Linear Motility Research Articles

Assessment of semen quality in Swamp Buffalo AI Bulls in Thailand

Characteristic of Thai swamp buffalo bulls semen used for artificial insemination (AI) in Thailand, aspects relevance in freezing and thawing of semen are review. Semen and sperm characteristics were evaluated included sperm count, motility (assessed subjectively and by CASA), morphology (using phase-contrast light microscopy and SEM), plasma membrane integrity (PMI) (using a hypo-osmotic swelling test [HOST]) and SYBR-14/propidium iodide [PI]), plasma membrane stability (PMS) (using Annexin-V/PI) and deoxyribonucleic acid (DNA) integrity (using SCSA and flow cytometry [FCM]). The average ejaculate volume was about 3.0–4.0 mL, with good viability (PMI measured by the HOST) and motility (<65% and <70%, respectively). Sperm concentration ranged from 1.1 to 1.2 billion/mL, being also affected by bull age. Whereas semen quality (including sperm output, pH and initial sperm motility) did not differ between the seasons. Few spermatozoa (>15%/ ejaculate) had abnormal morphology with abnormalities resembling those in other bovidae. In FT semen, PMI (using SYBR-14/PI) and PMS were highest in winter. Across seasons, ~50% of post-thaw spermatozoa depicted linear motility, a proportion that decreased to ~35% during incubation (38oC for 60 minutes), without marking any seasonal difference. The sperm DNA was hardly damaged (with <3% fragmentation, expressed as DNA fragmentation index [DFI]), among seasons.

Semen quality and sperm functional parameters in fertile Indian men

The reports of a decline in the reproductive health of men worldwide prompted the present study to be undertaken so that baseline semen parameters could be established in Indian men. Semen profile and sperm function parameters were evaluated in 368 Indian men of proven fertility, settled in Delhi. The results of the study were compared with available published information from Indian men. The mean sperm concentration and semen volumes were 68.22 +/- 15.14 x 10(6) ml(-1) and 3.20 +/- 0.94 ml, respectively. Rapid, linear progressive motility and sluggish linear motility were 40.95 +/- 9.15% and 24.95 +/- 7.01%, respectively. A comparison of the results of the present study with earlier published data did not support the contention of a decrease in the semen quality in Indian men.

Happily at Work

Happily at Work

Chemotaxis of capacitated rabbit spermatozoa to follicular fluid revealed by a novel directionality-based assay.

Precontact communication between gametes is established by chemotaxis. Sperm chemotaxis toward factor(s) in follicular fluid (FF) has been demonstrated in humans and mice. In humans, the chemotactic responsiveness is restricted to capacitated spermatozoa. Here, we investigated whether sperm chemotaxis to factor(s) present in FF also occurs in rabbits and, if so, whether only capacitated spermatozoa are chemotactically responsive. Chemotaxis assays were performed by videomicroscopy in a Zigmond chamber. We measured chemotactic responsiveness as a function of FF dilution by means of a novel directionality-based method that considers the ratio between the distances traveled by the spermatozoa both parallel to the chemoattractant gradient and perpendicular to it. A peak of maximal response was observed at 10(-4) dilution of FF, resulting in a typical chemotactic concentration-dependent curve in which 23% of the spermatozoa were chemotactically responsive. In contrast, the percentage of cells exhibiting FF-dependent enhanced speed of swimming increased with the FF concentration, whereas the percentage of cells maintaining linear motility decreased with the FF concentration. The percentages of chemotactically responsive cells were very similar to those of capacitated spermatozoa. Depletion of the latter by stimulation of the acrosome reaction resulted in a total loss of the chemotactic response, whereas the reappearance of capacitated cells resulted in a recovery of chemotactic responsiveness. We conclude that rabbit spermatozoa, like human spermatozoa, are chemotactically responsive to FF factor(s) and acquire this responsiveness as part of the capacitation process.

Effect of buffering systems on post-thaw motion characteristics, plasma membrane integrity, and acrosome morphology of buffalo spermatozoa.

