Beat Direction Research Articles

Branchial Ventilation in the Burrowing Crab,Atelecyclus Rotundatus

Atelecyclusburies itself by digging backwards into the sand, sometimes until only the antennae protrude above the sand. In this position it is difficult for the animal to maintain an adequate respiratory current through the branchial chambers by pumping water in the normal direction.Studies of the ventilatory behaviour ofAtelecyclushave shown that, when on the surface of the sand, water flows through the branchial chambers as in most other crabs i.e. water enters the chambers at the base of the limbs and the exhalant water leaves anteriorly. This direction of flow is also maintained when the crabs are only partially buried but when more deeply buried, the direction of water flow is reversed by the reversal of the direction of beat of the scaphognathites and water is drawn down the antennal tube and leaves the branchial chambers at the base of the limbs.InactiveAtelecyclusshow a well-defined rhythm of pumping in which regular periods of active pumping alternate with periods during which the scaphognathites stop beating and ventilation of the branchial chambers ceases. The rhythm of ventilation is regular but the duration of the ventilatory pauses (apnoea) may vary considerably between animals.During pausing, the activity of the scaphognathites and of the heart are closely synchronized. The cessation of scaphognathite activity is accompanied by a period of cardiac arrest, although single heart beats may sometimes occur during a pause.As in many decapods, the Po of the post-branchial blood was high (Pa, o = 92 torr, Pv, O = 21 torr) but during a pause P, o gradually declined.

Ca2+-dependent regulation of beat frequency of cilia in Paramecium.

Triton-extracted models of Paramecium cells prepared in the presence of Mg2+ and EGTA showed Ca2+ sensitivity not only in the direction of beat but also in the beat frequency of cilia. The beat frequency increased over a range of concentration of Ca2+ from 2 X 10(-7) to 4 X 10(-7) M, in which the model swam forwards. The frequency increased also above 10(-6)M-Ca2+, in which the model swam backwards. The increase in frequency was inhibited by calmodulin antagonists. Intracellular injection of a Ca2+ buffer giving a free Ca2+ concentration of 2 X 10(-7) to 5 X 10(-7) M in intact cells induced an increase in the beat frequency. Therefore, it is very likely that the beat frequency of cilia is regulated by the intracellular concentration of Ca2+.

Biochemical studies of the excitable membrane of paramecium tetraurelia. IX. Antibodies against ciliary membrane proteins.

The excitable ciliary membrane of Paramecium regulates the direction of the ciliary beat, and thereby the swimming behavior of this organism. One approach to the problem of identifying the molecular components of the excitable membrane is to use antibodies as probes of function. We produced rabbit antisera against isolated ciliary membranes and against partially purified immobilization antigens derived from three serotypes (A, B, and H), and used these antisera as reagents to explore the role of specific membrane proteins in the immobilization reaction and in behavior. The immobilization characteristics and serotype cross-reactivities of the antisera were examined. We identified the antigens recognized by these sera using immunodiffusion and immunoprecipitation with 35S-labeled ciliary membranes. The major antigen recognized in homologous combinations of antigen-antiserum is the immobilization antigen (i-antigen), approximately 250,000 mol wt. Several secondary antigens, including a family of polypeptides of 42,000-45,000 mol wt, are common to the membranes of serotypes A, B, and H, and antibodies against these secondary antigens can apparently immobilize cells. This characterization of antiserum specificity has provided the basis for our studies on the effects of the antibodies on electrophysiological properties of cells and electron microscopic localization studies, which are reported in the accompanying paper. We have also used these antibodies to study the mechanism of cell immobilization by antibodies against the i-antigen. Monovalent fragments (Fab) against purified i-antigens bound to, but did not immobilize, living cells. Subsequent addition of goat anti-Fab antibodies caused immediate immobilization, presumably by cross-linking Fab fragments already bound to the surface. We conclude that antigen-antibody interaction per se is not sufficient for immobilization, and that antibody bivalency, which allows antigen cross-linking, is essential.

