided that the source and grade of bioactivity is preserved, heparin maintains the efficiency of sperm penetration into matured oocytes.

Evaluation of nearest-neighbor methods for detection of chimeric small-subunit rRNA sequences.

Detection of chimeric artifacts formed when PCR is used to retrieve naturally occurring small-subunit (SSU) rRNA sequences may rely on demonstrating that different sequence domains have different phylogenetic affiliations. We evaluated the CHECK_CHIMERA method of the Ribosomal Database Project and another method which we developed, both based on determining nearest neighbors of different sequence domains, for their ability to discern artificially generated SSU rRNA chimeras from authentic Ribosomal Database Project sequences. The reliability of both methods decreases when the parental sequences which contribute to chimera formation are more than 82 to 84% similar. Detection is also complicated by the occurrence of authentic SSU rRNA sequences that behave like chimeras. We developed a naive statistical test based on CHECK_CHIMERA output and used it to evaluate previously reported SSU rRNA chimeras. Application of this test also suggests that chimeras might be formed by retrieving SSU rRNAs as cDNA. The amount of uncertainty associated with nearest-neighbor analyses indicates that such tests alone are insufficient and that better methods are needed.

Effects of neonatal decortication on the social play of juvenile rats.

The effects of radical neonatal decortication on the social play of juvenile rats, as well as the effects of neonatal ablation of frontal or parietal cortex, were examined in this series of experiments. When total decorticates were tested in like-lesioned pairs, the frequency of pinning was reduced by about 50% and their average pin durations were shorter. Nevertheless, the play of decorticates appeared essentially normal in general appearance, and did not differ from controls in a measure of overall play vigor using an electronic activity platform. Further, there were no differences in pin frequencies when controls and decorticates were paired together in cross-lesion testing. Separate tests of play solicitation behaviors did not detect any differences between controls and decorticates suggesting that play motivation was essentially intact after decortication. No deficits in pinning resulted from frontal ablations; however, pin durations were shorter in like-lesion testing. In cross-lesion testing, there was an increase in dorsal contacts and a trend toward shortening of pin durations. Parietal aspirations resulted in a 65% reduction in pin frequency, without substantially altering dorsal contacts. Anesthetization of the anterior surface of the animal's back with xylocaine reduced pinning in controls but eliminated pinning in parietals. Although the results generally indicate little participation of the neocortex in the instigation of rough-and-tumble play, the reliable numerical changes that were observed may be explained by apparent motor changes as well as reduced somatosensory sensitivity.