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R. D. BAKER and C. POLGE

Summary.

Boar spermatozoa fertilized only a small proportion of the follicular and ovulated eggs that were transferred into the uterus of artificially inseminated gilts. The possibility that spermatozoa had entered the oviduct or were influenced by tubal secretions before penetration cannot be excluded. Sperm penetration in utero was not observed when oviducts were ligated at the uterotubal junction before transferring eggs into the uterus.

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R. D. BAKER and A. A. DEGEN

Summary.

Twenty-four gilts were inseminated with live spermatozoa, dead spermatozoa (killed by rapid freezing and thawing) or a mixture containing an equal number of live and dead spermatozoa. Before insemination, the live spermatozoa were stained with fluorescene isothiocyanate. The females were killed 8 hr after insemination and spermatozoa were recovered from their reproductive tracts. Significantly more spermatozoa were recovered from the oviducts of gilts inseminated with live spermatozoa than from oviducts of gilts inseminated with either dead spermatozoa or the mixture. In gilts inseminated with the mixture, 54, 73, 71 and 100% of the spermatozoa recovered from the posterior half of the uterine horns, anterior half of the uterine horns, tubal flushings and the surface of ova, respectively, showed fluorescence.

In a second experiment, the left oviducts and right uterine horns were cannulated in eight gilts before cervically inseminating under anaesthesia with equal numbers of live-stained and dead spermatozoa. Fluid was collected from eight uterine and three tubal cannulae, the flow beginning within 1½ and 4½ min, respectively, after the onset of insemination. Both the volume of fluid collected and the concentration of spermatozoa in the fluid decreased with time after insemination, whereas the percentage of fluorescent spermatozoa increased. Dead spermatozoa were transported up the reproductive tract, but their transport was less efficient than that of live spermatozoa. Mechanisms of sperm transport are discussed.

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R. D. BAKER and P. J. DZIUK

Summary.

Pilocarpine (50 mg) administered to the boar 20 min before semen collection had no detectable effect on either ejaculation or semen composition. However, the volume of fluid semen, the total volume of semen, and the time required for ejaculation differed significantly (P<0·01) between breeds of boars.

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R. D. BAKER, N. L. VANDEMARK, C. N. GRAVES and H. W. NORTON

Summary.

To determine the effects of parasympathetic-influencing drugs on reproductive phenomena in bulls, four ejaculates were collected at 15-min intervals from each of six bulls beginning 30 min after a subcutaneous injection of 400 mg pilocarpine, 200 mg atropine, or physiological saline solution. Each treatment was administered twice, so that a total of 144 ejaculates was collected. Pilocarpine significantly (P<0·01) decreased the concentration of spermatozoa and increased the time required to mount, the duration of ejaculation, the volume of semen, the number of spermatozoa per ejaculate and the concentration of chloride in the semen. Atropine had the reverse effect on these characteristics and significantly (P<0·01) increased the concentration of fructose in the semen. These results demonstrate that atropine and pilocarpine, which are known to influence the parasympathetic system, alter the reaction time of the bulls, the secretion of one or more accessory sex glands, the passage of spermatozoa through the male reproductive tract, and the emission of semen during ejaculation.

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K S Sidhu, K E Mate, T Gunasekera, D Veal, L Hetherington, M A Baker, R J Aitken and J C Rodger

The phosphorylation of tyrosine residues in cellular proteins is a major signal transduction event during sperm capacitation. In this study protein phosphorylation was monitored using a fluorescein isothiocyanate (FITC)-labeled antiphosphotyrosine monoclonal antibody and a flow cytometric procedure optimized for sperm. Using this technique, the correlation between tyrosine phosphorylation and sperm capacitation was examined in two marsupial species, the brushtail possum and the tammar wallaby and compared with that of ram spermatozoa. The levels of tyrosine phosphorylation in sperm from all three species were increased by the addition of cyclic AMP (cAMP) and vandate, a phosphotyrosine phosphatase inhibitor and were decreased by the addition of the phosphotyrosine kinase inhibitor, staurosporine. Oviductal conditioned media (CM) induced a progressive increase in tyrosine phosphorylation in both marsupial species and also induced morphological transition from a streamlined to a ‘T’-shape configuration in brushtail possum spermatozoa but not in tammar wallaby spermatozoa. Transition to the ‘T’-shape orientation associated with capacitation in marsupial spermatozoa was observed by 2 h of incubation in both species when tyrosine phosphorylation was increased by higher levels of cAMP i.e. 5 mM dibutyryl cAMP plus 3 mM pentoxyphylline. Thus the tyrosine phosphorylation trigger with CM may differ in these two marsupial species. Ram sperm tyrosine phosphorylation could be increased by addition of lower levels of cAMP (1 mM). These results support the finding that tyrosine phosphorylation is associated with sperm capacitation in marsupials. Similar results were obtained by using SDS PAGE/Western blot analysis of tyrosine phosphorylation in the brushtail possum spermatozoa. The specificity, efficiency and sensitivity of the procedure described here make it applicable for routine assessment of capacitation in large numbers of samples and in other species.