Diet composition influences the fitness of the herbivorous crab Grapsus albolineatus

The tropical rocky shore crab Grapsus albolineatus feeds primarily on filamentous algae but eats animal matter whenever it is available. During the summer the crab's diet switches to encrusting algae due to a die-off of filamentous algae. As a result of the switch the nutrients in the diet of the crab vary seasonally and may influence the fitness of the crab. Maintenance, growth, reproductive performance and nutrient storage of crabs were examined under four dietary regimes of increasing nutritional value ranging from low organic to high protein content. The nutritional quality of these diets significantly affected crab survival and moulting. Crabs fed on the nutritionally superior diet of algae and meat exhibited enhanced growth, higher levels of energy in the reproductive organs and stored more energy in the hepatopancreas than did individuals on the shore and crabs fed only on algal diets in the laboratory. Filamentous algae were a better food source than other algae, resulting in fewer deaths and superior levels of maintenance and growth. Growth and maintenance can occur on a pure algal diet, but reproductive performance and nutrient storage require some degree of added nutrients in the form of animal matter in the diet. Crabs fed coralline or foliose algae had higher mortality and fewer successful moults than crabs fed the other two diets. The fitness of G. albolineatus appears to be limited by the amount of extra nutrients obtained from animal matter. The opportunistic consumption of animal material in the form of carrion, or of animals associated with dietary algae, could be a key factor in the reproductive success of this crab.     

Systematic revision of acoels with 9+0 sperm ultrastructure (Convolutida) and the influence of sexual conflict on morphology

We have used newly discerned morphological characters as well as molecular‐sequence data from 18S and 28S rDNA to revise the families recently designated as the ‘9+0’ acoels – what we call Convolutida. Characters from the ultrastructure of sperm, with their ‘9+0’ axonemes, are useful in delineating the Convolutida, but are either species‐specific or too conserved within the group to be used to infer relationships within it. Male genital organs, prostatoid organs, and sagittocysts, on the other hand, give a good phylogenetic signal for reconstructing relationships of such genera as Conaperta, Anaperus, and Achoerus; some features of the reproductive organs correlate with habitat and show how the Convolutida probably originated as epiphytic predators and radiated into the mesopsammon, pelagic, and coral‐associated realms. In this revision of the Convolutida we provide revised synopses of its families – which we restrict to the Anaperidae, Convolutidae, and Sagittiferidae – and describe a new species, Polychoerus gordoni, from New Zealand. We transfer the genus Adenopea from the Antroposthiidae to the Convolutidae; Conaperta, Neochildia, and Oxyposthia from the Convolutidae to the Anaperidae; Paranaperus and Praeanaperus from the Anaperidae to the Haploposthiidae. Convoluta aegyptica is synonymized with Convoluta boehmigi, Convoluta lacazii with Convoluta sordida, and the genus Picola (Convolutidae) with Deuterogonaria (Haploposthiidae). Amphiscolops blumi, A. carvalhoi, and A. langerhansi, all of which possess a cellular seminal bursa, are transferred to the genus Heterochaerus. Convoluta elegans and Pseudanaperus tinctus are classified as nomina nuda. We use our findings on the ultrastructure of female genital organs and spermatozoa to show that sexual conflict plays a major role in the evolution of diversity of these structures and that the phylogeny of the Acoela would comprise early forms without female genital organs and hyper‐ or hypodermal transfer of sperm through advanced forms with ever longer and narrower bursal nozzles and sperm with axial microtubules. Moreover, our results show that the acquisition of endosymbiotic algae happened at least twice within the Acoela.     

PREPARATION AND CHARACTERIZATION OF PROTOPLASTS OBTAINED FROM THE PRASINOPHYTE SCHERFFELIA DUBIA (CHLOROPHYTA)1

The prasinophyte genera Scherffelia and Tetraselmis are the only genera that form a cell wall by an extracellular fusion of scales called a theca. We established a protocol for the production of protoplasts from Scherffelia dubia Pascher emend. Melkonian et Preisig using 3 mM Ellman's reagent (5,5′‐dithio‐bis‐2‐nitrobenozoic acid [DTNB]). Protoplasts analyzed by EM lacked flagella and thecae but were otherwise similar to control cells. In response to treatment with DTNB, many protoplasts synthesized new thecal scales in the Golgi apparatus, indicating that cells attempted to regenerate new cell walls. However, complete regeneration of the thecae only occurred once DTNB was washed out from the medium. At higher DTNB concentrations (5 mM), two protoplasts were found within the parental cell wall and scales accumulated between the plasma membrane of the protoplasts and the original theca but failed to form a new theca.     