Negative Phototaxis in Blepharisma japonicum

ABSTRACTThe protozoan Blepharisma japonicum showed negative phototaxis caused by transient reversal of the direction of ciliary beat and changes of swimming velocity induced with varying intensities of light. The ciliary reversal occurred at 1–2 sec after a sudden increase in light intensity. When light intensity was decreased, no response was observed. Moreover, the ciliates swam fast in light areas but slowly in dark areas; the mean velocity of swimming was 80 μ m/sec at 5 × 102 lux but reached about 400 μMm/sec at 5 × 103 lux. In addition, the cell body elongated in response to light application; the mean length of the body was 308 μm at 5 × 102 lux, which increased to 397 μ m at 104 lux. Such body elongation seems to contribute to rapid swimming. Negative phototaxis may be an important behavior in B. japonicum because the organisms are killed by exposure to strong light.

Development of the larval nervous system of the sand dollar, Dendraster excentricus.

Transformation of the gastrula to the pluteus includes development of the ability of the larva to control the direction of ciliary beat and coordinate activities of the ciliary band with activities of the esophageal muscles (48-60 h, 15 degrees C). Glyoxylic acid-induced fluorescence shows several cells of the animal plate to contain catecholamines in the 36-h gastrula. As the ectoderm thickens to form the ciliary band (36 48 h), the catecholamine-containing cells increase in number and occur dispersed throughout the band. Tissues with the ultrastructural characteristics of nerves first became apparent associated with the ciliary band in 60-h larvae. The coincident development of coordinated behaviour and the appearance of cells with ultrastructural and histochemical characteristics of nerves suggests that the larval nervous system is derived at least in part from cells of the animal plate and develops in association with the ciliary bands.

Bending patterns of chlamydomonas flagella I. Wild-type bending patterns.

Using a uniflagellate mutant of Chlamydomonas and flash photomicrography at 300 Hz, we have obtained detailed information on the forward and reverse beating modes of Chlamydomonas flagella and on the relationship between rotation of the uniflagellate cell and the bending cycle of the forward mode. Flagella ranging in length from 5 to 15.5 micron were photographed. There is a decrease in wavelength and an increase in curvature in the principal bends when the length of the flagellum is less than the normal length of 12-13 micron, but these changes are not sufficient to maintain similarity of the bending pattern. In the reverse mode, the flagellum propagates symmetrical, planar, undulatory waves with a shear amplitude which is the same as in the forward mode; there is a 19% increase in beat frequency and a similar decrease in wave length. The reorientation of the flagellar beat direction towards the axis of the cell in the reverse mode is caused both by the decrease in asymmetry of beat and by activation of sliding in the principal bends at an earlier time in the beat cycle, relative to the time of activation of sliding in reverse bends. There are additional rare modes of beating which may be related to intermediate stages in the transition between forward and reverse beating modes.

The settlement and metamorphosis of the marine bryozoan Bowerbankia gracilis (Ctenostomata: Vesicularioidea)

The settlement and metamorphosis of the marine bryozoan Bowerbankia gracilis has been examined by light and electron microscopy. The period of rapid morphogenesis consists of the following sequence of morphogenetic movements: 1) eversion of the internal sac, 2) retraction of the apical disc, 3) coronal involution and exposure of the pallial epithelium, and 4) closure of the internal coronal cavity. The eversion of the internal sac at the onset of metamorphosis coincides with a sudden reversal of the direction of beat of the coronal cilia. The reversed beating of the coronal cilia wafts the adhesive secreted by the internal sac over the metamorphosing larva, forming the pellicle. The internal sac is subsequently internalized and histolyzed with the corona and the other transitory larval tissues, and the extensive pallial epithelium forms the epidermis of the ancestrular body wall (cystid). Type I mesenchyme cells form an incomplete somatic mesothelium beneath the differentiating cystid epidermis, and Type II mesenchyme cells become mobile phagocytes. The main body cavity develops by the histolytic enlargement of the internal cavity formed during coronal involution. The apical disc degenerates and the polypide develops from rudiments in the oral hemisphere of the larva. The distinctive larval morphology and metamorphosis of vesicularioid ctenostomes are compared with other bryozoans, and possible evolutionary trends are considered.

