Calaxin is required for asymmetric bend initiation and propagation in sperm flagella.

Regulation of waveform asymmetry in flagella is critical for changes in direction when sperm are swimming, as seen during the chemotaxis of sperm towards eggs. Ca2+ is an important regulator of asymmetry in flagellar waveforms. A calcium sensor protein, calaxin, is associated with the outer arm dynein and plays a key role in the regulation of flagellar motility in a Ca2+-dependent manner. However, the underlying mechanism of regulating asymmetric waves by means of Ca2+ and calaxin remains unclear. To clarify the calaxin-dependent mechanism for generating Ca2+-dependent asymmetric flagellar waveforms, we analyzed the initial step of flagellar bend formation and propagation in the sperm of the ascidian Ciona intestinalis. Our experiment used demembranated sperm cells, which were then reactivated by UV flash photolysis of caged ATP under both high and low Ca2+ concentrations. Here, we show that initial bends in the flagella are formed at the base of the sperm and propagate towards the tip during waveform generation. However, the direction of the initial bend differed between asymmetric and symmetric waves. When a calaxin inhibitor (repaglinide) was applied, it resulted in the failure of asymmetric wave formation and propagation. This was because repaglinide had no effect on initial bend formation, but it significantly inhibited the generation of the subsequent bend in the reverse direction. Switching of dynein sliding activity by mechanical feedback is crucial for flagellar oscillation. Our results suggest that the Ca2+/calaxin mechanism plays an important role in the switching of dynein activity from microtubule sliding in the principal bend into the suppressed sliding in the reverse bend, thereby allowing the sperm to successfully change direction.

Frequency of Diplopia after Intraoperative Nerve Disturbance in Trochlear Nerve Schwannoma: A Case Report and Systematic Review.

Schwannomas of the trochlear nerve are relatively rare, and most patients present with preoperative diplopia because of trochlear nerve palsy. We describe the case of a 61-year-old male patient with a trochlear nerve schwannoma and no pre- and postoperative diplopia, despite his trochlear nerve being cut during the operation. We aimed to investigate the frequency of postoperative diplopia associated with intraoperative trochlear nerve disturbance by reviewing previous case reports, wherein postoperative diplopia did not occur after the trochlear nerve was cut intraoperatively. We recorded the frequency of diplopia because of intraoperative trochlear nerve disturbance, such as the trochlear nerve being cut, in cases without pre- and postoperative diplopia. We searched the PubMed, Medline, and Google Scholar databases for works published from 1976 to 2020 and followed the preferred reporting items for systematic reviews and meta-analyses guidelines. We reviewed 36 publications and found 92 cases of trochlear nerve schwannoma. Surgical resection was performed for 43 patients, of whom 40 were kept under observation and 9 were treated with radiation therapy. Of the 43 cases, 9 without preoperative diplopia underwent gross total resection. We analyzed ten cases (including ours) without preoperative diplopia to check for postoperative diplopia. In total, four cases, including ours, did not display postoperative diplopia despite the trochlear nerve being cut. This may be attributed to the preoperatively acquired motor and sensory fusion in the patient's vision because of tumor progression. Our findings may benefit neurosurgeons who treat patients with schwannomas and help them predict patients' outcomes.

How female squid inseminate their eggs with stored sperm.

How sperm reach ova after mating is one of the central questions in reproductive biology. Many species copulate and store sperm in female reproductive organs until spawning [1]. The way females use stored sperm is closely associated with sperm competition and cryptic female choice. However, it is difficult to observe the process of fertilization in natural spawning, as fertilization usually occurs in some 'hidden place' within the female's body. Here, we report the fertilization process of a squid using a glass plate as a spawning substratum, enabling observation within the female arm crown where the sperm storage organ is located and where fertilization may occur. Additionally, we detail the distribution of sperm around newly spawned eggs. Our observations reveal that: the female places her sperm-storage organ (seminal receptacle) over an egg held within her arm crown and inseminates the eggs one-by-one during attachment to the spawning substratum; sperm pass through a pathway within the jelly layers surrounding an egg; and such direct insemination behavior and the pathway through the egg jelly enables a female squid to externally fertilize her eggs using relatively few sperm. This study is the first to reveal the fertilization process using stored sperm, under female control.

