Digestive tract measurements and histological adaptation in broiler lines divergently selected for digestive efficiency.

Two lines of broilers divergently selected for a high (D+) or a low (D-) AME(n) on a wheat-based diet were studied for morphological and histological characteristics of the digestive tract. A total of 630 birds of both lines were slaughtered after a 23-d feeding period. Digestive tract morphology and intestinal histology were investigated on a total of 24 birds to describe the consequences of divergent selection. Birds of the D+ line had 34% heavier gizzards (P < 0.001) and 22% heavier proventriculi than their D- counterparts. In contrast, intestines were 15 to 40% heavier in D- birds, mainly in the jejunum (P < 0.001) and ileum (P < 0.001). Intestinal segments were also longer (between 3 and 6%) in the D- birds. Intestinal villi were larger and longer in D- birds (P < 0.001), mainly in the jejunum (14 to 16%), and crypts were 10 to 15% deeper for the 3 intestinal segments in D- birds (P < 0.001). Muscle layers of the intestine were 17 to 24% thicker (P < 0.001) and goblet cells were 27 to 34% more numerous in the jejunum and ileum of D- birds (P = 0.027). This new characterization of the 2 lines shows that divergent selection based on AME(n) modified the morphology of the proventriculus and gizzard, suggesting greater activity of this compartment in D+ than in D- birds. Intestinal adaptation revealed by visceral organ weight and length and histological modifications in D- birds can be viewed as an attempt to compensate for the low functionality of the gastric area.

Protection from cytotoxic effects induced by the nitrogen mustard mechlorethamine on human bronchial epithelial cells in vitro.

The present study was undertaken to find potent molecules against the toxicity of nitrogen mustard mechlorethamine (HN2) on respiratory epithelial cells, using a human bronchial epithelial cell line (16HBE14o-) as an in vitro model. The compounds examined included inhibitors of poly(ADP-ribose) polymerase (PARP), sulfhydryl-group donors as nucleophiles, and iron chelators and inhibitors of lipid peroxidation as antioxidants. Their effectiveness was determined upon observance of metabolic dysfunction induced by HN2 following a 4-h exposure, using (3-(4,5-dimethylthiazole-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) reduction and ATP-level assays as indicators. Moreover, the fluorescent probe, monobromobimane (mBBr), and 2',7'-dichlorofluorescin-diacetate (H2DCF-DA) were used to assess intracellular sulfhydryl and peroxide level modifications by flow cytometry, respectively, following a 3-h exposure. At last, cell death was assessed by flow cytometry using the propidium iodide (PI)-dye-exclusion assay following 24-h exposure. PARP inhibitors (niacinamide, 3-aminobenzamide, 6(5H)-phenanthridinone), and two sulfhydryl-group donors (N-acetylcysteine, WR-1065) were found to be effective in preventing HN2-induced metabolic dysfunction when added in immediate or delayed treatment with HN2. Only N-acetylcysteine, however, was found to prevent cell death induced by HN2, though it must be present at the time of the HN2 challenge. Flow cytometric measurements of intracellular sulfhydryl levels strongly suggested that N-acetylcysteine and WR-1065 are preventive in alkylation of cellular compounds, mainly by direct extracellular interaction with HN2. PARP inhibitors prevent secondary deleterious effects induced by HN2, considering metabolism dysfunction as the endpoint. Elsewhere, the oxidative stress appears to be a side effect in HN2 toxicity only upon considering the inefficiency of several antioxidants.

Effects of hydroxyl radicals on outwardly rectifying chloride channels in a cultured human bronchial cell line (16HBE14o-).

Respiratory pathologies can result from the exposure of airway epithelial cells to oxidative stress. We studied the effects of the hydroxyl radical *OH, for which there is no natural intra- or extracellular scavenger, on an outwardly rectifying chloride channel (ORCC). In the human bronchial cell line 16HBE14o-, the cytoplasmic side of ORCC in inside-out excised membrane patches was exposed to *OH created by simultaneously superfusing Fe2+ and H2O2 in front of the patch-pipette. ORCC was activated by depolarizing voltage steps. Its open probability (Po) increased with bath [Ca2+] above 1 microM. Upon brief exposure to *OH, ORCC first closed and then alternated between periods of closure and normal activity. The duration of closure increased with the duration of *OH exposure but voltage steps could reopen the channel. After 10 min exposure to *OH, however, the channel closed irreversibly, regardless of the number of subsequent voltage steps or the duration of washing. Low [Ca2+] in the bath accelerated the irreversible closure of the channel in the presence of *OH. Intracellular application of *OH progressively inhibited ORCC activity by inducing long closure periods that increased with time. This might have important pathophysiological implications in the process of inflammation.

