Setogenesis and molting in planktonic crustaceans

The formation of new setae, termed setogenesis, is describedfor two taxa of planktonic crustaceans: euphausiids, Euphausiapacifica and Thysanoessa spinifera, and a calanoid copepod,Calanus marshallae. Characteristics of setal formation and eversionat ecdysis are described from two time-series of animals preservedat known intervals in their molt cycle. Results from these laboratoryreference series indicate that previous interpretations of setogenesisin the literature can by synthesized to describe a dynamic processof setal formation which is common to all crustacean taxa. The morphological characteristics of developing setae are usedto designate three specific phases in the molt cycle of planktoniccrustaceans (premolt, postmolt, and intermolt). This stagingtechnique may be used to study field-oriented problems relatedto molting in small planktonic crustaceans.     

Synaptic plasticity and humoral modulation of neuromuscular transmission in the lobster claw opener during the molt cycle

1. Neuromuscular properties of the lobster dactyl opener were studied at different stages of the molt cycle.2. Excitatory junctional potentials (EJPs) were found to be significantly larger in hard-shelled premolt lobsters than in soft-shelled postmolt animals.3. Inhibitory junctional potentials (IJPs) were larger in postmolt claw preparations than in premolt ones.4. Excitatory transmission was also monitored during superfusion with plasma samples obtained from lobsters in vartious stages of the molt cycle (postmolt = AB; intennolt = C; premolt = D₂).5. D₂ plasma increased EJP amplitude by an average of 28% relative to baseline levels (in saline), while AB plasma reduced EJP size by an average of 6%.6. Premolt and postmolt plasma produced opposite effects on quantal content (m); D₂; plasma increased m by a mean of 39%, whereas AB plasma caused a mean reduction by 10%, which suggests that humoral factors may act presynaptically to alter transmitter release.7. These results provide evidence of neuromuscular plasticity in the opener muscle during the molt cycle, and are consistent with changes in claw-opening behavior seen during this cycle.     

Silicification of the medial tooth in the blue crab Callinectes sapidus

Observations of cuticular structures mineralized with silica within the Crustacea have been limited to the opal teeth of copepods, mandibles of amphipods, and recently the teeth of the gastric mill in the blue crab Callinectes sapidus. Copepod teeth are deposited during premolt, with sequential elaboration of organic materials followed by secretion of silica into the tooth mold. The timing of mineralization is in stark contrast to that of the general integument of crustaceans in which calcification is completely restricted to the postmolt period. To determine the timing of molt‐related deposition and silicification of the teeth of the gastric mill, the medial tooth of the blue crab C. sapidus was examined histologically and ultrastructurally across the molt cycle. Histological data revealed deposition of the organic matrix of the epicuticle and exocuticle during premolt. No evidence of postmolt changes in the thickness of the epicuticle and exocuticle, or any deposition of endocuticle, was observed. Scanning electron microscopy revealed degradation of the outer surface of the old tooth during premolt. During premolt, epithelial structures resembling papilla appeared to secrete a fibrous web that coalesces to become the matrix of the new tooth. Semi‐quantitative elemental analyses indicated simultaneous deposition of silica and organic matrix, and demonstrated a homogeneous distribution of silicon throughout the epicuticle of the tooth at all stages. However, there is evidence of deposition (presumably silicification) during postmolt as spaces between the papillae become filled in. Thus, the pattern and timing of deposition and silicification of the tooth are different from both teeth of copepods and the general exoskeleton of decapods, and may facilitate rapid resumption of feeding and consumption of the exuvia in early postmolt. J. Morphol. 277:1648–1660, 2016. © 2016 Wiley Periodicals, Inc.     

Influence of molt cycle and beta-ecdysone on protein synthesis in the chelicerate arthropod, Limulus polyphemus.

1. The effect of molt cycle stage and beta-ecdysone on protein synthesis in the horsehoe crab, Limulus polyphemus, was examined. 2. A pronounced decline in protein specific radioactivity after incubation with 14C-leucine was noted in muscle, midgut gland and operculum from postmolt to intermolt to premolt and in gut and gill tissue from intermolt to premolt. 3. beta-Ecdysone injections produced an early stimulation of protein synthesis in the midgut gland followed by strong inhibition within 48 hr. 4. Results are compared with those obtained in mandibulate arthropods.     

