Pentoxifylline improves sperm capacitation and in vitro fertilization of oocytes in the golden hamster

Pentoxifylline (PF) is used to improve motility of spermatozoa from subfertile or nonfertile males to accomplish in vitro fertilization in humans. The possible adverse effect of PF on pre- and peri- implantation stage embryo development in a suitable rodent model, such as the golden hamster, is yet to be determined. In this study, hamster cauda epididymal spermatozoa were exposed to different concentrations (0.23 to 3.6 mM) of PF, and their quantitative [percentage of motility] and qualitative [Score 0 to 5] motility were assessed and values expressed as the sperm motility index. Upon addition of spermatozoa to dishes containing PF, an immediate increase in sperm motility and sperm motility index was evident, which increased up to 4 to 6 h and then declined. The sperm motility index increase by PF was dose-dependant, and greater than or equal to 1.8 mM PF was detrimental after 4 h. The optimum dose of PF was found to be 0.45 mM. To assess the fertilizing ability of PF-treated spermatozoa, in vitro fertilization was carried out. Fertilization rates for spermatozoa treated with 3.6 mM PF were lower (53.8 +/- 7.8) than for the controls (69.5 +/- 10.2), whereas, treatment with 0.45 mM PF increased the rates (91.6 +/- 4.3) compared with that of the controls (80.2 +/- 5.9). In conclusion, low concentrations (0.23 to 0.45 mM) of PF improve sperm capacitation and fertilization of oocytes in vitro in the golden hamster.

Susceptibility of sperm chromatin to acid denaturation in situ: A study in endogenous FSH-deprived adult male bonnet monkeys (Macaca radiata)

Acid denaturation of calf thymus DNA in vitro followed by acridine orange (AO) binding induced a 112% increase in the emission of red, a 58% decrease in green, and a consequential decrease in the ratio of green:red fluorescences from 1.7 to 0.9. This metachromatic property of AO on binding to DNA following acid denaturation was utilized to study the susceptibility of normal and ovine follicle-stimulating hormone (oFSH) actively immunized bonnet monkey spermatozoa voided throughout the year. For analyses, the scattergram generated by the emission of red and green fluorescences by 10,000 AO-bound sperm from each semen sample was divided into 4 quadrant zones representing percentage cells fluorescing high green-low red (Q1), high green-high red (Q2), low green-low red (Q3) and low green-high red. (Q4). Normal monkey sperm obtained during the months of July-December exhibited 76, 13, and 11% cells in Q2, Q3, and Q4 quadrants, respectively. However, during January-June, when the females of the species are markedly subfertile, noncycling, and amenorrhoeic, the spermatozoa ejaculated by the male monkeys exhibited 38, 39, and 23% sperm in Q2, Q3, and Q4, respectively, the differences being highly significant (p < .01-.001). FSH deprivation induced significant shifts in fluorescence emissions, from respective controls, with 39, 33, and 28% cells in Q2, Q3, and Q4, respectively, during July-December, and 15, 48, and 37% sperm in Q2, Q3, and Q4 quadrants, respectively, during January-June. It is postulated that the altered kinetics of germ cell transformations and the deficient spermiogenesis observed earlier following FSH deprivation in these monkeys may have induced the enhanced susceptibility to acid denaturation in sperm.

Report on Marine Mammal Unusual Mortality Event UMESE0501Sp: Multispecies Mass Stranding of Pilot Whales (Globicephala macrorhynchus), Minke Whale (Balaenoptera acutorostrata), and Dwarf Sperm Whales (Kogia sima) in North Carolina on 15-16 January 2005

