Sperm motility in three coral reef fish species

A method of computer assisted sperm analysis (CASA) is used to determine the parameters of sperm motility in three fish species from the coral reefs of the Nha Trang Bay, South China Sea, Vietnam: Zebrasoma scopas (Acanthuridae), Abudefduf sexfasciatus, and Dascyllus trimaculatus (Pomacentridae). The representatives of the families are characterized by different reproductive tactics and possess pelagic and demersal eggs, respectively. The main morphological parameters of spermatozoa have been measured. The average curvilinear velocity of spermatozoa movement along the real trajectory (VCL) after 1 min of sperm activation ranges from 15.3 to 74.5 μm/s in Z. scopas, it is comparatively low (12.7–21.6 μm/s) in A. sexfasciatus, and high (58.4–92.2 μm/s) in D. trimaculatus. The duration of progressive movements in more than 50% of spermatozoa at 25°C is 3–20, 5–11, and 6–9 min after sperm activation, respectively. Following storage of the sperm of three species at 4.5°C for 7, 20, and 28 h, respectively, spermatozoa retain the ability of progressive movements. The results are discussed based on the available information on the activity of sperm in fish.     

Sperm motility and spermathecal filling in lower diptera

A recently completed study of sperm transfer and spermathecal filling in Culicoides melleus (Ceratopogonidae) provided evidence that the spermathecae create an incoming current by fluid absorption. This current, rather than sperm motility, was thought to accomplish transfer of spermatozoa from spermatophore to spermathecae. This review discusses the approach used to assess the role of sperm motility, as applied critically to available observations in other lower Diptera. Aedes aegypti (Culicidae), Simulium decorum (Simuliidae) and Plecia nearctica (Bibionidae) are considered. The results suggest that sperm motility very probably does not contribute to spermathecal filling in these species, and that fluid absorbtion is the more likely mechanism. Certain physical properties of the female reproductive systems are discussed in the light of this conclusion.     

Sperm motility and cryopreservation of spermatozoa in freshwater gobies

Testicular sperm motility and methods for the cryopreservation of spermatozoa in the freshwater goby Rhinogobius sp. CB (Cross Band type) were examined. Spermatozoa were almost immotile upon dilution with 300 mOsm kg⁻¹ of NaCl, KCl and mannitol solutions but began to swim in solutions with concentrations <200 mOsm kg⁻¹. The highest percentage and longest duration of motility was obtained in the 0 and 100–200 mOsm kg⁻¹ solutions, respectively. The highest post-thaw motility, c. 50% of motility before cryopreservation, was obtained when spermatozoa were diluted with an extender of 10% methanol and 90% artificial seminal plasma, cooled at −10·0 ± 1·1° C min⁻¹ (mean ± s . e .) to −50° C and plunged into liquid nitrogen. Spermatozoa were cryopreserved in a 50 μl acrylic haematocrit tube to store the small amount of milt. As the cryopreservation method described above was applicable to the endangered Rhinogobius sp. BI (Bonin Island type), it is probable that this method can be used for other species of freshwater gobies.     

Sperm motility in the horseshoe crab,Limulus polyphemus L: I. Sperm behavior near eggs and motility initiation by egg extracts

Limulus spermatozoa are nonmotile when spawned and become motile only after encountering a sperm motility initiating factor (SMI) exuded by the egg. SMI extracts (produced by washing intact eggs with distilled water, lyophilizing the supernatant to dryness, and redissolving the dried extract in artificial seawater, ASW) initiate sperm motility in the absence of eggs. Utilizing such SMI extracts, sperm motility initiation was found to be unaffected by changes in temperature from 16 to 30°C, pH from 6.3 to 8.6, and salinity from 85 to 125% ASW. Within these ranges, sperm motility initiation was an “all-or-nothing” response, with greater than 99% of the spermatozoa becoming motile. Also, each sperm swam with apparently the same speed (at a given temperature) until spontaneously stopping within 10 min after the addition of SMI extracts. Evidence was found that SMI may bind irreversibly to a receptor, which is inactivated within a few seconds or minutes, leading to the observed cessation of motility. Observations of sperm behavior near intact eggs showed no evidence of chemotaxis. Spermatozoa observed to swim toward intact eggs progressed with a uniform speed and were motile less than 5 sec from initiation of motility until attaching to the egg. The presence of an all-or-nothing response to SMI, the independence of sperm motility to experimental parameters, and several other characteristics of the animal and its spermatozoa make Limulus a potentially excellent model animal for examination of sperm motility control mechanisms.     