Mechanisms of rapid morphogenetic movements in the metamorphosis of the bryozoan Bugula neritina (cheilostomata, cellularioidea). I. Attachment to the substratum.

The larval morphology, settlement behavior, and the rapid morphogenetic movements that occur during the first 60 sec of metamorphosis of the cellularioid cheilostome bryozoan Bugula neritina have been examined and analyzed by light and electron microscopy. The larva attaches to the substratum at the onset of metamorphosis by the eversion of the internal sac. At the same time, the coronal cilia reverse their direction of beat, spreading an adhesive secreted by the neck region of the everting sac over the metamorphosing larva. During attachment, the larva goes through several configurations that coincide with the sequential contraction and relaxation of certain larval muscles. Histological and ultrastructural evidence indicates that the neck and wall regions of the internal sac are everted by the contraction of the muscles in the equatorial plane of the larva at the same time that the roof region in pulled toward the larval equator by the contraction of the axial muscles. The subsequent relaxation of the axial muscles allows the roof region to be everted by the antagonistic force generated by the sustained contraction of the equatorial musculature. After the roof region attaches to the substratum, the apical disc is temporarily retracted by a second contraction of the axial muscles. The apical disc subsequently reextends as the axial muscles relax just before coronal involution. A comparison of the ontogenetic sequence of rapid morphogenetic movements in the metamorphoses of cheilostome and ctenostome bryozoans indicates that cellularioid cheilostomes have undergone peramorphosis in the aspect of development.

Failure of cilia of reprogram following segmental ampullary reversal of the rabbit oviduct.

Cilia exhibited unidirectional and coordinated movement within microsurgically reversed segments of rabbit ampulla when examined up to 13 months after surgery. The direction of ciliary beating was opposite that of the remainder of the oviduct.

Ciliary reversal without rotation of axonemal structures in ctenophore comb plates.

We have used a newly discovered reversal response of ctenophore comb plates to investigate the structural mechanisms controlling the direction of ciliary bending. High K+ concentrations cause cydippid larvae of the ctenophore Pleurobrachia to swim backward. High-speed cine films of backward-swimming animals show a 180 degree reversal in beat direction of the comb plates. Ion substitution and blocking experiments with artificial seawaters demonstrate that ciliary reversal is a Ca++-dependent response. Comb plate cilia possess unique morphological markers for numbering specific outer-doublet microtubules and identifying the sidedness of the central pair. Comb plates of forward- and backward-swimming ctenophores were frozen in different stages of the beat cycle by an "instantaneous fixation" method. Analysis of transverse and longitudinal sections of instantaneously fixed cilia showed that the assembly of outer doublets does not twist during ciliary reversal. This directly confirms the existence of radial switching mechanism regulating the sequence of active sliding on opposite sides of the axoneme. We also found that the axis of the central pair always remains perpendicular to the plane of bending; more importantly, the ultrastructural marker showed that the central pair does not rotate during a 180 degree reversal in beat direction. Thus, the orientation of the central pair does not control the direction of ciliary bending (i.e., the pattern of active sliding around the axoneme). We discuss the validity of this finding for three-dimensional as well as two-dimensional ciliary beat cycles and conclude that models of central-pair function based on correlative data alone must now be re-examined in light of these new findings on causal relations.

