Structure and Development of the Notochord ‘Elastica Externa’ and Nearby Components of the Elastic Fibre System of Agnathans

Between 15 days and 3 months in age, the ‘elastica externa’ of the notochord sheath of larval lampreys develops from patches of moderately dense and amorphous material into a thick, continuous and electron-dense layer. In both lampreys and hagfish, this layer stains strongly with Verhoeff's elastic stain and aldehyde fuchsin and is penetrated by collagen fibrils on both its outer and inner boundaries. Peroxidase labelling using an antibody raised against human elastin specifically labels both the notochord ‘elastica externa’ and the elastic fibre system of lampreys. The diameters of the microfibrils (10–13 nm) of the oxytalan, elaunin and elastic fibres of lampreys and hagfish are the same as those of higher vertebrates. The connective tissue immediately dorsal and ventral to the notochord of lampreys contains mainly oxytalan fibres in very young ammocoetes, a combination of oxytalan, elaunin and elastic fibres in older ammocoetes, and predominantly elastic fibres in adult lampreys. While the region of the endomeninx at the base of the spinal cord contains almost exclusively oxytalan fibres in young ammocoetes, it also possesses numerous elastic fibres in adult lampreys. These findings indicate that, as in higher vertebrates, the elastic fibres of lampreys develop from oxytalan fibres via elaunin fibres.     

Parasitic Attachments by Overwintering Silver Lampreys, Ichthyomyzon unicuspis, and Chestnut Lampreys, Ichthyomyzon castaneus

We present evidence that at least some parasitic-phase silver lampreys, Ichthyomyzon unicuspis, and chestnut lampreys, I. castaneus, remain attached to host fish during the winter. Lake sturgeon, Acipenser fulvescens, harvested through the ice by spearfishers in the Lake Winnebago system in Wisconsin may bear silver lampreys or fresh lamprey wounds, and sturgeon with lamprey marks were significantly larger than sturgeon without them. Silver lampreys collected on paddlefish, Polyodon spathula, in the Wisconsin River in March were not significantly longer than silver lampreys collected previously in late October, but they were significantly heavier, an indication that they were feeding to at least some extent during the intervening period. Other large fish species, including northern pike, Esox lucius, and flathead catfish, Pylodictus olivaris, have been collected or observed during the winter with silver or chestnut lampreys attached. Although energy and nutrient intake by parasitic lampreys may be reduced during the winter, lampreys attached to hosts may also benefit from the hosts' mobility and ability to avoid potentially harmful situations.     

Neuroendocrine and behavioral responses to weak electric fields in adult sea lampreys (Petromyzon marinus)

We characterized the behavioral and neuroendocrine responses of adult sea lampreys (Petromyzon marinus) to weak electric fields. Adult sea lampreys, captured during upstream spawning migration, exhibited limited active behaviors during exposure to weak electric fields and spent the most time attached to the wall of the testing arena near the cathode (−). For adult male sea lampreys, exposure to weak electric fields resulted in increased lamprey (l) GnRH-I mRNA expression but decreased lGnRH-I immunoreactivities in the forebrain, and decreased Jun (a neuronal activation marker) mRNA levels in the brain stem. Similar effects were not observed in the brains of female sea lampreys after weak electric field stimulation. The influence of electroreception on forebrain lGnRH suggests that electroreception may modulate the reproductive systems in adult male sea lampreys. The changes in Jun expression may be associated with swimming inhibition during weak electric field stimulation. The results for adult sea lampreys are the opposite of those obtained using parasitic-stage sea lampreys, which displayed increased activity during and after cathodal stimulation. Our results demonstrate that adult sea lampreys are sensitive to weak electric fields, which may play a role in reproduction. They also suggest that electrical stimuli mediate different behaviors in feeding-stage and spawning-stage sea lampreys.     