A MODE OF ENTRY OF BLUE-GREEN ALGAE INTO THE APOGEOTROPIC ROOTS OF MACROZAMIA COMMUNIS

The entry of blue-green algae into an aerial root of Macrozamia communis via a break in the dermal layers and leading through a continuous cortical channel into the algal zone is reported. Penetration of the algae appears to be mainly in the form of filaments. Intercellular migration of the algae is found to occur after dissolution of host cell middle lamellae, but host cortical cells are also destroyed near some of the algae. Intracellular migration is also suggested to be a likely pathway for the algae as they proceed to the algal zone.     

Growth inhibition of dinoflagellate algae in shake flasks: Not due to shear this time!

Large scale algae cultures present interesting challenges in that they exhibit characteristics of typical bacterial and animal cell cultures. One current commercial food additive, docosahexaenoic acid (DHA), is produced using the dinoflagellate algae, Crypthecodinium cohnii. Like animal cell culture, the perceived sensitivity of algae culture to hydrodynamic forces has potentially limited the agitation and aeration applied to these systems. However, the high density cultivation of C. cohnii required for an economically feasible process inevitably results in high oxygen demand. In this study, we demonstrated what first appeared to be a problem with shear sensitivity in shake flasks is most probably a mass transfer limitation. We subsequently demonstrated the limit of chronic and rapid energy dissipation rate, EDR, that C. cohnii cells can experience. This limit was determined using a microfluidic device connected in a recirculation loop to a stirred tank bioreactor, which has been previously used to repeatedly expose animal cells to high levels of EDR. Inhibition of cell growth was observed when C. cohnii cells were subjected to an EDR of 5.9 × 10⁶ W/m³ with an average frequency of 0.2/min or more. This level of EDR is sufficiently high that C. cohnii can withstand typically encountered hydrodynamic forces in bioprocesses. This result suggests that at least one dinoflagellate algae, C. cohnii, is quite robust with respect to hydrodynamic forces and the scale‐up of process using this type of algae should be more concerned with providing sufficient gas transfer given the relatively high oxygen demand. © 2009 American Institute of Chemical Engineers Biotechnol. Prog., 2010     

CONSPICUOUS ALGAL COLONIZATION OF THE ASH FROM MOUNT ST. HELENS1

Conspicuous growth of green and yellow-green algae developed in arid steppe communities in eastern Washington on the silt-sized ash from the 18 May 1980 eruption of Mount St. Helens. In a veneer on the ash, cell numbers per gram dry ash of these aerial immigrants equaled or exceeded the highest reported values for terrestrial chlorophytes (10⁸ cells · g^?1 dry soil). Such pronounced growth was restricted largely to three microsites (under the canopy of Artemisia tridentata, along the runways of the vole, Microtus montanus, and surrounding emergent agarics), apparently in response to water-holding properties of the ash, localized nutrient input and possible reduction of predation by nematodes. Highest algal counts were in Microtus runways where daily water and urea-N input per adult animal may be 8 mL and 23 mg, respectively. Cyanophytes, commonly considered early colonizers of volcanic ash, were extremely rare. The role of blue-green algae in succession on volcanic materials may be more restricted geographically than previously recognized.     

SCANNING ELECTRON MICROSCOPY OF THECAE OF THE DINOFLAGELLATE GENUS ORNITHOCERCUS1

Members of the genus Ornithocercus are all tropical oceanic species in which the theca is extended in the form of elaborate wing-like projections, the lists, supported by ribs. This paper illustrates the topography of 6 of the species. The theca is penetrated by numerous simple pores opening flush with the outer surface. On the inner side of the thecae the pores have a raised rim. The hypotheca of all species examined except O. splendidus have shallow depressions, the areolae, over most of the surface. Secondary thickening of mature cell walls deepens the appearance of the areolae, and increases their extent over the hypotheca in O. quadratus. The number of pores is not directly correlated with the areolae but seems rather to be a function of cell size. A comparison of the surface features confirms views that O. splendidus occupies a relatively isolated phenetic position. It also confirms the close similarity of O. steinii with O. thumii. Unexpectedly it suggested a phenetic closeness between O. magnificus and O. quadratus on the basis of hypothecal structure and rib features of the left sulcal list. O. heteroporus could not be subjected to the same degree of study and its position cannot be commented on. Some theoretical hydrodynamic and morphogenetic problems in Ornithocercus are discussed.     

