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Polyamines in ejaculated ram spermatozoa and their relationship with sperm motility.

Intracellular polyamine levels in ejaculated spermatozoa and seminal fluid from rams were determined by fluorescent spectroscopy of their dansyl derivatives. Relationships between the sperm polyamine content and sperm motility of six mature and eight pubescent rams were studied. Samples were collected from both groups once a month from August through October. Mature rams had a greater percentage of motile sperm cells than lambs (94% versus 73% in September and 92% versus 78% in October); higher spermidine content (36 versus 9 pmol/10(8) cells in September and 162 versus 55 pmol/10(8) cells in October); higher spermine content (984 versus 205 pmol/10(8) cells in September and 1,229 versus 414 pmol/10(8) cells in October); and higher total sperm polyamine content (1,021 versus 216 pmol/10(8) cells in September and 2,258 versus 973 pmol/10(8) cells in October). In the lambs, spermidine content increased (55 versus 9 pmol/10(8) cells); spermine content increased (414 versus 205 pmol/10(8) cells); and total sperm polyamine content increased (973 versus 215 pmol/10(8) cells) in October compared to September. Ejaculates with sperm motility higher than 85% had greater spermine (848 versus 234 pmol/10(8) cells in September and 1064 versus 449 pmol/10(8) cells in October), and total sperm polyamine content (882 versus 244 pmol/10(8) cells in September and 2,015 versus 1,008 pmol/10(8) cells in October) than ejaculates with less than 450 pmol total sperm polyamines/10(8) cells was 68% +/- 6% compared to 90% +/- 4% in cells with greater than 450 pmol (average for all ejaculates) total sperm polyamines/10(8) cells. These data suggest a positive relationship between sperm polyamine constant and sperm motility.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

Caffeine- and kallikrein-induced stimulation of human sperm motility: a comparative study.

Stimulation of human sperm motility by caffeine and kallikrein was compared in the same semen material. Caffeine, a cyclic nucleotide phosphodiesterase inhibitor, induced an immediate stimulation of sperm motility the intensity being relatively constant during the first two hours of incubation. Kallikrein, a kinin-releasing proteinase, induced a similar enhancement of total sperm motility, but showed a delayed type of reaction with maximum stimulation at 2 hours of incubation. In contrast to the effect of caffeine lasting some hours, enhancement of sperm motility induced by kallikrein was observed 24 hours. Simultaneous addition of serum (kininogen source) and kallikrein to semen samples led to a sitmulation of total sperm motility with higher mean values than those obtained by caffeine of kallikrein alone. However, the ratio of spermatozoa with very good forward progression was highest during caffeine stimulation. Simultaneous addition of caffeine and kallikrein led to a further improvement of sperm motility which was significantly above that produced by caffeine or kallikrein alone. This observation and the finding of a different response of the spermatozoa of two ejaculates towards caffeine of kallikrein indicate that caffeine (cyclic AMP) interferes quite differently in comparison to kallikrein (kinins) in stimulating sperm motility.

Caffeine↗

Fertility of a stallion with low sperm motility and a high incidence of an unusual sperm tail defect.

At the beginning of the breeding season an eight-year-old standardbred stallion had semen with virtually zero sperm motility and an approximately 90 per cent incidence of midpiece and tail defects. The motility of the sperm improved to 7 per cent when semen was collected daily but its morphology did not improve. Electron microscopy revealed that the defects consisted mainly of a loss of microtubules in the axoneme and of disorganised midpieces. A pregnancy rate of 24 per cent per cycle and 44 per cent for the season was achieved in 32 mares after the insemination of whole ejaculates collected from the stallion frequently. The fertility was much higher than would have been expected from the characteristics of the semen. It is concluded that this sperm defect, reminiscent of the 'Dag defect' in bulls and the defect in T-locus mice, does not render the animal infertile.

Animals↗

Methods for assessing rat sperm motility.

