Induction of direct endosome to endoplasmic reticulum transport in Chinese hamster ovary (CHO) cells (LdlF) with a temperature-sensitive defect in epsilon-coatomer protein (epsilon-COP)

In the present study we demonstrate that ricin, apparently without passing through the Golgi apparatus, reaches the endoplasmic reticulum (ER) and intoxicates cells in which the Golgi apparatus has been vesiculated by depletion of epsilon-COP, a subunit of COPI. LdlF cells contain a temperature-sensitive mutation in epsilon-COP. At the nonpermissive temperature epsilon-COP is degraded, and the Golgi apparatus undergoes a morphological change. To study ricin transport in these cells we used ricin sulf-2, a modified ricin molecule containing glycosylation and sulfation sites. Measurements of the incorporation of radioactive mannose into ricin sulf-2 showed that ricin reached the ER in cells depleted of epsilon-COP. Importantly, by investigating the glycosylation of ricin sulf-2 that was modified with radioactive sulfate in the trans-Golgi network, it was demonstrated that transport of ricin to the ER via the Golgi apparatus was severely inhibited. Moreover, we found that ricin was able to intoxicate ldlF cells depleted of epsilon-COP in the presence of brefeldin A. In contrast, control cells were completely protected against ricin by brefeldin A. In conclusion, our results suggest that in ldlF cells depleted of epsilon-COP ricin might be transported to the ER by an induced brefeldin A-resistant pathway that circumvents the Golgi apparatus.     

Weak geographical structure in sperm morphology across the range of two willow warbler Phylloscopus trochilus subspecies in Scandinavia

Sperm morphology is highly diversified among species and at higher taxonomic levels. In birds, there is also increasing evidence of geographical differentiation in sperm traits within species, especially in those with strong sperm competition. Geographical divergences in sperm traits might imply the formation of a reproductive barrier in a speciation process. Here we study sperm morphology variation of willow warblers Phylloscopus trochilus in a geographical context in Scandinavia, across the range of two subspecies that are differentiated in certain genetic markers, morphology and migratory routes. We describe geographical patterns in genotypes (two previously described single‐nucleotide polymorphism (SNP) markers and four polymorphic microsatellites); in wing length, tarsus length and body mass; and in sperm traits of 330 male willow warblers sampled at 33 localities across Norway (58°N–69°N). Birds were on average larger and longer‐winged in the north (spp. acredula) than in the south (spp. trochilus), and showed a sigmoid change in the SNP allele frequencies and body morphology around 65°N. We found no evidence of genetic structuring in the microsatellites. There was no geographical variation in sperm traits across Norway, except that sperm heads were on average longer in the south. Sperm head length was also associated with the two SNP markers, with longer sperm heads for the southern alleles, and midpiece length was weakly associated with one of the SNP markers. Similar among‐male variances in total sperm length among the 33 sampling sites indicate uniform levels of sperm competition across Norway. We conclude that sperm morphology remains a rather undifferentiated trait between the two willow warbler subspecies in Scandinavia, which is consistent with a pattern of a shallow genetic divergence. This indicates that sperm morphology is not a reproductive barrier maintaining the narrow hybrid zone.     

Which clinical and experimental data link temporal lobe epilepsy with depression?

The association of temporal lobe epilepsy with depression and other neuropsychiatric disorders has been known since the early beginnings of neurology and psychiatry. However, only recently have in vivo and ex vivo techniques such as Positron Emission Tomography, Magnetic Resonance Imaging and Magnetic Resonance Spectroscopy in combination with refined animal models and behavioral tests made it possible to identify an emerging pattern of common pathophysiological mechanisms. We now have growing evidence that in both disorders altered interaction of serotonergic and noradrenergic neurons with glutamatergic systems is associated with abnormal neuronal circuits and hyperexcitability. Neuronal hyperexcitability can possibly evoke seizure activity as well as disturbed emotions. Moreover, decreased synaptic levels of neurotransmitters and high glucocorticoid levels influence intracellular signaling pathways such as cAMP, causing disturbances of brain-derived and other neurotrophic factors. These may be associated with hippocampal atrophy seen on Magnetic Resonance Imaging and memory impairment as well as altered fear processing and transient hypertrophy of the amygdala. Positron Emission Tomography studies additionally suggest hypometabolism of glucose in temporal and frontal lobes. Last, but not least, in temporal lobe epilepsy and depression astrocytes play a role that reaches far beyond their involvement in hippocampal sclerosis and ultimately, therapeutic regulation of glial-neuronal interactions may be a target for future research. All these mechanisms are strongly intertwined and probably bidirectional such that the structural and functional alterations from one disease increase the risk for developing the other. This review provides an integrative update of the most relevant experimental and clinical data on temporal lobe epilepsy and its association with depression.     