This study was carried out to identify the suitable buffer for cryopreservation of buffalo semen. Semen was collected with artificial vagina (42 degrees C) from four buffalo bulls. Split pooled ejaculates (n=5), possessing more than 60% visual sperm motility, were extended at 37 degrees C either in tri-sodium citrate (CITRATE), Tris-citric acid (TCA), Tris-Tes (TEST) or Tris-Hepes (HEPEST). Semen was cooled to 4 degrees C in 2 h, equilibrated at 4 degrees C for 4 h, filled in 0.5 ml straws and frozen in a programmable cell freezer before plunging into liquid nitrogen. Thawing of frozen semen was performed after 24 h at 37 degrees C for 15 s. Sperm motion characteristics, plasma membrane integrity, and acrosome morphology of each semen sample were assessed by using computer-assisted semen analyzer (CASA), hypo-osmotic swelling (HOS) assay, and phase-contrast microscope, respectively. Analysis of variance revealed that percent post-thaw visual motility tended (P=0.07) to be higher in HEPEST (61.0+/-2.9) and lowest in CITRATE (48.0+/-2.5). Computerized motility did not vary due to buffering system. Percent post-thaw linear motility tended (P=0.09) to be higher in TCA (78.2+/-5.5) and lower in TEST (52.0+/-6.9). Circular motility (%) was significantly lower (P<0.05) in TCA (11.6+/-2.8) and higher in TEST (29.8+/-5.6). Curvilinear velocity (microm s(-1)) was lower (P<0.05) in TCA (69.4+/-2.0) than in CITRATE (79.0+/-5.8), TEST (87. 2+/-1.6) and HEPEST (82.6+/-3.0). Lateral head displacement (microm) was lowest (P<0.05) in TCA (1.7+/-0.2) and highest in TEST (3.7+/-0. 6). Plasma membrane integrity and normal acrosomes of buffalo spermatozoa did not differ due to buffering system and averaged 40. 0+/-2.7% and 61.4+/-4.6%, respectively. Based upon lower circular motility, curvilinear velocity, and lateral head displacement, it is concluded that post-thaw quality of buffalo semen can be improved using the Tris-TCA buffering system.

Diurnal variation of semen quality in human males.

The possibility of a diurnal variation in semen quality was tested in 54 human males attending our infertility clinic. Of the enrolled subjects, 24 were normozoospermic and 30 were suffering from oligo- and/or asthenozoospermia. Seminal fluid was collected by masturbation twice by each subject, once in the morning (7:00-7:30 a.m.) and once in the afternoon (5:00-5:30 p.m.). Abstinence from sexual intercourse for 3-4 days before each of the two collections was requested. Semen parameters were evaluated independently by two biologists before and after pellet swim-up. Beside similar macroscopic parameters, specimens collected in the afternoon showed a higher number (P < 0.01) and concentration (P < 0.01) of spermatozoa. Also, immediately (P < 0.05), and at 1 h (P < 0.02) and 2 h (P < 0.01) after pellet swim-up, the number of spermatozoa with progressive linear motility was higher in the afternoon than in the morning. These data are the first documenting a diurnal rhythm in sperm quality which may contribute to the reported variability in semen parameters, and may prove useful for spontaneous and assisted conceptions.

Sperm characteristics and zona pellucida binding in relation to field fertility of frozen–thawed semen from dairy AI bulls

