Culture of cells from tissues of adult and larval sea lamprey

Methods were developed for the culture of cells derived from tissues of the sea lamprey (Petromyzon marinus). Cultures were initiated from gill, liver, muscle and gut from larvae and newly transformed individuals and brain, heart, kidney and ovary from sexually mature adults. The lamprey cells were viable for up to six months in culture and cells from ovary, muscle, gut, gill and liver were propagated for multiple passages. For all cultures except liver, optimal cell attachment and spreading was obtained on surfaces coated with fibronectin and collagen. Optimal liver cell attachment was achieved on basement membrane. Cells synthesizing DNA were detected by precursor incorporation in five week-old cultures derived from adult and larval tissues. Metabolic labeling experiments with [³⁵S]-methionine demonstrated that cultures initiated from liver and ovary continued to synthesize and release proteins into the medium for several weeks. Ultrastructural examination revealed the presence of ciliated cells in cultures from brain and the accumulation of lipid in epithelial cells derived from liver and gill.     

Biology of the southern brook lamprey,Ichthyomyzon gagei

Synopsis Larval, metamorphosing and adult representatives of the southern brook lamprey,Ichthyomyzon gagei, were collected from Choclafaula Creek in Alabama to provide information on aspects of its life cycle. The proportional length of the prebranchial, branchial, trunk and tail regions changed throughout larval life in a pattern similar to that demonstrated in other lampreys. Body depth changed also throughout much of larval life. Oral disc, dorsal fin height, and eye diameter increased through all or most of metamorphosis and the adult interval. Length-frequency curves for larvae indicate a larval life of 3 1/4–4 1/4 years. Growth rate, reflected by change in total length, was constant throughout the year for each age group. Sex ratio of larvalI. gagei was 1: 1.02. Metamorphosis is initiated in early September and is completed between early February and mid March. Adult 1. gagei spawn between mid April and early May. The number of oocytes ranged from 820–2485 between 95–150 mm total length. The maximum gonadosomic ratio in males was 17.1 and 26.3% in females.     

Distribution, Feeding and Body Condition of Four Small Fish Species in the Near-shore and Central Areas of Liangzi Lake, China

Synopsis Small fish abundance is usually high in heavily vegetated habitats in Yangtze lakes, China. Visual and swimming barriers created by dense macrophytes beds could reduce feeding efficiency and growth of small fishes. We tested the hypothesis that small fishes in habitats with dense macrophytes would show decreased feeding efficiency and reduced growth rates by comparing feeding efficiency (measured as the relative weight of fore-gut contents), total length, and condition factor of four small young-of-the-year fishes collected in the near-shore (heavily vegetated) and central (less vegetated) areas of Liangzi Lake. Feeding efficiency, total length, or condition factor were each significantly reduced in the near-shore area compared with the central area for Ctenogobius giurinus, Pseudorasbora parva and Carassius auratus auratus. This supports our hypothesis that vegetation abundance may mediate feeding efficiency and growth of small fishes. Although Hypseleotris swinhonis did not show significant decreases in feeding efficiency or growth in the near-shore area, there was not any reversed tendency, i.e. increased feeding rate or growth in the near-shore area compared to the central area. Yibo Cui: Deceased     

Growth,fecundity, and egg size of least brook lamprey,Lampetra aepyptera

Synopsis Eight geographically-diverse (33°11N to 39°14N) populations of least brook lamprey varied significantly in density of larvae and mean length of metamorphosing and adult lampreys (101–152 mm). Absolute fecundity (503–5900 eggs) did not change significantly with stage of post-larval development, suggesting little post-metamorphic atresia; egg dry weight increased to stage 6, the penultimate stage of metamorphosis. Absolute fecundity increased with the cubic power of total length. Relative fecundity (250–1124 eggs per gram total body weight) decreased significantly with increasing length within two streams. Relative fecundity, adjusted to a common length, increased with mean length of the population. Conversely, mean egg dry weight (0.09–0.24 mg, stages 6–8) increased with maternal length within three streams, and decreased with length among streams. In populations where spawning occurs at a small body size, females produce comparatively fewer but larger eggs. Large eggs may be required to counter the effects of an unproductive environment and the accompanying reduction in relative fecundity is compensatory. No relationship was evident between latitude, annual thermal units, or density and relative fecundity or egg size. Mean length of a population was also unrelated to these environmental factors.     