Sperm from sneaker male squids exhibit chemotactic swarming to CO2.

Behavioral traits of sperm are adapted to the reproductive strategy that each species employs. In polyandrous species, spermatozoa often form motile clusters, which might be advantageous for competing with sperm from other males. Despite this presumed advantage for reproductive success, little is known about how sperm form such functional assemblies. Previously, we reported that males of the coastal squid Loligo bleekeri produce two morphologically different euspermatozoa that are linked to distinctly different mating behaviors. Consort and sneaker males use two distinct insemination sites, one inside and one outside the female's body, respectively. Here, we show that sperm release a self-attracting molecule that causes only sneaker sperm to swarm. We identified CO2 as the sperm chemoattractant and membrane-bound flagellar carbonic anhydrase as its sensor. Downstream signaling results from the generation of extracellular H(+), intracellular acidosis, and recovery from acidosis. These signaling events elicit Ca(2+)-dependent turning behavior, resulting in chemotactic swarming. These results illuminate the bifurcating evolution of sperm underlying the distinct fertilization strategies of this species.

Why small males have big sperm: dimorphic squid sperm linked to alternative mating behaviours.

BACKGROUND: Sperm cells are the target of strong sexual selection that may drive changes in sperm structure and function to maximize fertilisation success. Sperm evolution is regarded to be one of the major consequences of sperm competition in polyandrous species, however it can also be driven by adaptation to the environmental conditions at the site of fertilization. Strong stabilizing selection limits intra-specific variation, and therefore polymorphism, among fertile sperm (eusperm). Here we analyzed reproductive morphology differences among males employing characteristic alternative mating behaviours, and so potentially different conditions of sperm competition and fertilization environment, in the squid Loligo bleekeri. RESULTS: Large consort males transfer smaller (average total length = 73 µm) sperm to a female's internal sperm storage location, inside the oviduct; whereas small sneaker males transfer larger (99 µm) sperm to an external location around the seminal receptacle near the mouth. No significant difference in swimming speed was observed between consort and sneaker sperm. Furthermore, sperm precedence in the seminal receptacle was not biased toward longer sperm, suggesting no evidence for large sperm being favoured in competition for space in the sperm storage organ among sneaker males. CONCLUSIONS: Here we report the first case, in the squid Loligo bleekeri, where distinctly dimorphic eusperm are produced by different sized males that employ alternative mating behaviours. Our results found no evidence that the distinct sperm dimorphism was driven by between- and within-tactic sperm competition. We propose that presence of alternative fertilization environments with distinct characteristics (i.e. internal or external), whether or not in combination with the effects of sperm competition, can drive the disruptive evolution of sperm size.

Most fertilizing mouse spermatozoa begin their acrosome reaction before contact with the zona pellucida during in vitro fertilization.

To fuse with oocytes, spermatozoa of eutherian mammals must pass through extracellular coats, the cumulus cell layer, and the zona pellucida (ZP). It is generally believed that the acrosome reaction (AR) of spermatozoa, essential for zona penetration and fusion with oocytes, is triggered by sperm contact with the zona pellucida. Therefore, in most previous studies of sperm-oocyte interactions in the mouse, the cumulus has been removed before insemination to facilitate the examination of sperm-zona interactions. We used transgenic mouse spermatozoa, which enabled us to detect the onset of the acrosome reaction using fluorescence microscopy. We found that the spermatozoa that began the acrosome reaction before reaching the zona were able to penetrate the zona and fused with the oocyte's plasma membrane. In fact, most fertilizing spermatozoa underwent the acrosome reaction before reaching the zona pellucida of cumulus-enclosed oocytes, at least under the experimental conditions we used. The incidence of in vitro fertilization of cumulus-free oocytes was increased by coincubating oocytes with cumulus cells, suggesting an important role for cumulus cells and their matrix in natural fertilization.