Pregnancy, Ciliary Transport, and the Reversed Ampullary Segment of the Rabbit Fallopian Tube

Pregnancy following reversal of an ampullary segment in the rabbit oviduct has occurred only sporadically, and then usually after repeated mating. To explain this phenomenon it has been postulated that the direction of ciliary beat in the reversed ampullary segment must revert to the normal pro-uterine direction. To test this hypothesis a segment of the ampulla was reversed by microsurgery in 10 rabbits. These were then bred repeatedly for as long as 28 weeks. A single pregnancy, at 10 weeks, ensued. Observation of ampullary transport in vivo showed that muscular activity in the reversed ampullary segment may occasionally overcome the adverse, pro-ovarian ciliary currents that were found to persist in subsequent in vitro examinations. Scanning electron microscopy confirmed normal ciliation in the reversed segment. These observations may explain the occasional transport of an ovum across the reversed ampullary segment and an isolated pregnancy in the mated rabbit.

Calcium control of waveform in isolated flagellar axonemes of Chlamydomonas.

The effect of Ca(++) on the waveform of reactivated, isolated axonemes of chlamydomonas flagella was investigated. Flagella were detached and isolated by the dibucaine procedure and demembranated by treatment with the detergent Nonidet; the resulting axomenes lack the flagellar membrane and basal bodies. In Ca(++)-buffered reactivation solutions containing 10(-6) M or less free Ca(++), the axonemes beat with a highly asymmetrical, predominantly planar waveform that closely resembled that of in situ flagella of forward swimming cells. In solutions containing 10(-4) M Ca(++), the axonemes beat with a symmetrical waveform that was very similar to that of in situ flagella during backward swimming. In 10(-5) M Ca(++), the axonemes were predominantly quiescent, a state that appears to be closely associated with changes in axomenal waveform or direction of beat in many organisms. Experiments in which the concentrations of free Ca(++), not CaATP(--) complex were independently varied suggested that free Ca(++), not CaATP(--), was responsible for the observed changes. Analysis of the flagellar ATPases associated with the isolated axonemes and the nonidet- soluble membrane-matrix fraction obtained during preparation of the axonemes showed that the axonemes lacked the 3.0S Ca(++)-activated ATPase, almost all of which was recovered in the membrane-matrix fraction. These results indicate that free Ca(++) binds directly to an axonemal component to alter flagellar waveform, and that neither the 3.0S CaATPase nor the basal bodies are directly involved in this change.

Role of the Cilia Membrane in Control of Microtubule Sliding

It is currently believed that ciliary beating results from microtubule sliding which is restricted in regions to cause bending. Cilia beat can be modified to bring about changes in beat frequency, cessation of beat and reversal in beat direction. In ciliated protozoans these modifications which determine swimming behavior have been shown to be related to intracellular (intraciliary) Ca2+concentrations. The Ca2+levels are in turn governed by the surface ciliary membrane which exhibits increased Ca2+conductance (permeability) in response to depolarization. Mutants with altered behaviors have been isolated. Pawn mutants fail to exhibit reversal of the effective stroke of ciliary beat and therefore cannot swim backward. They lack the increased inward Ca2+current in response to depolarizing stimuli. Both normal and pawn Paramecium made leaky to Ca2+by Triton extrac¬tion of the surface membrane exhibit backward swimming only in reactivating solutions containing greater than IO-6M Ca2+Thus in pawns the ciliary reversal mechanism itself is left operational and only the control mechanism at the membrane is affected. The topographic location of voltage-dependent Ca2+channels has been identified as a component of the ciliary mem¬brane since the inward Ca2+conductance response is eliminated by deciliation and the return of the response occurs during cilia regeneration. Since the ciliary membrane has been impli¬cated in the control of Ca2+levels in the cilium and therefore is the site of at least one kind of control of microtubule sliding, we have focused our attention on understanding the structure and function of the membrane.