Ion Regulation in Crayfish: Freshwater Adaptations and the Problem of Molting

SYNOPSIS. Crayfish have a long evolutionary history in temperatefresh water (FW). Ion regulation is challenged by low externalconcentrations of Na, Cl, and Ca (<1 mM). In intermolt theprimary concern is Na and Cl balance; around ecdysis the emphasisswitches to Ca regulation as the cuticle is decalcified/calcified.Compared with marine crustaceans, intermolt crayfish maintaina reduced extracellular (EC) osmolality and have lower permeabilityto both ions and water. Hyperregulation involves active branchialuptake of Na and Cl and the unique ability to produce a hypotonicurine. Ion uptake involves apical electroneutral ion exchange(Na^$ for H^$; Cl^– for HCO₃–; counterions providedfrom CO₂ via carbonic anhydrase) followed by active basolateraltransport of Na via the Na pump, with Cl following passively.Reabsorption of 95% of filtered electrolytes at the antennalgland (kidney) involves similar subcellular mechanisms in amorphologically differentiated region of the distal tubule.Intermolt crayfish exhibit negative Ca balance (passive effluxunopposed by uptake) tolerable in view of the large cuticularCaCO₃ reserve. In premolt, cuticular Ca is reabsorbed. A smallamount is stored as gastroliths, the remainder is lost via branchialexcretion and in the discarded exuviae. At ecdysis, FW uptakegenerates the physical force for shedding, leaving the crayfishwith dilute hemolymph and a Ca deficiency. Levels of EC Na andCl are restored by intensive postmolt branchial uptake. Mineralizationof the soft exoskeleton involves remobilization of stored Caand branchial uptake of Ca and HCO₃. Transepithelial Ca transportinvolves Ca^2$ ATPase and Ca^2$/Na^$ exchange. The importance ofexternal electrolytes and pH in postmolt ion regulation is explored,as are some allometric considerations.     

Commitment of Mitotic Cells to Meiosis during the G2 Phase of Premeiosis

In the developing anther, archesporial cells that proliferateby mitotic division are converted into meiotic cells duringthe premeiotic interphase. Experiments with explanted microsporocytesof Lilium and Trillium were made to obtain evidence for theconversion of mitotic to meiotic cells during the premeioticperiod. Explanted premeiotic cells were cultured through thedivision cycle at relatively high division frequencies and showeda variety of division types with respect to chromosomal events.The type of division depended on the premeiotic stage at whichthe cells were explanted. Cells in the G₁, S and early G² phasesunderwent mitotic division and formed a diad or binucleate monad.Cells explanted at the late G₂ phase were cultured throughoutthe normal meiotic cycle, which resulted in typical tetrad configuration. In microsporocytes explanted during the main part of the G₂interval, centromere behavior was meiotic, but chromosome pairingand chiasma formation were disturbed. Thus, she G₂ intervalwas shown to be critical for the commitment of mitotic cellsto meiotic division. Detailed analysis showed that the intracellularchanges that commit the cells to meiosis begin shortly aftercompletion of premeiotic DNA synthesis and that these changesare progressive and cumulative. (Received February 2, 1982; Accepted May 24, 1982)     

Hormonal Effects on Calcium Metabolism in Crustacea

Cyclic shifts of calcium in the exoskeleton and soft tissues,as they are related to the intermolt cycle in crayfish, arereviewed. Regulatory factors, derived from the eyestalk, influencelevels of exoskeletal calcium; eyestalk extracts prepared fromanimals in premolt decrease shell calcium, while reciprocallyextracts from animals in intermolt increase it when these hormonalsources are injected into animals in the premolt stage (D₀-D₄). In addition, premolt eyestalk extract results in an increasein gastrolith calcium. In the exchange of calcium between theanimal and its environment there is evidence for differentialdepositionof recently available calcium in the exoskeleton. Further, intermoltand early premolt animals maintained in Ca⁴⁵-labelled waterfor 15 days concentrate it 4 and 3—fold in the exoskeletonand stomach, respectively. However, removal of a molt-inhibitingfactor through ablation of eyestalks results in a 20 and 40—foldincrease in incorporation inthese same sites relative to environmentalconcentrations. Treatment with mammalian parathyroid extract mobilizes bothexoskeletal and gastric calciumand leads to a rise in bloodcalcium. However, there is little or no effect on levels ofexoskeletal citric acid. Further, citric acid is higher in thecrayfish carapace during stage C, the period of mineralization,than in stage D, the period of demineralization. There are both similarities and differences between the effectsof crustacean and mammalianregulating factors with respect tothe direction and extent of mineralization. Biochemical studiesshould elucidate the mechanisms regulated by these hormones.     