The present study examined the relationship between bull sperm characteristics immediately post-thaw and some characteristics registered after swim-up, including the ability of spermatozoa to bind to homologous zona pellucidae (ZP) in vitro, and fertility after artificial insemination (AI) of 9426 females. Frozen-thawed semen from 22 AI bulls of the Swedish Red and White Breed, represented by 43 different frozen batches (1-4 batches/bull, 2 consecutive ejaculates/batch), was examined with the aim of determining concentration, motility patterns, morphology and membrane integrity. In addition, the frozen-thawed spermatozoa were subjected to a swim-up procedure and those separated in this way were tested with two assays of sperm-binding to the ZP of homologous oocytes in vitro (ZBA), using either a relative ZBA index against a control bull of proven high fertility or absolute binding (Absolute ZBA). The correlations of the various sperm traits and 56-day non-return rates (NRR) after field AI were retrospectively examined as single traits and as combinations of traits (combined measures), including regression analysis of significant traits. Among the sperm characteristics, positively significant (p < 0.01) correlations with NRR were found for linear motility post-thawing (r = 0.45-0.59) and the concentration of motile spermatozoa after swim-up (r = 0.43-0.63). Results obtained with the absolute ZBA approach were significantly (p < 0.05) correlated with NRR (r = 0.50), whereas the correlation between NRR and the ZBA index was not significant. The use of combined measures of sperm traits, including the ability to bind to ZP, showed a stronger predictive correlation with NRR (r = 0.68-0.75), compared with single traits. The results suggest that the combined analysis of sperm linear-motility patterns, swim-up separated sperm motility and absolute ZBA can provide a valuable assessment of the fertilizing capacity of AI bull semen.

Sperm from Spinal Cord Injured Men Lose Motility Faster than Sperm from Normal Men: The Effect is Exacerbated at Body Compared to Room Temperature

Purpose We investigated if sperm from spinal cord injured men lose motility and viability faster than sperm from normal men, and if there is an effect of temperature on these parameters. Materials and Methods Identical aliquots of fresh antegrade semen from 10 spinal cord injured men and 9 normal controls were stored at room temperature (23C) or body temperature (37C). Sperm motility and viability were measured at 0, 2, 4, 6 and 8 hours after semen collection. Results Sperm motility, particularly rapid linear motility, decreased faster in specimens from spinal cord injured men compared to normal controls. This effect was exacerbated at 37C. Heat did not affect the degradation of sperm motility in specimens from normal men. The percent of dead sperm in specimens from spinal cord injured men was significantly greater than that in normal men at all time points. However, the rate of cell death was not faster than normal with time or heat. Conclusions When using sperm from spinal cord injured men for purposes of insemination or experimentation the rapid loss of motility must be considered, particularly when stored at body temperature.

Evaluation of the effect of the absence of sperm with rapid and linear progressive motility on subsequent pregnancy rates following intrauterine insemination or in vitro fertilization.

The objective of this study was to investigate the association of rapid and linear progressive motility in seminal and Percoll-separated sperm with the outcome of intrauterine insemination (IUI) and in vitro fertilization (IVF) cycles. Motility was graded using the qualitative system proposed by the World Health Organization: grade A, rapid and linear, grade B, slow or nonlinear; grade C, non-progressive; or grade D, nonmotile. Absence of rapid and linear motility was defined as grade A sperm absent. Nine-hundred-fifty IVF and 1,448 IUI cycles were analyzed. In 7.9% (75) of the IVF cycles, grade A sperm were absent in the semen. Although the mean fertilization rate was lower in the absence of grade A sperm in the semen (44.5% vs. 63.4%, P < 0.05), the pregnancy rates were similar irrespective of their presence or absence (18.7% vs. 17.8%). In the cycles in which grade A sperm were absent following Percoll separation (26/950; 2.7%), the fertilization rate (29% vs. 62.8%) and the clinical pregnancy rate/retrieval were significantly lower (3.8% vs. 18.3%, P < 0.05). In 26.4% (382) of the IUI cycles, grade A sperm were absent in the semen and conception occurred in 30 (7.9%), compared to a pregnancy rate of 10.4% in the group with grade A sperm present in the semen. Following Percoll separation, only a 2.5% (2/80) pregnancy rate was observed in the group with no grade A sperm, compared to 10.2% in the group with grade A sperm (P < 0.05). The absence of rapid and linear motile sperm in the Percoll-separated sperm significantly reduced fertilization rates in vitro and pregnancy rates in both IUI and IVF cycles. The use of the total number of grade A sperm was also effective in predicting reduced fertilization in IVF and reduced pregnancy rates in IUI, but no better than the use of the mere presence/absence of grade A sperm. In a clinical situation, the simpler test is preferable. This type of evaluation is available to all centers as opposed to the more expensive computer-assisted semen analysis.

