Does nitric oxide derived from spemn function during the Ca^<2+>-transient at fertilization of sea urchin eggs?

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A transient increase of the intracellular Ca^<2+> concentration ([Ca^<2+>]_i)(Ca^<2+>-transient) at fertilization of sea urchin eggs is prerequisite for the egg activation and further development. We have showed the possibility that the ryanodine system via cGMP and cADPR exists more upstream than the IP_3 system in the signaling pathway inducing Ca^<2+>-transients. In some cells, NO is shown to stimulate the production of cGMP by activating soluble guanylyl cyclase. Kuo et al.(2000) have presented the hypothesis that NO is necessary and sufficient for the initiation of Ca^<2+>-transients. We measured the changes of [Ca^<2+>]_i, the intracellular NO concentration([NO]_i), the egg contents of cGMP and IP_3 during fertilization, and obtained the following results. 1)An inhibition of NO synthase slightly elongated the latent period of Ca^<2+>-transients, evidently delay the increase of cGMP and IP_3 contents, but could not abolish the Ca^<2+>-transient. 2)An inhibition of NO-sensitive guanylyl cyclase significantly elongated the latent period of Ca^<2+>-transients, abolished the initial increase of cGMP, delay the increase of IP_3, and diminished the peak of IP_3. 3)NO scavenger, oxyhemoglobin had little effects on the Ca^<2+>-transient. 4)Simultaneous measurements of [Ca^<2+>]_i and [NO]_i could not detect the increase of [NO]_i around the onset of fertilization potential or before the onset of Ca^<2+>-transient. 5)A block of Ca^<2+> channel, especially that of sperm, accompanied with the inhibition of NO-sensitive guanylyl cyclase or IP_3 receptor dramatically elongated the latent period of Ca^<2+>-transient. Thus, we could not obtained the supporting evidences for the hypothesis that NO is necessary and sufficient for the initiation of Ca^<2+>-transients, but confirmed again that cGMP was important in the Ca^<2+>-transient at fertilization of sea urchins.