Inter-population genetic variation in the freshwater leech Nephelopsis obscura in relation to environmental heterogeneity

The temporal distribution of populations of aquatic macroinvertebrates have been extensively investigated in temperate arctic regions, but little information is available for alpine regions in North America. This paper describes phenology patterns of the aquatic macroinvertebrates of a high elevation wetland (3593 m) in the Green Lakes Valley, Colorado Front Range.Abundant taxa were Chironomidae (primarily Orthocladius and the limnephilid trichopteran Asynarchus curtus (Banks). Temperature patterns were related to the seasonal timing of life stages of most species. Phenology of most alpine aquatic macroinvertebrates appears to be controlled by seasonal climatic patterns, particularly temperature patterns. All chironomid species and Asynarchus curtus (Banks) had univoltine life cycles. Multi-year life cycles were not observed at this alpine site, although they have been observed at some arctic sites with colder temperature regimes than observed in this study.     

Relative vulnerability of six freshwater gastropods to the leech Nephelopsis obscura (Verrill)

SUMMARY. 1. The vulnerability of six species of freshwater snails to the leech Nephelopsis obscura was determined in laboratory predation experiments.
2. Nephelopsis was unable to prey on prosobranch snails with an operculum, but did consume certain pulmonate species though predation rates were low, ranging from one to two snails per leech per night at 20°C. Apparently, Nephelopsis does not actively select prey, but merely consumes those species most easily handled.
3. Among pulmonates, leeches captured species in the following order: Physa gyrina (Say), Helisoma anceps (Menke). Lymnaea emarginata (Say) and Helisoma trivolvis (Say). Susceptibihty of size classes varied among snail species.
4. Neither environmental structure, such as cobble or macrophytes, nor gastropod escape behaviors, such as clamping to substrates or vertical migration away from bottom-feeding leeches, lowered capture rates by Nephelopsis.
5. In 2 years sampling, only one of seven lakes, however, had appreciable abundances of Nephelopsis. It is therefore suggested that the rarity, low feeding rates, and lack of strong gastropod preference limit the effect of Nephelopsis on field gastropod distributions.     

Cosexuality in the Leech,Nephelopsis obscura (Erpobdellidae)

Summary

Hermaphroditism of the erpobdellid leach Nephelopsis obscura is described on the basis of macroscopic and histological studies of the gonads. Perivitellogenic cells (female primordial germ cells) proliferate from the septa of segments XII and XIII in individuals of 26–30 mg and spermatogenic cells from segments XII to XIII of 20–25 mg specimens. The majority of hatch- lings develop functional testisacs when they reach 170 mg and functional ovisacs when they reach 250 mg. Oocytes proliferate only after the formation of stage 5–6 spermatozoa. Thus, N. obscura is a sequential cosexual hermaphrodite with spermatogenesis and mature spermatozoa development commencing before oogenesis. During the second cycle of gametogenesis observed in larger animals, both mature spermatozoa and ova co-occur and N. obscura exhibits simultaneous hermaphroditism.     

The effect of a predatory leech,Nephelopsis obscura,on mortality,growth, and production of chironomid larvae in a small pond

Summary The effect of a predatory leech, Nephelopsis obscura, on survivorship, growth, and production of chronomid larvae was studied by enclosure experiments carried out in a small pond. The prey population was composed almost entirely of the tubiculous, microphagous chironomid larvae, Chironomus riparius and Glyptotendipes paripes. Nephelopsis significantly reduced chironomid survivorship within the enclosures, and accounted for most of the measured mortality of fourth instar larvae. The cropping by Nephelopsis was not significantly biased toward either prey species. In long-term experiments (66 d) chironomid biomass in enclosures without leeches reached much higher levels than in enclosures containing Nephelopsis. This increase in biomass was due to growth of surviving larvae, rather than recruitment, since emergence and oviposition were not going on during the course of the experiments. The enhanced survivorship of larvae within leech-free enclosures was eventually accompanied by reduced growth and specific production (daily production/biomass) for C. riparius, which made up about 90% of the larval population. Growth and specific production of G. paripes (10% of larval population) was not affected. Short-term experiments (25 d) involving manipulation of densities and species ratio (9:1 CR:GP and 1:9 CR:GP) of larvae revealed that growth of the majority species was strongly influenced by larval density, whereas growth of the minority species was not. The same pattern was observed both in the presence and in the absence of Nephelopsis and was a result of differences in resource utilization between the two species. In the shortterm experiments, growth rates estimated for larvae exposed to leeches were significantly less than those for larvae in leech-free enclosures. This could be due either to size-biased consumption of larvae by Nephelopsis, or possibly a disturbance factor leading to reduced larval food intake and/or increased metabolic costs.     

