Summary. Pig oocytes were examined at hourly intervals after stimulation with hCG. Meiosis was resumed between 20 and 30 h after hCG. This coincided with a decline in the number of mitochondria and evidence is presented which indicates that this was due to fusion. The number of lipid droplets increased and the volume fraction of large vesicles decreased. Both these organelles maintained a close spatial relationship with the endoplasmic reticulum (ER). Mitochondria were clustered at the periphery of the cell before hCG injection but dispersed with maturation. The volume occupied by large vesicles, 'protein bodies' and Golgi also decreased at the edge of the oocyte with the progression of maturation.
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D. G. Cran
Summary. The administration of PMSG to sheep early in the oestrous cycle (Days 2 or 3) results in the formation of follicular cysts of varied morphology by Days 8 or 9. These may persist for up to 14 days after injection. If PMSG is given on Day 5 or later ~ 50% of such cysts ovulate. However, when PMSG is given at the beginning of the cycle (Day 2 or 3), the membrana granulosa is lost from the majority of the cysts and the theca interna luteinizes. The major hormone secreted by such luteinized follicles is progesterone. The structure of the steroid-secreting cells of the follicles is similar to that of large luteal cells of granulosa cell origin in cyclic corpora lutea. It is suggested that under suitable luteinizing conditions thecal cells may acquire many of the characteristics of granulosa cells.
K. Bamba and D. G. Cran
Summary. Boar spermatozoa acquired resistance to cold shock immediately after exposure to 2·0mmol butylated hydroxytoluene (BHT) l−1 when Beltsville thawing solution was used as a basic diluent, as judged by motility (the proportion of motile spermatozoa) and acrosomal integrity. The concentration of BHT could be reduced to 0·2 mmol l−1 without decreasing the protective action. However, motility was altered in the presence of >0·15 mmol BHT l−1. Beltsville freezing 5 (BF5) diluent was more effective than Beltsville thawing solution in protecting spermatozoa from cold shock, but addition of BHT to BF5 diluent did not affect the motility and acrosomal morphology of spermatozoa before or after cold shock. Dilution of BHT-treated spermatozoa with BF5 diluent did not restore motility and did not afford further protection against cold shock; it was detrimental to spermatozoa treated with 2 mmol BHT l−1 for > 15 min. Egg yolk or lecithin had a detrimental effect.
When spermatozoa were treated with 0·05–0·10 mmol BHT l−1 before slow cooling to 5°C, the progressive motility and acrosomal integrity were maintained better after storage for 6 days than in untreated spermatozoa.
Keywords: spermatozoa; cold shock; boar; butylated hydroxytoluene
K. Bamba and D. G. Cran
Summary. The effect of rapid dilution (1:8 with BTS or 1:6·5 with KRP) and temperature change on sperm morphology and physiology were studied using boar spermatozoa pre-diluted in BF5 diluent. Rapid dilution of cold semen (5°C) with a warm solution (37°C) caused marked acrosomal changes which were most prominent in the anterior region. The acrosomal damage appeared to be caused mainly by rapid warming. In contrast to rapid cooling, rapid warming had little effect upon motility, glutamic-oxaloacetic transaminase release and respiration.
K. Bamba and D. G. Cran
Summary. Rapid warming of semen from 5 to 37°C caused visible damage to the acrosomes of bull and rabbit spermatozoa. The degree and type of damage varied with the species, the bull being the more resistant. While vesiculation was observed in rabbit spermatozoa, neither warm nor cold shock resulted in this defect in the bull. Warm shock of bull spermatozoa caused acrosomal knobbing in an anterior region of the head. Spermatozoa with thread and/or droplet-like structures were frequently observed in bull semen after cold shock.
Keywords: bull; rabbit; rapid warming; spermatozoa; morphology
K. Bamba and D. G. Cran
Summary. Studies have been carried out to investigate factors related to the induction of warm shock in boar spermatozoa. Rapid dilution per se caused visible damage to acrosomes when the sample contained 7·5% or more glycerol. This dilution effect was greater at lower temperatures. Acrosomal damage was greatly reduced by raising the dilution temperature from 15 to 25°C, suggesting that a change in the physico-chemical characteristics of the acrosomal membrane occurred between these temperatures. During rapid dilution with warming, the dilution rate, the magnitude of the temperature change and the terminal temperature had a significant influence on acrosomal integrity; a terminal temperature of 35°C was much more detrimental than one of 25°C. The first sign of acrosomal damage was observed 15 sec after rapid dilution + warming and the damage was nearly maximal by 60 sec. An antioxidant, butylated hydroxytoluene (BHT), was effective against both rapid cooling and warming, while glycerol, dimethylsulphoxide and propylene glycol were ineffective in preventing warm shock.
