The rabbit alpha-like globin gene cluster is polymorphic both in the sizes of BamHI fragments and in the numbers of duplicated sets of genes

The alpha-like globin gene cluster in rabbits contains embryonic zeta- globin genes, an adult alpha-globin gene, and theta-globin genes of undetermined function. The basic arrangement of genes, deduced from analysis of cloned DNA fragments, is 5'-zeta 0-zeta 1-alpha 1-theta 1- zeta 2-zeta 3-theta 2-3'. However, the pattern of restriction fragments containing zeta- and theta-globin genes varies among individual rabbits. Analysis of BamHI fragments of genomic DNA from 24 New Zealand white rabbits revealed eight different patterns of fragments containing zeta-globin genes. The large BamHI fragments containing genes zeta 0 and zeta 1 are polymorphic in length, whereas a 1.9-kb fragment containing the zeta 2 gene and the 3.5-kb fragment containing the zeta 3 gene do not vary in size. In contrast to this constancy in the size of the restriction fragments, the copy number of the zeta 2 and zeta 3 genes does vary among different rabbits. No length polymorphism was detected in the BamHI fragments containing the theta-globin genes, but again the copy number varies for restriction fragments containing the theta 2 gene. The alpha 1- and theta 1-globin genes are located in a nonpolymorphic 7.2-kb BamHI fragment. The combined data from hybridization with both zeta and theta probes shows that the BamHI cleavage pattern does not vary within the region 5'-alpha 1-theta 1- zeta 2-zeta 3-theta 2-3', but the pattern genomic blot-hybridization patterns for the progeny of parental rabbits with different zeta-globin gene patterns shows that the polymorphic patterns are inherited in a Mendelian fashion. Two different haplotypes have been mapped based on the genomic blot-hybridization data. The variation in the alpha-like globin gene cluster in the rabbit population results both from differences in the copy number of the duplication block containing the zeta-zeta-theta gene set and from the presence or absence of polymorphic BamHI sites.      

Trimethyloxonium modification of single batrachotoxin-activated sodium channels in planar bilayers. Changes in unit conductance and in block by saxitoxin and calcium

Single batrachotoxin-activated sodium channels from rat brain were modified by trimethyloxonium (TMO) after incorporation in planar lipid bilayers. TMO modification eliminated saxitoxin (STX) sensitivity, reduced the single channel conductance by 37%, and reduced calcium block of inward sodium currents. These effects always occurred concomitantly, in an all-or-none fashion. Calcium and STX protected sodium channels from TMO modification with potencies similar to their affinities for block. Calcium inhibited STX binding to rat brain membrane vesicles and relieved toxin block of channels in bilayers, apparently by competing with STX for the toxin binding site. These results suggest that toxins, permeant cations, and blocking cations can interact with a common site on the sodium channel near the extracellular surface. It is likely that permeant cations transiently bind to this superficial site, as the first of several steps in passing inward through the channel.     

In vitro fertilization with normal development in the sheep

Ovine tubal (n = 87) and ovarian in vitro matured oocytes (n = 99) were fertilized in vitro with ejaculated spermatozoa capacitated for 8 h in modified defined medium buffered with Hepes. High levels of fertilization were obtained as assessed by development to two-to six-cell stage within 40 h (75. 8% for ovulated and 62. 6% for in vitro matured oocytes). Electron microscope analysis of oocytes 20–22 h after insemination indicated that in vitro fertilization approximated the in vivo events. Embryos (two- to six-cell) were transferred surgically to the oviducts of pseudopregnant rabbits. Three days later, 42 (from ovulated oocytes) and 15 (from in vitro matured oocytes) embryos were recovered; 26 (61. 9%) and 10 (66. 6%), respectively, had cleaved at least once. Embryos incubated in vivo (n = 20 from ovulated oocytes; n = 9 from in vitro matured oocytes) were transferred surgically to the uteri of seven and four recipient ewes resulting in four and two pregnancies, respectively, from which three and one, respectively, have been maintained ( > 3 months). The first lamb resulting from the in vitro fertilization of an ovulated oocyte was born. In addition, six embryos (two- to four-cell) from tubal oocytes and ten embryos (two- to six-cell) from in vitro matured oocytes were directly transferred to the oviducts of two and three ewes, respectively. Two pregnancies resulting from in vitro matured fertilized oocytes are in progress ( > 3 months).     

