Pre-hatching Stage Research Articles

Size relations and yolk utilization in embryonated ova and alevins of Atlantic salmon Salmo salar L. in various combinations of temperature and dissolved oxygen.

Relations among weight (W), surface (S), and volume (V) for freshly fertilized water-hardened Atlantic salmon ova from 5 to 7 mm in diameter are: S = log V2/3, S = 1.21 log W, and V = W/1.2.Fresh weights remained relatively constant from fertilization to the establishment of blood vessels in the yolk sac. Subsequently, there was a rapid increase to 110%, followed by a gradual decline to 105% shortly before hatching. Large blastodiscs weighed 1.2 mg and embryonal weight increased very gradually until late prehatching stages, when growth was accelerated rapidly. Embryos, at hatching, after continuous incubation at 5 and 10°C and three levels of dissolved oxygen, varied from 22.1 mg (5 °C, 100% air saturation) to 11.9 mg (5 °C. 30%) and From 14.5 mg (10 °C. 100%) to 10.0 mg (10 °C, 30%).Cumulative and mean daily yolk depletion declined with lowered oxygen supply, but they were considerably greater at 10 °C than at 5 °C. Gross conversion ratios (including uptake of exogenous materials) indicate that yolk is used more efficiently at 5 °C, and 100% air saturation. Alevins, kept in their original environments, became more efficient than they were in their encapsulated state, and, at 10 °C. 100% air saturation, they overtook those at 5 °C before the completion of yolk absorption. Those in lower oxygen supplies continued to lag, at both temperatures.

DNA, RNA, Protein, and Free Amino Acids During Ontogenesis of Rainbow Trout (Salmo gairdneri)

During the first 2 wk of incubation, the pattern of change in DNA concentration tended to precede similar changes in RNA concentration. Unfertilized eggs contained 23.2 and 77.2 μg of DNA and RNA, respectively. DNA concentration rose from 155.8 μg/fry (after 23 days of incubation) to 326.6 μg/larva (14 days after hatching), while RNA increased from 238.8 to 693.3 μg during the same period. Protein per ontogenetic unit was relatively constant during the prehatching stages but dropped markedly just prior to hatching. Of the protein in the unfertilized egg, approximately 20% was retained in the fry at hatching and approximately 50% was retained in the larva following yolk absorption. Total free amino acid concentrations declined just after fertilization, indicating the initiation of protein synthesis or a decline in yolk protein degradation. These concentrations increased to near initial values in the blastula and were nearly doubled in the yolk plus fry at hatching.

A "NORMAL" DEVELOPMENT OF DENUDED EGGS OF THE STARFISH, ASTERINA PECTINIFERA.

Experimental conditions that allow "normal" development of starfish eggs stripped of the fertilization membrane are reported in this paper. Four kinds of intercellular relation are distinguished during the pre-hatching stages of these eggs. Cells from 2- to 8-cell stages are hardly related to each other, while those from 16- to 128-cell stages are bound loosely together. After the 8th division (about 5.5 hr after insemination at 21°C) cells adhere closely and cooperate with each other to perform morphogenetic movement of "blastulation". This relation is taken over by that of a true multicellular system at about 10 hr after insemination. Closely after this, the function of cilia carries the embryo away from the substratum.

Histogenesis of the islets of Langerhans in insulin-treated chick embryos.

Chick embryos were exposed to a teratogenic dose of insulin at the 6-day incubation stage and the effect of this treatment on the histogenesis of the islets of Langerhans was studied throughout the incubation period. The exogenous insulin caused an initial delay in the differentiation of the beta islets and in the elaboration of the endogenous insulin. During the final prehatching stages, the endogenous insulin appeared to be elaborated more rapidly than in the control tissues. The insulin treatment caused an increased activity of the alpha cells and an increased production of glucagon. Histochemically, the ribonucleic acid content of both alpha and beta cells was reduced, especially during the early stages of development. Also, there was an initial increase in glycogen storage and alkaline phosphatase activity within the alpha cells. The beta islets in the insulin-treated tissues, except for the initial delay in their development, showed no significant differences in the glycogen and alkaline phosphatase patterns from the control.

A histological and histochemical study of the development of the pineal gland in the chick, Gallus domesticus.

The pineal gland of the chick is composed of vesicles during much of the incubation period. During incubation, these vesicles increase in size and number and reach maximum development at the 17-day prehatching stage. After day 17, the vesicular tissue decreases and, by 1 day posthatching, only a few vesicles remain. All vesicles have disappeared by 1 month posthatching and the gland has acquired a compact, lobed character. The vesicular cells are secretory in nature and react positively with tests for glycogen, glycoproteins, ribonucleic acid, acid mucopolysaccharides, and neutral lipids. In addition, the specialized apical border of these cells stains intensely for acid lipids. The vesicles contain a secretory product which includes a sulfated mucopolysaccharide component. Based on the intensities of the staining reactions, this secretion is present in greatest amounts during the 15- to 17-day prehatching period. No secretory product could be demonstrated after 1 day posthatching. It appears, therefore, that the development of the vesicular tissue is related to some secretory role and that this role is culminated during the 15- to 17-day prehatching period. After day 17, the decrease and eventual disappearance of the vesicles apparently marks the conclusion of this secretory phase for the gland.

