Embryonic Motility Research Articles

The emergence of patterned movement during late embryogenesis of Drosophila

Larval behavioral patterns arise in a gradual fashion during late embryogenesis as the innervation of the somatic musculature and connectivity within the central nervous system develops. In this paper, we describe in a quantitative manner the maturation of behavioral patterns. Early movements are locally restricted "twitches" of the body wall, involving single segments or parts of segments. These twitches occur at a low frequency and have low amplitude, reflecting weak muscle contractions. Towards later stages twitches increase in frequency and amplitude and become integrated into coordinated movements of multiple segments. Most noticeable among these is the peristaltic wave of longitudinal segmental contractions by which the larva moves forward or backward. Besides becoming more complex as development proceeds, embryonic movements also acquire a pronounced rhythm. Thus, late embryonic movements occur in bursts, with phases of frequent movement separated by phases of no movement at all; early movements show no such periodicity. These data will serve as a baseline for future studies that address the function of embryonic lethal genes controlling neuronal connectivity and larval behavior. We have analyzed behavioral abnormalities in two embryonic lethal mutations with severe neural defects, tailless (tll), which lacks the protocerebrum, and glial cells missing (gcm), in which glial cells are absent. Our results reveal prominent alterations in embryonic motility for both of these mutations, indicating that the protocerebrum and glial cells play a crucial role in the neural mechanism controlling larval movement in Drosophila.

Embryonic development ofPelteobagrus fulvidraco (Richardson, 1846)

For production enhancement and procedure upgrade, the developmental phases of laboratory-reared eggs of catfishPelteobagrus fulvidraco were investigated. Twenty mature females and 10 males were collected from Dadongmen wholesale fisheries market in Wuhan City on May 8, 2003. Zygotes were stripped from mature fish after hormone-induced ovulation, fertilized, and incubated through whole embryonic development. The fertilized eggs were stocked in density of 100 eggs/L in white square tanks of 10 L. Incubation water was dechlorinated tap water with continuous aeration. The tanks were lit directly with 60 W fluorescent bulbs with a 12 light: 12 dark photoperiod. Water temperature, dissolved oxygen and pH were 29.0±0.5°C, 6.7±0.4 mg/L and 7.4±2, respectively. The results showed that the eggs ofP. fulvidraco were yellow, sticky and contained much yolk. The mean diameter of fertilized eggs was 2.03 mm. At the water temperature of 29.0±0.5°C, the ontogenesis spent about 33 h after fertilization. From fertilization to hatching, the embryonic development can be divided into 30–40 phases, which varies in the emphasis and direction of development. The detailed embryonic movement was also described.

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Basic developmental rules and their implications for epilepsy in the immature brain