Sperm structure and motility of the freshwater teleost Cottus gobio

When motility of spermatozoa of Cottos gobio was initiated with distilled water, the motility rate decreased to 0% within 1 min, and significant signs of osmotic alterations were observed at the end of the motility period. By contrast, in 50 mmol 1⁻¹ NaCl solution, the motility rate persisted for 120–140 min. In both distilled water and in 50 mmol 1⁻¹ NaCl solution, the main swimming type of spermatozoa was linear motion during the whole motility period. The initial swimming velocity (50.0 ± 2.1 μm s⁻¹) measured 10 s after motility initiation was similar in both distilled water and in 50 mmol 1⁻¹ NaCl solution. In distilled water, the velocity decreased to <20 μm s⁻¹ (locally motile) during the first minute of the motility phase. In 50 mmol 1⁻¹ NaCl solutions, it remained at a constant level during the first 60 min of the motility period, but then started to decrease to <20 μm s⁻¹ after 120 min. When 5 mmol 1⁻¹ potassium cyanide, antimycin or atractyloside was added to the 50 mmol 1⁻¹ NaCl solution, the motility period was reduced to ≤2min. Ten millimoles per litre 2-deoxy-D-glucose, malonate or a mixture of 5 mmol 1⁻¹ atractyloside and 5 mmol 1⁻¹ carnithine did not effect the duration of the motility period. This indicates that sperm energy metabolism depends mainly on respiration rate and fatty acid metabolism.     

Sperm motility patterns and metabolism in Catalonian donkey semen

The Sperm-Class Analyzer detected four subpopulations of spermatozoa with different motility characteristics in the ejaculate of the Catalonian donkey. Significant differences (P < 0.001) in the distribution of these subpopulations, as well as in total sperm number and percentage total motility, were seen in the diluted semen of four sampled donkeys. All the ejaculates evaluated showed excellent semen quality characteristics; the sperm they contained was more rapid than horse sperm. Principal components analysis showed sperm l-lactate production to be a good predictor of semen condition. This, plus the characteristics of the motility patterns of the different sperm subpopulations, provides an excellent overall indicator of semen quality.     

Sperm transport and motility in the mouse oviduct: observations in situ

Sperm transport and motility were studied through the transparent walls of the mouse oviduct by direct microscopic observation and videomicrography. Observations were made on excised female tracts 1-2 h post-coitus (pc) and 1-2 h before and after the approximate time of ovulation. Motile sperm were seen at the uterine entrance to the uterotubal junction (UTJ) in all females at 1-2 h pc, but in fewer females at later times. The intramural UTJ was usually constricted and held few sperm. The extramural UTJ and adjacent lower isthmus contained many motile sperm at 1-2 h pc. Apparently, the column of sperm moved upwards because in some females, sperm were found in the upper isthmus and not in the UTJ at the later time points. Few sperm were seen in the ampulla in the periovulatory period, and none at 1-2 h pc. There appeared to be two mechanisms retaining sperm in the lower oviduct: immobilization and adherence to the epithelium. Columns of immotile sperm were seen in the lower isthmus of some females. Motile sperm usually appeared to adhere by their heads to the oviductal epithelium, only occasionally breaking free to move vigorously about the lumen.     

Sperm motility in the horseshoe crab. III. Isolation and characterization of a sperm motility initiating peptide

Sperm motility in Limulus is initiated by a sperm motility initiating factor (SMI) that emanates from Limulus eggs. This report describes the partial purification of SMI (greater than 230-fold purification with respect to protein content) with 40% recovery. SMI appears to be a hydrophobic peptide of 500–2,000 MW. Although probably not purified to homogeneity, SMI is estimated to be active at a concentration of less than 0.2 μM.     