Diet,intestinal morphology,and nitrogen assimilation efficiency in the damselfish,Stegastes lividus,in Guam

Synopsis Analysis of stomach contents of Stegastes lividus revealed a shift from omnivory in juveniles to herbivory in adults. Red algae, primarily Polysiphonia spp., Gelidiopsis intricata and Ceramium spp., formed the bulk of the diet in all size classes. Foraminifera and small crustaceans were of particular importance in the diet of juveniles. Net nitrogen assimilation efficiency of field-fed S. lividus was approximately 61%, and did not vary with fish size. The net assimilation efficiencies of fish fed on Enteromorpha in the laboratory ranged from size-class means of 36–79% for nitrogen and from 29–72% for total organic material, and increased from juveniles to adults. Apparently, juveniles compensate for a lower efficiency in assimilating plant food by including a higher percentage of animal material in their natural diets. The ratios of intestine length to standard length (IL/SL) and to intestine diameter (IL/ID) increased rapidly in juveniles and leveled off in adults. The retention time for ingested food items may only be about one-half as long in juveniles as in adults. The combination of observed IL/ID ratios and estimated retention times proved most valuable in the interpretation of ontogenetic changes in feeding habits and assimilation efficiency.     

Gut morphology,feeding rate and gut clearance in five species of caddis larvae

Feeding rate, the rate of movement of food through the gut and gut morphology of large larvae of five caddis species (Halesus radiatus, Hydropsyche instabilis, Polycentropus kingi, Rhyacophila dorsalis and Potamophylax cingulatus) were investigated in the laboratory. Following 72 hr starvation, P. cingulatus and H. radiatus larvae became satiated (refused prey offered directly to the mouthparts) after consuming 8–11 and 9–13 mayfly nymphs (Baetis rhodani, 3.5–4.6 mm) respectively. Hunger level affected prey consumption. In P. cingulatus, the number of prey consumed over 24 h (at 9.5–12 °C) increased with starvation periods from 0–72 h, but declined following longer starvation periods. Six clearly recognisable gut states (defined by the position of food material in various parts of the gut) can be identified at different times since commencement of a meal. As environmental temperature increased (from 8–12 °C to 15–17 °C), the rate of change of the gut state increased and the food retention time decreased in all species. Feeding periodicity (i.e. nocturnal/diurnal activity) in the field was estimated based on the evacuation rate and the gut state and environmental temperature at the known time of collection. Initiation of consumption of prey appeared to coincide with emptying of the foregut and proximal midgut, whereas actual feeding continued until complete satiation when some threshold fullness of the foregut had been reached. The length of time food was held in the foregut was positively correlated with increasing specialisation of the foregut (particularly elaboration of the proventriculus).     

Habitat selection by larvae of a fluvial lamprey, <Emphasis Type="Italic">Lethenteron reissneri</Emphasis>, in a small stream and an experimental aquarium

Habitat selection by fluvial lamprey larvae, Lethenteron reissneri (Petromyzontidae), was studied in a natural stream and an experimental aquarium to clarify microhabitat requirements for future conservation of natural populations. A gross collection survey of lamprey larvae in the Monbetsu River, southeastern Hokkaido, Japan, revealed a remarkable bias toward distribution in sandy-mud beds. An analysis using Jacobs' electivity index showed that the larvae selectively utilized spaces having regard to shallow water, weak current, deep sandy-mud, and fine substrate particles. A comparison of microhabitat use between small- (≤5 cm) and large-sized larvae (>5 cm) indicated that the latter utilized the space with greater ranges in both water depth and substrate particle size than the former. Both the field survey and laboratory experiments on larval selectivity of physical habitat variables clearly demonstrated that substrate particle size was the most important variable for small-sized larvae whereas both water depth and substrate depth were more important for large larvae. These findings should be applicable in directing attempts at fluvial habitat restoration for conservation of this endangered lamprey species. Received: November 1, 2000 / Revised: September 12, 2001 / Accepted: October 18, 2001     

Feeding dynamics of Palaemon peringueyi (Decapoda,Caridea) in the temporarily open/closed Kasouga Estuary,South Africa