Isolation of Aeromonas salmonicida from sea lamprey (Petromyzon marinus) with furuncle-like lesions in Lake Ontario

For the past six decades, parasitic sea lampreys (Petromyzon marinus) have caused devastating losses to salmonid fisheries in the Great Lakes. To reduce the number of sea lampreys, the Great Lakes Fishery Commission began a large-scale program based on trapping male sea lampreys, sterilizing them, and releasing sterile males back into streams to compete with fertile males for spawning females. The transfer of lampreys among lakes can potentially lead to the transfer of various pathogens, and this has raised major concerns regarding the possibility of resident fish populations becoming infected by introduced pathogens. During a health inspection of sea lampreys collected from Lake Ontario, lampreys with obvious furuncle-like lesions (1-2 cm in diameter) were noticed. Most of the furuncles occupied the dorso-lateral musculature, and Aeromonas salmonicida subsp. salmonicida was isolated from the kidneys. This bacterium was cultured from kidneys of 2.5% of the sea lampreys collected from two locations within the Lake Ontario watershed in 2004. The identity of bacterial colonies was presumptively verified with biochemical reactions and confirmed with polymerase chain reaction. This is the first report of A. salmonicida infection in sea lamprey in the Great Lakes basin associated with furunculosis.     

The role of dermal photoreceptors during the sea lamprey (<Emphasis Type="Italic">Petromyzon marinus</Emphasis>) spawning migration

Light avoidance in larval lampreys is mediated by dermal photoreceptors located in the tail. These photoreceptors continue to function in adults, but they seem redundant because post-metamorphic lampreys possess well-developed eyes. This study examined the role of dermal photoreceptors in adult sea lampreys by testing whether temperature-induced changes in refuge-seeking behavior are mediated by a reduction in dermal photosensitivity. In a lighted arena containing a single shaded refuge platform, lampreys at 22°C displayed five times less search activity and were less likely to attach beneath the refuge platform than lampreys at either 7 or 15°C. A behavioral assay for tail photosensitivity (locomotor response to tail illumination) revealed a corresponding reduction in dermal photosensitivity at 22°C. Moreover, the responses to head illumination (eyes and pineal) did not correspond with the observed light avoidance behaviors. The head was less responsive to light than the tail and was not influenced by temperature. These results provide strong evidence that the dermal photoreceptors continue to mediate light avoidance in adult lampreys, even though adults possess fully functional eyes. The fact that the eyes apparently do not take on this role suggests that there is functional specialization between these two light sensing systems.     

Shifting patterns of nitrogen excretion and amino acid catabolism capacity during the life cycle of the sea lamprey (Petromyzon marinus)

The jawless fish, the sea lamprey (Petromyzon marinus), spends part of its life as a burrow-dwelling, suspension-feeding larva (ammocoete) before undergoing a metamorphosis into a free swimming, parasitic juvenile that feeds on the blood of fishes. We predicted that animals in this juvenile, parasitic stage have a great capacity for catabolizing amino acids when large quantities of protein-rich blood are ingested. The sixfold to 20-fold greater ammonia excretion rates (J(Amm)) in postmetamorphic (nonfeeding) and parasitic lampreys compared with ammocoetes suggested that basal rates of amino acid catabolism increased following metamorphosis. This was likely due to a greater basal amino acid catabolizing capacity in which there was a sixfold higher hepatic glutamate dehydrogenase (GDH) activity in parasitic lampreys compared with ammocoetes. Immunoblotting also revealed that GDH quantity was 10-fold and threefold greater in parasitic lampreys than in ammocoetes and upstream migrant lampreys, respectively. Higher hepatic alanine and aspartate aminotransferase activities in the parasitic lampreys also suggested an enhanced amino acid catabolizing capacity in this life stage. In contrast to parasitic lampreys, the twofold larger free amino acid pool in the muscle of upstream migrant lampreys confirmed that this period of natural starvation is accompanied by a prominent proteolysis. Carbamoyl phosphate synthetase III was detected at low levels in the liver of parasitic and upstream migrant lampreys, but there was no evidence of extrahepatic (muscle, intestine) urea production via the ornithine urea cycle. However, detection of arginase activity and high concentrations of arginine in the liver at all life stages examined infers that arginine hydrolysis is an important source of urea. We conclude that metamorphosis is accompanied by a metabolic reorganization that increases the capacity of parasitic sea lampreys to catabolize intermittently large amino acid loads arising from the ingestion of protein rich blood from their prey/hosts. The subsequent generation of energy-rich carbon skeletons can then be oxidized or retained for glycogen and fatty acid synthesis, which are essential fuels for the upstream migratory and spawning phases of the sea lamprey's life cycle.     