Sperm motility and seminal fluid composition in the burbot, Lota lota

Sperm motility and composition of the seminal fluid in Lota lota were investigated. Fives after motility initiation, 88.2 ± 12.4% of the spermatozoa were motile, their mean average path swimming velocity was 61.6 ± 16.3 μm s^?1 and their principal swimming type the linear motion (77.4 ± 20.9%). In distilled water the rate of motile spermatozoa decreased to 0% in 40s. In 25–50 mosmol kg^?1 electrolyte (NaCl) or non-electrolyte (glucose, sucrose) solutions, motility was prolonged for 10s and these solutions can therefore increase the efficiency of artificial fertilization when used for sperm motility activation. When semen was diluted in electrolyte or non-electrolyte solutions with osmolalities higher than 50 mosmol kg^?1, sperm motility rates and swimming velocities decreased, and at osmolalities of 400 mosmol kg^?1 motility was completely suppressed. In the seminal fluid with an osmolality of 290.08 ± 45.22 mosmol kg^?1, sodium levels of 139.86 ± 23.79 mmol × 1^?1, potassium levels of 11.59 ± 2.45 mmol × 1^?1 and calcium levels of 0.20 ± 0.08 mmol × 1^?1, sperm motility was inhibited. Under in vitro conditions, artificial saline solutions resembling the seminal plasma composition and 400 mosmol kg^?1 NaCl or glucose solutions were useful as motility inhibiting solutions for predilution of semen. Sperm motility was not affected by pH 7.5–9.0, but at pH 6 the motility rate and the swimming velocity were reduced; seminal fluid pH was 8.47 ± 0.02. Therefore buffering of the artificial saline solutions can provide more stabile conditions for semen during storage and activation. Temperature optimum of semen was between 2 and 5°C. At higher temperatures semen became spontaneously motile. Therefore, controlled temperature conditions are an important factor for handling of semen. The qualitative, organical composition of seminal fluid was similar as in other fresh water teleosts.     

Bacterial motility confers fitness advantage in the presence of phages

Dispersal provides the opportunity to escape harm and colonize new patches, enabling populations to expand and persist. However, the benefits of dispersal associated with escaping harm will be dependent on the structure of the environment and the likelihood of escape. Here, we empirically investigate how the spatial distribution of a parasite influences the evolution of host dispersal. Bacteriophages are a strong and common threat for bacteria in natural environments and offer a good system with which to explore parasite‐mediated selection on host dispersal. We used two transposon mutants of the opportunistic bacteria, Pseudomonas aeruginosa, which varied in their motility (a disperser and a nondisperser), and the lytic bacteriophage ФKZ. The phage was distributed either in the central point of colony inoculation only, thus offering an escape route for the dispersing bacteria; or, present throughout the agar, where benefits of dispersal might be lost. Surprisingly, we found dispersal to be equally advantageous under both phage conditions relative to when phages were absent. A general explanation is that dispersal decreased the spatial structuring of host population, reducing opportunities for parasite transmission, but other more idiosyncratic mechanisms may also have contributed. This study highlights the crucial role the parasites can play on the evolution of dispersal and, more specifically, that bacteriophages, which are ubiquitous, are likely to select for bacterial motility.     

Sperm motility in the horseshoe crab. IV. Extracellular ions and intracellular pH are not mediators of motility initiation