Effects of aggregative behaviour on the bioenergetics of the freshwater predatory leech Nephelopsis obscura (Erpobdellidae)

1. Supplied with ad libitum prey, the effects of different group sizes (one, five, ten, twenty-five, fifty or 100) on the acquisition and allocation of energy were examined over the entire life cycle of the freshwater predatory leech Nephelopsis obscura , to evaluate the bioenergetic costs and/or benefits of aggregation.
2. In terms of growth, asymptotic biomass and ingestion, group sizes larger than one and less than ten are optimal. As group size increases from ten to 100, N. obscura has slower growth and lower weekly food ingestion, suggesting interference competition. Single leeches had lower food consumption than group sizes of less than 100 animals. With a lower asymptotic biomass, single leeches have lower fecundity and presumed lower fitness. Lipid deposition was unaffected by group size.
3. Size- and time-specific respiration rates declined with increasing group size. For group sizes between five and fifty, the same relative proportion of ingested energy was allocated to respiration. Above this threshold group size, leeches allocated a higher proportion of ingested energy to growth, i.e. had higher growth efficiency, as did leeches cultured individually.
4. In the largest group size, total time spent in activity was highest (reflecting the high degree of interference between conspecifics), but as a result of the lower hourly costs of respiration they allocated a lower proportion of ingested energy to total respiration.
5. Optimum group size of N. obscura in the laboratory is within the typical range of group sizes found on stony shores in the field.     

Effects of aggregative behaviour on the bioenergetics of the freshwater predatory leech Nephelopsis obscura (Erpobdellidae)

1. Supplied with ad libitum prey, the effects of different group sizes (one, five, ten, twenty-five, fifty or 100) on the acquisition and allocation of energy were examined over the entire life cycle of the freshwater predatory leech Nephelopsis obscura , to evaluate the bioenergetic costs and/or benefits of aggregation.
2. In terms of growth, asymptotic biomass and ingestion, group sizes larger than one and less than ten are optimal. As group size increases from ten to 100, N. obscura has slower growth and lower weekly food ingestion, suggesting interference competition. Single leeches had lower food consumption than group sizes of less than 100 animals. With a lower asymptotic biomass, single leeches have lower fecundity and presumed lower fitness. Lipid deposition was unaffected by group size.
3. Size- and time-specific respiration rates declined with increasing group size. For group sizes between five and fifty, the same relative proportion of ingested energy was allocated to respiration. Above this threshold group size, leeches allocated a higher proportion of ingested energy to growth, i.e. had higher growth efficiency, as did leeches cultured individually.
4. In the largest group size, total time spent in activity was highest (reflecting the high degree of interference between conspecifics), but as a result of the lower hourly costs of respiration they allocated a lower proportion of ingested energy to total respiration.
5. Optimum group size of N. obscura in the laboratory is within the typical range of group sizes found on stony shores in the field.     

Applications and limitations of measurement of 15-keto,13,14-dihydro prostaglandin E2 in human blood by radioimmunoassay

