Evidence that substance P may be a modulator rather than a transmitter of noxious mechanical stimulation

Substance P (SP), injected intrathecally, produced a dose-related increase in responsiveness (hyperalgesia) in a pressure test for nociceptive thresholds. Pretreatment with two doses of SP produced complete desensitization to this response but did not alter base-line responsiveness. The hyperalgesic response to SP and the lack of change in base line following desensitization to SP suggest a modulator rather than a transmitter role for SP in the transmission of noxious mechanical stimulation.     

An insect picornavirus may have genome organization similar to that of caliciviruses.

Computer-assisted analysis of the amino acid sequence of the product encoded by the sequenced 3' portion of the cricket paralysis virus (CrPV), an insect picornavirus, genome showed that this protein is homologous not to the RNA-directed RNA polymerases, as originally suggested, but to the capsid proteins of mammalian picornaviruses. Alignment of the CrPV protein sequence with those of picornavirus and calicivirus capsid proteins demonstrated that the sequenced portion of the insect picornavirus genome encodes the C-terminal part of VP3 and the entire VP1. Thus CrPV seems to have a genome organization distinct from that of other picornaviruses but closely resembling that of caliciviruses, with the capsid proteins encoded in the 3' part of the genome. On the other hand, the tentative phylogenetic trees generated from the VP3 alignment revealed grouping of CrPV with hepatitis A virus, a true picornavirus, not with caliciviruses. Thus CrPV may be a picornavirus with a calicivirus-like genome organization. Different options for CrPV genome expression are discussed.     

Japanese medaka Hox paralog group 2: insights into the evolution of Hox PG2 gene composition and expression in the Osteichthyes

Hox paralog group 2 (PG2) genes function to specify the development of the hindbrain and pharyngeal arch-derived structures in the Osteichthyes. In this article, we describe the cDNA cloning and embryonic expression analysis of Japanese medaka (Oryzias latipes) Hox PG2 genes. We show that there are only two functional canonical Hox genes, hoxa2a and b2a, and that a previously identified hoxa2b gene is a transcribed pseudogene, psihoxa2b. The functional genes, hoxa2a and b2a, were expressed in developing rhombomeres and pharyngeal arches in a manner that was relatively well conserved compared with zebrafish (Danio rerio) but differed significantly from orthologous striped bass (Morone saxatilis) and Nile tilapia (Oreochromis niloticus) genes, which, we suggest, may be owing to effects of post-genome duplication loss of a Hox PG2 gene in the medaka and zebrafish lineages. psihoxa2b was expressed at readily detectable levels in several noncanonical Hox expression domains, including the ventral aspect of the neural tube, the pectoral fin buds and caudal-most region of the embryonic trunk, indicative that regulatory control elements needed for spatio-temporal expression have diverged from their ancestral counterparts. Comparative expression analyses showed medaka hoxa2a and b2a expression in the 2nd pharyngeal arch (PA2) beyond the onset of chondrogenesis, which, according to previous hypotheses, suggests these genes function redundantly as selector genes of PA2 identity. We conclude that Hox PG2 gene composition and expression have diverged significantly during osteichthyan evolution and that this divergence in teleosts may be related to lineage-dependent differential gene loss following an actinopterygian-specific whole genome duplication.     

The human L-pipecolic acid oxidase is similar to bacterial monomeric sarcosine oxidases rather than D-amino acid oxidases

L-Pipecolic acid oxidase activity is deficient in patients with peroxisome biogenesis disorders (PBDs). Because its role, if any, in these disorders is unknown, the authors cloned the human gene to order to further study its functions. BLAST search of the translated sequence showed greatest homology to Bacillus sp. NS-129 monomeric sarcosine oxidase. The purified enzyme could use either L-pipecolic acid or sarcosine as a substrate. No homology was found to the peroxisomal D-amino acid oxidases. A further comparison of L-pipecolic acid oxidase to the two D-amino acid oxidases in peroxisomes showed that the proteins differed in many ways. First, both D-amino acid oxidase and L-pipecolic acid oxidase showed no enzyme activity in liver from Zell-weger syndrome patients; D-aspartate oxidase activity was unchanged from control levels. Although all were targeted to peroxisomes, their targeting signals differed. No L-pipecolic acid oxidase was found in brain or other tissues outside of liver and kidney. The D-amino acid oxidases were similarly and more widely distributed. Finally, although D-amino acid degradation is limited to peroxisomes in mammals, L-pipecolic acid can be oxidized in either mitochondria or peroxisomes, or both.     