Computer-assisted sperm analysis (CASA) systems are becoming more widely used. With this spread of technology come more data from toxicology studies, designed to determine if treatment with putative toxicants affects sperm motion parameters. While these CASA methods provide us with more ways to evaluate toxicity and thus perhaps increase our chances of successfully protecting human health, there is also a greater likelihood that different laboratories will use different methods of collecting data on sperm motility. Different systems used with different methods in different laboratories will inevitably generate data that are difficult to compare. In a prospective attempt to address this issue of comparability and limit the problems, a group of individuals using CASA systems to analyze rat sperm motility convened to discuss methodologic issues, share data, and try to reach a consensus about methods for performing these studies. This article shares those meetings and data in the hope that common methods will enhance interlaboratory comparisons.

Animals↗

Intrauterine insemination. Effect of wash media on pregnancy rates and sperm motility.

Testyolk Buffer has been shown to enhance sperm penetration in hamster penetration assays and increase fertilization of oocytes in in vitro fertilization. Based on these findings, we compared pregnancy rates and sperm motility in intrauterine inseminations done with sperm samples washed and resuspended in Ham's F10 as compared with Testyolk buffer. Charts were reviewed retrospectively from 1,098 husband and donor intrauterine inseminations performed at the University of Florida. Data were analyzed using life table analysis and the curves compared with the Mantel-Haenszel statistical test. In addition, sperm motility in fresh sperm was observed in samples incubated in Testyolk or Ham's F10, with motility counts performed at 0, 6 and 24 hours. Four hundred ninety-two Testyolk cycles and 579 Ham's F10 cycles were compared, with cumulative pregnancy rates at one year of 53% and 44%, respectively (P = .58). With donor sperm, 229 cycles with Testyolk and 314 cycles with Ham's F10 had cumulative pregnancy rates of 68% and 48%, respectively (P = .52). With husband insemination, 264 Testyolk and 253 Ham's F10 cycles had pregnancy rates of 37% and 35%, respectively (P = .23). Fresh sperm motility in 22 samples compared at 0, 6 and 24 hours in Ham's F10 (76%, 67.8%, 56.6%) versus Testyolk (76%, 67.7%, 58.8%) revealed no significant differences. There was also no difference in total motile sperm inseminated and postwash motility in 1,098 samples with Testyolk versus Ham's F10. This study demonstrates that there is no enhanced pregnancy rate or increased sperm motility when sperm are treated with Testyolk Buffer instead of Ham's F10.

Adult↗

Mitochondrial disease and reduced sperm motility.

Mitochondrial dysfunction reduces aerobic energy production and results in symptoms from various tissues, depending on metabolic demands. Mitochondrial adenosine triphosphate (ATP) is essential for sperm motility. Sperm motility was investigated in a patient with a mitochondrial disease caused by reduced activity of the mitochondrial enzyme complexes I and IV, and in two control subjects. Spermatozoa were cultured in media containing various energy substrates. Motility was judged by light microscopy, and ultrastructure by transmission electron microscopy. In the patient with mitochondrial disease, 12% of the spermatozoa were motile in the medium containing only glucose. There was a three-fold increase in motile spermatozoa when pyruvate and succinate were present together with glucose. In contrast, the spermatozoa of both control subjects had best motility in the presence of substrates for complex I, and no further increase was observed when succinate was added. Glucose and pyruvate enter the respiratory chain at complex I, and succinate at complex II. Electron microscopy of spermatozoa from the patient with mitochondrial disease revealed mitochondria with increased matrix, thickening of membranes, parallelization of cristae and lipid inclusions, which are characteristic findings in mitochondrial disorders. Abnormal mitochondria were also found in a spermatid, suggesting that the ultrastructural changes of mitochondria are primary rather than secondary to degeneration of the spermatozoa. The results indicate that mitochondrial dysfunction causes reduced sperm motility in some men.

Adenosine Triphosphate↗

Antisera to calreticulin inhibits sperm motility in mice.