It has been anticipated that the inherent limitations of radioimmunoassays for prostaglandin E (PGE) would be obviated by assays for its major circulating metabolite, 15-keto, 13,14-dihydro PGE₂ (KH₂-PGE₂) which has a longer half-life in blood. We examined the effects of PGE₂ infusion and alterations in lipolysis , and of clotting, prolonged storage and hemolysis , on KH₂-PGE₂ immunoreactivity in unextracted human plasma and serum samples. Indeed KH₂-PGE₂ levels rose several hundred fold during infusions of PGE₂ at doses which cause little or no increment in peripheral PGE levels. During stimulation of lipolysis by infusions of epinephrine, apparent KH₂-PGE₂ levels rose five-fold. However, the dilution curve of plasma obtained during stimulation of lipolysis was not parallel to the standard curve; furthermore, apparent KH₂-PGE₂ levels were correlated strongly with free fatty acid (FFA) levels, suggesting that FFA's cross-reacted in the RIA weakly but significantly due to their very high molar concentration in blood. Clotting and prolonged storage of samples, but not hemolysis, also caused marked apparent increments in KH₂-PGE₂ levels. Competition curves using dilutions of such samples were again not parallel to the standard curves in plasma or buffer, but resembled dilution curves of samples containing high levels of FFA. These results suggest that handling of human blood samples for KH₂-PGE₂ measurement must be carefully standardized to avoid significant artifacts which presumably are due in part to fatty acids released from triglyceride stores or from disrupted membrane phospholipids . Unextracted plasma appears to be unsatisfactory for use in this RIA.     

A synthetic analog of prostaglandin E(1) prevents the production of reactive oxygen species in the intestinal mucosa of methotrexate-treated rats

Administration of methotrexate to rats results in severe enterocolitis and death. Previous our studies showed that a synthetic analog of prostaglandin E(1), OP-1206 [17S, 20-dimethyl-trans-Delta(2)-prostaglandin E(1)] ameliorated the anticancer agent-induced enterocolitis of rats. In the current study, we have focused on the biochemical effect of OP-1206 on the methotrexate-induced intestinal inflammation implicating reactive oxygen species (ROS). Methotrexate (15 mg/kg body weight) was orally administered to rats once daily for 5 days. OP-1206 (0.5 microg/kg body weight) was orally administered to rats twice a day for 5 days. On the 6th day, the chemiluminescence from the jejunum was measured to evaluate the generation of ROS. Spontaneous chemiluminescence from the jejunum of the methotrexate-treated rats increased significantly, compared with the control. Luminol-enhanced chemiluminescence from inflamed mucosal scrapings from the jejunum of the methotrexate-treated rats indicated more remarkable enhancement than the control rats. The treatment of OP-1206 with methotrexate showed significantly lower chemiluminescence of both the jejunum and mucosal scrapings than those of the methotrexate-treated rats. The alkaline phosphatase (ALP) activity, as a marker of small intestinal differentiation, in the intestinal mucosa of the methotrexate-treated rats decreased remarkably, but that of the methotrexate and OP-1206-treated rats was significantly higher than that of the methotrexate-treated rats. Thus, OP-1206 may possibly help the anticancer chemotherapy by protecting the small intestine from the methotrexate-induced damage.     

The effects of different oxygen regimes on the feeding and vertical distribution of Nephelopsis obscura (Hirudinoidea)

The feeding rates and vertical distribution of the freshwater leech Nephelopsis obscura, fed on different densities of third instar Chironomus riparius, were determined in the laboratory at three different constant dissolved oxygen regimes (10%, 100%, 300% saturation) at 15 °C. The rate of feeding increased linearly with prey density up to maximal field prey densities, with no satiation response. Analysis of covariance showed that the rate (slope) at which prey were consumed was not significantly different among oxygen regimes but the amount eaten at any given prey density was higher in higher dissolved oxygen saturations. Analysis of the vertical distribution of N. obscura at different oxygen saturations and prey densities showed a higher proportion of N. obscura on or in the mud substrate compared to the water column at 100% and 300%. No correlation was found between prey density and the vertical distribution of N. obscura, but the numbers of prey eaten were correlated with the proportion of N. obscura on the mud substrate. These results indicate that higher percentage dissolved oxygen regimes increase the feeding of N. obscura and are of potential importance in explaining the seasonal depth distribution and microdistribution patterns recorded for N. obscura.     

