(Blackshaw, unpublished), nor ultraviolet absorbance by the sperm
heads (6,7) reflect changes consistent with the observed fertility
results,

Recently, we have turned our a:tention © other species Qli£13)..
Our work with the aging of rabbit spermatozoa before insemination
will be reported at this Congress by Miller and Blackshaw (13).

Our work with frogs (11), however, has provided us with the
first undeniable and direct experimental proof that the faulty em-
bryogenesis observed in diploid progeny produced by aged spermatozoa
was due to an impairment in the genetic information system (DNA —>
RNA —> protein) contributed by the spermatozoa to those embryos.
R. G. Hart in our laboratory found that spermatozoa of Rana pipiens
aged in a test tube for 24 % 1 hours at 18°C were able to fertilize
eggs, but about 30-35% of the embryos ceased development, though
living, at the gastrula stage of embryogenesis. Such arrested gas-
trulae when injected with RNA extracted from control, normal gastru-
lae (produced by fresh spermatozoa) were able to continue embryo-
genesis through gastrulation. Similar injections of normal gastru-
lae RNA extract subjected to RNase and RNA extracts from the arrest-
ed gastrulae did not correct the embryo arrest.

 

By serological tests the normal gastrulae were shown to pos-
sess two additional, distinctly different proteins than were found
in the arrested gastrulae produced by aged spermatozoa. The in-
jected RNA from normal, control embryos while permitting the arrest-
ed embryos to proceed through gastrulation did not, however, correct
all the genetic damage caused by the aging of the spermatozoa for
many different morphological anomalies were observed in the subse-
quent stages.

Since evidence is lacking that spermatozoa possess consist-
ently measurable and significant amounts of RNA, the age-induced ef-
fect must have been directly on the DNA the spermatozoa contain.
The physiological time-induced change in the fundamental component
of the spermatozoan genetic code and information transfer system,
presumably in the DNA, if a mutation, is of a dominant, death-deal-
ing kind. It is apparent from Dr. Hart's work with frogs that the
DNA change caused by spermatozoan aging is a drastic mutation cover-
ing by death of the embryo a number of lesser changes, which are
revealed only by the presence of a director of continued develop-
ment in the form of injected normal embryo RNA.

Our most recent observations in the study of the aging of bo-
vine spermatozoa prompt us to ask whether or not the changes in fer-
tility and embryo survival observed are due to a change in the
nucleotide base ratio, genetic code of the DNA per se or to a shift
in the degree of DNA-protein binding in the chromatin of the sperma-
tozoa. Recently, Dr. Radmilo Todorovic, in our laboratory,(24) has

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