Upon dilution into sea water, Limulus spermatozoa undergo a brief flurry of motility (duration < 60 sec), after which they are nonmotile until encountering a sperm motility initiating peptide (SMI) that emanates from eggs. Utilizing highly purified SMI extracts and simplified seawater formulations (from which individual ions have been deleted), we found that no specific extracellular ion is required for either dilution-initiated or SMI-initiated motility. Indeed, deletion of one ion (Na⁺) produced dilution-initiated motility of very long duration (several hours). When motility is initiated by SMI (in normal seawater) there is an increase in intracellular pH (pH_i), as indicated by the fluorescent probe, 9-amino acridine; however, this pH, change is not a trigger for motility. As a more general method examining ion movements, the fluorescent probe diS-C₃-(5) was used to qualitatively measure changes in the membrane potential of spermatozoa. Although crude SMI extracts caused membrane depolarization, further purification resulted in an almost complete separation of this activity from SMI, thus showing that SMI activation is apparently an electroneutral event. (The membrane-depolarizing factor has a molecular weight > 30,000 and does not initiate acrosome reactions.) Experiments utilizing the ionophore A23187 and Ca⁺²-blocking agents (verapamil and TMB-8) provided tentative evidence that mobilization of intracellular Ca⁺² may be required for motility initiation. These results show that neither changes in pH_i nor the influx of specific extracellular ions are direct mediators of SMI-initiated motility; however, experiments with pharmacologic agents indicate a possible role for intracellular Ca⁺².     

Sperm motility and calcium transport: a neurochemically controlled process

Elucidation of the functional significance of the presence of acetylcholinesterase in spermatozoa was deferred until considerably after the initial observations that eserine increased flagellar beat frequency in $\text{Mytilus}$ sperm (54). Discovered about forty years ago during surveys of seminal plasma constituents (2), the enzyme was later found to be associated with the particulate fraction (4). Even though characterized as a “true” acetylcholinesterase in the sperm flagellum (3), interpretations of the role of acetylcholine as a factor in motile processes remained speculative until a wide variety of cholinergic agents was found to affect sperm cell performance (32). Those agents which inhibit acetylcholine synthesis or block cholinergic receptors generally elicit a monophasic, inhibitory, dose-dependent response on sea urchin sperm swimming speed. These include hemicholinium, curare, α-bungarotoxin and decamethonium. Cholinomimetic substances and anticholinesterases (acetylcholine and nicotine; eserine, diisopropylfluorophosphate and neostigmine) act biphasically to increase the sperm swim rate at concentrations of 1 micromolar or less while higher concentrations slow or stop the forward motion of the sperm cells. Procaine (7), which competes with Ca²⁺ for cell surface binding sites, initially stimulates then completely inhinits sperm cell progression. Other substances which interfere with the cells′ access to Ca²⁺ or with the transport of Ca²⁺ into and within the cell adversely affect spermatozoan function. A conceptual model has been proposed, a) to relate the uptake of Ca²⁺ to cholinergic regulation of ionophoric channels through the sperm cell plasma membrane and b) for this Ca²⁺ to trigger release of sequestered calcium for coupling of excitation to flagellar wave propagation.     

Sperm motility is dependent on a unique isoform of the Na,K-ATPase

The Na,K-ATPase, a member of the P-type ATPases, is composed of two subunits, alpha and beta, and is responsible for translocating Na(+) out of the cell and K(+) into the cell using the energy of hydrolysis of one molecule of ATP. The electrochemical gradient it generates is necessary for many cellular functions, including establishment of the plasma membrane potential and transport of sugars and ions in and out of the cell. Families of isoforms for both the alpha and beta subunits have been identified, and specific functional roles for individual isoforms are just beginning to emerge. The alpha4 isoform is the most recently identified Na, K-ATPase alpha isoform, and its expression has been found only in testis. Here we show that expression of the alpha4 isoform in testis is localized to spermatozoa and that inhibition of this isoform alone eliminates sperm motility. These data describe for the first time a biological function for the alpha4 isoform of the Na,K-ATPase, revealing a critical role for this isoform in sperm motility.     

Sperm motility initiation factor is a minor component of the Pacific herring egg chorion

Polyclonal antibodies were generated to the 105 kDa herring sperm motility initiation factor (SMIF) and used to explore the role of SMIF in sperm-egg interaction. Using sodium dodecyl sulfate polyacrylamide gel electrophoresis and immunoblotting with SMIF antibodies, it was demonstrated that SMIF is present as a minor (4–7% of total chorion protein) component of the chorion. The major polypeptides in the chorion migrated at 117 kDa and in a grouping between 48–54 kDa, with other minor bands above and below. The only detectable glycosylated component was the 105 kDa band, which was resolved at two isoelectric points (8.22 and 8.31) after isoelectric focusing gel electrophoresis. Using antibodies to SMIF, fertilization was blocked, sperm motility was inhibited in vitro in the presence of solubilized SMIF and SMIF binding sites on sperm were localized. Lastly, SMIF was localized to the region of the herring egg that encircles the micropyle.     

