Immunolocalization of nitric oxide synthase (NOS) isoforms in ovarian follicles of the catfish, Clarias batrachus and its relation with ovarian activity

• NOS/NO system exists in the fish ovary.
• All NOS types express in theca and granulosa cells in developing ovarian follicles.
• iNOS expresses also in zona radiata of ovulated oocytes.
• Positive correlation between ovarian E2 and NO suggests estrogenic role of NO.

Nitric oxide, a gaseous molecule, is produced during the conversion of arginine to citrulline by the action of NOS isoforms (eNOS, iNOS or nNOS). Role of NO in regulation of mammalian reproduction is well established; however, practically no report is available on fishes. Hence, in the present study, expression of all three isoforms of NOS was worked out in the ovary of Clarias batrachus immunohistochemically during different phases of its reproductive cycle and its relation with ovarian activities. No immunoreactivity of eNOS was observed in the ovary of C. batrachus during the late-quiescence and early-recrudescence phases. While during the recrudescence phase (April and May) it expressed intensely in thecal and granulosa cells of the oocyte-II and III, but immune-intensity decreased in the late-recrudescence and spawning phases (June and July). Similar pattern of immunoprecipitation was also observed in case of iNOS. However, the immunoreactivity pattern of nNOS was quite varied, it expressed moderately only in the nucleus and cytoplasm of perinuclear and oocyte-I stages during late-quiescence phase. While during the early recrudescence phase, the expression of nNOS disappeared completely from the nucleus and cytoplasm, rather it expressed intensely in the thecal and granulosa cells, which declined in the late-recrudescence and spawning phases. Moderate immunoreactivity of iNOS could also be localized in the zona radiata of ovulated oocyte. The intense NOS immunoreactivity in the thecal and granulosa cells coincided with increased levels of ovarian NO and 17β-estradiol content. They exhibited statistically significant positive correlation amongst themselves, suggesting the involvement of ovarian NOS/NO system in oogenesis and steroidogenesis in the catfish.