Estrogen inhibition of noradrenaline release in the rabbit oviduct

In the present study, we investigated the influence of estrogen on 3H-noradrenaline (3H-NA) release induced in the oviductal isthmus by electrical stimulation, potassium and calcium. The fractional release of 3H-NA was measured in oviducts isolated from ovariectomized rabbits and from ovariectomized rabbits treated with estradiol cypionate, 70 micrograms/kg im 72 h before an experiment. Electrical field stimulation of the intramural nerves induced muscle contraction and augmented the release of labelled NA from the muscle. The 3H-NA release was reduced after estrogen treatment when reuptake of NA into the nerve terminals was blocked by desipramine, 10(-6) M. Estrogen also reduced the 3H-NA release evoked by exposure of the oviducts to 121 mM KC1 in the presence of calcium (2.5 mM) and in a high potassium, calcium-free medium upon the addition of 2.5 mM calcium. In the presence of desipramine a small fraction of 3H-NA was released in high potassium, calcium-free medium. This release was unaffected by estrogen. These results suggest that estrogen reduces the release of NA from the adrenergic nerves within the oviduct and that this action is exerted primarily on the calcium-dependent release. It therefore might be due to a reduction in the entry of calcium into the nerve terminal.     

Catecholamines within the rabbit oviduct at fertilization time

Dopamine, norepinephrine and epinephrine levels of rabbit oviductalfluids were measured in order to establish whether the ampullarysegment contained higher concentrations correlating with thesperm hyperactivity observed in the ampulla of different mammals.In this study, we found no statistically significant differencesbetween flushings from the ampulla or the isthmus at oestrus,ovulation or 72 h postcoitum (p.c). However, after correctingfor the volume secreted, the concentrations of catecholaminesare always lower in the ampulla than in the isthmus. In allcases the levels of biogenk amines were highest at oestrus thenfell at ovulation. Individual variation of these levels at thetime of ovulation was observed only in the ampulla, probablydue to some contribution from follkular fluid. During the earlyluteal phase (72 h p.c.) the catecholamine levels again increaseslightly, especially in the isthmus. We conclude that whiplashmotility cannot be explained by a higher level of catecholaminesin the ampulla.     

Periovulatory increase in temperature difference within the rabbit oviduct

BACKGROUND: Earlier studies demonstrated a small temperature difference between the sperm storage and fertilization sites within the oviducts of rabbits and pigs. Our aim was to reveal the time dependence of this temperature difference relative to ovulation, and to determine how this difference is generated-by temperature elevation at one of these sites or by temperature decrease at the other site. METHODS: The temperature at the sperm storage site (at the isthmus near the uterotubal junction) and at the fertilization site (the isthmic-ampullary junction) of rabbit oviducts were measured before, during, and after ovulation by two probes, connected to digital thermometers. Rectal temperature was constantly measured and served as a control for body temperature. RESULTS: The temperature difference between the fertilization site and the storage site was 0.8+/-0.2 degrees C before ovulation. This difference increased at ovulation, reaching 1.6+/-0.1 degrees C after ovulation (P<0.03). This increased difference was mainly due to temperature decrease in the sperm storage site. CONCLUSION: The temperature-difference increase within the rabbit oviduct is generated at ovulation by a reduced temperature at the sperm storage site. This temperature gradient may play a role in mammalian reproduction via sperm thermotaxis.     

Degeneration release of noradrenaline in skin flaps in rats

To study if noradrenaline released from degenerating sympathetic nerves may contribute to metabolic stimulation and low blood flow in an experimental skin flap, we have determined noradrenaline levels in such skin flaps. Attempts to use radioenzymatic methodology for the determination of noradrenaline in rat skin extracts were unsuccessful due to interference with the assay. High performance liquid chromatography with electrochemical detection was, however, found to give accurate estimates of rat skin noradrenaline contents. Rat skin contained approximately 2.7 nmol (455 ng) noradrenaline per gram dry weight. Extensive depletion of skin flap noradrenaline, occurring mainly between 6 and 24 h postoperatively, was found. At 48 h nearly all sympathetic nerves in the skin flaps had degenerated as evidenced by noradrenaline levels of 0.13 nmol/g. The degeneration of sympathetic nerve endings appears to proceed from the sides towards the middle of the flap, indicating a segmental distribution of the nerves. Noradrenaline released from degenerating sympathetic nerves may adversely affect the survival of critical skin flaps by causing vasoconstriction and metabolic stimulation.     