ABSTRACT The construction of the human brain with its 10 15 synapses follows a set of complex developmentally and environmentally regulated steps. A series of sequences have been described that are instrumental, in the sense that a failure of any one of them leads to dramatic, life‐long consequences. Hence the importance of determining the sequential maturation of neurons, synapses and cortical maps. It is also important to determine how network‐driven events become installed, as neuronal activity intervenes in all of these steps and modulates, for better or worse, the outcome. A fundamental consequence of these sequential events is that any disruption will have very different consequences depending on when it occurs, indeed, “when is as important as what”. An obvious aspect of these general features is related to seizures. In fact, the developing brain has both a higher incidence of seizures in human and animal models, and experiences seizures that can produce long‐lasting consequences that are also stage‐dependent. This seminar and the series of slides presented are an introduction to these issues, summing up several studies made notably by INMED researchers during the last two decades ( http://www.inmednet.com ). It concentrates on four basic developmental rules: i) the generation by very immature neurons, of very large currents mediated by the activation of receptors in neurons that bear no synapses. This is due to the release of GABA that diffuses to distal sites and acts as a paracrine factor; ii) the excitatory/inhibitory shift of the actions of GABA during development because of a progressive reduction in the intracellular chloride concentration; iii) the sequential formation of GABAergic synapses and networks before glutamatergic ones, implying that, at an early stage, all the excitatory drive will be GABAergic; iv) the presence, at an early stage, of a unique, primitive pattern in all developing structures, this pattern disappears when most GABAergic synapses have shifted to their adult configuration‐ .Several consequences of these sequences are described including: i) a control of neuronal migration by GABA‐acting drugs, and the possibility that migration disorders are also generated by environmental factors that include the effects of GABA‐acting agents; ii) If GABA excites immature neurons and inhibits adult one, then GABA‐acting agents will also produce different effects on the mother and the embryo; iii) early brain oscillations are generated by the periphery and propagate centrally – notably to the sensory‐motor cortex, suggesting that peripherally‐generated movements may provide an important signal for the formation of cortical maps, in keeping with the importance of embryonic movements; iv) “seizures beget seizures” in the developing brain. This has now been shown in a triple chamber with the two intact hippocampi that we developed, and with which it has been possible to show that only recurrent seizures that include high frequency oscillations can transform the naïve, contralateral hippocampus to an epileptic one that seizes spontaneously. Most interestingly, at an early developmental stage, when GABA excites many neurons and the density of glutamatergic synapses is not sufficiently high, purely glutamatergic seizures cannot lead to long‐term consequences, the additional excitatory drive provided by GABAergic synapses is needed. In other words, at that stage, blocking GABA synapses generates seizures, as in adults, but these do not lead to long‐term consequences. The mechanisms that underlie these differences is due to the need for high frequency oscillations (> 80 Hz or so), and these can only be generated when GABA synapses are operative in the developing brain: GABA receptor antagonists are ictogenic, but not epileptogenic. To facilitate teaching purposes the paper is published together with supplemental data (as a PowerPoint presentation included in the accompanying DVD), thus allowing an overview of important developmental steps and their implications. [ Published with supplemental data on CD ]

Effects of different incubation temperatures on the yolk‐feeding stage of Eupallasella percnurus(Pallas)

Embryos and yolk‐feeding larvae of lake minnow Eupallasella percnurus were reared at 13, 16, 19, 22 and 25° C with no access to external food. Time from egg activation to first embryonic movements, hatching, filling of swimbladder and final yolk resorption increased with decreasing temperature. At 13° C, c. 40% of larvae were unable to fill their swimbladder. The predicted lower temperature at which development and growth ceased (biological zero, t0) was the same for both processes, c. 7·5–10·5° C. There was no ontogenetic shift in the t0 value. Temperature coefficients for development (Q10dev.) ranged from 2 to 3 at 19–25° C, but were higher in hatched larvae at lower temperatures. Eggs of E. percnurus had a combination of small size, high hydration and low caloric value of fresh matter. Dry mass of larval tissue on yolk, percentage of dry matter in wet matter, and specific growth rate were maximized at 22 and 25° C. At 19–25° C, energy and matter contained in the initial eggs were converted to body tissue most efficiently. Temperatures from 22 to 25° C are considered optimal for E. percnurus embryos and yolk‐feeding larvae and are recommended for their indoor rearing.

Early effects of embryonic movement: 'a shot out of the dark'.