Effects of extracellular environment on the osmotic signal transduction involved in activation of motility of carp spermatozoa.

The mechanism by which a hypo-osmotic shock activates motility of carp spermatozoa was studied. The direct role of osmolality at the axoneme was investigated after demembranation of spermatozoa with Triton X-100 and reactivation in various ionic or anionic solutions containing Mg-ATP: demembranated spermatozoa remain motile in solutions of osmolality up to 550 mOsm kg-1 while non-demembranated spermatozoa are immotile when osmolality rises above 250 mOsm kg-1 with the same salt solutions as well as in non-ionic solutions. Suspension in hypo-osmotic saline solutions triggered the swelling of native carp spermatozoa. No motility or swelling occurred above 200-300 mOsm kg-1 and this osmolality is probably that of the cytosol. The swelling of carp spermatozoa is the result of an entrance of water but this was not affected by pCMBS, an inhibitor of the aquaporin CHIP28, or by various inhibitors of the co-transport of water with ions. Various pharmacological agents that affect the motility of different sperm species had no effect on carp sperm motility when used under similar conditions. However, prolonged exposure to a solution devoid of K+ or Cl- affects the activation of motility in a reversible manner, suggesting that these ions have a role in the perception or transduction of the osmotic signal. Altering the concentration of intracellular second messengers such as Ca2+ and cAMP, and the pH did not affect the motility of carp spermatozoa. However, DMSO at 1-20% (400-3200 mOsm kg-1) affects the motility of carp spermatozoa 3-4 min after mixing. These results show that the activation signal of carp sperm motility differs from that known for spermatozoa of other species of fish such as trout. Our results indicate that the activation mechanism may involve a co-transport of ions or specific 'stretch-activated channels' that are sensitive to osmotic pressure.

Role of free L-carnitine and acetyl-L-carnitine in post-gonadal maturation of mammalian spermatozoa.

Spermatozoa are produced in the testis and undergo post-gonadal modifications in the epididymis to acquire fertilizing ability. In epididymal plasma, high-molecular-weight proteins and such small molecules as free-L carnitine convert the gametes into "competent' and functional cells. This review summarizes the knowledge pertaining to L-carnitine and the significance of free L-carnitine uptake into the mature spermatozoa of mammals. We provide an overview of the function of free L-carnitine and carnitine esters in the metabolism of eukaryotic cells and review the role of the specific carnitine acyltransferases in mitochondrial transport of fatty acids and in modulating acyl-coenzyme A (CoA) pools in cellular organelles. In mammals, including man, free L-carnitine is taken from blood plasma and concentrated in the epididymal lumen. This epididymal secretion is beneficial for spermatozoa and is not merely an excretory waste. The uptake of free L-carnitine into the spermatozoa and its metabolic outcome are discussed first in in-vivo and then in in-vitro situations. Free L-carnitine goes through the sperm plasma membrane by passive diffusion. Free L-carnitine is acetylated in mature spermatozoa only. The excess acetyl-CoA from the mitochondria is probably stored as acetyl-L-carnitine and modulates the reserves of free CoA essential to the function of the tricarboxylic acid cycle. These properties of L-carnitine of buffering CoA in the mitochondrial matrix are known in somatic cells but are accentuated in this study of the male germinal cells. In the future, a precise measurement of the in-vivo and in-vitro concentrations of free CoA and acetyl-CoA in the cellular compartments of immature and mature spermatozoa might complete these data. The relationship between the endogenous pools of free and acetylated L-carnitine and the percentage of progressive sperm motility indicates a more important metabolic function related to flagellar movement. In conclusion, the potential to initiate sperm motility, which takes place in the epididymis, is probably independent of the carnitine system, while the energy properties of acetyl-L-carnitine can only be relevant in situations of "energy crisis'. The uptake of "cytoplasmic' free L-carnitine in mature spermatozoa must be a protective form of mitochondrial metabolism, useful to the survival of this isolated cell.

Morphological and kinetic changes of carp (Cyprinus carpio) spermatozoa after initiation of motility in distilled water.