Haptoglobin expression and release by rabbit oviduct and endometrium, its localization in blastocyst extra-embryonic matrix and fluid during preimplantation time

BACKGROUND: Evidence is emerging that haptoglobin, an acutephase protein with immunomodulatory properties, is expressedby the endometrium of various species. The present study describesan in-depth investigation of haptoglobin expression and releasein the rabbit reproductive tract and in preimplantation embryos.METHODS: The full-length cDNA sequence of rabbit haptoglobinwas determined by rapid amplification of cDNA ends PCR. Haptoglobinexpression was studied in the oviductal ampull, and isthmus,endometrium and embryos from the time of ovulation up to adhesion.These results were completed by western blot analysis of reproductivetract secretions and embryonic tissues. RESULTS: cDNA sequencingshowed a high homology between rabbit and human haptoglobin(84.1%). In oviductal tissues haptoglobin mRNA is clearly expressedfrom 6 h post-conception (p.c.) to day 3, and in the uteruson days 5 and 6. In the oviductal fluid highest haptoglobinprotein content was found between 6 h p.c and day 2, and inthe uterine fluid on days 5 and 6 p.c. Embryos do not expresshaptoglobin mRNA during preimplantation development. However,considerable amounts of maternal haptoglobin protein were detectedin the blastocyst coverings and in blastocyst fluid. CONCLUSIONS:Already during periovulatory time and oviductal passage, highamounts of haptoglobin are present in the microenvironment surroundingthe oocyte/embryo. Two days before implantation, again, highhaptoglobin levels are detectable in the embryo's environment.The incorporation of haptoglobin into the extra-embryonic matrixmay be of particular functional significance.     

Prostaglandin-mediated inhibition of noradrenaline release

The effect of indomethacin, a prostaglandin (PG) synthesis inhibitor, on some cardiovascular reflexes was studied in healthy subjects. Heart rate (HR) and respiratory sinus arrhythmia (RSA) in the basal state and during carotid stimulation (neck suction), Valsalva ratio, and changes in heart rate and blood pressure (BP) during an orthostatic test were measured before and one hour after administration of indomethacin (1.5 mg/kg). The efficacy of the PG synthesis inhibitor was monitored by analysis of platelet aggregation induced by arachidonic acid. Following indomethacin no change was observed in basal HR. Carotid stimulation depressed the HR and this effect was of the same amplitude before and after indomethacin. The amplitude of RSA was not affected by indomethacin, either in the basal state or during carotid stimulation. The Valsalva ratio and the changes in HR and BP during the orthostatic test were similar before and after the drug. Circulating levels of noradrenaline were unaffected by indomethacin. These data demonstrate that inhibition of PG bioformation in man does not affect major cardiovascular reflexes. Consequently they disfavour the hypothesis that endogenously formed PG would be involved in the normal activity of the afferent, central or efferent pathways for cardiovascular regulation.     

Electrical activity in the human oviduct after the menopause

Electrical activity was recorded in 18 morphologically normal human oviducts excised from 17 women during hysterectomy. All women were in the first five years after menopause. The first part of the oviduct to become inactive after menopause was the isthmus near the uterus or the ampullary-isthmic junction. The region which exhibited electrical activity last was the ovarian end of the tube. The typical wave-form of electrical activity in post-menopausal oviducts resembled that recorded during the follicular phase in fertile women: a single smooth wave lasting 3-5s. In three cases, over a bulbous swelling of the ampulla, this typical activity was replaced in continuous, high frequency activity. A hypothesis is provided that the observed changes in electrical activity may be related to certain clinical phenomena such as the increased incidence of tubal pregnancy in the pre-menopause.     