A histochemical study of the development of the mantle-edge and shell in the freshwater gastropod, Helisoma duryi eudiscus (Pilsbry)

Glycogen and ribonucleic acid are present in the mantle-edge during the prehatching period and in the adult. The cubocolumnar epithelium contains the largest stores of glycogen. Ribonucleic acid is most abundant in the mantle-edge gland, and the mucous and mucoprotein cells. Mucopolysaccharides and glycoproteins occur within the mantle-edge epithelium, excepting the mantle-edge gland, and within the shell ground substance. Mucous glands and the sheath surrounding the organic plates are rich in sulfated mucopolysaccharides. Alkaline phosphatase and calcium could not be demonstrated during the prehatching stages. In the adult, alkaline phosphatase reactions are intense along the distal border of the cubocolumnar epithelium, and the basal borders of the epithelia of the mantle-edge gland, median lobe, and ventral lobe. Calcium carbonate occurs as spherules in the connective tissue, in the extrapallial fluid, and within the organic plates and crystalline layers of the shell. In the adult, lipids are most plentiful in the dorsal lobe epithelium and yellow body cells. Vitamin A occurs only within the cubocolumnar and yellow body cells. Cytochrome oxidase is present within the mantle-edge epithelium and, in terms of relative amounts, reflects the activity of the various lobes. Similarly, the size of the Golgi element can be correlated with the activity of the mantle-edge epithelia.

A HISTOLOGICAL STUDY OF THE DEVELOPMENT OF THE MANTLE-EDGE AND SHELL IN THE FRESHWATER GASTROPOD, HELISOMA DURYI EUDISCUS (PILSBRY)

The mantle-edge originates as a peripheral thickening of the evaginated shell gland. During the prehatching stages, it differentiates to form a dorsal lobe including the mantle-edge gland, a median lobe, and a ventral lobe possessing numerous mucous glands. The connective tissue core contains yellow body cells, amoebocytes, and mucoprotein glands. The protoconch is composed of fibers embedded in an amorphous ground substance. Initially it is secreted by the entire mantle-edge but, as development advances, its secretion is progressively limited to the mantle-edge gland. The adult shell is composed of the periostracum, crystalline layer, and an inner layer. The periostracum is a highly tanned layer of fibers embedded in a ground substance. The crystalline layer consists of calcium crystals, the smaller of which contain a nucleus. The inner layer is composed of organic plates which possess amoebocyte nuclei, yellow bodies and calcium granules, and which are surrounded by a thick envelope containing sulfated mucopolysaccharides. These calcium crystals and organic plates are embedded in a ground substance. It is believed that the mantle-edge gland produces the periostracum, that the dorsal lobe contributes to the inner two shell layers, and that the mucous and mucoprotein glands, yellow body cells, and amoebocytes play a significant role in the deposition and calcification of the shell.

Experimental Hybridization of Pissodes strobi and P. approximatus (Coleoptera: Curculionidae)

Reciprocal matings of Pissodes strobi (Peck) collected from white-pine terminals and Pissodes approximatus Hopkins collected from red pine logs were made in the laboratory. No absolute reproductive isolating mechanisms were found. But there was greater mortality in the pre-hatch stages of the hybrids. Many anatomical features were measured to provide a basis for the identification of hybrids among natural populations.

Acid mucopolysaccharides in the development of the Pacific great skate, Raja binoculata

ABSTRACT Histochemical treatments specific for hyaluronic acid, C-S-A/C, C-S-B and heparin were applied to a series of Pacific great skate (Raja binoculata) embryos from cleavage to immediate prehatching stages, in order to characterize histochemically the AMPS present in the embryos and to study the events leading to AMPS localization in the adult. A progression was observed: (a) intracellular neutral polysaccharides in cleavage stages, (b) a combination of extracellular polysaccharides and weakly acidic AMPS (hyaluronic acid) associated with cell processes in neurulating stages, (c) extracellular strongly acidic AMPS (chondroitin sulphates) in later stages, particularly in areas of cartilage development. In neurulating embryos hyaluronic acid appeared in quantity between adjacent tissues, suggestive of some developmental significance for this compound. Hyaluronic acid also appeared after neurulation in close association with developing gut and mesonephros. Histochemical localization of these compounds in immediate prehatching stages agreed with previously reported biochemical analyses of shark skin and cartilage, i.e. C-S-B occurred in skin and C-S-A/C was a major component of cartilage matrix.

