Characterization of Ca2+ signaling in the external yolk syncytial layer during the late blastula and early gastrula periods of zebrafish development

Preferential loading of the complementary bioluminescent (f-aequorin) and fluorescent (Calcium Green-1 dextran) Ca2+ reporters into the yolk syncytial layer (YSL) of zebrafish embryos, revealed the generation of stochastic patterns of fast, short-range, and slow, long-range Ca2+ waves that propagate exclusively through the external YSL (E-YSL). Starting abruptly just after doming (similar to 4.5 h post-fertilization: hpf), and ending at the shield stage (similar to 6.0 hpf) these distinct classes of waves propagated at mean velocities of similar to 50 and similar to 4 mu m/s, respectively. Although the number and pattern of these waves varied between embryos, their initiation site and arcs of propagation displayed a distinct dorsal bias, suggesting an association with the formation and maintenance of the nascent dorsal-ventral axis. Wave initiation coincided with a characteristic clustering of YSL nuclei (YSN), and their associated perinuclear ER, in the E-YSL. Furthermore, the inter-YSN distance (IND) appeared to be critical such that Ca2+ wave propagation occurred only when this was similar to 8 mu m was coincidental with wave termination at shield stage. Treatment with the IP3R antagonist, 2-APB, the Ca2+ buffer, 5,5'-dibromo BAPTA, and the SERCA-pump inhibitor, thapsigargin, resulted in a significant disruption of the E-YSL Ca2+ waves, whereas exposure to the RyR antagonists, ryanodine and dantrolene, had no significant effect. These findings led us to propose that the E-YSL Ca2+ waves are generated mainly via Ca2+ release from IP(3)Rs located in the perinuclear ER, and that the clustering of the YSN is an essential step in providing a CICR pathway required for wave propagation. This article is part of a Special Issue entitled: 12th European Symposium on Calcium. 2012 Elsevier B.V. All rights reserved.