Cell junctions in early embryos of squid (Loligo pealei)

Summary Squid embryos examined by freeze-fracture and thin-section electron microscopy exhibit identifiable gap junctions during mid-cleavage stages (stages 7–8), and junctional complexes composed of adherent appositions, elaborate septate junctions and gap junctions at slightly later stages (stages 12–13). During germinal layer establishment (stages 12–13) cytoplasmic bridges frequently link the embryonic cells. The presence of gap junctions in cleavagestage embryos provides the morphological substrate for a demonstrated pathway of direct cell-cell communication that is modifiable by experimental treatments and may be physiologically regulatable. The existence of septate junctions and adherent contacts at later stages suggests that some functional specialization, perhaps the establishment of a strongly joined framework of cells at the surface of the embryo, accompanies the formation of germinal layers.     

The locomotory function of the fins in the squid Loligo pealei

Kinematic data of high spatial and temporal resolution, acquired from image sequences of adult long-finned squid, Loligo pealei, during steady swimming in a flume, were used to examine the role of fins and the coordination between fin and jet propulsion in squid locomotion. Fin shape and body outlines were digitized and used to calculate fin wave speed, amplitude, frequency, angle of attack, body deformation, speed, and acceleration. L. pealei were observed to have two fin gait patterns with a transition at 1.4-1.8 mantle lengths per second (L_m s^-1) marked by alternation between the two patterns. Fin motion in L. pealei exhibited characteristics of both traveling waves and flapping wings. At low speeds, fin motion was more wave-like; at high speeds, fin motion was more flap-like and was marked by regular periods during which the fins were wrapped tightly against the mantle. Fin cycle frequencies were dependent on swimming speed and gait, and obvious coordination between the fins and jet were observed. Fin wave speed, angle of attack, and body acceleration confirmed the role of fins in thrust production and revealed a role of fins at all swimming speeds by a transition from drag-based to lift-based thrust when fin wave speed dropped below swimming speed. Estimates of peak fin thrust were as high as 0.44-0.96 times peak jet thrust in steady swimming over the range of swimming speeds observed. Fin downstrokes generally contributed more to thrust than did upstrokes, especially at high speeds.     

Pigment Migration and Adaptation in the Eye of the Squid, Loligo pealei

The migration of the screening pigment was investigated in the retina of the intact squid. The action spectrum of pigment migration corresponds to the action spectrum of the visual pigment, rhodopsin, rather than to the absorption spectrum of the screening pigment. The total number of quanta required for a fixed criterion of pigment migration is the same, when the quanta are delivered over any period of time from 6 s to an hour or more. When less than 3–10% of the rhodopsin is isomerized, the screening pigment migrates out to the tips of the receptors with a time-course of 5–15 min, and back again over the same period of time. When rather more than 10% is isomerized, the outward migration takes 5–15 min, but the screening pigment does not migrate inwards, even after several hours in the dark. Indirect evidence suggests that the band of screening pigment, when it reaches the tips of the receptors, is approximately equivalent to a filter of 0.6 log units. The spectral sensitivity of the optic nerve response was measured, and was found to be broader than the absorption spectrum of squid rhodopsin in vitro; the broadness could be explained by self-screening, assuming a density of rhodopsin of 0.6 log units at 500 nm.     

An ultrastructural and cytochemical analysis of oogenesis in the squid, Loligo pealei

Oogenesis has been investigated utilizing both light and electron microscopical techniques in the squid, Loligo pealei. This complex process has been divided into five stages according to the structure of the follicle. Because of the highly coordinated differentiation of the follicle cells (and follicular syncytium) and the oocyte, their development is described in concert. Specific attention is given to the contribution of the follicular syncytium to vitellogenesis and the formation of the extracellular egg envelope or chorion. Our observations indicate heterosynthetic yolk production and the synthesis of the secondary envelope by the follicular syncytium.     

Topographic regulation of phosphorylation in giant neurons of the squid, Loligo pealei: role of phosphatases