Sperm performance in conspecific and heterospecific female fluid

Divergent sexual selection within allopatric populations may result in divergent sexual phenotypes, which can act as reproductive barriers between populations upon secondary contact. This hypothesis has been most tested on traits involved in precopulatory sexual selection, with less work focusing on traits that act after copulation and before fertilization (i.e., postcopulatory prezygotic traits), particularly in internally fertilizing vertebrates. However, postcopulatory sexual selection within species can also drive trait divergence, resulting in reduced performance of heterospecific sperm within the female reproductive tract. Such incompatibilities, arising as a by‐product of divergent postcopulatory sexual selection in allopatry, can represent reproductive barriers, analogous to species‐assortative mating preferences. Here, we tested for postcopulatory prezygotic reproductive barriers between three pairs of taxa with diverged sperm phenotypes and moderate‐to‐high opportunity for postcopulatory sexual selection (barn swallows Hirundo rustica versus sand martins Riparia riparia, two subspecies of bluethroats, Luscinia svecica svecica versus L. s. namnetum, and great tits Parus major versus blue tits Cyanistes caeruleus). We tested sperm swimming performance in fluid from the outer reproductive tract of females, because the greatest reduction in sperm number in birds occurs as sperm swim across the vagina. Contrary to our expectations, sperm swam equally well in fluid from conspecific and heterospecific females, suggesting that postcopulatory prezygotic barriers do not act between these taxon pairs, at this stage between copulation and fertilization. We therefore suggest that divergence in sperm phenotypes in allopatry is insufficient to cause widespread postcopulatory prezygotic barriers in the form of impaired sperm swimming performance in passerine birds.     

Limbic Structures Show Altered Glial–Neuronal Metabolism in the Chronic Phase of Kainate Induced Epilepsy

A better understanding is needed of how glutamate metabolism is affected in mesial temporal lobe epilepsy (MTLE). Here we investigated glial–neuronal metabolism in the chronic phase of the kainate (KA) model of MTLE. Thirteen weeks following systemic KA, rats were injected i.p. with [1-¹³C]glucose. Brain extracts from hippocampal formation, entorhinal cortex, and neocortex, were analyzed by ¹³C and ¹H magnetic resonance spectroscopy to quantify ¹³C labeling and concentrations of metabolites, respectively. The amount and ¹³C labeling of glutamate were reduced in the hippocampal formation and entorhinal cortex of epileptic rats. Together with the decreased concentration of NAA, these results indicate neuronal loss. Additionally, mitochondrial dysfunction was detected in surviving glutamatergic neurons in the hippocampal formation. In entorhinal cortex glutamine labeling and concentration were unchanged despite the reduced glutamate content and label, possibly due to decreased oxidative metabolism and conserved flux of glutamate through glutamine synthetase in astrocytes. This mechanism was not operative in the hippocampal formation, where glutamine labeling was decreased. In neocortex labeling and concentration of GABA were increased in epileptic rats, possibly representing a compensatory mechanism. The changes in the hippocampus might be of pathophysiological importance and merit further studies aiming at resolving metabolic causes and consequences of MTLE. Special issue article in honor of Dr. Frode Fonnum.     

Evaluation of prospective, routine application of Ki-67 immunoquantitation in early CIN for assessment of short-term progression risk

OBJECTIVE: To prospectively validate, in early cervical intraepithelial neoplasia (CIN), routine assessment of a previously developed prognostic Ki-67 immunoquantitative progression-risk model. STUDY DESIGN: Two hundred sixty-six consecutive cervical biopsies taken for an abnormal cytologic smear were routinely diagnosed by experienced pathologists as CIN. Ki-67 immunoquantitation was performed routinely by 3 technicians blinded to clinical and pathologic information. Progression of CIN 1-2 to CIN 3 in histologic follow-up biopsies was used as the intermediate end point. RESULTS: In 58 (22%) biopsies, technical shortcomings prevented Ki-67 immunoquantitation, and in 22 biopsies no follow-up was available. The routine diagnosis in the 186 remaining biopsies was CIN 1 = 24, CIN 2 = 56 and CIN 3 = 106. In 52 marker biopsies with expert review diagnosis of CIN 1-2 and adequate follow-up, histologic biopsies revealed CIN 3 in 9 (17%) cases: 9 of 34 (26%) of Ki-67 high-risk and 0 of 18 (0%) of Ki-67 low-risk lesions (log rank = 5.0, P = .03). Routine CIN grade (1 or 2) was not prognostic (P = .65). Eleven (55%) of 20 CIN 1 and 7 of 32 (22%) CIN 2 cases were Ki-67 low risk and none progressed, contrasting with 4 of 9 (44%) progressions of Ki-67 high risk CIN 1s and 5 of 25 (20%) high risk CIN 2s. Expert CIN grades were stronger prognostically than routine CIN grade, but Ki-67 was still stronger. CONCLUSION: Routine Ki-67 immunoquantitative progression prediction in CIN 1-2 is more predictive of CIN 3 in follow-up than are routine and review CIN grades.     

