Analysis of the Breakdown of Cortical Granules in Echinoderm Eggs by Microinjection of Second Messengers

Abstract

Breakdown of cortical granules was investigated by microinjecting Ca-EGTA buffers at various concentrations of Ca²⁺, inositol 1, 4, 5- trisphosphate (IP₃), guanosine triphosphate (GTP) and guanosine 5' -O- thiotriphosphate (GTPγS) into echinoderm (Clypeaster japonicus, Temnopleurus hardwicki, and Scaphechinus mirabilis) eggs using a video microscope. The breakdown started in the region near the injection site and then propagated over the entire surface of the egg. The cortical granules did not synchronously break down side by side, but instead underwent random breakdown, in the same manner as that upon fertilization. The reaction times of the breakdown of the cortical granule after the injection of Ca-EGTA buffers, IP₃, and GTPγS were 0.4 sec, 1.0 sec, and 2.7 sec, respectively, in C. japonicus eggs, and were in the same order in T. hardwicki and S. mirabilis eggs. It is inferred from this order of reaction times that GTP-binding proteins, IP₃ and Ca²⁺ act in this sequence upon fertilization in echinoderm eggs.