Mechanism of Fundulus Epiboly--A Current View

This paper summarizes evidence for the following picture ofFundulus epiboly, with an eye toward laying groundwork for futureinvestigation. The major force in epiboly is the yolk syncytiallayer (YSL). Prior to epiboly, it spreads well beyond the borderof the blastoderm to form the wide external YSL (E-YSL). Thishas contractile properties, which, however, are restrained priorto epiboly by the attached enveloping layer (EVL) of the blastoderm.Epiboly begins when the E-YSL contracts and narrows, throwingits surface into folds and pulling the internal YSL (I-YSL)and the attached EVL vegetally. When the narrowing of the E-YSLhas ceased, it is postulated that its contractility continuesas a circumferential wave of vegetally directed contractionthat moves over the yolk toward the vegetal pole, dragging theI-YSL and the attached EVL (and blastoderm) with it. The mostobvious visible manifestation of this wave is a marked marginalconstriction, where the YSL joins the yolk cytoplasmic layer(YCL). As this contractile wave passes over the yolk, cytoplasmfrom the YCL mingles with that of the advancing E-YSL, and YCLsurface adds to the already highly convoluted surface of theE-YSL. This folded surface is the site of a thin, highly localizedband of rapid endocytosis that encircles the egg and passesover it with the E-YSL in a wave throughout epiboly. This internalization,which is receptor independent and therefore somehow programmed,accompanies the putative contractile wave, and accounts forthe disappearance of the surface of the YCL. Since the YCL surfacestands in the way of the advancing YSL, its internalizationis part of the mechanism of epiboly. As the I-YSL expands inresponse to this marginal pull, its abundant microvilli graduallydisappear, providing surface for its epiboly. The firmly attachedEVL likewise expands toward the vegetal pole in response tothe pull of the autonomously expanding YSL. As epiboly of theEVL progresses, it adjusts to the geometric problems posed bya sheet expanding over a sphere by active cell rearrangementwithin the cell monolayer. Thus, epiboly of the EVL has an activeas well as a passive component. Deep cells are not causallyinvolved in epiboly, but move about in coordinated ways in theconstantly increasing space between the I-YSL and the EVL providedby epiboly and form the germ ring and the embryonic shield andeventually the embryo proper. An attempt is made to pull allof this together, and more, in order to achieve as comprehensivean understanding of epiboly as present evidence will allow.     

Programmed Endocytosis During Epiboly of Fundulus heteroclitus

A major question in the analysis of teleost epiboly is the fateof the yolk cytoplasmic layer. It diminishes during epibolyand eventually disappears at the completion of epiboly. Thispaper is concerned with the fate of the surface of the yolkcytoplasmic layer during epiboly. When gastrulae during epibolyare bathed in lucifer yellow (CH) and then observed with fluorescentmicroscopy or bathed in ferritin and then fixed and observedwith TEM, a thin circumferential ring of endocytic vesiclesis observed, confined to the external yolk syncytial layer justperipheral to the advancing margin of the blastoderm. Even thoughthe entire egg is immersed in the marker, endocytosis is confinedto this limited region. More precisely, this endocytosis occursonly within the region of the external yolk syncytial layer,where the surface is most folded. The endocytic vesicles thusformed move downward and settle on the surface of the membraneseparating the yolk from the cytoplasm in the yolk syncytiallayer. They do not join the surface of the internal yolk syncytiallayer; hence they do not contribute to its expansion. Priorto the onset of epiboly there is no such endocytosis at thesurface of the egg. Since this endocytosis occurs only duringepiboly and only at the surface of the external yolk syncytiallayer just peripheral to the advancing margin of the blastoderm,and in the absence of large molecules in the medium, we concludethat it is programmed. We, therefore, present this as a caseof programmed internalization of cell surface serving as themorphogenetic mechanism responsible for the disappearance ofthe surface of the yolk cytoplasmic layer during gastrulationof the teleost Fundulus heteroclitus     

Locomotion of Fundulus Deep Cells during Gastrulation

SYNOPSIS. Mechanism of locomotion of deep cells of Fundulusheteroclitus was studied in vivo during gastrulation with theaid of time lapse cinemicrography (Nomarski differential interferencecontrast optics), scanning electron microscopy of cells knownto be moving at the time of fixation, and cell culture. Theseare our findings. 1) Deep cells usually move rapidly, at about10–15 µ/min, regardless of whether they move byblebbing or spreading. Evidence suggests that this high speedis associated with weak adhesion of the trailing edge: it remainsrounded, without large retraction fibers, and it advances continuouslywith advance of the leading edge, not sporadically, as it wouldif it adhered strongly. 2) In contrast, when stationary cellsin close contact separate, they remain connected by retractionfibers, suggesting strong punctate adhesions. 3) Locomotionby shortening of a long lobopodium is really a form of spreadingmovement; the tip of a lobopodium always spreads. Also, sincespeed of shortening decreases with continuance, it may dependprimarily on elastic recoil rather than active contraction.4) Fundulus deep cells appear to move in two ways: a) protrusionof blebs, followed by much cytoplasmic flow; b) protrusion oflamellipodia, accompanied by filopodia and frequent cell shortening.5) Filopodia were not found except at the leading edge of aspreading lamellipodium and often spread themselves; perhapsfilopodia and lamellipodia are interconvertible. 6) A lamellipodialmargin may form undulations in vivo that move backward likeruffles in vitro. 7) At all times, whether stationary or moving,the surface of deep cells is smooth, raising unanswered questionsconcerning the source of surface for their rapid protrusiveactivity.