In previous studies of phosphorylation in squid stellate ganglion neurons, we demonstrated that a specific multimeric phosphorylation complex characterized each cellular compartment. Although the endogenous protein profile of cell body extracts (giant fiber lobe, GFL), as determined by Coomassie staining, was similar to that of axoplasm from the giant axon, in this study we show that the protein phosphorylation profiles are qualitatively different. Whereas many axoplasm proteins were phosphorylated, including most cytoskeletal proteins, virtually all phosphorylation in perikarya was confined to low molecular weight compounds (<6 kDa). Because phosphorylation of exogenous substrates, histone and casein, was equally active in extracts from both compartments, failure to detect endogenous protein phosphorylation in cell bodies was attributed to the presence of more active phosphatases. To further explore the role of phosphatases in these neurons, we studied phosphorylation in the presence of serine/threonine and protein tyrosine phosphatase (PTP) inhibitors. We found that phosphorylation of axonal cytoskeletal proteins was modulated by okadaic acid-sensitive ser/thr phosphatases, whereas cell body phosphorylation was more sensitive to an inhibitor of protein tyrosine phosphatases, such as vanadate. Inhibition of PTPs by vanadate stimulated endogenous phosphorylation of GFL proteins, including cytoskeletal proteins. Protein tyrosine kinase activity was equally stimulated by vanadate in cell body and axonal whole homogenates and Triton X-100 free soluble extracts, but only the Triton X soluble fraction (membrane bound proteins) of the GFL exhibited significant activation in the presence of vanadate, suggesting higher PTP activities in this fraction than in the axon. The data are consistent with the hypothesis that neuronal protein phosphorylation in axons and cell bodies is modulated by different phosphatases associated with compartment-specific multimeric complexes.     

Determination of Erythroid Cells in Unincubated Chick Blastoderm

Portions the size of 1/6 to 1/32 part of unincubated blastoderm cultured in an albumen-salineagar solid medium form erythroid cells under conditions which suppress normal co-ordinated movements for mesoderm induction. Anterior and posterior regions of unincubated blastoderm have the same potentiality to form erythroid cells. The marginal zone seems to be the contributor of prospective erythroid cells. Portion 1/16 part of unincubated blastoderm forms morphologically distinct erythroid cells of the primitive and definitive lines as in normal development in ovo. It seems progenitor cell(s) is pre-programmed to a particular pattern of differentiation and/or includes differentiation to various erythroid cell types as an obligatory step. This system provides novel experimental possibilities in the study of erythroid cell determination and differentiation.     

Bidirectional transport of fluorescently labeled vesicles introduced into extruded axoplasm of squid Loligo pealei

A reconstituted model was devised to study the mechanisms of fast axonal transport in the squid Loligo pealei. Axonal vesicles were isolated from axoplasm of the giant axon and labeled with rhodamine-conjugated octadecanol, a membrane-specific fluorescent probe. The labeled vesicles were then injected into a fresh preparation of extruded axoplasm in which endogenous vesicle transport was occurring normally. The movement of the fluorescent, exogenous vesicles was observed by epifluorescence microscopy for as long as 5 min without significant photobleaching, and the transport of endogenous, nonfluorescent vesicles was monitored by video-enhanced differential interference-contrast microscopy. The transport of fluorescent, exogenous vesicles was shown to be bidirectional and ATP-dependent and occurred at a mean rate of 6.98 +/- 4.11 micron/s (mean +/- standard deviation, n = 41). In comparison, the mean rate of transport of nonfluorescent, endogenous vesicles in control axoplasm treated with vesicle buffer alone was 4.76 +/- 1.60 micron/s (n = 64). These rates are slightly higher than the mean rate of endogenous vesicle movement in extruded axoplasm (3.56 +/- 1.05 micron/s, n = 40) not subject to vesicles or vesicle buffer. Not all vesicles and organelles, exogenous or endogenous, were observed to move. In experiments in which proteins of the surface of the fluorescent vesicles were digested with trypsin before injection, no movement of the fluorescent vesicles was observed, although the transport of endogenous vesicles and organelles appeared to proceed normally. The results summarized above indicate that isolated vesicles, incorporated into axoplasm, move with the characteristics of fast axonal transport. Because the vesicles are fluorescent, they can be readily distinguished from nonfluorescent, endogenous vesicles. Moreover, this system permits vesicle characteristics to be experimentally manipulated, and therefore may prove valuable for the elucidation of the mechanisms of fast axonal transport.     