Changes in motility of the rhizobacterium Azospirillum brasilense in the presence of plant lectins

The plant-beneficial bacterium Azospirillum brasilense can swim in liquids and swarm or migrate with the formation of microcolonies in soft media. To get closer to understanding the influence of natural environments on A. brasilense motility, we studied the individual and social movement of the bacterium in the presence of various plant lectins. The lectins with specificity for N-acetyl-beta-d-glucosamine oligomers (wheat germ, Solanum tuberosum and Ulex europeus agglutinins) decreased A. brasilense swimming speed and induced the formation of branched-granular colonies instead of the swarming rings. These effects seemed to be a consequence of specific interactions between the agglutinins and the lectin-binding polymers present in the A. brasilense cell envelope. Concanavalin A (with an affinity for terminal alpha-d-mannosyl and alpha-d-glucosyl residues) and Phaseolus vulgaris phytohemagglutinin P (with unknown specificity) almost did not affect the motility of A. brasilense.     

Sperm development and motility are regulated by PP1 phosphatases in Caenorhabditis elegans

Sperm from different species have evolved distinctive motility structures, including tubulin-based flagella in mammals and major sperm protein (MSP)-based pseudopods in nematodes. Despite such divergence, we show that sperm-specific PP1 phosphatases, which are required for male fertility in mouse, function in multiple processes in the development and motility of Caenorhabditis elegans amoeboid sperm. We used live-imaging analysis to show the PP1 phosphatases GSP-3 and GSP-4 (GSP-3/4) are required to partition chromosomes during sperm meiosis. Postmeiosis, tracking fluorescently labeled sperm revealed that both male and hermaphrodite sperm lacking GSP-3/4 are immotile. Genetic and in vitro activation assays show lack of GSP-3/4 causes defects in pseudopod development and the rate of pseudopodial treadmilling. Further, GSP-3/4 are required for the localization dynamics of MSP. GSP-3/4 shift localization in concert with MSP from fibrous bodies that sequester MSP at the base of the pseudopod, where directed MSP disassembly facilitates pseudopod contraction. Consistent with a role for GSP-3/4 as a spatial regulator of MSP disassembly, MSP is mislocalized in sperm lacking GSP-3/4. Although a requirement for PP1 phosphatases in nematode and mammalian sperm suggests evolutionary conservation, we show PP1s have independently evolved sperm-specific paralogs in separate lineages. Thus PP1 phosphatases are highly adaptable and employed across a broad range of sexually reproducing species to regulate male fertility.     

Sperm motility in the horseshoe crab,Limulus polyphemus L: II. Partial characterization of a motility initiating factor from eggs and the effects of inorganic cations on motility initiation

Egg extracts (obtained by washing intact Limulus eggs with either distilled water or artificial seawater, ASW) contain a sperm motility initiating factor (SMI). The SMI is heat stable (withstands boiling to dryness), passes through a dialysis membrane, and is retained by G-10 Sephadex (indicating a molecular weight of less than 700). Qualitative analysis (by X-ray fluorescence spectroscopy) and quantitative analysis (by atomic absorption spectroscopy) of SMI extracts revealed the presence of four divalent cations (Ca, Mg, Ni, and Cu) and one monovalent cation (K) that affect sperm motility. When assayed individually at high concentrations, all of the divalent cations initiate sperm motility and K⁺ inhibits motility initiation by the divalent cations. However, none of the divalent cations are present at concentrations high enough to produce the observed SMI activity, and since K⁺ is present when motility is initiated by SMI, K⁺ must not normally be an inhibitor. Therefore, if inorganic cations are involved in normal sperm motility initiation in Limulus, they are acting in conjunction with some other low molecular weight factor.