Studies on the flagellar movement of carp spermatozoa induced by dilution in distilled water allowed us to describe a sequence of early, rapid morphological and kinetic changes which begin at the very tip of the flagellum. They cause the progressive folding of the axoneme which ends stuck to the head within 90-120 seconds after the initiation of motility. However, the axonemal machinery remains functional as the folding can be reversed after transfer back into a high osmolality medium and partially folded flagella were able to propagate efficient waves along the non-folded proximal portion of the axoneme. The data also revealed that the membrane area of the terminal piece exhibits strong sensitivity to hypotonicity. These results suggest that in the normal freshwater medium, the brief swimming period allowing fertilization of oocytes is limited by the osmotic stress induced coiling of the carp sperm tail and not by ATP stores.

Changes in flagellar movement of rat spermatozoa along the length of the epididymis: manual and computer-aided image analysis.

The different patterns of motility of rat spermatozoa during epididymal transit were studied in vitro using high-speed videomicroscopy. The sperm images were analysed after manual tracing as well as with a computer imaging system. The present work is the first which reports both the swimming path of the sperm head and the characteristics of flagellation in this species. The hook-shaped head of the rat spermatozoa allowed us to demonstrate the two-dimensional (2D) swimming movement compared to the three-dimensional (3D) sperm motion which was mainly related to rotation of the head. Immotile spermatozoa entered the initial segment of the testis and showed rigid flagella. The potential for sperm motility occurred abruptly in the proximal caput region, and different patterns of flagellation were observed: vibrating, motile in place, motile with a static curvature of the midpiece resulting in a spinning motion or a circular path, and forward progressive movement with regular rotation of the head. The pattern of sperm movement became homogeneous in the distal cauda where the whole sperm population swam in a straight line. A static curvature appeared in the midpiece portion when the spermatozoa reached the proximal caput region. The formation of the static curvature was observed on both sides of the rat flagellum which were easily indicated by the head-shaped projection of the head and the axonemal side of the principal wave. As soon as they moved, the spermatozoa successively initiated principal (P) and reverse (R) waves, but the waves were visible only distal to the static curvature. The midpiece stiffness progressively decreased during the epididymal maturation; simultaneously the static curvature showed a larger radius and then disappeared. Consequently, the initiation of waves which was first seen in the distal part of the flagellum of immature cells occurred progressively near the junction with the head of maturing spermatozoa. These changes in sperm motion previously shown in rams and now in rats might be a general phenomenon in mammals. The high resolution of this computer imaging system applied tosperm motion showing a well-characterized "side of the flagellum" should allow sensitive detection of biochemical effects on flagellar beating.

Relationship between sperm ATP content and motility of carp spermatozoa.

Carp spermatozoa are immotile in seminal plasma or in saline solution of high osmolality (> 400 mosmol kg-1). These 'quiescent' spermatozoa initiate a progressive forward motility when transferred in freshwater or in saline solution with low osmolality (< 160 mosmol kg-1). In this study we investigated 'in vitro' the relationship between sperm ATP content (measured by bioluminescence) and sperm motility (analysed by videomicroscopy). Sperm ATP content remained high in the immobilizing medium (200 mM KCl, Tris 30 mM, pH 8.0) where no flagellar movement occurs. Dilution of these spermatozoa in the activating medium (45 mM NaCl, 5 mM KCl, Tris 30 mM, pH 8.0) triggered forward motility which varied with temperature. At 20 degrees C, sperm ATP content decreased rapidly during the progressive forward motility phase from 12 to 4 nmol/10(8) spermatozoa, concomitantly with decreases in velocity (130 to 10 microns s-1) and the beat frequency (50 to 7 Hz). An inhibitor of mitochondrial respiration (KCN 10 mM) produced a drop in sperm ATP content irrespective of the incubation medium (activating or immobilizing). A second phase of sperm motility in the activating medium was induced following a previous transfer of spermatozoa into a medium of high osmolality for a few minutes prior to the second phase. Within 10 minutes, spermatozoa recover 90% of the initial ATP level as well as forward motility. These results suggest that motility of carp spermatozoa depends on sperm ATP synthesized by mitochondrial respiration mainly stored before activation. In low osmolality conditions, the mitochondrial oxidative phosphorylation is unable to compensate for the ATP hydrolysis required to sustain motility.(ABSTRACT TRUNCATED AT 250 WORDS)

In vitro exposure of rabbit tracheal epithelium to SO(2): Effects on morphology and ciliary beating.

