Calcium release and development of heat-shocked porcine oocytes after nucleus-ooplasm reconstruction

We determined the effect of heat shock (HS) on the alterations of development and calcium releasing capacity of nuclear-ooplasmic reconstructed porcine oocytes stimulated by thimerosal. The non-HS (39 degrees C) and the HS2h (41.5 degrees C for 2 h) matured oocytes were enucleated and their spindles/chromosomes were exchanged between these two groups followed by parthenogenetic activation. In the Control group (Csp-Coop), the non-HS spindle (Csp) was transferred to the non-HS ooplasm (Coop). Blastocyst and cleavage rates were higher in both Csp-HSoop (non-HS spindle transferred to the HS ooplasm) and HSsp-Coop (HS spindle transferred to non-HS ooplasm) reconstructed oocytes, but no difference was detected in the average cell number per blastocyst. However, intracellular calcium concentrations ([Ca(2+)](i)) generally declined (p < 0.05) in the reconstructed HS oocytes, with a greater blastocyst rate after parthenogenetic activation. In the present study, time for the completion of spindle transfer in these oocytes was 1-2 h, during which some physiological remodeling or adaptation might have been occurred in the oocytes. Therefore, changes in heat-shock protein70 (HSP70) expression and developmental competence of the HS2h oocytes with 1 or 2 h of recovery time under normal culture temperature (39 degrees C) were examined. The results showed that the expression of HSP70 in the HS2h oocytes was higher (p < 0.05) than those had recovery incubation for 1 h (HC1h) after HS, but the cleavage and blastocyst rates were greater (p < 0.05) in the HC1h group. We demonstrated that a recovery period prior to activation of porcine oocytes and reconstructed oocytes is beneficial to further development. Heat shock to either the karyoplast or the ooplasm enhances embryonic development but reduces intracellular calcium release in the cloned porcine oocytes.