Effects of anoxia, hyperoxia, and salinity on neurons in the leech Nephelopsis obscura (Erpobdellidae): RNA redistribution by fluorescence histochemistry

The histochemical distribution of cytoplasmic RNA in ganglion cells of the freshwater leech Nephelopsis obscura has been studied using the fluorochrome acridine orange as a marker of nucleic acids. Two series of experiments, employing 50 adult animals, involved changes in oxygen tension in the water and changes in salinity. Normal leech neurons exhibit finely granular orange fluorescence uniformly distributed throughout the cytoplasm, with a perinuclear ring of especially strong fluorescence. After exposure to anoxic (0% O2), hypoxic (20% O2), or hyperoxic (200% O2) conditions at 20 degrees C for 1-15 days, the orange cytoplasmic fluorescence is no longer uniformly distributed; the redistribution is generally toward the periphery, leaving the perinuclear zone without RNA fluorescence, but irregular zones of cytoplasm devoid of RNA also occur not as a gradient. Leeches exposed to salinity of, or greater than, 2.5 ppt for 15 days exhibit similar changes. These alterations are confirmed by electron microscopy. Seasonal fluctuations in oxygen tension and salinity of lake water affect the distribution and abundance of organisms. The acridine orange method provides one measure of stress to the nervous system in freshwater invertebrates that might be applicable to ecological studies as well as to metabolic studies of individual animals.     

Effects of hypothalamic microinjection of PGE2 on body temperature and sympathetic nervous activities in the rabbit

The febrile response and sympathetic nervous response to hypothalamic microinjections of prostaglandin E₂ (PGE₂) were investigated in anesthetized rabbits. Microninjection of PGE₂ (500–1000 ng) caused an increase in rectal temperature of more than 0.3°C in 13 of 50 loci in the preoptic and anterior hypothalamic area (PO/AH). At 8 of these 13 loci, PGE₂ elicited response patterns in the sympathetic nervous system, such as an increase in cutaneous sympathetic nervous activity and decrease in renal sympathetic nervous activity. This pattern of sympathetic nervous responses was induced with a simultaneous increase in rectal temperature of more than 0.5°C. The 8 loci were distributed in the preoptic area, especially in the vicinity of the supraoptic nucleus. Electrolytic lesions of this region were made bilaterally, and intracerebroventricular injection of PGE₂ (8 µg/kg) was found to inhibit fever and sympathetic activity. The results demonstrate that the action of PGE₂ is responsible for the response patterns of sympathetic twigs during fever. The preoptic area, especially in the vicinity of the supraoptic nucleus, is most sensitive to PGE₂ for the patternized response of sympathetic neurons and fever.     

Correlation between methotrexate-induced intestinal damage and decrease in polyamine content

A synthetic analog of prostaglandin E(1), OP-1206 [17S, 20-dimethyl-trans-Delta(2)-prostaglandin E(1)] protects the small intestine from the methotrexate (MTX)-induced damage. The purpose of this study is to evaluate the protective effect of OP-1206 on the methotrexate-induced small intestinal damage in rats from the biochemical point of view. MTX (15 mg/kg body weight) was orally administered to rats once daily for 5 days. OP-1206 (0.5 microg/kg body weight) was orally administered to rats twice a day for 5 days, and on the 6th day biochemical components in the jejunal mucosa of the treated rats were determined. The contents of DNA, RNA, proteins and polyamines (spermine and spermidine) in the jejunal mucosa of rats were markedly decreased by the MTX treatment. The coadministration of OP-1206 with MTX prevented such decreases caused by the MTX treatment. The MTX treatment decreased the incorporation of 3H-thymidine into DNA in the jejunal mucosa, while the coadministration of OP-1206 with MTX prevented it. These results indicated that OP-1206 could protect the intestinal mucosa against the biochemical effects of MTX through a trophic action on intestinal villi. Further, it should be noted that polyamines may possibly play an important role of modulation action on intestinal mucosa.     

