The hatching process and its control by environmental oxygen pressures in three teleost species (Fathead minnow, Pimephales promelas; Muskellunge, Esox masquinongy; and walleye, Stizostedion vitreum)

The effects of environmental oxygen levels on the universal process of hatching in animals were examined in three teleost species, fathead minnow, Pimephales promelas, muskellunge, Esox masquinongy, and walleye, Stizostedion vitreum. Short-term exposure (1–2 hrs) to hypoxic conditions stimulated premature hatching as early as 2.6 days (70% of total embryonic development), 3.0 days (78% of total embryonic development), and 4.5 days (80% of total embryonic development) before normal hatching for fathead minnow, muskellunge, and walleye embryos, respectively. Hypoxia induced premature hatching in 50% of late stage fathead minnow, muskellunge, and walleye embryos within 10, 20, and 45 minutes, respectively. Compared to controls, hyperoxia delayed hatching for a minimum of two hours in muskellunge, seven hours in walleye and 50 hours in fathead minnows. Hypoxia did not induce hatching at early developmental stages (< 70%–80% of total embryonic development) before hatching gland cells (HGCs) have matured. HGCs were first identified by light and scanning electron microscopy during eye pigmentation stage in both fathead minnow and walleye embryos and during lens formation stage in muskellunge. The HGCs were located on the head and the anterior and dorsal regions of the yolk sac and were identified by light microscopy as granulated epidermal cells (8–4 μm) that disappeared after hatching. Hypoxia stimulates the release of the hatching enzymes from the HGCs shortly before hatching. Hatching enzymes were collected and electrophoretically separated from crude hatching media and caseinolytic activity was demonstrated on zymograms. Three caseinolytic bands were found in the hatching media of both the fathead (23 kDa, 24 kDa, 25 kDa) and muskellunge (23 kDa, 25 kDa, 30 kDa), and two bands were found in the walleye hatching media (23 kDa, 60 kDa). All enzymes were relatively thermostable and had a pH optima between 6 and 9. Preliminary purification of walleye hatching enzyme by sephadex gel filtration resulted in one SDS-PAGE band (23 kDa) with caseinolytic activity. Hypoxia as a universal physiological stimulant of hatching is discussed and a model for hypoxia-induced hatching is proposed.