On 15-16 January 2005, three offshore species of cetaceans (33 short-finned pilot whales, Globicephala macrorhynchus, one minke whale, Balaenoptera acutorostrata, and two dwarf sperm whales, Kogia sima) stranded alive on the beaches of North Carolina. The pilot whales stranded near Oregon Inlet, the minke whale in northern North Carolina, and the dwarf sperm whales near Cape Hatteras. Live strandings of three species in one weekend was unique in North Carolina and qualified as an Unusual Mortality Event. Gross necropsies were conducted on 16-17 January 2005 on 27 pilot whales, two dwarf sperm whales, and the minke whale. Samples were collected for clinical pathology, parasitology, gross pathology, histopathology, microbiology and serology. There was variation in the number of animals sampled for each collection type, however, due to carcasses washing off the beach or degradation in carcass condition during the course of the response. Comprehensive histologic examination was conducted on 16 pilot whales, both dwarf sperm whales, and the minke whale. Limited organ or only head tissue suites were obtained from nine pilot whales. Histologic examination of tissues began in February 2005 and concluded in December 2005 when final sampling was concluded. Neither the pilot whales nor dwarf sperm whales were emaciated although none had recently ingested prey in their stomachs. The minke whale was emaciated; it was likely a dependent calf that became separated from the female. Most serum biochemistry abnormalities appear to have resulted from the stranding and indicated deteriorating condition from being on land for an extended period. Three pilot whales had clinical evidence of pre-existing systemic inflammation, which was supported by histopathologic findings. Although gross and histologic lesions involving all organ systems were noted, consistent lesions were not observed across species. Verminous pterygoid sinusitis and healed fishery interactions were seen in pilot whales but neither of these changes were causes of debilitation or death. In three pilot whales and one dwarf sperm whale there was evidence of clinically significant disease in postcranial tissues which led to chronic debilitation. Cardiovascular disease was present in one pilot whale and one dwarf sperm whale; musculoskeletal disease and intra-abdominal granulomas were present in two pilot whales. These lesions were possible, but not definitive, causal factors in the stranding. Remaining lesions were incidental or post-stranding. The minke whale and three of five tested pilot whales had positive morbillivirus titers (≥1:8 with one at >1:256), but there was no histologic evidence of active viral infection. Parasites (nematodes, cestodes, and trematodes) were collected from 26 pilot whales and two dwarf sperm whales. Sites of collection included stomach, nasal/pterygoid, peribullar sinuses, blubber, and abdominal cavity. Parasite species, locations and loads were within normal limits for free-ranging cetaceans and were not considered causative for the stranding event. Gas emboli lesions which were considered consistent with or diagnostic of sonarassociated strandings of beaked whales or small cetaceans were not found in the whales stranded as part of UMESE0501Sp. Twenty-five heads were examined with nine specific anatomic locations of interest: extramandibular fat, intramandibular fat, auditory meatus, peribullar acoustic fat, peribullar soft tissue, peribullar sinus, pterygoid sinus, melon, and brain. The common finding in all examined heads was verminous pterygoid sinusitis. Intramandibular adipose tissue reddening, typically adjacent to the vascular plexus, was observed in some individuals and could represent localized hemorrhage resulting from vascular rete rupture, hypostatic congestion, or erythrocyte rupture during the freeze/thaw cycle. One cetacean had peracute to acute subdural hemorrhage that likely occurred from thrashing on the beach post-stranding, although its occurrence prior to stranding cannot be excluded. Information provided to NMFS by the U.S. Navy indicated routine tactical mid-frequency sonar operations from individual surface vessels over relatively short durations and small spatial scales within the area and time period investigated. No marine mammals were detected by marine mammal observers on operational vessels; standard operating procedure for surface naval vessels operating mid-frequency sonar is the use of trained visual lookouts using high-powered binoculars. Sound propagation modeling using information provided to NMFS indicated that acoustic conditions in the vicinity likely depended heavily on position of the receivers (e.g., range, bearing, depth) relative to that of the sources. Absent explicit information on the location of animals meant that it was not possible to estimate received acoustic exposures from active sonar transmissions. Nonetheless, the event was associated in time and space with naval activity using mid-frequency active sonar. It also had a number of features in common (e.g., the “atypical” distribution of strandings involving multiple offshore species, all stranding alive, and without evidence of common infectious or other disease process) with other sonar-related cetacean mass stranding events. Given that this event was the only stranding of offshore species to occur within a 2-3 day period in the region on record (i.e., a very rare event), and given the occurrence of the event simultaneously in time and space with a naval exercise using active sonar, the association between the naval sonar activity and the location and timing of the event could be a causal rather than a coincidental relationship. However, evidence supporting a definitive association is lacking, and, in particular, there are differences in operational/environmental characteristics between this event and previous events where sonar has apparently played a role in marine mammal strandings. This does not preclude behavorial avoidance of noise exposure. No harmful algal blooms were present along the Atlantic coast south of the Chesapeake Bay during the months prior to the event. Environmental conditions, including strong winds, changes in upwelling- to downwelling-favorable conditions, and gently sloping bathymetry, were consistent with conditions which have been correlated with other mass strandings. In summary, we did not find commonality in gross and histologic lesions that would indicate a single cause for this stranding event. Three pilot whales and one dwarf sperm whale had debilitating conditions identified that could have contributed to stranding, one pilot whale had a debilitating condition (subdural hemorrhage) that could have been present prior to or resulting from stranding. While the pilot and dwarf sperm whale strandings may have had a common cause, the minke whale stranding was probably just coincidental. On the basis of examination of physical evidence in the affected whales, however, we cannot definitively conclude that there was or was not a causal link between anthropogenic sonar activity or environmental conditions (or a combination of these factors) and the strandings. Overall, the cause of UMESE0501Sp in North Carolina is not and likely will not be definitively known. (PDF contains 240 pages)