Relevance of acrosome function in the evaluation of semen in vitro fertilizing ability

To determine whether or not acrosome evaluation can enhance the prediction of IVF results when associated to conventional semen parameters. Acrosome reaction, sperm concentration, motility, and morphology were recorded in 131 semen samples from patients undergoing an IVF attempt. Spontaneous acrosome loss after a 24-hour incubation in B2 medium and after induction by calcium ionophore A23187 and phorbol ester, phorbol 12-myristate 13-acetate 4-O-methyl ether (TPA). Statistically significant differences between fertilization failures and successes were found for concentration, viability, spontaneous and induced acrosome reaction, and most parameters of motility and morphology. However, none of the parameters could predict > 64% of IVF results when studied alone. A progressive discriminant analysis allowed to predict up to 83% of IVF results, by classifying sperms through their normal forms, rapid motility, spontaneous acrosome loss, enlarged heads, multiflagellar forms, vitality, linear motility, and acrosome response to TPA. The other parameters, including concentration and response to calcium ionophore, had no additive value. The study of acrosome function, through spontaneous acrosome loss and response to TPA, is of great interest in clinical practice when associated to some parameters of motility and morphology. However, it appears that response to calcium ionophore, one of the most studied parameters, is of poor practical interest.

Exposure of human spermatozoa to the cumulus oophorus results in increased relative force as measured by a 760 nm laser optical trap.

Spermatozoa change their movement characteristics in response to different environmental conditions. To investigate the relative force of spermatozoa exhibiting different motility patterns, a laser optical trap was used. A laser beam at 760 nm was directed through a microscope objective and focused above the spermatozoa to create a three-dimensional optical trap. Spermatozoa were trapped at 300 mW, and laser power was reduced until spermatozoa could escape. The force generated by the flagellar movement was proportional to the laser power at which the spermatozoa escaped from the trap. Three motility patterns were studied: linear, hyperactivated, and cumulus-related. Mean escape power for spermatozoa displaying linear motility was 59.5 +/- 43 mW, for hyperactivated motility 122.3 +/- 67 mW (P < 0.0001) and for cumulus-related motility 200.6 +/- 44.2 mW (P < 0.0001). In this study, we showed that human spermatozoa generated more relative force upon exposure to the cumulus mass. The combination of small-amplitude lateral head displacement and higher relative force may produce a 'drilling' effect which is synergistic with the enzymatic digestion of the cumulus matrix during the fertilization process.

Serum hormone levels affect sperm function

If hormone levels affect sperm motility during capacitation, then the serum added to samples prepared for artificial insemination could affect sperm fertilizability. We investigated various sperm functional movement characteristics (percent motile, progressive velocity, linearity, beat cross frequency, lateral head displacement, longevity, and hyperactivation) in specimens incubated with women's sera, as well as exogenous hormone preparations of estradiol (E2) and progesterone (P). Early follicular phase serum (low E2 and low P) maintained motility and longevity. Sperm in E2-treated medium exhibited higher progressive velocity, linear motility, and longevity. In contrast, the percent motility decreased as sperm exhibited hyperactivated motility with P.

Differential Response of Longitudinal and Circular Muscles of Intact Canine Colon to Morphine and Bethanechol

Strain transducers were permanently implanted on the proximal and distal portions of the colon of each of 12 healthy female beagles. This allowed the in vivo measurements of isometric muscle contraction in the healthy dog. The responses of the longitudinal and circular muscles as well as the proximal and distal portions of the colon were compared. Morphine and bethanechol were the stimuli. Morphine did not change the motor activity or tension of the longitudinal muscle but did increase the motor activity and tension of the circular muscle. Bethanechol increased the motor activity and tension of both longitudinal and circular muscles. This was true of both proximal and distal colon. Bethanechol caused a greater motor response of the longitudinal muscle of the distal than of the proximal colon. Both drugs caused a greater increase in the tension of the circular muscle of the distal than of the proximal colon. The linear displacement and motility index methods were both used to measure motility, because it was not clear from the literature which was preferable. Both methods were equally sensitive to change in motility. The linear displacement method had a smaller coefficient of variation and for this reason was preferable. The findings raise some interesting possibilities for studies of propulsion in the colon.