Alfvénic velocity spikes and rotational flows in the near-Sun solar wind.

The prediction of a supersonic solar wind1 was first confirmed by spacecraft near Earth2,3 and later by spacecraft at heliocentric distances as small as 62 solar radii4. These missions showed that plasma accelerates as it emerges from the corona, aided by unidentified processes that transport energy outwards from the Sun before depositing it in the wind. Alfvénic fluctuations are a promising candidate for such a process because they are seen in the corona and solar wind and contain considerable energy5-7. Magnetic tension forces the corona to co-rotate with the Sun, but any residual rotation far from the Sun reported until now has been much smaller than the amplitude of waves and deflections from interacting wind streams8. Here we report observations of solar-wind plasma at heliocentric distances of about 35 solar radii9-11, well within the distance at which stream interactions become important. We find that Alfvén waves organize into structured velocity spikes with duration of up to minutes, which are associated with propagating S-like bends in the magnetic-field lines. We detect an increasing rotational component to the flow velocity of the solar wind around the Sun, peaking at 35 to 50 kilometres per second-considerably above the amplitude of the waves. These flows exceed classical velocity predictions of a few kilometres per second, challenging models of circulation in the corona and calling into question our understanding of how stars lose angular momentum and spin down as they age12-14.

Cooling rate effect on outgrowth of Clostridium perfringens in cooked, ready-to-eat turkey breast roasts.

The potential for Clostridium perfringens spores to germinate and grow in cooked, ready-to-eat turkey products was evaluated to determine a safe cooling rate within the critical temperatures of 48.9 C (120 F) through 12.8 C (55 F). Raw turkey deli breast roasts were inoculated with a cocktail of C. perfringens spores (NCTC 8238, NCTC 8239, and NCTC 10388) and cooked in a steam oven to an internal temperature of 72 C. The sample roasts were then cooled through the critical cooling range at rates yielding cooling times of 6, 8, and 10 h. Turkey roasts were analyzed for spore growth and multiplication using tryptose-sulfite-cycloserine agar and anaerobic incubation at 37 C for 48 h. Cooling times of 6 and 8 h showed no proliferation of C. perfringens that would violate the USDA/Food Safety Inspection Service safe cooling standard criteria, which would allow no more than a 1 log10 multiplication between 48.9 and 12.8 C. A 9.6-h cooling period between the designated temperatures at a 95% confidence interval was determined to be adequate for nonproliferation of C. perfringens. On the other hand, a 95% tolerance interval would be more stringent in that it suggests no more than an 8.9-h cooling period. Tolerance intervals required that 95% of all our observations did not exceed the limit of 1 log10 increase in C. perfringens. This study indicated that in cooked, ready-to-eat turkey deli breasts, a cooling period between 48.9 C (120 F) and 12.8 C (55 F) of no greater than 8.9 h should be utilized to prevent possible C. perfringens foodborne outbreaks.

Microvasculature of preovulatory follicles: comparison of in situ and in vitro perfused rabbit ovaries following stimulation of ovulation.

Perifollicular vasculature undergoes significant morphologic changes as ovulation approaches. These vascular changes were observed in in vitro perfused and in situ rabbit ovaries by means of scanning electron microscopy of microcorrosion casts. Casts were made in in situ unstimulated ovaries, in situ ovaries stimulated with human chorionic gonadotropin, in vitro perfused unstimulated ovaries, and in vitro perfused ovaries after an ovulation-inducing dose of human chorionic gonadotropin, prostaglandin F2 alpha, histamine, or norepinephrine. Dilated vessels, extravasation of resin from weakened vessels, and filling defects at the apex of the follicle were observed in in situ ovaries 9 to 12 hours after stimulation and in in vitro perfused ovaries 4 to 6 hours after human chorionic gonadotropin. In vitro perfused ovaries stimulated with prostaglandin F2 alpha or histamine demonstrated dilated capillaries with extravasation of the resin and filling defects at the apex of large follicles. Norepinephrine-stimulated ovaries showed incomplete filling of vessels, although some large follicles showed extravasation of resin. The occurrence of dilated vessels, extravasation of resin, and filling defects is indicative of preovulatory vascular changes in both in situ and in vitro perfused ovaries, regardless of the ovulatory stimulus.

Examination of the role of calcium in ovulation in the in vitro perfused rabbit ovary with use of ethyleneglycol-bis(beta-aminoethyl ether)-n,n'-tetraacetic acid and verapamil.