Mouse sperm were rapidly immobilized when exposed to rabbit antisera against rat calreticulin. The inhibition of sperm motility was concentration dependent at dilutions 1:50-350. The velocity of sperm did not change significantly as long as they were motile. Neither motility nor velocity of sperm was affected by adding sheep antisera to bovine calmodulin. The antisera to calreticulin also inhibited in vitro fertilization of mouse eggs. Using the avidin-biotin-peroxidase complex method, the antigen was found to be localized in the acrosome of sperm. The results indicate that calreticulin is present in the acrosome of mouse sperm and may play an important role in sperm motility.

Animals↗

ATP concentration of human spermatozoa: lack of correlation with sperm motility.

The primary source of energy for sperm motility is intracellular adenosine 5'-triphosphate. We studied the relationship between the concentration of ATP and sperm motility in normal semen donors and in a series of male infertility patients. The ATP concentration was determined by the firefly luciferin-luciferase assay, and sperm motility was quantitated by the turbidimetric method. Although the patient population demonstrated a significantly lower sperm motility than the donor population, we found no significant difference between the two groups in the intracellular concentration of ATP. Thus, the data indicate that the poor sperm motility observed in some of the patients' specimens was not associated with abnormally low levels of ATP. In separate experiments the intracellular concentration of ATP decreased gradually with time at approximately the same rate at which sperm motility declined. Both factors had decreased by 50% approximately 3 hours after collection.

Adenosine Triphosphate↗

Tctex2-related outer arm dynein light chain is phosphorylated at activation of sperm motility.

When the motility of sperm is activated, only one light chain of flagellar outer arm dynein is phosphorylated in many organisms. We show here that the light chain to be phosphorylated was shown to be light chain 2 (LC2) in rainbow trout and chum salmon sperm and LC1 in sea urchin sperm. Molecular analyses of the phosphorylated light chains from sperm flagella of the salmonid fishes and sea urchin revealed that the light chains are homologs of the mouse t complex-encoded protein Tctex2, which is one of the putative t complex distorters. These results suggest that mouse Tctex2 might also be a light chain of flagellar outer arm dynein and that the abortive phosphorylation of Tctex2/outer arm dynein light chain might be related to the less progressive movement of sperm.

Amino Acid Sequence↗

Derivatives of 2-nitrofluorene cause changes of human sperm motility.

The effects on human sperm motility characteristics of 2-nitrofluorene and selected derivatives were studied in vitro, using computer aided sperm analysis (Cellsoft). Substances to be tested were dissolved in acetone and added to separated spermatozoa in culture media to final concentrations of 100 and 1000 microM. Aliquots were removed immediately (< 5 min.) and 24 hr after the addition and tested for sperm motility characteristics. Four of the substances tested; 2,4,7-trinitrofluoren-9-one (2,4,7-tNFO), 2,5-diaminofluorene (2,5-dAF), 7-hydroxy-2-nitrofluorene (7-OH-NF) and 2,7-diaminofluorene (2,7-dAF) showed strong detrimental effects on the sperm motility. Slight detrimental effects were also recorded using 2-nitrofluorene and 2,5-dinitrofluorene (2,5-dNF). Weak stimulatory effects were obtained using 2-acetoamidofluorene (AAF) and 2,7-dinitrofluorene (2,7-dNF). No significant effects were seen with 5-hydroxy-2-nitrofluorene (5-OH-NF), 2-aminofluorene (AF), 2-aminofluoren-9-one (AFO), 2-amino-9-hydroxyfluorene (9-OH-AF) or 9-hydroxy-2-nitrofluorene (9-OH-NF). The mechanism behind this effect is not known but it could be speculated that these lipophilic substances interact with the membranes or the cellular respiration.

Culture Media↗

Novel parameters of human sperm hypoosmotic swelling test and their correlation to standard spermatogram, total motile sperm fraction, and sperm penetration assay.

