| I have employed biochemical and molecular techniques to investigate the role and regulation of apoptosis during ovarian growth and atresia in teleost fish, using rainbow trout and goldfish as experimental models. These investigations have shown that apoptosis of somatic cells in the follicular envelope is a constitutive feature of follicular development in both species, from primary growth stages through to postovulatory regression. Furthermore, apoptosis can be attenuated in vitro by treatment with salmon gonadotropin, 17β-estradiol and epidermal growth factor, hormones that have previously been shown to promote cell proliferation in teleost follicles. Collectively, these data suggest multiple mechanisms by which selected hormones can influence follicular growth in teleosts. In contrast to the mammalian and avian ovary, teleost follicles do not show a pronounced increase in granulosa cell apoptosis at the onset of atresia. This finding led to a novel hypothesis to explain the biochemistry of follicular atresia in teleosts. This alternative hypothesis predicts that atretogenic factors induce enhanced proteolytic enzyme activity, in turn promoting the hydrolysis and resorption of the abundant yolk storage proteins sequestered within the oocyte. To test this hypothesis, lysosomal protease enzyme activities were measured in rainbow trout oocytes after prolonged serum-deprivation. The results showed a pronounced time-dependent increase in cathepsin L, but not cathepsin D, activity during serum-free culture, and a concomitant increase in oocyte free amino acid content that confirms enhanced yolk proteolysis. Over the same time period, only small and transient increases in follicular apoptosis were observed. The mechanistic basis of this increased protease activity was hypothesized to result from reduced expression of endogenous cysteine protease inhibitors. An oocyte-specific cysteine protease inhibitor (OCPI) was subsequently sequenced and characterised in the rainbow trout. Semi-quantitative RT-PCR revealed that mRNA expression of this putative inhibitor is greatly reduced in atretic follicles. Collectively, these data provide the foundation for a new model to explain the biochemical events characterizing follicular atresia in teleost fish, with potential relevance to other oviparous vertebrates. |