The in vitro perfused rabbit ovary preparation was used to examine the role of calcium in the ovulatory process. Two groups of rabbits were studied. In the first group, verapamil hydrochloride (10(-4) mol/L), a calcium channel blocker, was used together with human chorionic gonadotropin (50 IU) in the perfusate. Verapamil had no apparent effect on human chorionic gonadotropin-induced ovulation. Verapamil treatment, however, significantly reduced the percentage of ovulated ova that were mature (68.8%) in comparison to ovulated ova from human chorionic gonadotropin-treated control ovaries (95.0%). In a second experimental group, ethyleneglycol-bis(beta-aminoethyl ether)-n,n'-tetraacetic acid (2.0 mmol/L), a calcium ion chelator, was included in the perfusate with gonadotropin. The ethyleneglycol-bis(beta-aminoethyl ether)-n,n'-tetraacetic acid significantly reduced ovulatory efficiency (16.7% +/- 9.43%) in comparison to that of controls exposed to human chorionic gonadotropin alone (79.5% +/- 11.1%). In addition, ovulation occurred at an earlier time in ovaries perfused with ethyleneglycol-bis(beta-aminoethyl ether)-n,n'-tetraacetic acid; however, only four ovulations occurred in these ovaries. These four ovulated ova were immature, probably reflecting the early time of ovulation. Furthermore, both verapamil and ethyleneglycol-bis(beta-aminoethyl ether)-n,n'-tetraacetic acid blocked ovarian smooth muscle contractions during ovarian perfusion. These data provide additional support for the concept that calcium dynamics influence the processes of ovulation and ovum maturation. Furthermore ovarian smooth muscle contractions do not appear to be essential for ovulation in this model.

The relationship between prostaglandins and histamine in the ovulatory process as determined with the in vitro perfused rabbit ovary.

The process of follicle rupture has been described as an inflammatory reaction in which prostaglandins (PGs) and/or histamine may be involved. With an in vitro perfused rabbit ovary preparation, experiments were carried out for determination of whether a relationship exists among PGs, histamine, and ovulation. PGF2 alpha alone was capable of inducing ovulation when added to the perfusion fluid at 1, 10, and 100 ng/ ml. Effectiveness in achieving ovulation varied directly with the dosage; however, the ovulatory efficiency of PGF2 alpha-treated ovaries was lower than that of ovaries exposed to human chorionic gonadotropin (hCG, 100 IU). PGF2 alpha-induced ovulation could not be blocked by the H2 receptor antagonist, cimetidine. The PG synthesis inhibitor, indomethacin, did not prevent histamine-induced ovulation. Ovulation induced by hCG was partially blocked by the administration of indomethacin; however, the concomitant administration of cimetidine was not associated with further reduction in ovulation. In all but one experimental group, the majority of ovulated ova did not progress beyond the intact germinal vesicle stage unless the ovaries had been exposed to hCG. On the basis of these experiments, PGs and histamine do not appear to be interdependent in their effects on the ovulatory process in vitro.

Influence of calcium and magnesium deprivation on ovulation and ovum maturation in the perfused rabbit ovary.

The role of calcium (Ca++) and magnesium (Mg++) in the ovulation process was studied using in vitro perfused rabbit ovaries. Ovaries were perfused with or without human chorionic gonadotropin (hCG) in Ca++/Mg++-free medium (M199) alone or combined with standard M199 to yield varying concentrations of Ca++ and/or Mg++. In all ovaries perfused with hCG, ovulatory efficiency was similar regardless of the concentration of Ca++ and/or Mg++. In ovaries perfused in Ca++/Mg++-free medium without hCG, ovulatory efficiency was similar to that in ovaries perfused with hCG. As Ca++/Mg++ levels were increased without hCG, ovulatory efficiency declined. Ovulation time was significantly accelerated in ovaries perfused in Ca++/Mg++-free medium with or without hCG. Most ovulated ova from ovaries perfused without hCG were immature. With hCG, degree of ovum maturity was directly related to ovulation time. Ovarian smooth muscle contractions were undetectable in 3 ovaries perfused in Ca++/Mg++-free M199 despite occurrence of ovulation. Smooth muscle contractions were recorded in 2 of 3 ovaries perfused in standard M199 with hCG. These results indicate: 1) Ca++/Mg++ exclusion results in rapid follicle rupture and immature ova; 2) oocyte maturation appears to be gonadotropin-dependent; 3) ovulation occurs in the absence of ovarian smooth muscle contractions during perfusion with Ca++/Mg++-free medium.

Ultrastructure of ovarian follicles in in vitro perfused rabbit ovaries: response to human chorionic gonadotropin and comparison with in vivo observations.