Gap junctions and intercellular communication in the hepatopancreas of the crayfish (Orconectes propinquus) during molt

Summary The crustacean hepatopancreas is a major metabolic center intimately involved in molting and vitellogenesis. Cells of the hepatopancreas exhibit one of the richest endowments of gap junctions known and are thus presumed to be linked for intercellular communication. In order to monitor hepatopancreatic activity during the molt cycle of crayfish (Orconectes propinquus), the electrical coupling between cells of the hepatopancreatic tubules was measured during postmolt, intermolt and premolt. Samples of hepatopancreas from each of these stages were fixed and freeze-fractured to correlate morphologic features of gap junctions with electrophysiological data. Analysis of the data revealed that ionic coupling was greater in postmolt and premolt tubule cells than in cells of intermolt animals. Platinum replicas of hepatopancreatocyte plasmalemmata revealed that in postmolt, gap junction plaques were smaller and more numerous than those in intermolt and premolt; however, the total area of gap junction plaques per unit membrane area analyzed was approximately the same for hepatopancreatocytes from all molt stages. Although the hepatopancreatic gap junctions exhibited no quantitative differences, those from post- and premolt animals were rounded with tightly packed particles, while plaques from intermolt animals were generally pleomorphic with loosely packed particles. Results of this study suggest that cells of the crayfish hepatopancreas are more coupled in pre- and postmolt, with macular plaques of tightly packed particles, perhaps as a response to the increased metabolic demands of molt, and less well coupled, with irregular plaques of loosely packed junctional particles, during intermolt. The only recognizable morphological correlates of increased cell coupling were tight packing of junctional particles into rounded plaques, while decreased coupling corresponded to junctions with loosely packed irregular aggregates of particles.Supported by the Natural Sciences and Engineering Research Council of Canada (RRS)     

Effects of Seasonal Variation in Radiation and Temperature on Net Assimilation and Growth Rates in an Arid Climate

The net assimilation rate (E_A), relative growth-rate (R_w), andleaf-area ratio (F_A) were measured for rape (Brassica napus),sunflower (Hetianthus annuus), and maize (Zea mays) at varioustimes of year in an arid climate, using young plants grown widelyspaced on nutrient culture. Multiple regression analysis accountedfor 90–95 per cent of the variation in E_A and R_W in termsof two climatic variables: mean temperature and radiation receipt. E_A rose linearly with radiation in all three species; increasein E_A with temperature was greatest in maize and least (notsignificant) in rape. R_Wrose with radiation and temperature,the latter being the more important variable especially in coolweather; a temperature optimum was shown at 24° C in rape.F_A rose with increase in temperature or decrease in radiation;its variation was due to change in leaf area/leaf weight ratherthan in leaf weight/plant weight. Multiple regression analyses can lead to faulty interpretationif the independent variables are correlated (as are climaticvariables in nature), but conclusions can be checked by controlled-environmentstudies in which climatic factors are not correlated. The presentconclusions are supported by such studies. The regression equations, coupled with average weather records,indicate seasonal cycles of growth parameters. E_A is maximalnear midsummer and minimal near midwinter, following the radiationcycle. Maxima and minima in R_W are about a month later, becauseR_W is affected by the temperature cycle and this lags behindthe radiation cycle. F_A is maximal in autumn and minimal inspring. E_A is highest where radiation receipts near 750 cal cm^–2day^–1 coincide with high temperatures. This combinationoccurs only in clear midsummer weather at low latitudes, andis maintained over long periods only in arid regions. The fact that E_A rose linearly with radiation suggests thatleaf water deficits arising under high radiation had littleeffect on E_A and that saturating levels of light were very high.     

Molecular mechanism of molt-inhibiting hormone (MIH) induced suppression of ecdysteroidogenesis in the Y-organ of mud crab: Scylla serrata

The present study was focused on the regulation of ecdysteroidogenesis in the Y-organ of Scylla serrata during molting cycle. A strong expression of molt-inhibiting hormone (MIH) and phosphorylation of ERK was predominantly observed in the postmolt and intermolt stages of Y-organs, whereas protein kinase C, steroidogenic acute regulatory protein (StAR) and cytochrome P450(scc) activity were exclusively seen in the premolt stages. Interestingly, inhibition of ERK phosphorylation by PD98059 in the early postmolt (A), middle postmolt (B) and intermolt (C) stages resulted in the prominent expression of PKC and StAR in the postmolt stages. This result indicates that phosphorylation of ERK is required for suppression of ecdysteroid biosynthesis with the involvement of protein kinase C, and StAR protein.     

