Spatiotemporal characteristics and mechanisms of intracellular Ca2+ increases at fertilization in eggs of jellyfish (Phylum Cnidaria, Class Hydrozoa)

Abstract

We have clarified, for the first time, the spatiotemporal patterns of intracellular Ca(2+) increases at fertilization and the Ca(2+)-mobilizing mechanisms in eggs of hydrozoan jellyfish, which belong to the evolutionarily old diploblastic phylum, Cnidaria. An initial Ca(2+) increase just after fertilization took the form of a Ca(2+) wave starting from one cortical region of the egg and propagating to its antipode in all of four hydrozoan species tested: Cytaeis uchidae, Cladonema pacificum, Clytia sp., and Gonionema vertens. The initiation site of the Ca(2+) wave was restricted to the animal pole, which is known to be the only area of sperm-egg fusion in hydrozoan eggs, and the wave propagating velocity was estimated to be 4.2-5.9 mum/s. After a Ca(2+) peak had been attained by the initial Ca(2+) wave, the elevated Ca(2+) gradually declined and returned nearly to the resting value at 7-10 min following fertilization. Injection of inositol 1,4,5-trisphosphate (IP(3)), an agonist of IP(3) receptors (IP(3)R), was highly effective in inducing a Ca(2+) increase in unfertilized eggs; IP(3) at a final intracellular concentration of 12-60 nM produced a fully propagating Ca(2+) wave equivalent to that observed at fertilization. In contrast, a higher concentration of cyclic ADP-ribose (cADPR), an agonist of ryanodine receptors (RyR), only generated a localized Ca(2+) increase that did not propagate in the egg. In addition, caffeine, another stimulator of RyR, was completely without effect. Sperm-induced Ca(2+) increases in Gonionema eggs were severely affected by preinjection of heparin, an inhibitor of Ca(2+) release from IP(3)R. These results strongly suggest that there is a well-developed IP(3)R-, but not RyR-mediated Ca(2+) release mechanism in hydrozoan eggs and that the former system primarily functions at fertilization. Our present data also demonstrate that the spatial characteristics and mechanisms of Ca(2+) increases at fertilization in hydrozoan eggs resemble those reported in higher triploblastic animals.

Identification of jellyfish neuropeptides that act directly as oocyte maturation inducing hormones

“…Laboratory strains of Clytia hemisphaerica ("Z colonies"), Cladonema pacificum (6W, NON5, UN2), and Cytaeis uchidae (17) were maintained throughout the year (Deguchi et al, 2005;Houliston et al, 2010;Takeda et al, 2006). Wild specimens of Cladonema as well as…”

“…These peptides were dissolved in deionised water at 10 -2 M or 2x10 -3 M, stored at -20˚C, and diluted in ASW at 10 -5 -10 -10 M prior to use. TAMRA-WPRPamide (TAMRA-LEKRNWPRP-NH 2 ); was synthesised by Sigma and a 5x10 -4 M solution in H 2 O was injected at 2-17% of the oocyte volume, to give an estimated final oocyte concentration of 1 to 9x10 -5 M (Deguchi et al, 2005).…”

“…1B). These species are induced to spawn by dark-light or light-dark transitions respectively (Amiel et al, 2010;Deguchi et al, 2005;Houliston et al, 2010). Starting from the hypothesis that hydrozoan MIH might comprise neuropeptides, consistent with size filtration and protease sensitivity experiments (Ikegami et al, 1987), we screened synthetic candidate peptides predicted from gonad transcriptome data by treatment of isolated gonads (spawning assay) or isolated oocytes (MIH assay).…”

Comparative biology of cAMP‐induced germinal vesicle breakdown in marine invertebrate oocytes

“…Jellyfish of C. uchidae were obtained throughout the year by repetitive transplantation of strains of colonies, which were originally on shells of the gastropod Niotha livescens captured in the vicinity of Asamushi Marine Biological Station (Aomori Prefecture, Japan) (Hirai and Kakinuma, 1973; Deguchi et al, 2005). Sexually mature female jellyfish survived 1–3 months when fed with Artemia salina nauplii four or five times a week and released 20–50 eggs every day during the culture period, according to light‐dark cycles (Takeda et al, 2006).…”

Novel two‐step Ca2+ increase and its mechanisms and functions at fertilization in oocytes of the annelidan worm Pseudopotamilla occelata

“…Calcium Green‐1 conjugated to 3‐kDa dextran (CGD; Molecular Probes, Eugene, OR, USA) was selected as a Ca 2+ indicator, since it showed greater fluorescence changes in P. occelata oocytes than a 10‐kDa dextran form, which has been used in our previous studies (e.g. Deguchi et al . 2005).…”