Estrous sheep serum as a potent agent for ovine IVF: effect on cholesterol efflux from spermatozoa and the acrosome reaction

The aim of the present study was to evaluate the effect of heat-inactivated estrous sheep serum (ESS) on sheep IVF. When the capacitation and the fertilization media contained 20% ESS, a fertilization rate of 85% was achieved. The beneficial effect of ESS on sheep IVF was further demonstrated since the fertilization rate was null when ESS was omitted during sperm capacitation and fertilization. Estrous sheep serum supported both sperm capacitation and fertilization as shown by the results of experiments in which it was omitted during one of these steps: sperm capacitation in serum-free medium resulted in delayed sperm-oocyte penetration, while fertilization in serum-free medium significantly decreased the percentage of fertilized oocytes. To investigate the influence of serum on sperm ability to undergo the acrosome reaction, salt-stored follicular sheep oocytes were inseminated, and the acrosomal status of spermatozoa attached to zonae was examined by electron microscopy after a 4-h period of coincubation. Quantitative analysis on thin sections demonstrated that fewer acrosome-reacted spermatozoa were observed when the capacitation and insemination steps were carried out in DM-H medium without serum than in DM-H-SS supplemented with 20% ESS (0.08, [0; 0.34], (median, range)/100mum zona vs 1.32, [0.90; 2.28]/100mum zona; P < 0.01). Since a higher number of spermatozoa attached to the zona surface in DM-H medium, the proportion of acrosome-reacted spermatozoa was much lower (0.7%, [0%; 2.2%], (median, range) vs 54%, [25%; 100%]; P < 0.01) in the absence of serum. These results indicate that in our IVF system the development of the acrosome reaction depended on serum. Sperm cholesterol efflux during in vitro capacitation was measured on [(3)H] cholesterol labeled spermatozoa resuspended in DM-H or DM-H-SS medium. A time-dependent cholesterol removal was observed in the presence of serum (60 +/- 5%, mean +/- SD, after 5 h), whereas it was limited to 14 +/- 3 % in DM-H medium; hence addition of serum to the capacitation medium efficiently supports cholesterol efflux, which is thought to be a key-event in the capacitation process.     

Association between pygmy right whales (Caperea marginata) and areas of high marine productivity off Australia and New Zealand

Abstract

Opportunistic sightings and strandings of Caperea marginata (n=196) from the vicinity of Australia and New Zealand (1884 to early 2007) were used to relate geographic and temporal patterns to oceanographic and broad-scale climatic variability. Records were not uniformly distributed along the coast and more (69%) were from Australia than New Zealand. Seven coastal whale ‘hotspots’ were identified which accounted for 61% of records with locality data. Half of the hotspot records were from southeast (37) and northwest (20) Tasmania—others each had 9–15 events. Upwelling and/or high zooplankton abundance has been documented near all whale hotspots. Records of C. marginata occurred in all months, with 75% in spring and summer. Inter-annual variability showed broad agreement between increased whale records (usually in spring/summer) and strongly positive ‘Niño 3.4’ during 1980–1995 but not thereafter. Coastal upwelling and productivity increase during climatic phenomena such as El Niño and are likely to be quickly beneficial to plankton-feeding whales such as C. marginata.     

Egg-to-embryo transition is driven by differential responses to Ca(2+) oscillation number

Ca(2+) oscillations and signaling represent a basic mechanism for controlling many cellular events. Activation of mouse eggs entrains a temporal series of Ca(2+)-dependent events that include cortical granule exocytosis, cell cycle resumption with concomitant decreases in MPF and MAP kinase activities, and recruitment of maternal mRNAs. The outcome is a switch in cellular differentiation, i.e., the conversion of the egg into the zygote. By activating mouse eggs with experimentally controlled and precisely defined Ca(2+) transients, we demonstrate that each of these events is initiated by a different number of Ca(2+) transients, while their completion requires a greater number of Ca(2+) transients than for their initiation. This combination of differential responses to the number of Ca(2+) transients provides strong evidence that a single Ca(2+) transient-driven signaling system can initiate and drive a cell into a new developmental pathway, as well as can account for the temporal sequence of cellular changes associated with early development.     

Successful immunotherapy with matrix metalloproteinase-derived peptides in adjuvant arthritis depends on the timing of peptide administration

We have recently found that matrix metalloproteinases (MMPs) are targets for T-cell and B-cell reactivity in experimental arthritis. In the present article, we investigate whether modulation of MMP-specific T-cell responses could influence the course of adjuvant arthritis (AA). Lewis rats were treated nasally with MMP peptides prior to or after AA induction. Administration of the MMP-10 or the MMP-16 peptide prior to AA induction reduced the arthritic symptoms. In contrast, administration of the MMP-10 peptide after AA induction aggravated the arthritic symptoms. The present study shows the possible usefulness of MMP peptides for immunotherapy. However, a clear understanding of proper timing of peptide administration is crucial for the development of such therapies.     

Peptide fragments released from Phe-caseinomacropeptide in vivo in the rat

The aim of this study was to investigate the pharmacokinetics of bovine Phe-caseinomacropeptide (Phe-CMP) in the rat after oral administration. This polypeptide was monophosphorylated and mainly nonglycosylated: Phe-CMP-1P. During gastrointestinal digestion and absorption, Phe-CMP-1P was degraded. Intact Phe-CMP-1P and CMP-1P were rapidly released from the stomach. In contrast, partial hydrolysis by pancreatic enzymes was observed. In vitro hydrolysis by brush-border membrane vesicles also indicated that the peptide was degraded. In the blood, "CMP-immunoreactive material" appeared rapidly, reaching a maximum level of 5.5 microg/ml at 60 min.