The effect of partial removal of yolk on the chilling sensitivity of zebrafish (Danio rerio) embryos

The effect of partial removal of yolk on the survival of zebrafish embryos and the chilling sensitivity of yolk-reduced embryos were investigated at several stages of embryo development. Dechorionated embryos were punctured with a sharp microneedle and approximately 50 to 75% of yolk content was released following multiple punctures. The survival of yolk-reduced embryos was found to be stage dependent. Only 7.9% of 26-somite (24 h) embryos survived, whereas 56.7% of prim-6 (27 h), 62.4% of prim-15 (34 h), and 81.3% of high-pec (49 h) embryos survived after partial removal of yolk. For chilling sensitivity studies the yolk-reduced embryos at high-pec stage were cultured in embryo medium for 2, 6, or 24 h to allow embryo recovery before they were chilled at 0 degrees C for 6 h. No significant differences (P > 0.05) were seen in normalized survivals between control and yolk-reduced embryos following a 2- or 6-h recovery period. However, when the recovery period was extended to 24 h, the yolk-reduced embryos showed significant (P < 0.05) higher survival than that of chilled controls and the significance was more pronounced (P < 0.01) after a longer period (10 h) of chilling. Similar results were also obtained with embryos at prim-6 stage. These results indicated that after partial removal of yolk, zebrafish embryos at post-prim-6 stage can survive well and their sensitivity to chilling can be reduced. This may have significant implications in alleviating certain difficulties confronting the cryopreservation of fish embryos.