The feeding ecology of the caridean shrimp Palaemon peringueyi in the temporarily open/closed Kasouga Estuary, Eastern Cape, was examined during summer and winter employing stomach fullness index (SFI) analysis. Mean SFI values for male and female shrimp during summer ranged from 0.85% to 2.36% and from 0.56% and 2.61% body dry weight, respectively. During winter, SFI values for males and females were lower, and varied between 0.23% and 1.56% and 0.36% and 1.84% body dry weight, respectively. There were no significant differences in the SFI values between males and females during the two seasons. A peak in the feeding activity for both sexes was observed during the daytime during both summer and winter. For both males and females, non-linear regression analysis best explained the change in SFI over time. The gut evacuation rate constants (k) of males and females during summer were estimated at 0.43 h^?1 and 0.30 h^?1, respectively. These values correspond to a daily food intake equivalent to 16.2% body dwt for males and 11.1% body dwt for females. During winter, the gut evacuation rates of the shrimp were estimated at 0.35 h^?1 for males and 0.28 h^?1 for females. The daily ration for adult male and female shrimp during winter was estimated at 6.1% and 6.8% body dwt, respectively. The lower estimates obtained during winter appear largely to reflect the influence of the reduced water temperatures on the metabolic activities of the shrimp.     

Spawning ecology of the American brook lamprey, <Emphasis Type="Italic">Lampetra appendix</Emphasis>

Synopsis We observed spawning American brook lamprey, Lampetra appendix, in coldwater streams in Minnesota to assess various aspects of their spawning behavior and spawning habitat requirements. Spawning occurred during April and May, at water temperatures ranging from 8.7 to 15.5°C. Average adult lamprey length and mass differed significantly among streams, but there were no significant differences in length or mass between males and females. Overall sex ratio was 1:1, although one stream had significantly more males than females and one stream held significantly more females. Lampreys spawned in groups of 2–14 individuals, averaging 4.2 adults per nest across all streams. Nests were constructed in gravel and cobble substrate just upstream of riffles, spaced at an average density of three nests m⁻². The typical nest was 16 cm in diameter in water 31 cm deep with a bottom current velocity of 14 cm s⁻¹, and was excavated to a depth of 4 cm below the stream bottom; however, some nest characteristics varied significantly in a few streams. Nests were larger in streams with larger spawning groups, deeper water, and slower current velocities. American brook lamprey exhibited spawning behaviors and spawning habitat requirements similar to those of other species of lamprey in North America.     

Distribution of FMRFamide-like immunoreactivity in the brain and neurohypophysis of the lamprey,Lampetra japonica

Summary Distribution of molluscan cardio-excitatory tetrapeptide Phe—Met—Arg—Phe—NH₂ (FMRFamide) was determined by means of immunohistochemistry in the brain and neurohypophysis of the lamprey, Lampetra japonica. Many FMRFamide-like immunoreactive neurons were found in the periventricular nuclear region and in a region near the mammillary recess. Neurons situated in the former region were larger. The immunoreactive cell groups were shown to be located at sites differing from those of the AF-positive cell groups. The fibers of immunoreactive neurons extended in all directions within the brain and towards the spinal cord, some reaching the third ventricle and capillaries. Thus, FMRFamide-like immunoreactive peptides appear to function as neurotransmitters or neuromodulators and possibly also as neurohormones. FMRFamide-like immunoreactive material was rarely observed in the posterior neurohypophysis (neural lobe), but was noted to be present to a limited extent in the caudal part of the anterior neurohypophysis (median eminence). It would thus follow that FMRFamide-like immunoreactive neurons may not necessarily be related to the hypothalamo-neural lobe system, but may possibly be associated with the hypothalamoadenohypophysial system. The pineal body showed no FMRFamide-like immunoreactivity.     

Population genetics of a cyclostome species pair, river lamprey (Lampetra fluviatilis L.) and brook lamprey (Lampetra planeri Bloch)