Expression of prostaglandin E2 receptor subtypes, EP2 and EP4, in the rat hippocampus after cerebral ischemia and ischemic tolerance

We investigated the distribution and time course of expression of two subtypes of prostaglandin E₂ (PGE₂) receptors, EP2 and EP4, in a rat model of cerebral ischemia and ischemic tolerance. Adult male Sprague-Dawley rats were subjected to either lethal global ischemia (10 min) with or without sublethal ischemic preconditioning (3 min), or ischemia only (3 min). A short 3-min cerebral ischemia and a 3-min ischemia followed by a second lethal ischemia enhanced the expression of EP2 and EP4 receptors in CA1 pyramidal neurons of the hippocampus. In tolerance-acquired CA1 neurons, the immunoreactivities of EP2 and EP4 were upregulated after 4 h and 12 h, respectively. The immunoreactivities were most prominent at 3 days and were sustained for at least 14 days, consistent with results of immunoblotting experiments. However, immunoreactivities for these PGE₂ receptors increased in reactive glial cells in the vulnerable CA1 and hilar regions of rats subjected to lethal ischemia without ischemic preconditioning. Most of the EP2 immunoreactivity occurred in microglial cells and some astrocytes, whereas increased immunoreactivity for EP4 was found only in astrocytes. These data suggest that ischemia and the induction of ischemia tolerance have different regulatory effects on the expression of EP2 and EP4 receptors. Moreover, PGE₂ may exert its unique pathophysiological functions in relation to delayed neuronal death and ischemic tolerance induction in the rat hippocampus via specific PGE₂ receptors.This research was supported by a grant (M103KV010019 04K2201 01930) from the Brain Research Center of the 21st Century Frontier Research Program funded by the Ministry of Science and Technology of the Republic of Korea.     

Effects of prostaglandin E2 on the micronucleus formation in mouse bone marrow cells by various mutagens

The effects of prostaglandin E₂ (PGE)₂, as trigger of erythroid progenitor cells into the cell cycle, were studied on the induction of micronucleu by various mutagens; with mitomicin C (MMC) the optimal protocol was established. dose-response relationship between PGE₂ doses and micronucleus frequency were observed 30 h after injection of MMC to mice administered PGE₂ 24 h previously. Sensitazion by PGE₂ pretreatment was also found for other mutagens, such as vincristine, 5-fluorouracil, benzo[a]pyrene, 1,1-dimethylhydrazine and 2-naphthylamine. These results support the hypothesis that accelerating the erythropoiesis increases the frequency of micronucleic induced by mutagens.     

Angiotensin II and III suppress food intake via angiotensin AT(2) receptor and prostaglandin EP(4) receptor in mice

Intracerebroventricularly administered angiotensin (Ang) II and III dose-dependently suppressed food intake in mice and their anorexigenic activities were inhibited by AT(2) receptor-selective antagonist. Ang II did not suppress food intake in AT(2) receptor-knockout mice, while it did significantly in wild-type and AT(1) receptor-knockout mice. The suppression of food intake in AT(1) receptor-knockout mice was smaller than that in wild-type. The anorexigenic activities of Ang II and III were also blocked by a selective antagonist for prostaglandin EP(4) receptor. Taken together, centrally administered Ang II and III may decrease food intake through AT(2) receptor with partial involvement of AT(1) receptor, followed by EP(4) receptor activation, which is a novel pathway regulating food intake.     

Prostaglandin E2 receptors in asthma and in chronic rhinosinusitis/nasal polyps with and without aspirin hypersensitivity

Chronic rhinosinusitis with nasal polyps (CRSwNP) and asthma frequently coexist and are always present in patients with aspirin exacerbated respiratory disease (AERD). Although the pathogenic mechanisms of this condition are still unknown, AERD may be due, at least in part, to an imbalance in eicosanoid metabolism (increased production of cysteinyl leukotrienes (CysLTs) and reduced biosynthesis of prostaglandin (PG) E₂), possibly increasing and perpetuating the process of inflammation. PGE₂ results from the metabolism of arachidonic acid (AA) by cyclooxygenase (COX) enzymes, and seems to play a central role in homeostasis maintenance and inflammatory response modulation in airways. Therefore, the abnormal regulation of PGE₂ could contribute to the exacerbated processes observed in AERD. PGE₂ exerts its actions through four G-protein-coupled receptors designated E-prostanoid (EP) receptors EP1, EP2, EP3, and EP4. Altered PGE₂ production as well as differential EP receptor expression has been reported in both upper and lower airways of patients with AERD. Since the heterogeneity of these receptors is the key for the multiple biological effects of PGE₂ this review focuses on the studies available to elucidate the importance of these receptors in inflammatory airway diseases.

