PHYSIOLOGY OF INSECT DIAPAUSE. V. ASSAY OF THE GROWTH AND DIFFERENTIATION HORMONE OF LEPIDOPTERA BY THE METHOD OF TISSUE CULTURE

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Abstract
1. Spermatogonia and spermatocytes isolated from the testes of dormant pupae of the silkworms, Platysamia cecropia and Samia walkeri (Cynthia), promptly develop into well differentiated spermatids when cultured in hanging-drops of blood obtained from pupating larvae or developing adults of these species. Comparable development does not occur in cultures prepared in blood obtained from diapausing pupae or from mature larvae prior to a well-defined period before the onset of the prepupal stage. 2. The in vitro development of the sex cells signals the presence of a growth-promoting hormone. In inactive blood, as during the pupal diapause, this hormone is ordinarily in sub-threshold titer. 3. In addition to promoting the maturation of spermatocytes, the hormone causes the in vitro multiplication of blood cells and spermatogonia. 4. Pupal blood is active in promoting the development of germ cells in vitro after the initiation of adult development; the blood of mature larvae becomes active at a stage signalled by the spinning of the inner coat of the cocoon. This timing corresponds precisely with the periods during which the prothoracic glands secrete the growth and differentiation hormone which provokes the pupation of the larvae and the adult development of the pupa. 5. Evidence is presented that the development of the germ cells in vitro reflects the presence in the culture medium of the growth and differentiation hormone secreted by the prothoracic glands. The development of the sex cells in the tissue culture appears to be an exceptionally sensitive test for this hormone. 6. By the use of this test the hormone was found to be non-dialyzable and destroyed or precipitated by exposure to temperatures higher than 75° C. Accordingly, it is concluded that the growth and differentiation hormone is either a protein or a smaller molecule conjugated to a protein.