Involvement of protein kinase C in 5-HT-stimulated ciliary activity in Helisoma trivolvis embryos.
1. During development, embryos of the pulmonate gastropod, Helisoma trivolvis, undergo a rotation behaviour due to the co-ordinated beating of three bands of ciliated epithelial cells. This behaviour is in part mediated by the neurotransmitter serotonin (5-HT) released from a pair of identified embryonic neurons. Using time-lapse videomicroscopy to measure ciliary beat frequency (CBF) in response to pharmacological manipulations, we determined whether protein kinase C (PKC) is involved in mediating 5-HT-stimulated ciliary beating. 2. Diacylglycerol (DAG) analogues sn-1,2-dioctanoyl glycerol (DiC8; 100 microM) and 1-oleoyl-2-acetyl-sn-glycerol (OAG; 100 microM), partially mimicked the 5-HT-induced increase in CBF. In contrast, application of OAG in the absence of extracellular Ca2+ did not result in an increase in CBF. 3. 5-HT-stimulated CBF was effectively blocked by PKC inhibitors bisindolylmaleimide (10 and 100 nM) and calphostin C (10 nM). In addition, bisindolylmaleimide (100 nM) inhibited DiC8-induced increases in CBF. At a higher concentration (200 nM), bisindolylmaleimide did not significantly reduce 5-HT-stimulated cilio-excitation. 4. Two different phorbol esters, phorbol 12-myristate 13-acetate (TPA; 0.1, 10 or 1000 nM) and phorbol 12beta, 13alpha-dibenzoate (PDBn; 10 microM) did not alter basal CBF. TPA (1 microM) did not alter 5-HT-stimulated CBF. Likewise, the synthetic form of phosphatidylserine, N-(6-phenylhexyl)-5-chloro-1-naphthalenesulphonamide (SC-9; 10 microM), did not increase CBF, whereas a strong increase in CBF was observed upon exposure to 5-HT. 5. The results suggest that a DAG-dependent, phorbol ester-insensitive isoform of PKC mediates 5-HT-stimulated CBF in ciliated epithelial cells from embryos of Helisoma trivolvis. (+info)
Vaccination with cathepsin L proteinases and with leucine aminopeptidase induces high levels of protection against fascioliasis in sheep.
The potential of different parasite proteinases for use as vaccine candidates against fascioliasis in sheep was studied by vaccinating animals with the cathepsin L proteinases CL1 and CL2 and with leucine aminopeptidase (LAP) purified from adult flukes. In the first trial, sheep were immunized with CL1 or CL2 and the mean protection levels obtained were 33 and 34%, respectively. Furthermore, a significant reduction in egg output was observed in sheep vaccinated either with CL1 (71%) or with CL2 (81%). The second trial was performed to determine the protective potential of the two cathepsin L proteinases assayed together, as well as in combination with LAP, and of LAP alone. The combination of CL1 and CL2 induced higher levels of protection (60%) than those produced when these enzymes were administered separately. Those sheep that received the cocktail vaccine including CL1, CL2, and LAP were significantly protected (78%) against metacercarial challenge, but vaccination with LAP alone elicited the highest level of protection (89%). All vaccine preparations induced high immunoglobulin G titers which were boosted after the challenge infection, but no correlations between antibody titers and worm burdens were found. However, the sera of those animals vaccinated with LAP contained LAP-neutralizing antibodies. Reduced liver damage, as assessed by the level of the liver enzyme gamma-glutamyl transferase, was observed in the groups vaccinated with CL1, CL2, and LAP or with LAP alone. (+info)
Characterization of a cDNA encoding a precursor polypeptide of a D-amino acid-containing peptide, achatin-I and localized expression of the achatin-I and fulicin genes.
Achatin-I and fulicin, isolated from the ganglia and atria of the giant land snail Achatina fulica, are a tetrapeptide and pentapeptide containing a d-Phe and d-Asn at position 2, respectively. We succeeded in cloning a cDNA encoding a precursor of achatin-I from the Achatina ganglia, revealing that the d-Phe present in achatin-I is coded by a common l-Phe codon, TTT or TTC. The deduced polypeptide was found to comprise seven repeats of the achatin sequence GFAD and one analog GFGD flanked on both sides by the typical endoproteolytic site KR. Northern blot analysis of transcripts and Southern blot analysis of reverse transcription (RT)-PCR products demonstrated that achatin-I mRNA was localized in the subesophageal ganglia, whereas expression of fulicin mRNA was detected in the atrium as well as in the subesophageal ganglia. Furthermore, localization of the achatin gene transcript in the right and left pedal ganglia compartments was shown by in situ hybridization on sections of subesophageal ganglia, whereas the fulicin transcript was observed in the right and left parietal ganglia. These data suggested that achatin-I plays an essential role in the regulation of the heart as a neurotransmitter or neurohormone through production in the pedal ganglia and transport to the atrium, whereas fulicin serves not only as a neurotransmitter or neurohormone but also as a novel atrial hormone. (+info)
Binding of carbon monoxide to alpha-hemocyanin and beta-hemocyanin from Helix pomatia.