It has long been appreciated that studying the embryonic chick in ovo provides a variety of advantages, including the potential to control the embryo's environment and its movement independently of maternal influences. This allowed early workers to identify movement as a pivotal factor in the development of the locomotor apparatus. With an increasing focus on the earliest detectable movements, we have exploited this system by developing novel models and schemes to examine the influence of defined periods of movement during musculoskeletal development. Utilizing drugs with known neuromuscular actions to provoke hyperactivity (4-aminopyridine, AP) and either rigid (decamethonium bromide, DMB) or flaccid (pancuronium bromide, PB) paralysis, we have examined the role of movement in joint, osteochondral and muscle development. Our initial studies focusing on the joint showed that AP-induced hyperactivity had little, if any, effect on the timing or scope of joint cavity elaboration, suggesting that endogenous activity levels provide sufficient stimulus, and additional mobilization is without effect. By contrast, imposition of either rigid or flaccid paralysis prior to cavity formation completely blocked this process and, with time, produced fusion of cartilaginous elements and formation of continuous single cartilaginous rods across locations where joints would ordinarily form. The effect of these distinct forms of paralysis differed, however, when treatment was initiated after formation of an overt cavity; rigid, but not flaccid, paralysis partly conserved precavitated joints. This observation suggests that 'static' loading derived from 'spastic' rigidity can act to preserve joint cavities. Another facet of these studies was the observation that DMB-induced rigid paralysis produces a uniform and specific pattern of limb deformity whereas PB generated a diverse range of fixed positional deformities. Both also reduced limb growth, with different developmental periods preferentially modifying specific osteochondral components. Changes in cartilage and bone growth induced by 3-day periods of flaccid immobilization, imposed at distinct developmental phases, provides support for a diminution in cartilage elaboration at an early phase and for a relatively delayed influence of movement on osteogenesis, invoking critical periods during which the developing skeleton becomes receptive to the impact of movement. Immobilization also exerts differential impact along the proximo-distal axis of the limb. Finally, our preliminary results support the possibility that embryonic hyperactivity influences the potential for postnatal muscle growth.

Oncopeltus fasciatus zen is essential for serosal tissue function in katatrepsis

Unlike most Hox cluster genes, with their canonical role in anterior–posterior patterning of the embryo, the Hox3 orthologue of insects has diverged. Here, we investigate the zen orthologue in Oncopeltus fasciatus (Hemiptera:Heteroptera). As in other insects, the Of-zen gene is expressed extraembryonically, and RNA interference (RNAi) experiments demonstrate that it is functionally required in this domain for the proper occurrence of katatrepsis, the phase of embryonic movements by which the embryo emerges from the yolk and adjusts its orientation within the egg. After RNAi knockdown of Of-zen, katatrepsis does not occur, causing embryos to complete development inside out. However, not all aspects of expression and function are conserved compared to grasshopper, beetle, and fly orthologues. Of-zen is not expressed in the extraembryonic tissue until relatively late, suggesting it is not involved in tissue specification. Within the extraembryonic domain, Of-zen is expressed in the outer serosal membrane, but unlike orthologues, it is not detectable in the inner extraembryonic membrane, the amnion. Thus, the role of zen in the interaction of serosa, amnion, and embryo may differ between species. Of-zen is also expressed in the blastoderm, although this early expression shows no apparent correlation with defects seen by RNAi knockdown.

Promoting Epiboly

Although steroids have numerous functions--regulating electrolyte balance; carbohydrate, protein, and fat metabolism; the inflammatory response; and the development and maintenance of secondary sex characteristics--their role during early embryogenesis has been unclear. Hsu et al. used in situ hybridization to investigate the expression of the genes encoding Cyp11a1 (which converts cholesterol to pregnenolone, thereby initiating steroid biosynthesis) and Hsd3b (which converts pregnenolone into progesterone) during zebrafish embryogenesis. Shortly after fertilization, mRNA for both enzymes was present in the cytoplasm of blastomeres located above the yolk. However, during epiboly (a process during which the blastoderm envelops the yolk), cyp11a1 and hsd3b disappeared from the blastomeres and appeared in the yolk syncytial layer, which is composed of yolk syncytial nuclei and located at the junction between the blastomeres and the yolk. Injection of antisense morpholino oligonucleotides to cyp11a1 delayed epiboly without affecting cell fate. Whereas coinjection of cyp11a1 mRNA or incubation with pregnenolone partially rescued the delay in epiboly, progesterone worsened it. Yolk microtubules extend from yolk syncytial nuclei to drive epiboly, and yolk microtubules from embryos treated with morpholinos were more sensitive to disruption by nocodazole than those from wild-type embryos, an effect that was reversed by pregnenolone. Pregnenolone also increased the abundance of polymerized tubulin in taxol-treated extracts from morpholino-treated embryos. Moreover, fluorescently labeled pregnenolone colocalized with polymerized tubulin from morpholino-treated embryo extracts. Thus, pregnenolone appears to stabilize microtubules and thereby promote cell migration during epiboly, a role that likely involves binding to microtubules rather than association with nuclear receptors. H.-J. Hsu, M.-R. Liang, C.-T. Chen, B.-c. Chung, Pregnenolone stabilizes microtubules and promotes zebrafish embryonic cell movement. Nature 439 , 480-483 (2006). [PubMed]

