The induction of glutamine synthetase in cell aggregates of embryonic neural retina: correlations with differentation and multicellular organization

Abstract A rapid increase in the synthesis and accumulation of the enzyme glutamine synthetase (GS) in the neural retina of the chick embryo characterizes the functional differentiation and maturation of this tissue. A precocious increase of GS can be induced in the embryonic retina by hydrocortisone and related corticosteroids. This paper presents evidence that the responsiveness of neural retina cells to GS induction by the hormonal inducer is dependent on histotypic associations and organization. This was demonstrated, using retina from embryos of different ages, by comparing GS induction in cultures of intact retina tissue with that in aggregates of retina cells and in monolayer cultures of retina cells. At all embryonic ages tested, induced GS activities were always highest in intact retina tissue, intermediate in cell aggregates, and lowest in cell monolayers. While in the intact retina GS inducibility increased with embryonic age (as judged by enzyme levels attained after 24 hours of induction), in cell aggregates it declined with embryonic age; thus, GS activities in aggregates of retina cells from 10-day embryos were higher than aggregates of cells from older embryos. This reduction of GS inducibility in cell aggregates coincided with the age-dependent decline in cell aggregability, i.e., with the fact that suspensions of retina cells from older embryos formed smaller aggregates than younger cells under the same experimental conditions, and (as known from other work) they failed to reestablish in the aggregates patterns or organization similar to those found in aggregates of younger neuroretinal cells. It was demonstrated that the propensity of dissociated neuroretinal cells for histotypic reaggregation reflected inversely the state of differentiation of the cells at the time of their dispersion: the more differentiated were the cells at the time of dissociation, the lesser was their aggregability and the lower was their GS inducibility. The effects of embryonic age and of retina differentiation on the aggregation of dissociated retina cells were experimentally separated: using 12-day embryonic retina it was shown that cells from its least differentiated area formed larger and more inducible aggregates, while cells from the most differentiated formed smaller, much less inducible aggregates. It was established that the size of cell aggregates, as such, is not the decisive factor in GS inducibility: by increasing the speed of gyration of flasks with cell suspensions, the size of aggregates of 10-day retina cells were reduced to that of aggregates of 14-day retina cells, without significantly lowering their inducibility. It was possible to demonstrate that GS induction in older retina was no more sensitive to damage by trypsin than induction in younger retina, and therefore, trypsinization in itself could not account for the lower inducibility of aggregates of older retina cells. The evidence, as a whole, favors the interpretation that GS inducibility of neuroretinal cell aggregates is dependent on the multicellular organization of the cells, i.e., on the ability of the dispersed cells to become reassociated and reorganized in aggregates in a manner conducive to responsiveness to the inducer of GS. In coaggregates of 16-day embryonic retina cells with 10-day retina cells, GS inducibility of the 10-day cells was reduced, presumably due to interference by the older cells with the reorganization of the younger cells. The nature of the cellular associations and organization of retina cells in aggregates specifically required for GS induction are unknown; the general implications of the reported findings to regulatory mechanisms of enzyme induction in embryonic cells are briefly discussed.