Ovulation may be achieved and studied in an isolated perfused rabbit ovary upon inclusion of human chorionic gonadotropin (hCG) in the perfusion fluid. The ultrastructural features of the rabbit ovarian follicle prior to ovulation in vitro were compared with those in vivo. The perifollicular vasculature was also examined in in vitro perfused rabbit ovaries during the preovulatory interval. Granulosa cells of the preovulatory follicle share many ultrastructural features in vivo and in vitro; however, only small amounts of smooth endoplasmic reticulum (sER) were observed in granulosa cells in vitro after hCG. Ovulation after hCG in the in vitro preparation tends to occur earlier (6 hours) than in vivo (12 hours). Thus, there may be insufficient time and/or gonadotropin exposure to permit full functional development of granulosa cells, as reflected by reduced amounts of sER. Degradation of collagen fibrils was less prominent in the theca externa and tunica albuginea in vitro than in in vivo. Perifollicular capillaries became dilated after hCG, but interendothelial gaps were not observed. Disappearance of surface epithelium in the apex of follicles was similar in vitro and in vivo.

Prostaglandin F2a, an ovulatory intermediate in the in vitro perfused rabbit ovary model.

PGF2a has been proposed as a mediator of mammalian ovulation. To elucidate further the role of PGF2a in the process of ovulation, PGF and PGF2a metabolite were measured by radioimmunoassay in the perfusate of an in vitro perfused rabbit ovary preparation. Perfusion medium samples were collected over a 10 to 12 hour period from ovaries perfused with tissue culture M199 (total volume 150 ml, sample volume 3 ml) to which varying amounts of hCG had been added. [The PGF2a antisera a 40% cross reaction with PGF1a, hence total PGF was measured with this antisera.] Both PGF and PGF2a metabolite showed a linear increase with time and numbers of ovulations. PGF media accumulation was 575 pg/ovary/ovulation/hr and PGF2a metabolite accumulation was 367 pg/ovary/ovulation/hr. Medium prostaglandin content could be correlated with numbers of ovulations, ovulatory efficiency (number of ovulations/total follicles) but not total follicles. These data best fit a model of independent ovulatory units producing PGF2a without recruitment or interaction between them. We infer that PGF and PGF2a metabolites in this system can be used as a direct index of the ovulation process.

In-vitro fertilization of rabbit ova ovulated in vitro during ovarian perfusion.

During in-vitro perfusion of rabbit ovaries, ova that ovulated in response to hCG were recovered and cultured with spermatozoa capacitated in vivo. After culture for 30 h, 113 (48.8%) of the ova underwent cleavage and were not degenerate. Of these 113 zygotes, 75 were successfully transferred to the oviducts of 14 gonadotrophin-treated host rabbits. On Day 14 after transfer there were 14 implantation sites and 9 pregnancies proceeded to term. These data indicate that in-vitro ovulated ova can be fertilized in vitro and be used to establish normal pregnancies in a host rabbit.

Effect of histamine and histamine blockers on the ovulatory process in the vitro perfused rabbit ovary.

An increase in the content of histamine in the ovary following luteinizing hormone (LH) release and the inhibition of ovulation in the rabbit by antihistamines suggest that histamine may be involved in the ovulatory process. The effects of various doses of histamine and antihistamines on ovulation were investigated using the in vitro perfused rabbit ovary system. Histamine (100 ng/ml) added to the perfusate at hourly intervals induced ovulation, although at a rate below that observed following human chorionic gonadotropin (hCG) administration. Cimetidine (10 micrograms/ml), an H2 blocker, inhibited histamine-induced ovulation, while the H1 blocker, chlorpheniramine (66.7 micrograms/ml), failed to do so. Neither cimetidine nor chlorpheniramine was able to block ovulation following hCG (50 IU). In all experimental groups in which histamine was used to induce ovulation, both extruded ova and follicular oocytes remained in an immature stage and displayed little evidence of degeneration. In contrast, a high percentage of ova exposed to hCG were mature. Ovarian edema was increased in ovaries in which ovulation occurred, regardless of treatment. A linear correlation was noted between ovulatory efficiency and degree of ovarian edema. Histamine may be an intermediary in the mechanism of follicular rupture, but does not support ovum maturation. However, the inability of H1 and H2 antagonists to block hCG-induced ovulation raises questions regarding the role of histamine in the physiologic process of ovulation.

The effect of prostaglandin synthesis inhibition by indomethacin on ovulation and ovum maturation in the in vitro perfused rabbit ovary.

Prostaglandins (PGs) have been implicated in the mechanism of ovulation in several species through the use of PG synthesis inhibitors such as indomethacin. Studies of ovulation in the in vitro perfused rabbit ovary have aided in the delineation of the process of ovulation. This study was designed to determine the effects of indomethacin on follicle rupture and ovum development in the in vitro perfused rabbit ovary preparation. Indomethacin treatment (0.5 microgram/ml) significantly reduced the occurrence of ovulation in gonadotropin-treated ovaries. The percentage of in vitro ovulated ova and ova recovered from unruptured follicles during ovarian perfusion which achieved Metaphase II did not significantly differ between indomethacin-treated and control ovaries (hCG alone). However, increased degeneration of Metaphase II ova was associated with indomethacin treatment. In in vitro culture experiments this degeneration could be prevented by supplemental PGF2 alpha. These results indicate that inhibition of PG synthesis by indomethacin prevents follicle rupture but does not affect ovum maturation, thus providing further evidence that these two processes are distinct phenomena in vitro.