Behavioural patterns of sea lampreys' spawning migration through difficult passage areas, studied by electromyogram telemetry

Electromyogram telemetry (EMG) was used to study the behaviour of sea lamprey Petromyzon marinus during migration through difficult passage areas. When swimming on stretches free from obstacles, the EMG values of sea lampreys showed a constant pattern of activity. Conversely, when negotiating difficult passage areas their behaviour were characterized by alternating between short bursts of intense activity ( i.e . burst movements) and periods of motionless rest during which sea lampreys attached to convenient structures by means of the oral sucker. During blockstone weirs negotiation, tagged sea lampreys spent on average 23% of the time swimming rapidly by means of multiple bouts of burst movements of 31 s duration. To recover from each burst movement, the sea lampreys spent on average 2 min 25 s resting. Sea lampreys seemed affected by increasing fatigue, which probably resulted from initiating a new burst movement without fully recovering physiologically from the previous efforts.     

The presence of teleost-type angiotensin components in lamprey buccal gland suggests a role in endocrine mimicry

Previous characterization of a native lamprey angiotensin II (LpAng II) that possesses a different sequence and function than teleost-type angiotensin II (Ang II) has raised a question as to the role of teleost-type angiotensin peptides in lampreys. In this study, teleost-type angiotensin like-peptides were identified in the buccal gland of lampreys by immunoassays and immunohistochemistry. The possible sources of angiotensin like-peptides were investigated in lampreys by manipulating their choice of host and food. Ang II immunoreactivity (irAng II) was detected in the buccal gland and plasma of feeding phase sea lampreys exposed to Atlantic cod, but was mostly absent in fasting lamprey. Qualitatively, the HPLC profiles of irAng II observed in the plasma, when present, were highly similar to those in buccal gland, implying that the buccal gland could be a source of plasma Ang II. Japanese lampreys force-fed with dogfish blood had significantly elevated concentrations of irAng II in their buccal glands when compared to unfed individuals, suggesting that feeding stimuli may have enhanced buccal gland activity. Teleost-type Ang II-containing proteins, other than angiotensinogen, are present in the buccal gland as trypsinization generated Ang II in vitro, and the HPLC profile of these irAng II was highly comparable to those naturally present in the buccal gland. [Asn¹, Val⁵, Thr⁹]-Ang I that was identified in the buccal gland of Japanese lampreys has the same amino acid sequence to those previously isolated from the incubation of plasma and kidney extract, providing an alternative explanation for the previous isolation of teleost-type Ang I in lampreys. irAng I and irAng II were localized in the granule-like structures in the apical region of the secretory epithelia, suggesting that these peptides may be active components of lamphredin. The teleost-type angiotensin peptides in the buccal gland secretion suggested that these host-specific peptides could be part of the endocrine mimicry strategy used by lampreys to evade host immune responses and reduce immune-rejection.     

Profiles of the female-specific serum protein in the Japanese river lamprey, Lampetra japonica (Martens), and the sand lamprey, Lampetra reissneri (Dybowski), in relation to sexual maturation

The occurrence of female-specific serum protein (FSSP) was examined in the anadromous lamprey, Lampetra japonica, and a freshwater lamprey, Lampetra reissneri, by an immunodiffusion technique using antiserum raised against the serum of sexually mature female lamprey. A considerable amount of FSSP was consistently detected in the blood of vitellogenic, but not in that of nonvitellogenic, females of both species of lampreys. The relative amount of the FSSP tended to become larger in female lampreys at the spawning period. The FSSP may correspond to the yolk precursor protein which is incorporated into ovarian oocytes during the period of exogenous vitellogenesis in the lampreys.     

Icelandic lampreys (<Emphasis Type="Italic">Petromyzon marinus</Emphasis>): where do they come from?

The recent discovery of sea lamprey wounds on salmonids in Icelandic rivers prompted an investigation on the origin of sea lampreys in Icelandic waters. Using a mitochondrial DNA fragment, the origin of the lampreys examined was assigned to the European stock and not to the North American one.     