Actin in the green algae Coleochaete and Mougeotia

Summary Filaments associated with chloroplasts in cytoplasmic homogenates of the algae Coleochaete scutata and Mougeotia sp. bind rabbit skeletal muscle heavy meromyosin (HMM) to form arrowhead complexes that could be dissociated with ATP. This result suggests that the filaments are actin which may be involved in the characteristic chloroplast movements exhibited by these algae.     

Effect of Lipopolysaccharide on Progesterone Production during Luteinization of Granulosa and Theca cells In Vitro

The aim of this study is to examine the effect of lipopolysaccharide (LPS) on progesterone production during luteinization of granulosa and theca cells isolated from bovine large follicles. Granulosa and theca cells isolated from large follicles of bovine ovaries were exposed to LPS under appropriate hormone conditions in vitro. Progesterone (P4) production in theca cells, but not granulosa cells, was decreased by long‐term exposure of LPS. Long‐term exposure of LPS suppressed the gene expression of luteinizing hormone receptor in theca cells. Although long‐term exposure of LPS did not affect the expression of steroidogenic acute regulatory protein (StAR) and 3β‐hydroxy‐steroid dehydrogenase (3β‐HSD) genes, it did inhibit the protein expression of StAR and 3β‐HSD in theca cells. These findings suggest that theca cells, rather than granulosa cells, are susceptible to LPS during luteinization and that LPS inhibits P4 production by decreasing protein levels of StAR during luteinization of theca cells.     

ZONATION OF DEEP WATER BENTHIC ALGAE IN THE GULF OF MAINE1

Algal community structure is described for a deep-water rock pinnacle in the Gulf of Maine. Three depth zones of algal dominance were apparent consisting of 1) leathery macrophytes (to 40 m), 2) foliose red algae (to 50 m) and 3) crustose algae (fleshy crusts to 55 m and coralline crusts to 63 m). Microscopic filamentous and erect calcareous algae were also present but inconspicuous. Upright macroscopic filamentous and thin sheet-like forms were not observed on the pinnacle. Sea anemones (Metridium senile) dominated some vertical faces and abrupt prominences in the shallowest regions of the pinnacle (to 24 m) and locally appeared to set the upper vertical limits of kelp and possibly foliose reds. Laminaria sp. formed an open park-like canopy from 24 to 30 m whereas Agarum cribrosum, the deepest kelp, grew as isolated individuals to 40 m. Peyssonnelia sp. and Leptophytum laeve were the deepest occurring fleshy (to 55 m) and calcareous crusts (to 63 m), respectively. The occurrence of these algae at record depths for the Gulf of Maine and for cold water marine environments may be the result of an absence of large herbivores and the high productivity potential of the benthos in these relatively clear waters. By compiling data on depth distribution patterns world-wide, it is evident that the three zone structure of algal morphologies observed in the Gulf of Maine is a global phenomenon.     

THE ULTRASTRUCTURE OF MULTILAYERED STRUCTURES ASSOCIATED WITH FLAGELLAR BASES IN MOTILE CELLS OF TRENTEPOHLIA AUREA1

An ultrastructural study of motile cell development in the green alga Trentepohlia aurea has revealed the presence of multilayered structures (MLS) associated With flagellar bases. These MLS are ultrastructurally similar to MLS described in pteridophyte and bryophyte sperm and in the zoospore of the green algae Coleochaete and Klebsormidium. However, 2 MLS are found in each biflagellate motile cell of T. aurea, while other previously described MLS occur singly in biflagellate motile cells. In addition, the MLS of T. aurea consist of fewer microtubules and are structurally simpler than most other MLS described. The MLS of Trentepohlia may represent a stage in the evolutionary development of the MLS of land plants. The presence or absence of the MLS in motile cells of green algae may be a useful character in phylogenetic studies.     

