Phytohemagglutinin Induces Delays in Oral Regeneration in the Ciliate Stentor coeruleus1

Regenerating Stentor, exposed to the plant lectin phytohemagglutinin, are significantly delayed in completing oral regeneration. All of the delays are restricted to the earliest stages of regeneration prior to stage 3. The effects of phytohemagglutinin are reversible once the drug is removed. The addition of the presumed sugar receptor for phytohemagglutinin, N-acetylgalactosamine, at the start of oral regeneration could not reverse the effects of phytohemagglutinin, but the addition of excess (10 mM) extracellular calcium could. When the addition of the excess calcium was delayed for various times after phytohemagglutinin exposure, the effectiveness of the calcium in reversing the phytohemagglutinin-induced delays was reduced. Based on the mechanism of action of PHA in other cells, these results suggest that membrane proteins may be involved in controlling oral regeneration, possibly through mechanisms involving Ca⁺⁺.     

Fission in Heteropolar Tandem Grafts of the Ciliate Stentor coeruleus*

SYNOPSIS Stentor pairs fused by their posterior ends show persisting mutual adhesion, tho they eventually pull apart. When immobilized in methyl cellulose these grafts constricted in 2 in the line of heal. In no other type of stentor graft has such deliberate separation been observed.     

Cortical Control over the Direction of Macronuclear Elongation in the Heterotrich Ciliate Stentor coeruleus1

Earlier experimental work involving macronuclear implants in Stentor coeruleus has shown that the cytoplasmic cortex of the nuclear site 1) attracts the macronucleus and 2) holds it in place during interphase. Now experiments indicate macronuclei transferred with overlying cortex elongate in the direction of the transferred cortical pigment stripes, whether or not the transferred stripes realign in the direction of the host stentor's stripes. Therefore the third function of the cortex is to determine the direction of elongation and thus assure that both daughter cells at division receive part of the macronucleus.     

Visualizing Cytoplasmic Flow During Single-cell Wound Healing in Stentor coeruleus

Although wound-healing is often addressed at the level of whole tissues, in many cases individual cells are able to heal wounds within themselves, repairing broken cell membrane before the cellular contents leak out. The giant unicellular organism Stentor coeruleus, in which cells can be more than one millimeter in size, have been a classical model organism for studying wound healing in single cells. Stentor cells can be cut in half without loss of viability, and can even be cut and grafted together. But this high tolerance to cutting raises the question of why the cytoplasm does not simply flow out from the size of the cut. Here we present a method for cutting Stentor cells while simultaneously imaging the movement of cytoplasm in the vicinity of the cut at high spatial and temporal resolution. The key to our method is to use a "double decker" microscope configuration in which the surgery is performed under a dissecting microscope focused on a chamber that is simultaneously viewed from below at high resolution using an inverted microscope with a high NA lens. This setup allows a high level of control over the surgical procedure while still permitting high resolution tracking of cytoplasm.     

Mating Types in Stentor coeruleus

SYNOPSIS. Conjugation in Stentor coeruleus was investigated, using a standardized culture technic which yielded large numbers of mating pairs within a single culture. Spontaneous bursts of selfing occurred during a definite interval in the development of a culture. Structurally distinct preconjugator cells appeared immediately before as well as during the initial stages of a burst of conjugation. Mating pairs were formed by the union of 2 preconjugators.
Mixing 8 stocks in all possible combinations of pairs and observing their subsequent response revealed they were separable into 2 complementary mating types. The majority of mating pairs formed in mixtures of stocks consisted of individuals of different mating types.
It is suggested that control of mating types in the ciliate order Heterotrichida may be of a somewhat different nature from that found in other ciliates.     

Scanning Electron Microscopy of the Cortex of the Ciliate Stentor coeruleus. A View from the Inside*

SYNOPSIS. A microdissection procedure was developed which permits the viewing of the inside surface of the cortex of Stentor coeruleus with scanning electron microscopy. Parallel bands of myonemes cover the entire inner surface of the cortex. The myonemes of the stalk region are ribbon-shaped and lack cross connections. The myonemes of the anterior cortex are flattened against the surface and are interconnected by an extensive system of cross branches. The inner surface of the frontal field is covered with a regularly cross-branched myoneme system which follows the curved pattern of frontal field kinety. The observed branching patterns and shapes of the myonemes support the hypothesis that they cause contraction of the cell. The membranellar root system was examined. Each membranellar root makes a 90° counterclockwise twist along its vertical axis (viewed from the inside) as it descends into the cell. The outer edge of each root fuses with the inner edge of the adjacent one, forming a continuous fiber sheet linking the roots together.     

Microtubules and Filaments Beneath the Fission Furrow of Stentor coeruleus1

ABSTRACT. The ultrastructure of the cortex beneath the fission furrow of dividing Stentor coeruleus was examined using scanning and transmission electron microscopy. During division, basal bodies, axonemes, and km fibers beneath the furrow were absorbed near the moving primordial oral apparatus, and a circumferential band of microtubules and filaments was formed at the base of the furrow. The location and orientation of this fibrous band suggest that it may be an important component of the cytokinetic machinery. Treatment with vinblastine sulfate (4 × 10^-5 M) disrupted the circumferential microtubules and blocked division, which is consistent with this hypothesis.     

Single-cell analysis of habituation in Stentor coeruleus

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The Effects of Conjugation in Stentor coeruleus

SYNOPSIS. Culture methods favorable for conjugation in Stentor coeruleus are described and the effects of conjugation on over 12,000 conjugating pairs of S. coeruleus are presented.     

Light-adaptation in the photophobic response by Stentor coeruleus

Effects of preillumination on photophobic response (light-adaptation) and recovery of the photophobic sensitivity in the dark (dark-adaptation) in Stentor coeruleus were examined. When the cells were preilluminated with white light of 7.80 W/m² for 2 min, the fluence-rate response curve of photophobic response was shifted toward higher light intensities by half an order of magnitude compared to the one without preillumination. Preillumination with a higher light intensity resulted in a further shift of the fluencerate response curve. An action spectrum for light-adaptation showed a primary peak at 610 nm and secondary peaks at 540 and 480 nm which are almost identical to the peaks observed in the photophobic action spectrum.The light-adapted cells showed a recovery of their photophobic sensing ability following dark treatment. Dark-adaptation resulted in total recovery of photophobic sensing ability in 8 minutes for the most cases examined.     

Ciliary Proteins and Cytodifferentiation in Stentor coeruleus*

To elucidate further the molecular events required for cytodifferentiation in Stentor coeruleus, the effects of several chemical metabolic inhibitors were tested on the outgrowth in situ of the membranellar cilia of the oral feeding organelle. The chemicals used included several inhibitors of cytoplasmic and mitochondrial protein synthesis (cycloheximide, emetine, and chloramphenicol), and an antimitotic agent (colchicine). Ciliary growth was affected only by colchicine, suggesting that a pool of “ciliary proteins” exists in interphase Stentor of sufficient size to permit complete reformation of the membranellar cilia. The implication of these observations to an understanding of the more complicated process of oral regeneration is discussed.