Pathologic correlates of electronystagmographic tracings

Electronystagmographic tracings were correlated with the histopathologic findings in the peripheral vestibular system of 66 temporal bones of 34 patients who complained of dizziness. Although some findings (e.g., spontaneous nystagmus, unilaterally reduced caloric response, and directional preponderance) seemed to have doubtful value in localizing vestibular system disease, other findings indicated a peripheral site of the vestibular lesions. Bilaterally reduced or bilaterally absent caloric responses were consistently associated with pathologic changes in the peripheral vestibular system. Similarly, a unilaterally absent caloric response consistently indicated peripheral disease. On the other hand, positional nystagmus and bilaterally equal caloric responses seemed to indicate the absence of a peripheral site of disease in the vestibular system. Although there is no reliable way to differentiate end organ from vestibular nerve lesions, the latter were more commonly associated with positional nystagmus and absence of a caloric response. The direction of nystagmus beating correlated with the side of the diseased peripheral site in two of four patients with spontaneous nystagmus, in five of seven with positional nystagmus, and in all with unilateral reductions in or absence of caloric responses. Electronystagmographic findings in the present study seemed to reflect diffuse rather than localized pathologic changes in the peripheral vestibular system.

Directional change of ciliary beat effected with Mg2+ in Paramecium.

Two types of Triton X-100 extracted model of Paramecium caudatum were prepared and effects of the ionic compositions of the reactivating medium on the swimming behaviour of the extracted models were examined. Type I, prepared by the method of Naitoh and Keneko, did not change swimming direction depending on the concentrations of Mg2+ and K+ in the reactivating medium. Type II, prepared by a different method, however, reversed the swimming direction depending on the concentrations of Mg2+ and K+ in the reactivating medium, even when the concentration of Ca2+ was kept at an extremely low level (below 10-8M). Decreasing the concentration of Mb2+ induced the type II model to swim backwards. Intracellular injection of EDTA into a live paramecium cell induced reversal of the direction of ciliary beat. The concentration of Mg2+ at which the reversal of swimming direction occurred varied with the concentration of adenosine triphosphate (ATP) in a manner which suggested that the concentration of free ATP4- might be the critical determinant of swimming direction.

The role of cilia in fertility: An evaluation by selective microsurgical modification of the rabbit oviduct

Fertility, ovum transport, and ciliary activity were examined in 35 rabbits after either unilateral microsurgical reversal or double transection (surgical controls) of 1 cm. segments of distal, middle, or proximal ampulla. Fertility was also examined following similar segmental reversal at the level of the distal and proximal isthmus in eight additional rabbits. Reversal of 1 cm. segments of distal, middle, or proximal ampulla effectively prevented pregnancy. In contrast, control animals that underwent double transection without reversal experienced unaltered fertility. Reversal of 1 cm. segments of distal or proximal isthmus was also consistent with normal fertility. Ova placed on the fimbrial surface of oviducts that had undergone ampullary segmental reversal were arrested at the distal anastomosis of the reversed segment. When mechanically displaced into the reversed segment, such ova were transported toward the ovary and were again arrested at the distal anastomosis. Transampullary transport of ova was normal in doubly transected controls. Ova were transported across both anastomosis sites to the ampullary isthmic junction. Studies of ciliary activity confirmed that cilia within reversed segments of ampulla beat in the ovarian direction, counter to that in the rest of the oviduct. Direction of ciliary beat remained unaltered in surgical controls. These experiments demonstrate a critical role of tubal cilia in ovum transport through the rabbit ampulla. In contrast, other mechanisms control ovum transport into and through the tubal isthmus.

