Analysis of the Structure and Electrophysiological Actions of Halitoxins: 1,3 Alkyl-pyridinium Salts from Callyspongia ridleyi

Abstract

We have chemically characterized a preparation of halitoxins, (1,3 alkyl-pyridinium salts) isolated from the marine sponge Callyspongia ridleyi. At concentrations of 50 and 5 microg/ml the halitoxin preparation caused irreversible membrane potential depolarization, decreased input resistance and inhibited evoked action potentials when applied to cultured dorsal root ganglion neurones. Under whole cell voltage clamp the halitoxins produced an increase in cation conductance that was attenuated by replacing sodium with N-methyl-d-glucamine. Fura-2 fluorescence ratiometric calcium imaging was used to directly measure calcium flux into neurones after exposure to halitoxins. Calcium influx, evoked by the halitoxins, persisted when the neurones were bathed in medium containing the voltage-activated calcium channel antagonists cadmium and nickel. Experiments on undifferentiated F-11 cells showed little or no calcium influx in response to depolarizing concentrations of potassium and indicated that halitoxins evoked massive calcium influx in the absence of voltage-activated calcium channels. The halitoxins also produced transient increases in intracellular calcium when F-11 cells were bathed in calcium-free medium suggesting that the toxins could release calcium from intracellular stores. The pore-forming action of the halitoxins was identified when the toxins were applied to artificial lipid bilayers composed of phosphatidylcholine and cholesterol. Halitoxins evoked channel-like activity in the lipid bilayers, with estimated unitary conductances of between 145pS and 2280pS, possibly indicating that distinct channels could be produced by the different components in the preparation of halitoxins.

Two new alkaloids from marine spongeCallyspongiasp.

“…Marine sponges of the genus Callyspongia have been found to contain a variety of structurally unique metabolites with interesting biological activities, and a number of alkaloids have been isolated from this genus so far (Buchanan et al, 2007;Daviescoleman et al, 1993;Scott et al, 2000;Wang, Kuramoto, & Uemura, 1996). In our previous study on the sponge Callyspongia sp., one new and three known diketopiperazines were reported (Yang et al, 2009).…”

Chemical synthesis and biological activities of 3-alkyl pyridinium polymeric analogues of marine toxins

“…Intracellular Ca 2+ measurement Intracellular Ca 2+ transients [Ca 2+ ] i evoked in undifferentiated mouse ES cells by poly-APS (1) and its synthetic analogues APS12 (9a) and APS12-2 (9b) were measured as previously reported for studies on halitoxin [25] and poly-APS [20]. Cells were incubated in the dark for 1 h in NaClbased extracellular solution containing 0.01 mM fura-2AM (Sigma, 1 mM stock in dimethylformamide).…”

“…Matrix-assisted laser desorption ionisation-time of flight (MALDI-TOF) spectrometric analysis of 1 showed an ion cluster centred at m/z 5,520 Da with a low degree of polydispersity and a less intense and broader one at 18,900 Da [28]. Poly-APS (1) is related structurally to the epidermal growth factor (EGF)-active alkaloid from the sponge Callyspongia fibrosa [8] and to the halitoxins and amphitoxin isolated from several haplosclerid sponges [1,3,24,25]. Halitoxins and amphitoxin include polymeric compounds containing pyridinium rings with wide variations in chemical structure mainly with respect to the degree of polymerisation and to the monomer structure, which shows a linear, branched or unsaturated linking C5-C12 alkyl chains.…”

Mechanisms of Toxicity of 3-Alkylpyridinium Polymers from Marine Sponge Reniera sarai

“…Linear and cyclic 3-alkylpyridinium dimers and trimers are reported to be cytotoxic, antimitotic and inhibitory to muscarinic acetylcholine receptors and to some enzymes like histone deacetylase [reviewed in 3,4]. Polymeric compounds on the other hand, like halitoxins [10][11][12], amphitoxins [13], natural and synthetic epidermal growth factor (EGF)-active factors [14,15] and poly-APS [5] (Figure 1), show many more diverse and potent biological activities. Halitoxin was reported to be haemolytic and cytotoxic towards transformed cells, toxic to fish and mice, inhibitory to some Gram positive bacteria, antimitotic and neurotoxic [10,11], and amphitoxin shows antifeedant activity against fish [13].…”

“…Chemical structures of 3-alkylpyridinium polymers isolated from haplosclerid marine sponges. A, halitoxins [10,12]; B, amphitoxins [13]; C, EGF-active factors [14]; D, poly-APS [5]. A B…”