Fertilizability of ova ovulated and recovered from rabbit ovaries perfused in vitro.

Ovaries removed from New Zealand White rabbits were perfused and exposed to gonadotropin in vitro. The ova ovulated in vitro (N = 56) were recovered and cultured and then transferred to the oviducts of six previously mated Dutch Belted hosts. Twelve of the resulting 36 offspring (33.3 percent) were white. In control matings between 12 Dutch Belted females (six randomly selected and the six hosts) and New Zealand White males, only one of 80 (1.2 percent) offspring was white. These data indicate that ova ovulated in vitro can be transferred to the oviduct of a host rabbit where they may be fertilized and after implantation may develop into viable embryos.

The effect of dexamethasone and promethazine administration on adhesion formation, tubal function, and ultrastructure following microsurgical anastomosis of rabbit oviducts.

In a controlled study, 24 rabbits underwent bilateral division and immediate microsurgical anastomosis of the oviducts. Dexamethasone and promethazine were administered to 13 rabbits in a dose and route of administration similar to those used in clinical practice. Eleven control rabbits received a saline vehicle. Morbidity and mortality were encountered only in the dexamethasone and promethazine-treated group. Dexamethasone and promethazine did not appear to influence the formation of intraluminal and peritubal adhesions, histology, ultrastructure, patency, implantation, or pregnancy rates. The presence of intraluminal adhesions, as seen by scanning electron microscopy, was associated with relatively more efficient tubal function as measured by the ability of the oviducts to convey ova to intrauterine implantation sites. Collagen accumulation in the muscularis, as demonstrated by trichrome staining, was associated with relatively decreased tubal function.

Inhibitory effect of prolactin on ovulation in the in vitro perfused rabbit ovary.

Prolactin is best known for its effects on the breast, promoting mammary growth and lactation. In some species, including rat, mouse, hamster, sheep and rabbit, prolactin is necessary for the maintenance of the corpus luteum. Further, a relationship has been recognised between hyperpromus and or adenohypophysis. McNatty et al. have postulated a direct ovarian effect of prolactin, reporting that low prolactin levels were essential for progesterone production by preovulatory human granulosa cells cultured in vitro. However, high levels of prolactin decreased progesterone production. The present investigation sought to determine the effect of prolactin on ovulation using the in vitro perfused rabbit ovary preparation. We report that its effects are inhibitory.

In vitro reversal of indomethacin-blocked ovulation by prostaglandin F2alpha.

An in vitro perfused ovary preparation was used to study the role of prostaglandin F2alpha (PGF2alpha) in follicle rupture. The administration of PGF2alpha alone has been shown to restore indomethacin-blocked ovulation in rabbits and monkeys. In the model used, ovulation consistently occurred when human chorionic gonadotropin (hCG) was given to the intact rabbit prior to ovarian removal. hCG-induced ovulation was blocked in both perfused and in situ control ovaries by indomethacin (10 mg/kg intravenously 6 hours after hCG) given to the intact animal. The addition of PGF2alpha (1 mg/200 ml) to the perfusion fluid restored ovulation in the isolated ovary as compared with the in situ ovary (P less than 0.005) and with the perfused, untreated ovary (P less than 0.01). Following removal and perfusion of both ovaries from rabbits treated with indomethacin, ovulation occurred following the addition of PGF2alpha to the perfusate, but did not occur without PGF2alpha (P less than 0.05). These data indicate that indomethacin can block ovulation and that ovulation can be restored by the addition of PGF2alpha to the perfusion system, further supporting the significance of PGF2alpha in the process of follicular rupture.

Investigation of mammalian ovulation with an in vitro perfused rabbit ovary preparation.

An in vitro perfused rabbit ovary preparation has been developed to study local factors responsible for the ovulatory process. This preparation enables: (1) isolation of the ovary from systemic influences, (2) direct serial observations of follicle development and ovulation, (3) addition to the perfusing medium of substances that may influence ovulation acting at the level of the ovary, and (4) correlation of the times of individual follicle rupture with ovarian contractile patterns. With this model the temporal requirements for human chorionic gonadotropin in the process of ovulation have been studied. The direct effects on ovulation of prostaglandin F3 alpha (PGF 2 alpha), the antihistamine chlorpheniramine, and calcium deprivation were also investigated. Chlorpheniramine and EDTA each led to inhibition of ovulation and depressed ovarian contractility. The use of the isolated in vitro ovary preparation permits characterization of the local requirements for ovulation.