The aim of this in vitro study was to characterize the direct effects of short-term exposure to low concentrations of sulfur dioxide (SO(2)) on both the morphology and the physiology of rabbit tracheal primary cultures. Scanning electron microscopy (SEM) studies revealed that ciliated cells exposed for 1 hr to 10 ppm or 30 ppm SO(2) exhibited aggregated cilia. Transmission electron microscopy revealed numerous swollen mitochondria in cells exposed to 30 ppm SO(2) for 1 hr. This morphological damage to cells was coupled with physiological alterations. A 25% decrease in ciliary beat frequency (CBF) was measured in cells exposed to 30 ppm SO(2). This inhibition was partially reversible within 24 hr. This SO(2) concentration also induced a significant depletion of cellular ATP content which was completely restored after a 24-hr recovery period. A correlation was found between cellular ATP level depletion and CBF decrease.

Uptake and release of free L-carnitine by boar epididymal spermatozoa in vitro and subsequent acetylation rate.

In the male reproductive tract, very high concentrations (mmol l-1) of free L-carnitine and acetyl-L-carnitine are found in the epididymides, seminal plasma and spermatozoa. It has been reported that the uptake of free L-carnitine by spermatozoa might be related to the epididymal maturation of the sperm membrane, since a greater uptake was found by caput than by cauda spermatozoa in vitro. However, the free L-carnitine concentrations estimated inside the gametes were never greater than those of the surrounding medium. In this study, we investigated the mechanism of transport of free L-carnitine and its ester acetyl-L-carnitine, through the plasma membrane of mature and immature epididymal boar spermatozoa. In vitro, we found a passive diffusion of both compounds to the spermatozoa, whatever the maturation stage. The spermatozoa might progress in the epididymal lumen and accumulate high amounts of free L-carnitine. The active uptake of free L-carnitine occurs only across epididymal mucosa. These results are in agreement with those reported on cells of other organs that exchange pharmacological free L-carnitine concentrations (mmol l-1) by a passive mechanism through the plasma membrane. The acetylation of high amounts of free L-carnitine inside the spermatozoa was found only in caudal spermatozoa. This result suggests that oxidative metabolism (producing acetyl CoA) might be more active in mature cells. The acetyl-L-carnitine added to the incubation medium of boar spermatozoa was hydrolysed. Enzymatic activity of the sperm membrane is low and this may partially explain the low concentrations of acetyl-L-carnitine found in the caudal epididymal plasma.

Reversible intracellular ATP changes in intact rat spermatozoa and effects on flagellar sperm movement.

The initiation of motility and modification of energy metabolism of rat caudal epididymal spermatozoa can be induced by dilution in a saline medium. We have investigated in these cells the relationships between the energy reserve (sperm ATP content measured by bioluminescence) and flagellar movement (high speed videomicrography, 200 frames/sec). A steady state was observed in sperm ATP content, progressive velocity (Vp) and flagellar beat frequency (F) with sperm dilution in a medium with glucose, lactate, pyruvate and acetate substrates after 30 minutes of incubation. Without these substrates, changes in metabolic pathways occurred immediately and initially disturbed the relationship between ATP levels and F, suggesting differences in motility initiation when energy is from an endogenous origin via mitochondrial oxidative phosphorylation. This "energy crisis" was reversed by the addition of substrates to the medium. The three-dimensional flagellar movement observed in the presence of substrates quickly became two-dimensional in their absence. The flagellar beat envelope became more splayed, the mean amplitude of lateral head displacement increased and F decreased. The resulting high flagellar beat efficiency can be compared to that observed during hyperactivation which is a physiological event related to a fall in intracellular ATP level. In both media, the displacement of the flagellum in relation to the wave axis varied sinusoidally. The sine period increased with time when the spermatozoa were incubated in the medium without substrates. These results suggest a gradual slowing-down of the velocity of wave formation in the proximal part of the flagellum.

Catalase activity in human spermatozoa and seminal plasma.

Catalase activity was determined in human semen by measuring the oxygen burst with a Clark electrode, after H2O2 addition. Significant catalase activities (mean +/- SD) were found in migrated, motile spermatozoa (44 +/- 17 nmoles O2/min/10(8) cells) and in seminal plasma of normozoospermic men (129 +/- 59 nmoles O2/min/ml). It has been demonstrated that seminal catalase originated from prostate; however, its activity was not correlated with the usual prostatic markers (such as citric acid and zinc). Our data suggest a multiglandular function secreted by this organ. The catalase activities measured in seminal samples from asthenozoospermic, infertile men were found lower than those from normozoospermic subjects. The understanding of the relative contribution of the different enzyme systems against O2 toxicity (superoxide dismutase, catalase, glutathione peroxidase) seem to be a priority area of research to understand disturbances of sperm function.