中华绒螯蟹蜕皮过程中体壁结构和主要成分的变化

采用组织化学和原子吸收分析等方法, 研究了中华绒螯蟹蜕皮过程中体壁结构和主要成分的变化。结果显示: 中华绒螯蟹体壁分为上表皮、外表皮、内表皮和膜层, 糖类物质各层均有分布, 胶原纤维分布在除上表皮外的其他各层。在蜕皮前, 糖类、胶原纤维都被重吸收, 体壁上表皮和外表皮在蜕皮前形成, 内表皮和膜层在蜕皮后形成。体壁粗蛋白含量在蜕皮前期(D1-D3-4期)降低(P0.05), 蜕皮后A-B期含量极高(P0.05)。几丁质含量在蜕皮过程中变化不显著(P0.05), 只是在蜕皮前稍有上升。Ca2+和Mg2+含量在蜕皮前D1期显著低于蜕皮间期和蜕皮前其他时期(P0.05), 而蜕皮后A-B期降到最低(P0.05), 蜕下的甲壳中则含有较多的Ca2+和Mg2+ (P0.05)。Cu2+和Zn2+含量除蜕皮后A-B期升高外(P0.05), 其余时期变化不明显(P0.05)。这些研究结果表明, 中华绒螯蟹体壁结构和成分变化与蜕皮周期密切相关。       

Effects of Gibberellic Acid on the Activation, Synthesis, and Release of Acid Phosphatase in Barley Seed

Acid phosphatase activity was present in unimbibed barley seed,but rose during incubation of embryoless half-seeds and isolatedaleurone layers, and was further increased by 10^–6 M gibberellicacid (GA₃). Release of total acid phosphatase activity fromhalf-seeds and aleurone layers was markedly enhanced by GA₃.Inhibitor studies with cycloheximide and actinomycin D suggestedthat de novo synthesis of acid phosphatase occurred followingimbibition. Gel nitration, electrophoresis, and [¹⁴C]leucineincorporation studies revealed that a single molecular formof acid phosphatase was present in dry seed, whereas on incubationtwo further forms arose. A proportion of the three molecularforms of the enzyme was synthesized de novo. Gibberellic acidstimulated activation, but not de novo synthesis, of all threemolecular forms of acid phosphatase. Although a small amountof one of the molecular forms was secreted in the absence ofGA₃, the presence of gibberellin greatly increased secretionof the same form of acid phosphatase.     

Reciprocal changes in calcification of the gastrolith and cuticle during the molt cycle of the red claw crayfish Cherax quadricarinatus

Mobilization of calcium during the molt cycle from the cuticle to transient calcium deposits is widely spread in crustaceans. The dynamics of calcium transport to transient calcium deposits called gastroliths and to the cuticle over the course of the molt cycle were studied in the crayfish Cherax quadricarinatus. In this species, calcium was deposited in the gastroliths during premolt and transported back to the cuticle during postmolt, shown by digital X-ray radiograph analysis. The predominant mineral in the crayfish is amorphous calcium carbonate embedded in an organic matrix composed mainly of chitin. Scanning electron micrographs of the cuticle during premolt showed that the endocuticle and parts of the exocuticle were the source of most of the labile calcium, while the epicuticle did not undergo degradation and remained mineralized throughout the molt cycle. The gastroliths are made of concentric layers of amorphous calcium carbonate intercalated between chitinous lamella. Measurements of pH and calcium levels during gastrolith deposition showed that calcium concentrations in the gastroliths, stomach, and muscle were about the same (10 to 11 mmol l(-1)). On the other hand, pH varied greatly, from 8.7+/-0.15 in the gastrolith cavity through 7.6+/-0.2 in muscle to 6.9+/-0.5 in the stomach.     