Keywords: boar spermatozoa; warming and dilution; acrosome; structure
D. G. Cran and H. M. Dott
A.R.C. Unit of Reproductive Physiology and Biochemistry, University of Cambridge, U.K.
A defect of the sperm acrosome, called 'knobbing' has been associated with sterility in the bull (Teunissen, 1946; Blom, 1948; Hancock, 1952; Donald & Hancock, 1953; Blom & Birch-Andersen, 1962; Blom, 1972), and boar (Bane, 1961), and has been seen in stallion (Dott, 1975), ram, dog and impala (unpublished).
Donald & Hancock (1953) came to the conclusion that this was an autosomal sex-linked recessive characteristic in Friesian bulls. At the level of the light microscope it is seen as an apical, often eccentric swelling which folds back over the head. Blom & Birch-Andersen (1962) described this character as a swelling of the apical body to six to eight times normal. Much of the region was cystic and contained vesicles with inclusions. The ultrastructure of this defect in Friesian bulls is described in the present paper.
Semen was collected in
J. D. O'Shea, Mary F. Hay, and D. G. Cran
Summary. Sixteen antral follicles, 1·8–4·2 mm in diameter, at various stages of atresia, were studied by electron microscopy. Deletion of theca interna cells by condensation followed by fragmentation (apoptosis), with subsequent engulfment of the fragments by remaining thecal cells, was present at all stages, but was most marked during secondary and tertiary atresia. In primary and secondary atresia, the relative numbers of thecal cells whose cytoplasm was rich in tubular endoplasmic reticulum were higher than in non-atretic follicles of comparable size. During tertiary atresia the number of cell layers in the theca interna was reduced, and cells rich in tubular endoplasmic reticulum became proportionately less numerous. Degenerating cellular material was present within the lumina of thecal capillaries at all stages of atresia. Such material was rarely seen in primary atresia, and increased in incidence progressively in later stages. It was concluded that during atresia a large number of theca interna cells are deleted by apoptosis, and many thecal capillaries become blocked with cellular debris.
D. G. Cran, R. M. Moor, and Mary F. Hay
Summary. The structure and distribution of organelles within oocytes of developing antral follicles were studied qualitatively and quantitatively. In the smallest category of follicle (0·2–0·4 mm diam.) the Golgi was present in a peripheral position, the endoplasmic reticulum was distended and mitochondria were intimately associated with it. Processes from surrounding cumulus cells were in contact with the oolemma which was thrown up into slender villi, increasing the oocyte surface area 5-fold. In these and all subsequent follicles the oocyte cytoplasm contained numerous 30 nm particles or vesicles. As the follicles grew to about 2·0 mm the mitochondria became located in a peripheral band, interior to which were numerous large vesicles, and the villi became shorter and thicker. In follicles >2·0 mm the mitochondrial band was largely dispersed and cortical granules rested close to the oolemma. Little structural change was then observed until late oestrus. In oocytes from preovulatory follicles at this stage an internal dense zone was formed in the zona pellucida. Most of the cumulus cell processes degenerated and most mitochondria assumed a hooded appearance.
H. M. Dott, Mary F. Hay, D. G. Cran, and R. M. Moor
Summary. Follicles were obtained from the ovaries of four groups of 15 ewes. Ewes in the control group were ovariectomized on the 12th day of the oestrous cycle. The other ewes were all given PMSG on the 12th day of the cycle; some were ovariectomized 24 or 40 h later, the others were given prostaglandin followed by hCG and were ovariectomized 6 or 12 h after the hCG injection. All follicles >2 mm in diameter were measured and examined macroscopically for signs of atresia. Some were subjected to detailed morphological examination, the pattern of steroid secretion was determined in others. All the evidence from these three approaches suggested that, in vivo, reversal of the atretic process ('rescue') plays no part in the increase in the number of follicles observed following administration of PMSG.