Electronic supplementary material

The online version of this article (doi:10.1186/s12931-014-0100-7) contains supplementary material, which is available to authorized users.     

The use of oral prostaglandin E2 in the management of intrauterine fetal death

12 otherwise healthy patients with intrauterine fetal death 1 to 6 weeks earlier were treated with oral prostaglandin E₂. 9 of the 12 patients delivered within 48 hours after treatment began. 2 others delivered within 48 hours after unsuccessful treatment ceased. In a third patient the cervix relaxed after treatment, and the uterine contents were removed by curettage. No serious complications, such as hemorrhage occurred. The uterus seemed surprisingly responsive to oral prostaglandin E₂ in cases of intrauterine fetal death.     

Sexual behavior,insemination and development of the freshwater leech Helobdella stagnalis (Annelida,Hirudinea, Glossiphoniidae)

The reproductive biology of the Glossiphoniidae leech Helobdella stagnalis, under laboratory conditions, the structure of its eggs and its developmental stages were studied. Sperm transfer and insemination are made by hypodermic injection: one or numerous spermatophores were attached to the skin of the partner during copulation and sperm is injected through it. The leeches can copulate repeatedly with several partners. Each leech produces 4–8 cocoons containing 20–60 eggs whose are attached to the ventral side of the parent and carried around. At 22 °C, the developmental duration is 24 days from the oviposition until the juveniles leave the parent leech. Three (3) major stages have been distinguished: Eggs cleavage, germinal band generation, and juveniles hatching. The sexual behavior of Hellobdella stagnalis was described with a special attention given to parental care.     

Mechanotransduction in bone cells proceeds via activation of COX-2, but not COX-1

Cyclooxygenase (COX) is the key enzyme in the production of prostaglandins, which are essential for the response of bone to mechanical loading. We determined which COX-isoform, COX-1 or COX-2, determines loading-induced prostaglandin production in primary bone cells in vitro. Mouse and human bone cells reacted to 1 h of pulsating fluid flow (PFF, 0.6+/-0.3 Pa at 5 Hz) with an increased prostaglandin E(2) production, which continued 24 h after cessation of PFF. Inhibition of COX-2 activity with NS-398 abolished the stimulating effect of PFF both at 1 h and at 24 h post-incubation, while inhibition of COX-1 by SC-560 affected neither the early nor the late response to flow. PFF rapidly stimulated COX-2 mRNA expression at 1 h but did not affect COX-1 mRNA expression. COX-2 mRNA expression was still significantly enhanced 24 h after cessation of PFF. We conclude that COX-2 is the mechanosensitive form of COX that determines the response of bone tissue to mechanical loading.     

Gastric emptying in response to IAPP and CCK in rats with subdiaphragmatic afferent vagotomy

In the subdiaphragmatic vagal deafferentation procedure (SDA), the afferent fibers of the vagus are surgically severed unilaterally where they enter the brain stem. The technique includes a subdiaphragmal truncal vagotomy performed on the contralateral side. This procedure has been used to study the control of food intake, but it has not been used previously to investigate the role of vagal afferent fibers in the control of gastric emptying (GE). The current experiment studied the effect of SDA on the inhibition of GE by islet amyloid polypeptide (IAPP or amylin) and cholecystokinin (CCK) in awake, unrestrained rats with gastric cannulas. The experimental group underwent subdiaphragmatic vagal deafferentation; the control group had sham operations. All rats received 20-min intravenous infusions of IAPP (1, 3, 9, 27, and 81 pmol/kg/min), CCK (3, 30 and 90 pmol/kg/min), and normal saline in random order. Gastric emptying of saline was measured by the phenol red method during the last 5 min of each infusion period. CCK dose-dependently inhibited gastric emptying in both the control and SDA animals. The inhibition of GE by CCK was significantly attenuated by SDA (p<0.01). IAPP also inhibited gastric emptying dose-dependently, but the difference between the SDA and control groups was not significant. The current experiment, which used a different methodology than previous studies, provides support for the hypothesis that the inhibition of gastric emptying by CCK, but not by IAPP, is mediated partly by afferent vagal fibers.