Physiological Factors Influencing Ovulation, Fertilization, Early Embryonic Development and Establishment of Pregnancy in Pigs

Key reproductive processes in the female pig have been reviewed from the stage of pre-ovulatory resumption of meiosis until the commencement of implantation and prolongation of the lifespan of the corpora lutea. The time of ovulation is noted to be regulated by a specific surge of luteinizing hormone detectable in systemic blood at the onset of oestrus. Neither the time of ovulation nor the number of follicles ovulating is considered capable of significant modification by the respective procedures of double mating or nutritional 'flushing' imposed after the onset of oestrus. Sperm transport is assessed largely under conditions of natural mating, where regulation of the numbers of spermatozoa reaching the site of fertilization is seen as a vital function performed by the isthmus, in conjunction with its role as a reservoir for viable spermatozoa. Transport of male and female gametes in opposing directions in the Fallopian tubes is attributed in part to the direction of ciliary beat and progression of peristaltic contractions in the ampulla and isthmus. The relatively short fertilizable life of the egg is presented in the context of various abnormalities of fertilization, especially polyspermy, and the deleterious influence of delayed mating or insemination on conception rate and fecundity is stressed. Early embryonic development is noted to be dependent upon the secretory function of the reproductive tract, specific uterine proteins possibly facilitating the considerable growth and elongation of the blastocysts found during their redistribution throughout the cornua. Finally, the nature of the balance between luteotrophic and luteolytic factors that permits the establishment of pregnancy is described, together with an indication of the involvement of prostaglandin F 2α secretion into the uterine veins in both pregnant and non-pregnant animals.

Nonresonant coupling of ion acoustic waves in three dimensions

The slow nonresonant beat disturbance (ω2−ω1, k2−k1) generated by two obliquely propagating ion acoustic waves has been observed. The beat signal wavenumber, direction, and speed agree with predicted values. The beat signal amplitude, normalized to the product of the driving wave amplitudes, exhibits a maximum when the beat signal speed is approximately equal to one-half the ion thermal speed. The energy transfer between the two driving waves has also been observed.

Studies on the gill ciliation of the American oyster Crassostrea virginica (Gmelin).

Latero-frontal, para-latero-frontal, and frontal ciliary tracts on the gill filaments of Crassostrea virginica (Gmelin) were studied with light microscopy and scanning electron microscopy. Latero-frontal cirri are complex structures composed of varying numbers of paired cilia. The multiple pairs of cilia which constitute a single cirrus are closely appressed for a portion of their length; they then branch laterally from the central axis in a plume-like fashion. Latero-frontal cirri of adjacent gill filaments create a filtration sieve which should be capable of retaining particles smaller than 1 μm in diameter. Para-latero-frontal cilia are short, closely spaced cilia arranged as a staggered row along the frontal side of each tract of latero-frontal cirri. Latero-frontal cirri and para-latero-frontal cilia occur on ordinary, principal, and transitional gill filaments. Frontal ciliary tracts of ordinary filaments are divided into a central, ventrally directed coarse tract, flanked on either side by a dorsally directed fine ciliary tract. The coarse tract is covered by cirri which are comprised of five to eight cilia, while the fine frontal tracts are made up of individually functioning cilia. The frontal ciliary tracts of principal and transitional filaments bear only dorsally directed fine cilia. The unique direction of effective beat of the coarse frontal cirri of ordinary filaments, in combination with the action of fine frontal cilia and the strategic location of mucus producing cells, is used to describe a possible mechanism for the sorting of filtered particles.

Xylene Exposure Electronystagmographic and Gaschromatographic Studies in Rabbits

Complaints of vertigo from people who are exposed to industrial solvents have focused interest on their toxic effect on the nervous system. In order to evaluate the influence of an organic solvent, xylene, on the mammalian vestibular system, a series of rabbit experiments were performed. To achieve a constant concentration, the xylene was infused as a lipid emulsion. Blood concentrations were estimated by gas chromatography. Elec-tronystagmography in darkness revealed that at blood xylene concentrations of 30 ppm all rabbits had a positional nystagmus. The beat direction was the opposite of positional alcohol nystagmus. Another difference between the alcohol and the xylene reaction was that rotatory nystagmus responses were exaggerated. The relations between the present findings and the reactions and blood concentrations in people exposed to industrial solvents are discussed.