Extruded gallbladder mucoceles have characteristic ultrasonographic features and extensive migratory capacity in dogs.

Limited information is available on the ultrasonographic characteristics of extruded gallbladder mucoceles. The objective of this retrospective case series study was to describe the ultrasonographic features of extruded gallbladder mucoceles in a group of dogs. Medical records of four veterinary centers were searched between June 2010 and January 2018 for all dogs with a suspected extruded gallbladder mucocele based on abdominal ultrasound. Seven client-owned dogs met the inclusion criteria. All seven dogs had a discrete, heteroechoic, multilayered, stellate, or striated mass within the peritoneal cavity. Blood flow was not present in these structures. Four of these structures were identified in the caudal abdomen. In five cases, the gallbladder lumen was filled with material similar in appearance to the free-floating peritoneal mass. Four cases were confirmed surgically and one case was confirmed at necropsy. Histopathology of the gallbladder wall was evaluated in all surgical cases and analysis of the free-floating material was performed in one of the four cases. Extruded gallbladder mucoceles have common imaging characteristics that may be helpful in establishing a presumptive diagnosis. Additionally, these structures may have extensive migratory capacity. The results of the study justify the recommendation for exploration of the entire abdomen during cholecystectomy for gallbladder mucocele removal.

Evaluation of standard magnetic resonance characteristics used to differentiate neoplastic, inflammatory, and vascular brain lesions in dogs.

Magnetic resonance (MR) imaging characteristics are commonly used to help predict intracranial disease categories in dogs, however, few large studies have objectively evaluated these characteristics. The purpose of this retrospective study was to evaluate MR characteristics that have been used to differentiate neoplastic, inflammatory, and vascular intracranial diseases in a large, multi-institutional population of dogs. Medical records from three veterinary teaching hospitals were searched over a 6-year period for dogs that had diagnostic quality brain MR scans and histologically confirmed intracranial disease. Three examiners who were unaware of histologic diagnosis independently evaluated 19 MR lesion characteristics totaling 57 possible responses. A total of 75 dogs with histologically confirmed intracranial disease were included in analyses: 51 with neoplasia, 18 with inflammatory disease, and six with cerebrovascular disease. Only strong contrast enhancement was more common in neoplasia than other disease categories. A multivariable statistical model suggested that extra-axial origin, T2-FLAIR mixed intensity, and defined lesion margins were also predictive of neoplasia. Meningeal enhancement, irregular lesion shape, and multifocal location distinguished inflammatory diseases from the other disease categories. No MR characteristics distinguished vascular lesions and these appeared most similar to neoplasia. These results differed from a previous report describing seven MR characteristics that were predictive of neoplasia in dogs and cats. Findings from the current study indicated that the high performance of MR for diagnosing canine intracranial diseases might be due to evaluator recognition of combinations of MR characteristics vs. relying on any one MR characteristic alone.

CatSper-null mutant spermatozoa are unable to ascend beyond the oviductal reservoir.

Sperm hyperactivation is characterised by high-amplitude, asymmetrical flagellar bending and is required to penetrate the oocyte zona pellucida. It was proposed that hyperactivation also enables spermatozoa to reach the oocyte by assisting escape from the oviductal sperm reservoir. To test this hypothesis, the behaviour of CatSper-null mouse spermatozoa in the oviduct was compared with that of spermatozoa from heterozygotes. CatSper(-/-) males are infertile because their spermatozoa fail to hyperactivate, whereas spermatozoa from CatSper(+/-) males have normal amounts of CatSper proteins and can hyperactivate. Males were mated with wild-type females on the morning of ovulation. Oviducts were obtained 1 or 4 h later, and behaviour of spermatozoa was examined using transillumination. At 1 h, null mutant spermatozoa remained attached by their heads to oviductal epithelium in the reservoir, whereas spermatozoa from heterozygotes detached from the oviductal epithelium after performing deep asymmetrical flagellar bends. At 4 h, 50 to 200 CatSper(+/-) spermatozoa were still seen in the oviducts; in contrast, only one CatSper(-/-) spermatozoon was found. CatSper(-/-) spermatozoa were lost from the oviducts after failing to detach from the epithelium in a timely manner, thus demonstrating that hyperactivation is required by spermatozoa to ascend beyond the oviductal reservoir.

Calmodulin and CaMKII in the sperm principal piece: evidence for a motility-related calcium/calmodulin pathway.

