Effects of ulapualide A and synthetic macrolide analogues on actin dynamics and gene regulation

Several marine macrolide toxins act as potent and specific actin-severing molecules. Recent elucidation of their stereochemistries and modes of interaction with actin has allowed the syntheses of bioactive analogues. Here we used synthetic analogues in a structure-function analysis of ulapualide A, a trisoxazole-based macrolide. Ulapualide A harboured potent actin-depolymerising activity both in cells and in vitro. Its synthetic diastereoisomer was three orders of magnitude less active than the natural toxin and synthetic macrolide fragments lacked actin-capping/ severing activity altogether. Modulation of serum response factor (SRF)-dependent gene expression, as described for other actin-binding toxins, was also examined. Specific changes in response to ulapualide A were not observed, primarily due to its profound effects on cytoskeletal integrity and cell adhesion. Several synthetic fragments of ulapualide A also had no effect on SRF-dependent gene expression. However, inhibition was observed with a molecule corresponding to the extended aliphatic side chain of halichondramide, a structurally related macrolide. These findings indicate that side-chain derivatives of trisoxazole-based macrolides may serve to uncouple gene-regulatory events from actin dynamics. E. Vincent and J. Saxton: These two authors contributed equally Received 27 September 2006; received after revision 30 November 2006; accepted 8 January 2007     

Evidence for two populations of excitatory receptors for noradrenaline on arteriolar smooth muscle

We have recorded the responses of arteriolar smooth muscle cells to iontophoretically applied noradrenaline. Records of both muscle movement and muscle membrane potential were made. We found that two distinct types of response could be detected, depending on the position of the noradrenaline micropipette. One type of response consisted of a localised constriction near the noradrenaline source: this effect could be abolished by the alpha-antagonist phentolamine and was not associated with a change in arteriolar membrane potential. The other type of response was a depolarisation similar to the excitatory junction potentials (e.j.ps) produced by sumpathetic nerve stimulation. These observations suggest that there are two populations of receptors for noradrenaline on arterioles, and could explain the paradoxical failure of alpha-antagonists to block neuromuscular transmission at some sutonomic end organs such as the vas deferens, arteries and arterioles.     

Cholinergic properties of 1-methyl and 2-methylpentyltrimethylammonium salts

Résumé Étude des propriétés cholinergiques des iodures du méthyl-1 et du méthyl-2-pentyltriméthylammonium. Dans leur comportement agonistique, ces sels offrent des variations parallèles à celles de dérivés méthyles de l'acétylcholine. Le cation pentyltriméthylammonium est donc capable de s'associer aux récepteurs cholinergiques tels que l'acétylcholine.

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This investigation was supported by Grant No. MA-3359 to M.H. from the Medical Research Council of Canada.     

Identification of chicken lysozyme <Emphasis Type="Italic">g</Emphasis>2 and its expression in the intestine

Lysozyme is an important component of the innate immune system, protecting the gastrointestinal tract from infection. The aim of the present study was to determine if lysozyme is expressed in the chicken (Gallus gallus) intestine and to characterise the molecular forms expressed. Immunohistochemical staining localised lysozyme to epithelial cells of the villous epithelium along the length of the small intestine. There was no evidence for lysozyme expression in crypt epithelium and no evidence for Paneth cells. Immunoblots of chicken intestinal protein revealed three proteins: a 14-kDa band consistent with lysozyme c, and two additional bands of approximately 21 and 23 kDa, the latter consistent with lysozyme g. RT-PCR analyses confirmed that lysozyme c mRNA is expressed in 4-day, but not older chicken intestine and lysozyme g in 4- to 35-day chicken intestine. A novel chicken lysozyme g2 gene was identified by in silico analyses and mRNA for this lysozyme g2 was identified in the intestine from chickens of all ages. Chicken lysozyme g2 shows similarity with fish lysozyme g, including the absence of a signal peptide and cysteines involved in disulphide bond formation of the mammalian and bird lysozyme g proteins. Analyses using SecretomeP predict that chicken lysozyme g2 may be secreted by the non-classical secretory pathway. We conclude that lysozyme is expressed in the chicken small intestine by villous enterocytes. Lysozyme c, lysozyme g and g2 may fulfil complimentary roles in protecting the intestine.Received 4 August 2004; received after revision 1 September 2004; accepted 7 September 2004     

Analgesic effects of 3-carboxysalsolinol alone and in combination with morphine

Summary A biphasic dose-response pattern is generated by the isoquinoline, 3-carboxysalsolinol, in analgesia tests conducted in mice. Carbidopa pretreatment enhances this effect, as well as the morphine-induced analgesic increase by 3-carboxysalsolinol. Naloxone blockade of all of these responses suggests an interaction of the alcohol-based isoquinoline with central opiate receptors.We gratefully acknowledge the receipts of drugs from Merck and Co., West Point, Pennsylvania, and Endo Laboratories, Garden City, New York. A. M. is the recipient of a research scholarship from the Alcoholism and Drug Addiction Research Foundation, Toronto, Ontario, Canada.