Acetylcholine and calcium signalling regulates muscle fibre formation in the zebrafish embryo

Nerve activity is known to be an important regulator of muscle phenotype in the adult, but its contribution to muscle development during embryogenesis remains unresolved. We used the zebrafish embryo and in vivo imaging approaches to address the role of activity-generated signals, acetylcholine and intracellular calcium, in vertebrate slow muscle development. We show that acetylcholine drives initial muscle contraction and embryonic movement via release of intracellular calcium from ryanodine receptors. Inhibition of this activity-dependent pathway at the level of the acetylcholine receptor or ryanodine receptor did not disrupt slow fibre number, elongation or migration but affected myofibril organisation. In mutants lacking functional acetylcholine receptors myofibre length increased and sarcomere length decreased significantly. We propose that calcium is acting via the cytoskeleton to regulate myofibril organisation. Within a myofibre, sarcomere length and number are the key parameters regulating force generation; hence our findings imply a critical role for nerve-mediated calcium signals in the formation of physiologically functional muscle units during development.

Diminished embryonic movements of developing embryo by direct exposure of sidestream whole smoke solutions

Embryonic movements (EM) are considered to be the first sign of life and cigarette smoking during pregnancy has been linked to affect EM. Exposure to sidestream smoke, produced from the emissions of a smoldering cigarette, may result in poor pregnancy outcome and increased risk of serious perinatal morbidity and mortality. In this study, the chicken embryo bioassay was used to systematically assess the effects of short-term exposure to sidestream whole smoke solutions (SSWSS) on EM, recorded in real time by a video camera for 60 min and each EM was counted for every 3-min interval. Application of different types of SSWSS to the embryos caused significant changes in all types of EM from 15 to 18 min of recording time. Extensive reduction (P<0.001) and some time complete stoppage of swing-like movements and whole-body movements were observed in almost all treated embryos. Our data clearly link between exposure of SSWSS and substantial decrease in EM. It is unclear whether nicotine and/or other ingredients present in sidestream smoke are responsible for these alterations in EM. This article provides an outline of the relevance of SSWSS on EM for evolutionary developmental biology and this assay can be used to investigate the complex mixtures with regard to their effects on EM.

The internationalization of fire protection: in pursuit of municipal networks in Edwardian Birmingham

Through a case study of Birmingham fire brigade, this article examines the plethora of international networking activities undertaken during the late Victorian and early Edwardian period. Birmingham fire brigade, under the control of Alfred Tozer, led British municipal participation in early international fire networks, attending international congresses and exhibitions in Berlin and London, and also visiting continental cities to inspect fire brigades and their appliances. Locating the study firmly within historical debates concerning the embryonic international municipal movement, this article demonstrates that municipal institutions participated in networking activities as part of a policy learning and knowledge-transfer process.

Toxicological Effects of Mainstream Whole Smoke Solutions on Embryonic Movements of the Developing Embryo

Cigarette smoking is unrivaled among developmental toxicants in terms of total adverse impact on the human population. Maternal tobacco use during pregnancy adversely affects prenatal and postnatal growth and increases the risk of behavioral and developmental defects in children and adolescents. In the current study, the effects of different preparations of nicotine and mainstream whole smoke solutions (MSWSS) on embryonic movements during neonatal development were examined in vivo, using the chicken embryo model, recorded in real-time by a video camera. It was observed that low doses of nicotine induced hyperactivity and higher doses induced hypoactivity. Accordingly, a significant (p < 0.01) decrease in movements was observed by application of 10 µg of nicotine and different preparations of MSWSS. A dose-dependent decrease in embryonic movements was observed, which did not recover by the end of experiment. It was concluded that nicotine could alter embryonic movements, which are important during embryogenesis for differentiation and maturation of the body systems.