Cephalopods from the stomachs of sperm whales taken off California

Cephalopod remains (beaks, bodies, and parts of bodies) were collected from the stomachs of 157 sperm whales (Physeter macrocephalus) taken off central California (lat. 37°-39°N). At least 24 species representing 14 families were identified. Frequencies of occurrence of the six most numerous taxa were Moroteuthis robusta 72.0%, Gonatopsis borealis 66.2%, Histioteuthis dofleini 36.9%, Galiteuthis spp. (including G. phyllura and G. pacifica) 36.3%, Octopoteuthis deletron 35.0%, and Vampyroteuthis infernalis 27.4%. One find of two Mesonychoteuthis hamiltoni beaks strongly suggests transequatorial migration by one large male sperm whale. (PDF file contains 18 pages.)

The Voltage-Gated Sodium Channel Na(v)1.8 Is Expressed in Human Sperm

The role of Na+ fluxes through voltage-gated sodium channels in the regulation of sperm cell function remains poorly understood. Previously, we reported that several genes encoding voltage-gated Na+ channels were expressed in human testis and mature spermatozoa. In this study, we analyzed the presence and function of the TTX-resistant VGSC a subunit Na(v)1.8 in human capacitated sperm cells. Using an RT-PCR assay, we found that the mRNA of the gene SCN10A, that encode Na-v1.8, was abundantly and specifically expressed in human testis and ejaculated spermatozoa. The Na-v1.8 protein was detected in capacitated sperm cells using three different specific antibodies against this channel. Positive immunoreactivity was mainly located in the neck and the principal piece of the flagellum. The presence of Na-v1.8 in sperm cells was confirmed by Western blot. Functional studies demonstrated that the increases in progressive motility produced by veratridine, a voltage-gated sodium channel activator, were reduced in sperm cells preincubated with TTX (10 mu M), the Na-v1.8 antagonist A-803467, or a specific Na-v1.8 antibody. Veratridine elicited similar percentage increases in progressive motility in sperm cells maintained in Ca2+-containing or Ca2+-free solution and did not induce hyperactivation or the acrosome reaction. Veratridine caused a rise in sperm intracellular Na+, [Na+](i), and the sustained phase of the response was inhibited in the presence of A-803467. These results verify that the Na+ channel Na-v1.8 is present in human sperm cells and demonstrate that this channel participates in the regulation of sperm function.