151microtubule- containing structures in the cleavage plane, such as interzonal spindle
microtubules. The rapid expansion of new membrane sustained by the furrow
microtubule array is sometimes regarded as an embryonic amplification of the mid-
body-dependent localized exocytosis required for abscission of dividing cells of
organisms ranging from yeasts to plants and animals (Straight and Field 2000 ;
Glotzer 2001 ).
4.4.2.3 Protrusive Activity During Cleavage Furrow Closure
Various kinds of membrane protrusions develop along cleavage furrow margins
(Danilchik and Brown 2008 ; Danilchik et al. 2013 ). Long microvilli are seen associ-
ated with stress folds at the cortex, and lamellipodia and filopodia extend near the
furrow base itself. These protrusions are actively motile and can make relatively
robust contacts across the extracellular space separating the furrow margins.
Abrogating normal actin assembly by microinjection of constitutively active rho
and cdc42 disrupts normal furrow margin protrusive activity and cleavage furrow-
ing, suggesting that the protrusions play a role in normal furrow closure and blasto-
mere adhesion (Danilchik and Brown 2008 ).
4.4.2.4 Blastocoel Formation
In amphibian embryos, the initiation of blastocoel formation is evident well before
the first contractile ring has finished closing (Kalt 1971a, b). During advance of the
cleavage furrow, a new domain of plasma membrane becomes inserted in the plane
of the plasma membrane on either side of the contractile ring, resulting in two
expanding basolateral surfaces facing each other between separating blastomeres
(Bluemink 1970 ; Kalt 1971a, b). A similar phenomenon likely occurs in the zebraf-
ish (Feng et al. 2002 ; Urven et al. 2006 ; Eno et al. 2016 ). As discussed above, this
rapid expansion of new membrane depends on localized exocytosis of maternally
derived vesicles immediately behind the advancing contractile ring. The new cleav-
age planes express maternally encoded C- and EP-cadherins which facilitate adhe-
sion between sister blastomeres (Heasman et al. 1994a, b; Kühl and Wedlich 1996 )
and integrins. Each successive cleavage event inserts more basolateral surface
between dividing daughter cells; the blastocoel, resting at the intersection of all the
early cleavage planes, thus is entirely lined by basolateral surface and, with the
development of apical–basolateral tight junctions, becomes osmotically isolated
from the outside world. Although definitive extracellular matrix fibers, including
fibronectin, do not develop until mid- to late-blastula stage (Boucaut et al. 1984 ;
Davidson et al. 2004 , 2008 ), the volume of the blastocoel is nevertheless entirely
filled with extracellular matrix at all stages (Keller 1986 , Danilchik, unpublished).
The blastocoel undergoes continuous expansion and change in shape throughout the
cleavage stage, via osmotic uptake of water (Slack and Warner 1973 ; Han et al.
1991 ; Uochi et al. 1997 ), as well as the progressive epibolic thinning of the
4 Vertebrate Embryonic Cleavage Pattern Determination