Evidence that glucagon stabilizes rather than activates mitochondrial functions in rat liver

The present study is concerned with the question as to whether the acute treatment of intact rats or hepatocytes with glucagon and dibutyryl cAMP, respectively, leads to a stabilization or an activation of mitochondrial functions, such as state-3 respiration, succinate dehydrogenase activity and pyruvate carboxylase activity. For this purpose, the influence of various parameters of mitochondria preparation (isolation medium, washing steps, storage) as well as of phospholipase A inhibitors (cinchocain, chloroquine) on the expression of the hormone effect was examined. With regard to the above mentioned functions, the values displayed by control mitochondria were found to be considerably higher if mannitol instead of sucrose had been used for isolation. Accordingly, only small effects of hormone treatment became apparent. The addition of cinchocain or chloroquine to the sucrose medium yielded results similar to those obtained with mannitol. Furthermore, the hormone effect on state-3 respiration and succinate dehydrogenase activity was only small if the mitochondria had been prepared faster than usual and had been used without washing. Regarding pyruvate carboxylase, a considerably smaller glucagon effect was observed when it was assayed at 25 degrees C and not (as usual) at 37 degrees C. Our results indicate that glucagon application stabilizes rather than activates mitochondrial functions.     

Current status of sperm cryopreservation in biomedical research fish models: zebrafish, medaka, and Xiphophorus

Aquarium fishes are becoming increasingly important because of their value in biomedical research and the ornamental fish trade, and because many have become threatened or endangered in the wild. This review summarizes the current status of sperm cryopreservation in three fishes widely used in biomedical research: zebrafish, medaka, and live-bearing fishes of the genus Xiphophorus, and will focus on the needs and opportunities for future research and application of cryopreservation in aquarium fish. First, we summarize the basic biological characteristics regarding natural habitat, testis structure, spermatogenesis, sperm morphology, and sperm physiology. Second, we compare protocol development of sperm cryopreservation. Third, we emphasize the importance of artificial fertilization in sperm cryopreservation to evaluate the viability of thawed sperm. We conclude with a look to future research directions for sperm cryopreservation and the application of this technique in aquarium species.     

Parasite infection rather than tactile stimulation is the proximate cause of cleaning behaviour in reef fish.

Cleaning behaviour is a popular example of non-kin cooperation. However, quantitative support for this is generally sparse and the alternative, that cleaners are parasitic, has also been proposed. Although the behaviour involves some of the most complex and highly developed interspecific communication signals known, the proximate causal factors for why clients seek cleaners are controversial. However, this information is essential to understanding the evolution of cleaning. I tested whether clients seek cleaners in response to parasite infection or whether clients seek cleaners for tactile stimulation regardless of parasite load. Parasite loads on client fish were manipulated and clients exposed to cleaner fish and control fish behind glass. I found that parasitized client fish spent more time than unparasitized fish next to a cleaner fish. In addition, parasitized clients spent more time next to cleaners than next to control fish, whereas unparasitized fish were not attracted to cleaners. This study shows, I believe for the first time, which is somewhat surprising, that parasite infection alone causes clients to seek cleaning by cleaners and provides insight into how this behaviour evolved.     

Low transposition frequency of the medaka fish Tol2 element may be due to extranuclear localization of its transposase

Transposase proteins of some highly active DNA-based transposable elements, such as the maize Activator element, are known to possess nuclear localization signals (NLSs). We examined if this is also the case for the transposase of the medaka fish Tol2 element, a member of the hAT (hobo/Activator/Tam3) transposable element family, using human and mouse culture cells. Unexpectedly, the transposase-lacZ fusion protein, in which the lacZ is a location marker, was found to be present in the cytoplasm rather than in the nucleus, suggesting that the Tol2 transposase contains a signal for extranuclear localization. The same staining pattern was also observed with a fusion protein containing a 33-amino-acid region at about the center of the primary structure of the transposase. The Tol2 element might have a mechanism to control its transposition frequency that includes extranuclear localization of its transposase.     

Selective binding of zinc ions to heparin rather than to other glycosaminoglycans.