Toxicological Effects of Mainstream Whole Smoke Solutions on Embryonic Movements of the Developing Embryo

Cigarette smoking is unrivaled among developmental toxicants in terms of total adverse impact on the human population. Maternal tobacco use during pregnancy adversely affects prenatal and postnatal growth and increases the risk of behavioral and developmental defects in children and adolescents. In the current study, the effects of different preparations of nicotine and mainstream whole smoke solutions (MSWSS) on embryonic movements during neonatal development were examined in vivo, using the chicken embryo model, recorded in real-time by a video camera. It was observed that low doses of nicotine induced hyperactivity and higher doses induced hypoactivity. Accordingly, a significant (p < 0.01) decrease in movements was observed by application of 10 µg of nicotine and different preparations of MSWSS. A dose-dependent decrease in embryonic movements was observed, which did not recover by the end of experiment. It was concluded that nicotine could alter embryonic movements, which are important during embryogenesis for differentiation and maturation of the body systems.

Catecholaminergic and cholinergic regulation of swimming motility development in free embryos ofCichlasoma Nigrofasciatum

The divergence of progeny from the same spawners of Cichlasoma nigrofasciatum into two groups by duration of embryogenesis and the level of motor activity was studied close to the end of the embryonic period. Free embryos were also studied. During the study, eggs were treated with agents, modifying the activity of catecholaminergic and cholinergic systems. 3-Hydroxytyramine and L-Tyrosine were found to exert a weak influence on embryonic motility. After hatching, these substances modify swimming performance of free embryos, approximating movements of fish at later stages. 6-Hydroxydopamine and, still more, alpha-Bungarotoxin, decrease embryonic motility and postpone the hatching. The influence of these substances on the development of embryo motility increases during early ontogenesis, as indicated by decreased concentration of the substance, necessary for adequate reaction. Neither L-Tyrosine nor 6-Hydroxydopamine influenced the divergence of the progeny into two groups. Injection of the perivitelline fluid with high concentration of hatching enzyme from pre-hatching embryos into the perivitelline space of earlier embryos was found to induce the appearance of rotation movements, typical for more advanced embryos. Changes of correlation between the miogenic and neurogenic motor activity during early development of fish are discussed.

Patterns in Early Embryonic Motility: Effects of Size and Environmental Temperature on Vertical Velocities of Sinking and Swimming Echinoid Blastulae

Early embryonic swimming is widespread among marine invertebrates, but quantitative information about swimming behaviors is scarce. Swimming may affect encounters with predators, positioning in the water column, and nutrient absorption. Measured rates and patterns of swimming and sinking for blastulae of four eastern Pacific echinoid species show that sinking speeds equal or exceed swimming speeds. Swimming speed scaled negatively with embryo size, though sinking speed did not scale with size. Analysis of swimming paths of Strongylocentrotus franciscanus revealed a temperature dependency in swimming pattern that affected speed of upward movement. Sinking speeds were significantly greater at 10 degrees C than at 14 degrees C for blastulae of all four species examined. In Dendraster excentricus, killing the blastulae annulled this temperature effect, indicating an active density regulation by these embryos. Finally, measurements of particle velocities around sinking and swimming D. excentricus blastulae show that swimming creates a more localized disturbance than sinking. Embryonic swimming may therefore decrease rather than increase encounters with pelagic predators. Results from subsequent experiments in which embryos were reared in low-oxygen environments suggest that any oxygen-absorption advantages of swimming have little, if any, effect on the development of D. excentricus embryos.