Horizontal agarose gel electrophoresis of 24 allozyme loci in four species of Central European lampreys (321 Lampetra planeri , 83 L. fluviatilis , 11 Eudontomyzon mariae and nine Petromyzon marinus ) was used to study the 'paired species' L. fluviatilis and L. planeri . The genetic differentiation of the anadromous river lamprey ( L. fluviatilis ) from the stationary brook lamprey ( L. planeri ) was within the range of ingroup differentiation of the latter, but L. fluviatilis exhibited much greater population cohesion over a more extended geographic range: G _ST = 0.0537 versus G _ST = 0.3398, N _em = 4.402 versus N _em = 0.4856, mean genetic among-stock distances D = 0.0047 versus D = 0.0257. L. planeri populations coexisting geographically with L. fluviatilis in the Rhine and Elbe river systems were genetically more cohesive than L. planeri stocks from the Danubian basin where L. fluviatilis is absent. Danubian L. planeri populations exhibit a lower degree of heterozygosity than brook lampreys from the Rhine river system, but comprise deeper genetic lineages ( G _ST = 0.4629 versus G _ST = 0.2434), despite being sampled from a much more restricted area. Isolation-by-distance is observed for L. planeri from the Danubian but not from the Atlantic drainage basins. Transspecific gene flow between L. planeri from Atlantic drainage basins and the long-distant migrating L. fluviatilis is inferred, raising doubt on the validity of two separate biospecies. E. mariae and P. marinus are clearly differentiated from Lampetra spp. at several allozyme loci.     

Demonstration of rod and cone photoreceptors in the lamprey retina by freeze-replication and immunofluorescence

Summary In common with other cyclostomata, the Japanese river lamprey (Lampetra japonica) has a retina consisting of distinct types of photoreceptor cells called long and short photoreceptor cells. After freeze-fracture, disc membranes of these photoreceptor cells were characterized in common by a homogeneous distribution of intramembrane particles on the protoplasmic fracture faces, in contrast to those of the myeloid bodies bearing scattering particles.Immunofluorescent examination was applied to the retina with monoclonal antibodies raised against bovine and chicken rhodopsins. Positive immunoreactivity was found to be limited to outer segments of the short cell, leaving the entire body of the long cell and all other components of the retina negative. The results suggest that the short cell is more closely related to a rod-type photoreceptor cell characterized by rhodopsin as its visual pigment.     

Dissociation of photoreceptor cells from the pineal organ of the lamprey,Lampetra japonica

Summary Photoreceptor cells, nerve cells and supporting cells were dissociated from the pineal organ of the river lamprey, Lampetra japonica, by the use of 10 U/ml papain solution at 28° C for 20 min, followed by repeated trituration. With the aid of Nomarski interferencecontrast optics, photoreceptor cells, nerve cells and supporting cells were readily identified. Electron-microscopic examination revealed that isolated photoreceptor cells display an outer segment endowed with a few lamellar disks and connected to the inner segment (ellipsoid) via a connecting cilium. The structural features of the dissociated photoreceptor and supporting cells strongly resemble the morphology of the respective cellular elements in situ. We succeeded in culturing dissociated cells for time periods up to 48 h when the procedure described in detail was applied.     

Composition of speciose leaf litter alters stream detritivore growth, feeding activity and leaf breakdown

Leaf litter derived from riparian trees can control secondary production of detritivores in forested streams. Species-rich assemblages of leaf litter reflect riparian plant species richness and represent a heterogeneous resource for stream consumers. Such variation in resource quality may alter consumer growth and thus the feedback on leaf breakdown rate via changes in feeding activity. To assess the consequences of this type of resource heterogeneity on both consumer growth and subsequent litter breakdown, we performed a laboratory experiment where we offered a leaf-shredding stream detritivore (the stonefly Tallaperla maria, Peltoperlidae) ten treatments of either single- or mixed-species leaf litter. We measured consumer growth rate, breakdown rate and feeding activity both with and without consumers for each treatment and showed that all three variables responded to speciose leaf litter. However, the number of leaf species was not responsible for these results, but leaf species composition explained the apparent non-additive effects. T. maria growth responded both positively and negatively to litter composition, and growth on mixed-litter could not always be predicted by averaging estimates of growth in single-species treatments. Furthermore, breakdown and feeding rates in mixed litter treatments could not always be predicted from estimates of single-species rates. Given that species richness and composition of senesced leaves in streams reflects riparian plant species richness, in-stream secondary production of detritivores and organic matter dynamics may be related to species loss of trees in the riparian zone. Loss of key species may be more critical to maintaining such processes than species richness per se.     

