BIOLOGY OF EGGS AND IMPLANTATION 



815 



dish containing them. But after a short in- 

 terval in the new environment, the zonae 

 may become sticky and chng to the glass 

 surface of the dish or to the pipettes and 

 needles used in transporting them. Nonmo- 

 tile spermatozoa caught within or on the 

 zona pellucida have been pictured many 

 times in the eggs of the human (Shettles, 

 1953), the rhesus monkey (Lewis and Hart- 

 man, 1941), the guinea pig (Squier, 1932), 

 and the rabbit (Pincus, 1930). The same 

 phenomenon has been observed only on rare 

 occasions in rat eggs, again emphasizing dif- 

 ferences in the physical characteristics of 

 the zona from animal to animal. 



There is very little information as to the 

 permeability of the various membranes en- 

 closing the mammalian egg. Recently the 

 eggs of the rabbit, rat, and hamster were ex- 

 posed to dyes such as toluidine blue and 

 alcian blue and to a 1 per cent solution of 

 heparin and digitonin in order to test the 

 selectivity of the membranes (Austin and 

 Lovelock, 1958) . It was found that the zonae 

 pellucidae of all three animals were perme- 

 able to the dyes and digitonin but not to 

 heparin. 



There is too little known of the changes 

 which occur in the zona pellucida and other 

 egg membranes under varying environmen- 

 tal conditions to draw conclusions as to the 

 nature of its selectivity. Techniques whereby 

 invertebrate egg membranes are impaled 

 with microelectrodes have yielded new in- 

 formation as to membrane potentials and 

 resistance at varying stages of fertilization 

 (Tyler, Monroy, Kao and Grundfest, 1956). 

 Similar investigations on mammalian eggs 

 would be valuable in solving the problems 

 of selectivity of the egg membranes and in 

 evaluating the response of eggs to various 

 environmental fluids. 



The question should also be raised as to 

 whether or not the zona pellucida and/or the 

 mucin coating may present barriers to the 

 diffusion of gases and thus constitute a lim- 

 iting factor to the rate of development. 

 Fridhandler, Hafez and Pincus (1957) found 

 no differences in the 0^ uptake when com- 

 paring normal rabbit eggs and eggs in which 

 the mucin coat and zona pellucida had been 

 punctured. Other properties of the zona pel- 

 lucida will be considered later when the 



problem of the means by which spermatozoa 

 penetrate it is discussed. 



2. The Mucous or "Albuminous'' Layer 



Unlike the zona pellucida, which is formed 

 in the ovary, the "albumin" or mucous layer 

 is deposited on the zona by secretions of the 

 glandular cells in the oviducts or uterus and 

 is therefore classified as a tertiary mem- 

 brane. 



In the monotremes (Hill, 1933) and many 

 marsupials (Hartman, 1916; McCrady, 

 1938) an abundant albuminous coat is de- 

 posited on the zona pellucida as the egg 

 moves through the oviduct. A similar de- 

 posit, but composed principally of muco- 

 polysaccharides has been described for the 

 eggs of various animals forming the order 

 Lagomorpha (Cruikshank, 1797; Gregory, 

 1930; Pincus, 1936). A thinner but chenii- 

 cally identical coat has been described in the 

 ova of the horse and dog (Lenhossek, 1911; 

 Hamilton and Day, 1945). It is only in the 

 rabbit that the mucous coat has been 

 charged with limiting the period during 

 which the ovum can be penetrated by sper- 

 matozoa. A very thin layer of mucus has 

 been observed on rabbit eggs removed from 

 5 to 8 hours after ovulation (Pincus, 1930; 

 Braden, 1952). Furthermore, it has been 

 shown that the rabbit egg must be pene- 

 trated by a spermatozoon before the 6th 

 hour after ovulation if normal development 

 is to ensue (Hammond, 1934). That the 

 mucous membrane inhibits sperm penetra- 

 tion is confirmed by the fact that unferti- 

 lized rabbit ova may be stored in vitro for 

 48 to 72 hours without, in many instances, 

 losing their fertilizing capacity after being 

 transferred into the oviducts of properly 

 timed recipients (Chang, 1953). It has been 

 clearly demonstrated that the mucin is 

 stored in the secretory cells of the oviduct 

 and that estrogens are necessary for the 

 synthesis of the mucin granules (Greenwald, 

 1958a). Discharge of the mucin granules is 

 apparently controlled by progesterone. The 

 thickness of the mucin coat on rabbit eggs, 

 or glass beads placed in the oviduct, can be 

 either significantly increased by injecting 

 progesterone in properly conditioned fe- 

 males, or greatly reduced by injecting estro- 

 gens immediately after ovulation. 