Aspects of Lipid Metabolism in Crustaceans

Lipid is the predominant organic reserve of many crustaceansand is important in the metabolism of many of these animals.Ingested lipid is digested by gastric lipase and apparentlyabsorbed into depot-lipid as rß-monoglycerides. Thevariation in the content and composition of the depot-lipidis a function of both the external environment and internalcontrol systems. Evidence suggests that lipids from marine organismscontain more long-chain polyunsaturated fatty acids than doesthe lipid of fresh water organisms which in turn have a highproportion of C₁₆ and C₁₈ fatty acids. The fatty-acid compositionof the sub-tropical land crab,Gecarcinns lateralis, resemblesthat of the fresh-water crustaceans. In addition, our studiesindicate that aspects of lipid metabolism may be under endocrinecontrol. The induction of premolt by destalking markedly increasesthe synthesis of lipid from metabolic precursors and its subsequentincorporation into the depot-lipid of the hepatopancreas. Inthe late premolt stages there is a decrease in the lipid contentof the hepatopancreas. This occurs as the lipid is mobilizedfrom the hepatopancreas to meet the energy demands of all thoseprocesses resulting in ecdysis. This sinusoidal variation inthe lipid metabolism of the hepatopancreas is influenced byan eyestalk factor (s).     

Limitation to Photosynthesis in Water-stressed Leaves: Stomata vs. Metabolism and the Role of ATP

Decreasing relative water content (RWC) of leaves progressivelydecreases stomatal conductance (g_s), slowing CO₂ assimilation(A) which eventually stops, after which CO₂ is evolved. In somestudies, photosynthetic potential (A_pot), measured under saturatingCO₂, is unaffected by a small loss of RWC but becomes progressivelymore inhibited, and less stimulated by elevated CO₂, below athreshold RWC (Type 1 response). In other studies, A_pot andthe stimulation of A by elevated CO₂ decreases progressivelyas RWC falls (Type 2 response). Decreased A_pot is caused byimpaired metabolism. Consequently, as RWC declines, the relativelimitation of A by g_s decreases, and metabolic limitation increases.Causes of decreased A_pot are considered. Limitation of ribulosebisphosphate (RuBP) synthesis is the likely cause of decreasedA_pot at low RWC, not inhibition or loss of photosynthetic carbonreduction cycle enzymes, including RuBP carboxylase/oxygenase(Rubisco). Limitation of RuBP synthesis is probably caused byinhibition of ATP synthesis, due to progressive inactivationor loss of Coupling Factor resulting from increasing ionic (Mg²⁺)concentration, not to reduced capacity for electron or protontransport, or inadequate trans-thylakoid proton gradient (pH).Inhibition of A_pot by accumulation of assimilates or inadequateinorganic phosphate is not considered significant. DecreasedATP content and imbalance with reductant status affect cellmetabolism substantially: possible consequences are discussedwith reference to accumulation of amino acids and alterationsin protein complement under water stress.     

Digestive proteinases of red shrimp Pleoticus muelleri (Decapoda, Penaeoidea): partial characterization and relationship with molting

The present study describes the activity and some characteristics of proteinases in the hepatopancreas of red shrimp Pleoticus muelleri during the different stages of the molting cycle. Proteolytic activity was highest between pH 7.5 and 8. The hepatopancreatic protein content in the premolt stage was higher than in the other stages of the molting cycle (P<0.05). No significant differences were found in total proteolytic activity in the hepatopancreas when comparing molting stages. The proteolytic activity of the P. muelleri hepatopancreas enzyme preparations is the main responsibility of serine proteinases. TLCK, a trypsin inhibitor, reduced azocasein hydrolysis between 26% (intermolt) and 37% (premolt). TPCK, a chymotrypsin inhibitor, did not decrease hydrolytic activity, except for in postmolt. Low trypsin and chymotrypsin activities were found during intermolt, and increased in postmolt. The electrophoretogram of the enzyme extracts shows 12 bands of activity during intermolt (from 16.6 to 53.1 kDa). Some fractions were not detected in the postmolt and premolt stages. Three low molecular weight trypsin forms (17.4, 19.1 and 20 kDa) were found in all molting stages. One band of chymotrypsin (21.9 kDa) was observed in all molting stages. High molecular mass active bands (66-205 kDa) could not be characterized with inhibitors. Comparison of the protease-specific activity of the hepatopancreas of some species indicated a relationship between digestive enzyme activity and feeding habits of the shrimp. Omnivorous shrimp, such as Penaeus vannamei (syn: Litopenaeus vannamei) and Penaeus monodon, showed higher protease activity than the carnivorous shrimp, Penaeus californiensis (syn: Farfantepenaeus californiensis) and P. muelleri. In fact, the enzymatic activity in the hepatopancreas of P. muelleri showed variations in relation to feeding habit and molting cycle.