MOLECULAR-CLONING AND CELLULAR-LOCALIZATION OF TRKC IN THE CHICKEN-EMBRYO

Degenerate primers directed against conserved regions of the trk and t rkB amino acid sequences were used in the polymerase chain reaction to isolate a 455 bp fragment from embryonic day 3 chicken cDNA encoding the trkC. This fragment was subsequently used to synthesize an anti-se nse trkC cRNA probe which was used in a RNase protection assay of tota l RNA from chicken embryos. trkC mRNA was found in the E2 embryo with increasing levels later in development. In the E9 embryo highest level s were found in brain and spinal cord with intermediate levels in eye, heart, gut and muscle. Low levels were found in kidney, liver, skin a nd yolk sac. Using the 455 bp trkC fragment as a probe in RNA blot ana lyses of poly A+ RNA, a major transcript of 6.3 kb and two minor trans cripts of 3 kb and 10 kb were found. In situ hybridization was perform ed on embryos taken at three stages of development (embryonic day 3, 9 and 19), using a 48-mer antisense oligonucleotide probe for chicken t rkC. Within the sensory nervous system trkC mRNA expression at all age s was confined to the ventrolateral neurons of the spinal sensory and trigeminal ganglia as well as distal ganglia associated with the VIIth , IXth and Xth cranial nerves. Labelling for trkC mRNA was also observ ed within the developing CNS at E3 and the ganglion of Remak at E19. A barely detectable level of expression was observed in the sympathetic chain and no labelling was evident in the proximal ganglia of the cra nial nerves. These results suggest that neurons have a very early capa city to respond to neurotrophin-3 which continues throughout embryonic development. The early expression of trkC mRNA also support the growi ng evidence suggesting a role for neurotrophins in neuronal differenti ation.