CELL DIVISION IN THE SCALY GREEN FLAGELLATE HETEROMASTIX ANGULATA AND ITS BEARING ON THE ORIGIN OF THE CHLOROPHYCEAE

H. angulata is a scale-covered, asymmetrical green unicell with two laterally attached, anisokont flagella. In recent years it has been classified in the Prasinophyceae. The flagellar apparatus replicates, and the cell begins to cleave at the side opposite the flagella before the nucleus can be perceived to be in prophase. The flagellar apparatuses separate, and the extra-nuclear development of the spindle occurs from the regions occupied by rhizoplasts. Rhizoplasts or partial rhizoplasts lie at the flat metaphase spindle poles. By metaphase, the cell has already elongated to the extent that it is nearly twice as long as at interphase. The spindle and the cell itself elongate greatly during anaphase with a concomitant further separation of the flagellar pairs. Although the interzonal spindle persists during cytokinesis as in charophycean algae, H. angulata is similar in flagellar scale morphology and other characteristics to the chlorophycean Platymonas, which has a collapsing interzonal spindle at telophase, a phycoplast, and a wall-like theca, which develops by the fusion of small stellate scales. It is hypothesized that the collapsing telophase spindle and phycoplast evolved in green flagellates similar to Platymonas, in which cell and spindle elongation became restricted by a cell wall that evolved from stellate scales similar to those in Heteromastix. Such walled flagellates are then visualized as having eventually given rise to Chlamydomonas and to the entire range of chlorophycean algae with phycoplasts. It is pointed out that the hypothesis has a number of implications by which its validity could be judged when sufficient information becomes available.     

The effects of diet mixing on consumer fitness: macroalgae, epiphytes, and animal matter as food for marine amphipods

Herbivores are thought to achieve adequate nutrition by consuming numerous species of plants or by occasionally consuming animal tissue. Although active selection of diverse foods is common in nature, the relationship between diet mixing and consumer fitness is poorly understood, especially in marine environments. We studied the fitness-based consequences of dietary mixing in the sympatric amphipods Ampithoe marcuzzii, A. valida, Cymadusa compta, and Gammarus mucronatus by measuring survivorship, growth, and fecundity of these amphipods when they were offered single species of algae, a single animal food, a mixture of algal species, or a combination of algae and animal matter. For the more sedentary, tube-building amphipods A. marcuzzii, A. valida, and C. compta, fitness on mixed algal diets was matched by fitness on at least one of the monospecific algal diets, suggesting that they could benefit from preferential feeding on those algae in the field. The more mobile amphipod, G. mucronatus, survived and grew similarly on the mixed diets and on the filamentous brown alga Ectocarpus siliculosus. However, its fecundity was significantly higher when feeding on the algal and animal mixture than on Ectocarpus alone. Additionally, for G. mucronatus, fitness on mixed algae, mixed algae plus animal matter, and animal matter alone was equivalent, although female growth (but not gonad production) was slightly lower on animal matter alone than on the mixed algae combined with animal food. Thus the more mobile amphipod, G. mucronatus, was the only species able to perform well on animal food alone. In contrast, A. valida and C. compta experienced large negative effects when limited to consuming animal matter alone. For these two species, combining algae and animal matter did not enhance fitness over combining only algae. Fitness effects of specific algal diets showed some general similarities, but also considerable variance among the amphipods. For example, E. siliculosus was generally better food than other algae for all four amphipods, whereas Sargassum filipendula was generally poor. However, A. marcuzzii did not suffer negative effects of consuming only Sargassum. The red alga Polysiphonia sp. and the green alga Enteromorpha flexuosa decreased fitness in A. marcuzzii, C. compta, and G. mucronatus, but not A. valida, and the negative effects of Polysiphonia were considerably larger for A. marcuzzii than for the other amphipods. Our data show that nutritional requirements, even among related species (e.g., A. marcuzzii and A. valida), can be dramatically different. Diet mixing may benefit more mobile consumers like Gammarus that are better able to search for different foods, and may be less important for more sedentary herbivores like Ampithoe and Cymadusa that consume, and live in close association with, individual host plants. Received: 14 September 1998 / Accepted: 22 October 1999     

Phosphorus availability and nitrogenase activity in aquatic blue-green algae

Metabolically active phosphorus-starved cultures of blue-green algae assimilate ³²P rapidly in the light and in the dark. The uptake of phosphorus results in a rapid (within 15 min) stimulation in acetylene reduction by Anabaena cylindrica, A. flosaquae, Anabacnopsis circuiaris and Chlorogloea fritschii, with a response being obtained to less than 5 μg/1 of phosphorus. Uptake of phosphorus also causes a rapid increase in respiration in the dark but not in photo respiration, and the size of the cellular ATP pool and the ¹⁴CO₂ fixation rate both increase more slowly. The metabolism of phosphorus-sufficient cells, which assimilate phosphorus more slowly, shows little response when phosphorus is provided. Excess phosphorus is stored in the vegetative cells of blue-green algae as polyphosphate bodies which may form within 60 min of adding phosphorus to phosphorusstarved cells and which serve as a source of phosphorus for the algae when exogenous phosphorus is limiting. Preliminary results from Scottish waters suggest that urban effluents are important sources of available-phosphorus for algal growth and that the levels entering fresh waters from agricultural land are, per unit volume, lower. In both types of water the levels of available-phosphorus are rather similar to the levels of orthophosphate-phosphorus present. Most detergents tested serve as a source of phosphorus for nitrogen-fixing blue-green algae and cause a rapid stimulation in reduction when added to phosphorus-starved cultures. Of the detergents assayed, the biological types were richest in available phosphorus. The addition of detergents may result in a rapid increase in number of polyphosphate bodies present in the algae. Detergents in general also contain an inhibitor of algal metabolism. Whether a stimu-lation or an inhibition occurs depends on the quantities of detergent added and on whether or not the alga is phosphorus-deficient.     