Biology of the Ringneck Snake, Diadophis punctatus, in Florida

Two geographic color varieties of the ringnack snake, Diadophis punctatus punctatus (Linnaeus), occur in Florida. Snakes in the Peninsula are characterized by reddish subcaudal surfaces and by the curious trait of tail-coiling when disturbed; the snakes occupying extreme northern Florida have yellow subcaudal surfaces and do not tail-coil. In the intergrade zone the subcaudals vary from yellow through red and some yellow-tailed individuals will tail-coil. The color red and the associated tail-coil display are considered relict characters in Gulf Coastal ringneck snakes. Ways are enumerated whereby tail-coiling may serve an aposematic function. The almost universal occurrence of the color red in snakes that have tail-flash displays suggests the behavior may have been developed against birds; most other potential predators lack red vision. Diadophis punctatus has been found in practically all the major terrestrial habitats in Florida, although most commonly in low, wet, pine flatwoods. Main factors determining the species' ecological distribution and population density seem to be food and degree of moisture in the microhabitat. Significant size differences in snakes of two different populations may be related to availability of food. Microhabitat associates in two different habitats are considered, and known and supposed coactions discussed. Other snakes are probably the most important predators and competitors for food and space. The sex ratio in adults, at least, is 1:1. The gonads are bilaterally asymmetrical in position and usually in size. The testes are smallest in winter and spring. Spermatogenesis is initiated in the spring and cellular proliferation is greatest in summer. Spermatozoa released from the testis in late summer and fall are stored in the ductus deferens throughout most of the year. The largest ovarian eggs increased from approximately 3 mm in April to a near maximum of 24 mm in July when they were in the oviducts. Eggs are probably laid from late May or June into August; females probably lay each year; the time of mating is not known. Both sexes are thought to be sexually mature by their second spring. The male secondary sex characters-anal ridges and greatly enlarged kidney tubules-become apparent about when the spermatozoa are formed. A series of prehatching stages is briefly described. Males carry the hemipenes in tbe functional (everted) position until at least 28 days before hatching. Ventral markings are not apparent until after the dorsum becomes lightly pigmented. The unpigmented dorsal scale pits are the last structures to become discernible.

THE ROLE OF CREATINE AND ITS PHOSPHATE IN AMPHIBIAN DEVELOPMENT

1. A study has been made of the role of phosphorylated creatine in the energy metabolism of Rana pipiens and gastrula-blocked R. pipiens ♀ X R. sylvatica ♂ hybrid embryos.2. It is shown that the longest period of anaerobiosis that can be survived by R. pipiens embryos of all pre-hatching stages, with subsequent normal aerobic development, is of two hours duration, at 24° C.3. In hybrid embryos, creatine synthesis is deficient after stage 16, and the levels of phosphocreatine phosphorus and inorganic phosphorus are subnormal.4. In both types of embryos, two hours of anaerobiosis result in a decrease of phosphocreatine phosphorus, accomnpanied by a quantitatively similar increase of free inorganic phosphorus. Both changes are greater in normal embryos than in hybrids. In R. pipiens embryos, prolonged anaerobiosis (12 hours) results in the appearance of excess inorganic phosphorus, perhaps at the expense of ATP.5. A comparison is made of energy metabolism in muscle and embryos, and the role of energy metabolism in the development of hybrids is briefly discussed.

Walleye Egg Survival during Incubation on Several Types of Bottom in Lake Winnibigoshish, Minnesota, and Connecting Waters

Survival of naturally spawned walleye (Stizostedion vitreum vitreum) eggs to pre-hatching stage was determined on five bottom areas of Lake Winnibigoshish, Minnesota, and connecting waters over 4 years. These bottom types included soft muck, sand, gravel, rubble, and boulders. Egg sampling was done at intervals throughout the incubation period and estimates of egg abundance made shortly after the completion of spawning and at or near the eyed-egg stage. Initial fertility of walleye eggs was high (96-100 percent), but the percentage of live eggs declined steadily during incubation. During the eyed-egg stage, however, there was an apparent increase in the percentage survival because of disappearance of dead eggs. Egg survival was best on gravel-rubble, both as percentage survival and numbers of eggs surviving the eyed stage. Survival of eggs on improved gravel-rubble bottom was as high as 35.7 percent but on muck bottom was as low as 0.6 percent. Survival on better bottoms of gravel and rubble averaged 25 percent. Abundance of eggs on sand bottom to which gravel and rubble had been added was more than 10 times that observed previously, and survival of eggs on the improved bottom increased fivefold. Length of incubation period, which was determined by water temperature, was a factor in egg survival. Walleyes selected gravel bottom for spawning when it was available, and most eggs were deposited in water from 12 to 30 inches deep. Low water levels in the spring of 1958 made much former spawning area unavailable and may have been a factor in a weak 1958 year class.