In-vitro processing of sperm with autoantibodies: analysis of sperm populations.

Ejaculates from 51 infertile men with significant levels of anti-sperm antibodies were submitted to processing in vitro in order to increase the number of antibody-free spermatozoa. Rapid removal and washing of spermatozoa from antibody-containing seminal fluid resulted in a variable decrease in IgG and/or IgA binding; only IgG binding was significantly reduced. Those spermatozoa were allowed to swim-up into an overlaying medium to select those with the best progression. Unexpectedly, in the post-migration samples (PM), the proportion of progressively motile spermatozoa coated with IgG and IgA antibodies increased significantly. An efficient selection of antibody-free spermatozoa was achieved prior to swim up migration by immuno-binding to polystyrene Petri plates coated with anti-human immunoglobulin antibodies. Non-adherent populations were depleted of 27-100% of IgG and IgA antibody-coated spermatozoa. After migration, the percentages of antibody-coated spermatozoa of those populations (PMP) remained low. The comparison between PM and PMP populations shows that immunobinding increases the number of motile antibody-free spermatozoa. The fertility potential of both sperm populations is currently under investigation.

In-vitro processing of sperm with autoantibodies and in-vitro fertilization results.

In-vitro fertilization and embryo transfer were carried out in twenty infertile couples with significant levels of anti-sperm antibodies on the ejaculated spermatozoa. Over 21 cycles (15 couples), 95 mature oocytes were collected and inseminated with spermatozoa obtained by swim-up migration after rapid dilution and washing of the ejaculates. An overall fertilization rate of 38.9% was obtained with these post-migration (PM) preparations. When greater than 70% of the inseminated spermatozoa were covered with both IgG and IgA antibodies, only 14% of the 43 oocytes were fertilized. A higher fertilization rate was obtained with the PM preparations containing less than 70% of spermatozoa coated with one or both classes of antibodies. Under these conditions 60% of the 52 oocytes were fertilized. Fertility rate correlated better with IgG than with IgA antibody levels. In order to decrease the proportion of antibody-coated spermatozoa in the inseminated populations, washed spermatozoa were immuno-adsorbed on Mage's plates before swim-up migration. Over 11 cycles (10 couples) 52 mature oocytes were inseminated with these post-migration immuno-depleted sperm preparations (PMP) containing less than 65% of antibody-coated spermatozoa: 31% of the oocytes were fertilized. This rate compares favourably to IVF results obtained with the PM preparations with greater than 70% of spermatozoa coated with both classes of antibodies. In five couples who had different IVF attempts with the two sperm preparations, the immuno-depletion resulted in a slight increase in the fertilization rates: 10% for the PM preparations versus 26% for the PMP preparations. Sperm binding to the zona pellucida was decreased in the majority of unsuccessful attempts.(ABSTRACT TRUNCATED AT 250 WORDS)

[Acetylcarnitine and spermatozoa: relationship with epididymal maturation and motility in the boar and man ]. / Acétylcarnitine et spermatozoïdes: relation avec la maturation épididymaire et la mobilité chez le verrat et l'homme.

The level of carnitine and acetylcarnitine in spermatozoa of boar epididymal origin and of human ejaculates was demonstrated. In the epididymal fluid of boars, the concentration of carnitine (nmol/mg protein) began to increase from 20 in the distal caput to rise progressively to 700 in the distal cauda. By contrast, the carnitine content of spermatozoa only started to increase in the proximal cauda where the concentration of carnitine in the fluid was 200-300 nmol/mg protein, then gradually increased in spermatozoa from more distal sites. The increase in the acetylcarnitine content of spermatozoa paralleled that of the carnitine amount, represented 50% of total carnitine (carnitine + acetylcarnitine) and coincided with the acquisition of progressive motility. In two populations of human seminal spermatozoa selected by migration and characterised by a very large difference in their percentage of progressively motile cells, higher carnitine and acetylcarnitine contents (40%) were found in migrated spermatozoa compared to the residual population. These results suggest that accumulation of carnitine and its metabolite may be an important factor in the acquisition and the maintenance of progressive motility. Measurement of acetylcarnitine content of human seminal spermatozoa could be used as a marker of epididymal maturation.