STRUCTURE AND ASEXUAL REPRODUCTION OF THE ENIGMATIC CHAROPHYCEAN GREEN ALGA ENTRANSIA FIMBRIATA (KLEBSORMIDIALES,CHAROPHYCEAE)1

As the closest relatives of embryophytes, the charophycean green algae (sensu Mattox and Stewart) may reveal the evolutionary history of characters in this lineage. Recent molecular phylogenetic analysis indicates that the little‐known species Entransia fimbriata Hughes is a member of the charophycean order Klebsormidiales. In this study LM and EM were used to identify and describe additional structural characters of Entransia so that comparisons could be made with Klebsormidium and with other charophycean algae outside the order Klebsormidiales. Features that Entransia shares with various members of the genus Klebsormidium include cylindrical cells in unbranched filaments that may spiral, parietal chloroplasts that cover only part of the circumference of the cell, H‐shaped cross walls, and vegetative reproduction by both fragmentation and formation of zoospores or aplanospores. Among the characteristics that distinguish Entransia from Klebsormidium are a highly lobed chloroplast with multiple pyrenoids; a single large vacuole; short cells that die and collapse, apparently facilitating filament fragmentation; and germinating filaments with condensed adhesive at the base and a tapering spine at the tip. Although Entransia has sometimes been tentatively considered to be a member of the Zygnemataceae, the presence of a flagellate life history stage distinguishes Entransia from this group. The pyrenoids of Entransia are typical of those of charophycean algae in having traversing membranes and surrounding starch. Presence of multiple such pyrenoids in each chloroplast of Entransia supports the hypothesis that the common ancestor of charophycean algae and embryophytes had a single chloroplast with multiple pyrenoids.     

SEXUAL REPRODUCTION AND MEIOSIS IN PERIDINIUM INCONSPICUUM LEMMERMANN (DINOPHYCEAE)

In Peridinium inconspicuum Lemmermann, sexual reproduction occurs in both nitrogen-enriched and nitrogen-deficient media. In this homothallic strain, protoplasmic fusion begins between two thecate gametes; but zygote formation is completed in a space outside the fusing pair. This diploid cell can form a plated theca which is shed as the cell enlarges. This spherical zygote then forms a new non-plated theca. The process of ecdysis and the formation of a new non-plated theca is repeated several times. During this process the zygote gradually elongates and by cytoplasmic infurrowing becomes peanut-shaped. Eventually two cells are formed. The first and second meiotic divisions are greatly separated in time. The first meiotic division occurs in the spherical non-thecate zygote. The second meiotic division can occur in the peanut-shaped zygote before it completes cytokinesis. This meiotic division may not be synchronous, occasionally resulting in a trinucleate stage. Eventually four flagellated, haploid products are produced.     

Filter feeding in the hermit crab

Summary The hermit crab, Pagurus bernhardus is able to remove both Artemia nauplii and unicellular algae from suspension. Crabs with wet weights of 1.1–9.2 g consumed all of the 300 Artemia nauplii contained in 200 ml of sea water within 1 h. Crabs weighing 0.7–1.1 g wet weight filtered suspended Dunaliella algae (8 m) from concentrations of 10–350 million cells per liter at a rate of 26% and 47% within 2 and 5 h, respectively. A similar result was obtained with an 11 g crab. During filter feeding activity a water current is generated by the flagella of the exopods of the second and third maxillipeds. Artemia nauplii are caught by grasping movements of the endopods of the third maxillipeds, whereas filtering of unicellular algae is probably achieved by the two maxillae. It is assumed that filter feeding activity depends on the same structures and behavior as in deposit feeding. P. bernhardus is one more example of a benthic marine animal which may use any food source which becomes available in the course of the seasons.     