Attachments by parasitic lampreys within the branchial cavities of their hosts

Silver Lamprey (Ichthyomyzon unicuspis) in the Wisconsin River attached within the branchial cavity of Paddlefish (Polyodon spathula) significantly more often (6.5% of 1,578 attachments) than would be expected by chance on the basis of its relative surface area (3.7%), with as many as four lampreys found together within the branchial cavity of the same Paddlefish. Similar behavior has occasionally been noted for lampreys in other systems, but the phenomenon may be underreported when the lampreys are concealed from view. As suggested previously for some parasitic trychomycterid catfishes, lampreys in the branchial cavity may benefit from accessing blood under pressure, especially in the ventral aorta. In addition, Silver Lamprey within the branchial cavities of Paddlefish may be relatively protected from any negative effects that might result from Paddlefish breaching.     

High frequency of mating without egg release in highly promiscuous nonparasitic lamprey <Emphasis Type="Italic">Lethenteron kessleri</Emphasis>

Nonparasitic lampreys are highly promiscuous: a single female can mate over several dozen times with multiple males. It remains unknown why females mate so frequently despite presumed costs from an elongated spawning period. This paper documents that female Siberian brook lampreys mate without egg release (termed “sham mating”) at remarkably high frequencies. Females mated 20–196 times during a breeding experiment, of which sham mating comprised 35–90%. The number of eggs released may be physically constrained in each mating by the lamprey’s elongated body and behavior. Female lampreys might also control egg release depending on surrounding males.     

Comparative study of lipids and fatty acids in blood plasma of river lamprey Lampetra fluviatilis and brown frog Rana temporaria at the periods of elimination of exogenous feeding

Lipids of blood plasma of lampreys and frogs are composed of phospholipids, triglycerides, free fatty acids (FA), cholesterol, cholesterol esters, and waxes. The lipids content in plasma of frogs is markedly lower as compared with that of lampreys. However, the percentage of lipid components is represented by close values. Fluidity of triglycerides and phospholipids in lampreys is determined predominantly by monoenic acids and polyenoic acids of the ω-3-type, whereas that in frogs—by monoenic acids and polyenic acids of the ω-6-type. Free FA are represented mainly by saturated and monoenic acids.     

A second lamprey from the Lower Carboniferous (Namurian) of Bear Gulch,Montana (U.S.A.)

A second fossil lamprey is described from the Lower Carboniferous (Namurian) locality of Bear Gulch (Montana, U.S.A.), which had already yielded the type specimen of Hardistiella montanensis, the oldest and, probably, the most primitive of all known lampreys. This new specimen, which may possibly belong to the latter species, is badly preserved, but clearly shows the impressions of four to six branchial pouches which are relatively small and closely set, like those of the other Carboniferous lamprey Mayomyzon pieckoensis. They differ thus from extant lampreys, in which the branchial apparatus extends relatively far behing the eyeballs. This concentration of the branchial apparatus in early lampreys is regarded here as a primitive condition, which is also met with in many anaspids, as evidenced from their closely-set external branchial openings. The presence of an impression which recalls the loop of the trabecles in larval extant lampreys suggests that this specimen was a larval individual.     

Lamprey parasitism of sharks and teleosts: high capacity urea excretion in an extant vertebrate relic

We observed 10 sea lampreys (Petromyzon marinus) parasitizing basking sharks (Cetorhinus maximus), the world's second largest fish, in the Bay of Fundy. Due to the high concentrations of urea in the blood and tissues of ureosmotic elasmobranchs, we hypothesized that sea lampreys would have mechanisms to eliminate co-ingested urea while feeding on basking sharks. Post-removal urea excretion rates (J(Urea)) in two lampreys, removed from separate sharks by divers, were initially 450 ( approximately 9000 micromol N kg-1 h-1) and 75 times ( approximately 1500 micromol N kg-1 h-1) greater than basal (non-feeding) rates ( approximately 20 micromol N kg-1 h-1). In contrast, J(Urea) increased by 15-fold after parasitic lampreys were removed from non-ureosmotic rainbow trout (Oncorhynchus mykiss). Since activities of the ornithine urea cycle (OUC) enzymes, carbamoyl phosphate synthetase III (CPSase III) and ornithine carbamoyl transferase (OCT) were relatively low in liver and below detection in intestine and muscle, it is unlikely that the excreted urea arose from de novo urea synthesis. Measurements of arginase activity suggested that hydrolysis of dietary arginine made a minor contribution to J(Urea.). Post-feeding ammonia excretion rates (J(Amm)) were 15- to 25-fold greater than basal rates in lampreys removed from both basking sharks and rainbow trout, suggesting that parasitic lampreys have a high capacity to deaminate amino acids. We conclude that the sea lamprey's ability to penetrate the dermal denticle armor of sharks, to rapidly excrete large volumes of urea and a high capacity to deaminate amino acids, represent adaptations that have contributed to the evolutionary success of these phylogenetically ancient vertebrates.     