Intercellular Transport and Cytoplasmic Streaming in Chara hispida

The correlation between the velocities of cytoplasmic streamingand of translocation of ¹⁴C-photosynthate and ³²P-phosphateassociated radioactivity has been investigated in whole plantsof the green freshwater alga Chara hispida L. Tracer was suppliedto the plant's rhizoid system in a split-chamber. The velocityof cytoplasmic streaming of 52±3.3 µm s^–1compares with 57±10 µm s^–1 found for ¹⁴C-transportand 32±20 µm s^–1 found for ³²P-transport.There was no indication of intercellular translocation at avelocity faster than visible streaming. Cytochalasin B inhibitedthe translocation of ³²P and cytoplasmic streaming. CytochalasinB becomes fully effective in inhibiting streaming and transportafter an incubation time of at least 5 h. Key words: Chara hispida, Cytoplasmic streaming, Intercellular transport     

Mechanisms of Adhesion Among Cells of the Early Chick Blastoderm

Cells from the extraembryonic endoderm of the gastrulating chick embryo contain a β-d-galactoside-binding lectin inhibited by thiodigalactoside (TDG). TDG inhibits the aggregation of freshly prepared cells. In these fresh cell suspensions, adhesion is also inhibited when purified lectin is added to the aggregation assay. If these cells are incubated at 22° C their adhesion decreases. Associated with this is an increase in lectin activity in the cell supernatants. In these incubated cells aggregation is stimulated by TDG and desialyzed fetuin. These data suggest that the lectin may have a role to play in cellular adhesion. Under some experimental conditions extraembryonic endoderm cells from rosettes with trypsinized glutaraldehyde-fixed rabbit erythrocytes. This phenomenon is inhibited, to a certain extent, by TDG.     

Deltamethrin induced changes in choline transport and phosphorylation activities in synaptosomes from the optic lobe of squid, Loligo pealei

1. Deltamethrin, a powerful synthetic pyrethroid causes a significant change in choline transport in freshly prepared synaptosomes from squid optic lobes. 2. At resting state (nondepolarized) such an effect manifested as a reduction of 14C-choline uptake in a short term (1 min) uptake experiment. 3. At depolarized state, or under conditions where synaptosomes are subjected to osmotic, aging and other stress conditions, deltamethrin caused stimulation of 14C-choline uptake, resulting in elevation of the levels of total radiocarbons in synaptosomes. 4. Such changes are accompanied with changes in overall phosphorylation activities in synaptosomes.     

Arrangement of tubulin subunits and microtubule-associated proteins in the central-pair microtubule apparatus of squid (Loligo pealei) sperm flagella

This study provides a comprehensive, high-resolution structural analysis of the central-pair microtubule apparatus of sperm flagella. It describes the arrangement of several microtubule-associated sheath components and suggests, contrary to previous thinking, that microtubules are structurally asymmetric. The two microtubules of the central pair are different in several respects: the C2 tubule bears a single row of 18-nm-long sheath projections with an axial periodicity of 16 nm, whereas the C1 tubule possesses rows of 9-nm globular sheath components with an axial repeat of 32 nm. The lumen of the C2 tubule always appears completely filled with electron-dense material; that of the C1 tubule is frequently hollow. The C2 tubule also possesses a series of beaded chains arranged around the microtubule; the beaded chains are composed of globular subunits 7.5-10 nm in diameter and appear to function in the pairing of the C1 and C2 tubules. These findings indicate: that the beaded chains are not helical, but assume the form of lock washers arranged with a 16-nm axial periodicity on the microtubule; and that the lattice of tubulin dimers in the C2 tubule is not helically symmetric, but that there are seams between certain pairs of protofilaments. Proposed lattice models predict that, because of these seams, central pair and perhaps all singlet microtubules may contain a ribbon of 2-5 protofilaments that are resistant to solubilization; these models are supported by the results of the accompanying paper (R. W. Linck, and G. L. Langevin. 1981. J. Cell Biol. 89: 323-337.     

Phylogenetic characterization of epibiotic bacteria in the accessory nidamental gland and egg capsules of the squid Loligo pealei (Cephalopoda:Loliginidae)

Sexually mature female squid Loligo pealei harbour dense bacterial communities in their accessory nidamental glands (ANGs) and in their egg capsules (ECs). This study describes a molecular approach using the 16S rRNA gene (rDNA) to identify bacterial populations within the ANG and the ECs of the North Atlantic squid species L. pealei. Fluorescent in situ hybridization (FISH) and 16S rDNA analysis showed that predominantly alpha- and, to a lesser extent, gamma-proteobacteria were the predominant components of the ANG and EC bacterial communities. Sequencing results showed the presence of alpha-proteobacterial populations affiliated with the Roseobacter group and additional deep-branching alpha-proteobacterial lineages. In contrast, isolates from the ANG and ECs contained only a few alpha-proteobacteria of the Roseobacter group compared with several gamma-proteobacterial isolates, mostly Shewanella and Pseudoalteromonas species. Most of the ANG-associated bacterial populations were also found within the ECs of L. pealei. The molecular approach allowed the visualization of alpha-proteobacteria as major constituents of a bacterial symbiosis within the reproductive system of the Loliginidae.