OBJECTIVE: To improve the accuracy of human sperm hypoosmotic swelling test by introducing new morphological evaluation parameters for this test. DESIGN: Individual semen samples were processed, and the standard spermatogram, total motile sperm fraction, sperm penetration assay (SPA), and hypoosmotic swelling test were performed. SETTING: Male infertility clinic and andrologic laboratory in a university medical center. PATIENTS: One hundred eighteen subfertile men undergoing an infertility work-up. MAIN OUTCOME MEASURES: The results of hypoosmotic swelling test were subdivided into four groups (hypoosmotic swelling test 1, 2, 3, 4) according to the different shapes of hypoosmotically affected spermatozoa tails. RESULTS: The hypoosmotic swelling test 3 had the best correlation with the different sperm parameters: motility, total motile sperm fraction, concentration, and SPA. The hypoosmotic swelling test 1 showed a satisfactory interrelation for concentration and for total motile sperm fraction but no correlation for motility and SPA. The hypoosmotic swelling test 2 and the hypoosmotic swelling test 4 demonstrated no correlation at all with motility, concentration, morphology and SPA. CONCLUSIONS: The precision of hypoosmotic swelling test may be improved by using the hypoosmotic swelling test 3 type of spermatozoa tail shape as the parameter for a normally functioning spermatozoa.

Female↗

Inhibition of sperm motility and agglutination of sperm cells by free fatty acids in whole semen.

The effects of a serial dilution of linoleic acid on human spermatozoa in whole semen was tested on 21 semen samples obtained from 11 normal volunteers. The minimal concentration of linoleic acid required to stop the movement of at least 75% of the moving sperm ranged from 1 to greater than 100 mg/dl. Fifteen of 21 (71%) of the semen samples were inhibited by added free fatty acids (FFA) concentrations that were less than or close to the physiologic concentration ranges of FFA in blood plasma (1 to 30 mg/dl). The immobilized sperm often formed aggregates similar to those formed by the action of autoantibodies against sperm cells. Preliminary studies conducted on a variety of other FFA have indicated that oleic acid (18/1) was less toxic than linoleic acid (18/2) and that linolenic acid (18/3) was more toxic than linoleic acid. The saturated FFA palmitic acid (16/0) and stearic acid (18/0) at concentrations up to 100 mg/dl showed little or no toxicity to sperm cells. It is suggested that FFA toxicity be included among physiologic factors that affect the motility and spontaneous aggregation of sperm cells.

Ascorbic Acid↗

Change in intracellular K+ concentration caused by external osmolality change regulates sperm motility of marine and freshwater teleosts.

We previously demonstrated that osmolality isotonic to the seminal plasma suppresses sperm motility in marine and freshwater teleosts, and exposure of sperm to hypertonicity of sea water or hypotonicity of fresh water, respectively, induces the initiation of sperm motility at spawning. The motile sperm became immotile by return of osmolality to the isotonic osmolality both in a marine teleost, the puffer fish, and a freshwater teleost, the zebrafish. The initiation and termination of sperm motility could be repeated several times by changing surrounding osmolality in both species. In demembranated sperm, motility was suppressed by a K+ concentration equivalent to the seminal salt concentration in both puffer and zebrafish. Demembranated puffer sperm were reactivated when K+ concentration of the reactivating solution increased. Conversely, initiation of motility in the demembranated zebrafish sperm was induced by decreasing K+ concentration. The initiation and termination of the demembranated sperm were alternately repeated by changing K+ concentration of the reactivation solution in both species. Furthermore, intracellular K+ concentration rose when sperm motility of the puffer was initiated in hypertonic solutions. These results suggest that change in external osmolality is converted into change in intracellular K+ concentration, and that the change affects the flagellar axoneme as a signal to initiate or terminate sperm motility. The initiation and termination of motility in the demembranated puffer sperm were caused at a high pH and a low pH of the reactivating solution, respectively, suggesting the contribution of intracellular pH in the regulation of flagellar motility.

Animals↗

Identification of motile sperm in caput epididymis. Intraoperative observations and clinical correlations.

This retrospective review of patients who underwent vasoepididymostomy demonstrates that motile sperm may be identified in the most proximal portion of the epididymis, and implies that induction of sperm motility may be independent of transit through the epididymis. Although further studies are needed, the data imply that it may not be necessary to identify motile sperm in the epididymal fluid to achieve a successful vasoepididymostomy.