Both cyclic AMP (cAMP)/protein kinase A (PKA) and calcium (Ca(2+)) signaling pathways are known to be involved in the regulation of motility in mammalian sperm. Calmodulin (CaM) is a ubiquitous Ca(2+) sensor that has been implicated in the acrosome reaction. In this report, we identify an insoluble pool of CaM in sperm and show that the protein, in addition to its presence in the acrosome, is found in the principal piece of the flagellum. These findings are consistent with, though not proof of, the presence of a pool of CaM in the fibrous sheath. The Ca(2+)/CaM-dependent protein kinase IIbeta (CaMKIIbeta), which is a downstream target of Ca(2+)/CaM, similarly localizes to the principal piece. In addition, we confirm earlier reports that a CaM inhibitor decreases sperm motility. However, we find that this inhibition can be largely reversed by stimulation of PKA if substrates for oxidative respiration are present in the medium. Our results suggest that the Ca(2+)/CaM/CaMKII signaling pathway in the sperm principal piece is involved in regulating sperm motility, and that this pathway functions either in parallel with or upstream of the cAMP/PKA pathway.

GM1 dynamics as a marker for membrane changes associated with the process of capacitation in murine and bovine spermatozoa.

We previously showed that in live murine and bovine sperm heads, the ganglioside G(M1) localizes to the sterol-rich plasma membrane overlying the acrosome (APM). Labeling G(M1) using the pentameric cholera toxin subunit B (CTB) induced a dramatic redistribution of signal from the APM to the sterol-poor postacrosomal plasma membrane (PAPM) upon sperm death. We now show a similar phenomenon in the flagellum where CTB induces G(M1) redistribution to sterol-poor membrane subdomains of the annulus and flagellar zipper. Because sterol efflux from the plasma membrane is required for capacitation, we examined whether G(M1) localization might be useful to detect membrane changes associated with capacitation and/or acrosomal exocytosis. First, incubation of murine and bovine sperm with their respective stimuli for capacitation did not change G(M1) distribution in live cells. However, incubation of sperm of both species with specific stimuli for capacitation, followed by the use of specific fixation conditions, induced reproducible, stimulus-specific patterns of G(M1) distribution. By assessing changes in G(M1) distribution in response to progesterone-induced AE, we show that these patterns reflect the response of murine sperm populations to capacitating stimuli. These data suggest that G(M1) localization can be used as a diagnostic tool for evaluating sperm response to stimuli for capacitation and/or AE. Such information could be useful when deciding between technologies of assisted reproduction or when screening for male fertility. Furthermore, stimulus-specific changes in G(M1) distribution showed that sperm could respond to NaHCO(3) or mediators of sterol efflux independently, thereby refining existing models of capacitation.

Segregation of micron-scale membrane sub-domains in live murine sperm.

Lipid rafts, membrane sub-domains enriched in sterols and sphingolipids, are controversial because demonstrations of rafts have often utilized fixed cells. We showed in living sperm that the ganglioside G(M1) localized to a micron-scale membrane sub-domain in the plasma membrane overlying the acrosome. We investigated four models proposed for membrane sub-domain maintenance. G(M1) segregation was maintained in live sperm incubated under non-capacitating conditions, and after sterol efflux, a membrane alteration necessary for capacitation. The complete lack of G(M1) diffusion to the post-acrosomal plasma membrane (PAPM) in live cells argued against the transient confinement zone model. However, within seconds after cessation of sperm motility, G(M1) dramatically redistributed several microns from the acrosomal sub-domain to the post-acrosomal, non-raft sub-domain. This redistribution was not accompanied by movement of sterols, and was induced by the pentameric cholera toxin subunit B (CTB). These data argued against a lipid-lipid interaction model for sub-domain maintenance. Although impossible to rule out a lipid shell model definitively, mice lacking caveolin-1 maintained segregation of both sterols and G(M1), arguing against a role for lipid shells surrounding caveolin-1 in sub-domain maintenance. Scanning electron microscopy of sperm freeze-dried without fixation identified cytoskeletal structures at the sub-domain boundary. Although drugs used to disrupt actin and intermediate filaments had no effect on the segregation of G(M1), we found that disulfide-bonded proteins played a significant role in sub-domain segregation. Together, these data provide an example of membrane sub-domains extreme in terms of size and stability of lipid segregation, and implicate a protein-based membrane compartmentation mechanism.