The binding of carbon monoxide to alpha and beta-hemocyanin from the snail Helix pomatia was studied under equilibrium conditions. Homotropic interactions upon carbon monoxide binding were much weaker than upon the binding of oxygen. Heterotropic interactions (Bohr effect and calcium-ion effect), however, were just as strong as in the case of the binding of oxygen. For alpha-hemocyanin a linkage has been observed between the binding of carbon monoxide and a change in quaternary structure of the protein. (+info)
A special pattern of the smooth endoplasmic reticulum in the kidney of the snail Cryptomphalus aspersa.
A special structural pattern of the smooth endoplasmic reticulum (SER) has been observed in the kidney of the snail Cryptomphalus aspersa. Two types of cells (clear and dark) cover the foldings of the renal sac; the dark cells are by far the most numerous. A cisterna of SER enveloping the nucleus appears invariably in both types of cells, with no disruptions, or small ones (from 50 to 90 nm) along its profile. The layer of cytoplasm lodged between the external nuclear membrane and this cisterna is found invariably to be from 0-20 to 0-25 mum in width. Glycogen is abundant in the cytoplasm as alpha particles, and also in the nucleus, but as beta particles. It is noteworthy that absolutely no glycogen is present in the layer of cytoplasm lodged between the nuclear membrane and the surrounding SER envelope. Long profiles of SER are also observed closely approaching and parallel to the plasma membrane of the dark cells. Considering the role of SER in glycogen metabolism in the kidney of the snail, the possible function of these cisternae as a support system ofr enzymes involved in the metabolism of glucides is discussed. (+info)
Karyotypes on three species of Chinese mesogastropod snails, Semisulcospira libertina, S. dolichostoma and Viviparus rivularis.
Three species of the families Viviparidae and Pleuroceridae, the first intermediate host of paragonimiasis, metagonimiasis and echinostomiasis were studied cytologically. The observed diploid chromosome number was as follows: Semisulcospira libertina 36, S. dolichostoma 34, and Viviparus rivularis 64. The mitotic chromosome complement of S. libertina has nine metacentric pairs and nine submetacentric pairs, and S. dolichostoma has three metacentric pairs and 14 submetacentric pairs of chromosomes. Viviparus rivularis showed two metacentric pairs and 30 submetacentric pairs of chromosomes. (+info)
Macromolecular antimicrobial glycoprotein, achacin, expressed in a methylotrophic yeast Pichia pastoris.
A cDNA encoding achacin, an antimicrobial glycoprotein from the body surface mucus of giant African snail Achacina fulica Ferussac, was expressed in a methylotrophic yeast, Pichia pastoris, and recombinant achacin (rAch) was secreted in yeast minimal medium in a polyglycosylated form with 80 kDa. Carbohydrate analysis revealed that the glycosylated moiety of rAch was composed of 50 mol mannose and 2 mol N-acetylglucosamine residues. Antimicrobial activity using Escherichia coli and Staphylococcus aureus showed that the rAch had a behavior similar to its native counterpart. The rAch showed so wide an antimicrobial spectrum that 0.1 mg/ml rAch inhibited the growth of Pseudomonas fluorescens, Staphylococcus epidermidis, and Streptococcus faecalis in addition to E. coli and S. aureus, whereas it did not appreciably affect the growth of Proteus mirabilis, Bacillus cereus and Micrococcus luteus. The rAch was also effective in preventing growth of Vibrio anguillarum and Vibrio parahaemolyticus. The results suggested that the rAch had great potential of using as an antimicrobial agent. (+info)
Osmoregulation and FMRFamide-related peptides in the salt marsh snail Melampus bidentatus (Say) (Mollusca: Pulmonata).
The pulmonate snail Melampus bidentatus occupies the high intertidal zone of salt marshes in a nearly terrestrial environment. The hemolymph osmolarity of the snails collected in the field paralleled that of the adjacent water and was affected by the tides and precipitation. The snails initially gained or lost weight when submerged in hypo- or hyperosmotic media, respectively, but returned to their original weight after 24 h. The content of their immunoreactive (IR)-FMRFamide-Related Peptides (FaRPs) was measured in various tissues by radioimmunoassay, and IR-FaRPs were found in every tissue analyzed. The subesophageal part of the central nervous system (CNS) contained more IR-FaRPs than the supraesophageal part, and the kidney and the tissues of the reproductive tract contained more than other peripheral tissues. The levels of IR-FaRPs in the CNS, kidney, and hemolymph were higher in snails that were immersed in higher concentrations of seawater. Many IR neurons are present in all ganglia of the CNS except the pleural ganglia, and IR neurites are extensively distributed within the CNS and its connective tissue sheath. The visceral nerve from the visceral ganglion is immunoreactive and could be seen to innervate the kidney, which contains IR-varicosities. An osmoregulatory role for the FaRPs is suggested. (+info)