小鼠植入前胚胎对输卵管上皮细胞DNA甲基转移酶1表达的影响

目的 探讨小鼠植入前胚胎对输卵管上皮细胞DNA甲基转移酶1（DNA methyltransferase 1，Dnmt1）表达的影响。方法 应用免疫组织化学方法确定Dnmt1蛋白在小鼠输卵管的组织学定位；选取植入前胚胎的2细胞期、4细胞期和8细胞期作为3个观察点，采用实时逆转录-聚合酶链反应和Western印迹分别检测正常妊娠和假孕小鼠输卵管上皮Dnmt1 mRNA和蛋白表达水平的变化。结果 Dnmt1蛋白主要表达于上皮层；妊娠组小鼠各期Dnmt1 mRNA表达水平均显著性低于同期假孕组（P〈0．05），妊娠组4细胞期Dnmt1蛋白表达水平显著性低于同期假孕组（P〈0．05）。结论 小鼠植入前期的胚胎对母体输卵管上皮Dnmt1 mRNA和蛋白表达存在调节作用，提示胚胎可能通过DNA甲基化间接调控输卵管上皮某些基因的表达，Dnmt1可能参与胚胎-母体交流的部分机制。     

Mechanism of presynaptic actions of adenosine and acetylcholine on noradrenaline release in the guinea-pig heart

Isolated heart of the guinea-pig was stimulated transmurally (1 Hz), or by excess [K⁺] to evoke release of [³H]noradrenaline. The interactions of tetraethylammonium, acetylcholine and adenosine were investigated on [³H]noradrenaline overflow at different [Ca²⁺] concentrations. One mM tetraethylammonium doubled [³H]noradrenaline overflow, and at 30 mM the overflow was facilitated by about 50-fold. The facilitatory effect of 30 mM tetraethylammonium gradually decreased with reduction in [Ca²⁺] concentration. In 0.1 mM [Ca²⁺] overflow of [³H]noradrenaline was completely blocked. However, addition of 30 mM tetraethylammonium to 0.1 mM [Ca²⁺]-Krebs solution greatly facilitated the overflow. 35 mM K-induced [³H]noradrenaline overflow was facilitated by about 5-fold with 20 mM tetraethylammonium, and the facilitatory effect was blocked by 0.37 μM tetrodotoxin; 75 mM K-induced overflow was facilitated by only 1.5-fold by tetraethylammonium.Electrically induced overflow of [³H]noradrenaline was significantly reduced (30%) by 0.13 μM acetylcholine, and the effect increased up to about 75% with higher concentrations of acetylcholine (1.3 μM). 1.3 μM acetylcholine reduced 35 mM K-induced overflow by 30% and had no effect on 75 mM K-induced overflow. 0.37 μM tetrodotoxin had an almost similar effect on K-induced overflow. Ten-times higher concentrations of acetylcholine were needed to obtain over 70% inhibition of overflow produced by low or high K. The inhibitory effect of acetylcholine on [³H]noradrenaline overflow by 1 Hz was antagonized by increasing concentrations of tetraethylammonium (3 to 30 mM) in 2.5 mM Ca. The antagonism between acetylcholine and tetraethylammonium persisted even when Ca was lowered to 0.02 mM. However, if the overflow was enhanced by other means (high [Ca²⁺] plus phentolamine) to the same extent as with tetraethylammonium and low [Ca²⁺], then acetylcholine blocked overflow by 75%. 3.7 μM adenosine reduced [³H]noradrenaline overflow by about 54%, and this effect was completely prevented by 30 mM tetraethylammonium, either in 2.5 or 0.02 mM Ca.It is proposed that acetylcholine and adenosine interfere with nerve stimulation-evoked release of [³H]noradrenaline, presumably by altering electrical properties (i.e. conduction, resting membrane potential, duration of nerve action potential, etc.) of cardiac sympathetic nerve terminals. This primary action would subsequently lead to a reduction in the availability of [Ca²⁺] for the release process.     

Influence of impulse pattern on noradrenaline release from sympathetic nerves in cerebral and some peripheral vessels