Chromosomal differences between European and North American Atlantic salmon discovered by linkage mapping and supported by fluorescence in situ hybridization analysis

ABSTRACT: BACKGROUND: Geographical isolation has generated a distinct difference between Atlantic salmon of European and North American Atlantic origin. The European Atlantic salmon generally has 29 pairs of chromosomes and 74 chromosome arms whereas it has been reported that the North American Atlantic salmon has 27 chromosome pairs and an NF of 72. In order to predict the major chromosomal rearrangements causing these differences, we constructed a dense linkage map for Atlantic salmon of North American origin and compared it with the well-developed map for European Atlantic salmon. RESULTS: The presented male and female genetic maps for the North American subspecies of Atlantic salmon, contains 3,662 SNPs located on 27 linkage groups. The total lengths of the female and male linkage maps were 2,153 cM and 968 cM respectively, with males characteristically showing recombination only at the telomeres. We compared these maps with recently published SNP maps from European Atlantic salmon, and predicted three chromosomal reorganization events that we then tested using fluorescence in situ hybridization (FISH) analysis. The proposed rearrangements, which define the differences in the karyotypes of the North American Atlantic salmon relative to the European Atlantic salmon, include the translocation of the p arm of ssa01 to ssa23 and polymorphic fusions: ssa26 with ssa28, and ssa08 with ssa29. CONCLUSIONS: This study identified major chromosomal differences between European and North American Atlantic salmon. However, while gross structural differences were significant, the order of genetic markers at the fine-resolution scale was remarkably conserved. This is a good indication that information from the International Cooperation to Sequence the Atlantic salmon Genome, which is sequencing a European Atlantic salmon, can be transferred to Atlantic salmon from North America.     

The Cardiac Ventricular 5-HT4 Receptor Is Functional in Late Foetal Development and Is Reactivated in Heart Failure

A positive inotropic responsiveness to serotonin, mediated by 5-HT₄ and 5-HT_2A receptors, appears in the ventricle of rats with post-infarction congestive heart failure (HF) and pressure overload-induced hypertrophy. A hallmark of HF is a transition towards a foetal genotype which correlates with loss of cardiac functions. Thus, we wanted to investigate whether the foetal and neonatal cardiac ventricle displays serotonin responsiveness. Wistar rat hearts were collected day 3 and 1 before expected birth (days -3 and -1), as well as day 1, 3, 5 and 113 (age matched with Sham and HF) after birth. Hearts from post-infarction HF and sham-operated animals (Sham) were also collected. Heart tissue was examined for mRNA expression of 5-HT₄, 5-HT_2A and 5-HT_2B serotonin receptors, 5-HT transporter, atrial natriuretic peptide (ANP) and myosin heavy chain (MHC)-α and MHC-β (real-time quantitative RT-PCR) as well as 5-HT-receptor-mediated increase in contractile function ex vivo (electrical field stimulation of ventricular strips from foetal and neonatal rats and left ventricular papillary muscle from adult rats in organ bath). Both 5-HT₄ mRNA expression and functional responses were highest at day -3 and decreased gradually to day 5, with a further decrease to adult levels. In HF, receptor mRNA levels and functional responses reappeared, but to lower levels than in the foetal ventricle. The 5-HT_2A and 5-HT_2B receptor mRNA levels increased to a maximum immediately after birth, but of these, only the 5-HT_2A receptor mediated a positive inotropic response. We suggest that the 5-HT₄ receptor is a representative of a foetal cardiac gene program, functional in late foetal development and reactivated in heart failure.     

Development of Saccharina latissima (Phaeophyceae) kelp hatcheries with year-round production of zoospores and juvenile sporophytes on culture ropes for kelp aquaculture

Saccharina latissima is attractive for industrial cultivation for different usages, such as biofuels, feed supplements, and derivation of chemicals. A continuous supply of kelp sporelings throughout the year may ensure a year-round production of kelp juveniles on ropes. In this study, induction of sporangial areas (sorus portions) on the blade of S. latissima was performed throughout the year at three locations: Trondheim (Norway), Grenaa (Denmark), and Sylt island (Germany). The results indicate that a year-round sorus induction in S. latissima is possible and that this induction is controlled by applying short-day treatment of adult sporophytes throughout the year and by the removal of the basal blade meristem. The artificially induced and released zoospores formed viable sporelings at all seasons, but cultivation in the sea in Norway was successful only during autumn, winter, and spring, while the growth conditions were poor during the summer. The results are important for industrial scale-up and continuous production of kelp biomass.