PARTICIPATION OF EXTRACELLULAR NUCLEOTIDES IN THE WOUND RESPONSE OF DASYCLADUS VERMICULARIS AND ACETABULARIA ACETABULUM (DASYCLADALES,CHLOROPHYTA)1

As assayed by fluorescent reporter dyes, nitric oxide (NO) and H₂O₂, two downstream signaling agents induced by wounding in the alga Dasycladus vermicularis (Scop.) Krasser, can also be induced in unwounded Dasycladus cells by μM Adenosine 5′[γ‐thio]triphosphate (ATPγS) and Adenosine 5′‐[β‐thio]diphosphate (ADPβS), but not by Adenosine 5′‐O‐thiomonophosphate (AMPS). These nucleotide‐induced responses are blocked by pyridoxalphosphate‐6‐azophenyl‐2′,4′‐disulphonic acid (PPADS), an antagonist of animal purinoceptors, and by adenosine, a feed‐back inhibitor of extracellular nucleotide responses in animals. Similar nucleotide‐ and nucleotide‐antagonist responses were observed in Acetabularia acetabulum (L.) P. C. Silva. Significant levels of ATP released from Dasycladus cells were measured at wound sites by a sensitive luciferin‐luciferase assay. Additionally, the normal wound‐induced production of NO and H₂O₂ in Dasycladus can be blocked by pretreating the cells with PPADS. Our results indicate that nucleotides released from wounds can serve as a signal to trigger wound responses in algae, and that coordinated signaling between extracellular nucleotides and the NO pathway may have been established early during the evolution of plants.     

SELECTIVE INHIBITION OF BLUE-GREEN ALGAL GROWTH BY ETHIONINE AND OTHER AMINO ACID ANALOG1,2

Some metabolic analogs, including azaguanine, azathymine, azauracil, caffeine, 4-azaleucine, dl -ethionine, and dl -p-fluorophenylanlanine, were examined for their ability to repress the multiplication of algae from natural aquatic sources grown in defined or semidefined media. dl -ethionine, dl -p-fluorophenylalanine, and 4-azaleucine, in that order, inhibited the multiplication of blue-green but not other algal groups. The purine and pyrimidine analogs were not inhibitory. In chemically defined axenic media, dl -ethionine was about 100 times more inhibitory to the blue-green algae Synechococcus cedrorum and Anabaena cylindrica than to the eucaryotic algae Ochromonas danica and Euglena gracilis. The ciliate Tetrahymena pyriformis was at least 100-fold more resistant to ethionine than the algae. The unusual sensitivity of blue-green algae to ethionine and other amino acid analogs represents an exceptional phylelic character and may be useful in the control of these algae when they become a nuisance. Amino acid analogs such as ethionine may also serve to remove blue-green algae from cultures one may desire to render axenic.     

Theca cell monolayers that inhibit maturation of bovine oocytes show differences in their protein secretion pattern

A previous study reported that coculturing bovine cumulus-oocyte complexes (COCs) with theca cell monolayers maintained oocytes in meiotic arrest. The present study evaluated whether the protein secretion pattern in this system is different between theca cells and granulosa cells and whether the presence of COCs influences their pattern of secretion. Follicular cells were isolated from 2- to 5-mm follicles and cultured in TCM-199 supplemented with 10% fetal calf serum. Theca cell monolayers maintained COCs but not denuded oocytes (DOs) in meiotic arrest. Monolayers were incubated for 6 hr in medium supplemented with radioactive L-[³⁵S]methionine. The patterns of protein secreted in the medium were analyzed by electrophoresis SDS-PAGE 10%. These results showed that theca cell monolayers secreted two major proteins. This pattern was different from the protein pattern secreted by granulosa cell monolayers. The molecular weights of these proteins were estimated to be 214 and 190 kDa. Coculturing COCs with theca cell monolayers during the labeling revealed that COCs modulated the secretion of theca cell monolayers. When theca cells were grown on collagen-coated wells, the monolayers did not maintain the oocytes at the germinal vesicle (GV) stage. The secretion of the 214-kDa protein also decreased. Then, when theca cell monolayers are effective to maintain oocytes in meiotic arrest, the cells especially secreted the 214-kDa protein. In conclusion, the 214-kDa protein secreted by theca cell monolayers may play a role in the process of maintaining oocytes in meiotic arrest. Mol. Reprod. Dev. 50:200–206, 1998. © 1998 Wiley-Liss, Inc.