EXPRESSION OF A CHICKEN CILIARY NEUROTROPHIC FACTOR IN TARGETS OF CILIARY GANGLION NEURONS DURING AND AFTER THE CELL-DEATH PHASE

Ciliary ganglion (CG) neurons, like other neuronal populations, become dependent on their targets for survival during development. We have p reviously purified and cloned a secreted ciliary neurotrophic factor t hat was called growth-promoting activity (GPA). We report here the exp ression and purification of a highly active form of recombinant GPA, t he preparation of CPA-specific polyclonal and monoclonal antibodies, a nd the use of these antibodies to investigate the cellular location an d timing of GPA expression in tissues innervated by CG neurons. Virtua lly all of the trophic activity in extracts of embryonic eyes could be depleted by GPA-specific antibodies. GPA-like immunoreactivity was fo und in both targets of the CG: the arterial vasculature of the choroid layer and the ciliary body of the eye. In the choroid layer, GPA was localized to smooth muscle cells surrounding the choroid arteries. Sta ining in the choroid layer was first detectable at embryonic day (E) 1 0, or about 2 days after cell death has begun in the ganglion, then in creased in intensity through E19. Quantification of trophic activity f rom whole eye extracts at various ages showed a small increase in acti vity observed between E9 and E12 and at least a 10-fold increase betwe en E12 and E18. The presence of GPA protein in target cells of CG neur ons during the specific developmental period when these neurons underg o cell death is consistent with its proposed function as a target-deri ved ciliary neurotrophic factor. (C) 1996 Wiley-Liss, Inc.