Growth, dynamics, and distribution of a population of the brook lamprey Lampetra planeri in a South Swedish stream

Size and structure of a South Swedish population of Lampetra planeri larvae were studied during 5 yr. Population size fluctuated considerably within the studied section of stream with a general decline. The fluctuations were positively and strongly correlated (multiple R²= 0.85) with the population density at the beginning of each analysed period, while poorly correlated with such variables as heavy precipitation and temperature. Several factors suggest the entire larval population drifts downstream: fluctuations in the studied stream section, increasing mean size downstream, and incongruity in longitudinal distribution in surveys, performed with a 5-yr interval. Growth, estimated both in caged larvae and from length frequency data, occurred mainly in autumn. High density in cages reduced the growth rate of larvae.     

Membrane differentiation in the cytoplasmic-tubule system of the intestinal absorptive cells of the lamprey,Lampetra japonica

Summary Specific membrane differentiation occurs in the cytoplasmic-tubule system of the absorptive cells lining the mucosa of the lamprey anterior intestine. The absorptive cells are characterized by the presence of abundant mitochondria and a system of well-developed cytoplasmic tubules (120 nm in diameter). The cytoplasmic tubules open on to the basolateral cell surface and contain numerous lipoprotein particles (50–100 nm diam.) in their lumina. Lipoprotein particles are also observed in the endoplasmic reticulum and the Golgi complex, and they are transfered to the lateral intercellular space and lamina propria by way of the cytoplasmic tubules. Spirally-wound parallel rows of particles are found in the luminal surface of the cytoplasmic tubules. The rows are 17 nm apart and are wound spirally at a pitch of 210 nm. Freeze-fracture images of the tubule membranes also show spiral arrays of particles (9 nm in diameter) on the P-face, and complementary shallow grooves on the E-face. From these observations, it is suggested that the cytoplasmic-tubule system of the intestinal absorptive cells serves as a channel for the transport of synthesized lipoprotein into the interstitium, and is also the site of the ion and water exchange essential for the maintenance of ionic homeostasis.     

Stellate cells storing retinol in the liver of adult lamprey,Lampetra japonica

Summary Distribution, localization and fine structure of the stellate cells in the liver of lamprey, Lampetra japonica, were studied during the spawning migration by use of Kupffer's gold-chloride method, fluorescence microscopy for vitamin A (retinol) and electron microscopy. The stellate cells in the lamprey liver differ in some of their properties from those in mammalian livers. Stellate cells which store abundant retinol in lipid droplets, occur not only in the hepatic parenchyma, but also in the dense perivascular and capsular connective tissue of the liver and in the interstitium of pancreatic tissue. In the hepatic parenchyma these cells are located perisinusoidally or along thick bundles of collagen fibrils. The stellate cells display a number of large retinol-containing lipid droplets, granular endoplasmic reticulum, tubular structures, dense bodies, Golgi complex, microtubules, and microfilaments. In the space of Disse, the stellate cells and extracellular fibrilar components such as collagen fibrils and microfibrils (11–12 nm in diameter) are intervened between the two layers of basal laminae. Differentiation and possible functions of the stellate cells in the lamprey liver are discussed.     

Demonstration by lectin cytochemistry of rod and cone photoreceptors in the lamprey retina

Summary Lectin cytochemical analysis was undertaken to examine the distribution of glycoconjugates associated with the short and long photoreceptor cells in the lamprey retina. Concanavalin A bound preferentially to the outer segment region of the short cells. Wheat germ agglutinin bound weakly to both long and short cells. The outer segment regions of the long cells were stained intensely with peanut agglutinin. Pretreatment with neuraminidase to remove sialic acid resulted in decreased binding of wheat germ agglutinin throughout the retina and increased binding of peanut agglutinin to the outer segment region of the short cells and the region of myoid process of the long cells. These results suggest that there is a difference in the distribution of glycoconjugate residues between the long and short cells. A rod-like character of the short cell and a cone-like character of the long are tentatively discussed. Lectin-binding patterns in other retinal regions is also examined.     

Iodine distribution in the thyroid follicles of the hagfish,Eptatretus burgeri and lamprey,Lampetra japonica: Electron-probe X-ray microanalysis

Summary In the thyroid follicles of species of cyclostomes, a hagfish and a lamprey, the distribution of stable iodine was examined by electron-probe X-ray microanalysis. A high concentration of stable iodine, heterogeneously distributed, was observed in the follicular cells of hagfish thyroid follicles. In the lamprey a low concentration of iodine was seen in the follicular lumina. The relative values for stable iodine determined in this way corresponded to values obtained by a chemical analytical method.