Hardebo , J. E. 1992. Influence of impulse pattern on noradrenaline release from sympathetic nerves in cerebral and some peripheral vessels. Acta Physiol Scand 144 , 333–339. Received 18 June, accepted 7 November 1991. ISSN 0001–6772. Departments of Medical Cell research and Neurology, University of Lund, Lund, Sweden. Earlier findings suggested that the physiological firing rate in sympathetic nerves did not exceed 10 Hz. Later recordings have revealed that this is a net value: impulses are usually not continuous but occur in bursts of high frequencies separated by quiet periods. The effects of continuous and burst-like neurogenic activation in various isolated blood vessels were compared. In the first part of the study changes in vascular tone were registered. Electrical field stimulatory parameters were chosen to give tetrodotoxin-blockable, neurogenic responses. From dose-response experiments in rat caudal artery a net frequency of 6 Hz was chosen, for bursts usually designed as 30 Hz during 0.2 s followed by a quiet period of 0.8 s. In the rabbit ear artery the neurogenic contraction was enhanced by a mean of near 50% during stimulation with bursts. Now appeared a minor phentolamine-resistant portion, which was nor due to release of NPY, 5–HT, histamine or ATP. Also in the rat caudal artery and monkey pial artery a significant enhancement of contraction was seen during burst stimulation, whereas in the rabbit facial vein no significant difference in dilatation through β-receptor activation was obtained. In vessels that do not normally respond with purely neurogenic contractions/dilatations during continuous stimulation, like pial arteries from the rat and rabbit, not even bursts revealed a neurogenic response. In a second series of experiments the influence of continuous and burst-like nerve activation on the release of [¹⁴C]noradrenaline was studied in monkey, rabbit and rat pial arteries, rat caudal artery and rabbit central ear artery and facial vein. In all vessels except rabbit pial artery a significantly higher release was obtained with burst-like compared to continuous stimulation.     

Uptake, release, and modulation of release of noradrenaline in rabbit superior colliculus

The noradrenaline content, the uptake of [3H]noradrenaline, and the release of previously incorporated [3H]noradrenaline were studied in slices of rabbit superior colliculus. The concentration of endogenous noradrenaline was higher in superficial than in deep layers of the superior colliculus. Upon incubation with [3H]noradrenaline, tritium was accumulated by a mechanism that was strongly inhibited by oxaprotiline but little inhibited by 6-nitroquipazine. Electrical stimulation at 0.2 or 3 Hz increased the outflow of tritium from slices preincubated with [3H]noradrenaline; the increase was almost abolished by tetrodotoxin or a low calcium medium. Clonidine reduced the evoked overflow of tritium, whereas yohimbine increased it and antagonized clonidine. The evoked overflow was also reduced by the dopamine D2-receptor-selective agonists apomorphine and quinpirole, an effect antagonized by sulpiride. The preferential opioid kappa-receptor agonist ethylketocyclazocine produced an inhibition that was counteracted by naloxone. Nicotine accelerated the basal outflow of tritium; part of the acceleration was blocked by hexamethonium. The muscarinic agonist oxotremorine slightly diminished the electrically evoked overflow, and its effect was abolished by atropine. The oxaprotiline-sensitive uptake of [3H]noradrenaline as well as the tetrodotoxin-sensitive and calcium-dependent overflow of tritium upon electrical stimulation (presumably reflecting the release of [3H]noradrenaline) indicate that noradrenaline is a neurotransmitter in the superior colliculus. The release of [3H]noradrenaline is modulated through alpha 2-adrenoceptors as well as dopamine D2-receptors, opioid kappa-receptors and nicotine and muscarine receptors. No clear evidence was found for modulation through beta-adrenoceptors, D1-receptors, serotonin receptors, opioid mu- or delta-receptors or receptors for GABA or glutamate. Only the alpha 2-adrenoceptors receive an endogenous agonist input, at least under the conditions of these experiments. The pattern of presynaptic modulation resembles that found for noradrenaline release in other rabbit brain regions, suggesting that all noradrenergic axons arising in the locus coeruleus possess similar presynaptic receptor systems.     

Culture of epithelial cells derived from the oviduct of different species

This study proposes a procedure for the isolation and cultureof oviduct epithelial cells of several species. In-vitro cultureon such a feeder seems to allow full embryonic development andviability. The inner linings of Fallopian tubes from mouse,rabbit, cow and human were trypsinized and the epithelial cellswere enriched with Percoll gradient. Isolated cells, obtainedin high yield with good viability, were maintained in monolayerculture in B2-Menezo medium supplemented with serum, which alsosupports early embryonic development in vitro. The plated primarycultures reached confluence within 8 days, producting a monolayerof cohesive polygonalcells. Associated with this large epithelialcall population, ciliated cells as wellas polykaryotic cellsand few fibroblastic nestswere observed. After the first sub-culture,the ciliated cells disappeared and the epithelial cell monolayergrew rapidly to confluence with in 3 days and displayed contactinhibition. No epithelial cell growth could be obtained inculturein the absence of serum. The addition of oestrogens had no effecton any of the cultured oviductal epithelial cells. A sponotaneousalteration was observed in morphology and growth after severalpassages, the number of which depends mainly upon the species