The distribution of carnitine and acetylcarnitine in the epididymis and epididymal spermatozoa of the boar.

In the epididymal fluid of boars, the concentration of carnitine (nmol/mg protein) began to increase from 20 in the distal caput, then rose progressively to 700 in the distal cauda. By contrast, the carnitine content of spermatozoa only started to increase in the proximal cauda where the concentration of carnitine in the fluid was 200-300 nmol/mg protein then gradually increased in spermatozoa from more distal sites. The increase in the acetylcarnitine content of spermatozoa paralleled that of the carnitine amount and represented 50% of the total carnitine (carnitine + acetylcarnitine). We conclude that the acetylcarnitine content of epididymal spermatozoa may be used as a marker of maturation.

Sperm factors related to failure of human in-vitro fertilization.

Two groups of men were retrospectively selected according to their observed success in in-vitro fertilization. Seminal and post-migration sperm samples from a low fertilization rate group (less than or equal to 33% cleaved embryos) have been compared to results obtained from a high fertilization rate group (greater than or equal to 66%). It was found that a low mean value of the amplitude of lateral sperm head displacement and an increased percentage of abnormal acrosomes were related to in-vitro fertilization failure. None of the individual sperm factors studied was found to determine in-vitro fertilization success with certainty; only when they were considered in combination was it possible to predict the likelihood of successful in-vitro fertilization of human oocytes.

Semen abnormalities in patients with viral hepatitis B.

Semen parameters of 10 acute hepatitis and 5 chronic carriers were studies. The most common alterations were necrospermia (62% of the patients), teratospermia with irregular shape of the heads (46%), asthenospermia (36%), small volume (33%), and oligospermia (27%). The possible mechanisms of such anomalies are discussed.

Morphological factors influencing the penetration of human sperm into cervical mucus in vitro.

The efficiency of cervical mucus in filtering out single, multiple and associated abnormalities of human spermatozoa was determined. Twenty semen samples which gave a normal in vitro cervical mucus penetration test (CMPT) were analysed before and after migration using a detailed classification system (13 categories). The % of normal forms was significantly increased in cervical mucus (59.5 vs 33.2%), whereas the % of sperm with single, multiple or associated abnormalities of the midpiece or of the flagellum were found to decrease significantly in cervical mucus. Sperm with single or multiple abnormalities confined to the head migrated similarly to normal forms. The decrease in amorphous and elongated tapering sperm was explained by their more frequent association with other defects of the midpiece and/or of the flagellum.

The effects of centrifugation, various synthetic media and temperature on the motility and vitality of human spermatozoa.

The influence of various technical procedures and diluent media upon human spermatozoa has been tested in vitro. The percentage and velocity of motile spermatozoa were measured objectively using laser Doppler velocimetry. Vitality was determined by fluorescent staining. Centrifugation of semen (diluted 1 : 1 with Tyrode) at forces of 500, 800, 1 800 and 2 500 X g resulted in almost complete sedimentation of the spermatozoa. After resuspension in Tyrode's, spermatozoal motility was not changed significantly. The percentages of motile spermatozoa (measured at 37 degrees C) in Tyrode's and B2 medium remained the same as those in seminal plasma throughout an 8-hr period of incubation at 20 degrees C, while those in Locke's and phosphate-buffered saline (PBS) were significantly lower. The instantaneous modal velocities (Vc) of spermatozoa (also measured at 37 degrees C) in Locks's, Tyrode's and B2 were all significantly increased as compared to those observed in seminal plasma during incubation at 20 degrees C; spermatozoa in PBS showed a significantly reduced velocity compared to seminal plasma, but the difference was not significant. However, the rates of decline of the percentage of motile spermatozoa and of their velocities were indistinguishable in all four synthetic media and in the seminal plasma. No significant change in vitality was observed in any medium during the 8-hr incubation period at 20 degrees C. When spermatozoa were incubated at 37 degrees C, rapid declines in both percentage motile and in velocity were observed with incubation periods lasting more than 4-h. Subjecting the spermatozoa to a temperature of 4 degrees C for 1-h did not significantly change either of the motility parameters (measured at 37 degrees C), although prolonged exposure to 4 degrees C did greatly reduce their motility and velocity percentages. The decline in vitality was slow and uniform during incubation at either 4, 20 or 37 degrees C over periods of up to 10-hr, the best survival apparently being obtained at 20 degrees C and the worst at 4 degrees C, although there were no significant temperature differences at any time.