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Bovine spermatozoa have been shown to metabolize exogenous substrates both aerobically and anaerobically (Mann, 1964). Fructose (Mann, 1946) and sorbitol (King & Mann, 1958) have been unequivocally identified in seminal plasma, but evidence for the presence of metabolizable substrates in the cervical mucus rests on much less secure foundations. A number of reports that glucose is present in cervical mucus have appeared and indeed Doyle (1958) proposed to use the presence of glucose in human cervical mucus as an index of ovulation. This test has not attained widespread acceptance although there is a general impression that the sugar is present (see e.g. Moghissi, 1973).

The original authority for the presence of glucose in bovine cervical mucus at oestrus is Olds & VanDemark (1957) who used, as others have done since, measurements of reducing value

Summary.

Measurements were made of the swimming rate of bull spermatozoa at a concentration of 10,000 cells per μl in physiological saline solution and in saline solution buffered with 0·014 m-phosphate at pH values of 6·5, 7·5 and 8·5. The buffers at pH 6·5 and 7·5 gave results similar to those for unbuffered saline solution. At pH 8·5 there was an initial advantage, both in percentage motile and in swimming rate. By the end of 1 hr, the spermatozoa were, however, swimming more slowly, and fewer were surviving than in any other medium. The significance of these results in relation to the swimming of spermatozoa in the female tract is discussed.

Summary.

The swimming rate of bull spermatozoa diluted in various media was estimated by the stop-watch technique. At a dilution of 20,000 spermatozoa per μl in physiological saline solution, the mean of the mean swimming rates of spermatozoa from twenty-seven ejaculates from seventeen bulls was 155·±5·0 μ/sec (range of means 107·0 to 203·2 μ/sec). The rate fell progressively with time during incubation at 37° C. The rates at higher dilutions were progressively lower after only 5 min incubation, and the greater the dilution the less well was the rate maintained, so that the differences between different dilutions increased with time. The percentage motile followed the same pattern as the swimming rate, fewest non-motile being present at the highest cell concentration. Maintenance of rate was slightly better than in saline solution and the dilution effect was diminished, but not abolished, if semen was diluted with seminal fluid (from the same ejaculate), 'sperm extract' or Baker's medium. The rates were slightly higher in the last two media than in saline solution, but in seminal fluid the rate was only about two-thirds that in saline solution.

Spermatozoa swam more slowly in mucus from the uterine cervix of the cow than in any other medium tested. The mean of the mean swimming rates in forty-nine experiments with sixteen ejaculates and samples of mucus from ten cows was 62·7 ± 1·7 μ/sec. The rate could only be related to dilution by counting the spermatozoa present in the mucus at the end of the experiment, and dividing the experiments into categories accordingly. Although the final concentration of spermatozoa ranged from 28 to 115,000/μl, no dependence of swimming rate on concentration could be detected. At all dilutions in mucus, maintenance of rate was about equivalent to that for a 5000/μl dilution in saline solution. About one-half of the spermatozoa which had swum into the mucus were non-motile at the end of 1 hr, at all dilutions, though of those which were initially alive in saline solution at 5000 dilution, only 41% died during the 1st hour, with correspondingly fewer dying at higher cell concentrations. The possibility that the dilution effect on swimming rate might be absent in mucus was also tested by measuring the rates at which spermatozoa swim through a given microscope field of mucus from the moment the first one appeared until a swarm of several hundred had invaded it several minutes later. There was no difference in rate. The impossibility of carrying out an exactly comparable experiment in saline solution was discussed. If spermatozoa were allowed to swim through a strip of mucus into saline solution, the mean rate and frequency distribution of rates in this saline solution resembled those of controls directly diluted in saline solution (to a concentration of 10,000/μl) much more than they resembled the figures for swimming rates in mucus. It was concluded that mucus does not have a 'selecting' effect on spermatozoa with respect to swimming rate.

Summary. Cows of normal reproductive history were treated with progesterone for periods of 2–3 months. The uterine secretions yielded 5 major fractions of macromolecular components. Two of the fractions comprised serum proteins. In the other 3 fractions at least 9 non-serum proteins were observed, 7 in one fraction, but in only small amounts. Of the remaining 2 non-serum proteins one is an acid phosphatase recoverable in small amounts, and the other is lactoferrin and is the major non-serum protein present in the uterine secretions of progesterone-treated cows.

Capacitation is a pivotal event for mammalian spermatozoa, involving the loss of surface proteins known as decapacitation factors (DF) and consequent acquisition of fertilizing ability. Earlier studies showed that a mouse sperm DF binds to a receptor, DF-R, whose attachment to the sperm plasma membrane appears to involve a glycosylphosphatidylinositol (GPI) anchor. In the present study, purification and subsequent sequencing of DF-R has identified this ~23 kDa protein as phosphatidyletha-nolamine-binding protein 1 (PEBP 1). To obtain functional evidence that supports sequence homology data, purified recombinant PEBP 1 and PEBP 2 were evaluated for biological activity. While PEBP 1 was able to remove DF activity in solution at concentrations above ~1 nmol/l, PEBP 2 was ineffective, even at 600 nmol/l; this confirmed that DF-R is PEBP 1. Anti-PEBP 1 antiserum recognized recombinant PEBP 1 and a ~23 kDa protein in both mouse and human sperm lysates. Immunolocalization studies revealed that DF-R/PEBP 1 is located on the acrosomal cap, the post-acrosomal region and the flagellum of both mouse and human spermatozoa, with epitope accessibility being capacitation state-dependent and reversible. Treatment of cells with a phospholipase able to cleave GPI anchors essentially abolished immunostaining, thus confirming the extracellular location of DF-R/PEBP 1. We suggest that DF-R/PEBP 1 plays its fundamental role in capacitation by causing alterations in the sperm plasma membrane in both head and flagellum, with functional consequences for membrane-associated proteins. Obtaining more detail about DF ↔ DF-R interactions could lead to useful applications in both fertility treatments and new contraceptive approaches.

Summary.

Samples of pooled cervical mucus from groups of animals fed melengestrol acetate (MGA) for 14 days were taken 3 and 4 days after cessation of treatment. The mucus samples were separated into protein and glycoprotein portions and the epithelial glycoprotein was isolated. The protein fraction was examined by disc electrophoresis and the epithelial glycoprotein was characterized by chemical analysis and determination of the molecular weight. The cervical mucus of one of the treated groups yielded a glycoprotein which had a reduced sulphate content when compared with that of the controls and did not show the lowered neuraminic acid content characteristic of the glycoprotein from untreated animals at oestrus. Two other groups of MGA-treated animals which received HCG 3 days after withdrawal of the progestagen did not show these anomalies.

The protein in the MGA-treated samples of mucus contained considerably larger amounts of material migrating in the transferrin region of the electrophoretogram than did comparable controls.