Expression and development of the proenkephalin mRNA in the C cells of chicken ultimobranchial glands

Abstract

A large number of enkephalin-immunoreactive cells transiently appear in chick ultimobranchial glands during embryonic development. The expression and development of proenkephalin mRNA were examined in the ultimobranchial glands by in situ hybridization with digoxigenin (DIG)-labeled oligonucleotide probes, in comparison with those of calcitonin mRNA and enkephalin peptide. Proenkephalin mRNA, as well as calcitonin mRNA, appeared in some C cells at embryonal day 14 (E 14), and in many cells at E 16. Subsequently, there is a marked increase in the level of calcitonin mRNA around E 18–19; all C cells exhibited intense reaction for calcitonin mRNA. After hatching, intensity of calcitonin mRNA expression was more and more increased. Northern blot analysis with the calcitonin probe also indicated that calcitonin synthesis of the C cells progressively increased with developmental gradient, and reached to the adult level at 1 month after hatching. On the other hand, intensity of hybridization signal of proenkephalin mRNA was maintained moderately during development. In contrast to enkephalin immunoreactivity, which is markedly decreased after hatching, proenkephalin mRNA expression was consistently detected in many C cells of 1- and 2-month-old chickens. Reverse transcription–polymerase chain reaction (RT–PCR) analysis confirmed that proenkephalin mRNA was obtained in the ultimobranchial glands of not only embryos but also 1-day- and 1-month-old chickens. Furthermore, Northern blot analysis demonstrated that a single band for proenkephalin mRNA was obtained in the poly (A) +RNA isolated from the ultimobranchial gland of 1-day-old chicks. Thus, the present study evidences that proenkephalin mRNA is synthesized in almost all C cells of chicken ultimobranchial glands throughout life. Enkephalin may be essential for C cell function.