Mode of Action of some Stimulants of the Hatching Enzyme Secretion in Fish Embryos*

Mode of action of two stimulants of the hatching enzyme secretion, electric current (AC) and potassium cyanide, was analyzed by applying them to Medaka embryos in the presence or absence of suppressants of nervous system-mediated secretion, tetrodotoxin or MS–222. Electric current (AC) stimulated the secretion of the hatching gland of the embryos that had been treated with these suppressants, while potassium cyanide did not. These results strongly suggest that electric current acts as a stimulant of hatching enzyme secretion directly on the gland cell itself, while potassium cyanide stimulates the secretion indirectly, probably through nervous system of the embryo. In the present experiments, it was also shown that Ca²⁺ and ionophore, X-537A, when applied directly to the hatching gland extracellularly, induced a marked secretion-associated morphological change of the gland cells instantaneously. However, it was found that chum salmon prolactin did not induce the secretion-associated morphological changes in the hatching gland cells when it was applied directly to the gland cells in situ or indirectly through embryonic circulation.     

Presence of Active Hatching Enzyme in the Secretory Granule of Prehatching Medaka Embryos

Secretory granules of hatching gland were isolated from a 0.3 M sucrose homogenate of whole medaka embryos at prehatching stage by differential centrifugation, followed by a Percoll density gradient centrifugation. The obtained preparation was almost free of melanosomes and composed exclusively of the secretory granules of hatching gland (hatching enzyme granules), as judged by morphological as well as enzymological criteria.
The aqueous extracts of the purified secretory granules showed a specific choriolytic activity as high as about 40 times that of a partially purified secretory granule preparation, P_1,000, and represented a single protein band with molecular weight of about 21,000 on SDS-polyacrylamide gel electrophoresis. It was also revealed that a major component of the hatching enzyme preparation (P II–0.3 enzyme, 13) purified from the hatching liquid was identical with the 21,000 molecular weight band.
These results suggest that the hatching enzyme is present in the secretory granules of prehatching embryos in an active molecular form.     

Ion gradients in xylem exudate and guttation fluid related to tissue ion levels along primary leaves of barley

The concentration of ions in plant cells and tissues is an essential factor in determining physiological function. In the present study, we established that concentration gradients of mobile ions exist in both xylem exudates and tissues within a barley (Hordeum vulgare) primary leaf. For K⁺ and NO₃^?, ion concentrations generally decreased from the leaf base to the tip in both xylem exudates and tissues. Ion gradients were also found for Pi and Cl^? in the xylem. The hydathode strongly absorbed Pi and re‐translocated it to the rest of the plant, whereas Cl^? was extruded. The ion concentration gradients developed early during leaf growth, increased as the tissue aged and remained under both high and low transpiration conditions. Measurement of the expression profiles of Pi, K⁺ and NO₃^? transporters along the longitudinal axis of the leaf revealed that some transporters are more expressed at the hydathode, but for most transporters, there was no significant variation along the leaf. The mechanisms by which longitudinal ion gradients develop in leaves and their physiological functions are discussed.     

Mechanisms of Hatching in Fish: Secretion of Hatching Enzyme and Enzymatic Choriolysis

SYNOPSIS. Mechanisms of two constituent steps of the hatchingprocess, i.e., secretion of hatching enzyme from the gland cellsand enzymatic choriolysis, in the Medaka, Oryzias latipes, aredescribed. The ultrastructural changes of the hatching glandcells occurring at the initiation of electrically induced secretionas well as of natural secretion were the swelling of each glandcell and the separation of joints of the epithelial cells coveringthe gland cells, followed by a resultant exposure of the apicalpart of the gland cells. These changes, though their triggeringmechanisms are not sufficiently clarified, suggest an interventionof some mechanical stimuli in the initiation of secretion. Decreasein electron density of the secretory granules also occurredimmediately prior to the initiation of secretion. The secreted hatching enzyme was found to dissolve the innerlayer of chorion by attacking the scleroprotein of the innerlayer at some restricted sites and liberating a group of solubleglycoproteins of high molecular weights. This selective digestionappears to be the reason why choriolysis proceeds efficientlyduring a short period of time at hatching.