Adult↗

The lack of effect of tumor necrosis factor-alpha, interleukin-1-alpha, and interferon-gamma on human sperm motility in vitro.

Whether cytokines present in human peritoneal fluid reduce sperm motility, and thus contribute to infertility, is investigated. The human recombinant cytokines, tumor necrosis factor-alpha, interleukin-1-alpha, and interferon-gamma, were incubated with motile human sperm obtained from fertile men and separated by the swim-up technique. These cytokines, alone or in combination, in higher doses than those observed in vivo (greater than or equal to 25,000 U/ml), did not alter the percentage of motile sperm after 90 minutes, 24 hours, and 48 hours under standard culture conditions. Similarly, penetration of a column of bovine cervical mucus was unchanged after preincubation of the sperm with individual cytokines or combinations of several cytokines for 24 hours. In contrast to those given in previous reports, these dta do not support a direct effect of tumor necrosis factor-alpha, interleukin-1-alpha, or interferon-gamma on sperm motility, and suggest that other soluble factors are responsible for the observed effects of peritoneal fluid on sperm motility in vitro.

Humans↗

Role of ions and ion channels in the regulation of Atlantic croaker sperm motility.

The effects of the osmolarity and ionic composition of the external media and ion-channel blockers on the induction of sperm motility in the marine teleost, Atlantic croaker (Micropogonias undulatus) were investigated. An in vitro sperm motility assay was developed to determine treatment effects on the percentage of motile sperm, sperm speed, and turning rate, using a computer-aided motion-analysis system. Maximum activation of sperm motility occurred in saline with an osmolarity of 680 mOs/kg. Potassium caused a decrease in the percentage of motile sperm, but only at high nonphysiological concentrations in the presence of high amounts of calcium. Calcium caused an increase in sperm velocity and turning rate. Percent motility was inhibited by the potassium-channel blockers, 4-aminopyridine and veratrine, the calcium-channel blockers, verapamil, diltiazem, and nifedipine; the sodium-channel blocker, amiloride, and the chloride-channel blocker, ethacrynic acid. In addition, the calcium-channel blockers caused a decrease in sperm velocity and turning rate. These results provide evidence for the role of potassium, calcium, and possibly sodium, and chloride ions in Atlantic croaker sperm motility. In addition, this study demonstrates that membrane-bound ion channels are involved in the motility of sperm from a marine teleost.

Animals↗

The effects of the sperm motility activators 2-deoxyadenosine and pentoxifylline used for sperm micro-injection on mouse and human embryo development.

The sperm motility stimulants 2-deoxyadenosine (DOA) and pentoxifylline (PTF), used to improve the success of insemination and sperm micro-injection for low motility sperm samples, were studied for their effects on the developmental capacity of mouse and human oocytes. When human oocytes were micro-injected with spermatozoa in 3 mM DOA 80% of them became blocked at the 1-cell pronuclear stage. However, when spermatozoa in 3 mM PTF were used for micro-injection or when spermatozoa were washed to remove DOA before micro-injection only a few oocytes (9-10%) were blocked. Pregnancies occurred in five of 14 patients into whom cleaving embryos from all three treatments had been transferred, indicating that once cleavage was initiated, development of embryos occurred at expected rates. Exposure of mouse oocytes to DOA for a short period during insemination (4-6 h) or a longer period during the pronuclear cell cycle (18-20 h) significantly reduced cleavage beyond the 2-cell stage, resulting in a dramatic reduction in blastocyst formation. PTF did not significantly reduce mouse embryo development. Similar results were obtained for oocytes inseminated in vitro or micro-injected with a spermatozoon into the perivitelline space. Neither DOA nor PTF increased fertilization of mouse oocytes. PTF reduced fertilization, particularly in cumulus-enclosed oocytes and oocytes micro-injected with spermatozoa in PTF. We conclude that DOA is a potent inhibitor of embryo development and oocytes should not be exposed to DOA. Exposure of oocytes to PTF had little effect on their subsequent development but treatment of cauda epididymal mouse spermatozoa can reduce their fertilizing capacity.

Animals↗