Lactate Dehydrogenase Isozymes: Dissociation and Recombination of Subunits

Lactate dehydrogenase from beef tissues may be resolved electrophoretically into five isozymes each of which is a tetramer. These tetramers can be dissociated into monomers by freezing in 1M sodium chloride. On thawing, reassociation into functional tetramers occurs. On the basis of charge and amino acid composition there are two kinds of monomers. Lactate dehydrogenase-1 contains one kind of monomer and lactate dehydrogenase-5 the other kind. A mixture of equal quantities of these two isozymes, after dissociation and reassociation, leads to the production of all five isozymes in the expected proportions of 1:4:6:4:1.     

Sperm Morphology in the Domestic Cat, and its Relation with Fertility: A Retrospective Study

Knowledge about normal ranges in semen quality and the association between sperm morphology and fertility in felids is limited. The aims of this retrospective study were to (1) define a normal spermiogram in cats; (2) evaluate possible effects of season, age and breed on sperm morphology; and (3) evaluate the relationship between sperm morphology and fertility. Semen samples collected by electroejaculation from 52 cats were evaluated for sperm morphology. The cats constituted two groups: a general population of cats (n = 48) and cats examined because of poor breeding records (n = 4). The general population was divided into household (n = 20), pedigree (n = 19) and colony cats (n = 9) and into three age classes, <12 months, 12-59 months and >or=60 months. The median percentage of normal spermatozoa in the general population was 44.0% (range 1.0-91.0%). Criteria were tentatively set for what was considered a normal spermiogram. The mean percentage of normal spermatozoa was higher during February to July than during August to January (p < 0.05). Pedigree cats had a lower mean percentage of normal spermatozoa than did household cats (p < 0.05). Age had no effect on the percentage of normal spermatozoa but was positively correlated with the percentage of proximal droplets. Of the cats with <40% normal spermatozoa (n = 19), all those with known breeding records (n = 11) had produced litters. The four cats examined because of poor breeding results had higher percentages of different sperm abnormalities than tentatively stipulated for the normal spermiogram. In two of these cats both sperm morphology and fertility changed over time.     

Observations on the cooling and cryopreservation of pig oocytes at the germinal vesicle stage

This study examined the viability of pig oocytes at the germinal vesicle stage following cooling or cryopreservation. Cumulus-intact oocytes (n = 641) were collected from slaughterhouse pig ovaries and used in two experiments. In Exp. I the viability of 1) control, 2) cryoprotectant control (CC, 1.5 M glycerol/.5 M sucrose), 3) cooled (0 degrees C) and 4) cryopreserved (-196 degrees C) oocytes was assessed after no incubation or a 24-h incubation. Survivability was judged by morphological appearance, trypan blue exclusion and fluorescein diacetate staining. Survival rate of control oocytes (90%; based primarily on morphological appearance of the cumulus) incubated 0 h was greater (P less than .05) than that of all other groups, whereas survival rate of -196 degrees C oocytes (57%) was less (P less than .05) than that of all other groups. However, vital staining of 0 degrees C and -196 degrees C oocytes showed 0% survival rate as evidenced by trypan blue uptake and lack of fluorescence. The cumulus cells surrounding oocytes that were stored at 0 degrees C or -196 degrees C survived freezing as evidenced by trypan blue exclusion and intense fluorescence. Similar differences among treatment groups were found for oocytes incubated 24 h. Exp. 2 examined the temperature at which oocytes became sensitive to cooling. Oocyte death occurred when oocytes were cooled to 15 degrees C or lower. These results demonstrate that pig oocytes at the germinal vesicle stage did not survive cooling to 15 degrees C or below. When assessing the viability of cryopreserved cumulus enclosed oocytes it is important to use vital stains in conjunction with morphological appearance.     