The relative binding affinity of Zn2+ to several glycosaminoglycans was determined by gel-filtration chromatography. Binding was observed only between Zn2+ and heparin. No binding was observed between Zn2+ and chondroitin 4-sulphate, chondroitin 6-sulphate, dermatan sulphate of hyaluronic acid. All of the glycosaminoglycans contained carboxy groups, but only heparin bound Zn2+. This observation suggests that, contrary to a previously proposed hypothesis, simple electrostatic interactions between the negatively charged carboxy groups of the glycosaminoglycans and the positively charged Zn2+ cannot explain the observed binding.     

The Hox gene family in transgenic mice.

Evidence is accumulating which suggests that the vertebrate Hox homeobox gene family plays an important role in pattern formation, particularly in the specification of regional diversity. In the last year important advances in the understanding of their regulation and function have been provided using transgenic mice.     

Summer rain pulses may stimulate a CO2 release rather than absorption in desert halophyte communities

Background and aims

The response of plants and soil to rain pulses determines seasonal variations in the exchange of materials and energy at the ecosystem scale in arid and semi-arid regions. We assessed how the ecosystem carbon exchange (NEE) of desert halophyte communities of different plant functional-types responds to summer precipitation pulses in Tamarix and Haloxylon communities.

Methods

Plant water status, photosynthetic gas exchange, soil respiration and net ecosystem carbon exchange were measured to test the hypothesis that high physiological sensitivity may induce a greater changes in NEE resulting from the summer precipitation pulses in Haloxylon community.

Results

Plant water status and photosynthetic assimilation did not differ before and after summer precipitation pulses in either community. In contrast, soil respiration and NEE responded strongly to summer precipitation events in both communities. At the ecosystem level, precipitation pulses induced a pulse of CO₂ release, rather than absorption. The NEE response to summer precipitation was less in the deep-rooted Tamarix community, compared to the shallow-rooted Haloxylon community, which was even converted into a carbon source after summer precipitation inputs. As a result, the effect of summer precipitation inputs on soil respiration was more important than the plant (carbon assimilation) response in determining the ecosystem response to episodic precipitation pulses.     

Heavy chain variable region gene families evolved early in phylogeny. Ig complexity in fish

The V regions of channel catfish H chain cDNA clones have been analyzed. Based upon sequence relationships and hybridization analyses, five different groups of VH genes are identified whose definition is consistent with that of five different VH families. Genomic Southern blots indicate that as many as 100 different germ-line VH genes are likely represented by these families. The sequence diversity between identified members of these different families is similar in magnitude to the divergence represented between members of different human or mouse VH families. The FR regions are the most conserved regions when members of different catfish VH families are compared; specific amino acid positions appear to be highly conserved in phylogeny. Equally important is that diversity is represented in complementarity-determining regions CDR1 and CDR2 in members of the different families as well as in members of the same VH family. These results suggest that an extensive repertoire of VH genes can contribute to antibody diversity in this lower vertebrate. Sequence comparisons indicate that one of the catfish VH families shares considerable structural similarity to several higher vertebrate VH gene families--a relationship which suggests that this VH family may be ancestral to some VH gene families of higher vertebrates. Characteristic of the genomic organization of higher vertebrate H chains, catfish appear to have different VH families wherein a VH gene likely undergoes functional recombination with putative DH gene segments and one of apparently several different JH segments. The recombined V region is expressed with the same C region gene. These combined results suggest that bony fishes are the earliest known phylogenetic representatives to have evolved extensive V region gene families.     

Production of transgenic fish: introduction and expression of chicken delta-crystallin gene in medaka embryos

To produce a model of transgenic fish, recombinant plasmids containing chicken delta-crystallin gene were microinjected into the oocyte nucleus of a small teleost, medaka (Oryzias latipes). About 50% of the microinjected oocytes developed to 7-day-old embryos. By Southern blotting delta-crystallin gene was detected in 4 of 8 embryos, and, by Western blotting, delta-crystallin polypeptides in 5 of 16. In 1 of 6 examined histologically, delta-crystallin DNA was detected in all the tissues, and delta-crystallin polypeptides, in many of the tissues including the lens. Thus, the exogenous gene and/or its products were detected in 10 of 30 embryos examined. This is the first report of successful production of transgenic fish.