Hatching Events in the California Grunion, Leuresthes tenuis

This study provides the first detailed description of the events at hatching for an anamniotic vertebrate egg. California Grunion are ideally suited for this study, because these fish eggs incubate fully terrestrially and delay hatching until presented with an environmental trigger. Grunion eggs can be induced to hatch on demand within a few minutes, by mechanical agitation in seawater. We suggest that the process of hatching in grunion involves an enzymatic weakening of the chorion followed by the embryo's active efforts to escape. Using a dissecting microscope, microscope camera, and digital video camera, we compared embryonic movements within the chorion prior to hatching. Then, we recorded the exact moment of hatching with digital video to identify events that occur consistently. Prior to exposure to the hatching trigger, Grunion embryos move minimally within the egg. After the hatching trigger, embryos significantly increase the amount and types of movements within the egg. The chorion distorts in a location superior to the caudal region, and fluid escapes from the egg. Embryonic activity usually culminates in a vigorous tail lash that splits the chorion open. Ultimately, usually within two minutes, the Grunion larva emerges from the chorion into its new aquatic environment.

Defining boundaries during joint cavity formation: going out on a limb.

Whilst factors controlling the site at which joints form within the developing limb are recognised, the mechanisms by which articular element separation occurs during the formation of the joint cavity have not been determined. Herein, we review the relationships between early limb patterning, embryonic movement, extracellular matrix composition, local signalling events and the process of joint cavity formation. We speculate that a pivotal event in this process involves the demarcation of signalling boundaries, established by local mechano-dependent modifications in glycosaminoglycan synthesis. In our opinion, studies that examine early patterning and also focus on local developmental alterations in tissue architecture are required in order to help elucidate the fundamental principals regulating joint formation.

Steps during the development of the zebrafish locomotor network

This review summarizes recent data from our lab concerning the development of motor activities in the developing zebrafish. The zebrafish is a leading model for studies of vertebrate development because one can obtain a large number of transparent, externally and rapidly developing embryos with motor behaviors that are easy to assess (e.g. for mutagenic screens). The emergence of embryonic motility was studied behaviorally and at the cellular level. The embryonic behaviors appear sequentially and include an early, transient period of spontaneous, alternating tail coilings, followed by responses to touch, and swimming. Patch clamp recording in vivo revealed that an electrically coupled network of a subset of spinal neurons generates spontaneous tail coiling, whereas a chemical (glutamatergic and glycinergic) synaptic drive underlies touch responses and swimming and requires input from the hindbrain. Swimming becomes sustained in larvae once serotonergic neuromodulatory effects are integrated. We end with a brief overview of the genetic tools available for the study of the molecular determinants implicated in locomotor network development in the zebrafish. Combining genetic, behavioral and cellular experimental approaches will advance our understanding of the general principles of locomotor network assembly and function.

Embryonic motility: environmental influences and evolutionary innovation.

Embryos do not passively await hatching from their eggs or amnionic containments but begin active movement very early on in their development. The first muscle contractions in the chick embryo start on the third day of incubation and subsequently assume a characteristic pattern of increasing and decreasing motor activity (Fig. 1) (Hamburger 1963; Bekoff 1981, 1992). It is long known that embryonic motility represents an important epigenetic component of development. In vertebrates, active movement of the embryo is required for the correct development of cartilage, bone, and joints; of muscles, tendons and ligaments; and of connectivities in the central nervous system. The intrauterine disturbance of embryonic movements leads to severe malformations and functional disorders, such as pathological motor patterns and neurological deficits that are retained throughout adult life (Bos et al. 2001). It is equally known that embryonic activity depends on environmental conditions, both chemical and physical, and that changes of environmental parameters can strongly affect the motility patterns. However, the effect of such environmental influences on development, via the alteration of embryonic activity, is little explored. Even less is known about the possible evolutionary consequences of this kind of environment–development interaction. This article provides an outline of the relevance of environment dependent embryonic activity for evolutionary developmental biology.