Management of inappetant sheep during export by sea

In the first of 2 experiments, a simulated voyage was conducted to examine the effects of various treatments on bodyweight change and feeding frequency of inappetant sheep at the end of lot-feeding (non-feeders). The treatments, applied during simulated shipping, were: normal quantities of feed and length of troughs; extra trough length; extra feed. Adult Merino wethers (n = 108) were used in each treatment. A voyage to the Middle East was then conducted to establish whether shipboard mortality could be reduced by separating non-feeders (n = 305) from feeders (n = 5,620) late in the feedlot hase and housing the groups separately aboard ship. A control group of non-feeders (n = 215) mixed with feeders (n = 5,732) was used for comparison. Bars (marker bars), containing a coloured dye, were attached to feed troughs to mark sheep that fed. Most non-feeders (82%) began eating when placed in shipping pens in both studies. However, there was no significant difference in percentage of sheep that fed between non-feeders given extra trough length or extra feed compared with non-feeders given standard management at any stage of simulated shipping. There was no significant difference in mean bodyweights between treatment groups on days 1, 8 and 15 of simulated shipping. Differences in bodyweight on d 22 were probably associated with different levels of gut fill. Death rates were not significantly different in separated and control groups (1.1%, 0.9%, P = 0.6) in the voyage of 14 d to the Middle East.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of Oil Overlay on Inhibition Potential of Roscovitine in Sheep Cumulus‐Oocyte Complexes

Inhibitors of cyclin‐dependent kinases, as roscovitine, have been used to prevent the spontaneous resumption of meiosis in vitro and to improve the oocyte developmental competence. In this study, the interference of oil overlay on the reversible arrest capacity of roscovitine in sheep oocytes as well as its effects on cumulus expansion was evaluated. For this, cumulus‐oocyte complexes (COCs) were cultured for 20 h in TCM 199 with 10% foetal bovine serum (Control) containing 75 μm roscovitine (Rosco). Subsequently, they were in vitro matured (IVM) for further 18 h in inhibitor‐free medium with LH and FSH. The culture was performed in Petri dishes under mineral oil (+) or in 96 well plates without oil overlay (?) at 38.5°C and 5% CO₂. At 20 and 38 h, the cumulus expansion and nuclear maturation were evaluated under stereomicroscope and by Hoechst 33342 staining, respectively. No group presented cumulus expansion at 20 h. After additional culture with gonadotrophins, a significant rate of COCs from both Control groups (+/?) exhibited total expansion while in both Rosco groups (+/?) the partial expansion prevailed. Among the oocytes treated with roscovitine, 65.2% were kept at GV in the absence of oil overlay while 40.6% of them reached MII under oil cover (p < 0.05). This meiotic arrest was reversible, and proper meiosis progression also occurred in the Control groups (+/?). So, the culture system without oil overlay improved the meiotic inhibition promoted by roscovitine without affecting the cumulus expansion rate or the subsequent meiosis progression.     

Photoperiod induced off-season spawning of pink salmon (Oncorhynchus gorbuscha)

Four groups of pink salmon, which had been reared under artificial light, became sexually mature and produced viable gametes: 59 days prior to, and 19 – 32 days, 115 days and 220 days after their expected date of reaching sexual maturity. Altered times of sexual maturation were obtained by accelerating, leaving unchanged, or decelerating the rate of change of photoperiod which each group of fish would normally receive during its first year of life. All groups of fish were exposed to a normal rate of change in photoperiod during their final year of life. Mean fecundity was reduced from the 800–2000 ova observed in wild stocks, and ranged from 629 for the 59 day advanced fish, to 862 for the 115 day delayed fish. Egg mortality during the period from fertilization to eyeing was much greater in the three groups of fish subjected to accelerated or decelerated rates of change in photoperiod than in the fish subjected to the normal rate of change in photoperiod. Some of the progeny of the 220 day delayed fish, which were reared under artificial light with the normal rate of change in photoperiod set 220 days out-of-phase, became sexually mature 2 years after they had begun life as fertilized eggs.