Pineal-dependent locomotor activity of lamprey,Lampetra japonica,measured in relation to LD cycle and circadian rhythmicity

Summary Locomotor activity of the river lamprey, Lampetra japonica, was investigated under a light-dark (LD 1212) cycle and under continuous dark conditions. Intact lampreys were entrained to the light:dark cycle. They were active mainly in the early half of the dark period and inactive in light period. The light:dark entrainment continued in 72.7% of lampreys after the removal of bilateral eyes, but additional pinealectomy made the entrainment disappear in all lampreys. When lampreys were pinealectomized with their eyes intact, light: dark entrainment was abolished in most cases. The results indicate that the pineal organ of the lamprey is a photoreceptive organ responsible for synchronizing locomotor activity to LD cycle. Under continuous dark conditions, the locomotor activity began to free-run with a period of 21.3 ± 0.9 h (mean ± SD, n = 53). This circadian rhythmicity was not affected by the removal of lateral eyes but was abolished by pinealectomy. The pineal organ appears to function as an oscillator, or as one of the oscillators, for the circadian locomotor rhythm of lampreys.Abbreviations DD continuous dark - LD light:dark     

Allozyme Polymorphisms and Biochemical-Genetic Taxon Markers in Lampreys (Lampetra, Eudontomyzon, Petromyzon)

Variation at 23 putative enzyme-coding loci was scored in 424 lampreys, including 321 European brook lampreys (Lampetra planeri), 83 European river lampreys (L. fluviatilis), 11 Ukrainian brook lampreys (Eudontomyzon mariae), and nine sea lampreys (Petromyzon marinus). Twelve polymorphisms are described for Lampetra species (LDH*, SOD-2*, PNP*, AAT-1*, AK-1*, ES-2*, LGL*, MPI*, GPI-1*, GPI-2*, PGM*, IDHP-2*), and two each for E. mariae (GPI-1*, ME-2*) and P. marinus (MDH-1*, ME-2*). Diagnostic allozymes are presented for the discrimination of lamprey taxa, some of which are difficult to recognize from the morphology of ammocoetes larvae, the life stage usually encountered when collecting cyclostomes. The allelic markers described permit the clear allocation to a genus, except for the species L. fluviatilis and L. planeri, which are not differentiated by qualitative allozyme analysis.     

Steroid and thyroid hormone profiles following a single injection of partly purified salmon gonadotropin or GnRH analogues in male and female sea lamprey

The effects of exogenous administration of gonadotropin-releasing hormone (GnRH) analogues or of a partly purified salmon gonadotropin extract (GTH) on the duration of steroid and thyroid hormone levels were determined in female and male sea lampreys, Petromyzon marinus, tested under differing temperature and reproductive status. Plasma estradiol levels, but not androgens, were significantly elevated in response to the GnRH analogues or GTH injection compared to controls in female and male lampreys. Higher temperature and/or advance in time of maturation appeared to be inversely related to plasma estradiol levels. These data provide further evidence of hypothalamic control over reproductive function in lampreys. Plasma thyroxine was significantly elevated after female lampreys were treated with GTH, GnRHa (10 micrograms/lamprey) or GnRHa (1 microgram/lamprey) compared to controls. The present study is the first to demonstrate that the GnRH analogue stimulated in some way the pituitary-thyroid axis. These data suggest that a GnRH activity may activate both gonado- and thyrotropic secretion or that the endogenous hormone may itself have both functions in one of the most primitive vertebrates, the sea lamprey.